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https://f1000research.com/articles/12-1416/v2	08 Nov 23	{ "type": "Research Article", "title": "Politeness when native Japanese speakers and Uzbek Japanese learners give warnings about prohibited acts: a cross-sectional study", "authors": [ "Takashi Ninomiya", "Masaki Ono", "Munojot Umarova" ], "abstract": "Background Based on signs prohibiting certain actions, people warn others using imperative forms of the verb or give a soft warning using politeness strategies. This study investigates actual situations in which native Japanese speakers and Uzbek learners of Japanese give warnings about prohibitions.\n\nMethods\nThis study compared warnings given by native Japanese speakers and Uzbek Japanese learners regarding prohibited acts. After clarifying the difference between prohibition and warning of prohibitions, we proposed a classification scheme for the latter speech act in terms of politeness. Data were elicited using a discourse completion task.\n\nResults\nThe results showed that the Japanese speakers tended to mitigate warnings by implementing two politeness strategies: expression in hedged forms and positioning the warning as a rule. The Japanese speakers used expressions such as “It is prohibited” to indicate that they warned as a rule. Such expressions may cause the hearer to feel that the speaker is an overbearing person who wields authority. To avoid this risk, the Japanese speakers used hedges, such as “Looks like it’s prohibited,” or “It says it’s prohibited,” to make it clear that the right to forbid is not with them but with the public authority. In contrast, the Uzbek learners of Japanese tended to implement their politeness strategy as an apology to compensate for the explicit warning. They tended to apologize in situations where the Japanese speakers would not.\n\nConclusions\nThe present study presented a framework for categorizing warning of prohibitions. This framework can be applied to languages other than Japanese, Uzbek, and Japanese by Uzbek learners.", "keywords": [ "Warning of prohibitions", "representative", "directive", "politeness strategies", "native Japanese speakers", "Uzbek Japanese learners" ], "content": "Introduction\n\nThis study is interested in the relationship between signs indicating prohibitions and linguistic expressions in countries around the world, especially in Asian countries. We are interested not only in what kind of linguistic expressions are used in signs indicating prohibitions, but also in how a speaker of a language would warn someone who is committing a prohibited act after seeing a prohibition sign. For instance, they can order a person smoking in a non-smoking area not to smoke, request the person not to smoke, or inform the person of the fact that smoking is prohibited. We are interested in warning strategies in Japanese and other Asian languages. Interests include not only Japanese as a first language, but also Japanese as a second language. The purpose of this study is to clarify pragmatic features when native speakers of Japanese and Uzbek learners of Japanese give warnings about prohibited acts. Japanese and Uzbek, both of which are Asian languages, are agglutinative and have a subject-object-verb structure and similar usage of auxiliary verbs: e.g., both -te miru in Japanese and -b koʻrmoq in Uzbek mean “to try to Verb” using verbs derived from the word “to see” (Yamazaki 2017). For this reason, one might expect that Uzbeks could learn Japanese grammar relatively easily. However, even if learners are familiar with Japanese grammar and can produce grammatically correct sentences, they may not be able to successfully implement Japanese speech acts that include warning about prohibitions.\n\nThere are two reasons for focusing on Uzbek learners of Japanese. First, as Japanese language education has flourished in Uzbekistan in recent years (Iwasaki & Umarova 2019: 231), we expect research on Uzbek learners of Japanese will continue to develop. In order to avoid communicative misunderstandings between Uzbek Japanese learners and native Japanese speakers, it is necessary to clarify the pragmatic characteristics of learners. Second, our 2021 survey1 examined warning of prohibitions by native speakers of Japanese, as well as by Chinese, Korean, and Uzbek learners of Japanese, but failed to elucidate the characteristics appertaining to Uzbek learners. To address this oversight, this study raises the following research questions: 1) Which politeness strategies do native Japanese speakers and Uzbek Japanese learners use more frequently when warning of prohibitions? 2) What what pragmatic features other than the politeness strategies do the two language groups display?\n\nStudies of speech acts and politeness\n\nWarning of prohibitions is a speech act taken to avoid possible bad consequences after someone (the “hearer” or “addressee”) is observed performing a prohibited act. Searle (1976) proposed five basic kinds of illocutionary acts: representative, directive, commissive, expressive, and declaration. Of the five illocutionary acts, Erler (2020: 38) classified the speech act of prohibition as directive. Bach and Harnish (1979: 49) defined prohibition as follows:\n\n“The speaker believes that the utterance prohibits the hearer from doing something because of the authority the speaker has over the hearer; the speaker expresses the intention that the hearer should not do what is prohibited because the speaker says so.”\n\nFor example, a guard in a museum can prohibit a visitor from taking photos by saying “Photography is prohibited.” Even if the visitor’s friend says the same thing, they cannot prohibit the act. Since the friend has no authority, they give a warning, not utter a prohibition.\n\nBataineh and Aljamal (2014: 88) explained warning as follows: “warning refers to the different strategies used for getting the attention of the addressee and alerting him/her to a specific danger or bad consequences.” By saying “Photography is prohibited,” the friend tries to prevent a bad consequence, such as the museum guard scolding the other party. The same expression can either be a prohibition or a warning, depending on whether the speaker has authority or not. According to Searle (1976), warning can function not only as directive (1a), but also as representative (1b). Searle (1976: 22) maintained that warning “may be either telling you that something is the case (with relevance to what is or is not in your interest) or telling you to do something about it (because it is or is not in your interest).”\n\n(1) a. I warn you to stay away from my wife! (directive)\n\nb. I warn you that the bull is about to charge. (representative)\n\nWarning of prohibitions is a speech act that can cause the hearer to lose face. Although a speaker can baldly warn the hearer, they can also implement politeness strategies by apologizing for performing a face-threatening act (FTA). Brown and Levinson (1987) identified 14 positive and 10 negative politeness strategies. For example, a speaker can use questions or hedges or adopt the word “sorry” as a politeness strategy in warning situations.\n\nStudies of prohibition and warning of prohibitions\n\nHashimoto et al. (1992) investigated warning of prohibitions in eight languages including Japanese and proposed 10 categories of this particular speech act. In the “No photography” situation, the Japanese used the category “Value judgment about a rule” (e.g., Koko de satsuee-shite wa ikemasen “It’s wrong to take photos here”) most frequently. The study created different categories for warning of prohibitions and conducted a quantitative analysis. The study then formulated a classification system focusing on social parameters such as status, but did not analyze and discuss how warnings are given in terms of politeness. Previous studies on prohibition have often covered linguistic landscapes: e.g., signboards in public places in Japanese and Korean (Kim 2011), and signs on university campuses in China and Japan (Wang 2017). Kishie (2008) analyzed prohibition expressions in the “No dumping” situation in the Tokushima dialect. He established the following five categories: Direct Prohibition I (e.g., Suteruna “Don’t throw out the trash!”), Direct Prohibition II (e.g., Sutetara dame “It’s wrong to throw out the trash”), Indirect Prohibition (e.g., Suterarenaiyo “It’s not possible to throw out the trash”), Request I (e.g., Sutenaide “Don’t throw out the trash!”), Request II (e.g., Motte kaeriyo “Take your trash and go home!”), and Reason (e.g., Kyoo wa gomi no hijanaiyo “It’s not garbage day today”). Because his description of some categories was insufficient, the precise forms of the categories are not entirely clear. The definition of categories in Kishie (2008) requires some caution. For instance, concerning Request I, Kishie (2008: 41) gave only the imperative form, stating that it is used when the speaker orders the hearer not to throw garbage away. However, the study of Blum-Kulka et al. (1989) set nine levels of request in terms of indirectness. If Request I corresponds only to the imperative form (i.e., mood derivable) and does not include the other categories, it is not appropriate to label the category with the name Request. One should refer to it as the imperative. Also, despite the formal difference between Direct Prohibition I’s Verb-na (prohibited form) and Request I’s Verb-naide (negation + te-form), both are pragmatically classified as imperative in Blum-Kulka et al. (1989). Regarding Uzbek prohibition, Kobilova (2020) only enumerated prohibition expressions in Uzbek and English and did not discuss them. The study of prohibition in Uzbek did not indicate a category of prohibition.\n\n\nMethods\n\nSince the members of the pragmatics research project2 are university faculty members, we asked university students and university graduates, from whom it is relatively easy to collect data, to cooperate in the survey. Survey participants of this study fall into three groups: 36 native speakers of Japanese (Jap.), 36 Uzbek speakers who were learning or had learnt Japanese (UJap.), and 36 native speakers of Uzbek (Uzb.). We focused on Jap. and UJap. surveys and also analyzed the Uzb. data as a reference. With regard to the Jap. group, faculty members from Japanese universities (Dr. Ono and Dr. Ju) sent a questionnaire in Japanese to Japanese students and colleagues at their own universities, as well as to Japanese faculties and students at universities with whom they had a connection. Ms. Umarova and Ms. Turdiyeva, who teach at a university in Uzbekistan, collected the data on UJap. and Uzb. The UJap. data were obtained from Uzbeks who were studying or had studied Japanese at Uzbek or Japanese universities. The Uzb. data were obtained from Uzbek students enrolled at the university where Ms. Umarova and Ms. Turdiyeva teach from their Uzbek colleagues, and from Uzbek faculty members and students with whom they were acquainted. The UJap. participants’ Japanese language level was basically intermediate (i.e., level between N3 and N2 of Japanese language proficiency)3. The Uzb. data were collected and analyzed to investigate the possibility of Uzbek interference with the UJap. data.\n\nThe current study collected the data by means of a discourse completion task (DCT). Members of the above research project created scenarios for the DCT with university students and graduates as respondents. The project members assumed that the students would visit a museum and a hospital and would use warnings based on signs displaying prohibitions. Situations were set up in which a student sees two prohibition signs in a museum and two prohibition signs in a hospital. In order to investigate the social relationship between the speaker and the hearer, the members created eight scenarios, focusing on the social parameters of social distance (SD)4 and age as shown in Table 1 (see extended data for English translation of the scenarios).\n\nSee Situation 1-(i) and Situation 2-(i). Although the situations are the same, the social parameters with the hearers are different. In Sit. 1, the speaker warns a friend who is around the same age as the speaker (x=y). Also, in Sit. 2, the speaker warns a professor who is older than the speaker (x<y). In Sits. 3 and 4, the speaker warns a stranger (+SD). The respondents had to consider the social distance and age between himself/herself and the person who was about to commit the prohibited act. The DCT required the respondents to say something in response to the offender’s action, based on a sign that showed what was prohibited. After creating the scenarios, we created a questionnaire form in Google Docs (see extended data) and sent the link to the Jap., UJap., and Uzb. groups. The Jap. participants performed the task in Japanese, the Uzb. participants did the task in Uzbek, and the UJap. participants did the task in both Uzbek and Japanese. We gave the participants descriptions of the four situations to elicit warning expressions about a prohibition in oral conversation. At the beginning of the questionnaire, it was stated that only those who were willing to cooperate with the survey should respond and that we would take steps to protect their personal information. The response data from the Google form was exported to an Excel file (see underlying data). In cases where the data were incomprehensible, we contacted the respondent concerned and asked him/her to clarify his/her response (the incomprehensible data were marked in yellow on the sheet).\n\nWe created a scheme, based on request and prohibition studies comprising Blum-Kulka et al. (1989), Trosborg (1995), Kishie (2008), and Bella (2012). We presented two head acts and one supportive move. A head act is the minimal unit which can realize a warning of prohibition and is the core of the warning sequence. A supportive move is a unit external to the warning of prohibition, which modifies its impact by mitigating its force. The two head acts consist of the request-type and prohibition-type as presented by Kishie (2008). The request-type can be broken down into the following eight categories in terms of indirectness: Level 1 imperative “Don’t take photos!”; Level 2 explicit performative “I ask you not to take photos”; Level 3 hedged performative5 “I would like to ask you not to take photos”; Level 4 obligation statement “You should not take photos”; Level 5 want statement “I want you not to take photos”; Level 6 suggestory formula “How about not taking photos?/Let’s not take photos”; Level 7 query preparatory “Could you not take photos?”; Level 8 hint6 “You can take photos in another place.” We called these categories Directive-Warning (DW) because the request-type categories can only be interpreted as directive.\n\nAs the second head act, we proposed Representative-Directive-Warning (RDW), which corresponds to Kishie’s (2008) prohibition-type. This type can be broken down into the following categories: Level 1 explicit infringement-indication “Photography is prohibited”; Level 2 hedged infringement-indication “Looks like photography is prohibited”; Level 3 fact-checking “Isn’t it wrong to take photos?” As indicated in section “Studies of speech acts and politeness,” a warning functions as both a directive and representative act. The sentence “Photography is prohibited” functions as a directive in that the speaker stops the hearer from being about to take photos, but superficially it seems to be representative. Since there are sentences that have both representative and directive features, we made the RDW-type. In RDW-type expressions, the speaker points out that the hearer’s action is morally or legally problematic. As Blum-Kulka et al. (1989) classified explicit and hedged performatives as request strategies, this study classified explicit and hedged infringement-indications in the RDW-type categories. The RDW-type’s fact-checking corresponds to the DW-type’s query preparatory. After examining examples of this type from a different point of view than indirectness, we found that there are characteristic differences between Jap. and UJap. in the form of the predicates. We set up four predicate forms, “Prohibited to Verb”-form, “Not possible to Verb”-form, “Wrong to Verb”-form, and “Right to Verb”-form (Verb is hereinafter abbreviated as V), and found a difference in their utilization between the Jap. and UJap. groups.\n\nBased on Blum-Kulka et al. (1989) and Bella (2012), we made the following classification scheme for supportive moves: (i) preparator “I have a favor to ask. (Can you stop calling here?)”; (ii) getting a precommitment “Can I have a word with you? (Looks like drinking isn’t allowed)”; (iii) grounder “There is a sign that says, “Staff only!” (You can’t enter there)”; (iv) alternative “(You can’t enter there) I’ll show you to a restroom”; (v) apology “Sorry. (Photography is prohibited here).” One can place the first and second categories not only in the context of a request, but also in the context of a warning. The third category, grounder involves external mitigation where the speaker gives reasons, explanations, or justifications for his/her warning. The fourth alternative corresponds to Blum-Kulka et al.’s (1989) “promise of reward.” In the case of a request, the speaker may give the hearer a reward, but in the case of a warning, the speaker may suggest to the hearer an alternative plan to replace the prohibited act instead of giving a reward. Regarding the fifth category, Bella (2012), who investigated linguistic mitigation in Greek requests, set this category as one of the supportive moves.\n\nBased on the above, this study created two head acts, the DW-type (eight subcategories) and the RDW-type (three subcategories), and one supportive move (five subcategories). The RDW-type was also classified into four categories according to predicate form. In the utterance of a warning of a prohibition, either a head act of the DW or RDW-type is mandatory, but a supportive move is not necessarily mandatory.\n\nWe obtained the following data sets from the participants: The number of data sets was as follows: Jap. 288, Uzb. 288, and UJap. 288 (total 864). We labeled all the utterances pertaining to warning of prohibitions as follows.\n\n(2) Byooin desukara, koko de tabe tari, non dari wa ikemasen.\n\nGrounder Explicit infringement-indication (Wrong to Verb)\n\nThis is a hospital, so it’s wrong to eat or drink here.\n\nThe example (2) above is from Situation 3-i “No food or drink” in Uzbek Japanese. All labeling data is placed under the underlying data. In the example, there was one explicit infringement-indication for the RDW-type, and one grounder for the supportive moves. In the RDW-type, one “Wrong to V”-form was counted. We counted the number of subcategories of the DW-type, RDW-type, and supportive moves for each of the eight scenarios. We labeled the data that indicated no warning as “Don’t do FTA,” and their number was also recorded. We also calculated the percentages of the subcategories of the DW-type and RDW-type. The number of each subcategory was divided by the total number of head acts. To calculate the utilization rate of the four predicate forms of the RDW-type, the number of each predicate form was divided by the total number of RDW-types. To calculate the utilization of the three subcategories of supportive moves, the number of each subcategory was divided by the total number of supportive moves. In addition, we analyzed each social parameter. Since there were many apologies among the supportive moves, we counted the number of apologies in each social parameter (i.e., social distance and age), and divided the number of apologies by the total number of apologies for each social parameter. To obtain the utilization rate of “Don’t do FTA” for each social parameter, we counted the number of “Don’t do FTA” for each social parameter, the number of each social parameter was divided by the total number of “Don’t do FTA.” To obtain the utilization rate of the DW-type, RDW-type, supportive moves, and “Don’t do FTA,” the number of those four semantic formulae was divided by the total number of semantic formulae. Calculations were made using Microsoft Excel (see extended data).\n\n\nResults\n\nTable 2 indicates the DW-type categories and examples found in the Jap. and UJap. data. Contrary to our expectations, we did not find any examples of categories other than the imperative, want statement, and hint. Also, we did not find any examples of categories other than imperative, obligation statement, suggestory formula and hint in the Uzb. data7. Negative imperative forms that would stop the hearer’s action were classified as imperative. The desiderative form in which the speaker wishes the hearer to stop an action, were classified as want statements8. Vague warning without a specific form was classified as hint. We can say that these three languages lack diversity in relation to the DW-type.\n\nTable 3 shows the categories of the RDW type. We gave the auxiliary verb mitai “looks like” as an example of hedged infringement-indication in the table. In addition, there are also hedges such as omoimasu “I think that” and tte kaite aru “It says that” in both Jap. and UJap. verbal phrases. Regarding the category fact-checking, this study shows that Jap. had examples of this but UJap. did not.\n\nWhen the predicate of a warning sentence had any of the four predicates listed in the “Data analysis” section and the infringement was explicitly noted without hedging, the sentence was classified as explicit infringement-indication. When the predicate of infringement-indication was hedged, the sentence was classified as hedged infringement-indication9. For sentences in which the hearer is asked if his or her action is prohibited, the sentence was classified as fact-checking. The sentences in which explicit infringement-indication was interrogativized were classified into this category.\n\nFigure 1 presents the percentages of each category of the head acts for which at least one example was found. Regarding explicit infringement-indication, which is the most direct in the RDW-type, the UJap. group used it almost twice as much as the native groups. In regards to hedged infringement-indication, which is more indirect than explicit infringement-indication, the Jap. group used it about 1.5 times more than the other groups. The Jap. participants used very few imperative forms compared to the UJap. and Uzb. participants. Regarding hint, Uzb. used it the most and UJap. used it the least.\n\nAs shown in Table 4, we divided the three RDW-type categories into four in terms of predicate form. The first “Prohibited to V”-form means that the hearer’s action is not permitted according to a rule. The second “Not possible to V”-form has the meaning that the hearer cannot perform the action. Using the third “Wrong to V”-form and the fourth “Right not to V”-form, the speaker communicates that the hearer’s act is bad and not good based on the speaker’s value judgment10. If the speaker warns by using the second, third and fourth forms without adding an expression like “It’s a rule” or “There’s a no photography sign over there,” the basis for their warning is not due to a rule, but a value judgment. However, if the speaker uses the first form, they can warn the hearer on the basis of the rule, even without giving a reason for the warning. Concerning the third “Wrong to V”-form, we set two forms: dame and ikenai. As described below, we found differences in the utilization of these two forms between the Jap. and UJap. groups.\n\nFigure 2 shows the percentages of the four predicate forms in the RDW-type categories. The Jap. group used the “Prohibited to V”-form which conveys the fact of prohibition most frequently at about 70%. The Uzb. group used the “Not possible to V”-form most frequently at about 50%. The UJap. group used “Wrong to V”-form as often as “Prohibited to V”-form at about 40% for both forms. When subclassifing the “Wrong to V”-form in the Jap. and UJap. data, in the Jap. group, we found 36 words for dame and 8 words for ikenai, and in the UJap. group, we found 19 words for dame and 58 words for ikenai. In other words, for the “Wrong to V”-form, the Jap. group used the word dame more frequently, while the UJap. group used the word ikenai more frequently.\n\nAs Table 5 shows, Jap. and UJap. groups did not produce any examples of preparator and getting a pre-commitment in the supportive moves. The Uzb. participants also gave no examples of these categories. The DCT data revealed that an apology11 almost always preceded a head act.\n\nAs shown in Figure 3, apology was the most frequent in the supportive moves. The UJap. group used apology about 20% more frequently than the native groups.\n\nFigure 4 shows the number of apologies by social parameter. While Jap. participants apologized only to +SD, namely unknowns, UJap. participants apologized to not only +SD but also -SD, whom the speaker knows well. However, for -SD and x=y, the hearers who are close to the speaker, the UJap. group did not apologize as much.\n\nParticipants noted that they do not warn the hearer in some situations. We categorized the case as “Don’t do FTA.” Table 6 indicates the number and percentage of DW, RDW, supportive moves, and Don’t do FTA. The total number was highest for the UJap. group. The group used supportive moves more than the other groups, about twice as often. All the groups used the RDW-type more than the DW-type. In particular, the Jap. group used RDW about eight times more frequently than DW.\n\nFigure 5 indicates that all the groups implemented the “Don’t do FTA” strategy for +SD, namely unknowns. Among the three groups, the UJap. group had the least number of people who implemented “Don’t do FTA,” In other words, the learners’ group warned about prohibitions most frequently.\n\n\nDiscussion\n\nOne of the significant findings from this study was that Uzbek learners of Japanese tended to be verbose. Analysis of the data in Figure 3 and Table 6 revealed that the UJap. group used more supportive moves, especially apology, than the Jap. and Uzb. groups, although the native groups avoided verbose utterances by providing only the necessary information. The verbosity of the learner group would not have been influenced by their native language, but by the characteristics of the learner language. If Uzb. as well as UJap. had a higher total number of semantic formulae, the UJap. verbosity could have been said to be influenced by the native Uzbek language, but this is not likely. Blum-Kulka and Olshtain (1986: 177) asserted that lengthening of speech act patterns and addition of supportive moves in languages spoken by learners, especially advanced learners, are due to the learners’ lack of confidence. They want to get their message across but are not confident in their speaking ability. Cenoz (1995: 5) said that grounder, which provides reasons and explanations to justify the need to make a request, was the most common in the supportive moves that learners used in request. Ito (2002) reported that lengthening of speech act patterns (especially reasons for refusal) occurred in refusal expressions in Japanese made by groups of native Malay speakers. UJap. subjects tended to be at the intermediate level of Japanese language proficiency (N3-N2 levels), but as Blum-Kulka and Olshtain (1986: 177) previously noted, the subjects exhibited “lengthening of speech act patterns” found in advanced level learners.\n\nUnlike the previous studies of request and refusal, this study showed that apology was more common than grounder. In the scenario where the spaker warned about a prohibition, many Uzbek learners of Japanese may have assumed that the speaker does not need to actively explain the reason for the warning because the situation pertaining to the warning is clear and that the speaker expects the hearer to know that one is not allowed to take pictures in certain areas of a museum or use their cell phone in certain areas of a hospital. For this reason, UJap. participants probably implemented the negative politeness strategy of apology to express their feeling of being sorry for having to issue a warning rather than to give reasons for the warning. Also, UJap. participants apologized to both strangers and those who they knew, while Jap. participants apologized only to strangers. The more extensive use of apology in UJap. than in Jap. means that Uzbek learners of Japanese may apologize more readily than Japanese.\n\nAn analysis of the head acts, the core of warning of prohibitions, revelaed that Jap. warning was indirect and that it was a rule-based warning, while UJap. warning was direct and that it was based on the speaker’s value judgment. See the RDW-type’s categories in Figure 1. The results showed that the Jap. group used indirect hedged infringement-indication more frequently than the UJap. and Uzb. groups and that the UJap. group used direct explicit infringement-indication more frequently than the Jap. and Uzb. groups. To a limited extent, the DW-type’s data tended to indicate that Jap. was indirect and UJap. was direct. While the Jap. group did not use the highly direct imperative at all, the UJap. group used it only occasionally, as did the Uzb. group. Also, the Jap. group used almost twice as many hints with the highest level of indirectness compared to the UJap. group, although the overall number was small. The Jap. group tried to get the hearer to stop the prohibited act by moderately communicating the fact that the hearer’s action was not permitted. In contrarst, the UJap. group tried to get the hearer to stop the action by baldly communicating that based on their opinion, the hearer’s action was wrong. Generally, learners of a language have been found to be more direct than native speakers in some studies (Cenoz 1995: 5). For instance, Fukushima (1990) pointed out that offer and request by Japanese learners of English were more direct than the forms used by native English speakers. The Jap. group’s indirectness in this study was expressed by the negative politeness strategy of hedge. Many UJap. participants probably did not use hedged forms, because hedged forms in Japanese are difficult for the learners or because the learners do not know that the Japanese use the hedged forms as a politeness strategy.\n\nAs Figure 2 shows, the Jap. group used the “Prohibited to V”-form most frequently to convey the fact that the hearer’s action was not permitted according to a rule, while in addition to that form, the UJap. group also frequently used the “Wrong to V”-form to convey that the hearer’s rule violation was wrong. Many Jap. participants warned the hearer while gesturing towards the prohibition sign at the same time. This transferred the responsibility for warning to the museum or hospital. In contrast, about half of the UJap. participants themselves took on the responsibility that originally belonged to those institutions and conveyed the opinion that hearer’s action was wrong. The reason the UJap. participants frequently used the “Wrong to V”-form was not due to the influence of their native language, but the non-native language, because the Uzb. group used the “Not Possible to V”-form most frequently, but rarely used the “Wrong to V”-form. The UJap. group may have frequently used the “Wrong to V”-form due to the influence of a Japanese textbook. Of the “Wrong to V”-forms, Jap. participants used the word dame frequently. However, UJap. participants used the word ikenai, which appears early in Japanese language learning; the word ikenai appears in Lesson 15 of a major Japanese language textbook Minna no nihongo elementary I (Three A Network 2012: 126). The UJap. group may have wanted to give a warning by using the “Prohibited to V”-form but since the expression was difficult, the group may have been forced to use the more user-friendly word ikenai. All the language groups used the RDW-type more frequently than the DW-type (see Table 6). It is unclear whether or not the frequent use of the former type is due to the influence of a specific language, such as Japanese, Uzbek, or Uzbek-Japanese. If the language itself is unrelated to this phenomenon, then some individuals simply may not want to use the DW-type as a warning.\n\nWe found that the three language groups did not warn strangers (see Figure 5). When warning violators of the prohibited act, all the groups implemented the “Don’t do FTA” strategy for unknowns. Participants assumed that if they warned an unknown, this could result in a hostile response. A further analysis of the figure showed that the UJap. group was more likely not to implement “Don’t do FTA” for an unknown than were the Jap. and Uzb. groups. This feature indicates the possibility that Uzbek learners of Japanese may inadvertently warn in situations where native Japanese and Uzbek speakers do not.\n\nAlthough participants of the three language groups could show indirectness in the DW-type as well as in the RDW-type, they did not use any indirect strategies of the DW-type except for hint. While request categories and DW-type categories of warning were the same, the frequency of use was different. Harting (2008: 125) showed that in request situations, more than 50% of the Japanese participants used the highly indirect query preparatory and about 20% of them used the moderately indirect want statement. However, in the warning situations, none of the groups used such indirect DW strategies. The participants may have avoided using such indirect strategies because the strategies were too euphemistic or because directive speech acts were not appropriate for the warning of prohibition scenarios.\n\nNone of the three language groups used a preparator or attempted to get a precommitment in the supportive moves. In the warning of prohibition scenarios, although the speaker does not have the authority to prohibit, they can serve as a spokesperson for the authority. Because of this role, the participants may have thought that there was no need to use a preparator to inform the hearer that a warning was about to be given, nor does the speaker feel the need to try to secure a pre-commitment before the warning. The participants in all the language groups would not have used such circuitous strategies because the speaker does not need to adopt a humble attitude toward the hearer in warning situations.\n\nWhen compensating for warning, the Jap. group implemented the two politeness strategies of positioning the warning as a rule and of using hedged forms. Brown and Levinson (1987: 206) pointed out that in the strategy “State the FTA as a general rule,” one way of communicating that the speaker does not want to intrude but is merely forced by circumstances is to state the FTA as an instance of rule, regulation, or obligation. With the “Prohibited to V”-form, many Jap. participants did not want to warn, but may have wanted to imply that they had to warn due to the museum’s rule. Although the use of the “Prohibited to V”-form has the advantage of allowing the speaker to transfer the responsibility for the warning to an authority, there is a danger that the expression alone may lead the hearer to feel that the speaker is an overbearing person who enjoys wielding authority. To avoid this risk, the Jap. group also frequently implemented the second politeness strategy of hedge. As one of several hedge forms, Brown and Levinson (1987: 152) cited the hedge that may be used to distance the speaker from a command by indicating that the speaker’ speech act is attributed to a third-party command. For example, by saying, Satsuee-kinshi tte kaiteruyo “It says no photography,” the speaker can explicitly convey that a third-party, the museum, forbids the action. Also, the statement Soko, tachiiri-kinshi mitaidayo “There, looks like it’s off-limits,” indicates that the hearer’s action is an assumption and uncertainty on the part of the speaker. The speaker may be mitigating his/her accusation by the hedged form mitai. Since the UJap. group did not use those politeness strategies as much as the Japanese group, the UJap. participants appear to have issued blunt warnings. However, they implemented the politeness strategy of apology, feeling sorry for doing the FTA. Brown and Levinson (1987: 187, 189) suggested that by apologizing, the speaker communicates that he or she does not wish the hearer to lose face, thereby partially compensating for the infringement. Many UJap. participants may have believed that the Japanese typically use the apology as a politeness strategy in any situations, or may have thought that an apology was a way of saving the hearer’s face in a situation that could create the tension associated with giving a warning. The fact that the UJap. participants apologized more frequently than the Jap. participants indicates the possibility that Uzbek learners might apologize unnecessarily even in situations where the Japanese do not apologize. Japanese language teachers should tell learners that there is no need to apologize before giving a warning.\n\n\nConclusions\n\nReferring to prohibition and request studies, this study created a classification scheme for warning about prohibitions and compared the speech act used by native Japanese speakers and Uzbek Japanese learners. The Japanese participants frequently used hedged infringement-indication in the RDW-type. They implemented the politeness strategy of hedge to convey the content of a warning in a moderate manner. Also, by using the “Prohibited to V”-form, they implemented the politeness strategy in which one states the warning as a rule. Simply using the “Prohibited to V”-form runs the risk that the hearer feels that the speaker is an overbearing person who enjoys wielding authority. To avoid this risk, the Japanese speaker used the hedge’s politeness strategy (e.g., “It says it’s prohibited”) to make it clear that the right to prohibit rests with the public authority, not with themselves. On the other hand, the Uzbek learners of Japanese tended to apologize. Because of frequently using explicit infringement-indication, it seems as if they gave a blunt warning. To compensate for doing the FTA, the learners implemented the politeness strategy of apology. However, the learners’ overuse of apology means that they may apologize excessively in situations where the Japanese do not.\n\nWe found the two pragmatic features other than the politeness strategies. First, native speakers of Japanese, Uzbek learners of Japanese, and native speakers of Uzbek implemented the “Don’t do FTA” strategy only for unknowns; that is, none of the language groups warned complete strangers. However, as shown in Figure 5, more than half of the participants of all three groups did not respond in this way. Each of the DCT’s questionnaire statements did not ask whether the warning was given in the first place. In the DCT, if we ask not only “What do you say to the hearer to warn him/her?” but also “Do you warn the hearer in this situation?,” the majority of the participants may not warn unknowns. Regarding the second pragmatic feature, all the language groups used the RDW-type more than the DW-type. Although the participants could have chosen conventional indirect warning strategies among the DW-type, they did not use them at all. It is not clear whether speakers consider indirect expressions to be too euphemistic in the context of warning of prohibitions, or not. A future survey will allow us to obtain opinions on the use of indirect expressions in the DW-type.\n\nThe present study developed a classification scheme for warning about prohibitions, based on the Japanese and Uzbek Japanese data, and referring to the Uzbek data. In the future, we will use this framework to analyze this speech act of learners of Japanese other than Uzbeks and native speakers of Asian languages other than Japanese. By examining data from a large number of languages, we can develop a more general classification scheme, and will clarify why the RDW-type was used more frequently than the DW-type and the reason for the non-use of indirect strategies in the DW-type, as these issues were not fully addressed and clarified in this study. In response to the need to expand the study of Uzbek Japanese, we conducted a pragmatic study focusing on Uzbek learners of Japanese as well as native Japanese speakers. In the past, opportunities for Uzbeks to visit Japan were rare. They studied Japanese to learn about Japanese culture and to improve their Japanese language skills, including grammar, reading and listening comprehension, in preparation for the Japanese Language Proficiency Test. Pragmatic competence as an output may not have been much needed. However, in 2019, Japanese policy made it possible for Japanese companies to accept Uzbeks as technical interns, and many Uzbeks are now coming to Japan. In the future, it will be important to investigate the actual situation of Japanese language use by Uzbek learners from the viewpoint of pragmatics, including warning of prohibitions, and to apply the results to their Japanese language learning.\n\n\nEthics and consent information\n\nRetrospective ethical approval for this study on the 01/05/2023.\n\nAll participants provided implicit consent by completing the online questionnaire.", "appendix": "Data availability\n\nFigshare: All data on Politeness when native Japanese speakers and Uzbek Japanese learners give warnings about prohibited acts, https://doi.org/10.6084/m9.figshare.24179064.v1 (Ninomiya 2023).\n\nThis paper contains the following underlying data:\n\n- Raw data.xlsx\n\n- Labelling data: Japanese labelling.docx, Uzbek Japanese labelling.docx, Uzbek labelling.docx\n\nThis paper contains the following extended data:\n\n- Original scinario questionnaires: Japanese questionnaire.pdf, Uzbek Japanese questionnaire.docx, Uzbek questionnaire.pdf\n\n- English translation of questionnaires.docx\n\n- Data analysis.xlsx\n\n\nReferences\n\nBach K, Harnish RM: Linguistic communication and speech acts. Cambridge: MIT Press; 1979.\n\nBataineh RF, Aljamal MA: Watch out and beware: Differences in the use of warning between American and Jordanian undergraduate students. J. Theor. Linguist. 2014; 11(1): 87–110.\n\nBella S: Length of residence and intensity of interaction: Modification in Greek L2 requests. Pragmatics. 2012; 22(1): 1–39. Publisher Full Text\n\nBlum-Kulka S, Olshtain E: Too many words: Length of utterance and pragmatic failure. Stud. Second. Lang. Acquis. 1986; 8(2): 165–179. Publisher Full Text\n\nBlum-Kulka S, House J, Kasper G: Cross-cultural pragmatics: Requests and apologies. Norwood: Albex; 1989.\n\nBrown P, Levinson SC: Politeness: Some universals in language usage. Cambridge: Cambridge University Press; 1987.\n\nCenoz J: American vs. European requests: Do speakers use the same strategies? 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[A comparative study on the expression of prohibition in Japanese and Korean linguistic landscape-focusing on the place]. Meikai nihongo. [Meikai Japanese Language Journal]. 2011; 16(2): 53–62.\n\nKishie S: Shikoku hoogen ni okeru kinshi hyoogen to kinshi hyoogen koodoo. [Expressions and language behaviors of prohibition in Shikoku dialects].Yamaguchi Yukihiro hakase no koki o oiwaisuru kai [Organization to celebrate Dr. Yamaguchi Yukihiro’s 70th birthday], editor. Hoogen kenkyuu no zenei. [The vanguard of dialect studies]. Toyama: Katsura-Shobo; 2008; 29–46.\n\nKobilova NI: Speech acts of prohibition in the English and Uzbek languages. Materials of the republican 23-multidisciplinary online distance conference on scientific and practical research in Uzbekistan Part 7. 2020; 19–20.\n\nNinomiya T: All data on Politeness when native Japanese speakers and Uzbek Japanese learners give warnings about prohibited acts. Dataset. figshare. 2023. Publisher Full Text\n\nSearle J: A classification of illocutionary acts. Lang. Soc. 1976; 5(1): 1–23. Publisher Full Text\n\nThree A Network: Minna no nihongo elementary I. Tokyo: Three A Network; 2012.\n\nTrosborg A: Interlanguage pragmatics: Requests, complaints and apologies. Berlin/New York: Mouton de Gruyter; 1995.\n\nYamashita Y: “Shiyoo” no imi/yoohoo: “Hinan”/ “Ganboo hyooshutsu” no “Shiyooyo.” [The meaning and usage of shiyoo: “Criticism” and “desire expression”: Shiyoo-yo]. Nihongo/Nihongo-kyooiku-kenkyuu. [Studies in Japanese language and Japanese language teaching]. 2014; 5: 91–106.\n\nYamazaki M: Arutaishogo, choosengo to nihongo ni okeru shikaku dooshi no shikoosoo bunpooka yoohoo no tenkai. [Grammaticalization of the tentative usage of visual perception verbs in Altaic, Korean and Japanese]. Gengo shori gakkai dai 23-kai nenji taikai happyo ronbun-shuu. [Proceedings of the 23rd annual conference of the Association for natural language processing]. 2017; 70–73.\n\nWang T: Daigaku-kyanpasu niokeru kinshi-hyoogen no nitchuu taishoo kenkyuu. [The comparative analysis of the refusal expressions in Japanese and Chinese universities]. Nihongo kyooiku hoohoo kenkyuukai-shi. [Journal of Japanese language education methods]. 2017; 24(1): 28–29.\n\n\nFootnotes\n\n1 The results of this research were presented at the Symposium on Japanese Language Education at the University of Tsukuba on February 13, 2021: Ninomiya T., Li G., Lina A., Ju Hy., Gao, Y., Umarova M., Turdiyeva X, Li Ts. X., and Ono M. (2021) “Nihongo bogo washa nihongo gakushuusha ni yoru kinshi irai hyoogen no hyooka: Poraitonesu no kanten kara” [Evaluation of prohibitive and requestive expressions by native Japanese speakers and learners of Japanese: From the viewpoit of Politeness]. In: Shinpojiumu mirai shikoo no nihongo kyooiku 2.0. [Future-oriented Japanese language education 2.0].\n\n2 The presenters in footnote 1 are members of the research group.\n\n3 Of the 36 respondents, 27 indicated their level of JLPT: 0 were at N5, 4 at N4, 9 at N3, 12 at N2, and 2 at N1. The majority of the UJap. respondents were at N2 and N3, which correspond to intermediate level, while there were only a few at N5 and N4, which correspond to beginner level, and at N1, which corresponds to advanced level.\n\n4 According to Blum-Kulka et al. (1989: 15), social distance is associated with familiarity, which indicates the closeness of the relationship between speaker and hearer. If the relationship is not distant, it could be a family member or friend; if the relationship is distant, it could be a person one does not usually greet, or it could be a stranger.\n\n5 Regarding hedged performative, Blum-Kulka et al. (1989: 287) stated that the illocutionary verb denoting the requestive intent is modified by modal verbs or verbs expressing intention. Hedged performative in Blum-Kulka et al. (1989) is created not by inserting a lexical hedge adverb (e.g., somehow, kind of ) into the explicit performative’s sentence, but by adding an auxiliary verb or a verbal phrase to the explicit performative verb.\n\n6 According to Blum-Kulka et al. (1989) and Trosborg (1995), hint consists of strong hint and mild hint. Blum-Kulka et al. (1989) considered the strong hint to be more direct than the mild hint and placed them in separate categories, while Trosborg (1995) placed them in one category.\n\n7 We found one example of an obligation statement in the Uzb. data.\n\n8 In this study, only the form yameyoo, a morpheme of desire -yoo is added to the verb yameru “stop,” which functions as a want statement, was observed. That this morpheme implies desire was described in Yamashita (2014: 93, 98).\n\n9 As with hedged performative in footnote 5, sentences hedged by adverbs were not classified as hedged infrigment-indication.\n\n10 According to Hashimoto et al. (1992: 132), the word ikenai is the result of a value judgment about rules. Therefore, this study positioned the “Wrong to Verb”-form as being due to the speaker’s value judgment.\n\n11 This category’s examples consist of those that could be classified not only as apologizing for warning (e.g., Mooshiwake arimasen ga “I’m sorry but,” Sumimasen ga “Sorry but,”), but also as both apologizing for warning and alerters (e.g., Sumimasen “Sorry/Excuse me”). One cannot strictly classify the examples of “Sorry” as either an apology for a warning or as an alerter. This study labeled the examples that could be interpreted either way as an apology. Whether an apology for a warning or an alerter, the speakers probably said “Sorry” to convey something that was difficult for them to say." }	[ { "id": "250181", "date": "26 Mar 2024", "name": "Roswati Abdul Rashid", "expertise": [ "Reviewer Expertise Applied Linguistics" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nTakashi et al. al (2023) conducted a study to clarify pragmatic features when native speakers of Japanese and Uzbek learners of Japanese give warnings of prohibited acts. This study is executed based on a study that involves research on the classification scheme of prohibition in Uzbek and comparing this statement in Japanese society is very limited. This research design is very detailed, organized and successfully unravels the categorization based on the cultural norms that are influenced in the use of language from these two communities in uttering the warning of prohibition.\nHowever, there is a slight deficiency because this research refers to the research that has gone beyond research more than ten years ago. If Takashi et al. al (2023) have the opportunity, he can add the latest research to the Previous Studies segment to show the latest research on speech acts - warning of prohibition. However, it is not a problem with the existing reference because it is a reference to the theory of politeness and speech acts which is certainly not the latest in the era of 1980s.\nThis study is a qualitative study uses DCT as a tool to collect data. The item involves the situation of a visit to a museum and a hospital and would use warnings based on signs displaying prohibitions. Total respondents for the study were 108 consisting of 36 native speakers of Japanese (Jap.), 36 Uzbek speakers who were learning or had learned Japanese (UJap.), and 36 native speakers of Uzbek (Uzb.). Focus of the study is given to Jap. and UJap. surveys and analyse the Uzb. data as a reference. Japanese respondents need to give feedback in Japanese language and Uzbek respondents need to give feedback in both Uzbek and Japanese language.\nThe analytical framework is appropriate and can be used as a reference for future researchers to study other aspects in the field of warning of prohibition. Besides every analytical interpretation is appropriate against the data that has been identified. The identified data is reliable.\nOverall, the finding identified Japs implemented 2 politeness strategies, namely expression in hedged form and positioning the warning as a rule in uttering the warning of prohibitions. Both expressions are used as a strategy to avoid FTA and as a reference to regulations from the authorities. This is in line with the nature of Japanese society which always takes all rules seriously and does not add facts as stated and only provides important information in every communication.\nMeanwhile, what is interesting from the findings of this study is that instead UJaps uses the phrase apologize in situations while Japs and Uzebs do not use it. This shows that this language habit is influenced by learning the Japanese language from the Japanese society who are always polite in their speech. Whereas the Japanese society in uttering the warning of prohibition does not do so.\n\nOverall, this article has a high value in researching the classification scheme of politeness strategies in uttering warnings of prohibitions. It will be a reference to the field of pragmatics, especially Speech Acts - Prohibition and Request Studies and the learning and teaching of the Japanese language. The framework of this study can be used as a basis for the study of warnings of prohibition for other languages to have a better understanding.\nAnd it is strongly agreed that for future research, more detailed warning of prohibition research needs to be executed for the languages so that a universal classification scheme can be obtained. And the factors that underlies the application of RDW are much wider compared to the DW that will be obtained.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "250178", "date": "18 Apr 2024", "name": "Muhammad Alif Redzuan Abdullah", "expertise": [ "Reviewer Expertise Comparative Applied Linguistic" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nTitle \"Comparison of Politeness Strategies in Warning Speech Acts: A Cross-Sectional Study with Native Japanese Speakers and Uzbek Learners of Japanese\" This improved title is both descriptive and engaging. It succinctly conveys the focus, methodology, and comparative framework of the study while emphasizing its originality and novelty.\nAbstract 1. Lack of immediate clarity about key terms.\n\nThe distinction between \"prohibition of certain actions\" and \"warning about prohibitions\" is not immediately clear. This could confuse readers from the outset about the precise focus of the study. 2. Complexity of language and terminology.\n\nThe abstract uses terms such as \"prohibition',\" \"prohibition warning' \" and \"classification scheme\", which could be considered jargon by readers unfamiliar with the subject area. This complexity may deter some readers or make the study seem less accessible. 3. Lack of specificity in the description of the methodology.\n\nThe abstract could benefit from briefly explaining what the \"Discourse Completion Task (DCT)\" entailed. For example, what prompts or scenarios were used? How were participants instructed? 4. Limited details on the comparative analysis.\n\nWhile the abstract mentions the results of the native Japanese speakers and the Uzbek Japanese learners, more explicit comparisons could be made between the two groups. How do their strategies differ in detail and what might this tell us about cultural or linguistic differences? 5. Lack of implications or applications.\nWhilst the abstract suggests wider applicability, it could emphasize more clearly the possible implications of the findings. For example, how might understanding these politeness strategies benefit language learners, educators, or experts in intercultural communication? 6. Keywords do not fully reflect the abstract content.\nSome keywords such as \"representative\" and \"directive\" may not be directly representative of the primary focus of the study. In addition, more specific terms relating to the comparative nature of the study (e.g. \"comparative study\", \"cross-cultural communication\") could be included. To summarize, the abstract could benefit from clearer language, a more explicit description of the methodology and comparative analysis, and better alignment between the keywords and the content of the abstract. These improvements would enhance the clarity, accessibility, and overall impact of the abstract.\nIntroduction The introduction provides relevant background information, identifies the research gap, and outlines the purpose and significance of the study. However, there are areas where clarity and flow could be improved. 1. Flow and cohesion:\n\nThe introduction could benefit from more fluid transitions between the different points covered. For example, the transition from discussing global and Asian contexts to focusing on native Japanese speakers and Uzbek Japanese learners could be more seamless. 2. Clarity of research purpose:\n\nThe purpose of the study is mentioned but could be more explicitly linked to the identified research gap and issues. 3. Citation not critically reviewed:\n\nMany citations are provided in this section, but they are not critically reviewed and do not demonstrate the urgency of this research.\nLiterature Review\n\nThe literature review could be improved in terms of depth, critical discussion, and relevance to the current study. Here are some observations and suggestions: 1. Insufficient depth and critical discussion:\n\nRepetitive information:\n\nSome information, such as Searle's (1976) classification of illocutionary acts, is repeated without providing new insights or linking it to the study's focus on politeness in warnings. 2. Limited discussion of politeness strategies:\nBrown and Levinson's (1987) politeness strategies are mentioned, but there is no detailed exploration of how these strategies are used in warnings, especially in the context of the study's focus on native Japanese speakers and Uzbek Japanese learners. 3. Identification of gaps without detailed discussion:\n\nStudies such as Hashimoto et al. (1992) and Kishie (2008) are mentioned for their classification systems, but the researcher does not critically evaluate these classifications or discuss their limitations. 4. Lack of integration:\n\nThe literature review appears to list studies and their findings, rather than bringing them together into a coherent narrative that leads to the rationale for the current study.\nRather than listing numerous studies with brief descriptions, focus on a few key studies that are most relevant to the objectives of the current study and delve deeper into their methods, results, and implications. Connect these discussions to the study's focus on exploring the use of politeness strategies in warnings and provide a clear rationale for why the current study is necessary and how it builds on or differs from previous research.\nMethodology 1. Selection of participants:\n\nLack of criteria: The participants are described, but no specific criteria for the selection of participants are mentioned. What are the inclusion and exclusion criteria for participants? Sampling procedure: How were the participants selected from each group? Was it a random sample, a random sample, or some other method? 2. Data collection:\n\nDetails of the procedure: The description of the discourse completion task (DCT) is vague. What instructions were given to the participants? How were the scenarios developed and validated? Provide a detailed procedure of the DCT, including scenario development, participant instructions, and validation process.\n\nLanguage proficiency: For the Uzbek Japanese learners group, their Japanese language level is mentioned, but how was this level determined? Was there an assessment or a test?\n\nConsent and ethics: While it is mentioned that measures were taken to protect personal data, there is no mention of obtaining informed consent from participants or ethical considerations. 3. Data analysis:\n\nAnalysis procedures:\n\nThe section on data analysis is very dense and lacks a clear, step-by-step explanation of how the data were analyzed. How were the DW and RDW type categories assigned to the utterances? Explain the step-by-step process of coding and analyzing the data. Ensure transparency by providing the underlying data or extended data.\n\nTheory synchronization:\n\nThere is no discussion of how the selected theories (Blum-Kulka et al., Trosborg, Kishie, Bella) influenced the data analysis. How were these theories applied to the data? Were there any conflicts or challenges in applying these theories to the data? Validity and reliability:\n\nThere is no mention of how the validity and reliability of the coding scheme or data analysis process were ensured.\n\nTools and software:\nAnalysis tools: Microsoft Excel is mentioned for the calculations, but no qualitative data analysis software or tools were used to support the analysis. Documentation and transparency:\n\nDocumentation: the underlying data and extended data are mentioned but not provided. Transparent sharing of this data could help with the replication of the study. Clarity:\nSome sections are dense with technical terms and classifications that could be made clearer with visual aids such as flowcharts or tables.\nResults 1. Depth and context:\n\nThe frequency data presented in the results provide valuable insights but lack depth. Exploring the reasons for the observed patterns would enrich the interpretation and provide a more nuanced understanding of the results. 2. Comparative analysis:\n\nDirect comparisons between groups are essential to validate the observed differences. The inclusion of statistical tests or comparative analyses would strengthen the results and increase the rigor of the study. 3. Theoretical comparison:\n\nIncorporating the results into theoretical frameworks or previous studies would improve the theoretical contribution of the study. Discussing the extent to which the findings are consistent with or differ from existing research provides context and depth. 4. Presentation of data:\n\nWhile the quantitative data provides valuable insights, the inclusion of qualitative findings or examples would enrich the context and provide a more comprehensive understanding of the warning strategies used by the participants.\nDiscussion 1. Lack of comparative analysis:\n\nIn the discussion, the Uzbek Japanese learners are often compared to the native Japanese speakers and native Uzbekistan groups, but a clear comparative analysis is not always made. Statements such as \" Uzbek Japanese learners participants probably used the negative politeness strategy of apologizing\" lack a direct comparison with the native Japanese speakers group. The lack of a direct comparison makes it difficult to understand the significance of the observed differences and their effects. 2. Assumption of trust as the only reason for verbosity:\n\nIn the discussion, verbosity is attributed exclusively to a lack of confidence, without taking other factors into account. Blum-Kulka and Olshtain (1986) are cited in the discussion to support this claim. While lack of trust may be a factor, other factors such as cultural norms or level of language proficiency should also be considered. 3. Overemphasis on the apology strategy:\n\nThe discussion focuses largely on the apology strategy without examining other supportive measures in depth. The discussion repeatedly mentions the apology strategy but does not address other supportive measures used by learners. This narrow focus cannot fully capture the complexity of learners' communicative strategies. 4. Lack of contextual explanation for directness:\n\nIn the discussion, the directness of Uzbek Japanese learners is attributed solely to their non-native speaker status without examining other contextual factors. Statements such as \"The reason why the Uzbek Japanese learners participants often used the “Wrong to V” form was not the influence of their native language, but that of their non-native language\" lack contextual depth. Contextual factors such as the influence of textbooks or cultural differences have not been sufficiently researched. 5. Unclear influence of textbooks:\n\nThe discussion mentions the influence of Japanese textbooks but does not elaborate on this. The discussion cites the use of the word \"ikenai\" from a textbook without exploring how the content of the textbook may influence learners' language use. Understanding the role of textbooks could provide valuable insights into learners' language acquisition and use. 6. Lack of discussion of cultural factors.:\n\nThe discussion does not adequately address cultural factors that may influence learners' communicative strategies. While the differences between native and non-native groups are mentioned, there is no elaboration on how cultural differences can affect communication. Cultural factors can have a significant impact on communication strategies and their omission limits the depth of the discussion. 7. Lack of discussion of social parameters:\n\nSocial parameters such as warning strangers are briefly mentioned in the discussion, but this aspect is not explored in depth. Statements such as \"We found that the three language groups did not warn strangers\" are mentioned without discussing the implications or reasons for this result. Understanding the social parameters could provide valuable context for interpreting learners' communicative strategies. 8. Insufficient discussion of politeness strategies:\n\nThe discussion briefly touches on politeness strategies but does not elaborate on their role. The discussion mentions strategies such as \"Prohibited to V\" and hedging but does not address their effectiveness or Politeness strategies play a crucial role in intercultural communication, and their inadequate discussion limits the depth of the analysis.\nConclusion 1. Lack of Clear Implications:\n\nIn conclusion, the implications of the results for teaching or practical application are not clearly formulated. Whilst the study discusses the speech acts used by learners and native speakers, it does not provide any insight into how these findings could inform language teaching or intercultural communication. Without clear implications, the relevance of the study for language teaching or practical applications remains unclear. 2. Incomplete addresses of Pragmatic Features:\n\nTwo pragmatic features are mentioned in the conclusion, but their significance or implications are not fully explored. The study mentions the “Don’t do FTA” strategy and the use of the RDW type over the DW type but does not address why these findings matter or what they say about learners' pragmatic competence. A more detailed discussion of these pragmatic features could provide valuable insights into learners' communicative strategies and their development of pragmatic competence. 3. Lack of Direction to Future Research : While the conclusion points to future research, it does not mention specific directions or questions that future studies could address. The study mentions that the framework will be used to analyze other Japanese language learners and that a future survey will be conducted, but it does not specify what aspects these future studies will focus on. A clear direction for future research would help to build on the findings of the current study and overcome its limitations. 4. Limited Discussion of the Background of Uzbek Japanese Learners:\n\nThe conclusion briefly mentions the background of Uzbek Japanese learners but does not address how their background might influence their pragmatic competence. The study mentions that the Uzbek Japanese learners studied Japanese to gain cultural understanding and to pass the Japanese Language Proficiency Test, but does not address how these motivations or their background might influence their language use. Understanding the learners' background and motivations may provide valuable context for interpreting their language use and could help tailor language instruction to their needs. 5. Lack of Reflection on the Limitations of the Study : The conclusion does not reflect on the limitations of the study, nor does it mention areas where the study may fall short. Reflecting on the limitations of the study could provide a more balanced view of the findings and help guide future research. 6. Overemphasis on the Classification Scheme:\n\nThe conclusion emphasizes the development of a classification scheme but does not address its practical utility or limitations. The conclusion mentions the development of a classification scheme based on the data but does not address how this scheme might be applied or its limitations. While the development of a classification scheme is valuable, its practical utility and limitations should be discussed to provide a fuller understanding of its importance.\nReferences Delete the obsolete references. Cite more from current references - the last five years.\nOverall Evaluation While the paper offers valuable insights into the politeness strategies used by native Japanese speakers and Uzbek learners of Japanese when giving warnings, it could benefit from improvements in the title, abstract, introduction, previous studies, methodology, results, discussion, and conclusion sections. In particular, the paper could benefit from a clearer research question or hypothesis, a more comprehensive review of previous studies, a more detailed methodology section, a clearer presentation and interpretation of results, and a more thorough discussion of the implications of the findings. In addition, the paper could benefit from a clearer articulation of practical implications, a reflection on the limitations of the study, and clear directions for future research.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	2	https://f1000research.com/articles/12-1416
https://f1000research.com/articles/12-803/v1	10 Jul 23	{ "type": "Case Report", "title": "Case Report: An in-depth look into apocrine carcinoma of the axilla - a rare case presentation.", "authors": [ "Dr. Suhit Naseri", "Samarth Shukla", "Dr. Sourya Acharya", "Sunita Vagha", "Samarth Shukla", "Dr. Sourya Acharya", "Sunita Vagha" ], "abstract": "Apocrine carcinoma is an extremely rare malignant cutaneous neoplasm that usually arises in areas with a high density of apocrine glands.1 Diagnosis can be challenging as tumours share histological and immunophenotypic characteristics with them. At first evaluation, the disease is often assumed to be benign. There have been approximately 100 reports of apocrine neoplasms in the literature. A 48-year-old male presented with a right axillary mass which increased in size over a period of 2 years. The patient was reported to have had ayurvedic therapy, but his swelling remained unchanged. Axillary lymph nodes were palpable. USG axilla suggested a well-defined fungating solid isoechoic lesion. USG neck did not reveal any abnormality. The mass was surgically excised as a whole by removing the overlying skin with margins and lymph node excision. The patient was diagnosed with primary apocrine carcinoma after surgical excision. The differentials include adenocarcinoma of breast and prostate and apocrine adenoma. There are no established standards for the care of this form of carcinoma due to its rarity and the absence of clinical studies. A literature evaluation and further reporting will aid in developing diagnostic standards and the most efficient treatment options.", "keywords": [ "Apocrine carcinoma", "Axillary mass", "rare presentation", "apocrine neoplasms", "solid lesion", "lymph node excision", "malignant", "surgical excision." ], "content": "Introduction\n\nApocrine carcinoma of the axilla is a seldom-seen form of breast cancer that originates in the axillary sweat glands. This disease has been reported to be an invasive ductal carcinoma subtype, the most prevalent form of breast cancer among women. From a surgical perspective, apocrine carcinoma of the axilla may present as a palpable mass or a non-palpable abnormality on imaging studies, such as mammography or ultrasound. Treatment typically involves surgical excision of the tumor with clear margins, which may be followed by radiation therapy and/or systemic chemotherapy depending on the stage and biology of the cancer. From a pathological standpoint, Large, pleomorphic cells with an abundance of eosinophilic cytoplasm and prominent nucleoli are seen in axillary apocrine carcinomas. Apocrine carcinoma of the axilla is uncommon, but it can still be a clinically relevant diagnosis that has to be treated promptly and effectively to provide the patient the best chance of survival.\n\n\nCase report\n\nIn 2022, a case of a 48-year-old male of South Asian descent, working as a carpenter with an axillary mass presented to the surgery department with a lump in the axilla (Figure 1) diagnosed as primary apocrine carcinoma after surgical excision. The patient presented with a right axillary mass with inflammation of the overlying skin, with occasional serosanguinous discharge which was associated with mild pain in the affected axilla. The patient was reported to have had ayurvedic therapy, but his swelling remained unchanged. The patient had no family history of malignancy. A physical evaluation and USG for both breasts did not find any abnormalities. USG of the right axilla suggested a well-defined solid isoechoic lesion with multiple microcalcifications with prominent vascularity. The mass increased in size over a period of 1 year and measured 8×8×2.3 cm.\n\nSeven axillary lymph nodes were isolated, the largest lymph node was measured to be 4×2×1.5 cm.\n\nGrossly (Figure 2), the tumour mass was white, firm in consistency and measured 4×4×3.5 cm. On the cut section, solid, homogenous blackish areas were identified with the involvement of overlying skin.\n\nMicroscopically, sections from superior, anterior and posterior margins showed unremarkable squamous lining epithelium with unremarkable deeper tissue and adnexal structure with few distended ducts of histopathology. Section from the inferior margin (Figure 3) was positive for infiltration by malignant epithelial cells. Sections also show fibro-collagenous areas with minimal scattered inflammatory infiltrate. Sections from the tumour were also positive for perineural and lymphovascular invasion.\n\nSection from all seven lymph nodes shows histopathological features suggestive of metastatic deposits of epithelial malignancy.\n\nThe epidermis is visible in some portions of the tumour, which has a dermis with mostly papillary cystic architecture. Focal inflammation with numerous benign apocrine glands were noted.\n\nThe tumour with papillary architecture (Figures 4–6) was found to have fibrovascular cores lined by eosinophilic epithelial cells. The patient’s case was discussed in the hospital’s interdisciplinary tumour board and he was considered for adjuvant radiotherapy. A thorough follow-up plan was advised to the patient, but he did not follow through.\n\n\nDiscussion\n\nApocrine carcinoma is an extremely uncommon adnexal malignancy with limited data on histologic prognostic factors and patient outcomes.1 The axilla and adjacent medial upper arm are the most typical sites for apocrine carcinoma.2 The tumour, which derives from pleuripotent adenexal cells capable of eccrine and follicular development, had first been discovered by Goldstein in 1982.3 Reddish-purple subcutaneous nodules and solid or cystic masses are common characteristics of these tumours. Skin ulceration may be a comorbid condition, and they are frequently locally advanced when diagnosed. This tumour has a sluggish rate of growth, is locally invasive, and has the potential to spread to nearby lymph nodes, the lungs, the liver, the bone, and the brain.4 At the time of diagnosis, lymph node metastases were present in over fifty per cent of all reported individuals suffering from apocrine carcinoma. The recommended treatment for these lesions is wide local excision.5 Apocrine gland carcinomas and eccrine carcinomas are the two primary subtypes of sweat gland carcinomas. Apocrine carcinomas appear as hard, rubbery, cystic, solitary or numerous, non-tender masses with red to purple overlaying skin. Eccrine gland carcinomas lack distinguishing clinical characteristics, rendering gross examination diagnosis nearly difficult. They often only affect older individuals and present as quasi-tender, subcutaneous nodules.6 Although the tumour often arises de novo, it can potentially result from previously present benign tumours like apocrine hyperplasia or apocrine adenoma.7\n\nWide local excision is the preferred method of management of primary cutaneous ductal apocrine carcinoma. In addition to excision, chemotherapy as well as radiotherapy have been utilised, but they haven’t significantly reduced mortality or morbidity among patients with either localised or metastatic disease. The total number of reported instances and the amount of follow-up data that currently exists, both seem insufficient for determining the prognosis. Therefore, there is a need for further case accumulation.\n\n\nConclusion\n\nThe rare tumour referred to as apocrine carcinoma has a characteristic but non-specific histological appearance. For determining the prognosis and formulating particular therapy recommendations, the reported cases and follow-up data appear to be insufficient. Hence, additional cases needs to be collected.\n\n\nConsent\n\nWritten informed consent for publication of their clinical details and clinical images was obtained from the patient.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\n\nReferences\n\nRobson A, Lazar AJF, Ben Nagi J, et al.: Primary cutaneous apocrine carcinoma: a clinico-pathologic analysis of 24 cases. Am. J. Surg. Pathol. 2008; 32(5): 682–690. Publisher Full Text\n\nDhawan SS, Nanda VS, Grekin S, et al.: Apocrine adenocarcinoma: case report and review of the literature. J. Dermatol. Surg. Oncol. 1990; 16(5): 468–470. Publisher Full Text\n\nGoldstein DJ, Barr RJ, Santa Cruz DJ: Microcystic adnexal carcinoma: a distinct clinicopathologic entity. Cancer. 1982; 50(3): 566–572. PubMed Abstract \| Publisher Full Text\n\nHollowell KL, Agle SC, Zervos EE, et al.: Cutaneous apocrine adenocarcinoma: defining epidemiology, outcomes, and optimal therapy for a rare neoplasm. J. Surg. Oncol. 2012; 105(4): 415–419. PubMed Abstract \| Publisher Full Text\n\nChamberlain RS, Huber K, White JC, et al.: Apocrine gland carcinoma of the axilla: review of the literature and recommendations for treatment. Am. J. Clin. Oncol. 1999; 22(2): 131–135. Publisher Full Text\n\nMitts DL, Smith MT, Russell L, et al.: Sweat gland carcinoma: a clinico-pathological reappraisal. J. Surg. Oncol. 1976; 8(1): 23–29. PubMed Abstract \| Publisher Full Text\n\nMiyamoto T, Hagari Y, Inoue S, et al.: Axillary apocrine carcinoma with benign apocrine tumours: a case report involving a pathological and immunohistochemical study and review of the literature. J. Clin. Pathol. 2005; 58(7): 757–761. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "192585", "date": "24 Aug 2023", "name": "Rehan Zahid", "expertise": [ "Reviewer Expertise Plastic and Reconstructive Surgery" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nI appreciate the authors for presenting this rare carcinoma. The authors have duly noted the lack of literature of such rare cases which makes it even more important to provide as much detail as possible when presenting this case.\nI suggest starting with elaborating the presentation. Progression of disease including a timeline and more details of the history and physical findings. Rephrase the first sentence of the case report section.\nEmphasize on the treatment plan. Was a biopsy done to confirm the diagnosis before the final excision? If it was planned as an excisional biopsy before being identified as a carcinoma then was a wide local excision carried out? Or planned? Especially with a positive inferior margin. Please clarify the surgical treatment that the patient underwent including lymph node dissection.\n\nWhat were the surgical sequelae post excision? Was the defect closed primarily and was there any post surgical complication including wound healing delays or ulcerative margins? Although the patient was lost to follow up, it would be beneficial to provide as much data as available in terms of the outcome. It helps to identify the progression of disease and overall prognosis.\n\nWas any other work up planned to check for metastatic disease?\n\nFinally the authors should reflect on the findings of this case report and how it contributes to the existing literature on apocrine carcinoma. I feel there is more information that could be provided to contribute to the already scarce existing literature. Although it may not be practically feasible in many instances but all efforts should be made to pursue the patient for follow up and determine outcome of the disease.\n\nIs the background of the case’s history and progression described in sufficient detail? Partly\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? No\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Partly\n\nIs the case presented with sufficient detail to be useful for other practitioners? Partly", "responses": [ { "c_id": "10269", "date": "06 Oct 2023", "name": "Dr. Suhit Naseri", "role": "Author Response", "response": "1. Progression of disease including a timeline and more details of the history and physical findings. In 2022, a male aged 48, hailing from South Asian origins and employed as a carpenter, visited the surgery department due to the presence of a lump (Figure 1) in the axilla. The patient initially discovered a painless, indurated nodule in their right axilla around a year back. Over time, this nodule increased in size until the time of their presentation that is the size of 8 x 8 x 2.3 cm. The patient attempted ayurvedic treatment for a year, but the lesion continued to progress. On local examination, signs included inflammation of the skin covering the area, occasional discharge of a serosanguinous nature, and mild pain in the affected axilla. The patient did not experience any constitutional symptoms such as fever, weight loss, night sweats, or loss of appetite. The patient had no family history of malignancy. A physical evaluation and USG for both breasts did not find any abnormalities. USG of the right axilla suggested a well-defined solid isoechoic lesion with multiple microcalcifications with prominent vascularity. Additionally, the patient had a preexisting condition of hypertension and was taking antihypertensive medications. Patient revealed his past habit of smoking bidi once a day. He did not consume alcohol. 2. Emphasize on the treatment plan. Was a biopsy done to confirm the diagnosis before the final excision? If it was planned as an excisional biopsy before being identified as a carcinoma then was a wide local excision carried out? Or planned? Especially with a positive inferior margin. Please clarify the surgical treatment that the patient underwent including lymph node dissection. A Tru Cut Biopsy was carried out which revealed cells showing moderate amount of eosinophillic to pale vaccuolated cytoplasm, Eccentric nuclei showing mild hyperchromasia and significant mitotic activity. Another population of cells seen with squamoid feature intervening the stroma showing desmoplastic reaction. Biopsy findings were suggestive of Malignancy of Adnexal origin. Patient underwent wide local excision of the right axillary lesion with right axillary lymphadenectomy up to level III was carried out. 3. What were the surgical sequelae post excision? The patient's recuperation after surgery proceeded without any complications.Was the defect closed primarily and was there any post surgical complication including wound healing delays or ulcerative margins? A sufficient clearance margin of 1-1.5 cm was taken, and primary closure of defect was achieved. The patient's recuperation after surgery proceeded without any complications. 4. Was any other work up planned to check for metastatic disease? Considering the metastatic involvement in the lymph nodes and to exclude the possibility of distant metastasis, a comprehensive whole-body PET-CT scan was performed, revealing no signs of distant metastatic spread." } ] } ]	1	https://f1000research.com/articles/12-803
https://f1000research.com/articles/12-601/v1	05 Jun 23	{ "type": "Correspondence", "title": "Historical changes in baby names in China", "authors": [ "Yuji Ogihara" ], "abstract": "Based on previous research on names and naming practices, I propose three suggestions to Bao et al. (2021), which investigated historical changes in given names of Han Chinese in China between 1920 and 2005. Their study analyzed a one-shot cross-sectional survey conducted in 2005 and reported that unique names increased from 1920 to 2005. The authors concluded that China became more individualistic over time for the period. However, three questions have remained unanswered in Bao et al. (2021). First, were the samples of older birth cohorts truly representative? Second, did unique names increase only after the 1970s? Third, how are the historical changes in average name length interpreted? Answering these three questions would contribute to a further understanding of the historical changes in given names and their underlying psychological/cultural shifts in China.", "keywords": [ "name", "uniqueness", "historical change", "cultural change", "individualism", "China", "need for uniqueness", "culture" ], "content": "1. Were the samples of older birth cohorts truly representative?\n\nThe authors used a random subset of a one-shot cross-sectional survey conducted in 2005 (the 2005 China’s 1% Population Census) and analyzed given names of people born between 1920 and 2005. They emphasized that the sample is representative (e.g., “Using a large representative sample of Chinese names” in Abstract, “We used an unprecedentedly large representative sample of Chinese names, covering a longer period of time from 1920 to 2005” (p. 4) in Discussion, “To obtain a nationally representative sample of Chinese names covering a long period” (p. 2) in Method).\n\nHowever, the data is from a one-shot cross-sectional survey, not a cross-temporal survey (e.g., birth records). The authors investigated names of Chinese people aged from 0 (newborns) to 85 years. This indicates a possibility that the samples for some populations, especially older birth cohorts, may not be nationally representative (not including all the names given in a year in China). Considering that the average life expectancy in China in 2005 was approximately 73 years (72.99; United Nations, 2022), especially the data for older people would be systematically selected by death, yielding the selection effect. For example, economically wealthy people would live longer (despite diseases and aging, e.g., Wilkinson & Marmot, 2003; Jagger et al., 2008), and physically healthy people would be better suited to survive natural disasters at a higher rate, leading to the possibility that economically not wealthy and physically not healthy older people were underrepresented in the samples. In other words, although a subset of the 2005 China’s 1% Population Census would represent people who lived in 2005, older birth cohorts would not be representative, implying that the results for older years might not reflect the reality. To avoid these systematic biases, previous research examining historical changes in baby names analyzed cross-temporal data. Prior research in China (e.g., Cai et al., 2018), Japan (e.g., Ogihara, 2021a, 2022a; Ogihara et al., 2015; Ogihara & Ito, 2022), the United States (e.g., Ogihara, 2021d; Twenge et al., 2010, 2016), the United Kingdom (e.g., Bush, 2020; Bush et al., 2018), Germany (e.g., Gerhards & Hackenbroch, 2000), and France (e.g., Mignot, 2022) has used a series of yearly cross-temporal data of newborn baby names.\n\nIt would be necessary for the authors to clarify how they overcame these possible biases. The authors already stated that “because the sample sizes for birth years < 1920 were not sufficient, we limited the range of birth years to 1920~2005” (p. 2), but this issue is related to not only sample size but also sample characteristics (selection bias). The sample sizes for the earlier periods between 1920 and 2005 would not be sufficient to claim that the samples are representative, and the samples would be systematically selected and biased. Cross-sectional data should be carefully investigated to discuss cross-temporal changes (e.g., Cai et al., 2018; Ogihara, 2022b; Ogihara & Kusumi, 2020; Twenge, 2011; Twenge & Campbell, 2001).1\n\n\n2. Did unique names increase only after the 1970s?\n\nThe authors concluded that unique names increased in China between 1920 and 2005 and claimed that they replicated their previous study, which insists on an increase in unique names between 1950 and 2009 (Cai et al., 2018).\n\nHowever, all six indicators the authors analyzed consistently showed that the unique names did not increase from 1920 to 1969. Rather, the indicator of name-character uniqueness, which the authors “preferred” (p. 6) most and stated “the estimation would be more accurate” (p. 6) among all six indicators, shows a gradual decrease in uniqueness from 1920 to 1969 (Figure 2B in Bao et al., 2021). These results were inconsistent with their previous finding that insists on a continuous increase in unique names from 1950 to 2009 (Cai et al., 2018). The authors did not mention this point clearly.2 The study would be improved if the authors made efforts to explain why unique names did not increase between 1920 and 1969 and why the study did not replicate the previous finding.\n\nOne possible reason is the above-mentioned plausible biases in the samples. As I explained above, the samples in the older birth cohorts would likely include a higher proportion of more economically wealthy people. Previous research has demonstrated that people of high economic status tend to express more uniqueness (e.g., Ma et al., 2017; Snibbe & Markus, 2005; Stephens et al., 2007; Wang et al., 2020). Thus, the values of the uniqueness indicators in the older birth cohorts would be higher than the actual values and should be lower in reality. If this is true, an increase in unique names would be observed from 1920 to 1969 as well as from 1970 to 2005, showing that unique names would continue to increase from 1920 to 2005.\n\n\n3. How are the historical changes in average name length interpreted?\n\nThe historical changes in average name length (described in Figure 2F in Bao et al., 2021) were newly added to a previous study (Cai et al., 2018). They showed a different pattern of changes from those of character-based indices and seem to be divided into three periods: 1) 1920-1960: almost stable (maintained), 2) 1961-1990: sharp decrease, and 3) 1991-2005: sharp increase (Table 1).3\n\nHowever, the authors did not explain these changes and possible interpretations sufficiently. These drastic changes might be related to various changes in official rules regarding names, political policies, and so on (e.g., Ogihara, 2020). These changes in social, economic, and political aspects should also be considered when cultural changes are discussed.\n\nThe analysis shows that given names of Han Chinese in China typically consisted of two Chinese characters at least between 1920 and 2005 (Figure 1 in Bao et al., 2021). From 1920 to 1960, the proportions of one-character and three-character names did not change extensively, leading to the stability of the average name length. From 1961 to 1990, the proportion of one-character names remarkably increased (from approximately 10% to over 30%), but the proportion of three-character names did not vary, which decreased the average name length. It would be beneficial to investigate why only the proportion of one-character names remarkably increased during this period. From 1991 to 2005, the proportion of three-character names increased and the proportion of one-character names decreased, causing the increase in the average name length of this period. It would also be important to examine why the proportion of three-character names increased but the proportion of one-character names decreased.\n\n\nConclusion\n\nI propose three suggestions that would further increase the validity and impact of the article (Bao et al., 2021). First, it would be better to answer whether the samples of older birth cohorts were truly representative. Second, it would be preferrable to answer whether unique names increased only after the 1970s. Third, it should be clarified how the historical changes in average name length are interpreted. These suggestions would hopefully contribute to a further understanding of the historical changes in baby names and their underlying psychological/cultural shifts in China.\n\n\nEthics statement\n\nNot applicable.", "appendix": "Data availability\n\nNo data is associated with this article.\n\n\nReferences\n\nBao HWS, Cai H, Jing Y, et al.: Novel evidence for the increasing prevalence of unique names in China: A reply to Ogihara. Front. Psychol. 2021; 12: 731244. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBush SJ: Ambivalence, avoidance, and appeal: Alliterative aspects of Anglo anthroponyms. Names. 2020; 68: 141–155. Publisher Full Text\n\nBush SJ, Powell-Smith A, Freeman TC: Network analysis of the social and demographic influences on name choice within the UK (1838-2016). PLoS One. 2018; 13: e0205759. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCai H, Zou X, Feng Y, et al.: Increasing need for uniqueness in contemporary China: Empirical evidence. 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PubMed Abstract \| Publisher Full Text\n\nTwenge JM: The duality of individualism: Attitudes toward women, generation me, and the method of cross-temporal meta-analysis. Psychol. Women Q. 2011; 35: 193–196. Publisher Full Text\n\nTwenge JM, Abebe EM, Campbell WK: Fitting in or standing out: Trends in American parents’ choices for children’s names, 1880–2007. Soc. Psychol. Personal. Sci. 2010; 1: 19–25. Publisher Full Text\n\nTwenge JM, Campbell WK: Age and birth cohort differences in self-esteem: A cross-temporal meta-analysis. Personal. Soc. Psychol. Rev. 2001; 5: 321–344. Publisher Full Text\n\nTwenge JM, Dawson L, Campbell WK: Still standing out: Children’s names in the United States during the Great Recession and correlations with economic indicators. J. Appl. Soc. Psychol. 2016; 46: 663–670. Publisher Full Text\n\nUnited Nations: World Population Prospects 2022.2022. Reference Source\n\nWang X, Chen Z, Krumhuber EG: Money: An integrated review and synthesis from a psychological perspective. Rev. Gen. Psychol. 2020; 24: 172–190. Publisher Full Text\n\nWilkinson RG, Marmot M: Social determinants of health: the solid facts. World Health Organization; 2003.\n\n\nFootnotes\n\n1 Furthermore, because some people changed their given names, their names in 2005 are different from the names given at births. This possibility should also be considered when cross-sectional data are used to examine cross-temporal changes in names.\n\n2 Cai et al. (2018) also showed this possibility (unique names increased only after 1970). The average name character frequency per birth cohort of 1970-1979 was higher than that of 1960-1969 (Figure 2 in Cai et al., 2018). However, this might be solely due to small sample sizes (Ogihara, 2020).\n\n3 However, the possible bias in the samples, which I discuss before, should be considered here, too." }	[ { "id": "196699", "date": "18 Aug 2023", "name": "Gabriela Fatková", "expertise": [ "Reviewer Expertise Anthropology of names and naming", "gender", "kinship", "food", "memory" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe article discusses and criticizes a study (Bao et al. 2021) on historical changes in given names in China from 1920 to 2005. The author presents three key points of contention. Firstly, it questions whether the samples for older birth cohorts are truly representative due to potential selection biases. The cross-sectional nature of the data and the unsatisfactorily settled biases stemming from it could lead to inaccuracies in the study's conclusions. Secondly, it challenges the study´s claim of an increase in unique names only after the 1970s, pointing out that in the results presented it did not apply to all indicators. Lastly, the author aptly notes that the interpretation and introduction of broad social context (historical, societal, and policy changes) of any quantitative analysis is needed. The suggestions are reasonable and constructive.\nThe author recommends a deeper exploration of societal and policy transformations that might underpin the observed fluctuations. In addition to the last point, the broader reflection on the evolving functions of names within Chinese society would be very useful. Names are not mere symbols but integral to a dynamic social process, shaping identities and forging connections among individuals and groups on a daily basis. Name studies approaching names solely as indicators of single social processes, such as individualization in the case of Bao et al. (2021), is deemed shallow and insufficient. Consequently, the author´s critique proposes comprehensive considerations to enhance the validity and impact of the research, encompassing a nuanced examination of the multifaceted roles names play in the fabric of Chinese society.\n\nIs the rationale for commenting on the previous publication clearly described? Yes\n\nAre any opinions stated well-argued, clear and cogent? Yes\n\nAre arguments sufficiently supported by evidence from the published literature or by new data and results? Yes\n\nIs the conclusion balanced and justified on the basis of the presented arguments? Yes", "responses": [] }, { "id": "191660", "date": "22 Aug 2023", "name": "Shintaro Fukushima", "expertise": [ "Reviewer Expertise social psychology", "social survry", "culural psycholgy" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe claims of the author (Dr. Ogihara) are quite reasonable and convince me in general.\nHowever, the concept of uniqueness and some insistences are confusing or inadequately supported by the references.\nThe author insists on a selection bias on a sampled elderly; living elderly at the time when the national survey was conducted would be wealthier (and healthier) compared with the elderly who had already passed away, which results in more uniqueness of the names of the sampled living elderly.\nAlthough the fact that this systematic selection bias is stronger in the elderly is critical for the intergenerational comparison, the three variables A) unique names of the participants, B) uniqueness of the participants, and C) uniqueness of the parents of the participants seem to be used confusingly in the claims by the author.\nCan A) unique names of the participants be identified as B) psychological uniqueness of the participants? The author says that previous research has demonstrated that wealthy people tend to express more uniqueness, but does it really mean that wealthy people tend to have more unique names? As for the causal direction, money could get people to have freedom and uniqueness as demonstrated in Ma et al. (2017) which is cited in the present paper, but could unique-name also get people to be wealthy? In fact, some studies indicate that unique names limit future employment or social prospects (e.g., Kalist & Lee, 2009; Rogers-Anderson, 2017; Savage & Wells, 1948). In addition, the negative consequence of having unique names might relatively be seen in the interdependent countries such as China where people are expected to have social harmony or rejection avoidance rather than in the independent countries such as the U.S. where people are expected to have independence or uniqueness.\nFurthermore, A) unique names of the participants would be directly affected by C) uniqueness of the parents of the participants rather than by B) uniqueness of the participants themselves. If so, the selection bias the author pointed out might not be as critical as expected.\nOverall, I recommend the author to add empirical evidence which shows that having unique names results in low life expectancy directly or indirectly. In that case, individual-level (within culture) associations rather than social/group-level (between-culture) associations are preferable to be shown because between-culture association includes the mixed effects of A) unique names of the participants, B) uniqueness of the participants, and C) uniqueness of the parents of the participants, which would fail to explain the selection bias of the sampled individuals within China.\n\nIs the rationale for commenting on the previous publication clearly described? Yes\n\nAre any opinions stated well-argued, clear and cogent? Yes\n\nAre arguments sufficiently supported by evidence from the published literature or by new data and results? Partly\n\nIs the conclusion balanced and justified on the basis of the presented arguments? Yes", "responses": [ { "c_id": "10503", "date": "08 Nov 2023", "name": "Yuji Ogihara", "role": "Author Response", "response": "November 2nd, 2023 Dear Dr. Shintaro Fukushima, Thank you very much for reviewing my manuscript and providing a valuable comment. I have modified the manuscript according to your comment. I offer my response to the comment below. I have copied and pasted your comment without making changes. The claims of the author (Dr. Ogihara) are quite reasonable and convince me in general. However, the concept of uniqueness and some insistences are confusing or inadequately supported by the references. The author insists on a selection bias on a sampled elderly; living elderly at the time when the national survey was conducted would be wealthier (and healthier) compared with the elderly who had already passed away, which results in more uniqueness of the names of the sampled living elderly. Although the fact that this systematic selection bias is stronger in the elderly is critical for the intergenerational comparison, the three variables A) unique names of the participants, B) uniqueness of the participants, and C) uniqueness of the parents of the participants seem to be used confusingly in the claims by the author. Can A) unique names of the participants be identified as B) psychological uniqueness of the participants? The author says that previous research has demonstrated that wealthy people tend to express more uniqueness, but does it really mean that wealthy people tend to have more unique names? As for the causal direction, money could get people to have freedom and uniqueness as demonstrated in Ma et al. (2017) which is cited in the present paper, but could unique-name also get people to be wealthy? In fact, some studies indicate that unique names limit future employment or social prospects (e.g., Kalist & Lee, 2009; Rogers-Anderson, 2017; Savage & Wells, 1948). In addition, the negative consequence of having unique names might relatively be seen in the interdependent countries such as China where people are expected to have social harmony or rejection avoidance rather than in the independent countries such as the U.S. where people are expected to have independence or uniqueness. Furthermore, A) unique names of the participants would be directly affected by C) uniqueness of the parents of the participants rather than by B) uniqueness of the participants themselves. If so, the selection bias the author pointed out might not be as critical as expected. Overall, I recommend the author to add empirical evidence which shows that having unique names results in low life expectancy directly or indirectly. In that case, individual-level (within culture) associations rather than social/group-level (between-culture) associations are preferable to be shown because between-culture association includes the mixed effects of A) unique names of the participants, B) uniqueness of the participants, and C) uniqueness of the parents of the participants, which would fail to explain the selection bias of the sampled individuals within China. Thank you for your important comment. My explanation in the previous version of the article was insufficient. Thus, I have added an explanation as below. “Previous research has demonstrated that people of high economic status tend to express more uniqueness (e.g., Ma et al., 2017; Snibbe & Markus, 2005; Stephens et al., 2007; Wang et al., 2020). This leads that they tend to receive more unique names from their parents who are also more likely to be in high economic status. Thus, the values of the uniqueness indicators in the older birth cohorts would be higher than the actual values and should be lower in reality.” I provide one possible interpretation of the results, as I wrote in the previous version of the article. The important point is that all six indicators the authors analyzed consistently showed that the unique names did NOT increase from 1920 to 1969. Thus, I recommend that the authors of the target article examine this possibility. Thank you for your further consideration of this manuscript. I look forward to hearing from you at your earliest convenience. Sincerely, Yuji Ogihara, Ph.D. Department of Psychology, College of Education, Psychology and Human Studies, Aoyama Gakuin University Address: 4-4-25 Shibuya, Shibuya-ku, Tokyo, 150-8366, Japan E-mail: yogihara@ephs.aoyama.ac.jp Web: https://sites.google.com/site/yujiogiharaweb/english" } ] } ]	1	https://f1000research.com/articles/12-601
https://f1000research.com/articles/12-1445/v1	07 Nov 23	{ "type": "Systematic Review", "title": "Gender gaps in scientific research: a systematic review", "authors": [ "Rosario Violeta Grijalva Salazar", "Víctor Hugo Fernández-Bedoya", "Mónica Elisa Meneses La Riva", "Josefina Amanda Suyo Vega", "Martín Alexander Ríos Cubas", "Víctor Hugo Fernández-Bedoya", "Mónica Elisa Meneses La Riva", "Josefina Amanda Suyo Vega", "Martín Alexander Ríos Cubas" ], "abstract": "Background: The gender gaps present in the field of scientific and academic research generate discrimination and lack of equal opportunities for women, resulting in several barriers that significantly limit women's scientific productivity. The objective was to identify the main gender gaps in productivity and scientific research.\nMethod: The researchers conducted a systematic search for articles on gender disparities in women's scientific production in the SCOPUS and REDALYC repositories, taking into account manuscripts in English, Spanish, and Portuguese. Articles on gender gaps in scientific production were included, while empirical studies with other approaches to gender discrimination were excluded. Studies that did not address gender differences in scientific research, those that focused only on specific scientific disciplines without taking gender into account, and those that were not available in their entirety were excluded. The search and selection were conducted from May and June 2023. To avoid stumbling blocks, other methods were used, such as initially filtering titles based on a search equation and then excluding those that did not address gender differences in scientific research. Next, manuscripts were reviewed and those that did not meet the inclusion criteria were excluded. Finally, the remaining research was thoroughly reviewed to obtain the information needed for the study.\nResults: A total of 23 articles were analyzed, addressing various issues such as discrimination, lack of policies to support women, academic inequalities and other factors that make female participation more difficult.\nConclusion: The main findings revealed gender gaps that have an impact on worldwide female scientific production. The literature frequently focuses on low output without investigating the causes. When they approach, they just treat the surface. Future research should focus on gender disparities in production, as well as the daily challenges women face in research and scientific production.", "keywords": [ "Gender inequality", "Gender discrimination", "Female participation", "Equitable representation", "Equal opportunities." ], "content": "Introduction\n\nScientific research has long been the cornerstone of knowledge advancement and innovation in our society, contributing to improving the quality of life for the population and training professionals geared towards research (Delgado, 2021).\n\nAccording to Houssay (1960), the importance of scientific research lies in the fact that health, well-being, wealth, power, and independence of nations depend on it. Scientific research allows us to better understand biological processes and diseases, leading to advances in medicine and more effective treatments. Additionally, scientific research drives technological development and innovation, which in turn generates economic wealth and power for nations that invest in it.\n\nGender equity in scientific research is a topic of growing concern and debate worldwide. Despite significant progress in promoting gender equality in various spheres, notable gaps persist in the participation and recognition of women in scientific research.\n\nAccording to the European University Association (EUA), in universities across 48 European countries, only 15% of rectors were women, while the remaining 85% were men (EUA, 2020). Furthermore, it was highlighted that in 20 European countries, there were no women in the position of rector. Additionally, according to a report by the International Institute for Higher Education in Latin America and the Caribbean (IESALC) belonging to the United Nations Educational, Scientific and Cultural Organization (UNESCO), in 2020, only 18% of public universities in Latin America had women serving as rectors. This same report indicates that only 30% of university researchers worldwide were women, pointing to a significant gender gap in the field of research (IESALC, 2020, 2021).\n\nDespite the increase in the number of women entering universities, many choose to leave at higher levels, which are often necessary for a career in research (IESALC, 2020). This, in turn, leads to a continuing gap in terms of production and authorship of scientific publications. A report by Elsevier (2020) indicates that despite an overall increase in the representation of women in research, persistent inequality still exists. On average, female researchers have fewer publications (38%) than their male counterparts in all countries (68%).\n\nGender disparities in various aspects are profound, and the numbers alone are not sufficient to reflect the extent of the challenges women face in educational and professional settings. These women continue to report being in contexts that privilege the male gender, facing barriers such as gender-based wage disparities, as well as the threat and reality of harassment and sexual violence on campuses (UNESCO, 2021). There is also faster promotion of male gender, stricter review processes for women's research manuscripts, and the time constraints faced by female assistant professors with children, making their records less competitive (Chen and Crown, 2019).\n\nAs can be observed, numerous studies have revealed concerning patterns of inequality, from the underrepresentation of women in academic and scientific leadership positions to the disparity in funding and recognition opportunities. The gender gap in scientific research has profound implications. By excluding women from scientific decision-making, valuable perspectives and viewpoints are lost, and the diversity of ideas necessary for innovation is limited, resulting in a loss of talent and scientific potential.\n\nConsidering the commitment to gender equality and the empowerment of all women and girls, which is part of Sustainable Development Goal 5 (SDG), and the elimination of gender disparities in education mentioned in SDG 4 (United Nations, 2020), this research proposes a systematic review to analyze the evidence regarding gender gaps in scientific research. By identifying key factors contributing to this inequality, it seeks to contribute and provide a foundation for future research and actions aimed at closing the gender gaps in scientific research.\n\n\nMethods\n\nFor the development of this research, a search for scientific production associated with the topic of gender gaps in scientific production was conducted through a systematic review of previous studies. This method aims to identify, interpret, and evaluate different works carried out by scholars on a specific topic or field, based on the text (Pardal and Ochoa, 2017), with the intention of “providing the researcher and reader with clarifying information on a specific subject” (Pardal and Pardal, 2020, p.1). Based on this, the research question for the review was formulated as follows: What are the main gender gaps in productivity and scientific research addressed in research articles?\n\nIn this study, an exhaustive systematic review of scientific literature related to gender gaps in scientific research has been conducted. To carry out this review, the guidelines established in the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA, 2020) statement were followed. These guidelines are specifically designed to ensure the rigorous conduct of systematic reviews.\n\nThe first step in the systematic review process was to conduct initial searches in May 2023 in the Scielo, Redalyc, and Scopus databases. These searches yielded 151 results, providing a global overview of the breadth of the topic and confirming its significance. Initial search criteria included studies that specifically addressed gender differences in scientific research in general, studies that focused only on specific scientific disciplines without addressing the issue of gender gaps or differentiation were not included, and manuscripts that were not accessible in full text were also excluded.\n\nDue to the scarcity of results from Scielo and the absence of any studies that were not already included in the other two databases, it was decided to exclude Scielo from the systematic search. The final repositories for data collection were Redalyc and Scopus.\n\nThe search was conducted again in May 2023, using Redalyc and Scopus, resulting in a total of 147 papers. The search was limited to publications from 2009 to the present.\n\nThe combination of terms that yielded the best results in both databases was as follows: (“gender gap in scientific production”; “gender gap” AND “scientific production”) or the search strategy was adapted based on the specific requirements of each database.\n\nSpecifically, 147 results were obtained in Redalyc and Scopus. Before proceeding with the article selection, inclusion and exclusion criteria were defined.\n\nThe inclusion criteria were as follows:\n\n• Empirical studies and systematic reviews addressing gender gaps in scientific research.\n\n• Studies published in Spanish, English, and Portuguese.\n\n• Studies investigating topics related to gender discrimination, women's representation in science, obstacles and challenges faced by women in scientific research, policies and practices to address gender gaps, and other relevant aspects.\n\n• Studies published from 2000 to the cutoff date of our review (June 2023).\n\nThe exclusion criteria were as follows:\n\n• Studies that did not specifically address gender gaps in scientific research.\n\n• Studies solely focused on specific scientific disciplines without addressing the gender issue.\n\n• Studies that were not accessible in full text.\n\n• Study selection process\n\nThe selection of studies was conducted in three stages: an initial stage based on reading titles, a second stage based on reading abstracts, and a third stage based on a thorough review of the articles selected in the first stage. The selection was independently carried out by five researchers, and any discrepancies were resolved through consensus.\n\nFinally, 23 articles met the inclusion criteria and were selected for the systematic review.\n\nThe process for determining if an article met the criteria began with a review of the titles of each research project; this is where it was determined whether the article met the theme: Generic break. Following the first filter, all results were read and evaluated to see if they were inside the search variable. Finally, the whole contents of the manuscript were reviewed to complete the article. The filtering was done by three of the authors, and the examination of each registered and recovered document was done by each author. It is important to note that the distribution was done equitably based on the number of articles among the number of authors. Each paper was reviewed and analyzed individually, but at the end, all writers collaborated to create a final compilation.\n\nEach author thoroughly examined the content of each manuscript, meticulously assessing the variable ‘generic gap in scientific production’. The methodology used, the data collected, and the preliminary interpretations were all taken into account. Individual analysis allowed for detailed exploration of each manuscript, always leaving a variety of perspectives and experience in data interpretation.\n\nTo guarantee consistency and integration of the various studies, a collaborative review phase was carried out, during which the various methodologies and findings acquired were collated and compared. Constructive conversations and debates were encouraged to settle disagreements and ensure that the final interpretation was sound and well-supported. The procedure concluded in the formation of a final compilation containing the data and conclusions obtained by consensus from the different analyses.\n\n\nResults\n\nA total of 23 articles were analyzed, all of which met the inclusion criteria (Table 1). It should be noted that all studies discussing gender gaps in scientific production among women were included. All works addressing the topic were considered, regardless of the methodology used, population, or instrument.\n\nThe research and manuscript search (Table 1) revealed several gender gaps in science and academia that affect women's participation and advancement (Table 2). One of the main gaps found is the underrepresentation of women in high-level positions in scientific research, resulting in a lack of representation in scientific publications. This situation reflects the existence of invisible barriers and entrenched social norms that limit the progress of women in academia and science (Segovia et al., 2020; Cabrera and Saraiva, 2021; Boté et al., 2022).\n\nFurthermore, discrimination is observed in the National System of Researchers, where women have lower representation in higher levels and face difficulties in being accepted into the system. These findings suggest the presence of discrimination in the evaluation process (Cabrera and Saraiva, 2021; Carbonell et al., 2023).\n\nRegarding academic trajectories, differences between men and women are evident in terms of graduation time, training, and specialization development. Women face obstacles and ‘losses’ in their academic trajectory that men do not experience in the same way, contributing to inequality of opportunities (Aliaño et al., 2020; González and Mayo, 2021).\n\nInequality in recognition and rewards is also a significant gap. Although women may achieve individual merits earlier in their careers, men tend to be more rewarded and in less time for merits that rely on collective recognition. This inequality is attributed to institutional discrimination and entrenched cultural values (Carrillo and Flores, 2023; Frigi and Ávila, 2021; Vargas et al., 2016).\n\nAnother identified gap is professional and task pigeonholing. There is occupational gender segregation in certain fields, where the nursing field is heavily feminized, and the mathematics field is masculinized. This segregation contributes to limiting options and opportunities for women in choosing their careers (Osorio and Sokil, 2022).\n\nIn the field of scientific publication, there are barriers that affect women's participation. Women need to associate more with other women to publish, while men require less collaboration (Osorio and Sokil, 2022; Centeno et al., 2020). This reflects inequalities in co-authorship networks and opportunities to participate in publications (Lis and Bahos, 2016; Beigel and Gallardo, 2021).\n\nThe tensions and challenges associated with discrimination, lack of female academic and professional role models, integration into a male-dominated culture, and lack of interest and intrinsic motivation also contribute to gender gaps. These factors can lead to the exclusion of women in the information economy and hinder their access to scientific and technological fields (Palencia and Jiménez, 2016).\n\nFurthermore, domestic workload is identified as a factor that limits women's participation in academia and science. The traditional assignment of roles and responsibilities in society hinders women's access to positions of power, prestige, and responsibility.\n\nIn terms of academic opportunities, although women outnumber men in the demand for university education, there are still gaps in access to master's and doctoral degrees. This limits women's participation in higher levels of education and research (Carrasco, 2022; González and Mayo, 2021).\n\nWomen also face inequalities in their participation as first authors, corresponding authors, and conference speakers in the scientific field. This is reflected in less frequent citations, lower presence in specialized journals, and limited representation in scientific conferences (Tornero et al., 2020; López et al., 2021).\n\nFurthermore, the unequal presence of men in the evaluation and review systems of scientific journals is highlighted, which can influence gender gaps in scientific publication (Aquino et al., 2022). Women have limited representation in leadership roles and in higher-level decision-making instances in institutes, laboratories, and research teams (Betlloch, 2019; Giner et al., 2021).\n\nTo address these gender gaps, the importance of implementing policies to support the integration of women in higher education is emphasized. These policies should include the collection of gender-disaggregated data, collaboration between men and women, and strengthening the integration of women in academia and science (Carrillo and Flores, 2023; Cruz, 2021; Maldona et al., 2015).\n\nIt is critical to address the risk of failure due to insufficient results in each thesis and evaluation due to the nature of gender gaps. Given that gender disparities are frequently associated with underrepresentation and a lack of visibility for women in research, it is possible that some pertinent results were not recorded or published, which might cloud the overall picture of the situation. This selective data omission may obscure even further the full dimensions of gender gaps, leading to inexact or incomplete conclusions. As a result, it is critical to consider this potential source of data while analyzing and synthesizing available data.\n\nGiven the complexity of gender gaps and their impact on various aspects of scientific research, it is essential to be able to understand that the quality of the available evidence on the various gender gaps within research was clearly and systematically assessed.\n\n\nDiscussion\n\nThe research reveals various gender gaps in the scientific and academic field that hinder the participation and advancement of women. Multiple areas have been identified where these gaps are evident, calling for urgent attention to achieve gender equality and promote greater female representation in science and academia.\n\nThe underrepresentation of women in high-level positions in scientific research and the lack of representation in scientific publications reflect invisible barriers and entrenched social norms that limit their progress in academia and science (Segovia et al., 2020; Cabrera and Saraiva, 2021; Boté et al., 2022). This highlights the need to address these barriers and promote greater female participation and representation.\n\nDiscrimination in the National System of Researchers, where women face difficulties in being accepted into the system and have lower representation at higher levels, suggests the presence of discrimination in the evaluation process (Cabrera and Saraiva, 2021; Carbonell et al., 2023). It is essential to work towards eliminating biases and promoting fair and unbiased evaluation at all levels.\n\nDifferences in academic trajectories between men and women, including time to graduation, training, and specialization development, contribute to the inequality of opportunities (Aliaño et al., 2020; González and Mayo, 2021). These differences indicate the need to implement measures that support and promote equal opportunities for all individuals, regardless of gender.\n\nInequality in recognition and rewards is another significant gap, where it is observed that although women may achieve individual merits earlier in their careers, men tend to be rewarded to a greater extent and in less time for merits that depend on collective recognition (Carrillo and Flores, 2023; Frigi and Ávila, 2021; Vargas et al., 2016). This inequality reflects the existence of institutional discrimination and entrenched cultural values, emphasizing the need to promote equitable evaluation and recognition.\n\nProfessional and occupational segregation, where certain fields are feminized or masculinized, such as nursing and mathematics respectively, limit women's choices and opportunities in career selection (Osorio and Sokil, 2022). It is essential to promote greater diversity and eliminate gender stereotypes in all professional fields.\n\nRegarding barriers in scientific publishing, it is observed that women need to collaborate more with other women to publish, while men require less collaboration (Osorio and Sokil, 2022; Centeno et al., 2020). This reflects inequalities in co-authorship networks and opportunities to participate in publications, emphasizing the importance of fostering collaboration and support among women in the scientific field.\n\nThe tensions and challenges associated with discrimination, the lack of female academic and professional role models, integration into a male-dominated culture, and a lack of intrinsic interest and motivation contribute to gender gaps (Palencia and Jiménez, 2016). These factors need to be addressed through programs and policies that promote gender equality and encourage women's interest and participation in science.\n\nThe unequal presence of men in the evaluation and peer review systems of scientific journals, as well as in leadership roles in institutes, laboratories, and research teams, highlights the need to promote greater female representation in these areas (Betlloch, 2019; Giner et al., 2021). This involves implementing measures that foster gender equity and active participation of women in decision-making processes.\n\nLimited national incentive policies also need to be addressed. It is essential to develop policies that promote gender equality in the field of science and academia, including the collection of gender-disaggregated data, collaboration between men and women, and strengthening the integration of women in higher education (Carrillo and Flores, 2023; Cruz, 2021; Maldona et al., 2015). These policies are crucial to address gender gaps and promote greater participation and representation of women in science and academia.\n\nFor future research on this topic, it is essential not only to investigate why there is a difference between male and female production but also to explore the limitations that women face daily in the field of research and article production.\n\n\nConclusion\n\nIn conclusion, the findings of this research highlight the existence of various gender gaps in the scientific and academic fields. These gaps, including underrepresentation in high-ranking positions and research, barriers in scientific publishing, limited national incentive policies, unequal academic opportunities, lower likelihood of being primary or corresponding authors and limited participation as conference speakers, presence of men in evaluation and peer review systems of scientific journals, professional and occupational segregation, and discrimination, require urgent attention. It is essential to implement policies and measures that promote gender equality, eliminate biases and barriers, and foster greater participation and representation of women in science and academia.", "appendix": "Data availability\n\nZenodo. Gender Gaps in Scientific Research: A Systematic Review. https://doi.org/10.5281/zenodo.8240031 (Grijalva et al., 2023).\n\nThis project includes the following underlying data:\n\n• LEAKED MANUSCRIPTS - TOPIC - GENDER GAP.xlsx. (23 articles with variables).\n\nZenodo. Gender Gaps in Scientific Research: A Systematic Review. https://doi.org/10.5281/zenodo.8240031 (Grijalva et al., 2023).\n\nThis project includes the following extended data:\n\n• Manuscript inclusion and exclusion flowchart according to the PRISMA 2020 method.pdf. ( Figure 1 flowchart included in the manuscript).\n\nRepository: PRISMA checklist for ‘Gender Gaps in Scientific Research: A Systematic Review’. https://doi.org/10.5281/zenodo.8240031 (Grijalva et al., 2023).\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferencias\n\nAliaño M, Franco G, Gilsanz F: Diferencias de género en Anestesiología. ¿En qué punto nos encontramos en España? Resultados de una encuesta nacional. Rev. Esp. Anestesiol. Reanim. 2020; 67(7): 374–380. PubMed Abstract \| Publisher Full Text Reference Source\n\nAquino C, Chávez S, Benites C: Participación femenina en los comités editoriales de revistas médicas en Latinoamérica. Biomedica. 2022; 42(2): 355–363. PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source\n\nBeigel F, Gallardo O: Productividad, bibliodiversidad y biolingüismo en un corpus completo de producciones científicas. Revista Iberoamericana de Ciencia, Tecnología y Sociedad - CTS. 2021; 16(46): 41–71. Reference Source\n\nBetlloch M: Políticas de igualdad en el proceso editorial. Importancia de incluir el nombre de pila en las publicaciones científicas. Actas Dermosifiliogr. 2019; 110(10): 791–793. PubMed Abstract \| Publisher Full Text Reference Source\n\nBoté J, Ferrer M, Gorchs M: Gender differences in peruvian nursing: A bibliometric analysis in scopus and web of science. Telos. 2022; 24(2): 302–328. Publisher Full Text Reference Source\n\nCabrera M, Saraiva I: Principales problemáticas de las publicaciones científicas: un análisis en perspectiva latinoamericana. Revista e-Ciencias de la Información. 2021; 12(1): 188–210. Publisher Full Text Reference Source\n\nCarbonell S, Villodre C, Baeza A, et al.: Brecha de género en las publicaciones de Cirugía Española. Cir. Esp. 2023; 101(6): 453–456. Publisher Full Text Reference Source\n\nCarrasco G: Situación de la mujer en la Ciencia y tecnología: relaciones de poder al interior de una entidad académica pública con autonomía universitaria. Trilogía Ciencia Tecnología Sociedad. 2022; 10(19): 45–58. Publisher Full Text Reference Source\n\nCarrillo P, Flores M: Mujeres científicas en Yucatán: obstáculos, retos y experiencias durante sus trayectorias educativas. Revista Latinoamericana de Estudios Educativos. 2023; 53(1): 253–284. Publisher Full Text Reference Source\n\nCenteno D, Morales L, Lopez C, et al.: Mujeres científicas: características y factores asociados a la primera autoría y corresponsalía en revistas peruanas indizadas a SciELO, 2010-2015. Educación Médica. 2020; 21(1): 17–23. Publisher Full Text Reference Source\n\nChen J, Crown D: The Gender Pay Gap in Academia: Evidence from The Ohio State University. Am. J. Agric. Econ. 2019; 101(5): 1337–1352. Publisher Full Text\n\nCruz L: Diferencias y sesgos de género en la financiación de la investigación: un enfoque dinámico. Gestión y Análisis de Políticas Públicas. 2021; 26: 6–19. Publisher Full Text Reference Source\n\nDelgado J: La investigación científica: su importancia en la formación de investigadores. Ciencia Latina Revista Científica Multidisciplinar. 2021; 5(3): 2385–2386. Reference Source\n\nElsevier: The Researcher Journey Through a Gender Lens: An Examination of Research Participation, Career Progression and Perceptions Across the Globe. [Archivo PDF].2020. Reference Source\n\nEUA: Male vs Female University Leaders: The Hard Facts on International Women’s Day.2020. Reference Source\n\nFrigi P, Ávila M: Mulheres gestoras em CT&I: estudo de caso nas áreas espacial e do ambiente terrestre. Revista Iberoamericana de Ciencia, Tecnología y Sociedad - CTS. 2021; 16(46): 247–266. Reference Source\n\nGiner M, López O, Zabaleta E, et al.: Análisis bibliométrico de la autoría femenina en artículos originales en la revista Atención primaria. Aten. Primaria. 2021; 53(1): 12–18. PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source\n\nGonzáles A: La carrera profesional de las investigadoras jóvenes: un camino lleno de posibilidades. Revista Iberoamericana de Ciencia, Tecnología y Sociedad - CTS. 2009; 4(12): 31–54. Reference Source\n\nGonzález L, Mayo M: Análisis del género en las autorías de las publicaciones científicas del acta otorrinolaringológica española en la última década. Acta Otorrinolaringol. Esp. 2021; 72(4): 230–237. PubMed Abstract \| Publisher Full Text Reference Source\n\nGrijalva R, Fernández V, Meneses M, et al.: Gender Gaps in Scientific Research: A Systematic Review. Dataset. 2023. Publisher Full Text\n\nHoussay B: La investigación científica. Columba: 1960. Reference Source\n\nIESALC: Hacia el acceso universal a la educación superior: tendencias internacionales. [Archivo PDF].2020. Reference Source\n\nIESALC: Mujeres en la educación superior: ¿la ventaja femenina ha puesto fin a las desigualdades de género? [Archivo PDF].2021. Reference Source\n\nLis J, Bahos C: La participación femenina en publicaciones colombianas de economía y administración indexadas en Scopus (1974–junio de 2014). Revista Facultad de Ciencias Económicas: Investigación y Reflexión. 2016; 24(2): 183–213. Reference Source\n\nLópez D, García F, Arroyo A, et al.: Diferencias de género en las publicaciones originales de Archivos de Bronco-neumología en el periodo 2001-2018. Arch. Bronconeumol. 2021; 57(2): 107–114. Reference Source\n\nMaldona K, Guzmán A, Peredo F: La actividad inventiva de las mujeres en Brasil, 1997-2013. Economía: Teoría y práctica. 2015; 3: 53–81. Reference Source\n\nOsorio L, Sokil J: Producción científica sobre COVID-19 en Iberoamérica. Un análisis con perspectiva de género. Revista Iberoamericana de Ciencia, Tecnología y Sociedad. 2022; 17(49): 255–272. Reference Source\n\nPalencia R, Jíménez C: La brecha de género en la educación tecnológica. Ensaio: Avaliação e Políticas Públicas em Educação. 2016; 24(92): 743–771. Reference Source\n\nPardal J, Ochoa C: Revisiones sistemáticas. Revista ORL. 2017; 8(4): 197–203. Publisher Full Text\n\nPardal J, Pardal B: Anotaciones para estructurar una revisión sistemática. Revista ORL. 2020; 11(2): 155–160. Publisher Full Text\n\nPRISMA: 2020. Reference Source\n\nSegovia C, Briones E, Pastells R, et al.: Techo de cristal y desigualdades de género en la carrera profesional de las mujeres académicas e investigadoras en ciencias biomédicas. Gac. Sanit. 2020; 34(4): 403–410. PubMed Abstract \| Publisher Full Text Reference Source\n\nTornero S, Alonso I, García J, et al.: Desigualdades de género en la autoría de las principales revistas médicas españolas durante el año 2017. Anales de Pediatría, Anales de Pediatría. 2020; 93(2): 84–94. PubMed Abstract \| Publisher Full Text Reference Source\n\nUNESCO: Mujeres en la educación superior: ¿la ventaja femeninaha puesto fin a las desigualdades de género?2021. Reference Source\n\nUnites Nation: Objetivos de desarrollo sostenible.2020. Reference Source\n\nVargas D, Requena J, Caputo C: Género en la ciencia venezolana: Desvanecimiento de la brecha. Interciencia. 2016; 41(3): 162–170. Reference Source" }	[ { "id": "225300", "date": "22 Dec 2023", "name": "Linda Elizabeth Ruiz", "expertise": [ "Reviewer Expertise Genger studies in business", "management", "and entrepreneurship." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nCongratulations on your paper; you did a great job gathering all the information from different sources. Surprisingly, many of the documents came from somewhere other than English-speaking journals. I suggest you add some descriptives to your results; I know you added the table where you summarize all the articles, but it would be helpful if you added some statistics summarizing all the results. Another suggestion is adding information about the geography or location where the studies were performed; this is interesting since some areas have a more significant gender gap than others, so I would suggest trying to add that information. In your results section, you can also add subtitles as \"themes\" so you can describe your findings by identifying some patterns or trends.\n\nAre the rationale for, and objectives of, the Systematic Review clearly stated? Yes\n\nAre sufficient details of the methods and analysis provided to allow replication by others? Yes\n\nIs the statistical analysis and its interpretation appropriate? Partly\n\nAre the conclusions drawn adequately supported by the results presented in the review? Yes", "responses": [] }, { "id": "236639", "date": "15 Feb 2024", "name": "Xinyi Zhao", "expertise": [ "Reviewer Expertise gender disparity in science", "scholarly migration", "work-family conflict" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis paper provides a systematic review of gender gaps in science. The methods are clearly explained, and the conclusion, to some extent, summarizes the gender disparity in scientific production, which has some implications for future research work and also some decision-making for the inclusion of female researchers.\n\nHowever, this paper also has some flaws to be addressed: 1. The selected papers are highly skewed toward non-English publications and also toward the science system in Southern America. It is not representative enough for broader areas and lacks sufficient empirical evidence. I personally suggest considering more publications. 2. As far as I know, recently, an increasing number of publications have looked at the gender gaps in science from different dimensions. I was curious why only those 23 papers were selected for systematic review. For example, a lot of papers look at the gender gaps in scholar migration. 3. For each specific dimension, such as participation as the first author or presence in review, the discussion is too general, lacking sufficient details. I suggest authors go back to the literature, providing their main findings and also their limitations, which are unsolved currently.\n\nAre the rationale for, and objectives of, the Systematic Review clearly stated? Partly\n\nAre sufficient details of the methods and analysis provided to allow replication by others? Yes\n\nIs the statistical analysis and its interpretation appropriate? Not applicable\n\nAre the conclusions drawn adequately supported by the results presented in the review? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1445
https://f1000research.com/articles/12-1442/v1	07 Nov 23	{ "type": "Research Article", "title": "Clinical performance of light-cured orthodontic adhesives for bonding brackets – an in-vitro study.", "authors": [ "Sachin Tallani", "Ritesh Singla", "Nishu Singla", "Madhumitha Natarajan", "Jayaprakash Kukkila", "Sachin Tallani", "Nishu Singla", "Madhumitha Natarajan", "Jayaprakash Kukkila" ], "abstract": "Background The dental profession is seeing a constant influx of new adhesive systems from manufacturers, each claiming to be more dependable than the last. This study assessed the bond strength and adhesive remnants of different light-cured adhesives used for bonding metal brackets to teeth.\n\nMethods 80 extracted maxillary premolars with the sound crown structure were acid etched and bonded with brackets on their buccal surfaces utilizing primer and light-cured adhesives into four equal groups, which are Transbond XT, Heliosit, Enlight, and Bracepaste. Shear bond strength (SBS) for de-bonding the brackets were evaluated with Instron- testing machine after 48 hours. The de-bonded samples’ adhesive remnant index (ARI) scores were also measured.\n\nResults The maximum mean SBS was found for Transbond XT (12.91 ± 2.0 MPa), followed by Bracepaste (12.87 ± 1.59 MPa), Enlight (11.77 ± 1.87 MPa), and lowest for Heliosit (10.93 ± 1.71 MPa). According to the four point scale, adhesive remnant index (ARI), Transbond XT has the least adhesive residue left on the tooth, followed by Heliosit. Enlight and Bracepaste have a similar distribution of adhesive, with both having a maximum amount left.\n\nConclusion It can be inferred that all groups involved demonstrated a satisfactory level of bond strength from a clinical perspective. Transbond XT is the preferred orthodontic adhesive over the other three adhesives due to its superior SBS and ARI properties.", "keywords": [ "Orthodontic adhesives", "Light-cured adhesives", "Bracket bonding adhesives", "Shear Bond Strength", "Adhesive Remnant Index" ], "content": "Introduction\n\nDirect bonding procedures, which have become a fundamental technique in modern orthodontics, involve attaching orthodontic appliances to the teeth using adhesives.1 To ensure successful treatment, the bracket-to-tooth bond must be robust enough to sustain the pressures, stresses, and forces exerted during the treatment. The main objective of fixed orthodontic procedures is to achieve optimal bonding between the brackets and the teeth.2 According to Reynolds, force resistances in the 5.9–7.8 MPa range are adequate for withstanding occlusion and orthodontic forces.3 Clinical bracket failure rates vary from 2.7% to 29% for mandibular molars.4 Dislodging brackets during therapy is frustrating for an orthodontist as replacing the dislodged brackets requires additional time and expense and sometimes delays treatment completion.5 Thus, shear bond strength, often known as SBS, is essential when developing bonding materials.6 The type of adhesive used is vital in determining the effectiveness of the bracket’s bond with the enamel surface. Based on Gange’s review, the optimal adhesive for the future should be hydrophilic, removing the requirement for enamel acid etching and exhibiting an SBS value exceeding 20 MPa in dry or wet conditions.7\n\nAt the same time, the adhesive material used must not harm the enamel upon removal after the treatment. It’s essential to ensure that all adhesive remains are eliminated to prevent the accumulation of dental plaque along the remnant adhesive interface.8 Failure to do so may lead to an increased risk of tooth decalcification and the development and progression of caries lesions.9 The procedures of de-bonding and cleaning that are necessary after the treatment can consume a considerable amount of time. Furthermore, residual bonding material left on the tooth surface following removal can gradually become discolored, causing patient dissatisfaction.10 An adhesive remnant index (ARI) approach is a qualitative visual scoring method used to assess the adhesive residue left on teeth after de-bonding the brackets.11 Cehreli et al. and Kaneshima et al. have demonstrated that the ARI visual scoring method is a reliable alternative to scanning electron micrographs (SEM) analysis when evaluating the amount and location of adhesive remnants.12,13 The ARI method offers a reliable method to evaluate adhesive residue.\n\nWhen selecting an orthodontic bracket adhesive, it is essential to consider its bond strength and impact on the tooth’s enamel. The dental profession is seeing a constant influx of new adhesive systems from manufacturers, each claiming to be more dependable than the last.14 A perfect orthodontic adhesive should have sufficient bonding strength while preserving the enamel’s natural appearance after removing it from the tooth.15 Researchers have been putting much effort into developing adhesive materials for bonding orthodontic brackets that are both of the highest quality and least invasive.16\n\nThe performance of an adhesive agent was evaluated by checking its ability to form a strong bond with a tooth and identifying any adhesive material left on the tooth after removal in a controlled and standardized environment. This in-vitro research assessed and compared the SBS (shear bond strength) and ARI (adhesive remnant index) of four light-curing composite adhesives for bonding metal brackets to teeth. The adhesive materials derived from four distinct global manufacturers were Transbond XT (manufactured by 3M Unitek and located in Monrovia, California), Heliosit Orthodontic (manufactured by Ivoclar Vivadent and located in Liechtenstein), Enlight (manufactured by Ormco and located in Glendora, California), and Brace Paste (manufactured by American orthodontics and located in Sheboygan, Wisconsin).\n\n\nMethods\n\nThis was an in-vitro study conducted on 80 extracted maxillary premolars. The research’s permission was acquired from the Institutional Ethics Committee of Kasturba Medical College and Kasturba Hospital (IEC 254/2020). 80 intact maxillary premolars with a sound crown structure free of cavities, restorations, or fractures and no developmental anomalies were collected from patients undergoing fixed orthodontic treatment who have advised extraction in their treatment plan. All the teeth that were chosen were scrubbed with pumice that did not include fluoride to remove any debris or stains, and then they were placed in thymol 0.1% solution (weight/volume) until they were used. For experimentation, every tooth was mounted onto a cold acrylic block measuring 19 mm in diameter and 30 mm in height (Figure 1). The block was created using a cylindrical polyvinyl chloride (PVC) pipe as a template. The tooth roots were buried into the acrylic, and the crown extended outward.\n\nThe enamel of each tooth on the buccal surface was first treated with Kerr Gel Etchant 37.5% phosphoric acid for 15 seconds following the manufacturer’s instructions, followed by 10 seconds of rinsing of teeth with water and air drying. The enamel had a noticeable white frosty appearance distributed evenly over the tooth surface. Then, Transbond XT Light Cure Adhesive Primer was used on all the teeth. The primer was put on with a micro brush and cured for ten seconds with a 3M Elipar light curing unit.\n\nMetal brackets were affixed to these teeth with the following adhesive materials marketed by different brands. The teeth were divided randomly into four groups, each with twenty samples.\n\n○ Group A - Transbond XT (3M Unitek, Monrovia, California, USA)\n\n○ Group B - Heliosit Orthodontic (Ivoclar Vivadent, Liechtenstein)\n\n○ Group C - Enlight (Ormco, Glendora, California)\n\n○ Group D - Bracepaste Adhesive (American Orthodontics, Sheboygan, WI, USA)\n\nFor this study, 80 mini sprint upper first premolar stainless-steel brackets (McLaughlin Bennett 5.0) marketed by Forestadent were used. As per manufactures guidelines, the dimensions of this bracket base measured 3.0 mm mesio-distal, 3.0 mm occluso-gingival, and 1.9 mm bucco-lingual in height. The calculated bracket base area was 9.0 mm2. According to the MBT prescription, these brackets were fixed onto the labial surface of mounted acrylic teeth. The brackets were coated with adhesive specific to each group and carefully placed in appropriate positions on the teeth. The excess adhesive material was cleaned with a straight probe. The adhesive bonds were cured with a 3M S10 ELIPAR high-power LED light cure device (3M Unitek, Monrovia, CA, USA) with an intensity of 1200 mW/cm2 for 10 seconds, focusing on the gingival and occlusal or incisal features of the brackets. Afterward, the samples were submerged in distilled water at 37 degrees Celsius for 48 hours before de-bonding.\n\nIn this study, an Instron universal testing machine (Instron, Model no. 3366) (Figure 2) was utilized to evaluate the shear force needed to detach the brackets fixed on the labial surface of the mounted teeth. Each sample was mounted on the testing machine’s mounting jig to conduct the standardized tests. The acrylic block with mounted teeth with attached brackets was subjected to shear stress (Figure 3). The base of the brackets was maintained perpendicular to the shear load. The bracket base was subjected to a shear force for de-bonding at a crosshead speed of one millimeter per minute. This was done in an occluso-gingival orientation to simulate the typical forces experienced in the oral cavity. The machine measured the maximum force required to cause the bracket to fracture or de-bond in Newtons (N). To calculate mega-pascals (MPa), divide the value of N by the base area of the bracket (9 mm2).\n\nOnce the tests on the samples had been completed, scaled digital pictures of each bracket base were photographed33 using Canon DSLR 1500 D camera with a 100mm macro lens to take high-resolution photos needed to score ARI (Figure 4). Standardized images of the bases were taken at a distance of 30 cm and an angle perpendicular bracket base, with particular attention to ensuring that each image depicts the whole bracket base and any residual adhesive. The photos were loaded onto the computer and given arbitrary numbers before being saved as digital photographs in the JPEG file format; following this, the base of the brackets with residual adhesive was scanned. An automated count of the number of pixels was performed to compute the ARI percentages using Photoshop software (Adobe Systems inc, Mountain View, calif). This software program automatically calculated the bracket base’s adhesive residual surface area percentage (Figure 5). Based on this surface area percentage of residual adhesive, the teeth were assigned a score with Artun, and Bergland ARI on a 4-point scale. The ARI scale states that a score of 0 on the ARI index signifies the absence of adhesive on the enamel. A score of 1 indicates less than 50% of the adhesive remains, and a score of 2 indicates that more than 50% stays on the enamel. Meanwhile, a score of 3 on the ARI index is given when the entire adhesive remains on the enamel.\n\nThe study data were explored using the statistical software SPSS 26.0 (SPSS Inc., Chicago, IL) (RRID: SCR_002865). The Shapiro-Wilk test was employed to evaluate the normality of the data. It was found to follow a normal distribution, which enabled parametric tests to be conducted during data analysis. A one-way analysis of variance (ANOVA) and Tukey Posthoc test were conducted to compare the mean SBS Scores (± standard deviation) of the four groups. The comparison of the proportion of discrete ARI scores among the four groups was done with the Chi-square test. A P value less than 0.05 was considered statistically significant. Analyzing the data thoroughly and accurately to draw meaningful conclusions is essential.\n\n\nResults\n\nThe results of the one-way ANOVA test are presented in Table 1. The study revealed a significant variation in the mean SBS scores across the four groups (p<0.001). The group that used Transbond XT had the highest mean SBS with a value of 12.91 MPa ± 2.0 MPa. The Bracepaste Adhesive group followed closely with a value of 12.87 ± 1.59 MPa. The Enlight group had an SBS of 11.77 MPa ± 1.87MPa, while the Heliosit Orthodontic group had the lowest SBS at 10.93 MPa ± 1.71MPa. Table 2 analyzes the pairwise group comparisons of the mean SBS scores among the groups through the Post hoc test analysis. The mean SBS showed a significant statistical difference between Transbond XT and Heliosit (p = 0.004) and Bracepaste and Heliosit (p = 0.004). The results indicate that Transbond XT and Bracepaste demonstrated significantly higher shear bond strength (SBS) than Heliosit. However, there was no significant difference in the mean SBS values between the Transbond XT, Bracepaste, and Enlight groups.\n\n* A P value less than 0.05 was considered statistically significant.\n\n* A P value less than 0.05 was considered statistically significant.\n\nThe analysis in Table 3 utilized the Chi-Square test to explore the variation in ARI scores across the four groups. According to the study, there was a notable variation in ARI scores between the groups, with a statistical significance of p < 0.0001. In the Transbond XT group, it was discovered that two teeth (10%) had no residual adhesive on the enamel. Half of the teeth in the Transbond XT and Heliosit groups retained less than 50% of the adhesive on the enamel. Among the Enlight and Bracepaste groups, 75% of the samples retained more than 50% of the adhesive on the enamel. However, it was observed that none of the groups had complete retention of the adhesive on the enamel following bracket removal.\n\n* A P value less than 0.05 was considered statistically significant.\n\n\nDiscussion\n\nAn excellent bracket adhesive should have sufficient shear bond strength and be less invasive on the tooth to prevent enamel harm, treatment delays, additional costs, and patient or orthodontist annoyance. This study has analyzed and compared four light cure adhesives from four global manufacturers, i.e., Transbond - XT, Brace Paste, Enlight, and Heliosit. In concurrence with a study, as Reynolds stated, 5.9–7.8 MPa are appropriate to overcome oral forces.3 It has been found that all materials demonstrated optimal bonding strength in the range of 12.91 MPa – 10.93 MPa, capable of withstanding occlusal forces. The study utilized a three-step adhesive procedure, which included conditioning, primer application, and adhesive resin. The Transbond XT primer was included to seal micro-porosity to reduce the risk of demineralization and white spot lesions caused by acid etching on the enamel surface.\n\nThe current study quantitatively graded the adhesive residue on bracket bases by image analysis on digital photos. According to the research, the groups showed a notable variation in their ARI scores, which was statistically significant with a p-value of less than 0.0001. It is preferable to have a low Artun ARI score since it specifies that there is minimal residual adhesive remains on the tooth after debonding.11 This would make the finishing and polishing process easier and faster. Additionally, care must be taken to avoid inducing iatrogenic consequences such as cracks, scratches, and the loss of enamel sections while de-bonding the brackets.\n\nTransbond XT is widely used as an orthodontic adhesive agent with a high level of clinical acceptance.17 Extensive research has been conducted on Transbond XT through various studies, proving its superior bond strength and minimal invasiveness on teeth.18–22 According to this research, the mean SBS for the Transbond XT group was 12.91 MPa ± 2.0 MPa. This finding corresponds with previous studies showing that the SBS of Transbond XT falls within the range of 10.32 MPa to 15.5 MPa.20,21 Also, in harmony with the literature, the Transbond XT group was found to have the maximum SBS among the four groups. It was the least invasive on the tooth as per adhesive remnant index scores.22 In this group, it was observed that half of the sampled teeth retained less than 50% of the adhesive on the enamel. Additionally, two teeth (10%) had no adhesive retained on their enamel. This group is widely respected as the benchmark and frequently serves as a reference for comparison.17\n\nBracePaste is a relatively new adhesive used in orthodontics. Limited studies have been conducted to analyze its performance and efficiency.18,19 Based on the current study, both Bracepaste (12.87 ± 1.59 MPa) and Transbond XT (12.91MPa ± 2.0 MPa) demonstrated similar levels of shear bond strength. The findings align with the studies conducted by Katırcıoğlu and Büyükbayraktar, as well as Stephanie Becker, who also observed no notable variance in strength between the two groups.23,24 However, Shams et al. reported that Bracepaste had lower values than Transbond XT. According to a review by Irfan Eser et al., BracePaste and Transbond XT adhesive can be used interchangeably in clinical settings as they demonstrate similar shear stroke numbers.25 According to the manufacturer, BracePaste and Transbond XT possess similar bonding strengths owing to their common Bis-GMA and Quartz Silica ingredients and nearly identical filler content. However, BracePaste adhesive was found to have significantly higher ARI scores in this study, as 75% of the samples retained more than 50% of the adhesive on the enamel. In contrast, Stephanie Becker found that BracePaste was evenly distributed among ARI scores of “1” and “2”. Also, the ARI scores for BracePaste and Transbond XT were significantly similar.24\n\nIn the current study, Enlight Group demonstrated an optimal shear bond strength of 11.77 MPa ± 1.87 MPa. There was no statistically significant difference in the mean SBS between the Enlight group and the Transbond XT group, with a value of 12.91 MPa ± 2.0 MPa. Similar results were found in the studies conducted by Shaik M S et al., and Rai S et al.18,26 The ARI scores for Transbond and Enlight adhesives were almost identical in their research. However, in this study, the ARI scores of Enlight group were significantly higher than Transbond XT, with 75% of the samples in this group retaining more than 50% of the adhesive on the tooth. This contrasts with the finding that Verma G et al. reported that in most samples, more than 90% of their adhesive or all was left on the brackets.27 During the literature search, only a few studies were discovered that compared the performance of the light cure Enlight group with other existing bonding agents.\n\nAccording to this study, the Heliosit Orthodontic group exhibited a significantly lower mean SBS (10.93 MPa ± 1.71 MPa) than the other groups included in the research. This aligns with the studies conducted by Shaik M S et al. and Rai S et al.18,26 The use of Heliosit as a bonding agent for brackets has not been extensively researched. However, the research findings have consistently demonstrated that Heliosit offers inferior bond strength than Transbond XT. However, it has been established that Heliosit’s bond strengths for orthodontic bonding fall within the recommended range for clinical use.28–31 The current study found it less invasive on the teeth, similar to Transbond XT. This was evident from the evenly distributed adhesive remnant index scores between ARI scores of “1” and “2”.\n\n\nConclusions\n\nAfter analyzing the study results, it can be inferred that all groups involved demonstrated a satisfactory level of bond strength from a clinical perspective. The shear bond strength (SBS) was highest for Transbond XT, followed by Bracepaste, Enlight, and lowest for Heliosit. According to the four Point scale (ARI), Transbond XT has the least adhesive residue left on the tooth, followed by Heliosit. Enlight and Bracepaste have a similar distribution of adhesive, with both having a maximum amount left. Therefore, Transbond XT is the preferred orthodontic adhesive compared to the other three adhesives due to its superior SBS and ARI properties. However, it is essential to exercise caution when interpreting the findings of in vitro studies since these tests cannot precisely simulate the conditions within the oral cavity.", "appendix": "Data availability\n\nMendeley Data: Clinical Performance of Light-cured Orthodontic Adhesives for Bonding Brackets – An In-vitro Study, https://doi.org/10.17632/d7ydctfrsf.2. 32\n\nThis project contains the following underlying data:\n\nClinical Performance of Light-cured Orthodontic Adhesives for Bonding Brackets – Raw data.xlsx (Mendeley Data: Photographs data for Clinical performance of light-cured orthodontic adhesives for bonding brackets – an in-vitro study, https://doi.org/10.17632/fps6fhjdy6.1. 33\n\nThis project contains the following underlying data:\n\n- Group A (Folder containing all images from group A)\n\n- Group B (Folder containing all images from group B)\n\n- Group C (Folder containing all images from group C)\n\n- Group D (Folder containing all images from group D)\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nBozelli JV, Bigliazzi R, Barbosa HA, et al.: Comparative study on direct and indirect bracket bonding techniques regarding time length and bracket detachment. Dental Press J. Orthod. 2013; 18(6): 51–57. PubMed Abstract \| Publisher Full Text\n\nReynolds IR: A review of direct orthodontic bonding. Br. J. Orthod. 1975; 2(3): 171–178. Publisher Full Text\n\nReynolds IR, von Fraunhofer JA : Direct bonding of orthodontic brackets--a comparative study of adhesives. Br. J. Orthod. 1976 Jul; 3(3): 143–146. PubMed Abstract \| Publisher Full Text\n\nZachrisson BJ: A posttreatment evaluation of direct bonding in orthodontics. Am. J. Orthod. 1977 Feb; 71(2): 173–189. PubMed Abstract \| Publisher Full Text\n\nKachoei M, Mohammadi A, Esmaili Moghaddam M, et al.: Comparison of multiple rebond shear strengths of debonded brackets after preparation with sandblasting and CO(2) laser. J. Dent. Res. Dent. Clin. Dent. Prospects. 2016; 10(3): 148–154. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMing-Yi HE, Nie LP, Gui-Gang GU, et al.: Analysis of factors associated with bracket bond failure. Beijing J. Stomatol. 2008; 16(1): 29–31.\n\nGange P: The evolution of bonding in orthodontics. Am. J. Orthod. Dentofac. Orthop. 2015 Apr; 147(4 Suppl): S56–S63. PubMed Abstract \| Publisher Full Text\n\nPickett KL, Sadowsky PL, Jacobson A, et al.: Orthodontic in vivo bond strength: comparison with in vitro results. Angle Orthod. 2001; 71(2): 141–148. PubMed Abstract\n\nÖz AA, et al.: Assessment of the Confidence of the Adhesive Remnant Index Score With Different Methods. Turk. J. Orthod. 2013; 26: 149–153. Publisher Full Text\n\nDavid VA, Staley RN, Bigelow HF, et al.: Remnant amount and cleanup for 3 adhesives after debracketing. Am. J. Orthod. Dentofac. Orthop. 2002 Mar; 121(3): 291–296. PubMed Abstract \| Publisher Full Text\n\nArtun J, Bergland S: Clinical trials with crystal growth conditioning as an alternative to acid-etch enamel pretreatment. Am. J. Orthod. 1984 Apr; 85(4): 333–340. PubMed Abstract \| Publisher Full Text\n\nCehreli SB, Polat-Ozsoy O, Sar C, et al.: A comparative study of qualitative and quantitative methods for the assessment of adhesive remnant after bracket debonding. Eur. J. Orthod. 2012 Apr; 34(2): 188–192. Epub 2011 Jan 24. PubMed Abstract \| Publisher Full Text\n\nKaneshima EN, Berger SB, Fernandes TMF, et al.: Using UV light for adhesive remnant removal after debonding of orthodontic accessories. Braz. Oral Res. 2018 Sep 21; 32: e47. PubMed Abstract \| Publisher Full Text\n\nSofan E, Sofan A, Palaia G, et al.: Classification review of dental adhesive systems: from the IV generation to the universal type. Ann. Stomatol (Roma). 2017 Jul 3; 8(1): 1–17. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBayani S, Ghassemi A, Manafi S, et al.: Shear bond strength of orthodontic color-change adhesives with different light-curing times. Dent. Res. J (Isfahan). 2015 May-Jun; 12(3): 265–270. PubMed Abstract \| Free Full Text\n\nMandall NA, Hickman J, Macfarlane TV, et al.: Adhesives for fixed orthodontic brackets. Cochrane Database Syst. Rev. 2018 Apr 9; 2018(4): CD002282. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHellak A, Ebeling J, Schauseil M, et al.: Shear Bond Strength of Three Orthodontic Bonding Systems on Enamel and Restorative Materials. Biomed. Res. Int. 2016; 2016: 6307107–6307110. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nShaik MS, Pattanaik S, Pattanaik S, et al.: Shear Bond Strength of Different Adhesive Materials used for Bonding Orthodontic Brackets: A Comparative in vitro Study. Orthod. J. Nepal. 2015; 5: 22–26. Publisher Full Text\n\nShams S, Abela S, Andiappan M, et al.: Shear Bond Strengths of 3 Commonly Used Orthodontic Adhesives. Dentistry. 2020; 10: 568. Publisher Full Text\n\nShukla C, Singh G, Jain U, et al.: Comparison of Mean Shear Bond Strength of Light Cure, Self-Cure Composite Resins, Self-Etching, and Moisture-Insensitive Primers: An in vitro Study. J. Indian Orthod. Soc. 2012; 46(4): 254–257. Publisher Full Text\n\nHellak A, Ebeling J, Schauseil M, et al.: Shear Bond Strength of Three Orthodontic Bonding Systems on Enamel and Restorative Materials. Biomed. Res. Int. 2016; 2016: 6307107–6307110. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLinn BJ, Berzins DW, Dhuru VB, et al.: A comparison of bond strength between direct- and indirect-bonding methods. Angle Orthod. 2006 Mar; 76(2): 289–294. PubMed Abstract \| Publisher Full Text\n\nKatırcıoğlu A, Büyükbayraktar Z: Evaluation of the Shear bond strength of Light-Cured and Self-Cured Orthodontic Adhesives. Forum Ortodontyczne/Orthodontic Forum. 2022; 18(1): 18–23. Publisher Full Text\n\nBecker S: An in vitro Comparison of Shear Bond Strength between Two Orthodontic LightCurable Adhesive Pastes. Graduate Theses, Dissertations, and Problem Reports. 8124.2021.\n\nEser I, Cicek O, Ozkalayci N, et al.: Effect of Different Types of Adhesive Agents on Orthodontic Bracket Shear Bond Strength: A Cyclic Loading Study. Materials (Basel). 2023 Jan 11; 16(2): 724. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRai S, Prasad RR, Jain AK, et al.: A comparative study of shear bond strength of four different light cure orthodontic adhesives: An in vitro study. J. Contemp. Orthod. 2022; 6(3): 94–99. Publisher Full Text\n\nVerma G, Trehan M, Sharma S: Comparison of Shear Bond Strength and Estimation of Adhesive Remnant Index between Light-cure Composite and Dual-cure Composite: An in vitro Study. Int. J. Clin. Pediatr. Dent. 2013 Sep; 6(3): 166–170. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDurrani, et al.: In vitro comparison of shear bond strength of Transbond XT and Heliosit Orthodontic as Direct bracket bonding adhesives. Pakistan Oral Dent. J. 2008; 28(2): 203–206.\n\nBradburn G, Pender N: An in vitro study of the bond strength of two light-cured composites used in the direct bonding of orthodontic brackets to molars. Am. J. Orthod. Dentofac. Orthop. 1992 Nov; 102(5): 418–426. PubMed Abstract \| Publisher Full Text\n\nMurray SD, Hobson RS: Comparison of in vivo and in vitro shear bond strength. Am. J. Orthod. Dentofac. Orthop. 2003 Jan; 123(1): 2–9. PubMed Abstract \| Publisher Full Text\n\nYousry T, Abdel-haffiez S: Comparison of shear bond strength of brackets bonded with four different bonding protocols at different time intervals; An in vitro study. Egypt. Orthod. J. 2020; 58(December 2020): 1–18. Publisher Full Text\n\nSingla R, Singla N: Clinical Performance of Light-cured Orthodontic Adhesives for Bonding Brackets – An In-vitro Study. [Dataset]. Mendeley Data. 2023; V2. Publisher Full Text\n\nSingla R: Photographs data for Clinical performance of light-cured orthodontic adhesives for bonding brackets – an in-vitro study. [Dataset]. Mendeley Data. 2023; V1. Publisher Full Text" }	[ { "id": "226516", "date": "29 Dec 2023", "name": "Vincy Antony", "expertise": [ "Reviewer Expertise Orthodontic root response", "tooth movement", "in vitro studies of bond strength" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\n1. Overall the article was well-written 2. In the Methods section, 'cold acrylic block' was mentioned. Kindly correct/explain the term. 3. In the abstract methods section, '80 extracted maxillary premolars with the sound crown structure' . Keep it as '80 extracted maxillary premolars with sound crown structure' 4. The title will have to be changed- Since it is an in-vitro study, the phrase 'clinical performance' cannot be used.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "226515", "date": "22 Jan 2024", "name": "Kevser Kurt Demirsoy", "expertise": [ "Reviewer Expertise orthodontics", "orthognathic surgery", "3D print systems", "Laser in orthodontics" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis study assessed the shear bond strength and adhesive remnant index of different light-cured adhesives used for bonding metal brackets to teeth. The article is an in vitro study and aimed to evaluate the SBS and ARI scores of different orthodontic adhesive types. Although the subject is far from current, it seems to be designed to evaluate the SBS and ARI scores of the bracket adhesive under the brand name Heliosit, which has not been studied much before. Because there are many SBS and ARI studies conducted in the literature on other equivalent brands. Apart from this, improving the discussion section and adding the limitations section will contribute to the article. Also 33 references are written in the references section, but 31 references are shown in the text. The references section also needs to be revised.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "226513", "date": "22 Jan 2024", "name": "Saeed AlSamak", "expertise": [ "Reviewer Expertise Orthodontics", "Biomaterials" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nOrthodontic adhesive systems play a pivotal role in the success of bracket bonding during orthodontic treatments. This article evaluates the clinical performance of four commercially available orthodontic adhesives. The previous literature extensively investigated the clinical performances of Transbond XT, Heliosit, Enlight, and Bracepaste regarding shear bond strength and adhesive remanent index. The article well cites the current literature. The study design was appropriate to evaluate the mechanical performances of orthodontic adhesives through shear bond strength and adhesive remnant index. Several commercially available light-cured orthodontic adhesives were selected for this in-vitro study. Standardized brackets were bonded to enamel surfaces, simulating clinical conditions. The specimens were subjected to various assessments, including bond strength measurements, using universal testing machines. The adhesive remnant index was used to evaluate the amount of adhesive remaining on the enamel surface after debonding.\nHowever, the authors used an alternative way to assess the adhesive remnant index by calculating the amount of adhesive remaining on the bracket instead of the enamel surface, which differs from the original adhesive remnant index described by Artun and Bergland. During the test of shear bond strength, there is a possibility that the orthodontic adhesive will be debonded from both the bracket surface and the enamel surface, which may lead to errors in the calculation of the amount of adhesive remaining on the enamel surface. The statistical analysis was appropriate to reveal the study results. The study revealed significant variations in bond strength among the different adhesive systems tested. The observed differences in bond strength emphasize the need to select orthodontic adhesives carefully based on specific clinical requirements. This in-vitro study contributes valuable insights into the clinical performance of light-cured orthodontic adhesives for bonding brackets regarding shear bond strength and adhesive remnant index.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1442
https://f1000research.com/articles/12-29/v1	09 Jan 23	{ "type": "Review", "title": "Bakuchiol and its pharmacological benefits", "authors": [ "Nuder Nower Nizam", "Sohel Mahmud", "Mohammad Kamruzzaman", "Md. Kamrul Hasan", "Nuder Nower Nizam", "Sohel Mahmud", "Mohammad Kamruzzaman" ], "abstract": "Background and aims: Natural compounds extracted from medicinal plants have recently gained attention in therapeutics as they are considered to have lower toxicity and higher tolerability relative to chemically synthesized compounds. Bakuchiol is one such compound; it is a type of meroterpene derived from the leaves and seeds of Psoralea corylifolia plants. Natural sources of bakuchiol have been used in traditional Chinese and Indian medicine for centuries due to its preventive benefits against tumors and inflammation. It plays a strong potential role as an antioxidant with impressive abilities to remove Reactive Oxygen Species (ROS). This review has focused on bakuchiol’s extraction, therapeutic applications, and pharmacological benefits.\nMethods: A search strategy has been followed to retrieve the relevant newly published literature on the pharmacological benefits of bakuchiol. After an extensive study of the retrieved articles and maintaining the inclusion and exclusion criteria, 106 articles were finally selected for this review.\nResults: Strong support of primary research on the protective effects via antitumorigenic, anti-inflammatory, antioxidative, antimicrobial, and antiviral activities are delineated.\nConclusions: From ancient to modern life, medicinal plants have always been drawing the attention of human beings to alleviate ailments for a healthy and balanced lifestyle. This review is a comprehensive approach to highlighting bona fide essential pharmacological benefits and mechanism of action of therapeutic implications.", "keywords": [ "Bakuchiol", "Psoralea corylifolia", "medicinal plants", "health benefits" ], "content": "Introduction\n\nPlants have been used in traditional Indian and Oriental medicine for centuries. Despite improved access to essential medicines as supported by World Health Organization (WHO),1 a large population, especially in less developed countries, still relies on plant-based medications for primary health care needs due to ease of availability and high benefit to cost ratio.2 There is an increase in global demand for medicinal plants due to improved quality of life. Some compounds have been extracted from these beneficial plants to study their mechanisms of action that facilitate their beneficial effects.\n\nBakuchiol, extracted predominantly and traditionally from the Psoralea corylifolia L. plant, is one such compound. It is a meroterpene that has shown potent antimicrobial, anti-inflammatory, anti-osteoporotic, and antitumorigenic activities and exhibited other beneficial uses.3 It has also gained huge popularity in the beauty industry as a tolerable analogue of retinol in skin therapeutics.4 Studies have also shown bakuchiol’s protective effect on the liver, heart, bones, and other organs.5 Recent studies have focused on bakuchiol and its therapeutic effects, showing promise as a multi-targeting therapeutic agent.6–13\n\nBakuchiol is a naturally occurring prenylated phenolic isoprenoid (Figure 1). It is a type of meroterpene with a chiral tetra-alkylated (all-carbon) quaternary center.15 It was first extracted from the seeds of Psoralea corylifolia in 1966 when its plane structure was also determined, while its configuration of the quaternary center was determined in 1973.16 The formal chemical name of bakuchiol is 4-[(1E,3S)-3-ethenyl-3,7-dimethyl-1,6-octadien-1-yl]-phenol. Its naturally occurring form (S)-(+)-Bakuchiol has an enantiomer (R)-(-)-Bakuchiol, and its chirality influences their actions and effectiveness.\n\nThis figure has been adapted from Khuranna et al.14 under CC BY 4.0.\n\nThe source of bakuchiol varies between regions; it is commonly extracted from the seeds and leaves of Psoralea corylifolia (Babchi plant) in India, which belongs to the Leguminosae plant family, and is a major phytochemical present in the root and stem of Ulmus davidiana var.17 Japonica (Japanese Elm) is widely distributed in China, Japan, and Korea.6 These plants have been used in their regions of wide distribution in traditional medicine to treat inflammatory disorders and cancer. Piper longum (Long pepper), Psoralea glandulosa L, Otholobium pubescens, Prosopis glandulosa, Aerva sanguinolenta, Psoralidium tenuiflorum, Pimelea drupacea, Bridelia retusa, Spiraea formosana, Elaeagnus bockii and other sources of bakuchiol have been discovered in recent times.18–20\n\nMedicinal plants e.g.: Psoralea corylifolia where bakuchiol is derived have been used directly as traditional medication or via pharmaceutical preparations in modern medicine. Recent times have shown collective evidence highlighting the immense potential of traditionally used medicinal plants and their derived compounds.21 An increase in international demand and trade has also been observed, appreciating its low costs and tolerability. In addition, an increasing trend of antimicrobial resistance has been observed due to inappropriate dosage and lack of regulation of prescriptions. It is thus imperative to find new sources of antiviral and antimicrobial therapy to account for the increased resistance observed. Plants and phytochemicals are generally considered sources of tolerable, less toxic treatments.21 Bakuchiol has recently been studied in detail in research settings while being used in traditional medicine for centuries.22\n\nThis review article focuses on the potential pharmacological benefits of bakuchiol, focusing on its promising protective effects in controlling activities that lead to the amelioration of non-communicable diseases, as well as verifying its function as an antimicrobial and antiviral agent. The workflow of this review is illustrated in Figure 2. The knowledge gained from this review will shed light on the published research to pave the path for future research perspectives and considerations.\n\nThis figure is an original figure produced by the author(s) for this review article. The image of Psoralea corylifolia has been reproduced from Niu et al.23 under CC BY-NC 3.0.\n\n\nMethods\n\nThis review evaluates the pharmacological uses of bakuchiol, focusing on its promising protective effects on controlling both communicable and non-communicable diseases. Data for this study have been amalgamated from both primary and secondary data resources, including clinical trials (both randomized and non-randomized), as well as in vitro and in vivo studies, which have evaluated the use of bakuchiol as a potential pharmacological agent in controlling multiple disease conditions.\n\nSeveral inclusion and exclusion criteria were fixed while selecting articles for this review. The works done from 1990 to 2022 have been studied rigorously. Literature searches were done using the keywords such as “Psoralea corylifolia AND/OR Babchi AND/OR Bakuchiol” in various available online scientific databases. Articles that represent phytochemistry AND/OR pharmacological activity AND/OR health benefits of Bakuchiol published in PubMed, Web of Science, PMC, Google Scholar, ScienceDirect, and ResearchGate were incorporated in this review. From the evaluation of 106 articles, the effect of bakuchiol as a potential antitumorigenic, anti-inflammatory, antioxidative, antimicrobial, and antiviral agents, as characterized in several studies, was finally assessed for this review. In addition, clinical trial databases were also searched to find current registered clinical trials of the use of bakuchiol in multiple disorders, including diabetes, inflammatory disorders of the skin and other organs, oral disorders, cancer, and coronavirus disease 2019 (COVID-19). Only articles published in English were considered for this review. The databases were searched in a timeline from 2010 to date, except for articles related to its initial discovery and extraction, and only approved and published data were considered for this review unless otherwise mentioned. In most studies, the outcomes measured and considered for this review include the severity and comparative assessment of disease progression in bakuchiol-treated and untreated groups.\n\n\nResults\n\nTraditional plant-based medications work by grinding the interest portion and then extracting it into carrier oils or spirits.24 However, specific compounds cannot be distilled this way and there may also be the presence of potentially cytotoxic compounds in an extract made in traditional methods.25,26 While beneficial, the number of bioactive ingredients present in natural sources is generally meager, and extraction processes are time-consuming and lab-intensive, hindering the mass-scale use of natural products in drug selection and development. Current extraction methods include distillation, cold pressing for oils, and solvent extraction, the most commonly used method.27\n\nFactors that affect the ease of solubility and diffusivity are considered during extraction processes, including consideration of the properties of the solvent being used, such as the laws of inter miscibility as well as cost and safety, the size of the raw materials material being used, temperature and time duration.27 Generally, methanol and ethanol are used in solvent extraction for phytochemicals. Smaller particle sizes of raw materials are preferred for better extraction efficiency.28 Temperature is also an essential factor in controlling the extraction rate, as higher temperatures increase solubility and diffusion, but this is not ideal for highly volatile compounds as higher temperatures can cause the decomposition of thermolabile compounds.29\n\nBakuchiol is soluble in organic solvents such as ethanol and dimethyl sulfoxide (DMSO) and sparingly soluble in aqueous solutions.6 Its solubility can be increased by first making a stock solution with ethanol and then diluting it with aqueous buffers for experimental purposes. The most commonly reported method is extraction with 80% ethanol, followed by silica column chromatography.6 This method is also suitable for deriving other phenolic phytochemicals present in the sources of bakuchiol.\n\nThe first synthesis of the naturally occurring form of bakuchiol was done by Carnduff and Miller in 1967, with a Claisen rearrangement being a crucial part of the synthesis process; however, this was not enantioselective.20 Other variations in chemical synthesis methods have also been developed over the years, but current methods focus on concisely synthesizing enantioselective (S)-(+)-Bakuchiol. It is also synthesized chemically in four steps from (E)-geranic acid under aldol reaction conditions, with an overall yield of 53%, increasing commercial availability.30 Stock solutions of (S)-(+)-Bakuchiol synthesized from (E)-geranic acid can be made by dissolving the compound in DMSO.\n\nHowever, conventional extraction methods require a large solvent volume and longer durations. Separation techniques have developed and come a long way. More sophisticated techniques, including various forms of chromatography, have been developed to extract specific compounds from the plant source.14 Other methods (supercritical fluid extraction, pressurized liquid extraction, and microwave-assisted extraction) with much shorter extraction time and lower solvent consumption may be considered for mass extraction of bakuchiol.14 They may be used on a large scale for cheaper drug development. These methods have also been used in the extractions of natural products, so their efficiency in the extraction of bakuchiol needs to be evaluated.31\n\nMethods are being studied to increase the bioavailability and absorption of bakuchiol and its sources. Cho et al. (2011) studied the stability and physicochemical property of P. corylifolia extract encapsulated in 3 different vesicles (liposome, niosome, and transfersome) in nude mouse skin.32 The results of this study suggest that the use of niosome and transfersome could be a good bioavailability enhancement system (BAES) for P. corylifolia extract to improve skin permeation and stability, highlighting the importance of finding methods to increase the bioavailability of bakuchiol and other phenolic phytochemicals.32\n\nA large number of studies have evaluated the role of various phytochemicals and their pharmacological applications in communicable and non-communicable diseases.33 In addition, other potentially beneficial compounds have also been extracted from the plant sources of bakuchiol.31 However, since this review focuses on bakuchiol and its pharmacological benefits, other phytochemicals and compounds will not be reviewed in detail and will only be referred to as appropriate. A brief description of the extraction and solubility of bakuchiol precedes the comprehensive analyses of its various pharmacological benefits. The mechanism of actions, functions, and uses of the naturally occurring compound bakuchiol and its implications in therapeutic approaches are described below and illustrated in Figure 3.\n\nThis figure is an original figure produced by the author(s) for this review article.\n\nAnti-Inflammatory effects of bakuchiol\n\nInflammation is generally caused by the production of pro-inflammatory mediators and cytokines such as nitric acid, prostaglandin E2 (PGE2), tumor necrosis factor-alpha (TNF-α), and interleukin 6 (IL-6).34 The anti-inflammatory effects of bakuchiol have been evaluated in multiple studies, and it has been found that the anti-inflammatory effects of bakuchiol work via various mechanisms.7 It can inhibit the degranulation of neutrophils and decrease cell migration and myeloperoxidase activity (involved in producing oxidants such as hypochlorous acid) in inflammatory sites, generating multiple mechanisms of controlling leukocyte function and inflammation in various types of cells.35\n\nStudies have reported the ability of bakuchiol to inhibit nitric acid and PGE2 production, both pro-inflammatory mediators generated by inducible enzymes nitric oxide synthase and cyclooxygenase-2.6–8 A concentration of 50μM of bakuchiol has shown a reduction in nitric acid and PGE2 in macrophages by over 50% without exhibiting cytotoxicity.6 Bakuchiol can inhibit the expression of nitric acid synthase gene via the inactivation of nuclear factor kappa-light-chain-enhancer of activated B cells (NFkB), which can inhibit nitric acid production and IL-6 induced signal transducer and activator of transcription 3 (STAT3) activation, inhibiting IL-6 production in multiple cell lines.36 This is comparable to other natural compounds, such as aurantiamide acetate from the roots of the Baphicacanthus cusia.37\n\nBakuchiol can suppress inflammatory responses via downregulation of the p38 MAPK/ERK signaling pathway in microglia cells controlling neuroinflammation.7 Moreover, it can reduce the production of leukotriene B4 (LTB4), which contributes to disease severity in chronic inflammatory diseases in a dose-dependent manner.38 LTB4 can generate reactive oxygen species (ROS) via a Rac-dependent pathway, and a reduction in LTB4 production also reduces ROS production.39 Bakuchiol and its derivatives have also exhibited inhibitory activities in lipopolysaccharide (LPS) induced NO production in macrophages.40\n\nBakuchiol decreases the phosphorylation of ERK, p38, and NF-κB. It arrests the mRNA expression of pro-inflammatory molecules, including TNF-α and interleukin-1β (IL-1β), and inducible nitric oxide synthases (iNOS) in Methylglyoxal-administered mice, which reduces inflammation and helps in the prevention and control of diabetes.41\n\nAltogether, bakuchiol can provide beneficial therapeutic implications in treating inflammatory diseases such as nephritis, asthma, diabetes, skin inflammatory conditions such as psoriasis, and neurodegenerative disorders such as Alzheimer’s.40–42\n\nAntimicrobial effects of bakuchiol\n\nNature provides a great potential source for antimicrobial drug discovery. Phytochemicals extracted from natural sources are considered to have fewer side effects and a variety of functions, and some exhibit antimicrobial activity. In addition, natural compounds are much more accessible than synthetic compounds available for treatment; most developing countries depend on plant-based medications as the first line of treatment.43\n\nTerpenes such as thymol, carvacrol, eugenol, and menthol, most commonly used in fragrances and aromatherapy, show broad-spectrum antimicrobial activity through efflux pump inhibition and inhibition of bacterial growth.44–46 Bakuchiol is also a terpene that has recently been characterized to have multi-beneficiary effects on disease control, both communicable and non-communicable.\n\nThe antimicrobial effects of bakuchiol have been studied as a natural source of oral healthcare, and multiple studies have shown potent antimicrobial activity.11,47,48 Studies evaluating bakuchiol’s antimicrobial effect have shown potent antibiotic effects against Streptococcus aureus. Another study examining the antimicrobial effects of bakuchiol against Streptococcus mutans showed increased inhibition in the growth of colonies in a dose-dependent manner. Similar results were observed for Streptococcus sanguis, Streptococcus salivarius, Streptococcus sobrinus, Enterococcus faecalis, Enterococcus faecium, Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus casei, Lactobacillus plantarum, Actinomyces viscosus, Porphyromonas gingivalis and other bacterial strains that contribute to oral diseases.11\n\nConventional treatments for oral infections usually contain chlorhexidine, a gold standard.49 However, as it was toxic in large quantities and caused discoloration of the tooth surface, other natural-based alternatives were evaluated, some of which were successful inhibitors for antimicrobial growth.50–53 There is also a concern about emerging antimicrobial resistance, which has increased rapidly in the past decade and is now one of the leading causes of death.54,55 Given the increasing emergence of antimicrobial drug-resistant strains, finding nature-based alternatives to antibiotics is vital to help control the resistance.\n\nBakuchiol works by rupturing the cell membrane in bacterial strains, inhibiting DNA polymerase and other DNA replication enzymes, and reducing biofilm production by E. faecalis and E. faecium.56 These studies implicate the potential uses of bakuchiol in maintaining oral health care. It can potentially be used in combination with other natural pre-existing methods of oral healthcare to target the increasing demand in the market for natural healthcare products.\n\nBakuchiol also has antifungal effects, and ethanol isolates of bakuchiol from PF effectively inhibit the growth of dermatophytes in vitro.57 Oral Candida species have also been susceptible to bakuchiol treatment with minimum inhibitory concentration (MIC) ranging from 12.5 to 100 μg/mL, causing a reduction in growth rates, viable counts, cell metabolic activity and biofilm mass.58\n\nBakuchiol in the treatment of skin disorders\n\nBakuchiol has recently gained mass popularity as an active component in skin care products due to its functional similarity with retinoid compounds and relatively higher tolerability.59 It has been shown to work with melatonin and vitamin C derivatives to regulate genes involved in the modulation of angiogenesis, collagen biosynthesis, skin barrier function, and other aspects of skin cell biology.9 Bakuchiol also has retinol-like properties in modulating genes that regulate extracellular matrix production and the dermal-epidermal junction.60 It can help deter skin aging by enhancing human fibroblast cell activity and inhibiting the expression of matrix metalloproteinases by increasing tissue inhibitors of metalloproteinases-1 (TIMP-1), tissue inhibitors of metalloproteinases-2 (TIMP-2), collagen type I (Col I), and collagen type III (Col III) mRNA, and decreasing the expression of matrix metalloproteinase-1 (MMP-1) mRNA.61\n\nBakuchiol exhibits retinol-like functionality causing upregulation of types I, III, and IV collagen, which make up the skin’s extracellular matrix and basement membrane, in in vitro models.4 When formulated into a skincare product, regularly applied, and tested under clinical settings for 12 weeks, significant improvement in facial fine lines and wrinkles were observed, along with improvements in skin elasticity, firmness, and pigmentation, as well as a reduction in photodamage.4 This effect was observed in multiple studies on participants of varying races.4,59 In addition, other studies have also confirmed an increase in moisture retention from the regular application of products formulated with bakuchiol extract.62 It also works synergistically with other natural compounds to reduce inflammation and protect and maintain naturally aging skin.10\n\nThis higher tolerability allows bakuchiol to be an effective anti-aging treatment for people of all skin types. Furthermore, it can also be formulated for people with hyperpigmentation historically perceived as less tolerant of retinol.63\n\nUnlike retinoids, bakuchiol can be used daily due to its photostability.59 Combinations of retinoid and bakuchiol therapy also reduce oxidative stress induced by retinoids,64 as bakuchiol helps to stabilize retinol under photo-oxidative environments.\n\nBakuchiol works as an antioxidative agent due to its ability to interfere with free radical production, decrease the translocation of mitochondrial apoptosis-inducing factors, and its ability to quench superoxides and other radicals in vitro.13,49 Overall, it can be deduced that its naturally occurring phytochemical functions as a potent antioxidant that can help maintain cellular turnover and ensure protective functions for all organs. With higher tolerability, photostability, and ability to slow down skin aging, bakuchiol can be considered a suitable plant-based alternative to retinol, traditionally extracted from animal sources.\n\nAntitumorigenic effects of Bakuchiol and Hormone replacement therapy\n\nBakuchiol has traditionally been used in medicine for the treatment of various cancers. Early studies have shown that bakuchiol can inhibit cell proliferation, and further studies have recently evaluated its mechanisms of action in various cancer cell lines compared to its analogue resveratrol.12 It can induce ROS-related apoptosis in lung adenocarcinoma A549 cell line, S phase arrest, caspase 9/3 activation, p53 up-regulation, and B-cell lymphoma 2 (Bcl-2) downregulation, all of which contribute to anticancer activities.12 Bakuchiol has also shown inhibitory effects on DNA polymerase enzymes, while a related compound Bakuchicin inhibits the actions of topoisomerase II.65 Its effects on DNA replication enzymes and relatively much lower cytotoxic effects than its analogue resveratrol make it a potent tolerable therapeutic compound for treating non-small cell lung cancer.12\n\nHigh doses of Bakuchiol (>2 μg/mL) inhibited cell proliferation of breast cancer MCF-7 cells through actions via estrogen receptors (ER), inducing ERβ expression and suppressing ERα expression.66 S phase arrest was also observed in MCF-7 cells along with upregulation of ATM serine/threonine kinase (ATM), phosphorylated cell division cycle protein 2 (P-Cdc2), p21, myelin transcription factor 1 (Myt1), and downregulation of Cyclin B1, which implies the blocking of Cdc2 activation by upregulation of ERβ may play a role in the S phase arrest.66 It also induced apoptosis in MCF-7 cells with an increase in expression of pro-apoptotic Bcl-2 and cleaved caspase proteins, pointing towards its involvement in apoptosis via the intrinsic mitochondrial pathway, similar to that observed in the lung adenocarcinoma A549 cell line and even liver cancer cell lines.12,66\n\nIn addition, bakuchiol was found to effectively inhibit the activation of hypoxia-inducible factor 1 (HIF-1) and NFkB in gastric cancer AGS and cervical adenocarcinoma HeLa cell lines.67 It also inhibits proliferation, migration and invasion of androgen-independent prostate cancer cell line PC-3 by inactivating NF-κB signaling via Androgen receptor (AR) and ERβ expression in a time and dose-dependent manner. Bakuchiol’s cytotoxicity was considered in this assay via LDH assay, and the non-toxic concentration used in further experimentation was determined to be at 10μM.68\n\nBakuchiol-treated NUGC3 gastric cancer cells have also been shown to express reduced levels of phosphorylated protein kinase B (AKT) protein and increased p-extracellular signal-related kinase 1/2 (ERK1/2) and p-c-Jun N-terminal kinase (JNK) expression, indicating the induction of cell death was mitochondria-dependent, working via the MAPK/PI3K/AKT pathways.69\n\nFurthermore, bakuchiol was shown to induce estrogenic activity in vivo and in vitro study models.66 It activates the ERβ receptor and suppresses the ERα receptor, which reduces CDC2 activity and promotes S phase arrest, reducing tumor cell proliferation.66 This promotes bakuchiol as a phytoestrogen and anticancer drug, promoting safer hormone replacement therapeutics. Bakuchiol can also prevent bone loss and delay osteoporosis in post-menopausal women by activating the ERs.70 This is of increased significance as hormone replacement therapies usually use estrogen, which is linked with an increased risk of breast cancer.\n\nThere have also been other compounds extracted from Psoralea corylifolia L.that express similar pharmacological activities.71 Psoralidin, a natural phenolic coumarin, is one such compound that is beneficial in various diseases, including osteoporosis and hormonal cancers.72 This is elucidated by inducing oxidative stress and apoptosis in tumorigenic cells, which promote autophagy-dependent cell death and activate the ER signaling pathway. The use of bakuchiol or other anticancer natural compounds should be evaluated to further check their efficacy in preventing or controlling cancer development. This would provide a suitable cheaper alternative to current cancer chemotherapeutics in the market, which are not accessible equitably among patients due to high costs.\n\nAntioxidative effects of bakuchiol\n\nThere has been an increasing interest in identifying and characterizing antioxidant compounds to treat oxidative stress and related diseases, among which bakuchiol has been characterized as it has shown vital antioxidant activities in multiple studies.5,20,73 When studying the scavenging activity of bakuchiol against various oxidizing radicals, it was determined that the terpenoid chains present in bakuchiol play a role in preventing lipid peroxidation.13 It can protect against rat liver injury by inhibiting lipid peroxidation,74 with similar effects expressed by studies on other natural products such as pumpkin seeds and acanthoic acid.75 Bakuchiol can also inhibit liver fibrosis and show hepatoprotective effects by inhibiting oxidative stress while inducing apoptosis in myofibroblasts, relieving the hepatotoxicity of various toxicants.74,76 Bakuchiol reduced cell death in retinal ganglion cells (RGC-5) and reduced ROS-induced apoptosis and cell death in vitro.49\n\nIn vivo studies have demonstrated that bakuchiol reduces retinal degeneration following optic nerve injury.49 It has also been observed that bakuchiol can protect against sepsis and sepsis-induced acute kidney injury by significantly reducing inflammation and renal oxidative stress while inhibiting induced activation of NFkB and p38-MAPK signaling pathways in the kidneys.77 Its ability to block NFkB signaling also allows for cardioprotective functions, as seen in various mice models, allowing its application as a potential treatment for pathological cardiac hypertrophy.78\n\nIn another study, bakuchiol showed protective effects against early brain injury by reducing ROS, malondialdehyde (MDA), 4-Hydroxynonenal (4-HNE), 3-Nitrotyrosine (3-NT), and other biomarkers of oxidative stress produced by lipid peroxidation.79 Conversely, it causes increases in the enzyme activity of superoxide dismutase (SOD) and inducible glutathione peroxidase (GSH-Pxi), both of which play crucial roles in the body’s antioxidant defense system while reducing mitochondrial damage. As seen in this study, phosphorylation of AMP-activated protein kinase (AMPK) and thioredoxin 1 (Trx1) protein levels increased. In contrast, thioredoxin-interacting protein (TXNIP) levels decreased due to treatment with bakuchiol, which was thought to have occurred due to the regulation of Trx1 and TXNIP levels.\n\nBakuchiol may be considered a potential candidate for the prevention and treatment of insulin resistance as it can reduce induced insulin resistance and oxidative stress with reduced ROS expression and enhanced antioxidant enzyme expression.41\n\nBakuchiol can also reduce the severity of myocardial ischemia-reperfusion injury (IRI) by impairing mitochondrial oxidative damage through regulation of sirtuin 1/proliferator-activated receptor gamma coactivator 1-alpha (SIRT1/PGC-1α) pathway signaling via increased in the expression of SIRT1.80 It reduced mitochondrial oxidative damage by increasing the action of mitochondrial succinate dehydrogenase, cytochrome c oxidase, and mitochondrial SOD and decreased the production of malondialdehyde.79\n\nBakuchiol has also increased anti-apoptotic Bcl2 and decreased pro-apoptotic Bax and cleaved caspase 3, which helps control injury-induced cell death, and inhibitors of SIRT1 abolished the effects of bakuchiol further highlight the role of its interaction with SIRT1 signaling.78 Bakuchiol has also been found to reduce hyperglycemia-induced cardiomyopathy by activating the SIRT1/Nrf2 pathway, which reduces myocardial oxidative stress and elevates antioxidant production.73\n\nSimilar effects are observed in other natural products such as curcumin, melatonin, berberine, and icariin.80–84 These also reduce myocardial IRI via other pathways, highlighting the importance of studying the other potential signaling pathways regulated by bakuchiol, which leads to its ability to reduce injury-induced cell damage. Moreover, bakuchiol should be combined with other natural compounds with similar protective effects for potential synergistic activity.\n\nAntiviral effects of bakuchiol\n\nThe antiviral effect of bakuchiol has been evaluated in some studies.15,85,86 Viral diseases are constantly evolving in pathogenicity, and certain strains have been reported to be highly pathogenic to humans, as was seen in the influenza A pandemic in 1918, causing 50 million deaths,87 and in the current pandemic caused by the SARS-CoV-2 virus,88 which has infected more than 500 million people and has caused more than 6 million deaths since its first reported case on Dec. 31, 2019.89 The SARS-CoV-2 virus enters the host cell via the interaction between its receptor-binding domain (RBD) of spike glycoprotein with the angiotensin-converting enzyme 2 (ACE2) receptor found on the plasma membrane of the host cell.90 Given the rapid emergence of resistant viral strains, searching for potent and tolerable antiviral drugs is essential to combat and control viral infections.\n\nPlant-based natural compounds are being examined extensively for their therapeutic effects,91,92 and bakuchiol has also been evaluated, given its use in traditional medicine systems to treat a wide range of diseases. In in vitro studies on Madin-Darby canine kidney (MDCK) cells infected with influenza A H1N1 strain, the naturally occurring form of bakuchiol was found to inhibit influenza A growth and infection while reducing expression of viral mRNAs and proteins, decreasing viral load.15 In addition, it also was able to activate Nrf2 and two Nrf2-induced genes, NAD(P)H quinone oxidoreductase 1 and glutathione S-transferase A3, promoting activation of transcriptional regulation and regulating virus-induced host body oxidative stress response. To a lesser extent, these effects were also observed in its enantiomer form, highlighting the importance of chirality in designing potent antiviral drugs.\n\nFurther studies on related compounds such as cyclobakuchiols A, B, and C, derived from (+)-(S)-Bakuchiol as well as from its natural sources, have also established strong potential to inhibit viral growth, infection, and expression of viral mRNAs and proteins in influenza A virus-infected MDCK cells via similar mechanisms.85 Other natural compounds derived from natural sources show antiviral actions against Influenza A, such as aurantiamide acetate extracted from the plant’s roots Baphicacanthus cusia.37 These compounds work via similar mechanisms as bakuchiol, as observed in MDCK cells. In addition, these natural compounds also reduce virus-induced inflammatory responses via the suppression of NF-kB signaling pathways and pro-inflammatory cytokines.93\n\nIn a more recent study, bakuchiol effectively inhibited severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pseudovirus entry at concentrations of up to 100μM without toxicity. Furthermore, in HEK293 cell lines overexpressing human ACE2 receptors, bakuchiol also effectively blocked RBD-ACE2 binding at the cell membrane.86 This also implies that further evaluation of bakuchiol on its effect on other viral diseases is critical in the current pandemic setting caused by the SARS-CoV-2 virus to find effective antiviral treatments to reduce disease transmission and severity.\n\nOther natural compounds have also been evaluated in recent studies, among which epigallocatechin gallate (EGCG), 20(S)-ginsenoside Rg3 (SRg3), 20(R)-ginsenoside Rg3 (RRg3), isobavachalcone (Ibvc), and isochlorogenic A (IscA) were found to effectively inhibited pseudovirus entry at concentrations up to 100 μM.86,94\n\nAmong these compounds, EGCG and lbvc have shown comparable effects to bakuchiol administration by inhibiting the SARS-CoV-2-induced cytopathic effect and plaque formation and demonstrating a dual binding to RBD and ACE2.93 Another compound, 1,2,3,4,6-O-Pentagalloylglucose (PGG), also effectively inhibited virus binding and infection in ACE2 overexpressing human host cells, binding more with Spike-RBD than ACE2 receptors.95 It blocks the fusion of SARS-CoV-2 to hACE2 receptors on a dose-dependent level. Spike RBD PGG was also found to exhibit anti-influenza-virus activity by reducing the accumulation of nucleoprotein and viral hemagglutinin in plasma membranes at the late stage of the replication cycle and inhibiting the release of progeny virus from infected cells. PGG also works in ameliorating HBV, HCV, HIV infections.95 PGG may be a safe and potential antiviral agent against COVID-19 by blocking the fusion of SARS-CoV-2 spike-RBD.\n\nCombination effects\n\nMost studies evaluating the effects of bakuchiol used in combination with other molecules tend to be formulated by the cosmeceutical industry. It is also used in combination with retinol to help increase its photostability, or moisturizing molecules like squalene, working synergistically to improve skin elasticity and texture.\n\nBakuchiol can be combined with Vanilla tahitensis extract to inhibit skin photoaging and cause improvement in skin barrier function and elasticity, increasing cellular turnover and improving signs of aging.10 A study by Bacqueville et al. used an in vitro skin model made of human dermal fibroblasts treated with the compounds alone or in combination, after which they were exposed to an acute dose of UVA. Exposure to UVA-induced significant morphological changes and increased IL-8 and p16 expression in the control model (no treatment), suggesting inflammation and senescence. Compared to control, treatment with either compound alone prevented actin network alteration and IL-8 upregulation, while protecting against IL-8 and p16 overexpression in combination. This combination was also formulated into serum and tested in participants who applied it twice daily for 56 days. These compounds can work synergistically to reduce ptosis and skin deformation and improve the radiance of naturally aged skin in women.\n\nA face serum containing bakuchiol, palmitoyl tripeptide-38, hydrolyzed hyaluronic acid and a polyherbal and vitamin blend was tested in 55 healthy adults. Daily use for 3 months indicated improvements in skin as seen in in vitro studies and clinical trials on healthy volunteers.60 Protection of skin structure was observed in vitro with reduced collagenase activity and significant free radical scavenging activity as observed through increased gene expression of dermal collagen, elastin and hyaluronic acid synthesis. In addition, substantial improvements in skin elasticity, hydration, roughness (fine lines and wrinkles), and brightness occurred during the trial.\n\nIt also works marvelously in combination with melatonin and ascorbyl tetraisopalmitate to cause significant clinical anti-aging effects when applied once daily,96 with a statistically significant decrease in wrinkles and redness, an increase in skin firmness and overall improvement in skin quality and complexion as well as hydration.96 A combination of bakuchiol, Ginkgo biloba extract and mannitol was also shown to improve the efficacy of adapalene treatment in patients suffering from Acne Vulgaris.9 It has also been found to work exceptionally well with salicylic acid in managing P. acnes.97\n\nOutside of the dermatocosmetic industry, the use of bakuchiol can also improve the efficacy of various non-cosmetic treatments. Bakuchiol has been found to work in combination with tumor necrosis factor (TNF)- related apoptosis-inducing ligand (TRAIL) to inhibit the growth of TRAIL sensitive (HCT116) and resistant (HT-29) colon cancer cell lines.98 Combination treatment of bakuchiol with TRAIL on these cell lines significantly upregulated the expression of TRAIL cell death receptors DR4 and DR5 in a dose-dependent manner, as well as the expression of the pro-apoptotic proteins PARP and the cleaved caspases 3, 8 and 9, while suppressing the expression of survival proteins such as cFLIP, survivin, XIAP and Bcl2. Pretreatment of cells with JNK inhibitor SP600125 and ROS scavenger N-acetylcysteine and the depletion of DR4 or DR5 by small interfering RNA reduced the bakuchiol-induced cell growth inhibition Bakuchiol assists with TRAIL-induced apoptosis via the ROS/JNK pathway.\n\nWhen used alone or in combination with Allium sativum, it also demonstrated potent antimicrobial properties, with bakuchiol and Allium sativum showing synergistic effects when used in combination.99\n\nBakuchiol is generally used in the cosmeceutical field as a more tolerable version of retinol, having retinol functionality through retinol-like regulation of gene expression. However, it may initially cause some redness and peeling in sensitive skin,4 although chances are rare due to the established anti-inflammatory nature of bakuchiol. As it may increase cellular turnover, sunscreen is recommended for use after applying bakuchiol-containing products to reduce damage by UV radiation. Retinol and its derivatives are generally discouraged for use during pregnancy. No studies have been done to evaluate use safety in pregnant women, so use should be carefully conducted. It is difficult to quantify the benefits and side effects of bakuchiol usage as most studies have been done in vitro, potentially introducing a risk of bias.\n\nIt should be mentioned that studies have found bakuchiol to be non-toxic to cell cultures even in high concentrations of up to 5000 uG/mL,100 however, this was only observed in in vitro studies, and further studies are required to determine dosage toxicity. Recent clinical reports have indicated that treatment with Psoraleae Fructus (PF), an essential source of Bakuchiol, is associated with an increased risk of liver injury.25,26,100–104 A study by Guo et al. (2021) indicated that bakuchiol, among other constituents of Psoraleae Fructus induced oxidative stress and mitochondrial damage-mediated apoptosis, alleviated when Ethanol extracts of PF were used. Bakuchiol may induce cholestatic hepatotoxicity as treatment with Bakuchiol reduces mRNA expression of CYP7A1, HMG-CoA reductase, PPARα, and SREBP-2.105\n\nCare should be taken when using bakuchiol in combination therapy because it may cause an increase in cytotoxic effects of bakuchiol, and extensive studies should be conducted to ensure that metabolic toxicity does not occur. Bakuchiol can induce nephrotoxicity when it is used in combination with other natural ingredients such as Glycyrrhetinic acid (GA) found in licorice, which inhibits the CYP450 isoenzymes (CYP3A4, CYP2C9, CYP1A2) involved in metabolic detoxification of bakuchiol.106 The presence of GA altered the toxicokinetics of bakuchiol in rats, increased the internal exposure, suppressed the elimination of the bakuchiol prototype, and therefore may have enhanced the renal nephrotoxicity.\n\n\nConclusions\n\nThis review focused on the pharmacological benefits of the compound bakuchiol, traditionally isolated from the bakuchi plant and used in traditional medicine for centuries. Recent studies have highlighted its vital role in controlling several activities that lead to health depreciation and the onset of various non-communicable diseases. Additionally, bakuchiol has shown potent antimicrobial and antiviral responses against various pathogens in multiple studies, which allows for the development of potential novel cures and preventive strategies.\n\nMoreover, extensive research is required on the non-toxic extraction of the compounds, fully realizing its pharmacological and biochemical modes of action, and planning for sustainable ways of growing its sources to meet the increasing demands of the pharmaceutical industries. Further studies should also be done to evaluate the long-term effects of prolonged bakuchiol consumption, possibly via retrospective analyses, which have not been done yet.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nReferences\n\nImproving access to essential medicines: World Health Organization.2007 [cited 2022 Jun 3].Reference Source\n\nvan Wyk AS , Prinsloo G: Medicinal plant harvesting, sustainability and cultivation in South Africa. Biol. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTruong CS, Seo E, Jun HS: Psoralea corylifolia L. Seed Extract Attenuates Methylglyoxal-Induced Insulin Resistance by Inhibition of Advanced Glycation End Product Formation. Oxidative Med. Cell. Longev. 2019; 2019: 1–14. Publisher Full Text\n\nKim YJ, Lim HS, Lee J, et al.: Quantitative Analysis of Psoralea corylifolia Linne and its Neuroprotective and Anti-Neuroinflammatory Effects in HT22 Hippocampal Cells and BV-2 Microglia. Molecules. 2016 Aug; 21(8): 1076. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZhou P, Hua F, Wang X, et al.: Therapeutic potential of IKK-β inhibitors from natural phenolics for inflammation in cardiovascular diseases. Inflammopharmacology. 2020 Feb; 28(1): 19–37. PubMed Abstract \| Publisher Full Text\n\nHoller JG, Christensen SB, Slotved HC, et al.: Novel inhibitory activity of the Staphylococcus aureus NorA efflux pump by a kaempferol rhamnoside isolated from Persea lingue Nees. J. Antimicrob. Chemother. 2012 May; 67(5): 1138–1144. PubMed Abstract \| Publisher Full Text Reference Source\n\nNawrot R, Barylski J, Nowicki G, et al.: Plant antimicrobial peptides. Folia Microbiol. (Praha). 2014 Oct; 59(3): 181–196. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMahizan NA, Yang SK, Moo CL, et al.: Terpene Derivatives as a Potential Agent against Antimicrobial Resistance (AMR) Pathogens. Molecules. 2019 Jul; 24(14): 2631. PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source\n\nHsu PJ, Miller JS, Berger JM: Bakuchiol, an antibacterial component of Psoralidium tenuiflorum. Nat. Prod. Res. 2009; 23(8): 781–788. PubMed Abstract \| Publisher Full Text\n\nVahabi S, Najafi E, Alizadeh S: In vitro antimicrobial effects of some herbal essences against oral pathogens. J. Med. Plant Res. 2011 Sep; 5(19): 4870–4878. Reference Source\n\nKim KA, Shim SH, Ahn HR, et al.: Protective effects of the compounds isolated from the seed of Psoralea corylifolia on oxidative stress-induced retinal damage. Toxicol. Appl. Pharmacol. 2013 Jun; 269(2): 109–120. PubMed Abstract \| Publisher Full Text\n\nStan D, Enciu AM, Mateescu AL, et al.: Natural Compounds With Antimicrobial and Antiviral Effect and Nanocarriers Used for Their Transportation. Front. Pharmacol. 2021 Sep; 12. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCowan MM: Plant Products as Antimicrobial Agents. Clin. Microbiol. Rev. 1999; 12(4): 582. Free Full Text\n\nKhameneh B, Iranshahy M, Soheili V, et al.: Review on plant antimicrobials: a mechanistic viewpoint. Antimicrob Resist. Infect. Control. 2019 Jul; 8(1). Publisher Full Text Reference Source/\n\nGonelimali FD, Lin J, Miao W, et al.: Antimicrobial Properties and Mechanism of Action of Some Plant Extracts Against Food Pathogens and Spoilage Microorganisms. Front. 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Alternat. Med. 2015; 2015: 1–7. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDhaliwal S, Rybak I, Ellis SR, et al.: Prospective, randomized, double-blind assessment of topical bakuchiol and retinol for facial photoageing. Br. J. Dermatol. 2019 Feb; 180(2): 289–296. PubMed Abstract \| Publisher Full Text\n\nWest BJ, Alabi I, Deng S: A Face Serum Containing Bakuchiol, Palmitoyl Tripeptide-38, Hydrolyzed Hyaluronic Acid and a Polyherbal and Vitamin Blend Improves Skin Quality in Human Volunteers and Protects Skin Structure In vitro. Preprints. 2021 Jun.Reference Source1\n\nYu Q, Zou HM, Wang S, et al.: Regulative effect of bakuchiol on ESF-1 cells anti-aging gene. J. Chinese Med. Mater. 2014 Apr; 37(4): 632–635.\n\nDraelos ZD, Gunt H, Zeichner J, et al.: Clinical Evaluation of a Nature-Based Bakuchiol Anti-Aging Moisturizer for Sensitive Skin. J Drugs Dermatol. 2020 Dec; 19(12): 1181–1183. Publisher Full Text Reference Source\n\nWoolery-Lloyd HC, Keri J, Doig S: Retinoids and Azelaic Acid to Treat Acne and Hyperpigmentation in Skin of Color. J. Drugs Dermatol. 2013 Apr; 12(4): 434–437. Reference Source\n\nGimeno A, Zaragozá R, Vivó-Sesé I, et al.: Retinol, at concentrations greater than the physiological limit, induces oxidative stress and apoptosis in human dermal fibroblasts. Exp. Dermatol. 2004 Jan; 13(1): 45–54. PubMed Abstract \| Publisher Full Text\n\nSun NJ, Woo SH, Cassady JM, et al.: DNA polymerase and topoisomerase II inhibitors from Psoralea corylifolia. J. Nat. Prod. 1998 Mar; 61(3): 362–366. Publisher Full Text Reference Source\n\nLi L, Chen X, Liu CC, et al.: Phytoestrogen Bakuchiol Exhibits in vitro and in vivo Anti-breast Cancer Effects by Inducing S Phase Arrest and Apoptosis. Front. Pharmacol. 2016; 7(MAY): 128. /pmc/articles/PMC4877368. Publisher Full Text\n\nMajeed R, Reddy MV, Chinthakindi PK, et al.: Bakuchiol derivatives as novel and potent cytotoxic agents: A report. Eur. J. Med. Chem. 2012 Mar 1; 49: 55–67. Publisher Full Text\n\nMiao L, Yun X, Tao R, et al.: Bakuchiol exhibits anti-metastasis activity through NF-κB cross-talk signaling with AR and ERβ in androgen-independent prostate cancer cells PC-3. J. Pharmacol. Sci. 2018 Sep; 138(1): 1–8. PubMed Abstract \| Publisher Full Text\n\nLv L, Liu B: Anti-tumor effects of bakuchiol on human gastric carcinoma cell lines are mediated through PI3K/AKT and MAPK signaling pathways. Mol. Med. Rep. 2017 Dec; 16(6): 8977–8982. PubMed Abstract \| Publisher Full Text\n\nLim SH, Ha TY, Kim SR, et al.: Ethanol extract of Psoralea corylifolia L. and its main constituent, bakuchiol, reduce bone loss in ovariectomised Sprague-Dawley rats. Br. J. Nutr. 2009; 101(7): 1031–1039. PubMed Abstract \| Publisher Full Text\n\nAlam F, Khan GN, Asad MHH: Bin. Psoralea corylifolia L: Ethnobotanical, biological, and chemical aspects: A review. Phyther Res. 2018 Apr; 32(4): 597–615. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nXin Z, Wu X, Yu Z, et al.: Mechanisms explaining the efficacy of psoralidin in cancer and osteoporosis, a review. Pharmacol. Res. 2019 Sep; 147: 104334. PubMed Abstract \| Publisher Full Text\n\nMa W, Guo W, Shang F, et al.: Bakuchiol Alleviates Hyperglycemia-Induced Diabetic Cardiomyopathy by Reducing Myocardial Oxidative Stress via Activating the SIRT1/Nrf2 Signaling Pathway. Oxidative Med. Cell. Longev. 2020; 2020: 1–15.\n\nPark EJ, Zhao YZ, Kim YC, et al.: Protective effect of (S)-bakuchiol from Psoralea corylifolia on rat liver injury in vitro and in vivo. Planta Med. 2005 Jun; 71(6): 508–513. PubMed Abstract \| Publisher Full Text\n\nPark EJ, Zhao YZ, Kim YH, et al.: Acanthoic acid from Acanthopanax koreanum protects against liver injury induced by tert-butyl hydroperoxide or carbon tetrachloride in vitro and in vivo. Planta Med. 2004 Apr; 70(4): 321–327. PubMed Abstract \| Publisher Full Text\n\nPark EJ, Zhao YZ, Kim YC, et al.: Bakuchiol-induced caspase-3-dependent apoptosis occurs through c-Jun NH2-terminal kinase-mediated mitochondrial translocation of Bax in rat liver myofibroblasts. Eur. J. Pharmacol. 2007 Mar; 559(2–3): 115–123. PubMed Abstract \| Publisher Full Text\n\nWang J, Luo M, Shen J, et al.: Bakuchiol from Psoralea corylifolia L. Ameliorates acute kidney injury and improves survival in experimental polymicrobial sepsis. Int. Immunopharmacol. 2020 Dec; 89(Pt A): 107000. Publisher Full Text Reference Source\n\nWang Z, Gao L, Xiao L, et al.: Bakuchiol protects against pathological cardiac hypertrophy by blocking NF-$κ$B signaling pathway. Biosci. Rep. 2018 Oct; 38(5). Publisher Full Text Reference Source\n\nLiu H, Guo W, Guo H, et al.: Bakuchiol Attenuates Oxidative Stress and Neuron Damage by Regulating Trx1/TXNIP and the Phosphorylation of AMPK After Subarachnoid Hemorrhage in Mice. Front. Pharmacol. 2020 May; 11: 712. Publisher Full Text \| Free Full Text\n\nFeng J, Yang Y, Zhou Y, et al.: Bakuchiol attenuates myocardial ischemia reperfusion injury by maintaining mitochondrial function: the role of silent information regulator 1. Apoptosis. 2016 May; 21(5): 532–545. PubMed Abstract \| Publisher Full Text\n\nYang Y, Duan W, Lin Y, et al.: SIRT1 activation by curcumin pretreatment attenuates mitochondrial oxidative damage induced by myocardial ischemia reperfusion injury. Free Radic. Biol. Med. 2013; 65: 667–679. PubMed Abstract \| Publisher Full Text\n\nYang Y, Duan W, Jin Z, et al.: JAK2/STAT3 activation by melatonin attenuates the mitochondrial oxidative damage induced by myocardial ischemia/reperfusion injury. J. Pineal Res. 2013 Oct; 55(3): 275–286. Publisher Full Text Reference Source\n\nYu L, Li Q, Yu B, et al.: Berberine Attenuates Myocardial Ischemia/Reperfusion Injury by Reducing Oxidative Stress and Inflammation Response: Role of Silent Information Regulator 1. Oxidative Med. Cell. Longev. 2016; 2016.Reference Source\n\nWu B, Yu Feng J, Ming Yu L, et al.: Icariin protects cardiomyocytes against ischaemia/reperfusion injury by attenuating sirtuin 1-dependent mitochondrial oxidative damage. Br. J. Pharmacol. 2018 Nov; 175(21): 4137–4153. Publisher Full Text Reference Source\n\nShoji M, Esumi T, Tanaka N, et al.: Organic synthesis and anti-influenza A virus activity of cyclobakuchiols A, B, C, and D. PLoS One. 2021 Mar; 16(3): e0248960. Publisher Full Text Reference Source\n\nZhang D, Hamdoun S, Chen R, et al.: Identification of natural compounds as SARS-CoV-2 entry inhibitors by molecular docking-based virtual screening with bio-layer interferometry. Pharmacol. Res. 2021 Oct; 172: 105820. Publisher Full Text \| Free Full Text\n\nTaubenberger JK, Morens DM: 1918 Influenza: the Mother of All Pandemics. Emerg. Infect. Dis. 2006 Jan; 12(1): 15–22. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHossain A, Nasrullah SM, Tasnim Z, et al.: Seroprevalence of SARS-CoV-2 IgG antibodies among health care workers prior to vaccine administration in Europe, the USA and East Asia: a systematic review and meta-analysis. EClinicalMedicine. 2021 Mar; 33: 100770. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCOVID Live Update: 287,054,304 Cases and 5,449,037 Deaths from the Coronavirus - Worldometer.2021.\n\nDavidson AM, Wysocki J, Batlle D: Interaction of SARS-CoV-2 and Other Coronavirus With ACE (Angiotensin-Converting Enzyme)-2 as Their Main Receptor: Therapeutic Implications. Hypertension. 2020 Sep; 76: 1339–1349. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAli SI, Sheikh WM, Rather MA, et al.: Medicinal plants: Treasure for antiviral drug discovery. Phyther. Res. 2021 Jul; 35(7): 3447–3483. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBhuiyan FR, Howlader S, Raihan T, et al.: Plants Metabolites: Possibility of Natural Therapeutics Against the COVID-19 Pandemic. Front. Med. 2020 Aug; 7. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAl-Salihi SAA, Alberti F: Naturally Occurring Terpenes: A Promising Class of Organic Molecules to Address Influenza Pandemics. Nat. Products Bioprospect. 2021 Aug; 11(4): 405–419. Publisher Full Text \| Free Full Text\n\nMani JS, Johnson JB, Steel JC, et al.: Natural product-derived phytochemicals as potential agents against coronaviruses: A review. Virus Res. 2020 Jul; 284: 197989. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChen RH, Yang LJ, Hamdoun S, et al.: 1,2,3,4,6-Pentagalloyl Glucose, a RBD-ACE2 Binding Inhibitor to Prevent SARS-CoV-2 Infection. Front. Pharmacol. 2021 Mar; 12: 634176. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGoldberg DJ, Robinson DM, Granger C: Clinical evidence of the efficacy and safety of a new 3-in-1 anti-aging topical night serum-in-oil containing melatonin, bakuchiol, and ascorbyl tetraisopalmitate: 103 females treated from 28 to 84 days. J. Cosmet. Dermatol. 2019 Jun; 18(3): 806–814. PubMed Abstract \| Publisher Full Text\n\nChaudhuri RK, Marchio F: Bakuchiol in the Management of Acne-affected Skin. Cosmet Toilet. 2011 Jul; 126: 502–510.\n\nPark MH, Kim JH, Chung YH, et al.: Bakuchiol sensitizes cancer cells to TRAIL through ROS- and JNK-mediated upregulation of death receptors and downregulation of survival proteins. Biochem. Biophys. Res. Commun. 2016 Apr; 473(2): 586–592. PubMed Abstract \| Publisher Full Text\n\nSingh B, Sharma RA: Plant terpenes: defense responses, phylogenetic analysis, regulation and clinical applications. 3. Biotech. 2015 Apr; 5(2): 129–151. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLi YF, Cheng GF: Pharmaceutical Care for Patients with Liver Damage Induced by Traditional Chinese Medicine Fructus Psoraleae by Clinical Pharmacists. Eval. Anal. Drug-Use Hosp. Chin. 2018; 16(4): 571–573.\n\nTeschke R, Bahre R: Severe hepatotoxicity by Indian Ayurvedic herbal products: A structured causality assessment. Ann. Hepatol. 2009 Jul; 8(3): 258–266. PubMed Abstract \| Publisher Full Text\n\nSmith DA, MacDonald S: A rare case of acute hepatitis induced by use of Babchi seeds as an Ayurvedic remedy for vitiligo. BMJ Case Rep. 2014 Aug; 2014: bcr2013200958. Publisher Full Text Reference Source\n\nZhang LL, Huang JH: A Case of Drug-Induced Liver Injury Caused by the Combined Medication of Buguzhi Granules and Qubaibabuqi Tablets. Chin. J. Drug Appl. Monit. 2018; 15(4): 246–249.\n\nLi A, Gao M, Zhao N, et al.: Acute liver failure associated with Fructus Psoraleae: a case report and literature review. BMC Complement. Altern. Med. 2019 Apr; 19(1): 84. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGuo Z, Li P, Wang C, et al.: Five Constituents Contributed to the Psoraleae Fructus-Induced Hepatotoxicity via Mitochondrial Dysfunction and Apoptosis. Front. Pharmacol. 2021 Dec; 12. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLi A, Ma N, Zhao Z, et al.: Glycyrrhetinic acid might increase the nephrotoxicity of bakuchiol by inhibiting cytochrome P450 isoenzymes. PeerJ. 2016; 4(11): e2723. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "174263", "date": "30 Jun 2023", "name": "Amit Baran Sharangi", "expertise": [ "Reviewer Expertise Herbs", "spices", "medicinal and aromatic crops" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe article entitled \"Bakuchiol and its pharmacological benefits\" by Nizam et al., is a routine exercise wherein a systematic review approach summarized, analyzed, interpreted and concluded about the therapeutical values of the compound \"bachuchiol\" from the leaves and seeds of Psoralea corylifolia medicinal plants. While the potential of the same is pharmacologically great, the following are some of my observations on the article which needs immediate attention by the authors and rigorous revision before indexing:\nTo apprise the readers about the origin of the compound \"Bakuchiol\".....ie., whether it is synthetic or natural (plant based).....some appropriate words may be given in the title itself: -it may be like Bakuchiol from Psoralea corylifolia L. -it may be like Natural bakuchiol, etc\nThe above is for clarity in understanding only.\n\nIn the \"Introduction\", placement of Ref 17 is probably at a wrong position.\n\nIn the \"Results\", in the first subhead, the word 'material' has been repeated. Here, the last sentence may be separated into two.\n\nTherapeutic applications of bakuchiol: This portion seriously lacks in one or two summarized table containing some global research-based interesting information on In vivo Test, In vitro Test, Plant parts used, Visible / projected Effects, Model used(if any), References, etc\n\nJust before \"Side effects\", was it necessary to hyperlink the highlighted words?\n\nAfter \"Side effects\", some discussion was necessary on the Phytochemical screening and toxicity study of this very medicinal plants or the compound bakuchiol derived from it .....in a different subtitle along with contraindication, if any and of course, supplemented with a well organised table with convincing references was necessary [LD50 (mg/kg), Phytochemical constituents, References in the last column of the table].\n\nInstead of Conclusion, it may be replaced with Conclusion and Future research. In that case, there should be some restructuring of the text with inclusion of some promising upcoming areas of futuristic research in this direction.\n\nIs the topic of the review discussed comprehensively in the context of the current literature? Partly\n\nAre all factual statements correct and adequately supported by citations? Yes\n\nIs the review written in accessible language? Yes\n\nAre the conclusions drawn appropriate in the context of the current research literature? No", "responses": [ { "c_id": "10486", "date": "16 Nov 2023", "name": "Md. Kamrul Hasan", "role": "Author Response", "response": "The article entitled \"Bakuchiol and its pharmacological benefits\" by Nizam et al., is a routine exercise wherein a systematic review approach summarized, analyzed, interpreted and concluded about the therapeutical values of the compound \"bachuchiol\" from the leaves and seeds of Psoralea corylifolia medicinal plants. While the potential of the same is pharmacologically great, the following are some of my observations on the article which needs immediate attention by the authors and rigorous revision before indexing: To apprise the readers about the origin of the compound \"Bakuchiol\".....ie., whether it is synthetic or natural (plant based).....some appropriate words may be given in the title itself: -it may be like Bakuchiol from Psoralea corylifolia L. -it may be like Natural bakuchiol, etc. Authors’ response: Thank you. The revised version addressed this issue and fixed. The above is for clarity in understanding only. In the \"Introduction\", placement of Ref 17 is probably at a wrong position. Authors’ response: Addressed and fixed. Thank you. In the \"Results\", in the first subhead, the word 'material' has been repeated. Here, the last sentence may be separated into two. Authors’ response: Thank you. Addressed and fixed. Therapeutic applications of bakuchiol: This portion seriously lacks in one or two summarized table containing some global research-based interesting information on In vivo Test, In vitro Test, Plant parts used, Visible / projected Effects, Model used(if any), References, etc. Authors’ response: Thank you so much for the suggestion. The authors revised the manuscript as suggested. Just before \"Side effects\", was it necessary to hyperlink the highlighted words? Authors’ response: Thank you. Hyperlink removed. After \"Side effects\", some discussion was necessary on the Phytochemical screening and toxicity study of this very medicinal plants or the compound bakuchiol derived from it .....in a different subtitle along with contraindication, if any and of course, supplemented with a well organised table with convincing references was necessary [LD50 (mg/kg), Phytochemical constituents, References in the last column of the table]. Authors’ response: Thank you so much. The authors revised the manuscript by adding a table as suggested. Instead of Conclusion, it may be replaced with Conclusion and Future research. In that case, there should be some restructuring of the text with inclusion of some promising upcoming areas of futuristic research in this direction. Authors’ response: Thank you. Addressed and revised." } ] }, { "id": "177831", "date": "31 Aug 2023", "name": "Naoufal El Hachlafi", "expertise": [ "Reviewer Expertise Pharmacology", "essential oils", "bioactive compounds", "antimicrobial", "antioxidant" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe manuscript entitled: \"Bakuchiol and its pharmacological benefits\", presents interesting and valuable work, which is within the scope of the journal F1000Research. It is well-written. However, some minor corrections are needed to make this paper accepted for indexing. I encourage the authors to consider making the necessary changes.\nGeneral Considerations\nThe authors should carefully proof-read the entire manuscript to minimize typographical errors, especially with spellings, punctuation, unnecessary capitalizations, spaces, and units, as well as to ensure uniform expression of various special characters and abbreviations, terms, and phrases.\nResults and discussion The description of the results is clear; however, it would be helpful to provide more details in tables taken into account different in vitro and in vivo clinical studies carried out in this subject.\nConclusion\nConclusion should be improved by adding concluding remarks and future direction for further investigation in the field.\nReferences\nEnsure all references are relevant and up-to-date. It's essential to cite the most recent and pertinent research in the field to demonstrate a sound understanding of the current state of knowledge.\n\nIs the topic of the review discussed comprehensively in the context of the current literature? Yes\n\nAre all factual statements correct and adequately supported by citations? Yes\n\nIs the review written in accessible language? Yes\n\nAre the conclusions drawn appropriate in the context of the current research literature? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-29
https://f1000research.com/articles/12-1144/v1	13 Sep 23	{ "type": "Research Article", "title": "The proximate composition, amino acid profile, fatty acid content, and mineral content of scale flour from three fish species as potential feeds for fish fry", "authors": [ "Hafrijal Syandri", "Azrita Azrita", "Ainul Mardiah", "Netti Aryani", "Andarini Diharmi", "Azrita Azrita", "Ainul Mardiah", "Netti Aryani", "Andarini Diharmi" ], "abstract": "Background: Fish scale waste is highly valued both as a food additive and as a functional food ingredient. This study aimed to analyse the chemical composition, fatty acid profile, and mineral content in fish scale flour of Osphronemus (O) goramy, Cyprinus (C) carpio, and Oreochromis (O) niloticus as potential feed for fish fry. Methods: Fish scales were cleaned with 10% w/v NaCl solution at a ratio of 1:10 (w/w) for 24 hours at 4 °C. Agitation was used every eight hours to remove excess protein. Fish scales were evenly arranged in a cooker and cooked at 121 °C for 10 minutes with 15 psi pressure. After cooking, 100 grams of wet fish scales was dried at 50 °C for four hours. Dried fish scales were processed into flour for analysis of proximatel composition, amino acid content, fatty acid content, and mineral content. Results: The examined fish scale flour from three species displayed significant variations in chemical components, amino acids, and minerals (p<0.01). Crude protein content spanned 49.52% to 72.94%, and fat content ranged from 0.11% to 0.23%. Magnesium levels varied between 767.82 mg/kg and 816.50 mg/kg, calcium content ranged from 3.54 to 12.16 mg/kg, iron content was within 40.46 to 44.10 mg/kg, and zinc content ranged from 45.80 to 139.19 mg/kg. Predominantly, glycine emerged as the main free amino acid (FAA), varying from 13.70% to 16.08%, while histidine had the lowest content, at 0.39% to 0.71%. Conversely, fatty acid content was lowest among the species, ranging from 6.73% to 9.48%. Conclusions: Scale flour from three farmed fish types showed potential for fish fry feed due to its chemical composition and amino acid and mineral contents. To enhance the essential fatty acid content, enriching the flour with oils containing eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), and α-linolenic acid (ALA) is essential", "keywords": [ "Fish scale flour", "chemical composition", "amino acids", "mineral content", "fatty acid profile" ], "content": "Introduction\n\nAccording to the Food and Agriculture Organization of the United Nations (FAO), global fish production in 2018 reached approximately 179 million tonnes, with aquaculture contributing 46% of the total production.1 In Indonesia, the total aquaculture production was recorded at 16,032,122 metric tonnes (mt). Of this, 3,374,924 mt (21.05%) originated freshwater aquaculture production; 9,884,670 mt (61.65%) from marine water aquaculture production, including seagrass; and 2,772,568 mt (17.29%) from brackish water aquaculture production (CDSI; Central Data System Information).2 Approximately 10% of global fish production is currently discarded, while byproducts from fisheries constitute 70% of the total weight of fish production. Among these byproducts, fish bones and scales constitute 14 to 20% of the total weight, and these materials are also discarded.3\n\nThis significant quantity of byproducts is occasionally utilized as animal feed, fishmeal, oil, or plant fertilizer, but in most instances, it is discarded.4,5 In recent years, there has been a growing recognition of environmental sustainability and a heightened emphasis on harnessing the value of resources within green and blue economies.6,7 As a result, fish byproducts, including fish scale waste,3,8 have gradually been applied as raw materials for human consumption.9\n\nFish scales contain approximately 41-45% organic components, such as collagen, fat, lecithin, sclerotin, and vitamins, and 38-46% inorganic components and mineral elements, including magnesium, iron, zinc, calcium, and vitamins.10 Furthermore, fish scales possess antioxidant and antihypertensive properties.11 These components are also crucial for the growth and survival of fish fry. In recent years, fish fry feed has primarily been live feed, and expensive artificial feed is a bottleneck in aquaculture12,13 Therefore, exploring alternative ingredients for fish fry feed such as fish scale flour is of economic interest.\n\nA significant quantity of fish scale waste is readily accessible in the Indonesian market, encompassing fish scale waste derived from the death of farmed fish in Lake Maninjau.14 Regrettably, in the past decade, this waste has remained untapped as both a food source and an ingredient for fish fry feed. This research aims to assess the proximate, amino acid, fatty acids and mineral contents in freshwater fish scale flour, specifically flour from the scales of giant gourami (Osphronemus (O) goramy), carp (Cyprinus (C) carpio), and tilapia (Oreochromis (O) niloticus), with the potential to be utilized as feed ingredients for fish fry.\n\n\nMethods\n\nThe research was approved by the Research and Community Service Ethics Committee at Universitas Bung Hatta with an approval letter No.057a/LPPM/Hatta/VI-2023 dated June 23, 2023. Experiments were carried out in accordance with the guidelines outlined in the Standard Operating Procedure of Laboratory Aquaculture at Universitas Bung Hatta.\n\nTen O. goramy, C. carpio, and O. niloticus fish were obtained from a local fish market in Lake Maninjau, Indonesia. The fresh fish were carefully stored on ice and promptly transported to the laboratory.\n\nUpon reaching the laboratory, the fish were individually weighed (TW) using AD-600i scales with a precision of 0.001 grams and measured to their standard length (SL) and maximum height (H), with distance measured from the mouth to the end of the upper lobe of the caudal fin and height measured vertically, excluding the fins. Standard length and height were assessed using a meter ruler with an accuracy of 1 millimeter. The condition factor (CF) was calculated using the formula CF = (TW/SL3) × 100.\n\nUpon arrival at the laboratory, the fish scales from O. goramy, C. carpio, and O. niloticus were collected from each fish for further processing. Fish scales were collected with a stainless steel fish scaler cleaner of 17.5 cm × 3.5 cm × 1.5 cm.\n\nEvery 1,000 grams of wet scales of O. goramy, C. carpio, and O. niloticus were washed thoroughly to obtain 200 grams of dry scales. The fish scales were washed in a three-litres plastic jar with 10% w/v NaCl solution with a solution ratio of 1:10 (w/w). The washing procedure was conducted for a duration of 24 hours at a temperature of 4 °C. The washing process was repeated every eight hours to further improve the effectiveness of protein removal. This frequent repetition helped ensure that any remaining unnecessary proteins on the fish scales were thoroughly removed. By repeating the washing process at regular intervals, the purity of the fish scales was enhanced, preparing them for further processing (Figure 1).\n\nFurthermore, scales of O. goramy, C. carpio, and O. niloticus were washed three times in low mineral-content water at room temperature for 10 minutes and then drained. Subsequently, the scales were evenly arranged in a cooker that was equipped with a pressure control button and cooking time settings (Model: Classic Pressure Cooker, with a ø 20 cm, 5.5-litre capacity, named Culinart, Made in China). The heating process was carried out using a cooker until the temperature reached 121 °C with a pressure of 15 psi, as indicated by the temperature and pressure panel. At this point, the timer was set for 10 minutes. The cooking time was calculated as the time between when the pressure in the cooker reached 15 psi and that when the heat source for cooking was turned off.\n\nA total of 200 grams of scales from each species (O. goramy, C. carpio, and O. niloticus scales; wet weight) was dried using a 28 L stainless steel black digimatic oven tester at 50 °C for four hours until the moisture content reached 10%. The dried fish scales were then processed into flour using a Miller Powder Grinder with a 100-gram capacity. The resulting flour was then sieved using a mesh size of 60 μm to analyse the proximate composition and amino acid, fatty acid, and mineral contents (Figure 2).\n\nThe proximate composition of the fish scale samples was analysed using standard AOAC methods.15 The samples were dried at 105 °C until a constant weight was achieved. The crude protein content was analysed using the standard Kjeldahl method, calculated as N × 6.25. Crude lipids were analysed using the Soxhlet method with ether extraction. The ash content was determined by incinerating the samples at 550 °C for 16 hours. Gross energy was measured using a bomb calorimeter.\n\nTotal carbohydrates were determined by subtracting the sum of % crude protein (CP), % fat (F) and % ash contents (A) from 10016 by using the following equation: % Total carbohydrates = 100 – (CP + F + A). The gross energy value of each sample was determined by augmenting the percentage of crude protein (CP), fat (F), and total carbohydrate (C) contents with their respective energy values of 4, 9, and 4 kcal per 100 g of scale flour, respectively, to obtain the caloric values of the samples by using the following equation= (4CP + 9F + 4C)kcal/100 g weight.\n\nThe methods described by Ref. 17 were employed for amino acid analysis. The amino acid composition was determined using a high-performance liquid chromatography (HPLC) system, which consisted of a Waters 1525 binary HPLC pump, 717 autosamplers (Waters®), and Waters 2475 multi λ fluorescence detector optics (with wavelengths set at 250 nm for excitation and 395 nm for emission). The samples were hydrolysed in triplicate using 6 N hydrochloric acid for 24 hours at 11 °C.\n\nFor the analysis of mineral content (Na, Mg, Ca, K, P, Fe, and Zn), the ashed GCTS sample was dissolved in 1 ml of hydrochloric acid (35% v/v Suprapur® Merck). Subsequently, the sample was filtered using cellulose filter paper (Watchman No 1, International Ltd; Maidstone, UK) and appropriately diluted for each elemental mineral. Phosphorus (P) levels were analysed using a Perkin-Elmer AA spectrophotometer mod 3110 (Norwalk, CT, USA).\n\nThe fatty acid composition of fish scale flours was examined through gas chromatography-mass spectrometry (GC-MS) analysis. The total lipid extraction followed a modification of the method by Folch et al. (1957), as detailed by Rajion,18 employing a chloroform: methanol (2.1, v/v) solvent system. Transmethylation was carried out with 14% methanolic boron trifluoride.\n\nData analysis was conducted using Statistical Package for the Social Sciences (SPSS) 16.0 software (SPSS; Chicago, IL). The homogeneity of the data was assessed using the Levine test. One-way ANOVA was performed to determine the proximate and amino acid composition parameters and the mineral content for each fish scale flour of the three species. Post hoc analysis was carried out using Duncan’s multiple-range test.19 The results are reported as the mean values ± standard errors for each parameter.\n\n\nResults and discussion\n\nTable 1 displays the average standard length, wet weight, height, condition factor, and chemical composition of three fish species found in Lake Maninjau. Statistically significant variations were noted in the standard length, wet weight, height, and condition factor of the three examined fish species (p < 0.05; Table 1).46\n\n* p < 0.001; ns: non-significant.\n\nIn general, there was a significant difference (p < 0.05; Table 1) in the proximate content of fish scale flour between the three farmed fish species in Lake Maninjau. The water content, crude protein content, fat content, and energy values were higher in C. carpio scale flour than in O. goramy and O. niloticus scale flours (Table 1).\n\nThe highest protein content was recorded in the fish scale flour of C. carpio, and the content did not differ by more than 23% between the three fish groups. Huang et al.20 reported that tilapia fish scale flour contained 49.42% protein, 0.02% lipid, 45.18% ash, and 5.38% carbohydrates on a dry weight basis. Similarly, the protein content of spotted golden goatfish (Parupeneus heptacanthus) was 45.2%.21 On the other hand, protein valuation from demineralized fish scale gelatine and nondemineralized gelatine displayed protein purities of 57.19 g/100 g and 43.37 g/100 g, respectively.3\n\nHigher fat levels of C. carpio scale flour (0.23%) than in O. goramy (0.13%) and O. niloticus (0.11%) scale flour have also been observed for Labeo rohita22 and other species.23,24 This result could be due to various factors, including availability and dietary protein intake, fish size and age, and fish scale type.20,22\n\nFurthermore, there were significant differences in the mineral content of fish scale flour between species (p <0.05; Table 1). The magnesium, potassium, and iron levels were higher in the O. goramy fish scale flour than in the other scale flour samples. At the same time, the sodium, calcium, and phosphorous contents were higher in the O. niloticus fish scale flour. Moreover, zinc levels were higher in C. carpio fish scale flour (Table 1) than in the other scale flour samples. In general, the levels of the minerals analysed in this study are in alignment with the results of previous studies.25,26\n\nThe flour derived from fish scales of three farmed fish species exhibited a high mineral content, making it a potentially suitable choice for utilization as feed for fish fry. The inclusion of these minerals in fish feed is crucial because they serve as essential nutrients for the nourishment of fingerlings. As stated by Nagappan et al.,27 fibre, minerals, and vitamins are essential, albeit minimal, requirements for optimal fish growth performance. In the context of fish scale flour, it was observed that all three species contained elevated mineral levels, implying that the scale flour offers enhanced support for the growth of fish fry. Nevertheless, it is important to highlight that despite the relatively high mineral content in the feed, there was no significant impact observed on the development of experimental animals, as noted by Dominquez et al.28 and Wang et al.29\n\nThe FAA profiles for the scale flour of the three fish species are presented in Table 2. Statistically significant differences were recorded in the FAA for the scales of the three fish species studied (p < 0.05; Table 2). The three species of farmed fish showed higher levels of aspartic acid, glycine, and alanine and lower levels of serine, histidine, methionine, and isoleucine (Table 2). C. carpio scale flour of showed the highest total FAA content (62.74%) compared to that of farmed fish (O. goramy; 48.31% and O. niloticus; 41.58%). The differences in the FAA profile could be related to different aspects, such as fish species, wild or farmed fish origin, diet composition, feeding habits, animal size, and age.30–33\n\n* p < 0.001.\n\nIn all samples, glycine was the most abundant FAA, which is in accordance with other studies on fish scale collagen in tilapia, Oreochromis sp.,20 and the whole-body carcass of Hemibagrus nemurus34 and O. goramy.35 Glycine has been reported to be one of the essential components in the collagen molecule, helping maintain tissue strength and elasticity.36,37\n\nFollowing glycine, the most abundant FAAs were glutamic acid, alanine, and arginine. Furthermore, histidine, methionine, and isoleucine were present in all samples but in smaller quantities; these particular FAA are commonly found in greater proportions within aquatic organisms.38,39\n\nThe fatty acid composition in the fish scale flour of the three cultivated fish species is displayed in Table 3. This discovery is in line with data previously reported for Atlantic salmon (salmo salar) and Catla catla (Labeo catla),40,41 where the fatty acid content of both species is relatively low. Variations in the fatty acid composition were evident among the three cultured fish species, with statistically significant differences (p < 0.05) observed among the species. Nevertheless, only two out of the fourteen saturated fatty acids (SAFAs) were identified. In monounsaturated fatty acids (MUFAs), three out of eight were observed, whereas three of the ten polyunsaturated fatty acids (PUFAs) were found. This result is in contrast to the type of fatty acids detected in the carcasses of several species of fish.42–45\n\n*** p < 0.001.\n\nFish scale flour from O. niloticus exhibited the highest cumulative fatty acid content (9.48%) in comparison to farmed fish scale flour (C. carpio; 7.32% and O. goramy; 6.73%). These findings are very similar in structure to the results obtained from analysis of the composition of saturated fatty acids (SFAs), monounsaturated fatty acids (MUFAs), and polyunsaturated fatty acids (PUFAs) in the whole-body carcass of giant gourami and other fish species.30,32,43\n\nBased on the analysis of the composition of the fatty acids contained in the scale flour of three species of farmed fish, if it is intended as feed for fish fry, it is necessary to enrich this fish scale flour with compounds containing fatty acids. Some enrichment options to consider are fish oil, chia seed oil, flaxseed oil, and walnut oil. These four sources are rich in omega-3 fatty acids, especially eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), as well as alpha-linolenic acid (ALA).\n\n\nConclusion\n\nFish scale flour derived from O. goramy, C. carpio, and O. niloticus in the study region was identified as a valuable protein, amino acid, and mineral source. All fish scale flour samples across the three species contained amino acids and minerals but not fatty acids. Consequently, enriching fish scale flour with animal and plant oils rich in omega-3 fatty acids is vital. There is a lack of reliable data regarding the chemical composition, mineral, and fatty acid profiles of fish scale flours from the three local fish species in the study area. Therefore, the chemical, mineral, and fatty acid composition data presented in this study will be the groundwork for future research in fish scale flour chemistry, contributing to fish fry nutrition optimization.", "appendix": "Data availability\n\nFigshare: The Proximate Composition, Amino Acid Profile, Fatty Acid Content, and Mineral Content of Scale Flour from Three Fish Species as Potential Feeds for Fish Fry, https://doi.org/10.6084/m9.figshare.23954799. 46\n\nThis project contains the following underlying data:\n\n- Table 1. Raw biometric data of three species of farmed fish samples.\n\n- Table 2. Raw proximate composition data of fish scale flour from three Species (%W/W).\n\n- Table 3. Raw mineral content data of fish scale flour from three Species (mg/kg).\n\n- Table 4. Raw Amino Acid Composition Data of Fish Scale Flour from Three Species (%W/W).\n\n- Table 5. Raw fatty Acid Composition Data of Fish Scale Flour from Three Species (%W/W).\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nWe extend our gratitude to the Ministry of Education, Culture, Research, and Technology of the Republic of Indonesia for supporting this research.\n\n\nReferences\n\nFAO: The state of world fisheries and Aquaculture. Rome, Italy: Food and Agriculture Organization of the United Nations; 2020.\n\nCDSI: (Central Data System Information): Ministry of Marine and Fisheries Republic of Indonesia. 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Publisher Full Text" }	[ { "id": "208002", "date": "16 Oct 2023", "name": "Adhita Sri Prabakusuma", "expertise": [ "Reviewer Expertise Food safety and quality", "food science and technology", "food waste management", "and agri-food system" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe manuscript, entitled “The proximate composition, amino acid profile, fatty acid content, and mineral content of scale flour from three fish species as potential feeds for fish fry,” focuses on analyzing the chemical composition, fatty acid profile, and mineral content in fish scale flour of Osphronemus (O) goramy, Cyprinus (C) carpio, and Oreochromis (O) niloticus as potential feed for fish fry, provides insights into chemical properties influencing nutritional aspects in feed formulation for fish fry, and describes the enrichment of the fish scale flour with oils containing essential fatty acids. After careful review of the manuscript, I feel that it is an interesting study supported by a significant amount of data and contains an important topic within the scope of the Agriculture, Food, and Nutrition Gateway of F1000Research. However, there are some critical concerns that require attention to improve the quality of the manuscript before it is accepted for publication. The specific comments are as follows:\nPlease include a statement in the background section of the abstract about the potential of fish scale flour by-products as a potential source of farmed fish feed, not only as a food additive and a functional food ingredient, aligning it with the title and overall context of the paper.\n\nPlease include the analysis approaches for measuring tested parameters mentioned in the methods section of the abstract, as the authors outlined: proximate was analyzed using standard AOAC methods; amino acid composition was determined using high-performance liquid chromatography (HPLC); and fatty acid composition was examined through gas chromatography-mass spectrometry (GC-MS).\n\nPlease include the comparative study results of tested parameters (the proximate composition, amino acid profile, fatty acid content, and mineral content) of three fish species in the results section of the abstract.\n\nIn the introduction section, it is better to elaborate on the recent research on utilizing fish scales both for human consumption and fish feed formulation and to make a gap analysis by evaluating the effectiveness or performance of previous methods or results.\n\nIn the materials and methods section, put the relevant references to support the procedures for measuring the biometric properties of fish samples and preparing fish scales.\n\nSupport this statement “This frequent repetition helped ensure that any remaining unnecessary proteins on the fish scales were thoroughly removed,” with the relevant references.\n\nWhy did the authors select a temperature of 121 °C with a pressure of 15 psi to perform the heating process? Why not choose a higher temperature and more pressure for less than or within ten minutes?\n\nIn the results and discussion section, it is also better to elaborate on the recent research studies and compare those with the findings of this current study.\n\nIn all sub-sections, mention the function of chemical nutrition, amino acids, and fatty acids to support the growth of farmed fish, their effect on increasing the quality of fish carcasses, and their role in maintaining fish health.\n\nPlease double-check the use of English grammar in the manuscript.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10450", "date": "07 Nov 2023", "name": "Hafrijal Syandri", "role": "Author Response", "response": "Response to comments from Adhita Sri Prabakusuma Abstract: The author could only revise the introduction and method sections in the abstract due to the maximum abstract length of 300 words. We will change in the revised version. Background: Fish scale waste is highly valued as a functional food ingredient and a potential feed source for farmed fish. This study aimed to analyse the chemical composition, amino acids, fatty acids, and mineral content in fish scale flour of Osphronemus (O) goramy, Cyprinus (C) carpio, and Oreochromis (O) niloticus as potential feed for a fish fry. Methods: Fish scales were cleaned with 10% w/v NaCl solution at a ratio of 1:10 (w/w) for 24 hours at 4 °C. Agitation was used every eight hours to remove excess protein. Fish scales were evenly arranged in a cooker and cooked at 121 °C for 10 minutes with 15 psi pressure. After cooking, 100 grams of wet fish scales was dried at 50 °C for four hours. Dried fish scales were transformed into flour for proximate composition analysis via standard AOAC methods, amino acid and fatty acid assessment employing HPLC and GC-MS, while mineral content was determined using AAS. Results: Examined fish scale flour from three species showed significant variations in chemical components, amino acids, and minerals (p<0.01). Crude protein content ranged from 49.52% to 72.94%, while fat content ranged from 0.11% to 0.23%. Magnesium levels varied from 767.82 to 816.50 mg/kg, calcium content from 3.54 to 12.16 mg/kg, iron from 40.46 to 44.10 mg/kg, and zinc from 45.80 to 139.19 mg/kg. Predominantly, glycine was the main free amino acid (FAA), varying from 13.70% to 16.08%, while histidine had the lowest content, 0.39% to 0.71%. Conversely, fatty acid content was lowest among the species, ranging from 6.73% to 9.48%. Conclusion: Scale flour from three farmed fish types exhibits potential as fish fry feed, considering its chemical composition, amino acid, and mineral content. Enriching the flour with EPA, DHA, and ALA-containing oils is crucial to boost essential fatty acids. In the introductory part The author will add to the new revised version a sentence highlighted in yellow in paragraph 3 of the \"introduction\" section to explain recent research and gaps in evaluating the effectiveness or performance of previous methods or results. Fish scales contain approximately 41-45% organic components, such as collagen, fat, lecithin, sclerotin, and vitamins, and 38-46% inorganic components and mineral elements, including magnesium, iron, zinc, calcium, and vitamins10. Furthermore, fish scales possess antioxidant and antihypertensive properties11. Fish scales have been used as a culinary ingredient in baked bread 3, because they are a source of food that is rich in nutrients 5,7.These components are also crucial for the growth and survival of fish fry. In recent years, fish fry feed has primarily been live feed, and expensive artificial feed is a bottleneck in aquaculture12,13. Therefore, there is a technological gap in exploring alternative ingredients for fish fry feed, such as fish scale flour, which is a rich source of nutrients, has economic value, and provides an element of novelty in this study. Material and Methods Part In the revised version, in the materials and methods section, the author will include relevant references to support procedures for measuring biometry properties of fish samples and preparing fish scales. Upon reaching the laboratory, the fish were individually weighed (TW) using AD-600i scales with a precision of 0.001 grams and measured to their standard length (SL) and maximum height (H), with distance measured from the mouth to the end of the upper lobe of the caudal fin and width measured vertically, excluding the fins (Famoofo and Abdul, 2020). Standard length and width were assessed using a meter ruler with an accuracy of 1 millimeter. The condition factor (CF) was calculated using the formula CF = (TW/SL3) x 100 (Froese, 2006) This frequent repetition helped ensure that any remaining unnecessary proteins on the fish scales were thoroughly removed (Lou et al., 2020; Boronat et al., 2023). Why did the authors select a temperature of 121 °C with a pressure of 15 psi to perform the heating process? Why not choose a higher temperature and more pressure for less than or within ten minutes? The author ensures that at a temperature of 121°C with a pressure of 15 psi for 10 minutes, it is hoped that no damage will occur to the chemical composition of the fish scale flour to be analyzed. Discussion part In the new revised version, we agreed to add comparisons with current research findings in the discussion section. The author will add to the newly revised version information regarding the function of amino acids, fatty acids, and minerals in the context of increasing fish seed growth. English grammar in the manuscript Please double-check the use of English grammar in the manuscript: The language contained in this manuscript has been checked by the American Journal Experts (AJE) with the English version (United Kingdom English) and is accompanied by a certificate." } ] }, { "id": "212279", "date": "30 Oct 2023", "name": "Peter Vilhelm Skov", "expertise": [ "Reviewer Expertise Fish physiology", "energetics and nutrition" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nAbstract:\n\"Conversely, fatty acid content was lowest among the species, ranging from 6.73% to 9.48%.\" should be rephrased to \"Fatt acid content was low in all species examined, ranging from 6.73% to 9.48%.\"\n\"Conclusions: Scale flour from three farmed fish types showed potential for fish fry feed due to its chemical composition and amino acid and mineral contents. To enhance the essential fatty acid content, enriching the flour with oils containing eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), and α-linolenic acid (ALA) is essential\"\nI don't agree with this statement. An ideal ingredient for feed formulation need not contain FA (e.g. fishmeal is also low in FA. The conclusion must be that it is a suitable protein source fir fry feed. This should however be validated in a context of AA requirement. Is the AA content comparable to fish meal or another ideal protein source, and would it satisfy the AA requirements of farmed species.\nIntroduction:\nDo you means seagrass specifically, or seaweeds in general?\nMethods:\nCan you be more specific concerning \"low mineral content water?\"\n\"Mineral content analysis For the analysis of mineral content (Na, Mg, Ca, K, P, Fe, and Zn), the ashed GCTS sample was dissolved in 1 ml of hydrochloric acid (35% v/v Suprapur® Merck). Subsequently, the sample was filtered using cellulose filter paper (Watchman No 1, International Ltd; Maidstone, UK) and appropriately diluted for each elemental mineral. Phosphorus (P) levels were analysed using a Perkin-Elmer AA spectrophotometer mod 3110 (Norwalk, CT, USA).\"\nPhosphorous was analysed using the spectrophotometer, is there a reference for the method? How were the other minerals analysed following acid treatment? This information is missing.\nProximate composition and mineral content:\n\"The highest protein content was recorded in the fish scale flour of C. carpio, and the content did not differ by more than 23%\" - specify whether it is 23% in relative or absolute terms - insert \"absolute\" before content.\nConclusion:\nAs hinted at in the abstract, I would appreciate that as part of the assessment of the value of the protein, that you consider the AA composition relative to other protein sources used for fish feeds (e.g. fish meal, soy) or against the nutritional requirements of selected farmed species (e.g. tilapia, carp)\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10487", "date": "07 Nov 2023", "name": "Hafrijal Syandri", "role": "Author Response", "response": "Abstract: The authors agreed to change the sentence: \"Conversely, fatty acid content was lowest among the species, ranging from 6.73% to 9.48%, to Fatty acid content was low in all species examined, ranging from 6.73% to 9.48%. The author will add in the new version that scale flour from three types of cultivated fish has potential as fish seed feed because of its close composition, amino acid, and mineral content. However, this needs to be validated in the context of amino acid requirements. Is it comparable to fishmeal or other ideal protein sources that meet the amino acid requirements for cultured species. Introduction: Do you means seagrass specifically, or seaweeds in general? : The author will change seagrass to seaweed in general in new versions. Methods: Can you be more specific concerning \"low mineral content water?\". The author will added \"low mineral-content water (with a TDS of around 100 mg/L)\" in new versions. Phosphorous was analysed using the spectrophotometer, is there a reference for the method?: Phosphorus (P) levels were analysed using a Perkin-Elmer AA spectrophotometer mod 3110 (Norwalk, CT, USA) (Perkin-Elmer (1982) How were the other minerals analysed following acid treatment? This information is missing: The author will add new versions after the sentence dilute appropriately for each mineral element and finally analyze with a Perkin–Elmer AA mod 3110 spectrophotometer (Norwalk, CT, USA). Specify whether it is 23% in relative or absolute terms - insert \"absolute\" before content: The author state that the highest protein content was relatively recorded in C. carpio fish scale flour, and the content did not differ by more than 23%. Conclusion: In new revisions, in the conclusion part, the author will change the sentence \"Consequently, enriching fish scale flour with animal and plant oils rich in omega-3 fatty acids is vital\" to \"However, this needs to be validated in the context of amino acid requirements with fish meal or other ideal protein sources that meet the requirements for the cultured species.\"" } ] } ]	1	https://f1000research.com/articles/12-1144
https://f1000research.com/articles/12-1440/v1	07 Nov 23	{ "type": "Research Article", "title": "Recreational handball-based training for people with type 2 diabetes: a feasibility trial", "authors": [ "Martin Færch Andersen", "Allan Riis", "Henrik Foged Borup", "Astrid Dall", "Mie Torp", "Rikke Hareskov Elversøe", "Janus Laust Thomsen", "Peter Vestergaard", "Anne-Mette Lücke Dissing", "Allan Riis", "Henrik Foged Borup", "Astrid Dall", "Mie Torp", "Rikke Hareskov Elversøe", "Janus Laust Thomsen", "Peter Vestergaard", "Anne-Mette Lücke Dissing" ], "abstract": "Introduction Type 2 diabetes mellitus (T2DM) is recognized as a serious public health concern with a considerable impact on people suffering from the disease and the society. The benefit of physical activity in the prevention and treatment of T2DM are well documented, however, a considerable proportion of individuals with T2DM have an inactive and sedentary lifestyle. Although most people with T2DM are aware of the importance of exercise, many are not interested in joining traditional exercise options, and long-term adherence is poor for those who do. Thus, we aim to investigate the feasibility of recreational handball-based training (HBT) for people diagnosed with T2DM.\n\nMethods This single-arm feasibility trial included adults (over 30 years) with a clinical diagnosis of T2DM. They were invited to participate in a 12-week HBT consisting of two weekly 60-minute exercise sessions. The outcome was feasibility, determined by adherence, exercise intensity, adverse events, dropout rate, and metabolic parameters.\n\nResults From September to December 2021, 10 people were included in the study. One participant dropped out because of illness and one participant dropped out due to suspected atrial fibrillation. Eight participants concluded the intervention and participated on average 86% (range 14–23) of the HBT sessions exercising with a mean heart rate of 73.4% (standard deviation (SD) 10.2) of individual maximum heart rate.\n\nConclusions HBT for people diagnosed with T2DM was found feasible with a high attendance rate and clinically relevant exercise intensities. However, future randomized controlled trials about the effects of the handball intervention are needed.\n\nTrial registration This trial was registered in ClinicalTrials.gov (NCT05015946) on 23/08/2021.", "keywords": [ "Exercise Therapy", "Diabetes Mellitus", "Type 2", "Sports for Persons with Disabilities", "Treatment Adherence and Compliance", "Metabolism", "Quality of Life", "team handball training", "team sports" ], "content": "Introduction\n\nType 2 diabetes mellitus (T2DM) is recognized as a public health concern, in 2021 it was estimated that 537 million adults are living with diabetes.1 T2DM is associated with severe complications such as retinopathy, nephropathy, neuropathy, and ultimately premature mortality primarily due to cardiovascular events,2 and 35% of people with T2DM experience complications even before the time of diagnosis.3 As a consequence, T2DM influence individuals’ functional capacity and quality of life.4\n\nThe benefit of physical activity and exercise in the prevention and control of T2DM and its complications are well documented,5,6 and is a cornerstone in the first-line treatment. However, a considerable proportion of people with T2DM are inactive and have a sedentary lifestyle and do not adhere to clinical recommendations.7,8 Although most people with T2DM recognize the importance of exercise, many are not interested in joining formal, structured exercise programs and long-term adherence is poor for those who do.9 Thus, a need for new exercise opportunities that can effectively improve diabetes health, while endorsing high levels of motivation to ensure long-term adherence is needed.\n\nDuring the recent decades, there has been an increased interest in practicing adapted recreational team sports in the treatment and control of noncommunicable diseases.10 There are indications, that team sport is motivating for a sedentary population,11–13 and may promote internal motivation which is a strong predictor of fidelity and adherence.11 Most research in the field of recreational team sport has been conducted with a soccer-based intervention, however, recreational handball show similar demands to those described for recreational soccer.14,15 Former trials show15–20 that recreational team handball for untrained adult men and women is an intermittent high-intensity exercise mode in the range of those found to have a positive effect on aerobic, anaerobic and musculoskeletal fitness in adult individuals. Furthermore, the exercise intensity was unaffected by past experience with handball or other sports.19 A 16-week recreational team handball intervention for 67 postmenopausal women with no experience with the sport, found that handball produced moderate-to-vigorous aerobic intensities and two weekly 60 minutes handball sessions resulted in 10% improvements in cardiorespiratory performance.21 However, the feasibility of handball in a clinical population has yet to be investigated.\n\nWe aimed to study the feasibility of recreational handball-based training (HBT) for people diagnosed with T2DM.\n\n\nMethods\n\nThis trial was conducted according to the declaration of Helsinki and approved by the North Denmark Region Committee on Health Research Ethics (registration number N-20200006) on 17/06/2020. Participants provided written informed consent prior to baseline assessment.\n\nThis trial was registered in ClinicalTrials.gov identifier: NCT05015946. Registration date: 23/08/2021.\n\nSingle-arm feasibility trial.\n\nPeople older than 30 years with a clinical diagnosis of T2DM were eligible for inclusion. Potential participants who were prohibited by their general practitioner from participation in HBT were excluded. The local Diabetes Association in the municipality of Aalborg recruited study participants by advertising in their membership magazines.\n\nThe intervention was delivered in an indoor municipal sports arena and consisted of 12 weeks of recreational HBT with two 60 minutes sessions per week. Each session included 1) 20 minutes of standardized warm-up, 2) 20 minutes of handball specific exercises and 3) 20 minutes of small-sided matches. To avoid injuries, no hard tackles were allowed, and the handballs used were light and made of soft material. The intervention was facilitated by BHF, DA, TM and ERH as part of their physiotherapy bachelor project. The intervention is illustrated in Table 1.\n\nPrimary outcomes\n\nThe primary outcomes were feasibility of HBT, determined by adherence, adverse events, dropout rate, and the exercise intensity. Dropout was defined as individuals completing baseline tests but not the follow-up test. The exercise intensity was measured using heart rate (HR) monitors (Polar h10) at both sessions in week 2,7 and 12. Adverse events were recorded during the entire intervention period.\n\nFurther outcomes were change from baseline to 12 week on hemoglobin A1c (HbA1c) (HemoCue HbA1c 501), physical (PCSc) and mental health component (MCSc) measured with the Short Form 36 (SF36)22 and cardiovascular fitness measured with peak oxygen uptake (VO2peak) during an incremental cycling test following a standard protocol. A bike ergometer (Corival Lode, Groningen, The Netherlands) was used with a direct measurement of oxygen consumption by breath-by-breath on Jaeger Masterscreen CPX (Intramedic A/S, Lyngby, Denmark).\n\nSecondary outcomes\n\nSecondary outcome measures were change from baseline to 12 week on self-care behavior measured with Diabetes Intention, Attitude and Behavior Questionnaire (DIAB-Q)23 and physical activity levels measured with Physical Activity Scale 2 (PAS2),24 cholesterol (total, low-density lipoprotein (LDL), high-density lipoprotein (HDL), and triglyceride (TG) (Accutrend Plus Cholesterol Meter), body composition (body mass index (BMI), body fat, lean body mass) (Tanita MC-180MA, Tokyo, Japan), visceral adipose tissue (hip/waist ratio) and resting blood pressure (Omron, M4-I).\n\nAfter arriving at the test facility, the participants were informed about the procedure of the tests performed. Afterward, they were asked to rest for 10 min in a sitting position before applying the cuff on the left arm. Three measures were taken using an automatic blood pressure monitor and the lowest value of both diastolic and systolic blood pressure was used for calculations. HbA1c and cholesterol (total, LDL, TG, and HDL) were measured using a capillary blood sample. The fingertip was first disinfected. Then, blood was collected for both HbA1c and cholesterol. To measure waist circumference a standard measuring tape was placed between the lowest rib and the upper iliac crest after a normal exhalation. To find the hip circumference the measuring tape was placed at the widest part of the hip. A Tanita® digital scale with bioelectrical impedance was used to measure weight and body fat. The participants stepped barefooted onto the scale and were instructed to keep the handgrips slightly elevated from the torso. The VO2peak test was initiated with a 5-min warm-up period on 50 watts, followed by a 25-watt increase per minute until exhaustion. The test was considered a VO2max test if the respiratory exchange ratio (RER) value exceeded 1.10 and the O2-consumption reached a plateau. Otherwise, it was categorized as a VO2peak test.\n\nAdherence was defined as the percentage of training sessions attended of the scheduled 24 sessions. Exercise intensities were merged into three timepoints by calculating the mean of the two weekly sessions in week 2, 7 and 12. Percent of heartrate maximum (HRmax) was calculated using an estimated maximum heart rate 208−0.7×age.25 Statistics on adherence and intensities were conducted in Microsoft Excel (version 2302, Office 365). Results in within-group changes from baseline to 12 weeks are presented as median with minimum/maximum and analyzed using the Wilcoxon-rank test in SPSS (IBM SPSS Statistics 27) (RRID:SCR_002865). Statistical significance was set at p<0.05.\n\nFrom a convenience approach, a group of 10–15 participants was recruited in respect to the feasibility of small-sided matches on a single court.\n\n\nResults\n\nIn September 2021, 15 interested people with T2DM were recruited and screened for eligibility. Two were diagnosed with Type 1 diabetes, and one was under 30 years old and thereby not eligible for inclusion. Two decided not to participate due to time and transportation. In total, 10 participants were included. During the intervention, one participant dropped out due to illness not related to the intervention, and one participant stopped due to suspected atrial fibrillation detected in the baseline assessment. Eight participants contributed with follow-up data. A study flowchart is presented in Figure 1 and participants’ characteristics at baseline in Table 2.\n\nThe eight participants attended a mean of 20 (range 14–23) of 24 training sessions, corresponding to an attendance rate of 83%. Mean HR during HBT, based on six training sessions in weeks 2, 7 and 12, was 121.3 beats per minute (standard deviation (SD) 17) corresponding to 73.4% (SD 10.2) of individual maximal HR. Time spent with a HR over 70% was 61.1% (SD 30.1), and time spent in the intensity zones 70–80%, 80–90% and 90–100% was 31.9% (SD 18.5), 21.6% (SD16.1) and 7.6% (SD 10.6) of total training time, respectively. The mean % of maximal HR in the first 20 minutes (warm-up) was 65.5% (SD 6.5), the middle 20 minutes (handball specific) was 74.8% (SD 8.4) and the last 20 minutes (matches) was 79.4% (SD 10.2). Individual peak HR was 130 bpm (SD 4.7) corresponding to 89.6% of HRmax (SD 2.9) measured in week 12. Mean exercise intensities are displayed in Table 3 and individual exercise intensities with one-minute frequency for all participants is illustrated in Figure 2a–f.\n\nHRmax, heart rate maximum; ID, identification.\n\nThere was no statistical change from baseline to 12 weeks for HbA1c (p=0.14), VO2peak (p=0.4) or physical (p=0.11), and mental health (p=0.48). From baseline to follow-up there was a small increase in total cholesterol (p=0.09) mediated by an increase in LDL (p=0.02). There was a drop in body fat mass (p=0.04) and BMI (0.02) but no change in visceral fat (p=0.24). We found no change in systolic blood (p=0.54) pressure but a significant drop in diastolic (p=0.04) blood pressure. Lastly there were no changes in the participant’s physical activity levels (p=0.45) but higher levels of engagement in beneficial behaviors (p=0.03). All baseline to 12-week measures are displayed in Table 4.\n\nNo serious adverse events were detected. Two times a participant fell involuntarily during the training session but continued and finalized the sessions. One participant experienced a muscle strain and resumed handball training after a one-week rest period. There were no episodes with exercise induced hypoglycemia.\n\n\nDiscussion\n\nRecreational HBT was found feasible for people diagnosed with T2DM with high attendance rate and moderate to high exercise intensities. The HR recordings showed an average HR of 73% (SD 10.2) and almost 30% of the time with an HR above 80% of HRmax. These values are in line with those found in non-clinical inactive postmenopausal women playing recreational handball by Pereira et al.21 They reported a mean HRmax at 76% (SD 6), however duration above 80% of HRmax at 44% (SD 20) of the time which is higher than those found in the present study. This may be explained by the differences between the interventions, as they only measured exercise intensities during small-sided matches.21 The last part of the present intervention did produce the highest exercise intensities among the participants with a mean HR at almost 80% of HRmax and more than 45% (SD 37) of the time intensities above 80% of HRmax. Andersen et al.26 reported mean intensities at 82% of HRmax and almost 65% of total playing time with intensities above 80% of HRmax, among men diagnosed with T2DM playing small-sided football. The explanation for this may be that their participants were younger (mean 50.6, SD 7.1) compared to the present study and that they excluded people with co-morbidities and diabetic complaints that might reduce the potential of playing football with high intensities.26\n\nThe present study was not powered to detect significant changes in effects. However, there was a tendency towards improvement in most of the outcomes assessed. Previous studies investigating football in women and men with T2DM reported improvements of 10% after 12 and 24 weeks in VO2peak26,27 working at similar exercise intensities as in the present study. Similar results are found in studies investigating recreational handball in different populations16,21,28 suggesting that HBT for people with T2DM may be an effective way to improve cardiovascular function. The potential effect of recreational team sport in reducing HbA1c is under-investigated, however, exercise in general especially with higher intensities has been found to substantially decrease HbA1c levels in people with T2DM.29 Furthermore, team sport has been found effective to increase the quality of life and instinct motivation,30,31 which are important predictor of long-term adherence and fidelity.11\n\nNo serious adverse events occurred during the handball-based training which is in concordance with previous studies.17,20,21 Only one case of muscle injury happened and was managed conservatively. Few episodes with involuntary falls occurred, however, these falls did not cause any injury that prevented the participants from continuing the training. These falls may be a source of great concern and the presence of diabetes complications such as retinopathy and neuropathy may place people with T2DM at a greater risk of falling. Compared to non-clinical populations engaging in handball training, the present study did not report more cases of adverse events.17,20,21 This may be explained by the difference in the intervention as former study interventions only include small-sided matches, which only represented the last third part of present HBT. Furthermore, the present intervention included 20 minutes of warm-up comprising running, strength training, coordination exercises, balance training, and flexibility exercises followed by 20 minutes of handball skill training, which may reduce the risk of adverse events occurring during matches.\n\nSome deviations from the registration in ClinicalTrials.gov were made. Since the current trial is a feasibility trial, we changed our primary outcomes reported in ClinicalTrials.gov to feasibility outcomes and listed all effectiveness outcomes as secondary outcomes. Furthermore, we planned an aerobic endurance test, however this test was not completed as it was too demanding according to the participants’ physical condition.\n\nThe recruitment rate was lower than expected and there was a possibility of selection bias, as the present study appealed to participants motivated for handball. Participants were asked to stay on their current medical prescription and change in medicine may affect some outcome of interest. When reporting exercise intensities in warm-up, handball-specific exercises, and small-sided matches the categorization may not represent the actual content of the training session due to the pragmatic delivery of the intervention. Consequently, some overlap between the three parts is expected to be present. All assessments were pre-defined and the assessment of clinically relevant outcomes was conducted using validated instruments.\n\nFor patients with T2DM, adherence and fidelity to prescribed exercise are highly important. In the present study we observed a high attendance rate (>80%) and clinically relevant training intensities. Additionally, it is notable that HBT can be played without prior experience with handball and can be played with a limited number of participants. Furthermore, small-sided matches were played at a 13×20 meters indoor court which allows three of these to fit on a regular handball court, and thereby the possibility of triple the number of participants. Thus, HBT may be attractive and easily accessible for people with T2DM. The present study’s feasibility findings and preliminary results indicate that future studies need to investigate the efficacy and effectiveness of HBT for people with T2DM in randomized controlled designs.\n\n\nConclusion\n\nHBT was feasible for people with T2DM. The present study observed a high attendance rate of 83% and a mean training intensity of almost 75% of HRmax. More than 60% of the time participant trained with an intensity of 70-100% of HRmax. The highest training intensity was found during small-sided matches, with a mean at 80% of HRmax. Future randomized controlled trials are needed to determine the effect of HBT.\n\n\nAuthor contributions\n\nMartin Færch Andersen: Conceptualization, methodological inputs, project administration, intervention delivery, data collection, formal analysis, funding acquisition, writing the first draft, commenting in rounds of revision (lead), approved the final draft.\n\nAllan Riis: conceptualization, methodological inputs, writing the first draft, commenting in rounds of revision, approved the final draft.\n\nHenrik Foged Borup: Intervention delivery, methodological inputs, commenting in rounds of revision, approved the final version.\n\nAstrid Dall: Intervention delivery, methodological inputs, commenting in rounds of revision, approved the final version.\n\nMie Torp: Intervention delivery, methodological inputs, commenting in rounds of revision, approved the final version.\n\nRikke Hareskov Elversøe: Intervention delivery, methodological inputs, commenting in rounds of revision, approved the final version.\n\nJanus Laust Thomsen: Methodological inputs, oversighting test procedures, commenting in rounds of revision, approved the final version.\n\nPeter Vestergaard: Methodological inputs, commenting in rounds of revisions, approved the final draft.\n\nAnne-Mette Lücke Dissing: Conceptualization, Methodological inputs, Data collection (lead), Formal analysis, commenting in rounds of revisions, approved the final draft.", "appendix": "Data availability\n\nOpen Science Framework: Data repository: Recreational handball-based training for people with type 2 diabetes. https://doi.org/10.17605/OSF.IO/VZ6G5. 32\n\nThis project contains the following underlying data:\n\n• dataset_pub.xlsx (Sheet 1, attendance: Individual participant (ID) attendance in two weekly training sessions for twelve weeks. X marks when participants attended a training session. Sheet 2-7, exercise intensity: Individual participant (ID) exercise intensity at both training sessions in week 2,7 and 12. First row indicate time (minutes), first column indicates participant (ID), data is reported as percentage of individual heart rate maximum.)\n\nOpen Science Framework: CONSORT extension for pilot and feasibility trials for ‘Recreational handball-based training for people with type 2 diabetes: a feasibility trial’. https://doi.org/10.17605/OSF.IO/VZ6G5. 32\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nWe thank Vibeke Brinkmann Løite for her methodological contributions when planning this study.\n\n\nReferences\n\nIDF Diabetes Atlas 10th edition.[cited 2022 Apr 8]. 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Sports. 2014 Aug [cited 2019 Sep 23]; 24: 66–75. Publisher Full Text\n\nOttesen L, Jeppesen RS, Krustrup BR: The development of social capital through football and running: studying an intervention program for inactive women. Scand. J. Med. Sci. Sports. 2010 Apr 6 [cited 2019 Sep 23]; 20: 118–131. Publisher Full Text\n\nPedersen MT, Vorup J, Nistrup A, et al.: Effect of team sports and resistance training on physical function, quality of life, and motivation in older adults. Scand. J. Med. Sci. Sports. 2017 Aug [cited 2019 Sep 24]; 27(8): 852–864. Publisher Full Text Reference Source\n\nRanders MB, Nielsen JJ, Bangsbo J, et al.: Physiological response and activity profile in recreational small-sided football: no effect of the number of players. Scand. J. Med. Sci. Sports. 2014 [cited 2022 Mar 28]; 24(SUPPL.1): 130–137. Publisher Full Text Reference Source\n\nPóvoas SCA, Castagna C, Resende C, et al.: Physical and Physiological Demands of Recreational Team Handball for Adult Untrained Men. Biomed. Res. Int. 2017 [cited 2019 Sep 20]; 2017: 1–10. PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source\n\nPóvoas SCA, Castagna C, Resende C, et al.: Effects of a Short-Term Recreational Team Handball-Based Programme on Physical Fitness and Cardiovascular and Metabolic Health of 33-55-Year-Old Men: A Pilot Study. Biomed. Res. Int. 2018 Oct 3 [cited 2019 Sep 20]; 2018: 4109796. Reference Source\n\nHornstrup T, Wikman JM, Fristrup B, et al.: Fitness and health benefits of team handball training for young untrained women-A cross-disciplinary RCT on physiological adaptations and motivational aspects. J. Sport Health Sci. 2018 Apr [cited 2019 Sep 20]; 7(2): 139–148. PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source\n\nFristrup B, Krustrup P, Andersen JL, et al.: Effects of small-sided recreational team handball training on mechanical muscle function, body composition and bone mineralization in untrained young adults-A randomized controlled trial. PLoS One. 2020 Nov 1 [cited 2022 Mar 28]; 15(11): e0241359. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHornstrup T, Póvoas S, Helge JW, et al.: Cardiovascular and metabolic health effects of team handball training in overweight women: Impact of prior experience. Scand. J. Med. Sci. Sports. 2020 Feb 1 [cited 2022 Mar 28]; 30(2): 281–294. Publisher Full Text Reference Source\n\nHornstrup T, Løwenstein FT, Larsen MA, et al.: Cardiovascular, muscular, and skeletal adaptations to recreational team handball training: a randomized controlled trial with young adult untrained men. Eur. J. Appl. Physiol. 2019 Feb 6 [cited 2022 Mar 28]; 119(2): 561–573. PubMed Abstract \| Publisher Full Text\n\nPereira R, Krustrup P, Castagna C, et al.: Effects of a 16-week recreational team handball intervention on aerobic performance and cardiometabolic fitness markers in postmenopausal women: A randomized controlled trial. Prog. Cardiovasc. Dis. 2020 Nov 1 [cited 2022 Mar 28]; 63(6): 800–806. PubMed Abstract \| Publisher Full Text\n\nBjorner JB, Thunedborg K, Kristensen TS, et al.: The Danish SF-36 Health Survey: translation and preliminary validity studies. J. Clin. Epidemiol. 1998 Nov [cited 2023 Oct 2]; 51(11): 991–999. PubMed Abstract \| Publisher Full Text Reference Source\n\nTraina S, Mathias S, Colwell H, et al.: The Diabetes Intention, Attitude, and Behavior Questionnaire: evaluation of a brief questionnaire to measure physical activity, dietary control, maintenance of a healthy weight, and psychological antecedents. Patient Prefer. Adherence. 2016 Feb 29 [cited 2019 Oct 1]; 10: 213. Reference Source\n\nWare JE: SF-36 Health Survey.Maruish ME, editor. The use if psychological testing for treatment planning and outcomes assessment. Lawrence Erlbaum Associates Publishers; 1999; p. 1227–46.\n\nTanaka H, Monahan KD, Seals DR: Age-predicted maximal heart rate revisited. J. Am. Coll. Cardiol. 2001 [cited 2022 Mar 28]; 37(1): 153–156. PubMed Abstract \| Publisher Full Text\n\nAndersen TR, Schmidt JF, Thomassen M, et al.: A preliminary study: Effects of football training on glucose control, body composition, and performance in men with type 2 diabetes. Scand. J. Med. Sci. Sports. 2014 Aug [cited 2021 Mar 11]; 24(SUPPL.1): 43–56. Publisher Full Text\n\nDe Sousa MV, Fukui R, Krustrup P, et al.: Positive effects of football on fitness, lipid profile, and insulin resistance in Brazilian patients with type 2 diabetes. Scand. J. Med. Sci. Sports. 2014 [cited 2022 Apr 8]; 24(SUPPL.1): 57–65. Publisher Full Text\n\nPereira R, Krustrup P, Castagna C, et al.: Effects of recreational team handball on bone health, postural balance and body composition in inactive postmenopausal women — A randomised controlled trial. Bone. 2021 Apr 1; 145: 115847.\n\nLiu Y, Ye W, Chen Q, et al.: Resistance Exercise Intensity is Correlated with Attenuation of HbA1c and Insulin in Patients with Type 2 Diabetes: A Systematic Review and Meta-Analysis. Int. J. Environ. Res. Public Health. 2019 Jan 1 [cited 2022 Apr 8]; 16(1).Free Full Text Publisher Full Text \|\n\nPedersen MT, Vorup J, Nistrup A, et al.: Effect of team sports and resistance training on physical function, quality of life, and motivation in older adults. Scand. J. Med. Sci. Sports. 2017 Aug 1 [cited 2022 Apr 8]; 27(8): 852–864. Publisher Full Text\n\nNielsen G, Wikman JM, Jensen CJ, et al.: Health promotion: The impact of beliefs of health benefits, social relations and enjoyment on exercise continuation. Scand. J. Med. Sci. Sports. 2014 [cited 2022 Apr 8]; 24(SUPPL.1): 66–75. Publisher Full Text\n\nAndersen MF: Data repository: Recreational handball-based training for people with type 2 diabetes. [Dataset]. OSF. 2023. Publisher Full Text\n\nBizzini M, Dvorak J: FIFA 11+: an effective programme to prevent football injuries in various player groups worldwide-a narrative review. Br. J. Sports Med. 2015 May [cited 2019 Sep 24]; 49(9): 577–579. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "251454", "date": "17 Apr 2024", "name": "Jacob Uth", "expertise": [ "Reviewer Expertise Rehabilitation", "Physiotherapy" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis study aimed to assess the feasibility of recreational handball-based training (HBT) for individuals with T2DM. The trial involved 10 participants aged over 30 years with clinical T2DM diagnosis, engaging in 12 weeks of HBT sessions twice a week. The main finding was a high attendance rate (83%), and moderate to high intensity during training with an average heart rate of 75% of maximum heart rate and over 60% of the time spent training at intensities of 70-100% of maximum heart rate. There were no significant changes in HbA1c or cardiovascular fitness, but body fat mass, BMI and diastolic blood pressure decreased. The study reported no serious adverse events related to HBT, with only minor injuries occurring. Future randomized controlled trials are needed to determine the effectiveness of HBT in improving outcomes for individuals with T2DM. Specific comments: - Some references in the background are quite old. Please check for newer research e.g. exercise guidelines for T2DM. - In the Methods section: how was participants referred from the local Diabetes Association to the study staff, and who obtained informed consent from participants? - Sample size: It is stated that 10-15 participants were recruited. Should it state that 10-15 participants were planned to or intended to be recruited? -Please provide a reference for the fitness test protocol -In Table 4: The values for Vo2 peak seems to be in Liters and not ml. - If possible it would be beneficial to provide data on the fitness test (Time-to-exhaustion, Watts, RER values), possibly in an appendix. This would also clarify if the test were VO2 peak or VO2 max tests. - In the discussion, it is mentioned that there may have been a selections bias because the study appealed to participants motivated for handball, and under perspectives it is mentioned, that HBT can be played without prior handball experience. Is any data available regarding the participants prior experience with HBT? -It is concluded that HBT was feasible for people with T2DM. Are there specific lessons learned that should be considered when planning a randomized trial regarding, e.g., recruitment, testing and delivering of the intervention?\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1440
https://f1000research.com/articles/11-755/v1	07 Jul 22	{ "type": "Research Article", "title": "Performance of Malnutrition Universal Screening Tool and Patient-Generated Global Subjective Assessment in screening for cancer-related malnutrition in Nairobi, Kenya", "authors": [ "Caroline M.N. Auma", "Marshal M. Mweu", "Rose O. Opiyo", "Marshal M. Mweu", "Rose O. Opiyo" ], "abstract": "Background: Malnutrition is a common feature among oncology patients. It is responsible for poor response and tolerance to anticancer therapy, increased morbidity, and mortality. More than half of malnourished cancer patients remain undetected owing to lack of effective screening. Body mass index is the main indicator for assessing malnutrition in Kenyan public hospitals. However, it underestimates weight loss in patients with chronic illnesses. The Malnutrition Universal Screening Tool and Patient-Generated Subjective Global Assessment have been widely used in research and clinical practice and have both reported good validity and reliability. However, their diagnostic evaluation has not been performed in Kenya. Methods: A cross-sectional study was conducted among 138 and 76 cancer outpatients from Kenyatta National Hospital and Texas cancer treatment centres, respectively. Participants had a confirmed disease, stage 1-4 cancer, and aged 18 years and above. They were screened for malnutrition using both Malnutrition Universal Screening Tool and Patient Generated-Subjective Global Assessment. A separate study questionnaire was utilized to gather participant’s socio-demographic and clinical characteristics. A Bayesian latent class modelling framework was employed to infer the tests’ estimates based on participants ‘cumulative scores from the two tests. Results: The cut-off value of ≥ 1 and ≥ 4 gave the best combination of sensitivity and specificity of Malnutrition Universal Screening Tool and Patient Generated-Subjective Global Assessment. Both tests yielded statistically similar sensitivities and specificities. Predictive values were comparable between the tests and across the two populations. The posterior median true prevalences of malnutrition were high (˃ 54%) and numerically similar between the studied populations. Conclusions: The performance of both tests among patients with cancer is similar. Healthcare workers are therefore at liberty to use either of them to inform treatment. Given the high true prevalence of cancer-related malnutrition, routine screening is critical and should be included as part of cancer care.", "keywords": [ "Bayesian Latent Class Model", "Malnutrition", "Malnutrition Universal Screening Tool", "Patient Generated Global Subjective Assessment", "Test accuracy", "Cancer patients." ], "content": "Introduction\n\nMalnutrition is a common feature in oncology patients.1 Globally, malnutrition affects about 39 to 87% of patients with cancer2 depending on the assessment method3 and the type of cancer involved.4 For example, while breast cancer patients have a low risk of weight loss and subsequent malnutrition, patients with gastrointestinal, lung, and head and neck tumours have a high risk.5 In Kenya, 31% of patients with cancer are malnourished and the greatest burden lies among patients with gastrointestinal tumours at 49.1%.6\n\nCancer-induced malnutrition has significant clinical and economic burden. The malnutrition in these patients is associated with reduced responsiveness and tolerance to treatment, impaired immune function, increased risk of complications, reduced survival and quality of life, and high healthcare expenditures.4,7,8 Globally, 50% of patients dying from cancer are malnourished.9 About 20% of deaths in patients with cancer occur due to malnutrition-related complications rather than the direct effect of malignancy.10 Despite its high prevalence and adverse sequelae in cancer patients, malnutrition in more than half of these patients remains undetected and untreated owing to lack of effective screening.11–13\n\nTimely recognition of malnutrition is critical for appropriate and intensive nutrition support12 to stop or counteract weight loss and subsequent malnutrition.14,15 Currently, body mass index (BMI) is the major indicator for the assessment of malnutrition in Kenyan public hospitals.16,17 However, BMI when used independently is a less sensitive parameter in predicting malnutrition due to large tumours9 and short term fluid imbalance.18,19 Therefore, patients can suffer malnutrition regardless of BMI value because loss in lean body mass is masked by ascites and body fat.20,21 The commonly available screening tools for use in oncology setting are Patient Generated-Subjective Global Assessment (PG-SGA) and Malnutrition Universal Screening Tool (MUST). These screening tools assess patients’ nutrition status using a cluster of two or more nutrition-related parameters, thus are superior to BMI.4,19,20 PG-SGA and MUST have been widely used in both clinical practice22,23 and research.24 They have reported good sensitivity (Se) and specificity (Sp) in identifying the nutrition status of patients with cancer.25–30\n\nThe PG-SGA is a simple method employed in assessment of nutritional risk and identification of those who would benefit from nutritional support, thus considered the most appropriate tool for detecting malnutrition in cancer patients.31 The PG-SGA is a 4-in-1 instrument to screen, assess, and monitor malnutrition and its risk factors and to triage for interdisciplinary intervention.25,32 It is inexpensive, quick to complete33 and user-friendly.34,35 More to its merits, the first four boxes can be completed by a patient, which empowers them on matters related to their health and reduces the total time involved. In addition, PG-SGA contains more nutrition impact symptoms that are vital to cancer patients.18 Lastly, PG-SGA assess nutrition status on a continuous scale, which allows it to detect subtle changes in nutritional status occurring over a short period and triage patients for appropriate nutrition intervention.25,31,36 Overall, PG-SGA allows malnourished patients to be quickly identified and prioritized for care, thus a suitable screening tool.22,25\n\nThe MUST is designed to categorize adults as either malnourished, obese, or at risk of malnutrition and identify those who would benefit from appropriate nutrition intervention.37 It does not incorporate subjective/clinical parameters reflecting changes in nutritional risk and status and neither does it require time-consuming calculations.23 It is also quick, inexpensive, easy to apply and interpret, and hence acceptable to both patients and healthcare workers.23,38,39 MUST combines weight loss and BMI, which reduces misclassification errors, thus improving Se and Sp.19 Additionally, a patient can screen him/herself without the help of a health worker with a time response as low as 5 minutes.40\n\nAlthough these tools have been extensively validated in various countries, their performance has not been evaluated in Kenya. Additionally, in countries where they have been evaluated, evaluations were done against reference standards such as Subjective Global Assessment,25,26 Mini-Nutritional Assessment,27 Nutritional-Risk Screening-2002,41 Albumin and BMI.42 This may have plagued the estimates with information and selection bias.43 Nevertheless, the accuracy of two or more tests can be quantified without prior knowledge of the true disease status by using latent class models.43,44 Thus, the objective of the present study was to derive the Se and Sp together with the positive and negative predictive values of PG-SGA and MUST for malnutrition in head and neck, respiratory, and gastrointestinal cancer patients using the Bayesian Latent Class Model (BLCM).\n\n\nMethods\n\nA facility-based cross-sectional study was conducted to evaluate the performance of the MUST and PG-SGA in screening for malnutrition among cancer patients. This study design was most suitable due to the ease of recruitment of cancer patients presenting to the cancer clinic and its suitability for the descriptive nature of the study.\n\nThe study was conducted at Kenyatta National Hospital-Cancer Treatment Centre (KNH-CTC) and Texas Cancer Centre (TCC) in Nairobi, Kenya. KNH is the largest national public referral hospital in Kenya. It registers 80,000 inpatients and 500,000 outpatients annually. It offers comprehensive cancer services, which include diagnosis and treatment (chemotherapy, radiotherapy, surgical and hormonal therapy). The three most commonly diagnosed carcinomas are cervical, breast, and oesophageal cancer in that order. KNH-CTC provides both inpatient and outpatient services. Although nutrition screening of cancer patients is not conducted routinely, there is an in-house nutritionist whose services are sought under certain circumstances.\n\nThe TCC is the leading private cancer care and treatment centre in Kenya. It receives approximately 11000 patients annually. Apart from cancer diagnosis and treatment, TCC offers pain and palliative care services, counselling, physiotherapy, and rehabilitation services. With regards to nutrition services, currently, TCC does not have an in-house nutritionist, but under certain circumstances, one is outsourced. Consequently, nurses and doctors offering oncological services provide nutrition support. Weight and height are the measurements taken for BMI scoring but not routinely.\n\nThe study population consisted of all patients with head and neck, respiratory and gastrointestinal cancers attending outpatient oncology clinic at KNH-CTC and TCC. On eligibility, a patient had to have established disease, that is, stage 1-4 cancer, aged 18 years and above, physically stable, and have given informed consent. To obtain the study sample, a simple random sampling technique was applied to a sampling frame comprising of patients booked for evaluation in each facility on a given day. To ascertain the number of patients to be sampled from each facility, a probability proportional to size sampling45 was employed guided by the number of patients seen in a given facility in a particular month. As such, 138 and 76 cancer patients from KNH-CTC and TCC, respectively, were recruited.\n\nTwo research assistants with a medical background who had previously been trained on interviewing techniques recruited and interviewed participants. Upon obtaining informed consent from the participants, the MUST and PG-SGA (either English or Kiswahili version depending on the individual’s preference) were administered through a face-to-face interview in a private room within the Cancer Clinic. The MUST and PG-SGA tools were completed just before provision of oncology services. Aside from the nutrition status, the patients’ socio-demographic characteristics (age, sex, marital status, level of education, area of residence and employment status) and clinical characteristics (tumour site and cancer stage) were also recorded.\n\nThe required sample size was estimated using McNemar’s sample size formula for paired proportions.46\n\nThe study participants provided written informed consent prior to engaging in the study. Approval to conduct the study was granted by Kenyatta National Hospital and University of Nairobi joint Ethics and Research Committee (KNH-ERC/A/315).\n\nThis referred to the latent (unobserved) malnutrition status regardless of the severity level among head and neck, respiratory and gastrointestinal cancer patients that is targeted by MUST and PG-SGA.\n\nThe MUST employs three stand-alone criteria to classify patients’ nutritional status: BMI score (BMI ˃20 kg/m2 = 0, BMI 18.5-20 kg/m2 = 1, BMI <18.5 kg/m2 = 2), unintentional weight loss score in the last 3 to 6 months (weight loss < 5% = 0, weight loss 5-10% =1, weight loss ˃ 10% = 2) and cute disease effect score (add a score of 2 if there has been or is likely to be no nutrition intake for > 5 days). Each parameter is scored on a scale of 0, 1, or 2 giving an overall total score of six. Overall risk of malnutrition is classified as low risk if the score = 0, moderate risk if the score = 1, and a score of ≥ 2 signifies high risk.23 In this study, a patient with a total score of ≥1 was considered malnourished.\n\nThe PG-SGA relies on weight history, changes in patient’s dietary intake, presence of gastrointestinal symptoms, functionality, diagnosis, age, metabolic stress, and physical examination (subcutaneous fat loss, muscle wasting, and fluid status) to assess a patient’s nutritional risk. For each of the above-mentioned components, 0-4 points are awarded based on the relative impact on nutritional status. The overall total scores of PG-SGA range from 0-52. Based on the scores, patients were divided into three categories: (PG-SGA < 2, well-nourished/no risk), (PG-SGA ≥ 2 and < 9 suspected/moderate risk), (PG-SGA ≥ 9, severe risk). Three potential cut-offs were examined: individuals being categorised as malnourished if they had scores ≥2,9,26 ≥4,42,47 ≥9.25,48\n\nA Bayesian modelling framework was used to derive the prevalence, Se and Sp estimates along with the predictive values of the tests. The BLCM was fitted in OpenBUGS software (v 3.2.2)49 but called from R (v. 3.6.2) via the Brugs package (v0.9-0).50 Importantly, the model design and reporting were guided by the standards for reporting of diagnostic accuracy studies that use BLCMs (STARD-BLCM).51 The Bayesian code is available as Underlying data.52\n\nAccording to Hui and Walter,44 the BLCM has the following assumptions: (i) there must be two or more populations with differing prevalences. For this study, two populations of patients attending KNH-CTC and TCC facilities were established. Owing to the inherent socio-economic differences between the facilities’ catchment populations, the prevalences of cancer-related malnutrition were presumed to be distinct. (ii) The Se and Sp of both tools should be consistent across the populations. (iii) Given the disease status, there was conditional independence between the two tests. This was a reasonable assumption considering the two tools have separate symptom targets: MUST targets changes in body composition thus, assesses chronic malnutrition whereas PG-SGA assesses gastrointestinal symptoms preventing food intake thus, targets acute malnutrition.53 Consequently, the probability of a patient testing either positive or negative to one test is not affected by the result of the other test.\n\nIt was assumed that the different combinations of test results, for each population, observed as counts Ok follow a multinomial distribution:\n\nWhere SeiandSpi are the Se and Sp measures for the ithtest (i=1, 2) and pk represent the kth population’s prevalence. Probk, represents a vector of probabilities of observing the tests’ results’ combinations (such as +, +), while nk is the sample size used in population k. The probabilities are defined using the specific test characteristics (Se and Sp) and prevalence (p) of each population. For example, prob1for a person who tests positive to both tests is given by:\n\nSince there were two populations, the Latent Class Model contained six parameters: each test’s Se and Sp, as well as two population-specific prevalences. These six parameters were estimated from six degrees of freedom obtained from each of the two populations. As there was no reliable prior information on any of the parameters, uninformative priors (beta (1, 1)) were used.\n\nThe Positive Predictive Value (PPV) and Negative Predictive Value (NPV) for each test within population k were derived using the following formula:\n\nTo select the most optimal cut-off values, Youden indices54 were computed using the Se and Sp obtained at each of the following pairs of cut-off values: ≥1 for MUST paired with each of the following PG-SGA cut-off values (≥2, ≥4, and ≥9). The pair of cut-off points with the highest Youden index was chosen.\n\nTwo Markov Chain Monte Carlo (MCMC) chains with varying values were used to initialize the model. We ran 6000 iterations of the model with the initial 3500 discarded as the burn-in phase. Convergence of the MCMC chain was assessed by visual appraisal of the density plots and Gelman-Rubin Diagnostic plots. The posterior distribution of prevalence of each of the two populations, the test estimates, and their respective predictive values were reported as the median and their associated 95% posterior credible intervals (PCI).\n\n\nResults\n\nA total of 214 participants were selected, and 202 consented to participate, giving a response rate of 94.4%. However, of the 202 participants, 14 were excluded from analysis due to lack of records on previous anthropometric measurements, which rendered their total scores from MUST and PG-SGA unreliable.\n\nThe socio-demographic and clinical features of 188 participants are displayed in Table 1. Notably, the participants’ median age was 56.5 years (range: 18-81 years). Majority of the participants were males (64.36%). Only 46% of the participants had attained at least a secondary school certification. Clinically, patients with gastrointestinal cancers formed the largest proportion of the sample at 54.26%. Approximately 34% of the participants had stage 4 cancer.\n\nThe cross-classified counts of the two test results at various cut-off points are presented in Table 2. The cut-off value of ≥1 and ≥4 gave the best combination of the Se and Sp of MUST and PG-SGA (see Table 3). Thus, this cut-off was used to infer subsequent parameters. The PG-SGA registered a Se of (92.4; 95% PCI [81.2; 99.6]) and Sp of (72.5% 95% PCI [54; 97.2]) while MUST had a Se of (83.1 95% PCI [67.4; 98.9]) and a Sp (85.7; 95% PCI [71.4; 99.6]). Although the Se and Sp of the two tests differed numerically, they were statistically similar based on 95% PCIs.\n\na Positive.\n\nb Negative.\n\nOn predictive values, NPVs and PPVs were statistically similar between the tests and across the two populations (Table 4). In KNH, MUST had a NPV of 77.9% and a PPV of 89.7% while PG-SGA had a NPV of 87.2% and a PPV of 82.9%. In TCC, MUST had a NPV of 81.2% and a PPV of 87.8% while PG-SGA a NPV of 89.2% and a PPV of 80.1%. The posterior median true prevalences of malnutrition were high (˃54%) and numerically similar between the studied populations (Table 4).\n\nPPV=Positive Predictive Value; NPV=Negative Predictive Value.\n\n\nDiscussion\n\nIn this study, MUST registered a Se of 83.1% supported by finding from other studies28,29 and a Sp of 85.7% in agreement with findings from previous studies.28,30 However, in the literature, the Se of MUST has been shown to vary from 29%55 to 97%41 similar to its Sp from 48.9%56 to 94.5%.29 This inconsistency can be logically explained by the percentage weight loss, which is a crucial parameter in the MUST tool.28 For example, patients may have lost weight in the past, but their present weight recovery was not calculated since MUST does not give this provision.56 In addition, contrary to this study, in those studies, estimation of the test accuracy was based on the assumption that a perfect reference standard existed. This could have led to biased estimates. More so, MUST in its categorization of nutrition status, relies on the percentage of weight lost between 3 and 6 months. As a result, where patients were required to self-report the involuntary weight loss, there was a possibility of under-estimation hence affecting Sp. Ramboer, Verhamme57 in their study reflected the propensity for underestimation of weight loss in oncology patients. In this study, all patients with no previous records of anthropometric measurements were excluded from the analysis. Although MUST was found to be a suitable screening tool, it requires that patients be weighed for the determination of BMI and weight loss. Thus, inaccurate weight in patients with oedema or ascites could lead to underestimation of nutritional risk in some patients.33\n\nThe PG-SGA yielded a Se and Sp of 92.4% and 72.5%, respectively. The findings demonstrate that the PG-SGA is a highly sensitive malnutrition screening tool in the outpatient oncology population, consistent with findings from previous studies.27,25,26,42 An ideal screening tool would be 100% sensitive and specific. But, the necessity to categorize all malnourished patients (Se) correctly takes precedence over misclassification of well-nourished individuals (Sp).58 The Sp of PG-SGA varies greatly in published studies, with estimates ranging from 2.3%1 to 88.1%.27 The Sp of 72.5% observed in this study falls within the above range. The variability could be due to three things: First, use of different reference standards in deriving test estimates, which was not the case in this study, for example, specificities of 2.3% and 88.1% were based on albumin and Mini-Nutrition Assessment tool, respectively. Secondly, different administration methods, since the score from participant-versus researcher administered may not be comparable. This is because, the authors of PG-SGA speculate that patients may over-report symptoms based solely on their presence, regardless of their impact on food intake,58 thus underestimating Sp. For example, Sp of 21.8%59 and 82%25 were derived from patient and researcher administered studies, respectively. This study was researcher administered thus a good Sp was registered. Lastly, the scores derived from component Number Three of PG-SGA questionnaire (nutrition impact symptoms and other factors). In this section, any symptom impairing food intake reported by the patient is scored, and all points are cumulative (maximum 24 points).22 This could also rise the number of false positives thus underestimating Sp. However, this wide range of symptoms may help detect more patients at risk of malnutrition. As the detection of symptoms that impair nutritional intake in the early stages of the disease may be advantageous for proactively preventing cancer-related malnutrition.58 The observed high Se and the ability to determine what elements are influencing nutrition status make PG-SGA a suitable nutritional screening tool. However, its accuracy is dependent upon the experience of an observer.60 Therefore, it requires a trained healthcare professional to complete the assessment and score the tool.58\n\nIn this study, the two tests yielded comparable estimates of PPV and NPV across the two populations based on the observed 95% PCIs. Therefore, if each test is used singly to screen for malnutrition, it is reliable and can inform treatment. Hence, the tool of choice is dependent on the purpose of the assessment, prognosis or even on the response to nutritional intervention33 bearing in mind the inherent shortcomings of each.\n\nThe prevalence of malnutrition among patients with cancer was high (˃ 54%). The findings are not unusual since cancer patients, particularly those suffering from head and neck, lung, and gastrointestinal cancer, carry the greatest burden of malnutrition among hospital patients.61\n\nAlthough the findings of this study are similar to those observed in Spain (50%)62 and Italy (51%),63 higher prevalences have been reported elsewhere. For example, malnutrition was found to be present in 76% of ambulant cancer patients getting radiation therapy in Brazil25; In London 71% of cancer patients were malnourished,64 and in Ethiopia, a prevalence of 90.6% in ambulant cancer patients on treatment was reported.65 The prevalence of cancer-induced malnutrition is often cited as 40-80%,5,66 which largely depends on assessment method, clinical setting, and case-mix of patients. Notably, the lower and upper PCI limits in this study fall within the quoted range (40-80%). With respect to the parameters of this study, the study focused on specific malignancies, there was no restriction on cancer stage, and the analysis did not take into account whether or not patients received treatment. Moreover, this study used a combination of MUST and PG-SGA to estimate prevalence, unlike the above studies that employed one tool. The above factors could have been the cause of the disparity between the results of this study and the above-mentioned studies.\n\nIn Kenya, in contrast to this study, lower prevalences of 31%6 and 13.4%17 have been obtained using the same population. Although lower, 31% falls within the lower PCI limit obtained at TCC in this study. The disparity could be ascribable to the different PG-SGA classification methods used. In their study, Opanga, Kaduka6 utilized the categorical classification, while this study employed the scoring method. The scoring method is more sensitive than the categorical classification, thus capturing more patients and leading to a higher prevalence.47 For example, Ræder, Henriksen47 recorded a Se of 50.0% and 60.7% for PG-SGA categorical and scoring classification methods, respectively. With regards to Kaduka, Bukania,17 the disparity could be explained by the different assessment methods used. A prevalence of 13.4%17 was based on BMI < 18.5 kg/m2 while in this study it was based on both the PG-SGA and MUST, which are superior to BMI.\n\nThe major strength of this study was using a Bayesian Latent Class Modelling framework, which minimized bias in test estimates, as it does not rely on a reference standard. A few limitations are inherent in this study that readers should be aware of while interpreting the findings. First, the PG-SGA is in form of a questionnaire targeting symptoms occurring within two weeks or one month of the time of screening. Therefore, the study participants may have failed to recall accurately, leading to either over-reporting or under-reporting of their symptoms. These may have biased the tool’s Se and Sp. Second, due to the study’s focus on specific cancers, findings may not be generalizable to other ambulant cancer populations. Third, patients who rejected taking part in this study did so due to pain, nausea, and weakness or because they thought, the study was too burdensome. Since these patients were more likely to be malnourished, this could have resulted in sample bias.\n\n\nConclusions\n\nUsing the Latent Class analysis, we have estimated the Se, Sp, and predictive values of both PG-SGA and MUST. The two tests achieved an accepted professional standard for Se (≥80%) and yielded good specificities. In respect to predictive values, the two tests produced comparable estimates. As such, healthcare workers are at liberty to use any of the two screening tools. Considering the high true prevalence observed in the two study populations, malnutrition screening among cancer patients should be done at diagnosis, during treatment and follow-up.\n\n\nData availability\n\nRaw dataset for the study is kept under restricted access since it contains sensitive participant information. Access to the raw data is possible upon placing a formal request to the corresponding author (carolinemuseka@gmail.com). The replication data and analysis scripts for this manuscript are available from the Harvard Dataverse.\n\nHarvard Dataverse: Performance of Malnutrition Universal Screening Tool and Patient-Generated Global Subjective Assessment in screening for cancer-related malnutrition in Nairobi County, Kenya. https://doi.org/10.7910/DVN/HS5YM1.52\n\nThis project contains the following underlying data:\n\n• R_Code_maln.R (Rscript for analysis)\n\n• Maln_data_xlsx (Analysis dataset)\n\n• Questionnaire.pdf (The questionnaire used for data collection)\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).", "appendix": "Acknowledgements\n\nThe authors wish to express their sincere gratitude to the study participants for contributing to the research.\n\n\nReferences\n\nZhang YH, Xie FY, Chen YW, et al.: Evaluating the Nutritional Status of Oncology Patients and Its Association with Quality of Life. Biomed. Environ. 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PubMed Abstract" }	[ { "id": "143589", "date": "15 Aug 2022", "name": "Oscar Ngesa", "expertise": [ "Reviewer Expertise Statistical Modeling", "Bayesian analysis", "Malnutrition Analysis" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nBelow is my review of the paper titled “Performance of Malnutrition Universal Screening Tool and Patient-Generated Global Subjective Assessment in screening for cancer-related malnutrition in Nairobi, Kenya”.\n\nThe study aims at evaluating the performance of the Patient Generated-Subjective Global Assessment (PG-SGA) and Malnutrition Universal Screening Tool (MUST) in screening for malnutrition among cancer patients in Kenya. The cancers of interest in this setting are head and neck, respiratory and gastrointestinal cancer. Bayesian Latent Class Model (BLCM) was employed in the analysis. It is a preferred approach in the absence of a gold standard.\nGenerally, the paper is well written in an appropriate language and easy to follow.\n\nBelow are my specific comments and queries.\nIn the introduction, put 39% to 87% instead of 39 to 87%. All citations at the end of each sentence should be placed before the period/full stop rather than after. This needs to be corrected in the entire document. The noun citation styles Kaduka,Bukania,17 and Ramboer, Verhamme57 do not seem to be correct. Confirm the style. Can the authors check for model stability and robustness of the estimates via sensitivity analysis? Changing the different priors. The authors mentioned that they would assess convergence of the models using visual appraisal of the density plots and Gelman-Rubin Diagnostic plots. However, none of these is reported in the document.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10506", "date": "07 Nov 2023", "name": "Caroline Auma", "role": "Author Response", "response": "Response to comment 1 Revised as recommended. Now reads 39% to 87%. Response to comment 2 We have revised this as advised. However, It looks like this is an error from the journal editors because, from the Word document that I submitted, the citations were correctly placed at the end. Response to comment 3 This has been revised to align with the rest of the in-text citations. Response to comment 4 Based on the three assumptions of the latent class analysis, we only met the minimum criteria. In particular, we only had two populations thus the constancy of Se and Sp estimates could not be assessed. Secondly, the two tests (MUST and PG-SGA) target different symptoms thus we assumed conditional independence. Finally, there was no prior information on any of the parameters in particular the Se and Sp. Therefore sensitivity analysis by modifying priors wouldn't be possible. Response to comment 5 We have included this as the last part of the results section. It reads: On the convergence of the MCMC chains, the density plots of all posterior estimates yielded unimodal distributions. Similarly, the G elman-Rubin diagnostic statistics for the parameters produced unitary ratios. These results suggest convergence of the chains." }, { "c_id": "10545", "date": "09 Nov 2023", "name": "Caroline Auma", "role": "Author Response", "response": "More to Comment 2 Regarding citations coming after the full stops, I noticed the same issue after version 2 was published. I engaged the editors and they said the change couldn't be made due to F1000 journal style." } ] }, { "id": "196451", "date": "26 Oct 2023", "name": "Mariana Kruger", "expertise": [ "Reviewer Expertise Oncology", "nutrition", "paediatrics", "ethics" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nCheck spelling and edit. Professional grammar editing will improve the readability of the text. Avoid long sentences with high density of information. Check the following and correct: “….cute disease effect score…” under Malnutrition Universal Screening Tool.\n\nWrite out abbreviations when used for the first time e.g., Se and Sp.\n\nIndicated clearly under the section “Test outcomes, sensitivity and specificity” what cut-off point was used for which test; either by adding respectively or linking the cut-off point to the test.\n\nPlease add the time needed to use both tests as it seems to be quiet time-consuming for screening tests, especially if the PGA-SGA should be done by the treating physician.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10507", "date": "07 Nov 2023", "name": "Caroline Auma", "role": "Author Response", "response": "Response to comment 1 I confirm that proofreading has been done to correct the grammatical errors. Response to comment 2 We have included this in the second last line of paragraph 3 in the introduction section: Sensitivity (Se) and Specificity (Sp). Response to comment 3 We have revised this by linking the cut-off to the test. As such, it now reads: The cut-off value of ≥1 for MUST and ≥4 for PG-SGA gave the best combination of the Se and Sp. Response to comment 4 We have supplied the information on the time needed for the tests: PG-SGA and MUST in paragraph 4 and paragraph 5 of the introduction section, respectively." } ] } ]	1	https://f1000research.com/articles/11-755
https://f1000research.com/articles/11-1269/v1	08 Nov 22	{ "type": "Research Article", "title": "Research on factors affecting tourist involvement in coffee tourism after the COVID-19 pandemic in Thailand", "authors": [ "Warach Madhyamapurush" ], "abstract": "Background: The world economy was broken by the COVID-19 pandemic, which affected the coffee industry. The COVID-19 pandemic's financial effects might influence equity markets and personal lives. This includes financial commodities like coffee, which the pandemic is predicted to damage. Coffee tourism is an emerging new kind of tourism in Thailand, formed in response to growing demand from visitors with a particular affinity for the beverage. Coffee tourism may contribute considerably to the expansion of Thai tourism if given the proper guidance and assistance. Methods: As part of a coffee tourism experience focusing on first-hand activities and information, tourists can visit neighbouring sites while on a coffee plantation. This research uses a stochastic neuro-fuzzy decision tree (SNF-DT) to analyse coffee tourism in Thailand. The research surveys 400 international and Thai coffee tourists. According to studies, Thai visitors mostly visit coffee tourism locations in Thailand for enjoyment. They also wanted to visit coffee fields in order to get personal knowledge of coffee production and marketing. Based on the comments of Thai visitors, coffee tourism in northern Thailand looks to be highly and effectively handled. Due to the same factor, responses from foreign coffee tourists indicated that many of their journeys to coffee tourism destinations were made entirely for enjoyment rather than the business. They also wanted to meet local tour guides and acquire handmade and locally produced things to understand more about coffee tourism.\n\nResult: According to study results, coffee tourism management in northern Thailand looks well-received by international tourists. We also compare the suggested model to the traditional one to demonstrate its efficacy. The performance metrics are prediction rate, prediction error, and accuracy. The estimated results for our proposed technique are prediction rate (95%), prediction error (97%), and accuracy (94%).", "keywords": [ "Coffee Tourism", "COVID-19 pandemic", "Foreign Tourists", "Tourist Behaviors", "Stochastic Neuro-Fuzzy Decision Tree (SNF-DT)." ], "content": "Introduction\n\nOne of Thailand’s key economic sectors, tourism, has consistently shown promise and development throughout time. Many travelers are looking to satisfy their need for a coffee-related experience to satisfy their taste for coffee. Such visitor demands provide Thailand the chance to distinguish its tourism goods via a distinctive brand identity, promote its cultures and distinctive characteristics, and work towards achieving global recognition. This coffee tour should let travelers appreciate coffee tourism. Tourists enjoy fresh coffee from its origin, the hilly environment, coffee farms, harvesting operations, bean roasting, etc. Tourism might become a national brand. The COVID-19 epidemic has had a significant impact on many areas of society. As the COVID-19 pandemic spreads, the financial impact on commodities markets and human lives is expected to be enormous and far-reaching. As a result of the pandemic, cash crops like coffee are also projected to be adversely impacted. Farmers in over 52 nations depend on coffee to make a living.1 The epidemic also impacts cafes and restaurants that get their ingredients from these industries since they must adhere to rigorous food and hygiene regulations. Cafes and restaurants must follow current standards and newly updated COVID-19 health guidelines even though there have been no reported incidents of the virus being transmitted via food in any nation.2 Coffee shops, among other food and beverage businesses, feel the effects of the new regulations, which many economists and analysts say are already severe. Major global businesses like tourism have been harmed by COVID-19’s unprecedented effects.3 Figure 1 depicts factors of tourism experience.\n\nTourism and hospitality enterprises benefit from the coffee industry in this region. Risk perception and health guidelines have considerably impacted this, resulting in a decrease in coffee foot traffic and an increase in at-home usage. Furthermore, this pandemic has already had a severe impact on the worldwide coffee supply chain, which has resulted in a changing price for the beverage.4 There is little doubt that coffee is one of the most important cash crops in the world today. A claim has been made that coffee tourism is part of the culinary travel industry. The variety of cultural experiences, distinct coffee-related customs, and unique coffee processing facilities in each location fueled brand growth for the destinations. For coffee tourists, collecting and amassing coffee-related experiences is one of the main attractions.5 The potential of coffee tourism for sustainable livelihood and conservation is examined by Sustainable livelihood in southwest Ethiopia. The mixed-methods approach used the memorable tourism experiences scale (MTES) to analyze the tourism experiences offered by coffees in Taiwan. More qualitative research on coffee tourism is required.6,31 A case study method is used to examine how COVID-19 affects the coffee industry and the accompanying consequences for tourism, as well as crisis management techniques for a post-pandemic environment in Taiwan. This study’s inability to completely implement the framework of the strategic approach is one of its weaknesses.7,32 Qualitative research was not efficient for the analysis of coffee tourism. An empirical study to determine the role of coffee tourism in livelihood based on real-time data is not performed To overcome this issue, we introduce a machine learning approach in this study.\n\nAs stated in Ref. 6, the coffee tourism system in every tourist spot must be continually improved, and measures should be taken to share information about coffee growth in Northern Thailand’s tourism sustainability.\n\nA comprehensive literature review technique for tea and coffee tourism has been proposed in Ref. 7, which aims to identify the opportunities and obstacles of expanding such niche products for locations. This study’s methodology uses an English-language literature review of the previous studies on tea and coffee tourism. 31 papers on coffee tourism and 33 papers regarding tea tourism have been examined in total. Many studies have used a single stakeholder’s viewpoint or the viewpoint of a particular coffee/tea tourism setting, like buyers’ or manufacturers’, according to the findings of this investigation. Because this research is the first of its kind, it showed potential suggestions for future work in coffee and tea tourism.\n\nCommunity-based tourism (CBT) is a growing topic of study, and this study8 provides a framework to understand it grounded in experimentally verified socio-psychological, phenomenological, and cybernetic conceptions of behaviour. One of the most significant outcomes of this study is a more structured knowledge of how socio-psychological variables might support and enable a successful CBT business plan to achieve a desirable “sustainable” goal.\n\nAnalyzing the characteristics that influence the success of coffee agro-tourism and also designing an efficient mechanism for community-based coffee tourism have been the goals of this research. A total of 142 people from the Sukaratu District agrotourism sector took part in.9 Coffee agrotourism throughout the Galunggung tourism region was shown to be sustained by the coffee plantation, according to the study findings.\n\nUsing data gleaned from a number of researches on the Thai coffee market, the author of this thesis makes recommendations for resolving the market’s current issues. According to the limited capacities of Ref. 10, inadequate data and analysis of information, the facts and actual conditions could vary, therefore more complete inquiries and analyses of the current system and problems are required, so as to put up more effective remedies and proposals. Although the Thai coffee industry has been thoroughly examined, there are still issues that need to be addressed and solutions found.\n\nDepending upon that extended TPB framework,11 intends to analyze the factors. Using the TPB structure as a starting point, this study provides a preliminary examination of the elements that may influence Thai consumers’ purchase intentions for certified coffee. According to the findings, self-identification has a greater impact on attitude than social identity, whereas attitude has the greatest impact on consumer buying behavior consequently.\n\nBased on an extensive theoretical map that shows how an unfunded community member had carefully conceptualized a system of actions and endeavors that have a significant impact on society,12 provides a practical example of a conceptually rich strategic vision. It is feasible and increased by attempts to expand common values with buyers, communities, and other relevant parties to establish sustainability-oriented principles.\n\nAnother study13 found that customer happiness and consumer loyalty are directly linked to service quality. As a result, cost fairness and consumer loyalty are influenced positively by a company’s ability to keep its customers happy. Additionally, client satisfaction serves as a bridge connecting service quality and satisfaction. In addition, perceived price can operate as a bridge between a company’s ability to satisfy its customers and maintain their loyalty.\n\nThis research investigated the drivers of Norwegian senior tourists’ desire to visit Thailand and analyzed a path model of variables impacting their travel intention throughout the pre-visit stage. This research hypothesized that travel motivation, expectation, limitations, destination image, and electronics word of mouth (e-WOM) impact travel intention. The authors also explored participants’ travel sentiment about Thailand. Analysis was done on 500 samples of eligible responders. Using Linear structural relations (Lisrel) software version of 8.72, a structural equation model (SEM) was run to evaluate the factors affecting tourism. The impact investigation was carried out to see how study constructs had an impact. About 62% of such variation in tourist perception is explained by its predictors.14\n\nIt was the focus of Ref. 15 to examine the possibility and possibilities of coffee tourism throughout Ethiopia’s south-western highlands. Here, we are looking at how sustainable tourism may support rural community livelihoods while also helping to protect biodiversity, particularly the wild Arabica coffee gene pool, in the montane forest. According to the findings, a fragile livelihood strategy and a desire to increase revenue are to blame for expanding agriculture into the forest and destroying the unique wild Arabic coffee gene pool.\n\nThere has been a rise in the popularity of agritourism in developing nations, which has been under-researched by academics.16 They are helping to further the conversation about the advantages and disadvantages of coffee tourism by conducting a study based on the perspectives of many stakeholders. According to the study, farmers reap the most benefits from empowering and cooperating, diversifying their businesses, and creating a more sustainable environment.\n\nAs per Ref. 17, coffee tourism can potentially improve locals’ lives. This is because there are new markets to enter and benefits to be gained. Local communities and the environment benefit from strong government responsibilities, which should be supported.\n\nIn Ref. 18, there are several ways in which innovation and coffee tourism are linked in Gangneung, South Korea. By using coffee tourism as an example of food and drink tourism, they examined how previous research has linked meals and drinks with creativity before explaining how tiny towns like Gangneung, South Korea, have been able to thrive as coffee tourism operators and how their use of creative thinking contributed to its success.\n\nThe Cibulao landscape/environment was positively impacted by tourism.19 Strong interactions between actors, resources, and the local governing structure led to sound economic and social performance. Enhancing the economy’s performance is essential as a vital component of sustainable use of resources led by good local governance regulation. According to the study’s findings, Community-based Coffee Tourism (CbCT) is further expanded as a green tourism technique to help the community’s economy while also improving the environment.\n\nThe purpose of Ref. 20 is to discover the most important factors influencing people’s desire to visit sustainable coffee and tea estates. In addition, the study examined the impact of travelers’ fears about contracting COVID-19 on their decision to visit coffee and tea tourism hotspots. A survey of 302 eco-conscious Gen Y and Z customers was conducted online using the idea of planned behavior as a foundation. The data were analyzed using partial least squares. Buyers’ opinions regarding healthy coffee/tea tourism have been influenced by their desire to learn and unwind. Regarding sustainable coffee/tea tourism, risk and mindset are the most critical factors. Consumers’ attitudes and behaviors toward a rapidly expanding type of tourism, which is taking place amid historically unprecedented conditions, can now be better captured by the planned behavior by including contemporary factors.\n\nThe article21 outlines Lviv’s coffee tourism growth. The intended market for coffee tourism has been determined. Characteristics and significant components of the resource basis for the growth and operation of coffee tourism in Lviv are described. The local community and business work together to build and promote specialized infrastructure, tourist products (goods and services), and coffee tourism events. It aims to establish a pleasant urban hospitality area for both locals and visitors and to exhibit Lviv’s 200-year-old multiethnic coffee culture and distinctive coffee urban environment to the globe. The article includes representative statistics, data on Lviv’s coffee tourist offer, and survey findings.22\n\nIn the coffee business, a reengineering effort focuses on identifying current innovations, evaluating existing ones, and designing new ones that can be implemented in a contingency. Changes, production, and transmission of information necessitate structural reengineering. Depending on the number of assets and external pressures, this is a new bet that will be made to rebuild the coffee industry; new jobs must be created, including those of a leader, communicator, and strategist.\n\nResearchers23 are interested in learning more about how to boost revenue during the COVID-19outbreak era in Central Aceh Regency by increasing coffee tourism, both physically and non-physically, using existing local expertise as a basis. Researchers used a descriptive qualitative research design and gathered responses from 40 coffee shop owners and patrons in various tourist destinations. Results revealed that the Central Aceh Regency offers significant potential for developing a coffee tourism model depending on the local tradition and enhancing income, particularly for those actively engaged in ecotourism development and coffee growers.\n\nThe issues faced by Thailand’s coffee farmers are explored in this research. Weather variations, rising labor rates, and an increase in plant diseases have contributed to several failures throughout the decades. Comparing the state of coffee tourist experiences from both the supply and demand sides was the primary focus of the investigation. Tourists are unpredictable. A data-organized questionnaire was circulated to the respondents to analyze coffee farmers’ concerns. Our proposed model can handle numerical and category data and manage issues with several outputs. As more data points are utilized for training the tree, it increases the forecast data exponentially while comparing with other existing approaches.\n\n\n\n• Stop word removal, stemming, dimensionality reduction, Min-max normalization is used for data preprocessing.\n\n• Linear Discriminant Analysis (LDA) is used for feature extraction.\n\n• Bat Algorithm is used for feature selection.\n\n• Stochastic Neuro Fuzzy Decision Tree (SNF-DT) is used for data analysis.\n\n\nMethods\n\nThe COVID-19 pandemic might affect the coffee tourism industry in four ways: first, changes in demand in major markets, including short- and long-term changes. Secondly, effects on coffee production in the area from rules influencing production capacity, inputs, or employees. Third, hurdles and conflicts in getting coffee beans to markets such as transportation costs, border checks, etc. Furthermore, the crisis brought structural changes in the global economy, such as diminished accessibility to commerce. A quantitative method to investigate the effects of coffee tourism from COVID-19 was used for this research. The COVID-19 pandemic’s unexpected effect has influenced everyone in the coffee industry. Risks such as these are linked to adverse effects on health and mobility, and the economy. Hence deep analysis of the factors affecting coffee tourism after a pandemic is required. Figure 2 depicts the overall methodology used.\n\nA survey was created to learn more about the behaviours, demands, and satisfaction levels of visitors who engaged in coffee tourism in the northern region of Thailand. Validity testing was performed on the completed questionnaires. Following that, the data were encoded and examined using statistical analysis tools. Stop word removal, stemming, dimensionality reduction, and min-max normalization are used for data preprocessing. Linear Discriminant Analysis (LDA) is used for feature extraction. Bat Algorithm is used for feature selection. Stochastic Neuro Fuzzy Decision Tree (SNF-DT) is used for data analysis.\n\nThe Human Ethics Committee of the Faculty of Tourism and Faculty Management Program at University of Phayao gave its permission to the research protocol approval number 2.2/005/64; the date of approval was 03/23/2021. All participants also gave oral consent to take part in the study and received information about the research’s methodology. This method of consent was approved by the Human Ethics Committee.\n\n6,485,791 Thai and international visitors who traveled to the provinces of Chiang Rai, Chiang Mai, Mae Hong Son, and Lampang made up the population for this research. This study’s sample, chosen using accidental sampling and convenience sampling, consists of Thai and foreign tourists on vacation in coffee tourism destinations in Chiang Rai, Chiang Mai, Mae Hong Son, and Lampang provinces who participated in coffee-related tourism activities such as a coffee farm tour or coffee tasting. Yamane (1973) method was used to compute the sample size at a 95% confidence level with a 0.05 margin of error.24,34\n\nAlthough the result was 398, it was rounded up to 400 to make data gathering easier. Additionally, a percentage is used to show how many responders there were in each region.\n\nVisitors from Thailand were asked to complete the questionnaire in Thai, while tourists from English-speaking countries were asked to complete the questionnaire in English. The questionnaire was in pen and paper format, and participants were required to complete the entire questionnaire. 2 participants who were approached to take part declined. Table 1 shows the sample groups in each province.\n\nA survey was created to learn more about the behaviors, demands, and satisfaction levels of visitors who engaged in coffee tourism in the northern region of Thailand. The survey consists of closed-ended questions about the respondent’s general characteristics, behaviors, needs, and satisfaction with the destination(s) for coffee tourism in terms of those destinations’ potential, with an emphasis on the attractions, accessibility, amenities, available package, activities, and ancillary services as defined by Buhalis’ 6As framework. The questionnaire included many options (based on a five-point rating scale) to help respondents adequately answer the closed-ended questions.\n\nThe questionnaire’s validity was thoroughly investigated. Each item on the questionnaire, primarily assessed by the thesis adviser, was scrutinized by five specialists. An Index of Item Objective Congruence (IOC) was used to measure the content validity and language suitability. The acceptable range is shown by the IOC score of 0.926.32 After this, the questionnaire was modified following the expert’s advice, before being delivered to the thesis advisor for final revisions. A total of 30 visitors who were not part of the sample group completed the finished questionnaire as a test run. Cronbach’s alpha coefficient was used to gauge reliability. The output was 0.948, regarded as statistically valid.33\n\nThe researcher used the validated questionnaire to gather data from the samples between April 2021 and March 2022. The information was gathered in several districts throughout the provinces of Chiang Rai, Chiang Mai, Mae Hong Son, and Lampang, where coffee tourism activities were planned.\n\nValidity testing was performed on the completed questionnaires. Following that, the data were encoded and examined using statistical analysis tools.\n\nData preprocessing using stop word removal, stemming, term weighting, and dimension reduction are included.\n\nIn many natural language processing (NLP) applications, stop word removal is among the most often utilized test sets. The idea is to effectively remove words that exist in all of the corpus papers. Stop words frequently include adjectives and pronouns. Stop words are often eliminated from the text before deep learning and machine learning models are trained since stop words are plentiful and provide little to no unique information for classification or clustering. Stop words may help to reduce the size of your index and query by removing them. When it comes to performance, fewer words are usually better. Because stop words are semantically empty, they have no effect on relevance rankings for data preprocessing.\n\nStemming is the process of reducing words to their base by eliminating extraneous letters, generally a suffix, and removing inflection. Porter and Snowball are two examples of stemming models. The findings may be used to find similarities and links in massive datasets. Stemming is the text preparation normalization job associated with eliminating word affixes in natural language processing (prefixes and suffixes).\n\nThe dataset was preprocessed using min-max normalization to give each element in the dataset a value ranging between 0 and 1. It establishes a data range by subtracting the greatest and lowest values of each data element. The least value of data elements is subtracted from actual data element in the numerator. The normalization of data is carried out according to equation 1.\n\nWhere xi refers to the original value of input dataset, xi′ denotes the normalized value, xmax and xmin are the maximum and minimum values of the data respectively.\n\nData transformation into numerical features that may be handled while keeping the information in the original data set is known as feature extraction. LDA is one of the most widely used linear algorithms for feature extraction. LDA is used to discover a lower-dimensional space that best distinguishes between data from various categories. The optimal way to separate data from various categories is to utilise a lower-dimensional space found by LDA. With this approach, the Fisher criteria, which is an objective function, is aimed towards maximisation:\n\nEach of these is referred to as an Inter- or Intraclass scattering matrix. tx=mx/m is the prior probability of a sample belonging to class x with expectation W. E may be derived by solving E∗= argmaxY(E) in solution space Ls×t=E∈Qs×tEPE=X. This may be achieved by solving the following extended eigenvalues reduction issue: Dve=λDee. The label information on the samples is used directly by LDA to solve classification issues.\n\nA predictive model’s input parameters are narrowed down via a process known as ‘feature selection’. BAT algorithm is used for feature selection.\n\nResearchers from different professions have been fascinated by bats, which have a unique capacity to echolocate. Bats, especially microbats, use echolocation as a kind of sonar: they generate a loud and brief pulse of sound, wait for it to strike an object, and then wait for the echo to return to their ears. In this way, bats may calculate their distance from an item. As a result of this remarkable orienting system, bats can distinguish a barrier from a prey item even in full darkness, enabling them to hunt. Bat algorithm is a novel meta-heuristic optimization approach based on observations of bat behaviour. To mimic the behaviour of a colony of bats that uses echolocation to locate prey and food, such a system has been devised. The following are some of the idealized principles used to represent this method:\n\n• When it comes to detecting distance and distinguishing between food and background obstacles, bats employ echolocation in a remarkable manner.\n\n• Searching for prey, an unidentified bat tj flew in random direction yj and velocity wj while emitting sound at the fixed frequency fmin and loudness of B0 at the given location and time. They may automatically alter the wavelength λ (or frequency) of their produced pulses and the rate of pulse emission s∈01, based on the closeness of their target.\n\nWhile it’s possible for the loudness to fluctuate in numerous ways, imagine that the loudness ranges from a big (positive) B0 to a minimal constant Bmin.\n\nThe Bat algorithm is shown in Algorithm 1\n\nObjective purpose fy,y=y1…yn\n\nSet up the bat population yjand wj, j=1,2,…,n.\n\nDefine the frequency of pulses fjat yj,∀j=1,2,..,n.\n\nSet up the pulse rates sj and the volume of sound Bj,j=1,2,…,n.\n\nWhile t<T\n\nFor every bat tj, do\n\nMake novel solution by equation 5, 6, 7\n\nIf rand>si, then\n\nChoose one of the finest options from the list.\n\nCreate a local solution based on the best one.\n\nIf rand<Bjand fyj<fŷ, then\n\nExplore novel solutions.\n\nHigher si and lower Bj\n\nSort the bats & discover recent finest ŷ.\n\nThe movement of the digital bats is determined by updating their velocity and location using equations 5, 6, 7 for every time step u, where U is the maximum number of iterations are as follows:\n\nBetaβ - random integer created between the values zero and one.\n\nyjku - at time step u, the value of the decision parameter k for the bat.\n\nThe results of fj are utilised to regulate the speed and range of bat movement. The variable ŷk reflects the current global best position (solution) for the decision variable k, which is produced by comparing all of the n bats’ answers.\n\nTo increase the range of feasible options, random walks are introduced. Firstly, one of the finest solutions currently available is chosen, and every bat that accepts the criteria has its own solution generated using a random walk.\n\nB¯u - overall bat noise level at t.\n\nFor each iteration of the algorithm, the loudness Bj and the emission pulse rate sj are updated, as follows\n\nEvery iteration of the process, the Bj and sj parameters are modified, as follows:\n\nsj0 and Bj0 are frequently chosen randomly.\n\nIn general, Bj0∈12 and sj0∈01.\n\nData analysis is the act of analysing, cleaning, manipulating, and modeling data in order to identify usable data, inform conclusion, and assist decision-making. Stochastic neuro fuzzy decision tree (SNF-DT) is used for data analysis.\n\nIncorporating neural learning algorithms into the feedback loop of hierarchical FDT (fuzzy decision tree) is the goal of stochastic neuro fuzzy decision tree (SNF-DT). The approach considerably increases the accuracy of classification of FDT without sacrificing the comprehensibility of the FDT structure. By returning from each leaf node to the root node, the back propagation learning method may be directly applied to the SNF-DT structure. A single forward cycle of FDT induction precedes numerous rounds of back propagation to fine-tune the FDT parameters in SNF-DT (membership functions and leaf certainties). Because of this approach, the hierarchical structure of the FDT tree isn’t disrupted, and the tree parameters may be tuned efficiently while still being interpretable. Figure 3 depicts the structure of SNF-DT.\n\nTwo summing nodes have been added to the basic SNF-DT structure in Figure 3 in order to perform inference. To determinez1, the certainty factors for class 1 (Yes) from each leaf node are added together. In the same approach, to determinez2., the certainty factors corresponding to class 2 (No) are added together. To determine the strength of the mth class discharging at the nth leaf node in any pattern, use the formula\n\nOn the basis of the input characteristics accessible in traveling from the root node to the nth leaf node, each pathnn=2…6 is created. μpathnjis the pathn membership degree. Which may be estimated as\n\nThe degree of confidence with which pathn can categorise class m is βnm0≤βnm≤1=12.\n\nFirepower of all leaf nodes belonging to a specific class is combined to determine the prediction confidence zmjm=12 of the jth pattern using FDT\n\nPredictions may be made by using the formula:\n\nSegmentation to a unique class requires selecting the classes with the greatest degree of membership, such as classifying a given pattern to categorise m0.\n\nThe class with the most accurate predictions will be chosen, m0=argmaxm=1,2z1jz2j\n\nTo fuzzify input attributes, the method selects Gaussian membership (GM) functions out of many alternatives due to its differentiable property. For jth pattern membership degree of pathn can be calculated by\n\nWhere the centre and standard deviation of GM of the kth input attribute on thenth route of Gkn is represented by Dkn and σkn, respectively.\n\nThe technique specifies that the FDT’s error function is a differentiable function like the mean square error F.\n\nIn SNF-DT, n is the number of training patterns, dmj and zmj are required classes for the jth training set.\n\nFor the error to be minimised, all variations about the parameter’s Gaussian centre locations, widths, and confidence factors must disappear. The parameter update rule is a consequence.\n\nFDT structures with Gaussian membership functions are updated using the following update rules: centers, widths, and confidence factors.\n\n\nResults\n\nIn this research, we examined the factors affecting tourist involvement in coffee tourism after the COVID-19 pandemic in Thailand. Following a thorough analysis of the sample group, which included both Thai and foreign visitors who engaged in coffee tourism in Thailand’s northern area, the following conclusions may be drawn. The parameters are satisfaction level, coffee tourism rate, prediction rate, prediction error, accuracy. The existing methods are KNN (k-nearest neighbors),26 naïve Bayes,27 BGVAR (Bayesian global vector auto regressive model,28 ANN (artificial neural network),29 SNF- DT (stochastic neuro fuzzy decision tree) [Proposed].\n\nThe survey was completed by 275 travelers who came to Thailand to visit coffee plantations. According to the replies, more than 59% of respondents overall were female, which is in line with demographic data. Between 21-30 years old, around 60% of the population. Around 58% of people held bachelor’s degrees. About 29% of respondents were students or university students, and about 75% were unmarried. 38% of those who responded said it was their first time visiting the coffee tourist locations in northern Thailand. 40% of the participants were situated in Bangkok, and 28% earned between 10,001 and 20,000 THB each month. 52% of tourists learnt about places online, and 92% supported internet advertising. 48% of tourists drove privately. 52% spent 1,000-5,000 THB for transportation. Over 75% of participants spent little more than 1,000 THB. 56% spent less than 1,000 THB for meals and drink. 53% spent less than 1,000 THB for lodging. 64% spent less than 1,000 THB on souvenirs. 68% of respondents spent no more than 1,000 THB. The majority of Thai visitors surveyed (67%), came to the places for pleasure, followed by 22% who wanted an educational experience and to sample fresh coffee from the coffee manufacturing facilities. 54% of visitors stayed 1-2 days. 77% wanted to come during coffee harvest season. 75% wanted coffee goods, especially instant coffee. Almost 47% wanted to learn about coffee tourism by visiting farms and factories. Tours to coffee estates drew 81%of participants and more than 84% of satisfied coffee tourists wanted to re-visit. Almost 90% of the examined travelers would suggest coffee tourism to others.\n\nSecond, the findings revealed demographic data, habits, requirements, and levels of satisfaction for all 125 foreign coffee tourists who responded to the questionnaire. In terms of demographic data, the results revealed that women made up the majority of the respondents, or more than 57%. Nearly 45% of the population was aged 21-30. About 33% had completed high school. 80% of them were unmarried and 41% of the population was either students or university graduates. Around 23% of the population was based in the United States of America. 44% of workers earned less than 2,000 USD per month. It was about 65% of respondents’ first time visiting a coffee tourism site. Nearly 33% of travelers said their tour guides taught them about the places. Nearly 37% reached the destination by airways. 36% of the examined foreign tourists said they have spent between 501 and 1,000 USD on transportation. 48% spent up to 100 USD on activities in the places. Almost 42% spent between 101 and 500 USD on food and drinks. 45% spent between 101 and 500 USD on lodging. Half of them spent up to 100 USD on souvenirs. Around 18% of participants spent 100 USD on souvenirs and they spent between 101 to 500 USD on other expenses.35% visited the places for recreation. Over 47% stayed one to two days. 56% wanted to come during coffee pruning season. 54% wanted local coffee-related items and handicrafts. 56% wanted to learn about coffee tourism from local guides. 54% wanted to see coffee farms and factories. In terms of satisfaction, approximately 54% of foreign coffee tourists want to return to the coffee tourism sites. Nearly 73% said they’d suggest the coffee tourism sites they visited.\n\nThe degree to which an individual’s actions provide them joy and fulfillment is referred to as satisfaction. The following Figure 4 represents the satisfaction level. In this graph, it includes both Thai and foreign visitors who came to Thailand’s northern area to explore coffee tourism. The satisfaction level of visitors includes attractions, accessibility, amenities, available packages, activities, and ancillary services. The overall satisfaction level of foreign tourists is much greater than Thai tourists.\n\nThailand’s coffee tourism management seems to be a big hit with Thai visitors, according to the findings. Like the Thai visitors, most of the international tourists surveyed were extremely pleased with the country’s coffee tourism administration. Table 2 shows the satisfaction level of both Thai and foreign visitors.\n\nAs seen by the average score of 3.8, the findings generally imply that Thai visitors are quite satisfied with the administration of the coffee tourism industry in Thailand. In a similar vein, the foreign tourists who participated in the study are generally quite satisfied with the way that coffee tourism is managed in Thailand (mean = 3.93).\n\nThe coffee tourism industry recommends travel to coffee farms. Coffee tourism in Thailand has a lot of growth potential because of Thailand’s abundant capacity to host visitors and manage its coffee resources. The following Figure 5 represents the impacts of COVID-19 in coffee tourism.\n\nTable 3 shows the impacts of COVID-19 on the coffee tourism rate. Table 3 illustrates that prior to COVID-19, 90% of tourists were travelling to Thailand for the purpose of coffee tourism. In the midst of the COVID-19 epidemic, this dropped to a rate of 30%, but after the pandemic, it increased to a rate of 70%.\n\nThe parameters are satisfaction level, coffee tourism rate, prediction rate, prediction error, accuracy. The existing methods are KNN (k-nearest neighbors),26 naïve Bayes,27 BGVAR (Bayesian global vector auto regressive model,28 ANN (artificial neural network),29 SNF- DT (stochastic neuro fuzzy decision tree) [Proposed].\n\nThe prediction rate is the percentage of situations in which the test findings return positive. The following Figure 6 represents the prediction rate. We evaluated the k-nearest neighbors with a prediction rate of 55%, the Naive bayes with a prediction rate of 68%, the Bayesian global vector autoregressive with a prediction rate of 76%, the artificial neural network with a prediction rate of 88%, and we proposed SNF-DT with a prediction rate of 95%. The results of the comparisons reveal that the suggested approach is superior to each of the four methods that already exist.\n\nTable 4 shows the prediction rate for suggested and traditional methods\n\nThe following well-known equation may be used to compute the percentage prediction error:\n\nThe following Figure 7 represents the prediction error. We evaluated the k-nearest neighbors with a prediction error of 58%, the Naive bayes with a prediction error of 65%, the Bayesian global vector autoregressive with a prediction error of 77%, the artificial neural network with a prediction error of 85%, and the proposed SNF-DT with a prediction error of 97%. The comparative findings show that the proposed technique is lower than the four existing approaches.\n\nTable 5 shows the prediction error for suggested and traditional methods.\n\nThe effectiveness of a classifier may be measured by counting the number of true positives (TP), true negatives (TN), false positives (FP), and false negatives (FN).30 In performance evaluation, sensitivity and specificity are two metrics that are frequently utilised.\n\nIt determines how many samples are successfully categorized. It determines how exactly the outcomes correspond to the original outcome.\n\nThe following Figure 8 represents the accuracy. We evaluated the k-nearest neighbors with an accuracy rate of 52 percent, the Naive bayes with an accuracy rate of 68%, the Bayesian global vector autoregressive (BGVAR) with an accuracy rate of 79%, the artificial neural network with a accuracy rate of 86%, and we proposed SNF-DT with a accuracy rate of 94%. The comparative findings show that the proposed technique outperforms each of the four existing approaches.\n\nThe Table 6 shows the accuracy for suggested and traditional methods.\n\n\nDiscussion\n\nCoffee farms in Thailand were new experiences for most of the participants in the study.31-33 Participants in the survey were more inclined to visit coffee enterprises in Thailand to learn about coffee tourist destinations. Coffee tourism hotspots should set up their own websites, according to the report, as a way to better connect with visitors.31,34 As a result, it seems that the capacity to accommodate visitors by launching interesting coffee tourism activities that respond to known data on tourist satisfaction in this sector would likely gain their pleasure and assure return visits.35,36 In this regard, it is important to understand visitor behaviour and demands so that successful marketing strategies for each province can be developed and implemented, and the necessary objectives may be met. In addition, the costs in coffee tourism sites are not exceptionally expensive since they are considered alternative tourist attractions. Inadequate knowledge about activities available in coffee tourist locations led them to arrange a one-day trip. Visitors to coffee tourism sites were more interested in having fun than in learning about the process of making and consuming coffee. The majority preferred to travel in the winter during the coffee harvest season, followed by those who wanted to visit in the rainy season for crop rotation. Consequently, most visitors to the coffee tourism attractions were located in the central area of Thailand, particularly in Bangkok. Because to the region’s lowlands, humid temperature, and short winter season, most Central Thailand’s residents spend their winters seeking a change of scenery and visiting tourist sites in other parts of the country.37 Most Thai visitors want to learn more about coffee by visiting coffee estates and seeing coffee production and processing. A portion of the research team wanted to discover local culture from local tour guides. Tourists were interested in coffee farm excursions, coffee brewing demonstrations and tastings, visiting neighboring sights, seeing coffee processing facilities, and learning about local lives, cultures, and customs. Most want to return to coffee tourism hotspots and recommend them to others.38 A massive dataset does not work well with KNN.26 The probability outputs are not to be taken too seriously since Naive Bayes27 is also recognized for being a poor estimator. BGVAR28 cannot simulate distributions if it is difficult to calculate the next-symbol probability. The construction of artificial neural networks is not predetermined by any rule according to ANN.29 To tackle this issue, we propose stochastic neuro-fuzzy decision trees for analyzing the coffee tourism. The findings show that coffee tourism is a popular pastime for coffee aficionados, and that many tourists enjoy a cup of joe while on vacation. Coffee- related souvenirs, tour packages, and coffee history may all be developed to improve tourism, and tourists are aware of this potential.\n\n\nConclusion\n\nCoffee farms have been hit hard by the current pandemic-induced change in consumer perceptions of risk and behavior. Economic, social, and health repercussions have been noticed and felt by both coffee tourists and its producers and customers alike. These methods allowed for a rapid reaction to pandemic in the coffee tourist. Data on the sample group’s visitor numbers in each province is collected and preprocessed. The linear discriminant analysis is used to extract the features, and the Bat method can be used to select the features. Stochastic neuro-fuzzy decision trees were utilized to analyze the coffee tourist data. Future research will focus on developing recommendations for coffee tourism management for local communities, adopting coffee tourism-related identities such as Robusta Coffee in the south of Thailand, and conducting a comparative study of coffee tourism administration among ASEAN and Asian countries.\n\n\nReporting guidelines\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).", "appendix": "Data availability\n\nfigshare: Research on factors affecting tourist involvement in coffee tourism after the COVID-19 pandemic in Thailand – Questionnaire. https://doi.org/10.6084/m9.figshare.21310734.v3\n\nThis project contains the following underlying data:\n\n- Coffee Tourism – Raw Data.xlsx\n\nfigshare: Research on factors affecting tourist involvement in coffee tourism after the COVID-19 pandemic in Thailand – Questionnaire. https://doi.org/10.6084/m9.figshare.21310734.v3\n\nThis project contains the following extended data:\n\n- Coffee Toursim - Questinnaire to Participants.docx\n\nfigshare: Research on factors affecting tourist involvement in coffee tourism after COVID-19 pandemic in Thailand. https://doi.org/10.6084/m9.figshare.20417568.v4\n\nThis project contains additional extended data.\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\ndel Rio-Chanona RM , Mealy P, Pichler A, et al.: Supply and demand shocks in the COVID-19 pandemic: An industry and occupation perspective. arXiv preprint arXiv:2004.06759.2020.\n\nWoolway M: State-by-state: COVID-safe requirements for dine-in.2020.Reference Source\n\nBaldwin R, Mauro BW: Economics in the Time of COVID-19. Centre for Economic Policy Research;2020.\n\nHernandez-Aguilera JN, Gómez MI, Rodewald AD, et al.: Quality as a driver of sustainable agricultural value chains: The case of the relationship coffee model. Business Strategy and the Environment. 2018; 27(2): 179–198. Publisher Full Text\n\nSetiyorini H, Chen T, Pryce J: Learning from the COVID-19 Pandemic: Media Representations of Responsible Coffee Tourism Practices in Indonesia. Community Empowerment, Sustainable Cities, and Transformative Economies. 2022; pp. 315–336. Publisher Full Text\n\nSmith N, Suthitakon N, Gulthawatvichai T, et al.: Creating a coffee tourism network in the North of Thailand. Local Economy. 2019; 34(7): 718–729. Publisher Full Text\n\nChen SH, Huang J, Tham A: A systematic literature review of coffee and tea tourism. International Journal of Culture, Tourism and Hospitality Research. 2021.\n\nTan CC, Sitikarn B: Coffee-and-Tea based and Social Entrepreneurship-Oriented Community-based Tourism (CBT) in Northern Thailand: Contributing towards a Theory.2018.\n\nJafaruddin N, Noor TI, Karyani T: Variables Influencing the Potency of Community Based Coffee Agro-Tourism in Mount Galunggung, Tasikmalaya, Indonesia. Pelita Perkebunan (a Coffee and Cocoa Research Journal). 2020; 36(3): 267–276.\n\nChuqian W: A STUDY ON THE SITUATION AND DEVELOPMENT OF THE COFFEE INDUSTRY IN THAILAND (Doctoral dissertation, SIAM UNIVERSITY).2018.\n\nUt-Tha V, Lee PP, Chung RH: Purchase Intention of Certified Coffee: Evidence from Thailand. The Journal of Asian Finance, Economics and Business. 2021; 8(8): 583–592.\n\nArayawut S: A Community-Based Case Study of Coffee Farming in Thailand. International Journal of Multidisciplinary in Management and Tourism. 2020; 4(2): 103–116.\n\nSan V, Kijkasiwat P, Abbasi A: UNDERSTANDING SERVICE QUALITY AND PRICE FAIRNESS TO CUSTOMER LOYALTY IN THE COFFEE SHOP INDUSTRY IN THAILAND. International Journal of Social Science Research. 2022; 4(1): 505–518.\n\nKvarme UK: The Effect analysis of influence factors on Norwegian senior tourists’ travel intention to Thailand. Journal of Rangsit Graduate Studies in Business and Social Sciences. 2020; 6(1): 167–184.\n\nWoyesa T, Kumar S: Potential of coffee tourism for rural development in Ethiopia: a sustainable livelihood approach. Environment, Development and Sustainability. 2021; 23(1): 815–832. Publisher Full Text\n\nCandelo E, Casalegno C, Civera C, et al.: A ticket to coffee: Stakeholder view and theoretical framework of coffee tourism benefits. Tourism Analysis. 2019; 24(3): 329–340. Publisher Full Text\n\nSetiyorini HD: Coffee tourism development potential: benefit and consequences. 3rd International Seminar on Tourism (ISOT 2018). Atlantis Press;2019, June; (pp. 154–157).\n\nSeo US: 2019. Coffee tourism as creative tourism: implications from Gangneung’s experiences. A Research Agenda for Creative Tourism.\n\nPrihayati Y, Veriasa TO: Developing green tourism to create the sustainable landscape: evidence from Community-based Coffee Tourism (CbCT) in Puncak, Bogor, Indonesia. IOP Conference Series: Earth and Environmental Science. IOP Publishing;2021, October; (Vol. 879(No. 1): p. 012027).\n\nYeap JA, Ooi SK, Ara H, et al.: Have coffee/tea, will travel: assessing the inclination towards sustainable coffee and tea tourism among the green generations. International Journal of Culture, Tourism and Hospitality Research. 2021; 15: 384–398. Publisher Full Text\n\nRutynskyi M, Kushniruk H: Coffee Tourism in Lviv in the, ontext of World offeeTourism. Annales UMCS, Geographia, Geologia, MineralogiaetPetrographia.2020; 75.\n\nHernández-Gracia TJ, Guillén JC, García-Lirios C: Reengineering in The Entrepreneurship of the Coffee Industry and Tourism in Central Mexico. Advances in Mechanics. 2021; 9(2): 63–81.\n\nSatria DI, Yusra M, Hilmi H: COFFEE TOURISM DEVELOPMENT STRATEGY BASED ON LOCAL CULTURE AS AN EFFORT TO INCREASE INCOME DURING THE COVID 19 PANDEMIC IN ACEH CENTRAL REGENCY. International Journal of Educational Review, Law And Social Sciences (IJERLAS). 2021; 1(2): 197–206. Publisher Full Text\n\nYamane T: Statistics: An Introductory Analysis. 3rd Edition.New York:Harper and Row;1973.\n\nSmith N, Suthitakon N, Gulthawatvichai T, et al.: The circumstances pertaining to the behaviors, demands and gratification in tourist engagement in coffee tourism. PSAKU International Journal of Interdisciplinary Research. 2019; 8(1).\n\nNamahoot CS, Panawong N, Brückner M: A tourism recommendation system for Thailand using semantic web rule language and K-NN algorithm. International Information Institute (Tokyo). Inf. Dent. 2016; 19(7B): 3017.\n\nNuritha I, Arifiyanti AA, Widartha VP: Analysis of Public Perception on Organic Coffee through Text Mining Approach using Naïve Bayes Classifier. 2018 2nd East Indonesia Conference on Computer and Information Technology (EIConCIT). 2018, November; (pp. 153–158). IEEE.\n\nSong H, Qiu RT, Park J: A review of research on tourism demand forecasting: Launching the Annals of Tourism Research Curated Collection on tourism demand forecasting. Annals of Tourism Research. 2019; 75: 338–362. Publisher Full Text\n\nChang JH, Tseng CY, Hwang RH, et al.: Using ANN to Analyze the Correlation Between Tourism-Related Hot Words and Tourist Numbers: A Case Study in Japan. 2017 IEEE 7th International Symposium on Cloud and Service Computing (SC2). 2017, November; (pp. 132–137). IEEE.\n\nSeyitoğlu F, Alphan E: Gastronomy tourism through tea and coffee: travellers’ museum experience. International Journal of Culture, Tourism and Hospitality Research. 2021; 15: 413–427. Publisher Full Text\n\nWang MJ, Chen LH, Su PA, et al.: The right brew? An analysis of the tourism experiences in rural Taiwan’s coffee estates. Tourism Management Perspectives. 2019; 30: 147–158. Publisher Full Text\n\nTeerasorn S: Basic Research. Bangkok:Chulalongkorn University Press;2009.\n\nKaiwan Y, Palaprom K: Basic of research. 5th ed.Bangkok:The Media Center for Bangkok;2010.\n\nHickey G, Grant S, Dunning J, et al.: Statistical primer: sample size and power calculations-why, when and how? European Journal of Cardio-Thoracic Surgery. 2018; 54(1): 4–9. PubMed Abstract \| Publisher Full Text\n\nNakamura RY, Pereira LA, Costa KA, et al.: BBA: a binary bat algorithm for feature selection. In 2012 25th SIBGRAPI conference on graphics, patterns and images.2012, August; (pp. 291–297). IEEE.\n\nJuvan E, Omerzel D, Maravić M: Tourist Behaviour: An Overview of Models to Date. (A paper presented at the MIC 2017 Conference Proceedings - Managing the Global Economy, Monastier di Treviso, Italy, 24-27 May 2017).2017.\n\nThe World Tourism Organization: UNWTO Annual Report 2011.2012.Reference Source\n\nPrince S: “Cohen’s Model of Typologies of Tourists.” In The SAGE International Encyclopedia of Travel and Tourism.2017. Publisher Full Text" }	[ { "id": "160144", "date": "01 Feb 2023", "name": "Ahmad R. Albattat", "expertise": [ "Reviewer Expertise Hospitality", "Events and Tourism Expert" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nPlease remove the word \"research on\" from the title\n\nAdd the recommendation / contribution at the end of the abstract\n\nKindly avoid using the very short sentences in the writing\n\nMust use the in-text citations in all the writing, don't leave any writing without citation\n\nRe-write the problem statement and no need to highlight the title \"Problem statement\" just add it to the end of the introduction and followed with the research objectives\n\nYou have two titles; methods and methodology, you need to use only one title to discuss your methodology part\n\nEthics not needed here, may use it in the acknowledgment section\n\nYou have mentioned that the data collection was made using the paper and pen; we are in the COVID situation, how was that possible?\n\nJustify why the validation was high for the instrument, what was the sources of the instrument?\n\nAdd recommendation to your conclusion\n\nUpdate the references\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "9317", "date": "14 Feb 2023", "name": "Warach Madhyamapurush", "role": "Author Response", "response": "Response to the Reviewer Please remove the word \"research on\" from the title Answer: Thank you for the comments. Based on the comments, the word research has been removed from the title and updated the revised title. Add the recommendation / contribution at the end of the abstract Answer: Thank you for the comment. Based on the suggestion, the recommendation and contribution of the study has been updated in the abstract. \"Major global businesses such as tourism have been harmed by COVID-19’s unprecedented effects. This study attempts to determine the role of coffee tourism in livelihoods based on real-time data using a machine-learning approach. More research is needed to analyze the factors of the coffee tourism experience using different machine learning approaches.\" Kindly avoid using the very short sentences in the writing Answer: Thank you for the comment. Based on the suggestion, the short sentences in the manuscript have been rewritten for flow and readability. Must use the in-text citations in all the writing, don't leave any writing without citation Answer: Thank you for the comment. Based on the comment, entire article has been checked for the in-text citations. Re-write the problem statement and no need to highlight the title \"Problem statement\" just add it to the end of the introduction and followed with the research objectives Answer: Thank you for the comment. Based on the comment, the problem statement word has been removed, and the paragraph was added after the introduction section. You have two titles; methods and methodology, you need to use only one title to discuss your methodology part Answer: Thank you for the comment. Based on the recommendation, I have used only “Methods” title to discuss the methodology part and updated in the manuscript. Ethics not needed here, may use it in the acknowledgment section Answer: Thank you for the comment. Based on the comment, the ethical section has been moved to the acknowledgement section in the revised manuscript. You have mentioned that the data collection was made using the paper and pen; we are in the COVID situation, how was that possible? Answer: Thank you for your comments. I would like to clarify that the work was not conducted during the peak pandemic but was undertaken during the relaxation phase and after the slow influx of tourists. The data collection was made using a paper and pen, we were wearing masks and was following social distancing and other COVID safety precautionary methods. Data collection was performed using strict COVID protocols. Justify why the validation was high for the instrument, what was the sources of the instrument? Answer: Thank you for the comments. Based on the comments, the details of validation have been updated in the manuscript. \"This study examined the conceptual model by collecting data from a self-administered questionnaire. For the purpose of certifying its validity, the survey instrument was pretested. For the purpose of ensuring content validity, 50 national and international tourists were tested. All constructs were tested for reliability (Cronbach's alpha greater than 0.70) and survey questions were confirmed to be reliable.\" Add recommendation to your conclusion Answer: Thank you for the comment. Based on the suggestion, the recommendations have been updated in the conclusion. \"Coffee tourism is becoming an increasingly popular travel experience, with people seeking to find the tastes of different cultures. Machine learning approaches offer the potential to explore the coffee tourism experience further. Using appropriate algorithms, researchers can gain a better understanding of consumer preferences and the experiences of those visiting coffee plantations. A predictive analytics approach can be used to analyze consumer data and identify patterns in purchasing behavior. This would allow researchers to understand better how tourists engage in coffee tourism. Additionally, sentiment analysis can be employed to analyze customer reviews and gain insight into the emotional aspects of their experiences.\" Update the references Answer: Thank you for the comments. References have been checked and updated." } ] }, { "id": "158786", "date": "01 Feb 2023", "name": "Vanessa G. B. Gowreesunkar", "expertise": [ "Reviewer Expertise Tourism management", "community based tourism", "island tourism", "marketing", "communication", "entrepreneurship" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis is an interesting piece and the topic of study is well justified. However, the way the paper is presented makes me doubt whether it qualifies for indexing. The work is not appropriately presented and structured as a research paper.\n\nThe aim and objective of the study are not clearly stated. There is no research question that guides the research problem. The literature review section is mixed with the introduction section and there is no clarity on the key themes that are examined (eg coffee tourism, the profile of the case study etc). If authors claim to have used case study as their research design, they do not justify their choice. Likewise, readership might be interested to know more about Thailand. The profile of the case study is therefore missing. The methodology section is another weak section which is not appropriately presented. The work is concluded in 5 lines.\n\nOverall, the work lacks substance and sounds like a mish-mash of information. In my opinion, authors need to do a major revision and re-organise the whole work.\n\nIs the work clearly and accurately presented and does it cite the current literature? No\n\nIs the study design appropriate and is the work technically sound? No\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? No\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [] } ]	1	https://f1000research.com/articles/11-1269
https://f1000research.com/articles/12-43/v1	11 Jan 23	{ "type": "Research Article", "title": "An analysis of the possibilities and challenges of long stay tourism in Thailand", "authors": [ "Warach Madhyamapurush" ], "abstract": "The tourism sector is significant in emerging nations like Thailand. The cost of lodging is a significant component of practically every trip, thus it is important to consider accommodation development while trying to draw in visitors from other nations. The long-stay tourism industry is crucial since longer visitor stays result in more revenue. Following this, other research on long-term lodging of all forms has been conducted, with an emphasis on both the tourist and real estate sectors. The best tourist option in Thailand is long-term travel. As evidenced by the American, European, and Japanese visitors, the target market is tourists from nations with high costs of living, frigid climates, and aging populations. Therefore, it is anticipated that the tourist demographic will change in future, leading to the emergence of the retirement home niche market as a part of long-stay tourism. The characteristics of long-stay tourism in Thailand are examined in this paper, and we assess the theoretical and conceptual framework as an analysis of Thailand's tourism. Examining the current situation of the Thai long-stay tourist business is the initial and main objective of this study. There is currently no perfect answer, but various alternatives from comparable markets in representative nations have been used as examples to subsequently create tourism accommodation in Thailand for long-stay tourism.", "keywords": [ "lon-stay tourism", "Thailand", "possibilities and challenges", "current status", "tourist business", "tourist attractions" ], "content": "1. Introduction\n\nOne type of tourism that has been popular for a long time is long-stay travel. Spain is without a doubt the most popular travel destination, but due to the increased globalization of the travel industry, growing regions like Southeast Asia are already showing remarkable market shares for long-stay travel.1 Apart from leisure, long-term travel is heavily influenced by the concept of a “second home,” which is common parlance among visitors with disposable income. As a result, the word “retirement home” is also closely associated because of the global socio-demographic trend, which foresees a significant shift in the proportion of the population that is older in several nations, including Japan and Scandinavian countries.2 The Japanese are a prominent foreign ethnic minority in Thailand and have a long-term presence there for a variety of reasons, including employment, travel, education, and retirement.1 Figure 1 indicates Thailand's international tourist arrivals.\n\nDue to the concern for an aging population, the retiree group is significant. Additionally, the Japanese government and the Japan & Thailand International Relations Organization (JTIRO) both strongly support this group. Japanese visitors and senior citizens have stayed in a variety of lodging options, including hotels, apartments for rent, and condominiums. In contrast to the Japanese, Europeans do not provide any kind of assistance to their elderly tourists. However, due to their potential and active efforts in reserving long-term lodgings in Thailand, European visitors, particularly those from Scandinavia, are a significant segment. Although there is no official database of long-term lodgings, many coastal tourist destinations have evidence of their presence along the shore. The Thai government investigated and conducted several efforts to encourage long-stay tourism, particularly the retirement segment, after realizing the importance of the sector. The 2004 campaign was unsuccessful thus far for a variety of reasons, and it is necessary to revise it to draw tourists and compete with other nations. In this study, the terms “long-stay accommodation” and “long-stay tourism” will be used synonymously for convenience.2 After the global economic crisis of 2008, it became abundantly clear that the tourist industry was a mainstay of the Thai economy. When the crisis first started, it didn't have much of an impact on Thailand. This was due largely to the fact that the tourism industry was responsible for a sizable portion of the country's GDP, and so provided a large number of people with employment opportunities.3,4 Thailand is a nation whose economy heavily relies on the travel and tourism sector. Thailand's natural beauty in Southeast Asia has earned it the nickname “Land of Smiles.” Many individuals in Thailand depend on the tourism sector to provide a living. Thailand, which is ranked as the ninth nation with the most money generated by tourism, has received THB 2000 yearly since 2014, according to the Tourism Authority of Thailand (TAT). Thailand also drew 32.6 million tourists globally in 2016, placing ninth overall and second in Asia. Thailand's foreign visitors have greatly expanded, and because of this quick expansion, the country placed fourth globally in terms of incoming tourists in 2015.5 Thailand's capital, Bangkok, continued to rank second among the most popular cities for international travelers. The fourth quarter of 2016 saw challenges for Thailand's tourism industry as a result of the Thai government's efforts to combat illegal immigration and impose various entertainment-related regulations. The growth of the tourist sector, however, remained steady. Additionally, the foreign tourist receipts for Thailand's tourism business were $49.9 billion in 2016 and $44.9 billion in 2015. In 2016, these earnings made up 12.3% of Thailand's GDP. This significant portion of the income from foreign visitors, primarily from China and Europe, is broken down as follows: Europe (THB 460 billion), China (THB 439 billion), America (THB 102 billion), CLMV2 (THB 97 billion), the Middle East (THB 5 billion), and other regions (THB 489 billion).3 The original and primary goal of this review is to examine the state of the Thai long-stay tourism market today. The link between the growth of long-stay accommodation and tourist trends will be studied to comprehend the market. Additionally, all appropriate tourist options will be illustrated to encourage long-stay tourism in the market.\n\nThe remaining section of the article is divided into five sections: Section 2 – Methods, Section 3 – Results, Section 4 – Discussion, and Section 5 –Conclusions.\n\n\n2. Methods\n\nThis section introduces prior research pertinent to the goals and objectives of the thesis. This section presents four components. First, the purpose of tourism reveals the nature and fundamental components of tourism.6 The role of demographic shift is then illustrated to highlight the anticipated future trend in tourism. Additionally, as lodging is a fundamental component of tourism, long-term lodging ownership is demonstrated. Domestic and foreign travel were both prohibited during the lockdown and preventive measures' implementation period, severely reducing earnings from tourism-related business. This revenue accounted for around 12% of Thailand's GDP and 50% of Thailand's export GDP, the latter being further impacted by the coronavirus disease (COVID-19) induced global slump.7 Finally, this section explains why government policy must regulate the real estate market. The conceptual foundation for the study is briefly illustrated as seen in Figure 2.\n\nThe 1992 Earth Summit in Brazil, which gave rise to the present worldwide emphasis on sustainable development, has acted as a driving force for the growth of tourism in three important areas. Protecting the natural resources and the environment is essential, followed by the need to continuously research the tourist industry, and finally, the need to develop human resources. All of these important factors have driven alternative tourism development to meet not only the recognized demands, but also to replace current tourism growth. Long-stay tourism is seen as an example of alternative tourism since it emphasizes language acquisition and intercultural interactions between visitors, who are viewed as guests, and locals, who are viewed as hosts. Because lodging is one of the three fundamental components of tourism—the other two being transportation and service—the tourist sector is related to real estate acquisition. The shared traits of the notion of a second home, which is based on two primary components, may explain long-stay tourism. Tourists may stay in privately operated, leased, or even free accommodations in this category. Second, they regularly go back to the same vacation spot. Their returns show that they are extremely knowledgeable of, devoted to, and appreciative of the location. To attract and keep visitors, tourism service information must be current, accurate, and easily accessible via a variety of digital mediums.8 As illustrated in Figure 3, Hall has offered three categories of tourism.8 Every visit, however, has the potential to serve many purposes and to straddle multiple categories, as in the case of a tourist who spends their winter vacation at their second home in a warmer country due to forced temporary migration (factors not related to tourism, academic and job opportunities).9 Accommodations that enable Muslim visitors to do the five necessary morning prayers, that must be done at specified times of day (before dawn, in the afternoon, in the afternoon, at sunset, and at night), are a much-appreciated addition to urban landscapes. With many prayer times spread out throughout the day, it's helpful for Muslim travelers to have a clean, well-maintained place to worship before continuing with their day.10\n\nOne month or more is regarded as a period of lengthy residence.11 Due to this circumstance, senior or pensioner travelers have emerged as one of the major target demographics for long-term travel. They are finished with their family caring duties and have no further work obligations.12 Additionally, they get regular income from pension funds granted by their government, including elderly visitors from Japan, the United States (US), and Europe. In a nutshell, the following are some elements that favor its development:\n\n• A general increase in household income\n\n• Longer vacations and more free time\n\n• Liberalization of financial flows\n\n• More affordable and effective transportation easing of border restrictions and facilitation of international travel.\n\nThe European Union is the second-largest market, with US visitors accounting for the largest portion of second-home tourism.13 There is a rise in demand for such residences in southern European nations, and northern Europeans frequently acquire them.14 One of the frequent customs in the area, like Scandinavian nations, is the ownership of second homes that are essentially household spaces. However, due to frequent travel in some nations, buying second houses or vacation homes overseas has grown in popularity. Spain is a favorite destination for tourists planning extended stays because of its proximity to the Mediterranean Sea, its cultural diversity, and its continued European character.15 At the same time, South American and Central American vacation spots like Costa Rica, Panama, and Brazil are prioritized by travelers from the US. More marketplaces exist now outside of western nations. The primary driver of the initial demand was the need for improved climatic conditions. Transferring to Asian nations, particularly Southeast Asian nations like Malaysia, the Philippines, Singapore, and Thailand, is a significant development area for this industry.16\n\nThe second-home vacation sector is dominated by travelers from the US, with visitors from the European Union making up the second-smallest marketplace. The demand certainly rises in the nations of southern Europe, while northern Europeans are the most common buyers of such properties (Pedro, 2006). One of the things that people in the region like Scandinavian nations have in common is the culture of having a second house that is mostly in the domestic area. On the other hand, because of the frequent travel required in some nations, purchasing a vacation or second home in other countries has become an increasingly common practice. Spain is frequently cited as one of the best places in the world for extended vacations due to the country's proximity to the Mediterranean Sea, its diversity of cultural traditions, and its preservation of a European ambiance. At the same time, travelers from the US tend to prioritize locations in South and Central America, such as Brazil, Costa Rica, and Panama. In today's world, there are a growing number of marketplaces that are located outside of western nations. The primary impetus behind initial demand was consumers' desire to experience more favorable weather conditions.14 The relocation of operations to Asian nations, particularly those in Southeast Asia like Malaysia, the Philippines, Singapore, and Thailand, is one of the most significant drivers of growth in this industry.\n\nAfter the homecoming of American soldiers from World War II, the term “Baby Boomer” was first used to describe Americans born between 1946 and 1964. Despite efforts to contain the COVID-19 epidemic by limits on international travel, these restrictions cannot be seen as viable long-term answers to the situation. When people are unable to freely explore a country, it can have devastating effects on the economy, including a deepening recession and a drop in tourism revenue in Thailand.17 The U.S. birth rate skyrocketed during that period, and they today make up an age range between 42 and 65 years. The word has been used widely since then to refer to those who were born during that period worldwide, not only in the US. While they are currently experiencing relatively slow demographic development, Europe and Japan will certainly experience some population loss in the first part of the twenty-first century. According to the United Nations, an estimated world population of up to 2300 was estimated, despite not being carried out by the primary area or specific nations. Except for Northern America, almost all areas predicted decreased growth rates. It is believed that the average lifespan will continue to increase indefinitely. Population aging emerges as a key demographic trait outside of the demographic window. Thus, it could cause international migration. While some seniors look for retirement destinations to maintain their pensions, the influx of foreign workforces will address the population and new generational decline.18 Analyzing the recent growth of Thailand's golf tourism sector reveals a market shift in the direction of focusing on improving the tourist industry's management mechanisms and expanding its use of environmentally friendly practices. Golf tourism in Thailand has been around longer than in China, and its development method and talents are more refined.14 There are several ways in which tourism affects the economies of host countries, including the creation of jobs, the building of infrastructure, the expansion of the tourism-related value chain, and other societal and economic ripples across the local population.19\n\nBased on research conducted by the Population Division, DESA, and the United Nations.17 The Japanese are a clear example of the rising elderly population. The government has a program to relocate most elderly pensioners to locations that are suited for them to live out their remaining years with a fixed amount of pension. They willfully purchase products and services that support independent living. Their worries about aging and death have sparked the creation of novel goods and services. The creation of specialized goods and services in the fields of health and medical care, home care, real estate, construction, financial services, education and learning, cuisine, cosmetics, travel, and entertainment are all part of the silver market. Most of the real estate and personal financial assets are owned by seniors, and when these assets are passed down through inheritance, there will be a greater need for new financial management services that include more investment than saving and making use of personal financial assets. Thailand, Malaysia, and the Philippines are three popular travel destinations. Given that Japan is a sizable trading partner and a popular tourist destination, it seems to sense that some of them would choose Thailand as their retirement residence. Currently, a bilateral company oversees other Japanese retirement complexes, and some Japanese families also maintain their private residences for both short-term accommodation and retired living.\n\nThe real estate sector, like many other economic sectors, is an essential component and the foundation of the tourism business. In terms of use patterns, the lines between real estate and tourism are becoming blurred. For instance, hotels operate as both long-term dwellings and workplaces for a mobile workforce of businesspeople, and resort areas provide both residential and recreational facilities to a frequently global populace. The hotel sector is a major topic of empirical examination of the globalization of tourism, particularly the real estate implications. Figure 4 represents the analogy of a timeline of the duration of stay.\n\n\n3. Results\n\nAs mentioned, the service sector, which is a key engine for the area, includes both the construction industry and tourism.20 Thailand's tourist prospects are shared by those of its neighbors. Despite the changes throughout this time, the TAT reports that a rise in the number of foreign visitors occurred practically year between 2010 and 2017. Figure 5 illustrates the international tourists from 2010 to 2017.\n\nFour distinct market categories have been singled out by the TAT as possible long-stay travelers visiting Thailand. The four groups of long-term visitors are as follows:\n\n• Retirees from different nations, with a focus on the Japanese market; the European market, including those from Britain, Germany, the Netherlands, and Scandinavia; the Chinese industry abroad, as well as from other countries.21\n\n• Snowbirds are visitors from chilly nations who will arrive in course of the wintertime. These folks have previously traveled to Thailand to spend two to three weeks taking part in beach activities, experiencing mountain and rural hill tribal life, and engaging in thrilling and daring events. Some people are choosing Thailand as a launching point to visit neighboring nations like Myanmar, Laos, and Cambodia (Indochina).22\n\n• Included in this category are international students and trainees. Numerous education systems choose to establish their affiliation or subsidiaries there to train foreign students in a range of areas due to Thailand's financial advantages. Even though students may not be big spenders, their participation will increase national income.\n\n• Those who go to sports training camps belonging to the fourth category. To conduct their programs at various locations around the nation, several sports training associations have traveled to Thailand with their coaches and players.\n\nAs a result, there are four kinds of long-stay accommodation in Thailand:\n\n• A resort or hotel\n\n• Apartments homes and serviced residences\n\n• a targeted health promotion effort\n\n• Accommodations for a certain Group\n\n3.2.1 Market overview for retail\n\nThe retail sector in Thailand may be divided into seven major groups based on size, characteristics, products sold, and price, according to a Colliers Thailand analysis.23 After the fourth quarter of 2004, the Bangkok retail market declined. It then rose in 2007, before sharply declining in response to the most recent financial crisis and the protests in 2008–2010. Despite reduced interest rates, the household continued to save due to anxiety over expenditures. Due to planned enhancement, the new future supplies are anticipated to be more fiercely competitive.24 The outdated malls are undergoing renovations or refurbishing to maintain their client base, which would likely cause a decline in occupancy throughout these changes. Figure 6 depicts the Thailand retail sales index. Bangkok's overall retail supply increased to 5.46 million m2, up 0.7% quarter over quarter (Q-o-Q) and 1.5% year over year (Y-o-Y).25 Nevertheless, a rise in customer confidence was accompanied by higher rental rates of 92% (4.97 million m2) accounting for 0.3% Q-o-Q but was still a drop of 1.5 % Y-o-Y. The following figures demonstrate as previously stated. Figure 7 indicates the demand for occupancy by region.\n\n3.2.2 Demand analysis\n\nMaterial obtained from the latest statistics is updated using estimates, while forecasts try to gauge potential changes. Increases in population and employment, facility relocations, speculative demand, and other factors are a few examples of demand drivers. A demand estimate for long-term housing is correlated with the number of visitors and retirees, just like in the tourism sector. A forecast will produce a projection using the number of tourists and demographic changes. The number of long-stay visitors is tracked by the arrival and departure stamps, but it is unclear how many long-stay lodgings there are because some properties are unwittingly acquired as private assets. Real estate items that developers had initially released for their locals but later became well-known in the tourism sector started to be purchased by tourists.26 International visitors and investors have undoubtedly been provided with local coastline and island tourism projects during the previous ten years. Since Thai owners are included in the total, it does not, however, provide an accurate indication of the level of demand from international buyers.\n\n3.2.3 Supply analysis\n\nAccording to TAT's four categories of long-term accommodations, most real estate items for sale fall into the “hotel and resort” and “condominium and service apartment” categories.27\n\n3.2.3.1 Resort and hotels\n\nIn conclusion, the amount of tourist lodging in this category is based on the “number of lodging places for tourists,” which adds up to four different types of lodging: hotel, resort, guesthouse, and bungalow. The following are succinct summaries of these four types:\n\n➢ A hotel is a type of housing that is especially designed, separated into rooms, equipped with useful services for tourists, and charges per room.\n\n➢ A resort is a location where guests may rent individual rooms or entire units while enjoying the natural surroundings.\n\n➢ A guest house is a home that has been altered or added to, with split rooms used for accommodation and a rent collection system.\n\n➢ A bungalow is a type of housing where guests can stay in groups, including businesses and tourists, and where rent is charged.\n\nFigure 8 represents the number of places available for tourist lodging from 2016 to 2020. We assess the BKK (BKK is IATA airport code for Bangkok) and vicinities, central, northern, eastern, northeastern, and southern. Figure 9 indicates the permits for high-rise residences, indicating the number of structures from 2016 to 2020.28\n\n\n4. Discussion\n\nUltimately, the study has demonstrated the core tourism potential of Thailand, including its tourist destinations, infrastructure, and lodging. Demand and supply for long-stay accommodations are anticipated to follow the tourist sector as an extraction from long-stay travel, as it formerly did with Spain's Mediterranean coast. Demographic factors are more likely to cause international migration. Tourists, especially frequent tourists, were accustomed to stealing foreign property. However, because of the lodging records' hazy ownership, the study was unable to determine how many accommodations were inhabited by foreign long-stay visitors. Due to ownership restrictions, the names of Thai spouses or Thai companies are frequently used as the owners of properties, which results in this issue. In our case study on Scandinavian visitors, the figure reveals that the number of visitors has been rising in recent years despite minor volatility over the previous 10 years. Interviews with current developers and examples of specific projects have demonstrated the ongoing business. Although Thailand has been a popular destination, it cannot claim to be the most prosperous country.29 The nation has recently been dealing with significant challenges including political unrest, economic fluctuations in the Thai baht, and rising oil prices.30 To further understand how people on minimum wage make sense of Simplified Employee Pension (SEP) and incorporate it into their conceptions of a livable wage, a qualitative study was conducted. Twelve workers at a riverside resort in Thailand were asked to participate in the study because they were representative of employees who had internalized SEP in the workplace.31 These challenges have the consequence that political instability prevents the implementation of consistent policies, such as those that promote long-term travel or even concentrate on senior housing. Additionally, political uncertainty reduces the trust of foreign investors, which reduces investment in other industries. Additionally, currency fluctuations will have an impact on travelers’ decisions to invest in foreign real estate. Growing building costs will also result in higher home prices and everyday living expenses due to rising inflation in Thailand or rising oil prices. As a result, in our example study, we can observe residential constructions from Scandinavian developers.32 In addition to tourists from Scandinavia,2,5 reports a noticeable increase in visitors from Russia, who also have a strong purchasing power; this is encouraging for upcoming tourist and lodging initiatives.33 In more detail, the need for standardized accommodations is brought on by the fact that potential tourists hail from a variety of nations and that many of them demand excellent quality and security due to the length of time they will spend in long-stay lodgings. The government has never established or initiated the initiative on its own, although it now offers conventional guarantee services and some market promotion.34 In some cases, tourism can be a significant factor in revitalizing a rural economy. To alleviate poverty in a remote community in Thailand's northeast, this article suggests implementing a tourism micro cluster model. Using principles gleaned from a comprehensive literature survey, the project also addressed the theoretical underpinnings of a tourism micro cluster model for a rural hamlet in Thailand.35 Developers with qualified operations related to tourism accommodations, such as hotels, retirement homes, care centers, dedicated health facilities, and long-stay businesses, can get investment incentives from responsible sectors like Board of Investment (BOI).36 Throughout this case study of Thailand, the author places special emphasis on the presenting of findings from quantitative and qualitative studies of urban tourism in the cities of Chiang Mai and Phuket.37 Only the private sector creates initiatives and engages in active marketing. Once more, for visitors from wealthy nations with considerable purchasing power, such as Scandinavian nations, the cost of a house is not their major issue; rather, it is the process of buying a property. In this study, we evaluated the theoretical and conceptual frameworks and examined the Thailand tourist industry\n\n\n5. Conclusions\n\nThis study promotes the notion of inviting long-stay tourists by outlining significant prospects and stated challenges to offer solutions to these issues. Ultimately, an analysis is meant to produce some helpful recommendations for practitioners and, ideally, future policymakers. Finally, it is necessary to gather data to develop a central database for long-stay accommodations; subsequent studies will then be able to use a more exact quantitative technique. Additional research should concentrate on significant aspects of public policy, such as the length of time a visa is valid, who owns the land, how tourist information is provided, and how tourism affects public relations. It is recommended that some case studies be researched, such as Thailand's neighbors Malaysia and Singapore, not only because these countries are Thailand's rivals but also because they are excellent examples. It is anticipated that the continued popularity of tourism in Thailand will be maintained by a development in this sector, which will in turn promote the economy of the nation.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\n\nReferences\n\nSopha C, Jittithavorn C, Lee TJ: Cooperation in health and wellness tourism connectivity between Thailand and Malaysia. Int. J. Tour. Sci. 2019; 19(4): 248–257. Publisher Full Text\n\nSunanta S:Globalizing the Thai 'high-touch’ industry: Exports of care and body work and gendered mobilities to and from Thailand. Thai-Western Mobilities and Migration. 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Publisher Full Text\n\nÇakmak E, Lie R, McCabe S: Reframing informal tourism entrepreneurial practices: Capital and field relations structuring the informal tourism economy of Chiang Mai. Ann. Tour. Res. 2018; 72: 37–47. Publisher Full Text\n\nSun K, Fang CC, Du H: Research on the Competitiveness of Thailand Golf Tourism Industry Based on Big Data. 2020 International Conference on Data Processing Techniques and Applications for Cyber-Physical Systems. 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Publisher Full Text\n\nEbrahim AH, Ganguli S: A comparative analysis of medical tourism competitiveness of India, Thailand and Singapore. Tourism: An International Interdisciplinary Journal. 2019; 67(2): 102–115.\n\nStatham P, Scuzzarello S, Sunanta S, et al.: Globalising Thailand through gendered ‘both-ways’ migration pathways with ‘the West’: cross-border connections between people, states, and places. J. Ethn. Migr. Stud. 2020; 46(8): 1513–1542. Publisher Full Text\n\nPhinaitrup BA: Health Tourism Management: The Case of Thai Health Tourism Industry. NIDA Development Journal. 2018; 58(3): 221–245.\n\nMayakul T, Kiattisin S, Prasad R: A sustainable medical tourism framework based on the enterprise architecture design: The case in Thailand. J. Green Eng. 2018; 8(3): 359–388. Publisher Full Text\n\nChantamart W, Thabhiranrak T: Factors Affecting Travel Behaviour of Thai Tourists at Don Wai Floating Market, Nakhon Pathom Province, Thailand. Proceedings of 17th Global.2018.\n\nDummanonda T, Nuangjamnong C: The influence of social media advertising value on consumer behavior in renting apartment rooms in Bangkok, Thailand. International Research E-Journal on Business and Economics. 2021; 6(1).\n\nLaw CC: The Relationship Between Air Transport and Rural Tourism in Thailand. Tourism: An International Interdisciplinary Journal. 2021; 69(3): 395–405. Publisher Full Text\n\nPratt G, Johnston C: Dementia care for Europeans in Thailand: A geography of futures. Am. Behav. Sci. 2022; 66: 1880–1895. Publisher Full Text\n\nLiu Y, Li Y, Parkpian P: Inbound tourism in Thailand: Market form and scale differentiation in ASEAN source countries. Tour. Manag. 2018; 64: 22–36. Publisher Full Text\n\nYodsuwan C, Pianluprasidh P, Butcher K: Against the flow: challenges in tourism development for a small-border town in Thailand. Managing Asian Destinations. Springer;Singapore:2018; (pp. 107–123).\n\nLai P, Jang H, Xu C, et al.: The impact of low-cost carriers on inbound tourism of Thailand. International Journal of Supply Chain Management. 2019; 8(3): 846–853.\n\nYoelao D, Mohan KP, Sombatwattana P: A qualitative construction of sufficiency living wage in Thailand based on the sufficiency economy philosophy. International Perspectives in Psychology: Research, Practice, Consultation. 2019; 8(4): 227–239. Publisher Full Text\n\nPitakdumrongkit K, Lim G: Neo-liberalism, the rise of the unelected and policymaking in Thailand: The case of the medical tourism industry. J. Contemp. Asia. 2021; 51(3): 447–468. Publisher Full Text\n\nLane LG: International Tourism as a Threat to Public Health in Thailand. Alpenglow: Binghamton University Undergraduate Journal of Research and Creative Activity. 2020; 6(1): 10.\n\nPalang D, Tippayawong KY: Performance evaluation of tourism supply chain management: the case of Thailand. Bus. Process. Manag. J. 2018; 25: 1193–1207. Publisher Full Text\n\nTapachai N: Applying a tourism micro cluster model to rural development planning: a case study of Kaeng Ruang village in Thailand. Часописсоціально-економічноїгеографії. 2019; 26: 45–54.\n\nHuang HONGYI: Research of the tourism industry development strategy in Bangkok of Thailand. Unpublished master’s thesis]. Siam University, Thailand.2019.\n\nKoodsela W, Dong H, Sukpatch K: A Holistic Conceptual Framework into Practice-Based on Urban Tourism Toward Sustainable Development in Thailand. Sustainability. 2019; 11(24): 7152. Publisher Full Text" }	[ { "id": "159823", "date": "06 Feb 2023", "name": "I Gusti Bagus Rai Utama", "expertise": [ "Reviewer Expertise Tourism Management" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nAbstract\nIn the abstract it is necessary to include the method used in this study. and also need to include the conclusions of the results of this study.\nIntroduction\nThe introduction has explained the fact that tourists with long stays are the dream of destination managers.\n\nConceptual framework\nTheoretical study is sufficient, it is only necessary to explain the difference between tourists on vacation and immigrants with the reason for traveling with the definition set by the UN-WTO.\nResults\nNeed to explain why the data analyzed is data from 2010 to 2017? This is to ensure that the data is still feasible for analysis and is still relevant to current issues related to length of stay.\nConclusions\nIn the concluding section, it is necessary to explain the contribution of the results of this research to science, and to tourism actors.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "159822", "date": "06 Feb 2023", "name": "Thanam Subramaniam", "expertise": [ "Reviewer Expertise Tourism experiences", "tourists behaviour", "tourism economics", "pro-environmental behavior", "ecotourism", "sustainable tourism", "place attachment." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIt is an interesting topic for the current tourism situation which is in the rejuvenation phase. However, there are a few limitations in the study where the need of the study (problem statement) is unclear.\nWhy has the author selected long-stay tourism in Thailand and what is the need for it?\n\nThe methodology of the study can be improved by clearly explaining how the data was obtained. Is this content analysis?\n\nThere is no literature review and underpinning theories in the study.\n\nSuggested having clear research objectives and answering the objectives through various outcomes.\n\nRecommended to have constructive arguments in literature review and outcome discussion.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "159819", "date": "06 Feb 2023", "name": "Anila Thomas", "expertise": [ "Reviewer Expertise Tourism Planning and Development", "Community-Based Tourism", "Sustainability practices and Tourism Development" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThere is a widespread belief that the expenditure of lodging is an important factor of nearly every vacation; thus, it's essential to think about accommodation expansion when attempting to attract visitors from other countries. Long-stay tourism is important relatively because long visitor stays contribute more revenue generation. The main target are tourists from countries with high living costs, extreme cold climates, and ageing populations. The tourist demographic patterns may expect to change in the coming years, causing the development of new markets, especially, home for retired communities, as an important segment of long-stay tourism.\nThe author emphasizes the presentation of findings from quantitative and qualitative studies of urban tourism in the cities of Chiang Mai and Phuket. The current research promotes the idea of encouraging long-stay tourists by identifying vast potential of the region and also indicated the various challenges faced by the service providers and communities in order to provide solutions to the mentioned problems. The sustained prominence of tourism sector in Thailand is expected to be balanced by further expansion of infrastructure and implementation of policy initiatives, that might enhance the country's economic growth.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-43
https://f1000research.com/articles/12-866/v1	21 Jul 23	{ "type": "Research Article", "title": "Multivariate data analysis: Validation of an instrument for the evaluation of teaching digital competence", "authors": [ "Andrés Santiago Cisneros-Barahona", "Luis Marqués-Molías", "Nicolay Samaniego-Erazo", "Catalina Mejía-Granizo", "Gabriela De la Cruz-Fernández", "Luis Marqués-Molías", "Nicolay Samaniego-Erazo", "Catalina Mejía-Granizo", "Gabriela De la Cruz-Fernández" ], "abstract": "Background: Technology plays a fundamental role to achieve higher education key learning objectives. Digital competence (DC) is defined as a set of skills, knowledge, abilities, and attitudes in technological aspects. It is necessary to employ an effective training action plan in higher education institutions to advance towards a level of teaching digital competence (TDC). The objective of this study was to validate the COMDID A instrument to assess Teaching Digital Competence (TDC) of active teachers, through a confirmatory factor and internal reliability analysis. Methods: The research was developed within a descriptive-correlational scope and a non-experimental-cross-sectional design to validate the dimensionality and reliability of the COMDID A instrument and evaluate the self-perceived digital competence of active teachers. The population was made up of 690 professors who were part of the teaching staff of the National University of Chimborazo, Ecuador, in the first academic period of the year 2021. The sample was probabilistic, in a simple random scheme, the percentage of potential error admitted was 3%. The representativeness of the sample was 50%, and the confidence level was 97%. A total of 511 teachers completed the questionnaire compared to the 452 individuals needed. Results: The instrument was robust, and it was reliable for the calculated sample. There were correlations between the variables, and the statistical calculation ensured the development of the multivariate analysis to validate the dimensionality of the instrument. Moreover, the correct dimensionality was determined through a confirmatory analysis and high reliability of the instrument. Conclusions: The calculated factorial scores were defined in order for further studies to be carried out. It is important to apply confirmatory factor analysis in educational technology research to validate the dimensionality of data collection instruments.", "keywords": [ "technological literacy", "teacher training", "educational research", "university teachers", "confirmatory factor analysis" ], "content": "Introduction\n\nIn 21st century society, which is digitally rich, the construction of a comprehensive and inclusive higher education is essential, one which attends to various actors’ needs and linking the university with society (Domingo-Coscollola et al., 2020). To accomplish this task, it is necessary for academic personnel to reach at least a medium DC level so universities must invest time and resources in the training of their professors (Amaya Amaya et al., 2018; Cisneros-Barahona, Marqués-Molías, et al., 2022b; Reyes, Cárdenas, 2018; Sánchez-Caballé et al., 2020).\n\nNowadays, there is a gap between teacher's skills and the deficient academic training they receive to achieve them. This is due to the confusion about the conceptualization of digital competences and then to the limitations of developing efficient digital training plans (Biel & Ramos, 2019; Malagón Terrón & Graell Martín, 2022), through public policies that strengthen the inclusion and treatment of these capacities in initial and continuous teachers’ training (Cabero-Almenara et al., 2021; Garita-González et al., 2019; Guillén-Gámez et al., 2021; Silva et al., 2019).\n\nThe importance of information and communication technologies (ICTs) in higher education lies in the improvement of teaching and learning processes through the inclusion of DCs, which also improve students’ training and professional performance (Fernández-Márquez et al., 2018; García-Ruiz et al., 2023; Juárez Arall & Marqués Molías, 2019).\n\nDC is defined as the integration of knowledge, skills, attitudes, capacities (Rangel, 2015; Vivar, 2014; Zepeda et al., 2019), whose purpose is the use of digital technologies in a responsible, safe, and critical manner (Ferrari, 2013).\n\nSystematic literature reviews have been carried out, supported by meta-analysis and bibliometrics, to explain the concept of teaching digital competence (TDC) and to categorize theoretical aspects that make it possible to interpret the evaluation effects easily and improve these skills (Cisneros-Barahona, Marqués-Molías, et al., 2022a; Cisneros-Barahona, Marqués Molías, et al., 2023a; Cisneros-Barahona, Marqués-Molías, Samaniego-Erazo, Uvidia-Fassler, & De la Cruz-Fernández, 2023d; Cisneros-Barahona, Marqués-Molías, Samaniego-Erazo, Uvidia-Fassler, De la Cruz-Fernández, et al., 2023; Delfín & Pirela, 2017; Gisbert Cervera et al., 2016; Marqués-Molías et al., 2016; Verdú-Pina et al., 2022).\n\nIn this regard, models with dimensions, standards, and indicators have been designed to evaluate DC levels from various perspectives using various instruments (Almås & Krumsvik, 2007; Beetham et al., 2009; Butcher, 2019; Caena & Redecker, 2019; Campo et al., 2013; CDEST, 2002; Elliot et al., 2011; INTEF, 2017; ISTE, 2000, 2008; Lázaro-Cantabrana et al., 2019; Palau et al., 2019; Redecker, 2017; Trilling, 2002), that have enabled, on the one side, the appreciation of the problems related to the deficiency of the formative aspects in teachers, as part of the strategies implemented to reach adequate levels of TDC (Angulo et al., 2015; Cisneros-Barahona, Marques-Molías, et al., 2022; Fernández Cruz & Fernández Díaz, 2016; Fernández-Diaz et al., 2021; Gutiérrez & Cabero-Almenara, 2016; Morales Capilla et al., 2014; Ramírez García & González Fernández, 2016; Silva Quiroz & Miranda Arredondo, 2020); and, on the other side, to generate more competent professionals (Juárez Arall & Marqués Molías, 2019).\n\nAccording to the relationships that the TDC has with other variables, there are studies that point out the importance of age (Cabero-Almenara et al., 2020; Usart Rodríguez et al., 2020a), generation (Basantes-Andrade et al., 2020) or gender (De la Iglesia et al., 2020; Zhao et al., 2021). In contrast, there are studies in which the variables of gender (Guillén-Gámez et al., 2021), age, and type of degree are considered inconclusive, and the relevance is assigned to the variable of the teachers’ attitude toward the use of technological tools (Galindo-Domínguez & Bezanilla, 2021).\n\nOn the other hand, the evaluation instruments must be reliable and valid to generalize their use in any context (Hernández Sampieri et al., 2014b; Larraz, 2013). Reliability is the degree to which repeated application of an instrument produces the same results, while validity is the degree to which an instrument measures a variable for real.\n\nThe COMDID A self-perception instrument (Lázaro & Gisbert, 2015a; Lázaro-Cantabrana et al., 2018), is a self-perception rubric that characterizes the teacher, relating him to the level of TDC. The instrument has four dimensions: 1. Didactic, curricular, and methodological (six indicators); 2. Planning, organization and management of spaces and digital technological resources (five indicators); 3. Relational, ethical and security (five indicators), and 4. Personal and professional (six indicators) and proposes five response options related to a rating scale for each of the 22 indicators (0, 25, 50, 75 and 100).\n\nThe COMDID A instrument has been through several design and development stages (Usart Rodríguez et al., 2020b): 1. Literature review on which the instrument is based; 2. Items design that are part of the questionnaire; 3. Validation by experts and, 4. Validation of factorial structure and internal reliability in relation to age, gender and access to the university in the version for initial teacher’s training (Lázaro & Gisbert, 2015b; Lázaro-Cantabrana et al., 2018).\n\nCronbach's alpha is an internal consistency measure and makes it possible to quantify the correlation that exists between the items that compound a scale (Cervantes, 2005; Cronbach, 1951; González & Pazmiño, 2015).\n\nFactorial analysis is a multivariate statistical technique, which seeks to obtain a reduced set of unobserved or abstract variables (common factors), which reproduce or represent the correlation shared by the observed variables. In other words, it makes it possible to facilitate the interpretation of a group of observed variables by reducing their number to a few that represent the common causes shared by the original variables, without losing the information (Mateos-Aparicio & Hernández Estrada, 2021).\n\nThis study validates dimensionally and in its internal reliability the COMDID A instrument to evaluate the self-perceived TC in active teachers.\n\n\nMethods\n\nEthical approval was obtained on December 23rd, 2021 from the Society and Environment Ethic Research Committee (CEIPSA (in Spanish)), Universitat Rovira i Virgili, CEIPSA-2021-PR-0035. All participants were asked to sign a written informed consent before enrolment.\n\nThe study aims to validate the COMDID A instrument to assess active teachers’ DC through a confirmatory factor analysis and internal reliability.\n\nThe scope was descriptive-correlational, and the design was a cross-sectional non-experimental (Arias, 1999, 2012; Arnal et al., 1992; Bisquerra, 1989; Bisquerra et al., 2009; Hernández Sampieri et al., 2014b; Ramos-Galarza, 2020).\n\nEquation 1 was applied to calculate the sample size to estimate the portion of the desired population with a known confidence interval (Badii et al., 2008):\n\nWhere:\n\nn: required minimum sample size.\n\nN: Population size.\n\nz: Z statistic for a level of confidence.\n\np: Expected proportion.\n\nq: Expected proportion.\n\nE: margin of error.\n\nThe population for this study was made up of 690 professors who were part of the teaching staff of the National University of Chimborazo, Ecuador, during the first academic period of 2021. The sample is probabilistic in a simple random scheme (Hernández Sampieri et al., 2014a; Kerlinger & Lee, 1985), the admitted potential error rate was 3%. The representativeness of the sample was 50%, and the confidence level was 97%. A total of 511 teachers completed the questionnaire, compared to the 452 individuals needed, according to the sample requirement (Badii et al., 2008) and, above the five samples per item required to confirm structures (110 samples according to the 22 items) (Hair et al., 2010).\n\nA scale from 0 to 100 defined the level of development of TDC in each dimension, with intervals: 1. Not started (N0), 2. Beginner (N1), 3. Medium (N2), 4. Expert (N3) and 5. Transformer (N4).\n\nThe reliability of the instrument was calculated through Cronbach's Alpha (Cronbach, 1951) using IBM SPSS Statistical Software, version 28.0.1.1(15). The dimensional constructs of COMDID A for active teachers were validated through confirmatory factor analysis, which also identified the latent factors that simplified the relationships established in the set of observed variables (López-Aguado & Gutiérrez-Provecho, 2019).\n\nThe intention was to confirm the structure of four factors that were related to the construction and theoretical validation of the instrument, through the principal component extraction method, with Kaiser-Meyer-Olkin (KMO) measure and Bartlett's test of sphericity; on a set of 22 indicators or items to try to reduce the amount of data observed and, thus, identify the four theoretical dimensions. A Varimax rotation was also used since they were orthogonal factors. The sample was 511 individuals, above the five samples per item required for this type of analysis (110 samples) (Hair et al., 2010).\n\n\nResults\n\nThe Cronbach coefficient was used to validate the instrument’s reliability as a statistic to estimate the reliability of any compound obtained from the sum of several measurements (Cronbach, 1951).\n\nThis validation was used as an analysis technique, in the second period of 2021, with the sample of 511 teachers. Results can be seen in Table 1, according to the dimensions of the instrument:\n\n• Dimension 1 (D1): Teaching, Curricular and methodological\n\n• Dimension 2 (D2): Planning, organization and management of digital technological spaces and resources\n\n• Dimension 3 (D3): Relational, ethical and security\n\n• Dimension 4 (D4): Personal and professional\n\nThe stages of analysis are (Mateos-Aparicio & Hernández Estrada, 2021):\n\n• Stage 1. Prior assumptions of the analysis.\n\n• Stage 2. Extraction of factors.\n\n• Stage 3. Rotation of factors.\n\n• Stage 4. Determination of factorial scores.\n\nStage 1. Prior assumptions of the analysis\n\nTable 2 shows the Kaiser-Meyer-Olkin (KMO) measure with a sampling adequacy of 0.974. Bartlett's test of sphericity presents the statistic value (7025.987), because of the low value of significance (0). Figure 1 shows the correlation matrix and its determinant close to 0 (8.310x10−7) (Cisneros-Barahona et al., 2023b).\n\nStage 2. Extraction of factors\n\nThe confirmatory factor analysis through the principal component extraction method and with the extraction criterion of a fixed value of 4 explained the variance value of 65.31%, see Table 3. Figure 2 shows the scree plot that indicates that four factors were viable according to the fall contrast criterion. Figure 3 reveals the Measure of Sampling Adequacy (MSA) index, through the values of the main diagonal of the anti-image matrices.\n\nStage 3. Rotation of factors\n\nAn orthogonal rotation is applied, using the Varimax method. In Table 3, it can be seen how the value of the total variance explained is the same for the non-rotated matrix and for the rotated matrix (65.316), even though the accumulated variances of each factor do not hold.\n\nStage 4. Determination of factorial scores\n\nThe communalities coefficients are shown in Table 4. Table 5 notes the score obtained in each of the cases of the extracted components to estimate factors.\n\nPrincipal component analysis: with one Varimax rotation for one extraction of four principal components, the rotation has converged in six iterations, and an explained variance greater than 65.316% was obtained (Table 3). Table 6 states the rotated components ordered according to the instrument factors. Values less than 0.3 are excluded for samples superior to 350 people by using SPSS (Hair et al., 2010).\n\na Rotation converged in six iterations.\n\n\nDiscussion\n\nInternal consistency reliability is a way to estimate the equivalence of the components among themselves, and it indicates the inner correlation between the variables of the instrument by separating the variation of the common factors and the variation of the unique factors of each item. In this sense, the reliability of the instrument was evaluated through the calculation of Cronbach's Alpha coefficient for the complete instrument, understanding an alpha calculation for each of the dimensions (Campo-Arias, 2006; Ledesma et al., 2002; Merino Soto & Lautenschlager, 2003; Torres, 2021), which gave the following results: For Dimension 1. Didactic, curricular, and methodological (α=0.836); for Dimension 2. Planning, organization and management of spaces and digital technological resources (α= 0.871); for Dimension 3. Relational, ethics and security (α=0.857), and Dimension 4. Personal and professional (α=0.891). The α of the complete instrument was 0.956, data that allows us to confirm that the instrument has high internal reliability.\n\nWhen observing the correlation matrix of the indicators, it was difficult to define for certain the number of correlation coefficients greater than 0.5 (Mateos-Aparicio & Hernández Estrada, 2021); because of that the determinant of the correlation matrix was used. If this value is closer to 0, it will imply a more significant association of the variables with each other, reaching the total dependence if it is 0 (all the elements of the matrix to 1). In the study, it was necessary to calculate the determinant since not all the values of the matrix were 1 (determinant = 0, total dependency), nor the values of the main diagonal at 1 and the rest at 0 (identity matrix) (determinant = 1, total independence). In our case, we have the relation to 1 on the diagonal of the correlation matrix for each variable with itself, and outside of this diagonal, the correlation coefficients of each pair of variables, with a calculated determinant of 8.310 x 10-7. At first glance, the determinant is quite close to 0. However, considering that the information comes from a sample and, to define an adequate degree of correlation between the variables, the Bartlett test of sphericity was calculated.\n\nBartlett's sphericity test proves the null hypothesis that the variables analyzed are not correlated in the sample, which means that it contrasts with the hypothesis that the correlation matrix is the identity matrix (the intercorrelations between the variables are zero, except for the main diagonal, which is 1). If this were true, there is no correlation between variables, and it would not make sense to do a factor analysis. Visually, the null hypothesis is rejected, since the correlation matrix is not the identity matrix, in fact, it is significantly different, which implies that there are high values of association. However, if the null hypothesis is not rejected for a level of significance, the variables would not be sufficiently correlated, and it would not make any sense to do a factor analysis. The high value of the statistic (7025.987) indicates that it belongs to the critical region, data that is confirmed with the low value of significance (0); these values allow the rejection of the null hypothesis. However, having a sample size greater than 100, the null hypothesis is always rejected since the sample size is predominant when calculating the statistic. To solve this problem, we chose the KMO measure, which compares the observed correlation coefficients with the partial correlation coefficients for all variables (Garmendía, 2007; Mateos-Aparicio & Hernández Estrada, 2021).\n\nThe structure of the instrument fitted the sample through hypothesis contrasts. The KMO measure showed a sampling adequacy of 0.974 that allowed us to be sure that the sample data were appropriate to perform a factor analysis (if it was higher at 0.90, it would be considered excellent sample adequacy of factorial data matrices (Kaiser, 1970)), between 0.8 and 0.9 means that the analysis is good or very good (Mateos-Aparicio & Hernández Estrada, 2021). Additionally, the value of the determinant of the correlation matrix was close to 0, which allowed us to confirm that the intercorrelation degree of the variables was quite high.\n\nWhen inspecting the sedimentation graph (Figure 2), it was observed that four factors (dimensions) were viable according to the falling contrast criterion since the inflection point was located where the eigenvalues stop forming a slope and begin to generate a low inclination fall from the fifth factor (Cattell, 1966; Hair JR et al., 2010; Pérez & Medrano, 2010).\n\nIn the anti-image matrix of Figure 3, the values of the complete matrix indicate the coefficients of partial relationships and explain the correlations not explained by the common factors. MSA is based on KMO; therefore, the interpretation of MSA in the main diagonal is like the coefficient. In this case, all the values were greater than 0.9, so the elimination of any variable was not considered, in addition to the fact that the elements outside the diagonal were less than 0.5 (Mateos-Aparicio & Hernández Estrada, 2021). It implies that the application of factor analysis was adequate in this sample (Garmendía, 2007).\n\nThe initial communalities in Table 4 measure the percentage of variance in a variable explained by all the factors together, and it can be interpreted as the reliability of the indicator (Garmendía, 2007). They appear in 1 because, in the principal component analysis (PCA), as many factors are calculated as original variables; this means that the total variance of the original variables is reproduced. The communalities are also observed after the extraction; the greater the communality, the better the variables will be represented by the factorial model. In this case, all the communalities were greater than 0.5, which means that they reproduced more than half of their variance, data that indicate that our variables were very well represented (Mateos-Aparicio & Hernández Estrada, 2021).\n\nA confirmatory factor analysis was developed, with the principal component extraction method, as it is the most appropriate method to estimate the factorial model and because of having the advantage of always providing a solution (López-Aguado & Gutiérrez-Provecho, 2019) as the factors explain the total variance correctly. The extraction criterion was a fixed value of 4 at the rate of each one of the dimensions of the questionnaire and through a Varimax rotation (it is the best known and applied method (Mateos-Aparicio & Hernández Estrada, 2021)) to minimize the number of variables with high load in each factor and to simplify the interpretation of the factors. This means that it simplifies the components to have high correlations with few variables and it is one of the properties of the Varimax method since the total variance explained before and after rotating is maintained, but not the total variance of each factor.\n\nAn explained variance effect of 65.31% was obtained to see the original structure of the instrument in the sample, with four factors in Table 3 (with a reduction of dimensionality from 22 to 4). This implies that there are enough factors (greater than 60%) (Hair et al., 2010) to determine that the factorial structure is correct for the sample, rediscovering the four theoretical dimensions. Thus, the importance of the application of confirmatory factor analysis in educational technology research is determined to validate the dimensionality of the data collection instruments.\n\nThe confirmatory factorial analysis determines that the factorial structure is correct for the sample, rediscovering the four theoretical dimensions (1. Didactics, Curricular and methodological; 2. Planning, organization, and management of spaces and digital technological resources; 3. Relational, ethics and security, and 4. Personal and professional (Lázaro et al., 2018; Lázaro & Gisbert, 2015b).\n\nIt was observed that not all the items had the necessary weights to be located univocally in a factor; this is due to the high association that the dimensions have concerning the formative aspects of teachers, the organization and management of resources, and strategic area (Usart Rodríguez et al., 2020b).\n\nFactorial scores were calculated (Table 5) for each case and in each of the extracted components to estimate factors, to carry out subsequent studies, and to replace the set of original variables with the set of principal components that represent them (reduced). In addition, it was observed that the instrument was robust for evaluating the DC of active teachers.\n\n\nConclusions\n\nThe reliability of the instrument was evaluated through the calculation of Cronbach's Alpha coefficient for the complete instrument, and we can confirm that the instrument is reliable for the calculated sample.\n\nBartlett's sphericity test explains the existence of correlations between the variables, additionally to the high statistical value, the significance was close to 0, and the value of the KMO measure and the MSA values ensure the possibility of developing a confirmatory factorial analysis to validate the dimensionality of the instrument.\n\nThe confirmatory factorial analysis determines that the factorial structure is correct for the sample, rediscovering the four theoretical dimensions (1. Didactics, Curricular and methodological; 2. Planning, organization, and management of spaces and digital technological resources; 3. Relational, ethics and security, and 4. Personal and professional) dimensionality (Lázaro et al., 2018; Lázaro & Gisbert, 2015b).\n\nIt is important to apply confirmatory factor analysis in educational technology research to validate the dimensionality of data collection instruments.\n\n\nConsent\n\nWritten informed consent for publication of the participants’ details was obtained from the participants.", "appendix": "Data availability\n\nZenodo: Underlying data for ‘Multivariate data analysis: Validation of an instrument for the evaluation of teaching digital competence. https://doi.org/10.5281/zenodo.8075442 (Cisneros-Barahona et al., 2023b).\n\nThe project contains the following underlying data:\n\n• Data File 1: spss data.sav\n\n• Data File 2: excel data.xlsx\n\n• Data File 3: data project factorial.xlsm (data from the principal component extraction method.)\n\n• Data File 4: data project reliability.xlsm (data showing the reliability of the instrument.)\n\n• Figure 1. jpeg\n\n• Figure 2. jpeg\n\n• Figure 3. jpeg\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nAlmås AG, Krumsvik R: Digitally literate teachers in leading edge schools in Norway. 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Reference Source\n\nRamírez García A, González Fernández N: Competencia mediática del profesorado y del alumnado de educación obligatoria en España. Comunicar: Revista Científica Iberoamericana de Comunicación y Educación. 2016; 49: 49–58.\n\nRamos-Galarza CA: Alcances de una investigación. CienciAmérica. 2020; 9(3): 1–6. Publisher Full Text\n\nRangel A: Propuesta De Un Perfil Digital Teaching Skills: a Profile. Pixel-Bit. Revista de Medios y Educación. 2015; 235–248.\n\nRedecker C: European framework for the digital competence of educators: DigCompEdu. Joint Research Centre (JRC) Science for Policy report. 2017. Publisher Full Text\n\nReyes J, Cárdenas M, Díaz Ocampo E: Las Competencias Digitales: una necesidad del docente Ecuatoriano del siglo XXI. Evista Dilemas Contemporáneos: Educación, Política y Valores. 2018; 12–26. Reference Source\n\nSánchez-Caballé A, Gisbert-Cervera M, Esteve-Mon F: The digital competence of university students: a systematic literature review. Aloma: Revista de Psicologia, Ciències de l’Educació i de l’Esport. 2020; 38(1): 63–74. Publisher Full Text\n\nSilva J, Usart M, Lázaro-Cantabrana J-L: Teacher’s digital competence among final year Pedagogy students in Chile and Uruguay. Comunicar. 2019; 27(61): 33–43. Publisher Full Text\n\nSilva Quiroz J, Miranda Arredondo P: Presencia de la competencia digital docente en los programas de formación inicial en universidades públicas chilenas. Revista de Estudios y Experiencias En Educación. 2020; 19(41): 149–165. Publisher Full Text\n\nTorres J: Fiabilidad de las escalas: interpretación y limitaciones del Alfa de Cronbach.2021. Reference Source\n\nTrilling B: 21st CENTURY STUDENT OUTCOMES.2002. Reference Source\n\nUsart Rodríguez M, Lázaro Cantabrana JL, Gisbert Cervera M: Validation of a tool for self-evaluating teacher digital competence. Educación XX1. 2020a; 24(1). Publisher Full Text\n\nUsart Rodríguez M, Lázaro Cantabrana JL, Gisbert Cervera M: Validation of a tool for self-evaluating teacher digital competence. Educación XX1. 2020b; 24(1): 353–373. Publisher Full Text\n\nVerdú-Pina M, Usart M, Grimalt-Álvaro C: Report on the process for evaluating and certifying Teacher Digital Competence An international perspective.2022. Reference Source\n\nVivar DM: Ramón Cózar Gutiérrez y Ma del Valle de Moya Martínez (coords.) (2013) Las TIC en el aula desde un enfoque multidisciplinar. Aplicaciones practices. Barcelona: Octaedro. ENSAYOS. Revista de La Facultad de Educación de Albacete. 2014; 29(2): 181–182. Publisher Full Text\n\nZepeda H, Méndez ME, Galván H: Evaluación de la Competencia Digital en Profesores de Educación Superior de la Costa Norte de Jalisco. Revista Iberoamericana de Producción Académica y Gestión Educativa. 2019; 6(11).\n\nZhao Y, Pinto Llorente AM, Sánchez Gómez MC, et al.: The impact of gender and years of teaching experience on college teachers’ digital competence: an empirical study on teachers in gansu agricultural university. Sustainability (Switzerland). 2021; 13(8): 4163. Publisher Full Text" }	[ { "id": "200607", "date": "12 Sep 2023", "name": "Yu Zhao", "expertise": [ "Reviewer Expertise ICT", "digital competence", "language learning and teaching." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis paper focused on the validation of an instrument for the evaluation of teaching digital competence. It is an interesting topic and the article is easy to read.\nHowever, there are some issues that need further improvement and revision:\nWhat's the difference between digital competence and teaching digital competence? Since COMDID focuses mainly on teaching digital competence, it should be mentioned in the introduction part.\n\nThe COMDID tool has been mentioned and used in several works, why did the authors choose this tool for validation?\n\nThe objective of this study is to validate the COMDID to assess TDC, but the findings do not seem to respond well to the purpose of the study.\n\nThe limitations of the study need to be mentioned.\n\nThe manuscript requires thorough proofreading.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nI cannot comment. A qualified statistician is required.\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10481", "date": "29 Nov 2023", "name": "Andres Santiago Cisneros Barahona", "role": "Author Response", "response": "What's the difference between digital competence and teaching digital competence? Since COMDID focuses mainly on teaching digital competence, it should be mentioned in the introduction part. In the introduction section, the difference between the definition of DC and TDC has been added. The COMDID tool has been mentioned and used in several works, why did the authors choose this tool for validation? In the Methods section, the justification for the choice of the instrument has been added The objective of this study is to validate the COMDID to assess TDC, but the findings do not seem to respond well to the purpose of the study. In the Results and Conclusions section, the relevant aspects that fulfill the research objective have been added. The limitations of the study need to be mentioned. The limitations of the study have been added in the Methods section. The manuscript requires thorough proofreading. The manuscript has undergone a comprehensive review." } ] }, { "id": "191131", "date": "22 Sep 2023", "name": "Francesc M. Esteve-Mon", "expertise": [ "Reviewer Expertise Educational technology", "digital competence and teacher training." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis paper discusses a relevant and important topic on the evaluation of teaching digital competence. It is well written and in general very clear.\nStill I have comments:\nThe authors provide a well-founded conceptual and state-of-the-art overview. However, there are some references that should be revised in the Introduction section. There are some citations that, although they may be related to the research, are a bit forced. These citations are not directly related to the authors' argument and should be reviewed. Furthermore, in the theoretical framework (Introduction), the link between digital competence (DC) and teaching digital competence (TDC) should be reviewed.\n\nAlso, the purpose of the article should be made clearer as the final part of the Introduction. Right now the aim is in the Ethical Considerations section of the Method. It should be restructured. In addition, the details of the instrument should be in the Method section.\n\nBeyond the analysis and discussion of statistical results, it is important to broaden the discussion in terms of progress and impact in the field of TDC. Overall, the manuscript makes some interesting points, but conclusions and implications could be extended.\nMinor comments:\nPlease review the acronyms used and define them at the first mention. For example, digital competence (DC).\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10482", "date": "29 Nov 2023", "name": "Andres Santiago Cisneros Barahona", "role": "Author Response", "response": "The authors provide a well-founded conceptual and state-of-the-art overview. However, there are some references that should be revised in the Introduction section. There are some citations that, although they may be related to the research, are a bit forced. These citations are not directly related to the authors' argument and should be reviewed. Furthermore, in the theoretical framework (Introduction), the link between digital competence (DC) and teaching digital competence (TDC) should be reviewed. Definitions linking TDC and DC have been added to the Introduction section. Also, the purpose of the article should be made clearer as the final part of the Introduction. Right now the aim is in the Ethical Considerations section of the Method. It should be restructured. In addition, the details of the instrument should be in the Method section. The research objective has been added to the end of the Introduction section. Additionally, in the Methods section, details regarding the chosen instrument have been included, along with the justification for its selection. Beyond the analysis and discussion of statistical results, it is important to broaden the discussion in terms of progress and impact in the field of TDC. Overall, the manuscript makes some interesting points, but conclusions and implications could be extended. The Results and Conclusions sections have been expanded to highlight the relevance of the study in relation to TDC. Minor comments: Please review the acronyms used and define them at the first mention. For example, digital competence (DC). The acronyms for Digital Competence (DC) and Teacher Digital Competence (TDC) are defined in the abstract of the article" } ] }, { "id": "191135", "date": "05 Apr 2024", "name": "Amaia Arroyo Sagasta", "expertise": [ "Reviewer Expertise Education", "Digital Competence", "Teacher Digital Competence", "Technopedagogy" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis article explains the validation process of the COMDID A instrument for the assessment of Teaching Digital Competence (TDC). It describes deeply the method used for reaching that validation, based on a descriptive-correlational scope and a non-experimental-cross-sectional design. The main conclusion highlights that the reliability is confirmed, also the dimensionality.\n\nAs a contribution to the article, I would add some more information in the introduction and conclusion sections. In the introduction, some more information about the COMDID framework would be appreciated for those researches who don't know much about it. In the case of the conclusion section, the last key idea can be developed and some other research lines for the future can be cited.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nI cannot comment. A qualified statistician is required.\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-866
https://f1000research.com/articles/12-385/v1	12 Apr 23	{ "type": "Research Article", "title": "Assessment of the antibacterial effect of Barium Titanate nanoparticles against Staphylococcus epidermidis adhesion after addition to maxillofacial silicone", "authors": [ "Yasir Mohammed Kareem", "Thekra Ismael Hamad", "Thekra Ismael Hamad" ], "abstract": "Background: Maxillofacial silicones are the most popular and acceptable material for making maxillofacial prostheses, but they are not perfect in every sense. To enhance their effectiveness, more improvements to their properties are required, such as their antimicrobial efficiency. This study assess the antibacterial effect of barium titanate nanoparticles in various percentages against staphylococcus epidermidis biofilm adhesion after addition to maxillofacial silicone. Methods: Barium titanate nanoparticles were added into VST-50 platinum silicone elastomer in four weight percentages (0.25wt%, 0.5wt%, 0.75wt% and 1wt%). 50 specimens were prepared and categorized into five groups; one control group and four experimental groups. All conducted data was statistically analyzed using (one-way ANOVA) analysis of variance, and Games-Howell multiple comparison test (significant level at p < 0.05). Shapiro-Wilk and Levene’s tests were used, respectively, to evaluate the normal distribution and homogeneity of the data. Result: One-way ANOVA test revealed a highly significant difference between all groups, and Games-Howell test revealed a highly significant difference between the control group and the four experimental groups. The 0.25wt% and 0.5wt% groups revealed a highly significant difference between them and with the (0.75%wt and 0.1%wt) groups. While the 0.75wt% group revealed a significant difference with 1wt% group. Conclusions: The addition of barium titanate to VST-50 maxillofacial silicone enhanced the antibacterial activity of silicon against Staphylococcus epidermidis, and this activity seems to be concentration dependent. FTIR analysis demonstrated no chemical interaction between the Barium Titanate and the VST-50 maxillofacial silicone elastomer. SEM pictures show that the barium titanate nanopowder was effectively dispersed inside the maxillofacial silicone matrix.", "keywords": [ "barium titanate (BaTiO3)", "VST-50 silicone elastomer", "bacterial adhesion", "Staphylococcus epidermidis." ], "content": "Introduction\n\nThe restoration of abnormalities resulting from inherited or acquired causes, such as cancer or trauma, has often utilized prosthetic devices. Due to the location and extent of the lesion, surgery may not always be a solution; hence, the need for prosthetic rehabilitation has grown correspondingly.1\n\nAny abnormality that develops in the body, especially in the head and neck area, has a negative impact on the patient’s attractiveness, function, social acceptability, and psychological confidence. The most vital and difficult part of these individuals’ therapy is rehabilitation. Any rehabilitation process attempts to reintroduce the patient into society in a condition that is close to normal. A facial prosthesis preserves the tissues of a defect, restores normal anatomy and appearance, and offers the patient significant psychological benefits.2\n\nMaxillofacial prosthetics can be made from a variety of materials, such as chlorinated polyethylene, polyvinyl chloride, polyurethanes, polymethyl methacrylate, and polydimethylsiloxane.\n\nSilicone elastomers have become more important in medicine and the construction of maxillofacial prostheses because of their strength, durability, ease of manipulation, esthetics, and flexibility.1,3 Currently, no facial prosthetic material, including silicone, satisfies all the requirements for a satisfactory prosthesis. The primary cause of routine facial prosthetic replacement is deterioration in appearance caused by changes in physical characteristics and color.3 Therefore, silicone maxillofacial material requires reinforcement.\n\nDuring the development of the nanoparticle industry, nanoparticles have been incorporated into the polymer matrix as fillers to provide a modified polymer characterized by improved features gained from the reinforcing action of the nanoparticles. The expected mechanical, physical, and biological properties of a silicone elastomer depend on the type and amount of filler added to the polymer. These little additions could enhance certain characteristics of the material.1\n\nBiofilm formation on the surface of maxillofacial prostheses is one of the most critical problems. Biofilms are formed due to various reasons, such as fungal, bacterial, and commensal microflora. These microbes have a clear association with reports of bacterial dermatitis and endophthalmitis.5 Among the different species found, the most frequent have been Staphylococcus aureus and Staphylococcus epidermidis. The major limitation of maxillofacial silicone is that it has numerous porosities on its surface that are colonized by these microorganisms.4,5\n\nStaphylococcus epidermidis is the most prevalent commensal bacteria on human skin. Although S. epidermidis defends us against foreign invasion, it also takes advantage of human weakness when it has the chance. Such chances appear in immunocompromised people or when biomedical implants provide a chance for surface colonization and biofilm formation.6\n\nThe physical rubbing or brushing of maxillofacial prostheses is one method of disinfection, although it is not perfectly advised since the repetitive cleaning might roughen the material’s surface. Similar chemical immersion, for example, repeated use of chlorhexidine gluconate CHX, may change the physical and mechanical characteristics of maxillofacial silicone elastomers, resulting in roughness, color change, and an increase in microhardness.5\n\nIncorporation of a nanoparticle such as Barium Titanate (BaTiO3) may enhance the antimicrobial and other properties of maxillofacial silicon. Barium titanate (BaTiO3), a dielectric/ferroelectric semiconductor, is the most extensively used photocatalyst in environmental and medical applications due to its low cost, chemical stability, biocompatibility, and non-toxicity. BaTiO3 has been proven to accelerate osteogenesis, and the same material in nanoparticle form acts as a second harmonic generation (SHG) probe to identify Osteogenesis Imperfecta.7,8\n\nBaTiO3 had shown antibacterial activity against numerous types of bacteria when added to different materials such as polyvinylsiloxane, hydroxyapatite, and implants.9,23 This study aimed to evaluate the effect of BaTiO3 on S. epidermidis biofilm adhesion after addition to VST-50 maxillofacial silicone in various weight percentages.\n\n\nMethods\n\nBarium Titanate (BaTiO3) (Sky Spring Nanomaterials, USA) and VST-50 room temperature vulcanized silicone (Factor II Inc., USA) were used.\n\nParticle size analyzer was used to verify that the BaTiO3 particles are at the nanoscale, and the effective diameter was (59.4 nm).\n\n50 specimens were prepared and categorized equally into five groups: one control group (0wt% BaTiO3) and four experimental groups (0.25wt%, 0.5wt%, 0.75wt%, and 1wt% BaTiO3) 10 specimens for each group.\n\nThree clear acrylic sheets (the matrix, bottom, and cover) with 2 ± 0.05 mm thickness were created. The matrix sheet was designed with 10 mm disk-shaped perforations and was glued to the bottom sheet by chloroform (glue material) to avoid its moving while silicone was being poured. Using a computer’s software (CorelDraw 2020) to design the mold and a CNC machine to fabricate it. Clamps, screws, and nuts were also used for further tightening at the edges,1,10 an alternative open-source software is FreeCAD.\n\nIn accordance with the manufacturer’s instructions, the VST-50 maxillofacial silicone is mixed at a ratio of 10:1 (10 parts base to 1 part catalyst). A vacuum mixer had been used to prevent air entrapment.\n\nSpecimens for the control group were mixed by using an electronic digital balance (to 3 dp) for weighing the base and catalyst, then mixed for 5 minutes.\n\nFor the experimental groups, BaTiO3 filler was first weighted and added to the bowl, followed by the addition of the base part to the filler. The mixture was mixed for 3 minutes without vacuum to avoid suction of the filler, followed by 7 minutes of mixing with air suction. The vacuum pressure is set to -10 bar (-28 inch Hg), and the speed is set to 140 ± 10 rpm. The mixture was then allowed to cool for 5 minutes. The catalyst was then added to the base-filler mixture and mixed for 5 minutes.11,12\n\nThe mixture was poured into the mold, and the cover part was sealed over it. The mold was tightened by screws, nuts, and G-clamps. The mixture was left to set at (23°С ± 2°С) for 24 hours according to the manufacturer’s instructions. The specimens were stored at 20-25°C, 50 ± 10% humidity, and for 16 hours according to ISO 23529:2016 (Figure 1F).13\n\nS. epidermidis was isolated from three patients using sterile transport cotton swabs. By rotating the transport swab across the contaminated skin region, necrotic tissue was avoided.14 It was then inoculated into blood agar and mannitol salt agar prepared according to the manufacturer’s instructions in an aerobic condition at 37 °C for 48 hours (Figure 1A and B).15 Identification of S. epidermidis: they form grayish-white, elevated, round, smooth, cohesive, 1–2 mm in diameter non-hemolytic colonies. They showed positive results in the catalyst test, and bacterial species were verified using the VITEK 2 compact identification system.\n\nBacterial adherence test\n\nThis test was performed to evaluate the antibacterial activity of BaTiO3 against S. epidermidis, depending on optical density (OD) measurement using a spectrophotometer (APEL PD-303, Japan) set at 600 nm.16 Brain heart infusion broth was used to grow and create the bacterial suspension. It was prepared according to the manufacturer’s instructions by suspending 34.5 grams of powder in one liter of distilled water and dissolving it completely, then autoclaved at 15 lbs. of pressure (121°C) for 15 minutes. Then a suspension of 107 colony forming units (CFU/ml) (0.5 McFarland standards) was prepared using a McFarland densitometer. The silicone specimens were sterilized for 20 minutes in an autoclave at 121°C. The sterile silicone specimens were placed in a sterile plastic dish containing the produced bacterial solution and incubated at room temperature for one hour (Figure 1D).10 Following completion of the incubation time, the specimens were withdrawn from the suspension, rinsed twice with phosphate-buffered saline for one minute with gentle rocking to remove any non-adherent bacterial cells, and dried on filter paper.10 The specimens were then stained by 1% crystal violet for 10 minutes and rinsed well in phosphate-buffered saline (Figure 1C).17 Each specimen was immersed in 3 ml of 96% ethanol alcohol for 3 minutes; this solution was then used to confirm the optical density of each specimen (Figure 1E).18\n\nFourier transforms infrared spectroscopy (FTIR)\n\nFTIR (IRAffinity-1 laser product, Shimadzu, Japan) was utilized to verify if silicone material and the BaTiO3 nanoparticles interacted chemically. Three samples, one from each group, were examined. (Control, 0.5wt% and 0.75wt%).\n\nField emission scanning electron microscope (FE-SEM)\n\nThe scattering of BaTiO3 nanoparticles within the silicone specimen matrix was evaluated using a FE-SEM (FEI, Netherland) machine. Three samples were tested, one from each group (control, 0.5wt% and 0.75wt%).\n\nThe statistical analysis was performed using one-way ANOVA (analysis of variance) and post hoc tests (Games-Howell) by statistical analysis software (IBM SPSS Statistics 23, a proprietary free alternative we can suggest is PSPP). The Shapiro-Wilk test was used to discover the normality distribution of data, and Levene’s test was used to discover if the variances were homogenous.\n\nThe probability (P) value was considered non-significant statistically (NS) when (P > 0.05), while P value was considered statistically significant (S) when (P ≤ 0.05), and P value was considered highly significant (HS) when (P ≤ 0.01).\n\n\nResults\n\nFTIR Results: There was no change in the spectra range of VST-50 silicone by the incorporation of BaTiO3 (no chemical interaction) as shown in Figure 2.\n\nFE-SEM result: the BaTiO3 nanoparticles were evenly distributed throughout the VST-50 silicone matrix in the FE-SEM images, with slight agglomeration as filler loading increased, as shown in Figure 3. FE-SEM showed reduced silicone porosity.\n\nThe Shapiro-Wilk test revealed a normal distribution of data around the mean (P value ˃ 0.05) (Table 1).\n\nThe descriptive statistic revealed a decrease in the mean of optical density (OD) as the concentration of BaTiO3 increased, which represented a decrease in bacterial adhesion (Table 2), as shown in Figure 4.\n\nOne-way ANOVA test revealed a highly significant difference in the mean values among all groups (P < 0.01) (Table 3).\n\nTo choose the type of multiple comparison post hoc test and assess the homogeneity of variances, Levene’s test was used (Table 4).\n\nGames-Howell test, revealed a highly significant difference between groups (P < 0.01). except there was a significant difference between 0.75wt% group and 1wt% group at (P < 0.05) (Table 5).\n\n\nDiscussion\n\nLong-term usage of maxillofacial prostheses encourages the colonization of microorganisms on the silicone surface and spreads infection to nearby tissues; similarly, biofilm may transfer from infected skin to the prosthesis.4\n\nAs previously stated, prolonged physical and chemical immersion disinfectants may result in material deterioration and color change, and the removal of bacterial accumulation is essential for external prostheses.\n\nAdditionally, it’s essential to discover a cleaning technique that is both effective in preventing infections and silicone prosthesis degeneration.19 Due to the potential for a toxic or adverse effect, the use of any antimicrobial must be limited. The development of bacterial antibiotic resistance is one of the most urgent problems facing worldwide health care. In recent years, due to fewer side effects and effective antimicrobial activity, the use of oxides instead of chemical or synthetic medicine has increased.20–22\n\nIn this investigation, it was shown to have an antibacterial action against the aforementioned bacteria since, as shown in Table 2, the percentages of bacterial cells adhering to the silicone specimens were dramatically reduced when compared to the control group.\n\nResult of this study agreed with,9 as they found a long-term antibacterial effect of BaTiO3 against S. epidermidis at 24 hours when added to Polyvinylsiloxane (PVS) between 5% and 15%. They also stated that the antibacterial activity was due to the release of Ba2+ and the formation of TiO2, resulting in slightly acidic environments. Then, when Ba2+ and TiO2 interact with water, they both help to create hydroxyl radicals (OH) and free radicals (O2-) that destroy nucleic acids, bacterial cell walls, and other molecular structures.\n\nSwain et al. found that the positively charged hydroxyapatite-BaTiO3 composite revealed antibacterial activity against S. aureus, E. coli, and P. aeruginosa with a remarkable inhibition zone. Positively polarized HA-BT composites rupture the bacterial membrane in vitro.23 Additionally, many studies have shown that barium titanate has antifungal activity.24,25\n\nFTIR measurements were performed both prior to and following the addition of BaTiO3 nanoparticles. As the spectral range remained unchanged both prior to and following the addition, there was no chemical reaction. The only interaction in this case is described as a physical reaction (hydrogen bond or Van der Waals bond), and it results from fillers interacting with silicone. This interaction manifested as a slight change in the vibration of preexisting bonds and a change in the silicone matrix’s light transmittance. This confirms that the antibacterial activity is related to BaTiO3, as no new chemical material was produced, and explains the difference in antibacterial activity between the control and experimental groups.\n\nFE-SEM revealed well dispersion of BaTiO3 inside the silicone matrix with some agglomeration as the filler percentage increased, and this agreed with.1,26 And disagreed with other studies because they utilized different fillers in varying quantities and agglomeration was only noticeable at higher percentages. This could be because surface-treated silicon dioxide nanoparticles were used; surface treatment impacts the dispersion of the nanofiller inside the matrix by decreasing the probability of nanoparticle aggregation.27,28\n\nAnother factor that affected the reduction in bacterial adhesion was reduced porosity. Many studies confirm that the addition of non-filler materials to various materials reduces porosity since the filler fills the space inside the matrix of materials.29,30 FE-SEM results showed reduced porosity of the silicone matrix, which reduces the opportunity for bacterial adhesion.\n\n\nConclusions\n\nWith respect to the limitations of this study, it can be concluded that the addition of BaTiO3 powder to VST-50 maxillofacial silicon elastomer will enhance the antibacterial activity of silicon against Staphylococcus epidermidis, and this activity seems to be concentration dependent. For further study we could evaluate the effect of the addition of BaTiO3 nanoparticles on the fungal biofilm’s adhesion to the maxillofacial silicones and study the effects of adding BaTiO3 nanoparticles to pigmented VST-50 RTV silicone elastomers. Evaluating the artificial aging of VST-50 RTV maxillofacial silicone after the addition of BaTiO3 nanopowder is another suggestion that could be explored.", "appendix": "Data availability\n\nFigshare. Antibacterial effect of Barium Titanate, https://doi.org/10.6084/m9.figshare.22336786.v1. 31\n\nThis project contains the following underlying data:\n\n• Raw data. (optical density of bacterial test)\n\n• FTIR data. (for BaTiO3 and for silicone before and after addition of BaTiO3)\n\n• FE-SEM data (pictures for BaTiO3 and for silicone before and after addition of BaTiO3)\n\n• VITEK 2 Microbiology Chart Report\n\n• Pictures of steps of bacterial test\n\n• Particle size analyzer report of barium titanate\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nAhmed AS, Ali MM: Effect of Strontium Titanate Nano Powder Addition on Some Mechanical Properties of Room Temperature Vulcanized Maxillofacial Silicone. J. Res. Med. Dent. Sci. 2021 Dec; 9(12): 59–65.\n\nLanzara R, Viswambaran M, Kumar D: Maxillofacial prosthetic materials: current status and recent advances: A comprehensive review. Int. J. Appl. Dent. Sci. 2021; 7(2): 255–259. Publisher Full Text\n\nManjula N, Rao SP: Properties of Maxillofacial Silicone Materials: A Literature. IOSR Journal of Dental and Medical Sciences (IOSR-JDMS). 2019 Oct; 18(10): 42–45.\n\nKumar A, Seenivasan MK, Inbarajan A: A Literature Review on Biofilm Formation on Silicone and Poymethyl Methacrylate Used for Maxillofacial Prostheses. Cureus. 2021 Nov 30; 13(11). Publisher Full Text\n\nde Azevedo MN , Marques NT, Fonseca MF, et al.: Disinfectant effects of Brazilian green propolis alcohol solutions on the Staphylococcus aureus biofilm of maxillofacial prosthesis polymers. J. Prosthet. Dent. 2021 May 13; 128: 1405–1411. Publisher Full Text\n\nSkovdal SM, Jørgensen NP, Meyer RL: JMM Profile: Staphylococcus epidermidis. J. Med. Microbiol. 2022 Oct 28; 71(10): 001597. Publisher Full Text\n\nMaji M, Kivale P, Ghosh M: A novel therapy to combat non-small cell lung carcinoma (A549) using platinum (IV) and barium titanate conjugate. J. Drug Deliv. Sci. Technol. 2022 Sep 1; 75: 103617. Publisher Full Text\n\nTavangar M, Heidari F, Hayati R, et al.: Manufacturing and characterization of mechanical, biological and dielectric properties of hydroxyapatite-barium titanate nanocomposite scaffolds. Ceram. Int. 2020 May 1; 46(7): 9086–9095. Publisher Full Text\n\nMarin E, Boschetto F, Sunthar TP, et al.: Antibacterial effects of barium titanate reinforced polyvinyl-siloxane scaffolds. Int. J. Polym. Mater. Polym. Biomater. 2021 Apr 13; 70(6): 425–436. Publisher Full Text\n\nIbrahim HI, Abdul-Ameer FM: Influence of kappa-carrageenan powder addition on staphylococcus epidermidis adhesion on the room temperature vulcanized maxillofacial silicone. Pak. J. Med. Health Sci. 2021; 15: 359.\n\nFatihallah AA, Alsamaraay ME: Effect of polyamide (Nylon 6) micro-particles incorporation into RTV maxillofacial silicone elastomer on tear and tensile strength. J. Baghdad Coll. Dent. 2017 Dec 15; 29(4): 7–12. Publisher Full Text\n\nAlanssari BF, Khalaf BS: Effect of Addition of Composite Polyamide Micro Particles and Silicone Dioxide NanoParticle on Some Mechanical Properties of Room Temperature Vulcanized Maxillofacial Silicone Elastomer Before and after Artificial Aging. Indian J. Forensic Med. Toxicol. 2020 Jan 16; 14(1): 1013–1019.\n\nISO/TC 45/SC 2 Testing and analysis: Rubber — General procedures for preparing and conditioning test pieces for physical test methods. 3rd ed. ICS: 83.060 Rubber ; 2016. Reference Source\n\nCross HH: Obtaining a wound swab culture specimen. Nursing. 2020; 44(7): 68–69.\n\nMicrobiology Services UK Standards for Microbiology Investigations. Bacteriology. 2015; 9(3): 1–27\n\nXu P, Yang H, Tian L, et al.: Function and safety evaluation of Staphylococcus epidermidis with high esterase activity isolated from strong flavor Daqu. LWT. 2023 Jan 31; 176: 114534. Publisher Full Text\n\nEbert C, Tuchscherr L, Unger N, et al.: Correlation of crystal violet biofilm test results of Staphylococcus aureus clinical isolates with Raman spectroscopic read-out. J. Raman Spectrosc. 2021 Dec; 52(12): 2660–2670. Publisher Full Text\n\nVolpe V, Giacomodonato MN, Sordelli DO, et al.: Ciprofloxacin loaded o/w microemulsion against Staphylococcus aureus. Analytical and biological studies for topical and intranasal administration. J. Drug Deliv. Sci. Technol. 2020 Jun 1; 57: 101705. Publisher Full Text\n\nGoiato MC, Rossatti Zucolotti BC, Mancuso DN, et al.: Care and cleaning of maxillofacial prostheses. J. Craniofac. Surg. 2010; 21(4): 1270–1273. PubMed Abstract \| Publisher Full Text\n\nGudkov SV, Burmistrov DE, Serov DA, et al.: A mini review of antibacterial properties of ZnO nanoparticles. Front. Phys. 2021 Mar 11; 9: 641481. Publisher Full Text\n\nNaseem T, Durrani T: The role of some important metal oxide nanoparticles for wastewater and antibacterial applications: A review. Environ. Toxicol. Chem. 2021 Jan 1; 3: 59–75. Publisher Full Text\n\nParham S, Kharazi AZ, Bakhsheshi-Rad HR, et al.: Antioxidant, antimicrobial and antiviral properties of herbal materials. Antioxidants. 2020; 9(12): 1–36. Publisher Full Text\n\nSwain S, Padhy RN, Rautray TR: Polarized piezoelectric bioceramic composites exhibit antibacterial activity. Mater. Chem. Phys. 2020 Jan 1; 239: 122002. Publisher Full Text\n\nSasikumar M, Ganeshkumar A, Chandraprabha MN, et al.: Investigation of Antimicrobial activity of CTAB assisted hydrothermally derived Nano BaTiO3. Materials Research Express. 2018 Nov 23; 6(2): 025408. Publisher Full Text\n\nMontoya C, Kurylec J, Baraniya D, et al.: Antifungal effect of piezoelectric charges on PMMA dentures. ACS Biomater Sci. Eng. 2021 Oct 1; 7(10): 4838–4846. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTukmachi MS, Safi IN, Ali MM: Evaluation of mechanical properties and cytotoxicity of maxillofacial silicone material after incorporation of zirconia nanopowder. Mater. Today: Proc. 2021 Jan 1; 42: 2209–2217.\n\nTukmachi M, Moudhaffer M: Effect of nano silicon dioxide addition on some properties of heat vulcanized maxillofacial silicone elastomer. IOSR-JPBS. 2017; 12(3): 37–43. Publisher Full Text\n\nAtta Allah J, Muddhaffer M: Influence of artificial weathering on some properties of nano silicon dioxide incorporated into maxillofacial silicone. Int. J. Sci. Res. 2017; 6(5): 423–428.\n\nGad MM, Fouda SM, Al-Harbi FA, et al.: PMMA denture base material enhancement: a review of fiber, filler, and nanofiller addition. Int. J. Nanomedicine. 2017; 12: 3801–3812. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRahman HA: The effect of addition nano particle ZrO2 on some properties of autoclave processed heat cure acrylic denture base material. J. Bagh. Coll. Dent. 2015; 27: 32–39.\n\nKareem YM: antibacterial effect of Barium Titanate. Dataset. figshare. 2023. Publisher Full Text" }	[ { "id": "192100", "date": "30 Aug 2023", "name": "Razia Z. Z. Adam", "expertise": [ "Reviewer Expertise Dental biomaterials", "application of nps" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis article addresses the application of nanoparticles to improve the properties of maxillofacial silicone. Barium Titanate is explored as a potential modifier which will improve the antibacterial effect of the silicone. Prepared nps are tested against S Epidermidis specifically with five groups including a control group. Characterization test confirm the nps and SEM also clearly indicates the dispersion in the material. Antibacterial activity was tested and yielded promising results.\nThe Methodology is well explained in detail. The results and discussion are appropriate.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nI cannot comment. A qualified statistician is required.\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "184170", "date": "30 Aug 2023", "name": "Kadhim A. Hubeatir", "expertise": [ "Reviewer Expertise Material science and Nanotechnology", "Laser application in medicine specially in dentistry" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nI think this research was an experimental and revealed a new results in dentistry especially to enhance the prevention and to improve the efficiency of dentin against antibacterial effect using a Barium Titanate nanoparticles.\nA good statistically analysis using ANOVA method and another different methods which was give a highly significant difference between them. Also FTIR analysis shows no chemical reactions between the Barium Titanate and Maxillofacial silicon elastomer. The pictures of SEM shows the nano powder of barium titanate dispersed inside the maxillofacial silicon.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "201564", "date": "12 Sep 2023", "name": "Nidambur Vasudev Ballal", "expertise": [ "Reviewer Expertise Microbiology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis manuscript constitutes an attempt to assess the antibacterial effect of Barium Titanate nanoparticles against Staphylococcus epidermidis adhesion after incorporation to maxillofacial silicone. The study is relevant to the journal and well performed. Some of the specific queries have been addressed below.\nIntroduction:\nMention the rationale of performing this study. There are several antibacterial nanoparticles like Silver, Chitosan, Zinc, Titanium etc. which have incorporated into silicone maxilla-facial prosthesis to exhibit antibacterial activity (Chong et al., (2022)1; Cevik et al., (2023)2). Why specifically, Barium Titanate nano-particles were tested in this study? Mention in detail.\n\nMention the hypothesis of the study tested.\nMethodology:\nA positive control group of chlorhexidine should have been used.\n\nHow was sample size estimated?\n\nWhy in control group, was silicone material mixed using digital weighing balance when compared to test groups which were mixed with vacuum pressure?\n\nDid the skin of the patient from which the swab obtained have any lesions or disease? If so, mention it.\n\nHow was biofilm formation on the silicone material confirmed prior to the testing of antibacterial efficacy of test agents?\n\nIn FTIR analysis, mention the resolutions at which the spectra were obtained.\n\nIn FESEM analysis, mention how was samples prepared for the analysis and also at what magnification the images were captured.\n\nIn FTIR and FESEM analysis, why samples treated with 0.25 and 1% BaTiO3 were not evaluated?\nResults:\nIn FESEM image, in control group, what are those particles seen? Also, how was the even distribution of the filler particles assessed? In test group, only few filler particles are seen.\n\nIn FTIR and FESEM analysis, why statistics was not performed? Was it only qualitative data obtained?\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10401", "date": "16 Oct 2023", "name": "Yasir Mohammed Kareem", "role": "Author Response", "response": "Dear Nidambur Vasudev Balla, Thank you for taking the time to review our article titled “Assessment of the antibacterial effect of Barium Titanate nanoparticles against Staphylococcus epidermidis adhesion after addition to maxillofacial silicone” and providing us with your insightful feedback to improve the manuscript. We have included answers in the manuscript to the questions put forth. Please see below the point-by-point responses to the questions. Introduction: \"Mention the rationale of performing this study. There are several antibacterial nanoparticles like Silver, Chitosan, Zinc, Titanium etc. which have incorporated into silicone maxilla-facial prosthesis to exhibit antibacterial activity (Chong et al., (2022)1; Cevik et al., (2023)2). Why specifically, Barium Titanate nano-particles were tested in this study? Mention in detail.\" Response: Thank you for your comment, As mentioned in the introduction in the fourth paragraph \"the primary cause of routine facial prosthetic replacement is deterioration in appearance caused by changes in physical characteristics and color\". So not only the antibacterial effect of BaTiO3 is important. The other mechanical and physical properties of BaTiO3 absolutely differ from those of silver, chitosan, zinc, titanium, etc. This may be superior or inferior. This study can be followed by other studies to evaluate the mechanical and physical properties of BaTiO3, as mentioned in the suggestion. \"Mention the hypothesis of the study tested.\" Response: The null hypothesis (H0): Suggested that adding BaTiO3 Nanoparticles will not affect bacterial adherence. The alternative hypothesis (H1): Suggested that adding BaTiO3 Nanoparticles will significantly reduce bacterial adherence. Methodology: \"A positive control group of chlorhexidine should have been used.\" Response: Chlorhexidine is antibacterial, has been approved in many studies, and needs no more approval. Additionally, we need to discover the effects of BaTiO3 on the physical and mechanical properties of silicon, not only its antibacterial activity. For this reason, we compared the silicone without a positive control. If the antibacterial property is the only property that we need, we would use chlorhexidine as a positive control to compare the antibacterial activity of BaTiO3 with it. \"How was sample size estimated?\" Response: The sample size estimated from previous studies, such as Ibrahim HI, Abdul-Ameer FM: Influence of kappa-carrageenan powder addition on staphylococcus epidermidis adhesion on the room-temperature vulcanized maxillofacial silicone. Pak. J. Med. Health Sci. 2021; 15: 359 Additionally, statistically, the minimum sample size can be 7 to 10 to be acceptable. \"Why in control group, was silicone material mixed using digital weighing balance when compared to test groups which were mixed with vacuum pressure?\" Response: The digital balance was used for all groups (control and test) and only for weighing the components that would be mixed. and the vacuum mixer was also used for all groups (control and test) to avoid air entrapment. The weighing and mixing for all groups were done in the same way. I am sorry, but you may have had a misunderstanding while reading the mixing part. \"Did the skin of the patient from which the swab obtained have any lesions or disease? If so, mention it.\" Response: Yes, the skin patients have a lesion, and they had a facial prosthesis. \"How was biofilm formation on the silicone material confirmed prior to the testing of antibacterial efficacy of test agents?\" Response: It was confirmed using scanning electron microscope. \"In FTIR analysis, mention the resolutions at which the spectra were obtained.\" Response: the resolutions was at 400-4000 cm-1 \"In FESEM analysis, mention how was samples prepared for the analysis and also at what magnification the images were captured.\" Response: According to the FE-SEM device used, the sample was prepared as follows: Cutting small pieces with scissors from the sample and coating them with gold using a sputter coater device for 2 minutes. The magnification was 1000, 4000, 13000, 25000, and 50000. \"In FTIR and FESEM analysis, why samples treated with 0.25 and 1% BaTiO3 were not evaluated?\" Response: The FTIR and FE-SEM were used to obtain qualitative data. Because the FTIR was used only to confirm if there was a chemical reaction or not, evaluating one test sample was enough to compare it with the control group. Also, FE-SEM was used to evaluate the distribution of BaTIO3 in the silicon matrix, and we took two different percentages. and they were enough to show the distribution of filler and some agglomeration as filler concentration increased. Results: \"In FESEM image, in control group, what are those particles seen? Also, how was the even distribution of the filler particles assessed? In test group, only few filler particles are seen.\" Response: These particles are sputter coating materials related to the device, and they are seen only at low magnifications. and some surface roughness was also seen. the even distribution of filler assessed by a observation The even distribution of filler was assessed by observation. It seems uniform, and in some areas there are some agglomerations. Your comment that \"only a few filler particles are seen\" could be due to the very tinny size of the nanoparticles (59.4 nm). and could appear at greater magnifications. These few particles could be a cluster of more than one particle. and the white, tiny particles are the single ones. \"In FTIR and FESEM analysis, why statistics was not performed? Was it only qualitative data obtained?\" Response: Yes, it was only qualitative data obtained to confirm the type of reaction and distribution of filler. Finally, we thank you again for your valuable and insightful comments." } ] } ]	1	https://f1000research.com/articles/12-385
https://f1000research.com/articles/12-483/v1	11 May 23	{ "type": "Research Article", "title": "Assessment of quality of life in patients with surgically treated maxillofacial fractures", "authors": [ "Sunil S Nayak", "Srikant Gadicherla", "Sreea Roy", "Muskaan Chichra", "Shriya Dhaundiyal", "Vanishri S Nayak", "Vinayak Kamath", "Sunil S Nayak", "Sreea Roy", "Muskaan Chichra", "Shriya Dhaundiyal", "Vanishri S Nayak", "Vinayak Kamath" ], "abstract": "Background: The complex nature of maxillofacial injuries can affect the surgical treatment outcomes and general well-being of the patient. To evaluate the efficiency of the surgical treatment, assessment of the quality of life (QOL) of the patients is of vital importance. Due to the absence of an exclusive QOL assessment tool for maxillofacial fractures, we introduce the ‘Twenty-point quality of life assessment in facial trauma patients in Indian population'. The aim of this study was to assess and evaluate the QOL following surgical management of maxillofacial trauma patients based on the severity of the injury. Methods: The study consisted of 182 subjects divided into two groups of 91 each (Group A: severe facial injury and Group B: mild to moderate facial injury). The Facial Injury Severity Scale (FISS) was used to determine the severity of facial fractures and injuries. The twenty–point quality of life assessment tool includes Zone 1 (Psychosocial impact) and Zone 2 (Functional and aesthetic impact), with ten domains each to assess QOL. Results: In Zone 1, the mean scores for Group A and Group B were 38.6 and 39.26, respectively. In Zone 2, Group B (44.56) had higher mean scores compared to Group A (32.92) (p< 0.001). Group B (83.8) had higher mean scores compared to Group A (71.58) when the total of both Zone 1 and Zone 2 were taken into consideration (p<0,001). In Group A, 9 out of 91 patients had a total score of 81- 100 compared to 68 in the same range in Group B. Conclusions: Proper surgical management with adequate care to the hard and soft tissues can improve the QOL by reducing postoperative psychosocial and functional complications. Aesthetic outcomes play an important role in determining the QOL. Mild/ Moderate injuries show better QOL compared to severe maxillofacial injuries.", "keywords": [ "Quality of life", "Maxillofacial injuries" ], "content": "Introduction\n\nMaxillofacial trauma and associated injuries can cause severe physical deformity and psychological consequences.1 Disability following major trauma has gained significant attention due to these related complications.2 Poor quality of life in injured patients with a risk of developing psychiatric problems has been observed by some studies.3–6 In the present era, to evaluate the impact of disease and the efficiency of the treatment, assessment of the quality of life of the patients is of vital importance.1 Unfortunately, studies that assess the quality of life (QOL) after surgical treatment of maxillofacial fracture surgeries are rare and so we decided to investigate this.\n\nSpecific factors that influence the outcomes of trauma are the site of the fracture, the type of fracture, and the patient's age.7 An associated fear factor with the treatment of fractures warrants the need to look into the psychological issues related to fracture surgery.8–10 Though there are many quality of life (QOL) assessment tools in literature, an exclusive QOL assessment tool for maxillofacial fractures is lacking. We have devised an exclusive, first-of-its-kind QOL assessment tool that looks into both physical and psychological aspects following maxillofacial trauma surgery. The ‘Twenty-point Quality of life assessment in facial trauma patients in Indian population' assessment tool was developed and used to determine the QOL in this study. The aim of this study was to assess the QOL following surgical management of maxillofacial trauma patients based on the severity of the injury. The objectives of the study were:\n\n1. To determine the domains that effect the QOL of a surgically treated individual.\n\n2. To assess the efficiency of the surgical treatment based on the QOL results.\n\nAlso, the authors intend to introduce a new assessment tool to determine QOL precisely from a maxillofacial injury perspective.\n\n\nMethods\n\nThe study was held in the Oral and Maxillofacial Surgery Department and Kasturba Hospital, Manipal, from January 2020 to October 2021 after approval from the Institutional Ethics Committee (IEC: 924/2019) on 19/11/2019. Written informed consent was obtained from each of the study participants.\n\nThe study included patients between 18-70 years of age. The study participants were consecutive patients reporting to our unit with maxillofacial trauma who underwent maxillofacial fracture surgery. The patients were assessed 8-12 weeks after surgical intervention. A data schedule was prepared to document age, sex, and fracture type. The sex of the participants was determined based on self-report. A total of 188 patients were seen during the study period and were divided into 2 groups according to their consecutive arrival (Group A: severe facial injury and Group B: mild to moderate facial injury) based on the severity of maxillofacial fractures and facial injury. Six patients (three from each group) were lost at follow-up. The final study consisted of 182 subjects divided into two groups of 91 each. Patients with associated diseases like cysts or tumors of the jaw bones, pregnant women, and those with underlying psychological issues were excluded from the study.\n\nWe followed the Facial Injury Severity Scale (FISS)11 to determine the severity of facial fractures and injuries prior to evaluating the QOL. The face is divided horizontally into the mandibular, mid-facial, and upper facial thirds. Fractures in these thirds are given points based on their type (Table 1). Injuries with a total score above 4.4 were considered severe facial injuries (Group A), and those with a total score below 4.4 were considered mild/moderate facial injuries (Group B). The QOL was compared between the two groups.\n\n\n\n• Dentoalveolar/Condyle/Coronoid\n\n\n\n• Body/Ramus/Symphysis\n\n\n\n• Dento Alveolar\n\n\n\n• Le Fort I\n\n\n\n• Le Fort II\n\n\n\n• Le Fort III\n\n\n\n• Naso-Orbital Ethmoid (NOE) 3 points\n\n\n\n• Zygomatico Maxillary Complex (ZMC) 1 point\n\n\n\n• Nasal\n\n\n\n• Orbital roof/rim\n\n\n\n• Displaced frontal sinus/bone fractures\n\n\n\n• Non-displaced fractures\n\n\n\n• Over 10 cm long\n\nMeticulous management of hard and soft tissue injuries in our state-of-the-art tertiary care hospital was implemented. All elective cases were surgically treated at least 72 hours after the initial trauma. The facial fractures were adequately reduced and fixed with high–end titanium miniplates and screws (AO Principles of Fracture Management). Soft tissue injuries were managed by wound debridement, removal of foreign bodies, and layered wound closure. Adequate pain-relieving medication was prescribed to the patients postoperatively for effective pain control.\n\nThe QOL of the subjects was assessed using the 'Twenty-point Quality of life assessment in facial trauma patients in Indian population' assessment tool. The development was based on the WHOQOL-BREF scale and was modified from a maxillofacial trauma point of view. The twenty-point QOL assessment contains 20 questions (domains) and uses a five-point Likert response scale, and includes two zones: Zone 1 (Psychosocial impact) (Table 2) and Zone 2 (Functional and aesthetic impact) (Table 3), with ten questions (domains) each. The scores for each question ranged from 1-5, with a higher score denoting better quality of life. Accordingly, the score in each zone for a patient ranged from 10-50, and the total scores of both zones were recorded to determine the QOL. The sum of both zones determined the prognosis following surgery (Table 4).\n\n\n\n1. You are satisfied with your overall physical health\n\n\n\n2. You can perform your daily activities without any medicines or medical/surgical aid\n\n\n\n3. You can concentrate on your life and enjoy it as same as before\n\n\n\n4. You are confident to mix with your friends and family in the same way as before\n\n\n\n5. You are able to get good sleep\n\n\n\n6. Your personal relations have not experienced a set back\n\n\n\n7. You are satisfied with the support you received from friends/family/colleagues during treatment and recovery phase\n\n\n\n8. You don’t have aversion or hatred towards life? You never had suicidal feelings\n\n\n\n9. You can accept changes in your facial appearance\n\n\n\n10. You are happy with the medical services provided to you during your treatment\n\n\n\n1. You are happy with decline in pain level\n\n\n\n2. You are happy with the reduction in amount of swelling\n\n\n\n3. You can enjoy your daily meals like before\n\n\n\n4. You can open your mouth as wide and easily as before\n\n\n\n5. The surgical hardware does not cause you any irritation\n\n\n\n6. You can feel the same sensation of touch on your cheeks/chin as before\n\n\n\n7. You don’t feel any discomfort in swallowing\n\n\n\n8. You are completely satisfied the way you talk\n\n\n\n9. You feel that you look good as before\n\n\n\n10. Your vision is as good as before\n\nThe Test-retest method was used to determine the reliability and the correlation coefficient (r) values was above 0.7 The scale was piloted previously but 2 questions (1 each from each zone) were modified due to the poor understanding by the participants:\n\n• Question 3: You can concentrate on your life and enjoy it as same as before (the earlier version was ‘Can you focus well in your life?’)\n\n• Question 13: You can enjoy your daily meals like before (the earlier version was ‘Are your tongue movements normal at present?’)\n\nThe data collected was entered into a Microsoft Excel 365 MSO (Version 2301 Build 16.0.16026.20002) 64-bit spreadsheet and analyzed in the form of frequency and percentage for categorical variables, and in the form of mean, median, standard deviation, and quartiles for continuous variables. A non-parametric test was used and QOL scores were compared using SPSS Statistics, Version 22 (Armonk, NY: IBM Corp). Descriptive data were presented in between the study groups using the Mann-Whitney U test. P value < 0.05 was considered statistically significant.\n\n\nResults\n\nThe study group comprises 182 subjects (145 males and 37 females) (Table 5). 160 were below 65 years of age (Table 5). The causes of fractures were road traffic accidents (84.3%), violence (5.3%), falls (8.2%), and sports activities (2.2%). In Zone 1, the mean scores for Group A and Group B were 38.6 and 39.26, respectively. In Zone 2, Group B (44.56) had higher mean scores compared to Group A (32.92), and this was statistically significant (p< 0.001) (Table 6). Group B (83.8) had higher mean scores compared to Group A (71.58) when the total of both Zone 1 and Zone 2 were taken into consideration, and this was statistically significant (p<0.001) (Table 6).\n\n* p<0.05 statistically significant.\n\np>0.05 non-significant (NS), A=Group A, B=Group B.\n\nIn Zone 1, the domains of 'Satisfaction in daily activities' (Question 2) and 'Acceptance of post-trauma facial appearance' (Question 9) had the lowest mean scores in Group A. In Group B, 'the ability to interact with family and friends' (Question 4) domain showed the lowest mean score (Table 7, Figure 1). In Zone 2, the domains of 'aesthetics' (Question 9) and 'mastication' (Question 3) had the lowest mean scores in Group A, and in Group B, the 'aesthetics' domain (Question 9) had the lowest mean score (Table 8, Figure 2). On comparison of individual domains between the two groups in Zone 1, except for domains of 'Quality of sleep,' 'Lack of suicidal tendencies,' and 'Acceptance of post-trauma facial appearance,' all other domains showed statistically significant differences (Table 7). In Zone 2, on comparison of the two groups, all domains showed statistically significant differences (p<0.001) (Table 8).\n\n* p<0.05 statistically significant.\n\np>0.05 non significant (NS), A=Group A, B=Group B.\n\n* p<0.05 statistically significant.\n\np>0.05 non significant (NS), A=Group A, B=Group B.\n\nIn Group A, 9 out of 91 patients had a total score of 81-100. Three patients scored below 60, and the remaining 79 scored in the 61-80 range. Two out of 16 female participants had excellent prognosis and the remaining 14 showed good prognosis. In Group B, 68 patients scored in the 81-100 range, and the remaining 23 scored in the 61-80 range. 14 out of 21 female participants showed excellent prognosis and the remaining seven showed good prognosis (Table 9).\n\n\nDiscussion\n\nThe inclusion of assessment of QOL is essential in treating patients with maxillofacial fractures to determine psychological well-being and patient satisfaction.12 Unfortunately, the evaluation of QOL of people who had been surgically treated for maxillofacial fractures is not practiced routinely.13 While assessing QOL in maxillofacial injuries, it is essential to consider the severity of the injury. Maxillofacial injuries occur in various combinations, and individual fractures require specific descriptions. We used the FISS to classify severe and moderate/mild injuries. The FISS is a valuable tool for maxillofacial trauma assessment. This scale can reliably predict the severity of maxillofacial injuries and is easily calculated.11 We consider 4.4 as the average demarcating score to classify severe and moderate/mild injuries based on the study by Bhageri et al.11\n\nSeveral assessment tools are available to determine the well-being and QOL of patients, but maxillofacial injuries are unique due to the disfigurement and dysfunction they cause. These injuries can lower the person's self-esteem and adversely affect daily activities and social relationships, ultimately affecting QOL.13,14 We have devised an exclusive QOL assessment tool for facial trauma patients to assess post-surgery patients from a maxillofacial trauma perspective. The new assessment tool determines both the psychosocial and functional and aesthetic impacts due to maxillofacial fracture surgery.\n\nWHO's Quality of Life (WHOQOL-100) and its shorter version, the WHOQOL-BREF, are used in various settings to determine patients' quality of life.15 The WHOQOL-BREF, more popular among the two, contains four domains: physical health, psychological health, social relationships, and environment.13 Mood disorders, body image disorders, and a poor QOL are often exhibited in aesthetic and functional disturbances associated with maxillofacial trauma.16 A disfiguring maxillofacial injury can make an individual withdraw from social interaction.1 Also, a lack of support from family and friends affects an individual's physical and emotional well-being.16 All these observations were considered in the new assessment tool (The Twenty-point Quality of life assessment in facial trauma patients in Indian population), which we devised. The psychosocial impact questionnaire evaluated the patient's satisfaction with their overall physical health, the confidence of the individual in performing daily activities and interacting with family and friends, and the ability to concentrate and sleep well (Zone 1, Questions 1-5) (Table 2). Also, the personal relationships of the patient with people around them, the support by their near and dear ones, suicidal tendencies, if any, ability to accept changes in facial appearance, and satisfaction with medical services provided can be assessed (Zone 1, Questions 6-10). Hence the four domains of the WHOQOL-BREF evaluation tool are given adequate importance in the new assessment tool presented by the authors.\n\nMoreover, we also determined the functional and aesthetic impact on subjects after maxillofacial surgery (Zone 2) (Table 3). The progress of pain, swelling, mouth opening, and paresthesia can be assessed by this questionnaire (Zone 2, Questions 1, 2, 4, and 6). Evaluation of daily activities like eating, swallowing, talking (Zone 2, Questions 3, 7, 8), and discomfort due to the hardware (Zone 2, Question 5) are also evaluated. This assessment tool also considers patient concerns regarding facial appearance and visual disturbances (Zone 2, Questions 9, 10).\n\nPrevious studies have documented that patients with facial trauma subsequently exhibit poor QOL outcomes.17–20 The inflammatory response caused by a facial injury results in increased vascular permeability, vasodilation, and infiltration of monocytes and polymorphonuclear leukocytes in the area of injury. These changes take place within a few days following an injury. Hence an early fracture surgery can lead to an unfavorable outcome due to a lack of initial blood supply.21 In our study, most surgical treatment of fractures was done at least 72 hours after injury to achieve favorable outcomes.\n\nComplications in all surgically treated patients accounted for 6.6%. Alcohol abuse, smoking, and plating procedures are some factors significantly associated with complications.22 The findings of our study suggest that adequate reduction and fixation of fractures with high-end titanium miniplates and screw systems (AO Principles of Fracture Management) and efficient management of soft tissue injuries greatly enhance the outcomes of facial trauma patients. Our unit's soft tissue injury management included adequate debridement of devitalized tissue, layered closure of the wounds, and aesthetic reconstruction of soft tissue injuries with tissue loss. By following these measures, the outcomes of the surgical intervention can be significantly enhanced, as shown in our study. Moreover, severe injuries are associated with extensive soft tissue injuries, which can lead to poor QOL. In our study, the mean total scores (Zone 1+ Zone 2) show a good outcome for Group A subjects with severe injuries (71.58) and an excellent outcome for Group B subjects with mild/moderate injuries (83.8) (Table 6). These favorable outcomes in our study can be attributed to the use of state-of-the-art titanium hardware for fixation, excellent soft tissue care, restoring the functional ability of the patient, such as chewing and mouth opening, and adequate control of postoperative pain and edema.\n\nA lack of improvement in QOL after surgery was attributed to appearance, pain, and mood issues in the postoperative period.23 Also, the primary concern of patients with maxillofacial trauma compared to other types of trauma is their appearance.24 These findings correlate to our study in which the ‘facial appearance’ domain in Zone 1 and the ‘aesthetics’ domain in Zone 2 had low mean scores (Tables 7 and 8). As both these domains are related to the cosmetic appearance of the patient, cosmetic defects can adversely affect QOL in patients. Severe maxillofacial injuries can cause cosmetic defects. Cosmetic defects caused by maxillofacial trauma can, in turn, lead to depression and affect QOL.25 Moreover, trauma leading to difficulty in chewing and functional impairment such as paresthesia and diplopia can also cause depression in individuals.26 Hence, a correlation exists between psychological and aesthetic/functional components resulting from severe maxillofacial trauma. These findings relate to our study wherein the QOL was comparatively better in mild to moderate injuries compared to severe ones. In our study, better QOL was seen in Group B (mild/moderate facial injuries) compared to Group A (severe facial injuries) (Figure 3). There was statistical significance in Zone 2 scores and the combined Zone 1 + Zone 2 scores (Table 6).\n\nPatients who underwent open reduction and internal fixation of fractures recorded higher pain scores post-surgery.27–29 Pain reduces the QOL of patients, and hence it is essential to employ adequate pain reduction protocols post-surgery to improve QOL.30 Our study also shows the effect of pain on the QOL of the patient. The average mean score for decline in pain (Zone 2, Question 1) was 3.38 and 4.89, respectively, for severe and mild/moderate injuries (Figure 2). As higher scores denote better QOL, it is evident that mild/moderate injuries show better QOL compared to severe injuries. Adequate analgesics are to be prescribed to patients for effective pain control postoperatively. Moreover, we believe that meticulous and gentle handling of soft and hard tissues during the surgical procedure is vital in decreasing postoperative pain, edema, and patient discomfort. Anggayanti et al. reported significant improvement in QOL within 14 days of surgical intervention.31 Open reduction and fixation in association with excellent management of postoperative complications restore the normal configuration of anatomic structures, enhance stability, and establishes normal function.32 Efficient surgical management, good postoperative care, and due consideration of facial aesthetics were employed in our unit to obtain favorable outcomes. Both sexes showed excellent outcome in Group B compared to Group A (Table 9). The higher male:female ratio of the participants can be attributed to the fact that motorcycles in India are predominantly driven by males. Due to this higher male:female ratio, only a descriptive analysis based on sex of the individual has been demonstrated.\n\nThe study had certain limitations. As we had used a new diagnostic tool, its credibility could have been enhanced by advocating it to assess the pre-operative QOL and comparing the outcomes with the present findings. Also, comparing postoperative outcomes at different periods could have shed light on the possibility of improvement in the QOL over a period of time after surgery.\n\n\nConclusion\n\nProper surgical management with adequate care to the hard and soft tissues can improve the QOL by reducing postoperative psychosocial and functional complications. Aesthetic outcomes play an important role in determining the QOL. Mild/Moderate injuries show better QOL compared to severe maxillofacial injuries.", "appendix": "Data availability\n\nfigshare: Quality of Life in Patients with Surgically Treated Maxillofacial Fractures, https://doi.org/10.6084/m9.figshare.21702023.v4. 33\n\nThis project contains the raw data file: QoL Life Data.xlsx\n\nfigshare: Quality of Life in Patients with Surgically Treated Maxillofacial Fractures, https://doi.org/10.6084/m9.figshare.21702023.v4. 33\n\nThis project contains the ‘Twenty-point quality of life assessment in facial trauma patients in Indian population' questionnaire.\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nReferences\n\nUkpong DI, Ugboko VI, Ndukwe KC, et al.: Health-related Quality of life in Nigerian patients with facial trauma and controls: A preliminary survey. Br. J. Oral Maxillofac. Surg. 2008; 46: 297–300. Publisher Full Text\n\nHolbrook TL, Hoyt DB: The impact of major trauma: quality-of-life outcomes are worse in women than in men, independent of mechanism and injury severity. J. Trauma. 2004; 56: 284–290. Publisher Full Text\n\nMayou R, Bryant B, Duthie R: Psychiatric consequences of road traffic accidents. BMJ. 1993; 307: 647–651. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMayou R, Bryant B: Outcome in consecutive emergency department attenders following a road traffic accident. Br. J. Psychiatry. 2001; 179: 528–534. PubMed Abstract \| Publisher Full Text\n\nMayou R, Bryant B: Outcome 3 years after a road traffic accident. Psychol. Med. 2002; 32(6): 671–675. Publisher Full Text\n\nMayou RA, Ehlers A, Hobbs M: Psychological debriefing for road traffic accident victims. Three year follow up of a randomised controlled trial. Br. J. Psychiatry. 2000; 176: 589–593. PubMed Abstract \| Publisher Full Text\n\nAnyanechi CE, Chukwuneke FN: Prognosis of teeth in the line of mandibular fracture: 5-year clinical and radiological follow-up. Niger. J. Med. 2013; 22: 61–63.\n\nGironda MW, Der-Martirosian C, Belin TR, et al.: Predictors of depressive symptoms following mandibular fracture repair. J. Oral Maxillofac. Surg. 2009; 67: 328–334. PubMed Abstract \| Publisher Full Text\n\nDe Sousa A: Psychological issues in oral and maxillofacial reconstructive surgery. Br. J. Oral Maxillofac. Surg. 2008; 46: 661–664. Publisher Full Text\n\nRogers SN, Gwanne S, Lowe D, et al.: The addition of mood and anxiety domains to the University of Washington quality of life scale. Head Neck. 2002; 24: 521–529. PubMed Abstract \| Publisher Full Text\n\nBagheri SC, Dierks EJ, Kademani D, et al.: Application of a facial injury severity scale in craniomaxillofacial trauma. J. Oral Maxillofac. Surg. 2006 Mar; 64(3): 408–414. PubMed Abstract \| Publisher Full Text\n\nShepherd J: Victims of personal violence: the relevance of Symonds' model of psychological response and loss-theory. Br. J. Soc. Work. 1990; 20: 309–332.\n\nSomoye MS, Adetayo AM, Adeyemo WL, et al.: A comparative study of Quality of life of patients with maxillofacial fracture and healthy controls at two tertiary healthcare institutions. J. Korean Assoc. Oral Maxillofac. Surg. 2021; 47: 351–359. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nUkpong DI, Ugboko VI, Ndukwe KC, et al.: Health-related Quality of life in Nigerian patients with facial trauma and controls: a preliminary survey. Br. J. Oral Maxillofac. Surg. 2008; 46: 297–300. Publisher Full Text\n\nKrägeloh CU, Henning MA, Hawken SJ, et al.: Validation of the WHOQOL-BREF Quality of life questionnaire for use with medical students. Educ. Health (Abingdon). 2011; 24: 545. PubMed Abstract\n\nDe Sousa A: Psychological issues in oral and maxillofacial reconstructive surgery. Br. J. Oral Maxillofac. Surg. 2008; 46: 661–664. PubMed Abstract \| Publisher Full Text\n\nHull AM, Lowe T, Devlin M, et al.: Psychological consequences of maxillofacial trauma: a preliminary study. Br. J. Oral Maxillofac. Surg. 2003; 41: 317–322. PubMed Abstract \| Publisher Full Text\n\nLento J, Glynn S, Shetty V, et al.: Psychologic functioning and needs of indigent patients with facial injury: a prospective controlled study. J. Oral Maxillofac. Surg. 2004; 62: 925–932. PubMed Abstract \| Publisher Full Text\n\nShepherd JP, Qureshi R, Preston MS, et al.: Psychological distress after assaults and accidents. BMJ. 1990; 301: 849–850. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGirotto JA, MacKenzie E, Fowler C, et al.: Long term physical impairment and functional outcomes after complex facial fractures. Plast. Reconstr. Surg. 2001; 108: 312–327. PubMed Abstract \| Publisher Full Text\n\nHermund NU, Hillerup S, Kofod T, et al.: Effect of early or delayed treatment upon healing of mandibular fractures: A systematic literature review. Dent. Traumatol. 2008; 24: 22–26. Publisher Full Text\n\nFurr AM, Schweinfurth JM, May WL: Factors associated with long-term complications after repair of mandibular fractures. Laryngoscope. 2006; 116: 427–430. PubMed Abstract \| Publisher Full Text\n\nBoljevic T, Vukcevic B, Pesic Z, et al.: The Quality of life of patients with surgically treated mandibular fractures and the relationship of the posttraumatic pain and trismus with the postoperative complications: A prospective study. Medicina. 2019; 55(4). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRahtz E, Bhui K, Hutchison I, et al.: Are facial injuries really different? An observational cohort study comparing appearance concern and psychological distress in facial trauma and non-facial trauma patients. J. Plast. Reconstr. Aesthet. Surg. 2018; 71(1): 62–71.\n\nRanganathan V, Panneerselvam E, Chellappazham S, et al.: Evaluation of depression associated with post-traumatic stress disorder after maxillofacial injuries- a prospective study. J. Oral Maxillofac. Surg. 2018; 76(6): 1282.e1–1282.e9. Publisher Full Text\n\nSen P, Ross N, Rogers S: Recovering maxillofacial trauma patients: the hidden problems. J. Wound Care. 2001; 10(3): 53–57.\n\nOmeje KU, Rana M, Adebola AR, et al.: Quality of life in treatment of mandibular fractures using closed reduction andmaxillomandibular fixation in comparison with open reduction and internal fixation - A randomized prospective study. J. Craniomaxillofac. Surg. 2014; 42(8): 1821–1826. PubMed Abstract \| Publisher Full Text\n\nOmeje KU, Adebola AR, Efunkoya AA, et al.: Prospective study of the Quality of life after treatment of mandibular fractures. Br. J. Oral Maxillofac. Surg. 2015; 53(4): 342–346. PubMed Abstract \| Publisher Full Text\n\nAtchison KA, Shetty V, Belin TR, et al.: Using patient self-report data to evaluate orofacial surgical outcomes. Community Dent. Oral Epidemiol. 2006; 34(2): 93–102. PubMed Abstract \| Publisher Full Text\n\nSharma G, Kaur A: Quality of life after orbito-facial trauma. Orbit. 2017; 36(6): 407–410. PubMed Abstract \| Publisher Full Text\n\nAnggayanti NA, Sjamsudin E, Maulina T, et al.: The Quality of life in the treatment of maxillofacial fractures using open reduction: A prospective study. Bali Med. J. 2020; 9(3): 757–761. Publisher Full Text\n\nZoghbi Y, Gerth DJ, Tashiro J, et al.: Open versus closed reduction of maxillary fractures: Complications and resource utilization. J. Craniofac. Surg. 2017; 28(7): 1797–1802. Publisher Full Text\n\nNayak S, Nayak V: QoL Life Data.xlsx. [Dataset]. figshare. 2022. Publisher Full Text" }	[ { "id": "173201", "date": "08 Jun 2023", "name": "Amit Sethi", "expertise": [ "Reviewer Expertise Minor oral surgery", "medicine", "anesthesia", "oral pathology" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors have done well to do “first of its kind QOL survey” for facial trauma patients. The objectives, research methodology and results have been well presented in the paper. I will highlight some of the major and minor points regarding the study.\nMajor points\nThe large sample size (182 patients) with Indian patients further make the research unique and relevant.\n\nThe age range of patients is wide (18-70 years) and covers young as well as old patients.\n\nAuthors have studied patients with both mild-moderate and severe facial injury and have covered both psychosocial and esthetic-functional components.\nMinor points\nAuthors have tried to establish a relation between psychological and esthetic and functional outcomes.\n\nAdequate pain control which is so important in post operative course yet remains undervalued has been emphasized well by the authors\n\nPoor cosmetic results and possible resultant depression and effect on QOL has also been discussed\nPoints that may need further clarification\nWhat was the reason for doing QOL assessment after 8-12 weeks ? There is a 4 week difference in 8 and 12 weeks. 4 weeks can influence healing and QOL. How did authors account for this difference?\n\nThe authors do talk about post op analgesics. Were the patients prescribed post op antibiotics or in the interim 72 hrs (“most patients were treated after 72 hrs”) and did this have any bearing on outcomes?\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "198716", "date": "04 Sep 2023", "name": "Gabriele Canzi", "expertise": [ "Reviewer Expertise Facial trauma" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis paper focuses on a very interesting and extremely important topic: The personal perception on QOL of patients treated for facial trauma. Most facial traumatologists stress the radiological accuracy of their obtained results, forgetting that the real final outcome is the satisfaction and fully health of their patients.\nThe work is clear, well structured, the statistics are simple but effective. I appreciated the intuition of dividing the population into Severe and Mild-to-Moderate facial injuries. This makes the results extremely convincing. Facial trauma is often not categorized in scientific research and the results leave open the question of the true homogeneity of the samples under study.\nThe FISS represents a milestone, since 2006, for the severity classification of facial trauma.\nFISS stratifies the severity of facial trauma based on the anatomical sites involved, but does not distinguish the true fracture characteristics (i.e. compound or displaced fractures). Therefore it does not allow to predict the type of treatment needed (i.e. ORIF vs closed management) and the final prognosis.\n\nAs you stated soft tissue injuries heavily influence the outcome of your study; the sot tissues consideration using the FISS (based only on the length of the wounds) is reductive.\nIn the end the stratification of patients in Mild, Moderate and Severe facial trauma, using the FISS, is not statistically validated. The arbitrarily established limit value of 4.4 could be reached by a patient with LeFort II and compound nasal bone fracture (FISS=5), who can be treated with MMF and an excellent restitutio ad integrum.\nIn 2019, the Comprehensive Facial Injury (CFI) score was proposed, which exceeded the limits of the FISS from which it derives, while maintaining its simplicity of use. Stratification of patients into Mild, Moderate and Severe facial trauma, using CFI score, was yet statistically validated and published. I therefore invite you to optimize your excellent work using this tool, suggesting you add a table describing the characteristics (site and type) of the fractures examined and their assigned CFI score.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "11022", "date": "13 Apr 2024", "name": "Sunil Nayak", "role": "Author Response", "response": "The reservations observed by the reviewer (Gabriele Canzi, Niguarda Hospital, Milan, Italy) have been addressed, and the manuscript modified accordingly." } ] } ]	1	https://f1000research.com/articles/12-483
https://f1000research.com/articles/11-1280/v1	09 Nov 22	{ "type": "Brief Report", "title": "A human mesenchymal spheroid prototype to replace moderate severity animal procedures in leukaemia drug testing", "authors": [ "Aaron Wilson", "Sean Hockney", "Jessica Parker", "Sharon Angel", "Helen Blair", "Deepali Pal", "Aaron Wilson", "Sean Hockney", "Jessica Parker", "Sharon Angel", "Helen Blair" ], "abstract": "Patient derived xenograft (PDX) models are regarded as gold standard preclinical models in leukaemia research, especially in testing new drug combinations where typically 45-50 mice are used per assay. 9000 animal experiments are performed annually in the UK in leukaemia research with these expensive procedures being classed as moderate severity, meaning they cause significant pain, suffering and visible distress to animal’s state. Furthermore, not all clinical leukaemia samples engraft and when they do data turnaround time can be between 6-12 months. Heavy dependence on animal models is because clinical leukaemia samples do not proliferate in vitro. Alternative cell line models though popular for drug testing are not biomimetic – they are not dependent on the microenvironment for survival, growth and treatment response and being derived from relapse samples they do not capture the molecular complexity observed at disease presentation. Here we have developed an in vitro platform to rapidly establish co-cultures of patient-derived leukaemia cells with 3D bone marrow mesenchyme spheroids, BM-MSC-spheroids. We optimise protocols for developing MSC-spheroid leukaemia co-culture using clinical samples and deliver drug response data within a week. Using three patient samples representing distinct cytogenetics we show that patient-derived-leukaemia cells show enhanced proliferation when co-cultured with MSC-spheroids. In addition, MSC-spheroids provided improved protection against treatment. This makes our spheroids suitable to model treatment resistance – a major hurdle in current day cancer management Given this 3Rs approach is 12 months faster (in delivering clinical data), is a human cell-based biomimetic model and uses 45-50 fewer animals/drug-response assay the anticipated target end-users would include academia and pharmaceutical industry. This animal replacement prototype would facilitate clinically translatable research to be performed with greater ethical, social and financial sustainability.", "keywords": [ "Animal replacement", "3D models", "Preclinical models", "cancer research", "leukaemia" ], "content": "\n\n\n\nScientific benefit\n\nA 3D spheroid-based approach with improved clinical relevance for ex vivo co-culture of clinical leukaemia samples for further investigation into cancer biology such as blast-niche interactions, blast proliferation and treatment resistance.\n\n3Rs benefit\n\nTo replace moderate severity animal (mice) procedures in leukaemia research and drug testing.\n\nPractical benefit\n\n3Rs approach that yields drug response data quickly and is more ethically, socially and financially sustainable than its in vivo counterparts.\n\nCurrent applications\n\nExploration of leukaemia biology such as blasts proliferation, blast-niche interactions, niche-impacted treatment resistance and obtain drug response data.\n\nPotential applications\n\nAdapt the approach to include other haematological cancers as well as bone cancers.\n\n\nIntroduction\n\nLeukaemia management is hindered by two chief obstacles: 1. Treatment still involves toxic chemotherapy drugs1,2 2. 15-20% of patients go into relapse at which point the disease can be treatment resistant.3 The nature of relapse disease highlights the need for safer and improved treatments such as targeted therapy. While many novel therapeutics have been identified, high drug attrition rates have been a major hindrance against anti-cancer drug development. 95% of bench side drugs that make it to phase 1 clinical trial never reach patients.4–6 Three key areas5 have been identified that must be addressed in order to reduce attrition rates: 1. Increase in scientific collaboration 2. Preclinical studies and clinical trials must be tumour biology driven for the drug to be more therapeutically effective. 3. Lack of robust and translatable preclinical cancer models that could be used to screen out many unsuccessful therapeutics before going to trial.5\n\nThe gold standard preclinical model in leukaemia research is a patient derived xenograft model.7–9 Recent Home Office annual returns show that 195,407 mice are used annually in cancer research. Indeed, cancer research remains the most common applied sciences area to use animal procedures. Given leukaemia research is heavily dependent on mouse models combining these metrics with financial data from major cancer funders in the UK suggest that 5% of these mice or at least 9000 mice are used in leukaemia research every year. Although leukaemia is a very aggressive disease, leukaemia cells do not maintain viability outside the body.9 This has led to the development of patient-derived xenograft (PDX) models where leukaemia cells are engrafted into the BM of immunocompromised mice. PDX models are currently the most clinically relevant models in blood cancer preclinical studies. Mouse PDX models (which are regarded as moderate severity procedures) are used to amplify patient-derived leukaemia cells, to study leukaemia biology and perform translational studies.8,10 Patient-derived leukaemia cells can take anywhere between 2-12 months to engraft in mice before any preclinical drug testing can be carried out, hence these mice experiments are very time consuming. In addition, most of these procedures are considered to be moderate severity and often involve mice developing significant adverse effects such as weight loss, fur ruffling, back hunching as well as increased white cell counts.\n\nThe pipeline for preclinical testing using mouse PDX models are as documented in literature is: 1. Toxicity analysis to determine no observed adverse effect limit (NOAEL)/maximum tolerated dose (MTD): 15-20 mice per drug.11 2. Pharmacokinetic studies to determine drug maximum plasma/tissue levels and kinetics of drug clearance:15-30 mice per drug.12 3. Drug efficacy studies and pharmacodynamics assays on tumour cell response: 5-10 mice needed per drug treatment group.13 Per drug group a total of 40-50 animals are needed for single drug assays and twice as many for drug combination assays.11–13 In addition, in vitro cancer models preceding in vivo validation experiments rely on cell lines that show limited in vivo predictability. Cell lines retain cytogenic characteristics of acute lymphoblastic leukaemia (ALL), but being derived from patients with relapsed disease these samples do not represent the molecular complexity of disease at presentation.14,15 Cell lines have also artificially adapted to grow in suspension culture which eliminates a key feature of leukaemia biology: microenvironment and its role in treatment resistance. Furthermore, these models are restricted by limited in vivo predictability and often lead to in vitro to in vivo drug attrition rates. This in turn results in unnecessary animal experiments which could have been avoided for these failed drug candidates, i.e., false positive data from cell line models.\n\nALL is a disease primarily of the bone marrow (BM) microenvironment (or leukemic niche), a compartment within the bone composed of cell types including mesenchymal stroma cells (MSC), osteoblasts, osteoclasts, endothelia, immune cells and supporting perivascular cells. Difficulty in growing and maintaining patient derived samples once they removed from the patient alludes to the vital role that the niche plays in cancer biology. Study into blast-niche interaction has unveiled some of the supportive roles the niche plays in disease, including cell-cell signalling, growth factors, cell adhesion and evidence has shown the niche providing an element of chemoprotection.16,17 The niche particularly mesenchyme-blast interaction gives rise to two lymphoblast subpopulations, the first being a slow cycling, dormant population and the second being actively cycling disease propagating blasts.14,18 Many treatments are effective in reducing the populations of active cycling blasts; however, it is reported that the dormant populations can survive treatment, seeking refuge within the niche. These dormant cells can later shift into the cycling state post treatment, leading to relapse disease.15,18\n\nTargeting of niche-blast interactions opens a door into a variety of potential therapeutic approaches. An example would be CXCL12, a chemokine secreted by stroma cells and upregulated in disease, it was discovered that CXCL12 induces a non-cycling state within LSCs, reducing the effectiveness of tyrosine kinase inhibitors. Mesenchymal cell CXCL12 knockout reversed this change and caused a phenotypic switch into disease propagating cycling blasts, temporarily increasing disease load but allowing once chemo resistant blasts to be treated.19 In preceding studies, we have developed ex vivo 2D co-culture platforms,14,15 the purpose of which has been to 1. conduct preclinical drug screening using patient-derived leukaemia cells thereby reducing dependence on cell lines and consequently minimising in vitro to in vivo drug attrition rates and 2. Explore the impact of human BM cells in driving key elements of cancer biology such as leukaemia proliferation, dormancy and treatment resistance.14 Importantly, to build proof of confidence in application we show that such human relevant ex vivo platforms have 1. the potential to detect therapeutically exploitable targets that disrupt leukaemia-BM cell interactions conferring treatment resistance to the cancer cells. For example, using our ex vivo approach we reveal that CDH2 drives niche-mediated treatment resistance in leukaemia and that this interaction can be targeted via a drug ADH-1. In addition, we validate that CDH2 is indeed significantly upregulated in human leukaemia bone marrow thereby corroborating the strength of our ex vivo model in predicting human/clinical data. Such models also have greater experimental accessibility than in vivo procedures and thus mitigate reliance on mouse experiments to perform such mechanistic studies 2. We use our ex vivo approach to conduct ADH-1 drug testing on 15 different patient-derived clinical samples (i.e., biological replicates. We used previously cryopreserved PDX clinical samples generated historically in older projects) and drug combination testing on 3 patient-derived clinical samples. These ex vivo experiments informed our in vivo validation experiments so these could be streamlined to achieve optimal animal replacement. Although such 2D studies aim to successfully capture interaction of patient-derived leukaemia cells with cell components of the human BM niche/leukaemia microenvironment, 2D models do not portray functional biomimicry and consequently improved clinical relevance displayed by 3D models.\n\nRecent advances in medicine have utilised 3D models as miniature in vitro representations of specific organs, they are produced in 3D and retain a realistic microanatomy, allowing a representational human model.20 3D models recapitulate their respective organ function and have been proven to be useful models of human disease, particularly in cancer research.21 2D models cannot recapitulate the leukemic niche or the complex and varied interactions that influence leukemic cell growth.22 Current medicine is making use of organ-on-a-chip systems which have led to advances in liver and lung drug screening, providing more accurate toxicological reports than 2D systems.20 The complex and dynamic nature of ALL and the bone marrow mesenchymal microenvironment makes it an ideal candidate for a more representative drug screening platform, this paper aims to do this by developing mesenchymal spheroids, which will support the culture of patient derived leukaemia samples allowing patient-specific therapeutic investigation.23\n\n\nMethods\n\nEthical approval\n\nPatient-derived leukaemia blasts were obtained from the Newcastle Biobank (REC reference number 07/H0906/109+5). All samples were obtained following written informed consent. Animal data shown here reflect unpublished data that were existing from past experiments. PDX samples used in this project are from our cryopreserved PDX bank where the samples were generated in previous/earlier projects. All animal studies were carried out in accordance with UK Animals (Scientific Procedures) Act, 1986 under project licence P74687DB5 following approval of Newcastle University animal ethical review body (AWERB).\n\nPatient derived ALL samples were cultured on 2D monolayers of MSC cells. MSC were seeded on Matrigel coated 48 well plates at 2×104 cells/0.5 mL/2 cm2 in their respective media (MSC media, Table 1). After 24 hours, the media was aspirated and the cells rinsed with 1 mL SFEM, Leukaemia blasts were then plated at a density of 0.25×105 cells/mL per well suspended in SFEMII.\n\nMSC cells were transferred to a 6-well low adhesion plate for suspension culture and cultured in 2 mL MSC media (Table 1) 3D for 48 hours, after which, MSC spheres were seen to form. Spheroids obtained thus were irregular in shape and therefore optimised experiments using Aggrewell400 plates were used to create uniform MSC spheres. Aggrewell plates were prepared by rinsing each well with 0.5 mL of Aggrewell rinsing solution, spinning at 1500 g for 5 minutes and aspirating. MSC cells were seeded at varying concentrations depending on desired sphere size. Cells were split using 1x trypsin solution and seeded in single cells on Aggrewell plates at desired concentration in MSC media. Aggrewell plates were spun at 100g for 3 minutes to ensure all cells reached the bottom of the microwells. After 48 hours, MSC spheroids were ready for harvest and transferred manually for culture onto low adhesion plates. Using a stereomicroscope fitted within a Class II biological safety cabinet the spheroids were transferred using a Gilson p1000 pipette tip with the end cut off using sterile scissors.\n\nLeukaemia cells from PDX samples were co-cultured with the MSC spheres in SFEMII at 2×105 cells/mL in 5 ml of media in low adhesion 48 well plates over a 5 day period. On day of harvest, the cells and spheroids were harvested into a 30 ml conical flask. This was centrifuged at 1500 RPM for 5 minutes. After aspirating the supernatant, 0.5 ml of X1 Trypsin EDTA was added and cultures incubated at 37°C, 5% CO2 for 2 minutes. Cells were mechanically dissociated into single cells by resuspending with a pipette. To the cells 4ml of FBS was added and this was centrifuged at 1500 RPM for 5 minutes. Supernatant was aspirated and cells were resuspended in 1ml of fresh SFEM media. During cell count leukaemia cells were distinguished from MSC based on their morphology and size difference, the MSC are 3-4 times the size of leukaemia cells.\n\nInvestigation into Dexamethasone mediated treatment resistance was carried out through cell fate tracing. 10 million blasts were span at 500 g and resuspended in 10 mL of 5 uM CellTrace violet, incubated for 20 minutes at 37 degrees Celsius, 1 mL FBS was added and cells were span once more at 500 g for 5 minutes and resuspended in fresh SFEMII. Cells were then plated on 3D MSC spheres in 48 well plates as described above. 5 nM Dexamethasone (stock dissolved in ethanol to generate 10 mM Dexamethasone concentration, working concentration of dexamethasone was made by performing serial dilutions using SFEM media) was added to desired wells and blasts were cultured for 5 days. Experiments include 3 biological replicates with each replicate including 3 experimental replicates.\n\nRNA was purified from cell pellets (viable cells centrifuged at 5000 g × 5 minutes followed by aspiration of supernatant media) using the RNeasy mini kit. Cells were resuspended in buffer RLT plus (containing 10 μl beta-mercaptoethanol/10 ml RLT) at either <5×106 cells/350 μl buffer RLT plus or 1×107 cells/600 μl buffer RLT plus. 1 volume of 70% ethanol was added to the RNA lysate and 700 μl of the cell suspension was transferred to a RNeasy mini spin column placed in a 2 ml collection tube, this was centrifuged at maximum speed for 30 seconds and the flow through discarded. The RNase-free DNase kit was used to remove contaminating DNA. 10 μl DNase I stock solution was added to 70 μl buffer RDD and mixed by gently inverting the tube, this solution was added directly to the column membrane and the sample incubated at room temperature for 15 minutes. 350 μl buffer RW1 was added and centrifuged at maximum speed for 30 seconds, the flow through was discarded. Next, 500 μl buffer RPE was added to the spin column, centrifuged at maximum speed for 30 seconds, and flowthrough discarded. Another 500 μl buffer RPE was added to the spin column, centrifuged at maximum speed for 2 minutes and the flow through discarded. The RNeasy spin column was transferred to a new 2 ml collection tube and centrifuged at maximum speed for 1 minute with the column lid open to dry the membrane. 30 μl RNase free water was then added to elute the sample, centrifuged for 1 minute at maximum speed and the flow through was collected. The RNA yield and quality was checked using the nanodrop ND-1000 spectrophotometer.\n\nRevertAidTMH Minus First Strand cDNA Synthesis Kit was used to synthesise cDNA from the RNA isolated. 500 ng RNA was collected and added to RNase/DEPC free water to a final volume of 11 μl. 1 μl (dN)6 (200 mg/l) random hexamers was added, mixed gently by inverting the vial and briefly centrifuged. Using a GeneAmp PCR system 2700 the sample was incubated at 65°C for 5 minutes, after which the sample was immediately placed on ice, 8 μl of the master mix (Table 2) was added, the samples were vortexed and briefly centrifuged. The samples were placed back in the PCR machine to incubate at 25°C for 10 minutes, 42°C for 60 minutes and 75°C for 10 minutes to terminate the reaction. The product was either used immediately for RT-PCR or stored at -20°C for short term storage.\n\nUpon receipt, primers were reconstituted in RNase/DNase free water to a final stock concentration of 100 μM. PCR Mix (Table 3) was loaded onto PCR plate in triplicates. The plate was sealed and centrifuged for 1 minute at 1000 RPM and placed in an applied Biosystems 7900HT Sequence Detection System. ViiA7TM System was used to run the qRT-PCR plate and cycle threshold (Ct) and melting curves were obtained for analysis.\n\nCostings of in vitro models were estimated using list prices of media, reagents and other materials associated with developing and using the models for a 2 drug combination dose finding assay. This included 5 biological replicates, 3 experimental repeats with each having 3 technical repeats with a total number of experiments of 45. These costs were divided into the following categories: Production of clinical/patient-derived leukaemia samples; in vitro CRISPR/RNAi organoid generation; Dose/IC50 finding for 2 drug combination; and evaluation of drug combination.\n\nIn vivo costings were calculated using figures obtained from the Comparative Biology Centre, Newcastle University. This included fixed costs including mouse purchase from an NSG in-house colony, housing and IVIS use. Comparative in vivo equivalents to the in vitro design accounted for: production of PDX engrafted tissue (5 PDX, 6 NSG mice/PDX); in vivo CRISPR/RNAi (control and treatment groups, 5 NSG mice each); Dose finding for 2 drug combination (3 mice per sex); and evaluation of the combination (4 arms, 6 mice per arm). All total costings were calculated using Microsoft Excel and figures were created using Microsoft Excel.\n\nMicroscopy images were analysed using the Windows download bundled with Java 8. Version 1.4.3. Real time qPCR data were captured and analysed using the StepOne Plus Real-Time PCR System (ThermoFisher Catalog No: 4376600) and Software package StepOne Software v23.\n\n\nResults\n\nWe first show that BM-MSC/leukaemia 2D co-culture platform generates drug dose response data that is comparable with drug response observed in vivo in mouse models (Figure 1).24 These data where we compare our in vitro data with in vivo data (unpublished data existing in the lab from earlier studies) show the predictive capacity of our 2D co-culture platform against mouse experiments. Improved in vivo predictability is important in ex vivo animal replacement models if these platforms are to meaningfully replace mouse experiments. We further perform pilot studies where we show that such ex vivo co-culture platforms have the ability to bring about significant replacement of animals for PDX models, which are the current gold standard preclinical leukaemia model (Figure 2). In order to fully recapitulate the complexity of the leukaemia niche, a 3D Model with improved biomimicry is required. 3D BM-MSC-spheroids were created by subculturing cells onto low adhesion non-tissue culture treated plates (Figure 3a,b). These spheroids were tested for expression levels of their respective niche markers using QT-PCR. It was found that both niche sphere types expressed mesenchymal markers CD90, CDH2 and Nestin (Figure 3c) similar to their 2D counterparts.\n\nLeukaemia cells from patient A is a responder to ABT199 (targets BCL2 in BCL2 positive ALL) and Dexamethasone drug combination in vitro and in vivo. On the other hand leukaemia cells from patient B do not respond to this drug combination in vitro and subsequently poor response is also noted in vivo using PDX mice models. Error bars shown refer to standard error (SE) using 4 mice per treatment group. Total flux is a surrogate of blast number in vivo. Dexamethasone and ABT-199 are used in the clinics to treat ALL.\n\nKey benefits of the animal replacement organoid models include species specificity and consequently higher biomimicry.\n\nDevelopment and validation of MSC spheroid cultures through 3D suspension culture. Aggrewell plates allow for the creation of uniform 3D spheroids from single cell suspension of the desired cell-type. SD from three independent experiments (using MSC from same patient) are plotted as error bars. 2D (a), scale bar = 100 μM and 3D MSC (b), scale bar = 200 μM after 96 hours of growth. (c) mesenchymal marker levels quantified through qPCR and normalised to GAPDH. (d,e) MSC spheroids obtained through Aggrewell plate culture. Scale bar = 1000 μM (f) Cell seeding densities directly correlate to size of spheroids obtained. Data captured following 48 hours of seeding.\n\nAggrewell400 plates are designed for higher throughput and more uniform production of MSC spheres. Each well on these plates house 400 conical microwells, funnelling single cells and encouraging them to conglomerate into spheroids (Figure 3d). These microwells were utilised to create uniform niche spheres allowing standardisation of the technique when producing assays involving 3D spheres (Figure 3e). A direct correlation was observed with seeding cell density and size of spheroids with higher seeding densities resulting in generation of more uniformly sized spheroids (Figure 3f).\n\nALL blasts from three different clinical samples were co-cultured with BM-MSC in routine 2D cultures and with 3D spheroids. Cell proliferation was monitored over a 5 day period through tryphan blue cell counts. We observed that when co-cultured with BM-MSC-spheroids the blasts showed 2-fold higher cell proliferation compared to 2D co-cultures (Figure 4a-c). Next, we repeated the co-cultures under treatment with the steroid Dexamethasone which is used to treat ALL. We observed that blasts showed marked reduction in sensitivity to Dexamethasone in 3D MSC organoid co-cultures compared to 2D co-cultures (Figure 4d-f). These data show the advantage of 3D organoid based co-culture systems over routine 2D cultures in two respects: 1. Achieving blast cycling which is essential for the action or testing of majority of anti-cancer treatments 2. Modelling treatment resistance in the laboratory – a major current day clinical challenge in cancer management. Finally, we perform a costs analysis and show improved financial sustainability in our ex vivo organoid platform compared to animal models (Figure 5).\n\nN= 3 Error bars = SD. Increased growth kinetics of L49120 (a,d), MS40 (b,e) and L707D (c,f) across a 5 day period on 3D MSC feeder cells compared to a 2D monoculture. L49120, MS40 and L707D refer to three different patient samples, i.e., 3 biological repeats. Dexamethasone treatment is less effective in a 3D microenvironment (d,e,f).\n\n\nDiscussion\n\nRelapse is still a major concern within the treatment of leukaemia, relapse disease is often chemo resistant, meaning many therapies cease working in patients with relapse disease leading to an increased mortality.3 Differing cytogenic abnormalities in this disease are known and patient stratification and precision medicine are implemented in an effort to treat individual disease in the most effective way possible, allowing weaknesses in tumour biology to be exploited therapeutically. However, high drug attrition rates remain a challenge when developing such improved treatments in ALL. High drug attrition has been attributed to preclinical models not being clinically translatable. Key barriers hindering bench to bedside translation when using complex in vivo models include species specificity, high financial costs, and lengthy experiments. Patient samples do not always engraft in mice. Samples that do engraft form successful xenograft models in 3-6 months at which point drug testing can be started which takes another 3-4 weeks.\n\nA 3D BM-mesenchyme-leukaemia spheroid model will provide researchers with a biomimetic platform where clinical samples can be cultured within the context of their microenvironment, and patient specific drug testing performed within a week. Besides delivering drug response data within clinically relevant timeframes the relative simplicity of ex vivo spheroid models mean that they are tractable, transferrable and sustainable. Consequently, such models have wide applications in translational research in academia and industry alike. These models can be set up in laboratories and SMEs locally, nationally and globally that do not contain infrastructure for animal procedures. Following further extensive validation, such spheroid biobanks also have the potential to be embedded within clinical trials and healthcare systems for the purposes of detecting responders as well as to aid risk stratification.\n\nIn this paper we show that our 2D model itself brought about significant animal replacement locally. Leukaemia research at Newcastle requires 3600 animal procedures every year. Local metrics confirmed that using the 2D pilot approach we replaced mice in 67 drug tests last year. Given minimum of 40 animals are needed per drug test we replaced 2680 animals (out of 3600) which constituted a minimum 73% local animal replacement. To improve biomimicry and consequently translatability and transferability of our model, we develop 3D BM-mesenchyme spheroids. We show that these 3D spheroids show superior ability in supporting culture of leukaemia patient samples. We also show sensitivity of leukaemia cells to drugs such as dexamethasone is reduced when these cells are being co-cultured with 3D spheroids. This means that compared to 2D MSC cultures, MSC spheroids provide superior protection to leukaemia cells against dexamethasone treatment. are appropriate in modelling treatment resistance which remains a major clinical challenge in cancer management. This simple and tractable 3D prototype will form the first steppingstone in developing next generation 3D preclinical models with improved in vivo and patient drug response predictability. Such sustainable ex vivo models will ultimately replace existing moderate severity procedures in leukaemia research with models that successfully impact clinical outcome.", "appendix": "Data availability\n\nMendeley Data. A human mesenchymal spheroid prototype to replace moderate severity animal procedures in leukaemia drug testing. https://doi.org/10.17632/56npmkbpfb.1. 24\n\nThis project contains the following underlying data:\n\n- 3D spheres.tif\n\n- 3D Sphere.tif\n\n- drug combination.xlsx\n\n- in vivo Vs in vitro costs.xlsx\n\n- Raw Figure Data.xlsx\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nThis study was funded by NC3Rs grants to DP. We thank NECCR for funding technical assistance at Newcastle University. We thank Professor Josef Vormoor, Professor Olaf Heidenreich and Dr. Kenneth Rankin, Newcastle University for provision of samples.\n\n\nReferences\n\nO'Connor D, et al.: Infection-related mortality in children with acute lymphoblastic leukemia: an analysis of infectious deaths on UKALL2003. Blood. 2014; 124: 1056–1061. 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PubMed Abstract \| Publisher Full Text\n\nBaryawno N, et al.: A Cellular Taxonomy of the Bone Marrow Stroma in Homeostasis and Leukemia. Cell. 2019; 177: 1915–1932.e16. PubMed Abstract \| Publisher Full Text\n\nDuan CW, et al.: Leukemia propagating cells rebuild an evolving niche in response to therapy. Cancer Cell. 2014; 25: 778–793. Publisher Full Text\n\nEbinger S, et al.: Characterization of Rare, Dormant, and Therapy-Resistant Cells in Acute Lymphoblastic Leukemia. Cancer Cell. 2016; 30: 849–862. PubMed Abstract \| Publisher Full Text\n\nAgarwal P, et al.: Mesenchymal Niche-Specific Expression of Cxcl12 Controls Quiescence of Treatment-Resistant Leukemia Stem Cells. Cell Stem Cell. 2019; 24: 769–784.e6. PubMed Abstract \| Publisher Full Text\n\nSkardal A, Shupe T, Atala A: Organoid-on-a-chip and body-on-a-chip systems for drug screening and disease modeling. Drug Discov Today. 2016; 21: 1399–1411. PubMed Abstract \| Publisher Full Text\n\nDrost J, Clevers H: Organoids in cancer research. Nat Rev Cancer. 2018; 18: 407–418. Publisher Full Text\n\nGarreta E, et al.: Fine tuning the extracellular environment accelerates the derivation of kidney organoids from human pluripotent stem cells. Nat Mater. 2019; 18: 397–405. Publisher Full Text\n\nShih YR, et al.: In vivo engineering of bone tissues with hematopoietic functions and mixed chimerism. Proc Natl Acad Sci U S A. 2017; 114: 5419–5424. PubMed Abstract \| Publisher Full Text\n\nPal D, Wilson A, Blair H, et al.:A human mesenchymal spheroid prototype to replace moderate severity animal procedures in leukaemia drug testing. [Dataset]. Mendeley Data. 2022; V1. Publisher Full Text" }	[ { "id": "158193", "date": "19 Dec 2022", "name": "Simon E. Richardson", "expertise": [ "Reviewer Expertise Leukaemia biology", "disease modelling", "stem cells", "epigenetics", "preclinical drug testing." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis manuscript by Wilson et al provides a rationale, methodological details and pilot data supporting the use of 3D human mesenchymal stroma cell (MSC) spheroid co-cultures in the preclinical testing of drugs for acute leukaemia. The authors provide a detailed rationale for the potential benefits of this system, including: i) reduction in animal use/suffering; ii) reduced costs; and iii) the ability to model the protective impact of bone marrow micro-environmental influences in abrogating drug efficacy in vitro, reducing the number “false positive” drug hits taken forward to in vivo model systems. The authors provide data showing the equivalence of 2D in vitro drug sensitivity with PDX models (n=2) and compare 2D vs 3D co-culture of human leukaemias (n=3) in proliferation and dexamethasone sensitivity assays, showing consistently enhanced proliferation and reduced dexamethasone sensitivity in the 3D system. They also present methodological data on the establishment of MSC spheroids as well as modelling on the potential reduction in animal use and costs based on local experience.\nOverall, the rationale to develop more physiologically relevant in vitro co-culture systems is a very worthwhile endeavour with clear potential benefits for animal welfare, costs and in streamlining drug development. It also has the potential advantage of being used in the study of leukaemias that do not engraft in current immunosuppressed mouse models.\nIn my opinion the work has three major limitations that would benefit from either additional data or acknowledgement in the discussion.\n\nWhilst this work demonstrates the feasibility of the approach, it is not possible from the data presented to conclude that 3D systems perform better than either 2D co-culture or PDX systems. Specifically, the data presented uses a small number of patient samples (n=2 or 3) and drugs (Dexamethasone ± ABT199). The three systems are not directly compared and no example of an in vitro drug hit that proves “false-positive” when tested in vivo is tested in parallel 2D/3D co-culture.\n\nThere are a number of potential technical limitations of the co-culture approach that are not acknowledged: i) the requirement for access to a source of MSCs which are not readily available to most labs; ii) certain drugs (notably cytotoxic chemotherapy) are likely to have direct toxicity to the MSCs themselves; iii) the lack of a vascular component to this system might result in artificially high levels of physical protection from drug exposure by the 3D niche (i.e. potential for false negatives).\n\nThe potential costs and animal savings presented are feasible, but likely at the upper end of the spectrum and will differ between institutions. For example, the time to generate a novel PDX is indeed many months, but once established PDX models in ALL engraft much more quickly and reproducibly and established PDX material is readily available. In vivo toxicity testing is not needed for established drugs being re-purposed; conversely novel chemical agents will still need some in vivo toxicity data prior to regulatory approval for first-in-human studies.\n\nMinor points:\nIn the introduction it would be helpful to specify the precise leukaemia being studied and at which age. In particular, relapse rates in adult B-ALL are much higher than those presented, which are indicative of the paediatric setting.\n\nIn the 2D co-culture methods, please provide a reference on how to process MSCs and specify the final volume of media per well used to plate the leukaemia cells.\n\nFor the response data presented in figures 1 and 4, please clarify in the legend or methods the nature of the readouts and timepoints used.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate? Not applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10408", "date": "28 Nov 2023", "name": "Deepali Pal", "role": "Author Response", "response": "This manuscript by Wilson et al provides a rationale, methodological details and pilot data supporting the use of 3D human mesenchymal stroma cell (MSC) spheroid co-cultures in the preclinical testing of drugs for acute leukaemia. The authors provide a detailed rationale for the potential benefits of this system, including: i) reduction in animal use/suffering; ii) reduced costs; and iii) the ability to model the protective impact of bone marrow micro-environmental influences in abrogating drug efficacy in vitro, reducing the number “false positive” drug hits taken forward to in vivo model systems. The authors provide data showing the equivalence of 2D in vitro drug sensitivity with PDX models (n=2) and compare 2D vs 3D co-culture of human leukaemias (n=3) in proliferation and dexamethasone sensitivity assays, showing consistently enhanced proliferation and reduced dexamethasone sensitivity in the 3D system. They also present methodological data on the establishment of MSC spheroids as well as modelling on the potential reduction in animal use and costs based on local experience. Overall, the rationale to develop more physiologically relevant in vitro co-culture systems is a very worthwhile endeavour with clear potential benefits for animal welfare, costs and in streamlining drug development. It also has the potential advantage of being used in the study of leukaemias that do not engraft in current immunosuppressed mouse models. In my opinion the work has three major limitations that would benefit from either additional data or acknowledgement in the discussion. Whilst this work demonstrates the feasibility of the approach, it is not possible from the data presented to conclude that 3D systems perform better than either 2D co-culture or PDX systems. Specifically, the data presented uses a small number of patient samples (n=2 or 3) and drugs (Dexamethasone ± ABT199). The three systems are not directly compared and no example of an in vitro drug hit that proves “false-positive” when tested in vivo is tested in parallel 2D/3D co-culture. Response: We provide additional data comparing 2D co-culture and in vivo data with 3D sphere co-culture data. We find that patient-derived leukaemia cells show reduced sensitivity in 3D co-culture settings compared to 2D co-cultures, potentially indicating the superior ability of 3D models to screen out 2D “false-positive” hits. Furthermore, we show that patient-derived leukaemia cells from a leukaemia cytogenetic subgroup, subgroup A, showing high sensitivity to dex-ABT-199 combination in vivo, shows high combination sensitivity in 3D in vitro co-cultures. Similarly, we show that patient-derived leukaemia cells from the subgroup B shows reduced sensitivity to dex-ABT-199 combination both in vivo and in vitro, in 2D and 3D co-culture settings. We add the following text within the main manuscript: Co-cultures of 3D MSC with patient-derived leukaemia cells how high in vivo predictivity Improved in vivo predictivity is a key requisite for ex vivo animal replacement models, if these platforms are to effectively replace mouse experiments. We confirmed that BM-MSC/leukaemia 2D co-culture platform generated drug dose response data that was largely comparable with drug response observed in vivo in PDX-mouse models (Fig 4.a). Furthermore, we reveal that when leukaemia cells are co-cultured with 3D BM-MSC-spheroids they show greater reduced sensitivity, compared to the 2D co-culture arm, against dexamethasone, ABT-199 and the combination in a sample subgroup that exhibited reduced in vivo efficacy (previously unpublished data existing in the lab from earlier studies). This reveals the ability of our 3D co-culture model in being able to screen off false-positive hits derived from 2D models, consequently minimising in vitro to in vivo attrition rates. We further confirm that when co-cultured with 3D MSC spheres, ALL cells expectedly in 3D-MSC spheroid-co-cultures, however, fail to increase in cell counts on day five, in the dexamethasone treatment arm (Fig.4.b). Moreover, we show that neither dexamethasone, nor ABT-199, a molecularly targeted therapy against BCL-2 positive blood cancers, have any effect on survival of MSC, when treated for five days within a 3D sphere format (Figure 4.c). 2. There are a number of potential technical limitations of the co-culture approach that are not acknowledged: i) the requirement for access to a source of MSCs which are not readily available to most labs; ii) certain drugs (notably cytotoxic chemotherapy) are likely to have direct toxicity to the MSCs themselves; iii) the lack of a vascular component to this system might result in artificially high levels of physical protection from drug exposure by the 3D niche (i.e. potential for false negatives). Response: We provide additional data and show that neither dexamethasone, nor ABT-199 have any effect on survival of MSC when treated for five days within a 3D sphere format. We include the following text within Results, Co-cultures of 3D MSC with patient-derived leukaemia cells how high in vivo predictivity: Moreover, we show that neither dexamethasone, nor ABT-199, a molecularly targeted therapy against BCL-2 positive blood cancers, have any effect on survival of MSC, when treated for five days within a 3D sphere format (Figure 4.c). We address remaining of points i), ii) and iii) in an additional “Study Limitations” section. Study Limitations Here we use primary human MSC, which indeed may be difficult to source, and which may lead to donor variability. Using induced pluripotent stem cell (iPSC) derived MSC to co-culture patient-derived leukaemia14 may be an attractive alternative, and would furthermore enable assay standardisation, thereby improving data reproducibility. Moreover, although we show that MSC showed no adverse effect to dexamethasone and ABT-199 treatment, many anti-cancer treatments notably cytotoxic chemotherapy may cause direct toxicity to the MSC. However, this further emphasises the need to include key microenvironment cell types when conducting anti-cancer drug testing, especially given cancer treatments influence interactions between leukaemia and their surrounding niche, with many cytotoxic treatments affecting both leukaemia and bone marrow cells in patients. Furthermore, we acknowledge that an ideal representation of 3D biomimetic human cell-based model would include capturing both mesenchymal and vascular bone marrow components within an immune-responsive context. 3. The potential costs and animal savings presented are feasible, but likely at the upper end of the spectrum and will differ between institutions. For example, the time to generate a novel PDX is indeed many months, but once established PDX models in ALL engraft much more quickly and reproducibly and established PDX material is readily available. In vivo toxicity testing is not needed for established drugs being re-purposed; conversely novel chemical agents will still need some in vivo toxicity data prior to regulatory approval for first-in-human studies. Response Indeed, the time taken to generate a novel PDX sample can span between months to a year, and this time is shortened when PDX samples are serially transplanted in subsequent mouse models. Nevertheless, some samples, particularly those belonging to good risk groups, such as high hyperdiploid samples, still require between 6 months – 1 year to engraft successfully, before any drug response testing can begin. We agree that in vivo toxicity is not needed for drugs being re-purposed, unless these involve testing of new combinatorial regimens, and furthermore note recent announcement by the U.S. Food and Drug Administration (FDA) that new drugs would not be mandated to be tested in animals prior to first-in-human trials. We have reflected this in the manuscript within Discussion via the following additional text Not all leukaemia samples engraft in mice. We have shown, in a previous study, that the 2D MSC co-culture system cultured acute myeloid leukaemia (AML) cells from t(8;21)-positive samples8, which failed to engraft any mouse model available at the time. Samples that do engraft form successful xenograft models, yielding PDX samples in 3-6 months at which point drug testing can be started which takes another 3-4 weeks. Indeed, although the time taken to generate a novel PDX sample can span between months to a year, this time is often shortened when cells from established PDX samples are serially transplanted in subsequent mouse models, to aid generation of PDX lines. Nevertheless, some samples, particularly those belonging to good risk groups, such as high hyperdiploid samples, still require between 6 months – 1 year to engraft successfully before any drug response testing can begin. Moreover although in vivo toxicity assays are not needed for drugs being re-purposed, indicating this to be an area where animal-replacement technologies may perhaps not be relevant, animal testing has been required prior to regulatory approval being given for first-in-human studies. Furthermore, toxicity studies which use animal models are indicated when testing new combinatorial treatment regimens comprising repurposed drugs. This is because tolerability of drug combinations is distinct from that of component single drugs. Nevertheless, the U.S. Food and Drug Administration (FDA) has recently announced that new drugs would not be mandated to be tested in animals prior to first-in-human trials, and an FDA-wide program is furthermore prioritising development of human-relevant methods, with high in vivo predictivity, to replace, reduce and refine animal models in drug testing. This further corroborates the timeliness of prioritising development of transformative human cell-based, non-animal technologies towards preclinical drug testing. Most specifically, a key advantage of our prototype approach would be in medium-high throughput screening of single and combinatorial compounds, particularly as pre-in vivo screens, to prevent “false positive” 2D in vitro hits, such as that observed in Figure 4, from progressing towards in vivo drug efficacy testing. Minor points: In the introduction it would be helpful to specify the precise leukaemia being studied and at which age. In particular, relapse rates in adult B-ALL are much higher than those presented, which are indicative of the paediatric setting. Response: We have addressed this both in the abstract and in the introduction via the following text: Abstract: Using childhood acute lymphoblastic leukaemia (ALL) as a proof-of-concept paradigm, here we develop an in vitro approach to co-culture patient-derived leukaemia cells with 3D mesenchymal stroma cell spheroids, MSC-spheroids. Introduction: Acute lymphoblastic leukaemia (ALL) management in children is hindered by two chief obstacles: 1. Treatment toxicity1,2 2. 15-20% of patients go into relapse, when the disease can be treatment resistant3. In the 2D co-culture methods, please provide a reference on how to process MSCs and specify the final volume of media per well used to plate the leukaemia cells. This has been provided as below: 2D Niche-Blast co-culture Patient derived ALL samples were cultured on 2D monolayers of MSC cells using a protocol published in an earlier study9. MSC were used between passages ranging from p2 – p5. MSC were seeded on Matrigel coated 48 well plates at 1x104 cells/0.5mL/cm2 in their respective media (MSC media, Table 1). After 24 hours, the media was aspirated, and the cells rinsed with 1mL SFEM. Leukaemia cells were then plated onto the MSC layers, at a density of 0.25x105 cells/1 mL per well, suspended in SFEM media. For the response data presented in figures 1 and 4, please clarify in the legend or methods the nature of the readouts and timepoints used. Figure 1, now Figure 4. Readouts have been clarified in legend as follows: “In vitro data readout includes viable cell counts; in vivo data (right hand graphs) is shown as bioluminescence imaging from luciferase expressing PDX cells in total flux(p/s). Total flux is a surrogate of blast number in vivo.” Figure 4, now Figure 3. Readouts have been clarified in legend as follows: Figure 3. 3D MSC-spheroids show confer higher proliferation and improved survival onto patient-derived ALL. Growth kinetics, measured via viable cell counts of ALL samples (a,d) L49120, (b,e) MS40 and (c,f) L707D across a 5 day period, with and without low dose dexamethasone (10nM) pressure, as co-cultures with 3D MSC spheres versus 2D MSC-co-culture. L49120, MS40 and L707D refer to three different patient samples, i.e., 3 biological repeats. (d,e,f) Dexamethasone treatment, data shown for 5nM treatment, is less effective in a 3D microenvironment. N= 3. g. Flow cytometry data, representative plots, showing propidium iodide, PI, positive ALL cells (sample L707) following five days of 10nM dexamethasone treatment. The column graph shows % ALL cells stained positive for PI. N = 2" } ] }, { "id": "159730", "date": "31 Jan 2023", "name": "Helen Wheadon", "expertise": [ "Reviewer Expertise Stem cells", "leukaemia modelling", "signalling", "pre-clinical drug testing." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe manuscript by Wilson et al. provides a convincing argument for the requirement of additional in vitro models to improve drug testing in acute lymphocytic leukaemia (ALL) and reduce the reliance on Xenograft mouse models. Preliminary data is presented supporting the use of 3D human mesenchymal stroma cell (MSC) spheroid/primary ALL co-cultures for the use in preclinical testing of drugs. Proof of principle data is provided showing that simple 2D co-culture on MSCs gives a similar range of drug responses to dexamethasone and the BCL2 mimetic drug ABT199 as observed in in vivo studies for 2 primary ALL patient samples. The authors then show optimisation data using Aggrewell plates, demonstrating uniform spheroid formation and a corresponding increase in spheroid diameter when cell seeding density is increased. The final figure then compares primary ALL cell proliferation in 2D and 3D culture and drug responses to dexamethasone. Using 3 independent patient samples they show that MSC spheroids support cell proliferation over a 5 day time frame and provide protection to dexamethasone. These are important observations and support the introduction of a more physiologically relevant humanised system for drug screening prior to conducting in vivo studies. Another potential advantage would be if the authors had data demonstrating that the system supports patients' samples which do not engraft, as there is a high engraftment failure rate in Xenograft models with not all patient samples being successful. This could potentially lead to more robust data on patient’s responses. Given patient heterogeneity mouse models may be biasing certain traits/genetics due to the engraftment limitations observed.\nOverall, the system if adopted by other researchers could potentially reduce animal numbers and suffering, reduce the cost and select the most promising drugs for additional testing. It would therefore have impact on replacement and reduction of mouse experiments.\nHowever, there are some limitations in the results shown that would be strengthened by additional experiments:\nThe MSC are characterised by gene expression, it would be good to include flow cytometry data; CD90+105+73+34-45- are key markers demonstrating stemness. The culture conditions using FCS can also lead to changes in the MSCs so it would be good to include passage number. Figure 3 live/dead staining of the spheroids especially for the size used in the drug treatment assays in Figure 4 would be important to include.\n\nData showing that dexamethasone and ABT199 do not detrimentally affect the MSC monoculture and MSC spheroids would also be important to include.\n\nFigure 4 looks at proliferation, apoptosis assays would be good to include to show whether cell death was occurring.\n\nData showing similar results in ALL patient samples known to fail to engraft in vivo would strengthen the rationale for adopting the 3D system.\n\nAlthough the potential costs and animal savings presented are feasible, the system is quite simplistic and is more likely to be incorporated as an additional test rather than be adopted as a replacement. It would be more attractive if the time frame could be extended and more details on the application for high throughput screening development was discussed. Also, in vivo toxicity testing of novel compounds will still be required in order for regulatory approval for any Phase 1 clinical trial in humans to be progressed.\nMinor corrections:\nResearch highlights - blasts would only be appropriate for certain types of leukaemia, either need to define it’s ALL specifically or change to stem/progenitor.\nScientific benefit: A 3D spheroid-based approach with improved clinical relevance for ex vivo co-culture of primary patient leukaemia samples for further investigation into cancer biology such as stem/progenitor cell-niche interactions, stem/progenitor cell proliferation and treatment resistance.\n3Rs benefit: To replace moderate severity animal (mice) procedures in leukaemia research and early pre-clinical drug testing.\nCurrent applications: Exploration of leukaemia biology such as stem/progenitor cell proliferation, stem/progenitor cell -niche interactions, niche-impacted treatment resistance and obtain drug response data.\nIntroduction:\nNeed to define at the start that you are talking about ALL as it reads like a general overview which means some of the % incidence rates etc. are not correct. It would be good to mention what type of ALL as well, is it B-ALL, T-ALL, childhood, adult?\n\nParagraph 3 page 3: Most research groups would use primary patient samples in vitro prior to moving to in vivo models. This should be included.\n\nParagraph 2 Page 4: Remove blasts and replace with LSCs also include disease type chronic myeloid leukaemia.\n\nPage 11 ‘such spheroid biobanks’: do they actually exist? Or are you planning on constructing a biobank?\n\n‘MSC spheroids provide superior protection to leukaemia cells against dexamethasone treatment. are appropriate in modelling treatment resistance which remains a major clinical challenge in cancer management’. Is this statement correct, not really looking at resistance, just reduced sensitivity to the drugs at that dose when in 3D rather than 2D culture. Need to amend and extend to include how you would look at resistance i.e. relapsed patients and the potential to interrogate resistance mechanisms.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate? Not applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10409", "date": "28 Nov 2023", "name": "Deepali Pal", "role": "Author Response", "response": "The manuscript by Wilson et al. provides a convincing argument for the requirement of additional in vitro models to improve drug testing in acute lymphocytic leukaemia (ALL) and reduce the reliance on Xenograft mouse models. Preliminary data is presented supporting the use of 3D human mesenchymal stroma cell (MSC) spheroid/primary ALL co-cultures for the use in preclinical testing of drugs. Proof of principle data is provided showing that simple 2D co-culture on MSCs gives a similar range of drug responses to dexamethasone and the BCL2 mimetic drug ABT199 as observed in in vivo studies for 2 primary ALL patient samples. The authors then show optimisation data using Aggrewell plates, demonstrating uniform spheroid formation and a corresponding increase in spheroid diameter when cell seeding density is increased. The final figure then compares primary ALL cell proliferation in 2D and 3D culture and drug responses to dexamethasone. Using 3 independent patient samples they show that MSC spheroids support cell proliferation over a 5 day time frame and provide protection to dexamethasone. These are important observations and support the introduction of a more physiologically relevant humanised system for drug screening prior to conducting in vivo studies. Another potential advantage would be if the authors had data demonstrating that the system supports patients' samples which do not engraft, as there is a high engraftment failure rate in Xenograft models with not all patient samples being successful. This could potentially lead to more robust data on patient’s responses. Given patient heterogeneity mouse models may be biasing certain traits/genetics due to the engraftment limitations observed. Overall, the system if adopted by other researchers could potentially reduce animal numbers and suffering, reduce the cost and select the most promising drugs for additional testing. It would therefore have impact on replacement and reduction of mouse experiments. However, there are some limitations in the results shown that would be strengthened by additional experiments: The MSC are characterised by gene expression, it would be good to include flow cytometry data; CD90+105+73+34-45- are key markers demonstrating stemness. The culture conditions using FCS can also lead to changes in the MSCs so it would be good to include passage number. Figure 3 live/dead staining of the spheroids especially for the size used in the drug treatment assays in Figure 4 would be important to include. Response: Additional data addressing these points have been added in Figure 2. We include flow cytometry data showing MSC in 3D spheroid format express CD105, CD90 and CD73. We include additional data showing the expression of casein Calcein (live cell dye) and Hoechst-33342 (live and dead cell dye) in MSC spheroids at a diameter of 100 µM, spheroid size used in subsequent experiments. MSC passage number used was between p2 and p5, this has been clarified within Methods 2D Niche-Blast co-culture Patient derived ALL samples were cultured on 2D monolayers of MSC cells using a protocol published in an earlier study9. MSC were used between passages ranging from p2 – p5. Data showing that dexamethasone and ABT199 do not detrimentally affect the MSC monoculture and MSC spheroids would also be important to include. Response: We provide additional data and show that neither dexamethasone, nor ABT-199 have any effect on survival of MSC when treated for seven days within a 3D sphere format. We include the following text within Results, Co-cultures of 3D MSC with patient-derived leukaemia cells how high in vivo predictivity: Moreover, we show that neither dexamethasone, nor ABT-199, a molecularly targeted therapy against BCL-2 positive blood cancers, have any effect on survival of MSC, when treated for five days within a 3D sphere format (Figure 4.c). Figure 4 looks at proliferation, apoptosis assays would be good to include to show whether cell death was occurring. Response We provide additional data showing propidium iodide, PI staining in ALL cells following treatment with dexamethasone. We include following text within Results, Patient-derived leukaemia cells co-cultured with 3D MSC spheres show superior proliferation and enhanced treatment resistance. “We find that ALL cells show greater reduction in sensitivity to dexamethasone when co-cultured with 3D BM-MSC (Fig.3.d-f). This could be owing to potentially improved functional cell-cell contacts and physiologically relevant cell polarity being achieved in a 3D format. This might have caused increased 3D versus 2D MSC-conferred ALL proliferation and/or improved ALL survival. However, reduced sensitivity could also be due to altered drug availability within 3D spheroids. These data highlight the advantage of 3D organoid-based co-culture systems over routine 2D cultures in two respects: 1. Achieving higher ALL cell cycling which is essential for the action or testing of majority of anti-cancer treatments 2. Obtaining reduced drug sensitivity, possibly owing to improved treatment resistance– a major current day clinical challenge in cancer management. We further investigate the ability of MSC-spheroids to reduce ALL cell death and find that when treated with a higher dose of dexamethasone, that is, at 10nM, then both 2D and 3D MSC-spheroids support survival of leukaemia cells (Fig.3.g.). We now progress to reveal proof-of-confidence-in-application data, justifying the suitability of our platform in conduction combinatorial drug testing.” Data showing similar results in ALL patient samples known to fail to engraft in vivo would strengthen the rationale for adopting the 3D system. Response Unfortunately, we do not have ALL samples that fail to engraft. Nevertheless, we have previously shown the ability of the 2D MSC co-culture system to culture primary AML samples from t(8;21)-positive patients, which would not successfully engraft into mouse models available at the time. Given our finding that 3D-MSC-spheres provide superior support in in vitro proliferation of leukaemia cells, this study strengthens the rationale for adopting in vitro MSC-based co-culture systems. Text entered in Discussion: “Patient samples do not always engraft in mice. We have shown, in a previous study, that the 2D MSC co-culture system successfully cultured acute myeloid leukaemia (AML) cells from t(8;21)-positive samples8 which did not engraft any mouse model available at the time.” Although the potential costs and animal savings presented are feasible, the system is quite simplistic and is more likely to be incorporated as an additional test rather than be adopted as a replacement. It would be more attractive if the time frame could be extended and more details on the application for high throughput screening development was discussed. Also, in vivo toxicity testing of novel compounds will still be required in order for regulatory approval for any Phase 1 clinical trial in humans to be progressed. Response We have aimed to successfully address these points in the discussion, via the additional text: Not all leukaemia samples engraft in mice. We have shown, in a previous study, that the 2D MSC co-culture system cultured acute myeloid leukaemia (AML) cells from t(8;21)-positive samples8, which failed to engraft any mouse model available at the time. Samples that do engraft form successful xenograft models, yielding PDX samples in 3-6 months at which point drug testing can be started which takes another 3-4 weeks. Indeed, although the time taken to generate a novel PDX sample can span between months to a year, this time is often shortened when cells from established PDX samples are serially transplanted in subsequent mouse models, to aid generation of PDX lines. Nevertheless, some samples, particularly those belonging to good risk groups, such as high hyperdiploid samples, still require between 6 months – 1 year to engraft successfully before any drug response testing can begin. Moreover although in vivo toxicity assays are not needed for drugs being re-purposed, indicating this to be an area where animal-replacement technologies may perhaps not be relevant, animal testing has been required prior to regulatory approval being given for first-in-human studies. Furthermore, toxicity studies which use animal models are indicated when testing new combinatorial treatment regimens comprising repurposed drugs. This is because tolerability of drug combinations is distinct from that of component single drugs. Nevertheless, the U.S. Food and Drug Administration (FDA) has recently announced that new drugs would not be mandated to be tested in animals prior to first-in-human trials, and an FDA-wide program is furthermore prioritising development of human-relevant methods, with high in vivo predictivity, to replace, reduce and refine animal models in drug testing. This further corroborates the timeliness of prioritising development of transformative human cell-based, non-animal technologies towards preclinical drug testing. Most specifically, a key advantage of our prototype approach would be in medium-high throughput screening of single and combinatorial compounds, particularly as pre-in vivo screens, to prevent “false positive” 2D in vitro hits, such as that observed in Figure 4, from progressing towards in vivo drug efficacy testing. Minor corrections: Research highlights - blasts would only be appropriate for certain types of leukaemia, either need to define it’s ALL specifically or change to stem/progenitor. Scientific benefit: A 3D spheroid-based approach with improved clinical relevance for ex vivo co-culture of primary patient leukaemia samples for further investigation into leukaemia biology such as stem/progenitor cell-niche interactions, stem/progenitor cell proliferation and treatment resistance. 3Rs benefit: To replace moderate severity animal (mice) procedures in leukaemia research and early pre-clinical drug testing. Current applications: Exploration of leukaemia biology such as stem/progenitor cell proliferation, stem/progenitor cell -niche interactions, niche-impacted treatment resistance and obtain drug response data. Response We have replaced “blasts” with “stem/progenitor” as appropriate. Introduction: Need to define at the start that you are talking about ALL as it reads like a general overview which means some of the % incidence rates etc. are not correct. It would be good to mention what type of ALL as well, is it B-ALL, T-ALL, childhood, adult? Response This has been clarified as below: Using childhood acute lymphoblastic leukaemia (ALL) as a proof-of-concept paradigm, here we develop an in vitro platform to establish co-cultures of patient-derived leukaemia cells with 3D bone marrow mesenchyme spheroids, BM-MSC-spheroids. Paragraph 3 page 3: Most research groups would use primary patient samples in vitro prior to moving to in vivo models. This should be included. Response This has been included as below: Although several research groups would likely use primary patient for in vitro studies before moving into in vivo models, in vivo models are regarded as gold standard preclinical models, and are used as key models to expand primary samples as vital research resources. Paragraph 2 Page 4: Remove blasts and replace with LSCs also include disease type chronic myeloid leukaemia. Response We apologise for using “blasts”, all of these have been replaced with “leukaemia cell” or “ALL”. Disease type chronic myeloid leukaemia has been added to the following sentence Cell lines retain cytogenic characteristics of acute lymphoblastic leukaemia (ALL), and other diseases like chronic myeloid leukaemia, but being derived from patients with relapsed disease these samples do not represent the molecular complexity of disease at presentation14,15. Page 11 ‘such spheroid biobanks’: do they actually exist? Or are you planning on constructing a biobank? Response We apologise for the ambiguity and have now re-worded the section below: Following further extensive validation, such 3D-based preclinical models would furthermore have the potential to be embedded within clinical trials and healthcare systems for the purposes of detecting responders as well as to aid risk stratification. ‘MSC spheroids provide superior protection to leukaemia cells against dexamethasone treatment. are appropriate in modelling treatment resistance which remains a major clinical challenge in cancer management’. Is this statement correct, not really looking at resistance, just reduced sensitivity to the drugs at that dose when in 3D rather than 2D culture. Need to amend and extend to include how you would look at resistance i.e. relapsed patients and the potential to interrogate resistance mechanisms. Response This statement has now been clarified: This means that compared to 2D MSC cultures, MSC spheroid-based models, might be promising to study the biology of leukaemia cells from relapsed disease, consequently helping interrogate treatment resistance mechanisms, a major clinical challenge in cancer management." } ] } ]	1	https://f1000research.com/articles/11-1280
https://f1000research.com/articles/10-991/v1	30 Sep 21	{ "type": "Method Article", "title": "Using multiple outcomes in intervention studies for improved trade-off between power and type I errors: the Adjust NVar approach", "authors": [ "Dorothy V. M. Bishop" ], "abstract": "Background The CONSORT guidelines for clinical trials recommend use of a single primary outcome, to guard against the raised risk of false positive findings when multiple measures are considered. It is, however, possible to include a suite of multiple outcomes in an intervention study, while controlling the familywise error rate, if the criterion for rejecting the null hypothesis specifies that N or more of the outcomes reach an agreed level of statistical significance, where N depends on the total number of outcome measures included in the study, and the correlation between them. Methods Simulations were run, using a conventional null-hypothesis significance testing approach with alpha set at .05, to explore the case when between 2 and 12 outcome measures are included to compare two groups, with average correlation between measures ranging from zero to .8, and true effect size ranging from 0 to .7. In step 1, a table is created giving the minimum N significant outcomes (MinNSig) that is required for a given set of outcome measures to control the familywise error rate at 5%. In step 2, data are simulated using MinNSig values for each set of correlated outcomes and the resulting proportion of significant results is computed for different sample sizes,correlations, and effect sizes. Results The Adjust NVar approach can achieve a more efficient trade-off between power and type I error rate than use of a single outcome when there are three or more moderately intercorrelated outcome variables. Conclusions Where it is feasible to have a suite of moderately correlated outcome measures, then this might be a more efficient approach than reliance on a single primary outcome measure in an intervention study. In effect, it builds in an internal replication to the study. This approach can also be used to evaluate published intervention studies.", "keywords": [ "intervention", "methodology", "statistics", "correlated outcomes", "power", "familywise error rate", "multiple comparisons" ], "content": "The case against multiple outcomes\n\nThe CONSORT guidelines for clinical trials (Moher et al. 2010) are very clear on the importance of having a single primary outcome:\n\nAll RCTs assess response variables, or outcomes (end points), for which the groups are compared. Most trials have several outcomes, some of which are of more interest than others. The primary outcome measure is the pre-specified outcome considered to be of greatest importance to relevant stakeholders (such a patients, policy makers, clinicians, funders) and is usually the one used in the sample size calculation. Some trials may have more than one primary outcome. Having several primary outcomes, however, incurs the problems of interpretation associated with multiplicity of analyses and is not recommended.\n\nThis advice often creates a dilemma for the researcher: in many situations there are multiple measures that could plausibly be used to index the outcome. A common solution is to apply a Bonferroni correction to the alpha level used to test significance of individual measures, but this is over-conservative if, as is usually the case, the different outcomes are intercorrelated. Alternative methods are to adopt some process of data reduction, such as extracting a principal component from the measures that can be used as the primary outcome, or using a permutation test to derive exact probability of an observed pattern of results. Here I explore a further, very simple, option which I term the “Adjust NVar” approach. The idea is that if one has a suite of outcomes, instead of adjusting the alpha level, one can adjust the number of outcomes that are required to achieve significance at the conventional alpha level of .05 to maintain an overall familywise error rate of 1 in 20 or less.\n\nTo illustrate the idea with a realistic example, suppose we are reading a report of a behavioural intervention that is designed to improve language and literacy, and there are 6 measures where we might plausibly expect to see some benefit. The researchers report that none of the outcomes achieve the Bonferroni-adjusted significance criterion of p < .008, but two of them reach significance at p < .05. Should we dismiss the trial as showing no benefit? We can use the binomial theorem to check the probability of obtaining this result if the null hypothesis is true and the measures are independent: it is 0.033, clearly below the 5% alpha level. But what if the measures are intercorrelated? That is often the case: indeed, it would be very unusual for a set of outcome measures to be independent. A thought experiment helps here. Suppose we had six measures that were intercorrelated at .95 - in effect they would all be measures of the same thing, and so if there was a real effect, most of the measures should show it. Extending this logic in a more graded way, the higher the correlation between the measures, the more measures would need to reach the original significance criterion to maintain the overall significance level below .05.\n\nA simulation script was developed to test these intuitions and to obtain estimates of:\n\ni) the minimum number of outcome variables in a suite that would maintain the overall familywise error rate at 1 in 20, if each individual measure was evaluated at the significance criterion of .05. This we term MinNSig.\n\nii) the power to detect a true effect, if the criterion for rejection the null hypothesis was based on the value of MinNSig identified at step A.\n\n\nMethods\n\nCorrelated variables were simulated using in the R programming language (R Core Team 2020) (R Project for Statistical Computing, RRID:SCR_001905). The script to generate and analyse simulated data is available on https://github.com/oscci/MinSigVar. Initially, two approaches to modeling correlated variables were compared, but differences between them proved to be trivial, and so only one is reported here.\n\nThe mvrnorm function of the MASS package (Modern Applied Statistics with S, RRID:SCR_019125) was used to generate a set of 12 outcome variables with a specified covariance matrix. For simplicity, all variables were simulated as random normal deviates with SD of 1, and the covariance matrix had a prespecified correlation, r, in all off-diagonal elements. The correlation varied across runs from 0 to .8 in steps of .2, and the number of simulated cases varied from 20 to 110 in steps of 30. Outcomes for Intervention (I) and Control (C) groups differed only in terms of the mean, which was always zero for the group C, and a given effect size, e, for group I. The average observed effect size for all measures in a given condition was computed and used as the basis for comparisons of efficiency between single and multiple measure scenarios.\n\nThis method is simple but can lead to unrealistic data: in particular it is possible to have a set of outcomes that are independent of one another (r = 0) yet all having the same effect size. In real-world data, one would expect outcomes to be correlated, especially those that all showed an impact of intervention. Conversely, if a set of outcomes was very highly intercorrelated, then we would expect them all to show a similar intervention effect.\n\nAn alternative approach was evaluated to consider such cases, in which the set of 12 outcome measures are simulated as indicators of an underlying latent variable, which mediates the intervention effect. This can be achieved by first simulating a latent variable, with an effect size of either zero, for group C, or e for group I. Observed outcome measures are then simulated as having a specific correlation with the latent variable - i.e. the correlation determines the extent to which the outcomes act as indicators of the latent variable. This can be achieved using the formula:\n\nwhere r is the correlation between latent variable (L) and each outcome, and L is a vector of random normal deviates that is the same for each outcome variable, while E (error) is a vector of random normal deviates that differs for each outcome variable. Note that when outcome variables are generated this way, the mean intercorrelation between them will be r2. Thus if we want a set of outcome variables with mean intercorrelation of .4, we need to specify r in the formula above as sqrt(r) = .632. Furthermore, the effect size for the simulated variables will be lower than for the latent variable: to achieve an effect size, e, for the outcome variables, it is necessary to specify the effect size for the latent variable, el, as e/r2. It was found that when this is done, the results with this method were closely similar to those obtained using MASS, for the range of correlations and effect sizes considered here. The exception is for the case where r = 0, which is not computuable with this method - i.e. it is not possible to have a set of outcomes that are indicators of the same latent factor but which are uncorrelated. As noted above, the case where r = 0 is unrealistic in any case, and so for the simulations reported here, the lowest value of r that was included was r = .2.\n\nThe size of the suite of outcome variables entered into later analysis ranged from 2 to 12. For each suite size, principal components were computed from data from the C and I groups combined, using the base R function prcomp from the stats package (R Core Team, 2020). Thus, PC2 is a principal component based on the first two outcome measures, PC4 based on the first four outcome measures, and so on.\n\nPower of analyses based on the principal components was compared with power obtained using the Adjust NVar approach, as specified below.\n\n10,000 simulations were run for each combination of:\n\n- sample size per group, ranging from 20 to 110 in steps of 30\n\n- correlation between outcome variables, ranging from .2 to .8 in steps of .2\n\n- true effect size, taking values of 0, .3, .5, or .7.\n\nThe data generated from each combination of conditions was used to derive results for different sizes of suites of outcome variables, ranging from 2 to 12. Thus, the analysis was first conducted on the first 2 outcome measures, then on the first 3 outcome measures, and so on.\n\nFor each set of conditions, on each run, a one-tailed t-test was conducted to obtain a p-value for the comparison between C and I groups, assuming C would be lower. The p-values for outcome measures were rank ordered for each run and each suite size.\n\nTo obtain MinNSig, the results were filtered to include only the runs where the null hypothesis was true, i.e. effect size = 0. Then, the proportion of p-values less than .05 was calculated for each rank for each number of outcome variables, to find the highest rank at which the overall proportion was less than .05. This is the MinNSig.\n\nTable 1 gives a toy example of the logic, using the case where we have either 2 or 4 outcome measures. Columns V1 to V4 show p-values for the t-test comparing the two groups each of the 4 outcome measures. Columns r2.1 and r2.2 show the same p-values rank ordered for just the first two measures; columns r4.1 to r4.4 show the p-values rank ordered for all 4 outcomes. We can then count the number of p-values that are below .05 for all runs (1000 in this case) for each ranked position. With 2 outcomes, if we take just the first ranked (lowest) p-value, the proportion lower than .05 is around .10. For the 2nd ranked p-value, the proportion drops below .05, to .002. Thus, we set MinNSig to 2.\n\nV1 to V4 are p-values from one-sided t-test, one row for each run of simulation in a given condition. Columns with prefix r2 or r4 show the same p-values rank ordered for either the first two columns, or the first four columns. The final two rows show the number and the proportion of values falling below .05 for that column.\n\nWe can then turn to the case where we have four outcomes: the proportion of the 1st ranked p-values below .05 is .185; the proportion of the second ranked below .05 is .014. Thus again, we set MinNSig to 2. As noted above, when the correlation between variables is zero, we can use the binomial theorem to compute values in the final row; however, when variables are intercorrelated, more p-values will be below .05, and so MinNSig may be higher.\n\nBecause MinNSig moves in quantum steps, the effective familywise error rate is often lower than .05. For instance, in the example above with a suite of four outcome measures, MinNSig is set to 2, but this gives p = .014, rather than .05.\n\nFor each run of the simulation, and each number of outcome measures, we take the value of MinNSig from the previous step and compute the proportion of p-values below .05, depending on the effect size, sample size and correlation between measures. For effect sizes above zero, this proportion corresponds to the statistical power.\n\nPower using Adjust NVar can be compared to:\n\n- power obtained with a single outcome measure for the same effect size and sample size\n\n- power obtained by using the principal component extracted for this set of outcome measures\n\n\nResults\n\nTable 2 shows results from a simulation of the Adjust NVar approach, with the values in the body of the table showing MinNSig, the minimum number of measures that would maintain the overall familywise error rate at 1 in 20, if each individual measure was evaluated at the significance criterion of .05. Because the t-test statistic used to determine p-values is adjusted for sample size, these values are independent of numbers of subjects. In principle, researchers could use Table 2 to specify in their research protocol the minimum number of outcomes that would reach their significance level in order for the null hypothesis to be rejected.\n\nEntries in body of table show smallest N variables reaching p < .05 that preserve familywise error rate at .05 or less. N prefix denotes suite size for a set of outcomes. Corr indicates correlation between outcomes.\n\nFull tables of results for all combinations of parameters are provided in Extended Data. Figures 1 to 3 plot power vs familywise error rate for different sizes of suite of outcome measures.\n\nSymbols denote sample size per group, and colours denote correlation between outcomes (see Key). Vertical dotted lines show power for single outcome at different sample sizes. Horizontal line shows type I error rate of .05.\n\nSymbols denote sample size per group, and colours denote correlation between outcomes (see Key). Vertical dotted lines show power for single outcome at different sample sizes. Horizontal line shows type I error rate of .05.\n\nSymbols denote sample size per group, and colours denote correlation between outcomes (see Key). Vertical dotted lines show power for single outcome at different sample sizes. Horizontal line shows type I error rate of .05.\n\nFor these plots, we see power for small, medium and large effect sizes (corresponding to Cohen’s d of .3, .5 and .7). An efficient method is one that gives power of .8 or above, and a familywise error rate of .05 or less, i.e. the results should cluster in the bottom right quadrant. Power, which depends on sample size, is shown for a study with a single outcome in the vertical dotted lines, with an alpha of .05 shown in the horizontal dotted line. We can compare by eye how well Adjust Nmin with multiple outcomes compares with a single outcome for the same sample size. With just two outcome measures we obtain a very low familywise error rate but power is generally worse than for the single outcome case, except when the effect size is large. This is because when using Adjust NMin with two outcomes, both outcomes have to achieve p < .05. With three outcomes, again Adjust NMin requires we have at least two individual outcomes with p < .05: this gives power equivalent to that of a single variable, but a lower familywise error rate is achieved. When the number of outcomes is four or more, the benefit of Adjust NMin over a single outcome becomes more evident, with higher power coupled with a lower familywise error rate. The specific results depend also on the intercorrelation between outcomes (which in turn influences the MinNSig value, see Table 2): a moderate level of intercorrelation (between .4 and .6) generally gives an efficient measure.\n\nFigures 4 to 6 give equivalent plots for power from principal components.\n\nSymbols denote sample size per group, and colours denote correlation between outcomes (see Key). Vertical dotted lines show power for single outcome at different sample sizes. Horizontal line shows type I error rate of .05.\n\nSymbols denote sample size per group, and colours denote correlation between outcomes (see Key). Vertical dotted lines show power for single outcome at different sample sizes. Horizontal line shows type I error rate of .05.\n\nSymbols denote sample size per group, and colours denote correlation between outcomes (see Key). Vertical dotted lines show power for single outcome at different sample sizes. Horizontal line shows type I error rate of .05.\n\nThe Principal Components plots show all points are to the right of the vertical line denoting power from a single outcome, i.e. this method achieves higher power than a single outcome measure for each size of suite of outcomes. Familywise error rate clusters around .05. If we compare how Adjust NVar compares with Principal Components, with three or more outcomes the power is generally slightly lower, but the tradeoff between power and familywise error (expressed as a ratio) is higher for Adjust NVar.\n\n\nDiscussion\n\nThe logic of conventional multiple testing is turned on its head with the Adjust NVar approach, in that instead of adjusting the p-value used for significance (as in the Bonferroni correction, or methods based on False Discovery Rate), we adjust the number of individual outcome measures that we need to reach the intended significance criterion. This value can be easily computed using the binomial theorem for a given suite size of outcomes if the measures are uncorrelated, but in the context of intervention trials uncorrelated measures is an unrealistic assumption.\n\nOne advantage of this approach is that it is more compatible with trials of interventions that are expected to affect a range of related processes, as is common in some fields such as education or speech and language therapy. In such cases, the need to specify a single primary outcome tends to create difficulties, because it is often unclear which of a suite of outcomes is likely to show an effect. Note that the Adjust NVar approach does not give the researcher free rein to engage in p-hacking: the larger the suite of measures included in the study, the higher the value of MinNSig will be. It does, however, remove the need to put all one’s eggs in one basket by pre-specifying one measure as the primary outcome.\n\nA second advantage is that in effect, by including multiple outcome measures, one can improve the efficiency of a study, in terms of the trade-off between power and familywise errors. A set of outcome measures may be regarded as imperfect proxy indicators of an underlying latent construct, so we are in effect building in a degree of within-study replication if we require that more than one measure shows the same effect in the same direction before we reject the null hypothesis.\n\nThe comparison with power and familywise error rate from principal components shows that the latter approach is more consistent in improving power over a study with a single outcome than the Adjust NVar approach, regardless of the size of the suite of outcomes, but it does not influence familywise error rate. Variation in familywise error rate is a consequence of the quantum nature of the adjustment with Adjust NVar, where the same value of MinNVar may be used with varying sizes of outcome suite, which can lead to values of familywise error rate well below .05. For instance, obtaining two p-values below .05 in a suite of two outcomes is a more unusual circumstance than obtaining two values this extreme in a suite of three or four outcomes. Nevertheless, the ratio of power to familywise error is generally higher for Adjust NVar than for principal components.\n\nA possible disadvantage of Adjust NVar over principal components is that this approach is likely to tempt researchers to interpret specific outcomes that fall below the .05 threshold as meaningful. They may be, of course, but this simulation demonstrates that when we create a suite of outcomes that differ only by chance, it is common for only a subset of them to reach the significance criterion. Any recommendation to use Adjust NVar should be accompanied by a warning that a suite of outcomes should be selected as representative of the underlying construct the intervention is designed to influence, in effect serving as replicate measures, all of which should be equally promising as indicators of an intervention effect. If a subset of outcomes show an effect of intervention, this could be due to chance. It would be necessary to run a replication to have confidence in a particular pattern of results.\n\nIt is also worth noting that results obtained with this approach depend crucially on assumptions embodied in the simulation that is used to derive predictions. Outcome measures simulated here are normally distributed, and uniform in their covariance structure. It would be possible to generate datasets with different underlying covariance structures to be tested in the same way, but that is beyond the scope of this paper.\n\nPerhaps the main advantage of this approach is that once the values of MinNSig have been specified (as in Table 2), the method is very simple to apply, and could be used in two ways. First, it can be used a priori to specify in a protocol the number of outcomes that would need to achieve the conventional .05 level of significance, in order for the intervention to be deemed effective. This assumes that the researcher already has a rough idea of the degree of intercorrelation between outcome measures, but a range somewhere between .4 and .6 is a reasonable assumption for many behavioural studies. Pre-registering a specific level of MinSigN would help guard against a tendency to explore different kinds of correction for multiple hypothesis testing only after viewing the data (Lazic 2021).\n\nSecond, the simplicity of the approach makes it useful for evaluating published studies that report multiple outcomes but may not have been analysed optimally. We started with the example of a study where there were six outcome measures, none of which met the Bonferroni-corrected significance level of .05/6 = .008, but two of which met p < .05. From Table 2 we can see that to be confident in a true intervention effect, assuming correlated outcomes, then at least three out of six outcomes need to be significant at the .05 level. In this case, therefore, we do not reject the null hypothesis.\n\nIn sum, the Adjust NVar method shows how inclusion of multiple outcomes can be a positive strategy in intervention studies and can give stronger statistical evidence than a single outcome, provided that attention is paid to the need for several outcomes to reach a significance threshold.\n\n\nData availability statement\n\nOSF: Adjust NVar. https://doi.org/10.17605/OSF.IO/5T4SE. (Bishop, 2021).\n\nThis project contains the following extended data.\n\n• Extended data.docx (Word version of powertab_methodM.csv, showing power for each combination of parameters, with separate subtables for individual variables (Figures 1-3) and principal components (Figures 4-6).\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nSoftware availability\n\nThe script to generate and analyse simulated data is available on https://github.com/oscci/MinSigVar.\n\nThe OSF project contains the following files.\n\n• toybit.csv - corresponds to Table 1\n\n• MinNSig_methodM.csv - corresponds to Table 2\n\n• p_1sided_methodM_allN_allES_allcorr_maxn12_nsim10000_nstep1.csv - output of simulation of 10000 runs of Multiple_outcomes.Rmd script\n\n• Extended data.docx - word version of powertab_methodM.csv\n\n• powertab_methodM.csv - csv version of Extended Data table, with individual variables in columns N1-N12, and principal components in columns PC2-PC12", "appendix": "References\n\nBishop DVM: Adjust NVar.2021, September 24. Reference Source\n\nLazic SE: Why Multiple Hypothesis Test Corrections Provide Poor Control of False Positives in the Real World. arXiv:2108.04752 [q-Bio, Stat] . 2021; (August). Reference Source\n\nMoher D, Hopewell S, Schulz KF, et al.: CONSORT 2010 Explanation and Elaboration: Updated Guidelines for Reporting Parallel Group Randomised Trials. BMJ (Clinical Research Ed.) . 2010; 340(March): c869. Publisher Full Text\n\nR Core Team: R: A Language and Environment for Statistical Computing . Vienna, Austria: R Foundation for Statistical Computing; 2020. Reference Source\n\nVenables WN, Ripley BD: Modern Applied Statistics with S . Fourth ed. New York: Springer; 2002.0-387-95457-0." }	[ { "id": "96192", "date": "12 Oct 2021", "name": "Kristin Sainani", "expertise": [ "Reviewer Expertise Statistics", "Sports Medicine" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe paper presents a method for controlling the familywise error rate when testing multiple outcomes: Adjust NVar. Unlike multiple testing adjustments that lower the p-value threshold, the idea behind Adjust NVar is to require a minimum number of p-values (MinNSig) to meet a nominal p<.05 threshold.\nWhen outcomes are independent, MinNSig is defined as follows, where M is the number of outcomes: X~binomial(M, 0.05) MinNSig is the minimum x for which P(X>=x)<.05 or, equivalently, P(X < x)>.95. For example, if M=6, then: P(X=0)=.95*6=0.735 P(X<=1)=P(X<2)=0.735+6.95*5.05=.232+.735=.967 Thus, MinNSig=2.\nWhen outcomes are correlated, there is no simple formula for obtaining MinNSig, so the author has used a simulation approach to account for varying correlation structures.\nThe idea is novel and has several merits. In particular, the approach closely matches what many researchers already do in the published literature. Many published studies apply a p-value cutoff of 0.05 to multiple outcomes without correcting for multiple testing (or designating a primary outcome). I can envision the AdjustNVar approach being a useful heuristic for re-evaluating published studies that used multiple outcomes but failed to account for multiple testing in any way. For example, if an RCT reported 10 outcomes with only 1 significant result (p<.05), readers can easily recognize that this is compatible with a chance finding. But what if the trial found 2 significant results or 3? And what if the outcomes are moderately correlated instead of independent? I can envision researchers using a table such as Table 2 of the AdjustNVar paper to make a quick assessment based on the number of outcomes reported and a rough guess at the correlation structure.\nHowever, I am less convinced of the value of AdjustNVar as a formal tool for controlling the familywise error rate in a planned study. At a minimum, further development and a broader set of simulations would be required to support such a recommendation. The current manuscript describes three alternatives to specifying a single primary outcome in an RCT: (1) Bonferroni adjustment, (2) permutation tests, and (3) use of PCA to derive a single composite outcome. But this ignores existing p-value adjustment methods that are less conservative than Bonferroni. For example, the “M-effective” (Meff) approach accomplishes many of the same goals as AdjustNVar (see: Cheverud (2001)1, Nyholt (2004)2, and Derringer (2018)3). In the Meff approach, one adjusts the p-value threshold by dividing by the effective number of outcomes (Meff) rather than the actual number of outcomes (M). Meff is based on the eigenvalues of the correlation matrix of the outcomes. Where Eigen is the observed vector of eigenvalues from the correlation matrix of the outcomes, Meff is calculated as:\nMeff = 1 + (M-1)(1-(Var(Eigen)/M))\nBonferroni threshold = alpha/M Meff threshold = alpha/Meff\nLike AdjustNVar, Meff is simple and accounts for correlated outcomes. But I believe it has several advantages over AdjustNVar: (1) Meff precisely controls the Type I error rate, whereas AdjustNVar has varying Type I error rates that cannot be precisely controlled by the investigator; (2) Meff accounts for the correlation structure observed in the data, whereas AdjustNVar requires the investigator to guess at the correlation structure; if this guess is far off (which could easily be the case), this would lead to poor Type I error control.\nThis paper has numerous strengths, including the novelty of the idea; the potential use as a heuristic for re-interpreting flawed published papers; the concision of the writing; and the availability of all code and data. The major limitations of the paper are: (1) it presents an overly narrow set of simulations that do not capture most realistic situations, but then makes overly broad claims based on these simulations. (2) it does not compare AdjustNVar to existing approaches that are less conservative than Bonferroni, such as Meff and (3) it does not address the different reasons why researchers may be including multiple outcomes, but these different reasons lead to markedly different correlation structures.\nSpecific comments:\nThe paper would benefit from further consideration of the reasons why researchers may include multiple outcomes. The paper focuses on the case where an intervention is “expected to affect a range of related processes.” The simulations make assumptions that match this case, assuming equal correlations across outcomes and equal true effects for each outcome. But researchers may include multiple outcomes for many other reasons, such as: (a) they aren’t sure which process the intervention will affect, (b) they believe the intervention may affect two different processes but they measure each process with several different measurements to “hedge their bets”, or (c) they include a “soft” endpoint in addition to a “hard” endpoint because the “hard” endpoint may occur too rarely. Each of these cases corresponds to different assumptions for the simulations. For example, (b) would be expected to have two clusters of highly correlated variables that are only weakly correlated with each, which will affect MinNSig.\n\nThe paper suggests that AdjustNVar could be used in study planning—researchers would guess at the correlation structure and set a MinNSig ahead of time. But if they guess the correlation structure incorrectly, such as underestimating the true correlation, then they may choose a MinNSig that does not adequately control Type I error.\n\nThe “quantum nature” of AdjustNVar is not a desirable characteristic. The researcher is unable to precisely control the Type I error rate. In planning a study in which the correlation is expected to be 0.4, for example, Table 2 would suggest that the researcher should then always choose 9 outcomes over 5-8 outcomes, since 9 maximizes the chances of getting at least 3 p-values <.05. This is one reason I prefer Meff, which precisely controls the Type I error rate.\n\nThis description is misleading: “Should we dismiss the trial as showing no benefit? We can use the binomial theorem to check the probability of obtaining this result if the null hypothesis is true and the measures are independent: it is 0.033, clearly below the 5% alpha level.” The description gives the misleading impression that one would be justified in re-evaluating a paper that used a Bonferroni correction by instead applying the criterion of at least two p-values <.05. But doing so would inflate the Type I error rate. Results would have been declared significant if EITHER at least one p-value met the Bonferroni threshold OR at least two p-values were <.05 — leading to an effective Type I error rate of 7% (assuming independent outcomes). Note that this example reappears in the discussion and also mistakenly implies that had three p-values been <.05, we would have been able to reject the null hypothesis. But this is not the case because the results were already subjected to Bonferroni, and additionally subjecting them to AdjustNVar makes the effective Type I error rate higher than 5%. AdjustNVar should be applied only to re-evaluate studies that failed to incorporate any adjustments for multiple testing originally.\n\nI found Table 1 confusing on first read, and I would recommend simplifying it by focusing on a single number of outcomes rather than both 2 outcomes and 4 outcomes and by removing discussions of ranking p-values. (The p-value ranking isn’t important — this is just part of the mechanics of how the algorithm is calculating MinNSig, so I don’t think it’s needed.) For example, you could just focus on calculating MinNSig for 6 outcomes. Show 6 columns of p-values for the 6 outcomes, and then show a single final column that tabulates the number of p-values <.05 for each simulated trial. Then show a frequency table of how many simulations out of 1000 resulted in 0 p-values <.05, 1 p-value <.05, 2 p-values <.05, etc. Then indicate that MinNSig occurs at one number above when the cumulative frequency crosses 95%.\n\nI’m unclear as to why the paper focuses on one-tailed tests, which are less common in the literature. I think it would be more useful to present two-tailed tests in Table 2 or to present two tables — one for one-tailed tests and one for two-tailed tests. This makes a difference in a few MinNSig values.\n\nFigures 1-3: These figures compare AdjustNVar to a single study outcome. I think the logic behind this comparison is flawed, however. It is comparing apples to oranges. The simulation assumes that, when applying AdjustNVar, ALL variables studied have a true effect. This, in effect, stacks the deck on statistical power for any* method that considers multiple outcomes rather than a single outcome. For example, I ran a simulation comparing Bonferroni with 6 outcomes compared to a single outcome with n=50 per group. When the correlation is <0.8, Bonferroni also has more power than the single outcome. And, when I compared the Meff strategy to a single outcome in a variety of scenarios, Meff was always more powerful than a single outcome. I think a more useful comparison would be to directly compare different methods that handle multiple outcomes (e.g., PC to AdjustNVar to Meff).\n\nRelated to comment (7), I don’t think the paper is justified in making this broad claim: “The Adjust NVar approach can achieve a more efficient trade-off between power and type I error rate than use of a single outcome when there are three or more moderately intercorrelated outcome variables.” This conclusion is true only when the intervention truly affects ALL outcomes, which is a narrow and arguably unrealistic case. A more realistic scenario is where the intervention works only on a subset of outcomes. In this case, the single variable strategy will be more statistically powerful than the multiple-variable strategies if you choose the right variable.\n\nFigures 4-6. Same issue as for Figures 1-3: comparing the principal components composite variable strategy (PC) to a single outcome is flawed because the simulation “stacks the deck” for PC by assuming that all outcomes have a true effect. I believe that Figures 1-6 should focus instead on comparisons of different methods for handling multiple outcomes.\n\nThe article claims that power is only “slightly lower” for AdjustNVar compared with the PC strategy. However, when I run simulations comparing PC to AdjustNVar, I get consistently higher statistical power for PC and I would characterize the difference as more than just “slight”. For example, with N=50, global corr=0.6, global ES=0.3, and 6 outcomes (two-tailed test), I get power of 35% for AdjustNVar versus 37% for Meff versus 46% for PC.\n\nPC is always more powerful than AdjustNVar and Meff when we assume that all outcomes have a true effect. However, PC is not more powerful when we assume that only a subset of outcomes have true effects. For example, if I tweak the above simulation so that only three outcomes out of six have true effects of 0.3, this changes the power to 15% for PC and AdjustNVar, and 28% for Meff. This illustrates why one narrow simulation is insufficient for making general conclusions about the tradeoffs in performance between the different methods.\n\nIn my simulations, Meff consistently has higher statistical power than AdjustNVar, which is a function of the fact that Meff always has a 5% Type I error rate whereas that of AdjustNVar is variable and sometimes lower than 5%. I don’t view it as a strength that AdjustNVar results in arbitrarily lower Type I error rates. It is better for the investigator to be able to precisely control the tradeoff between Type I and Type II error. Meff allows this whereas AdjustNVar does not.\n\nFigures 1-6: I found these graphs hard to read as they don’t have a clear take-home point. I would suggest removing the single outcome comparisons altogether and then reformatting so that the graphs directly compare AdjustNVar to Meff and PC (three lines). In one graph, hold effect size, sample size, and N outcomes constant, and then show the power of the three methods as a function of increasing correlation. In another graph, hold correlation, sample size, and effect size constant, and show the power of the three methods as a function of N outcomes. And so on. All methods aim to control the familywise error rate at 5%, so this should not be a variable. The fact that AdjustNVar sometimes results in a lower Type I error rate is incidental — it arises as a quirk of the method not as the intent of the researcher.\n\nWhere AdjustNVar may have an advantage over a method like Meff is for re-evaluating flawed published studies where researchers used multiple outcomes but did not account in any way for multiple testing. An AdjustNVar table such as Table 2 would give readers a quick way to reevaluate such studies without the need for any calculations or access to raw data. I would focus the paper more on this application.\n\nIs the rationale for developing the new method (or application) clearly explained? Partly\n\nIs the description of the method technically sound? Partly\n\nAre sufficient details provided to allow replication of the method development and its use by others? Yes\n\nIf any results are presented, are all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions about the method and its performance adequately supported by the findings presented in the article? Partly", "responses": [ { "c_id": "9034", "date": "13 Jan 2023", "name": "Dorothy Bishop", "role": "Author Response", "response": "Thanks for the very useful evaluation of this paper, which has prompted further thoughts and a revised manuscript. There was some convergence of views of the two reviewers, although the specific recommendations varied. I thank the reviewer for engaging so thoroughly with the manuscript and for helping improve it. General point A: AdjustNVar as a heuristic. Reviewer 1 writes: The idea is novel and has several merits. In particular, the approach closely matches what many researchers already do in the published literature. Many published studies apply a p-value cutoff of 0.05 to multiple outcomes without correcting for multiple testing (or designating a primary outcome). I can envision the AdjustNVar approach being a useful heuristic for re-evaluating published studies that used multiple outcomes but failed to account for multiple testing in any way. For example, if an RCT reported 10 outcomes with only 1 significant result (p<.05), readers can easily recognize that this is compatible with a chance finding. But what if the trial found 2 significant results or 3? And what if the outcomes are moderately correlated instead of independent? I can envision researchers using a table such as Table 2 of the AdjustNVar paper to make a quick assessment based on the number of outcomes reported and a rough guess at the correlation structure. Response: After exploring Meff, I decided not to proceed with the AdjustNVar approach, as I think a lookup table derived from MEff would achieve the same effect but without the problems arising from the quantal nature of N variables. General Point B: Need to consider MEff Reviewer 1 notes that the MEff approach accomplishes many of the same goals as AdjustNVar, and is preferable in many respects. Response: I had been unaware of MEff, and having consulted the references, I agree it is an elegant solution to the problem I was trying to tackle that avoids some of the limitations of AdjustNvar, particularly the need to assume a given correlation structure. I initially thought this approach had only been used in genetics and not been applied in psychology, but the final reference pointed to the work of Derringer, who has done a great job in a preprint that provides a tutorial in its use. I don’t think it is much used in intervention research, which is the context I am particularly interested in, and so I think it is worthwhile updating the article so that it serves as a basic introduction to MEff, with discussion of factors affecting power. Accordingly, I have changed the focus to compare different methods with a focus on MEff. Specific comments: 1. The paper would benefit from further consideration of the reasons why researchers may include multiple outcomes. The paper focuses on the case where an intervention is “expected to affect a range of related processes.” The simulations make assumptions that match this case, assuming equal correlations across outcomes and equal true effects for each outcome. But researchers may include multiple outcomes for many other reasons, such as: (a) they aren’t sure which process the intervention will affect, (b) they believe the intervention may affect two different processes but they measure each process with several different measurements to “hedge their bets”, or (c) they include a “soft” endpoint in addition to a “hard” endpoint because the “hard” endpoint may occur too rarely. Each of these cases corresponds to different assumptions for the simulations. For example, (b) would be expected to have two clusters of highly correlated variables that are only weakly correlated with each, which will affect MinNSig. Response: This really got me thinking and I have now incorporated some further simulations where correlations are not uniform, as also added more discussion of this issue 2. The paper suggests that AdjustNVar could be used in study planning—researchers would guess at the correlation structure and set a MinNSig ahead of time. But if they guess the correlation structure incorrectly, such as underestimating the true correlation, then they may choose a MinNSig that does not adequately control Type I error. Response: Agreed. This is now dropped. 3. The “quantum nature” of AdjustNVar is not a desirable characteristic. The researcher is unable to precisely control the Type I error rate. In planning a study in which the correlation is expected to be 0.4, for example, Table 2 would suggest that the researcher should then always choose 9 outcomes over 5-8 outcomes, since 9 maximizes the chances of getting at least 3 p-values <.05. This is one reason I prefer Meff, which precisely controls the Type I error rate. Response: Agreed. AdjustNVar now dropped 4. This description is misleading: “Should we dismiss the trial as showing no benefit? We can use the binomial theorem to check the probability of obtaining this result if the null hypothesis is true and the measures are independent: it is 0.033, clearly below the 5% alpha level.” The description gives the misleading impression that one would be justified in reevaluating a paper that used a Bonferroni correction by instead applying the criterion of at least two p-values <.05. But doing so would inflate the Type I error rate. Results would have been declared significant if EITHER at least one p-value met the Bonferroni threshold OR at least two p-values were <.05 — leading to an effective Type I error rate of 7% (assuming independent outcomes). Note that this example reappears in the discussion and also mistakenly implies that had three p-values been <.05, we would have been able to reject the null hypothesis. But this is not the case because the results were already subjected to Bonferroni, and additionally subjecting them to AdjustNVar makes the effective Type I error rate higher than 5%. AdjustNVar should be applied only to re-evaluate studies that failed to incorporate any adjustments for multiple testing originally. Response: thanks for this clarification. As AdjustNVar is now omitted, this no longer applies. 5. I found Table 1 confusing on first read, and I would recommend simplifying it by focusing on a single number of outcomes rather than both 2 outcomes and 4 outcomes and by removing discussions of ranking p-values. (The p-value ranking isn’t important — this is just part of the mechanics of how the algorithm is calculating MinNSig, so I don’t think it’s needed.) For example, you could just focus on calculating MinNSig for 6 outcomes. Show 6 columns of p-values for the 6 outcomes, and then show a single final column that tabulates the number of p-values <.05 for each simulated trial. Then show a frequency table of how many simulations out of 1000 resulted in 0 p-values <.05, 1 p-value <.05, 2 p-values <.05, etc. Then indicate that MinNSig occurs at one number above when the cumulative frequency crosses 95%. Response: this no longer applies as tables are redone 6. I’m unclear as to why the paper focuses on one-tailed tests, which are less common in the literature. I think it would be more useful to present two-tailed tests in Table 2 or to present two tables — one for one-tailed tests and one for two-tailed tests. This makes a difference in a few MinNSig values. Response: One-tailed tests have a bad reputation because so often they are misused to just require a lower level of significance when there are no directional predictions, but there are contexts in which they are entirely appropriate, and that includes the kinds of intervention study that is the focus of attention here. You can reasonably predict that an intervention will improve performance rather than worsen it. Reviewer 2 wrote a blogpost about this which I find convincing: http://daniellakens.blogspot.com/2016/03/one-sided-tests-efficient-and-underused.html. I have now explained this further. 7. Figures 1-3: These figures compare AdjustNVar to a single study outcome. I think the logic behind this comparison is flawed, however. It is comparing apples to oranges. The simulation assumes that, when applying AdjustNVar, ALL variables studied have a true effect. This, in effect, stacks the deck on statistical power for any method that considers multiple outcomes rather than a single outcome. For example, I ran a simulation comparing Bonferroni with 6 outcomes compared to a single outcome with n=50 per group. When the correlation is <0.8, Bonferroni also has more power than the single outcome. And, when I compared the Meff strategy to a single outcome in a variety of scenarios, Meff was always more powerful than a single outcome. I think a more useful comparison would be to directly compare different methods that handle multiple outcomes (e.g., PC to AdjustNVar to Meff). Response: Thanks again for helping clarify what is being simulated here. I hope this is clearer in the current article. The figures have been redrawn and I hope are now clearer. Perhaps one takeaway point is that, in being concerned to control type I error, the CONSORT recommendations appear to ignore the gains in power that can be achieved with multiple outcomes. 8. Related to comment (7), I don’t think the paper is justified in making this broad claim: “The Adjust NVar approach can achieve a more efficient trade-off between power and type I error rate than use of a single outcome when there are three or more moderately intercorrelated outcome variables.” This conclusion is true only when the intervention truly affects ALL outcomes, which is a narrow and arguably unrealistic case. A more realistic scenario is where the intervention works only on a subset of outcomes. In this case, the single variable strategy will be more statistically powerful than the multiple-variable strategies if you choose the right variable. Figures 4-6. Same issue as for Figures 1-3: comparing the principal components composite variable strategy (PC) to a single outcome is flawed because the simulation “stacks the deck” for PC by assuming that all outcomes have a true effect. I believe that Figures 1-6 should focus instead on comparisons of different methods for handling multiple outcomes. Response: same as for point 6; this is clearly a key issue, and I think it is clearer now that the alternative models (L2 and L2x) are included. 9. The article claims that power is only “slightly lower” for AdjustNVar compared with the PC strategy. However, when I run simulations comparing PC to AdjustNVar, I get consistently higher statistical power for PC and I would characterize the difference as more than just “slight”. For example, with N=50, global corr=0.6, global ES=0.3, and 6 outcomes (two-tailed test), I get power of 35% for AdjustNVar versus 37% for Meff versus 46% for PC. Response: thanks for pushing back on this – this is fair comment. 10.PC is always more powerful than AdjustNVar and Meff when we assume that all outcomes have a true effect. However, PC is not more powerful when we assume that only a subset of outcomes have true effects. For example, if I tweak the above simulation so that only three outcomes out of six have true effects of 0.3, this changes the power to 15% for PC and AdjustNVar, and 28% for Meff. This illustrates why one narrow simulation is insufficient for making general conclusions about the tradeoffs in performance between the different methods. Response: agreed. 11. In my simulations, Meff consistently has higher statistical power than AdjustNVar, which is a function of the fact that Meff always has a 5% Type I error rate whereas that of AdjustNVar is variable and sometimes lower than 5%. I don’t view it as a strength that AdjustNVar results in arbitrarily lower Type I error rates. It is better for the investigator to be able to precisely control the tradeoff between Type I and Type II error. Meff allows this whereas AdjustNVar does not. Response: fair comment 12. Figures 1-6: I found these graphs hard to read as they don’t have a clear take-home point. I would suggest removing the single outcome comparisons altogether and then reformatting so that the graphs directly compare AdjustNVar to Meff and PC (three lines). In one graph, hold effect size, sample size, and N outcomes constant, and then show the power of the three methods as a function of increasing correlation. In another graph, hold correlation, sample size, and effect size constant, and show the power of the three methods as a function of N outcomes. And so on. All methods aim to control the familywise error rate at 5%, so this should not be a variable. The fact that AdjustNVar sometimes results in a lower Type I error rate is incidental — it arises as a quirk of the method not as the intent of the researcher. Response: Agreed. Graphs reformatted as suggested and I hope are now much easier to interpret. 13. Where AdjustNVar may have an advantage over a method like Meff is for re-evaluating flawed published studies where researchers used multiple outcomes but did not account in any way for multiple testing. An AdjustNVar table such as Table 2 would give readers a quick way to reevaluate such studies without the need for any calculations or access to raw data. I would focus the paper more on this application. Response: Agreed. The paper has been revised to make this point." } ] }, { "id": "97181", "date": "10 Nov 2021", "name": "Daniel Lakens", "expertise": [ "Reviewer Expertise Applied statistics" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe author discusses more optimal ways to control error rates than the Bonferroni correction when researchers use multiple measures in a study that are positively correlated. In these cases the authors proposed to specify the number of variables that should be significant at the default alpha level (e.g., 0.05) to make sure the overall Type 1 error rate does not exceed 0.05.\nThe difficulty with correcting for multiple comparisons based on the number of variables that are significant is that you want to prevent a situation where a researcher performs a Stroop effect test, a Simon effect test, and a test for precognition, and conclude the hypothesis is supported because 2 out of 3 tests are significant. The solution the author proposes is to only use this approach if 1) the same latent variable is measured in different measures, and 2) the correlation between these variables can not be zero. I believe this is a valid approach. These assumptions are discussed enough in the article, but on page 11 (of the pdf version), one might want to discuss that whether different measures actually are ‘replicants’ could be a matter of debate among peers. One could imagine a situation where researchers in a field who disagree about measures would also disagree about whether different measures are replicants.\nThe comparison against the Bonferroni correction is one interesting baseline, but there is a large literature on how to correct for multiple comparisons that is also much more efficient than a Bonferroni correction, and which is the more interesting comparison. A strength of the Bonferroni correction is that it makes no assumptions about the variables that are corrected. But if one is willing to make assumptions, most importantly about the correlation between variables, more efficient approaches are available. How does this approach compare against other correction approaches? Although there are many correction approaches (this seems to be a particularly active field in neuroscience, where multiple comparisons when analyzing brain activation are common, and measures are strongly correlated), the following two references provide a starting point.\nFan, J., Han, X., & Gu, W. (2012). Estimating False Discovery Proportion Under Arbitrary Covariance Dependence. Journal of the American Statistical Association, 107(499), 1019–10351\nYekutieli, D., & Benjamini, Y. (1999). Resampling-based false discovery rate controlling multiple test procedures for correlated test statistics. Journal of Statistical Planning and Inference, 82(1), 171–1962\nThere are several things to consider. First, the proposed simulation based approach fixes the alpha level, and selects the number of variables that need to be significant. Numbers of variables are relatively crude, in that we can pick 1, 2, 3, 4, etc, but not 1.23 variables. Alternative approaches in the literature lower the alpha level. A benefit of these approaches is that the alpha level can be set at any value (e.g., 0.0352) to exactly control the Type 1 error rate. The consequences of this are also clear when we look at the figures and compare the principle component approach with the Adjust NVar approach. The familywise error rate for the Adjust NVar approach is often well below 0.05, while it is controlled at 0.05 in the principle component approach (and it is controlled at 0.05 in other approaches in the literature that lower the alpha level). The author discusses this (e.g., page 7 of the pdf version) in some detail, but the author seems slightly biased towards their own approach, stating that “the tradeoff between power and familywise error (expressed as a ratio) is higher for Adjust NVar.” It is not clear this ratio is a fair evaluation of the methods, and the information is difficult to distill from the figures (a Table with Type 1 error rates and Type 2 error rates would be more useful for this). This ratio is not so easy to summarize in a single sentence, I feel. If a design has 99.9% power, lowering the alpha from 0.05 to 0.02 has little effect on power, but if power is 0.8, lowering the alpha has a greater effect. Typically, the evaluation is done on the required sample size – which type of correction would require the smallest sample size? And then it becomes important to take a long more modern corrections for correlated variables as well.\nI could imagine other modern corrections are a bit more efficient, but the author makes a good point that the simplicity of the methods allows one to evaluate published studies. This might be a useful application. However, then I would like to urge the author to add an example. It would be good to see how this approach should be applied in a real use case (e.g., read a justification for why the measures are measuring the same latent construct, and how to make an assumption about the correlation) and how to interpret the results, depending on how many tests are significant.\nThe current simulation is somewhat limited. The idea that all correlations are identical will not be true in practice, and this will complicate the application of the proposed technique. What is the recommendation in practice? Should authors choose the largest correlation or the smallest correlation? Which approach is more or less conservative? What is the effect of differences in standard deviations, and does it matter if measures with low correlations have larger standard deviations? I am not sure all these factors will have a large influence but users will most likely need to know what to do.\nIt was very nice to see the paper was written in Rmarkdown. I was able to reproduce the results computationally (I did not repeat the simulations). As a minor comment, skipsim was no longer on line 207 but on 222. The files would be clearer if all simulation code was a separate R script that is run to generate the simulation data. The datafiles generated can then be read in at the top of the Rmd file. The ‘skipsim’ workaround does not improve clarity, and I had a hard time figuring out where the code reads in the data. In row 561, for example, the toybit file is written, but if I read it in, this is not needed. Separating the creation of data, and reading in data, makes this cleaner. I needed to create an ‘Images” folder to run the plot code on line 679 – this folder could be uploaded to the github repo perhaps?\nTo conclude, I believe this current version of the manuscript needs some additional work, which could include a more extensive discussion of other corrections in the literature, an exploration of additional simulations, and a practical example of how to use this approach when analyzing published studies.\n\nIs the rationale for developing the new method (or application) clearly explained? Partly\n\nIs the description of the method technically sound? Yes\n\nAre sufficient details provided to allow replication of the method development and its use by others? Yes\n\nIf any results are presented, are all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions about the method and its performance adequately supported by the findings presented in the article? No", "responses": [ { "c_id": "9033", "date": "13 Jan 2023", "name": "Dorothy Bishop", "role": "Author Response", "response": "I thank the reviewer for such a comprehensive and constructive review. The two reviews complemented one another very nicely, and helped me see how to restructure and revise the paper to make it clearer. As background, I should explain that this paper grew out of an attempt to write a primer on how to analyse/evaluate published intervention studies for students/professionals in allied health professions/education. My concern was that Bonferroni correction is often too conservative, but explanations of other methods are typically highly complex, so I was looking for a simpler rule of thumb that could be useful in interpreting published studies. I have emphasised this rationale more in the revised paper. Response to specific points. 1. The difficulty with correcting for multiple comparisons based on the number of variables that aresignificant is that you want to prevent a situation where a researcher performs a Stroop effect test, a Simon effect test, and a test for precognition, and conclude the hypothesis is supported because 2 out of 3 tests are significant. The solution the author proposes is to only use this approach if 1) the same latent variable is measured in different measures, and 2) the correlation between these variables can not be zero. I believe this is a valid approach. These assumptions are discussed enough in the article, but on page 11 (of the pdf version), one might want to discuss that whether different measures actually are ‘replicants’ could be a matter of debate among peers. One could imagine a situation where researchers in a field who disagree about measures would also disagree about whether different measures are replicants Response: This is an important point, which is similar to issues raised by reviewer 1, so I have now extended the simulations to take into account situations where outcomes may not all be indicators of one factor. 2. Alternative methods for correction for multiple comparisons. Response: The reviewer recommends two references with alternative approaches to multiple comparison correction. As I noted above, one goal was to keep things simple: so I am reluctant to do a full comparison of all methods, especially since it is clear that I need to consider different underlying data structures. I like the relative simplicity of the MEff approach recommended by reviewer 1, and so I am now presenting a comparison of Bonferroni, PCA and MEff. 3. I could imagine other modern corrections are a bit more efficient, but the author makes a good point that the simplicity of the methods allows one to evaluate published studies. This might be a useful application. However, then I would like to urge the author to add an example. It would be good to see how this approach should be applied in a real use case (e.g., read a justification for why the measures are measuring the same latent construct, and how to make an assumption about the correlation) and how to interpret the results, depending on how many tests are significant. Response: Again, this converges nicely with the recommendations of reviewer 1, and I have now restructured the paper to focus more on this aspect and to give a real-life example. 4. The current simulation is somewhat limited. The idea that all correlations are identical will not be true in practice, and this will complicate the application of the proposed technique. What is the recommendation in practice? Should authors choose the largest correlation or the smallest correlation? Which approach is more or less conservative? What is the effect of differences in standard deviations, and does it matter if measures with low correlations have larger standard deviations? I am not sure all these factors will have a large influence but users will most likely need to know what to do. Response: I now contrast three models with different patterns of intercorrelation between intervention and outcome. Potentially, there is no end to the options to consider, but I found it reassuring that with the MEff approach, the adjusted alpha was similar for different correlated variables, provided the mean off-diagonal correlation was constant. 5. It was very nice to see the paper was written in Rmarkdown. I was able to reproduce the results computationally (I did not repeat the simulations). As a minor comment, skipsim was no longer on line 207 but on 222. The files would be clearer if all simulation code was a separate R script that is run to generate the simulation data. The datafiles generated can then be read in at the top of the Rmd file. The ‘skipsim’ workaround does not improve clarity, and I had a hard time figuring out where the code reads in the data. In row 561, for example, the toybit file is written, but if I read it in, this is not needed. Separating the creation of data, and reading in data, makes this cleaner. I needed to create an ‘Images” folder to run the plot code on line 679 – this folder could be uploaded to the github repo perhaps? Response: I have now separated the simulation and generation of corresponding figures from the code to write the paper." } ] } ]	1	https://f1000research.com/articles/10-991
https://f1000research.com/articles/12-1439/v1	06 Nov 23	{ "type": "Research Article", "title": "Exploring students’ performance using lingua franca in science education: a study of grade ten students in Capiz, Philippines", "authors": [ "Dirk Diestro" ], "abstract": "Background Currently global competitiveness is the main thrust of the country’s education department, raising the quality of education in the Philippines has become a priority for government officials, who see it as one way to address other teaching and learning challenges. Hence, this quasi-experimental research aimed to determine the influence of lingua franca-based and English-based instruction on the science performance of Grade ten students.\n\nMethods The respondents of this study were forty-six Grade ten students out of the total population of fifty-three. Out of the forty-six respondents, twenty-three students were assigned to the experimental group and were taught using lingua franca-based instruction, and the remaining twenty-three were taught using English-based instruction. A researcher-made test was prepared and underwent validity and reliability tests. Mean was used to determine the science performance of the students, t-Test was used to test the significant difference between the students’ science performance before and after teaching using lingua franca-based instruction and English-based instruction, and Cohen’s d was used to compare two means in determining the effect size.\n\nResults It was found that both groups exhibited fairly satisfactory science performance in the pre-test, and in the post-test, the control group reveals satisfactory Science performance while the experimental group shows very satisfactory science performance. No significant difference was determined between the pre-tests of both groups and significant differences were determined between the pre-test and post-test of both groups, and in the post-test of both groups, a large size effect was obtained in using the lingua franca-based medium of instruction on the science performance of the students.\n\nConclusions Hence, the practice of Lingua Franca-based instruction exhibits positive and desirable results in improving the performance of the students.", "keywords": [ "Instruction", "Language", "Lingua Franca", "Science", "Students’ Performance" ], "content": "Introduction\n\nIn many countries, the medium of instruction has long been a source of contention among educators, particularly in nations that were once subject to colonial rule. Although the nations gained their independence, their legacy still exists in one form or another. Language is the most notable of these legacies. The colonization made by the American regime in the Philippines left its traces in terms of language. In this present time where global competitiveness is the main thrust of the educative process, raising the quality of education in the Philippines has become a priority for government officials, who see it as one way to address other teaching-learning challenges.\n\nThe preference for English as the main medium of instruction in Philippine schools has left students from marginalized groups with little opportunity to learn and grow. Looking at the scenario nowadays, it is a known fact that students learn and acquire language differently, so it may come as easy as mimicking. While others need more time and tedious processes. With this, the challenge lies in the hands of the teacher. The teacher may either make or break a meaningful and fruitful teaching and learning process.\n\nLanguage is a very important factor in the teaching-learning process since there is a mandate for the utilization of the medium of instruction. However, recent orders and memoranda present that students should be taught in a way that would suit the learners’ personal and national needs. There have been arguments and debates made in forums and seminars towards the best language of instruction in the teaching scenario and this gives a huge question in the field of the educative process that, could the utilization of bilingual education increase higher level learning and retention? Hence, this leads Philippine education to decide on the appropriate medium of instruction.\n\nIn the abovementioned scenario, the researcher contemplated the idea presented by Nisar and Ahmad (2011) that language is the medium of instruction by which all content in any discipline or level of learning is delivered to students. The question of which language should be used to teach particular subjects sparks controversy every time a new school system is put into place, since it challenges long-standing cultural norms Furthermore, the medium of instruction which refers to the language used holds a vital role in changing the teaching-learning process and making it easy or difficult for a student. The acquisition of learning among students is greatly influenced by their language since language is an important facet of culture. Therefore, it is considered an important instrument through which the change of cultural values is made easier and according to Amamio (2010), the individual develops his character from the perspective of his own cultural patterns, including languages.\n\nIn linguistics, the lingua franca refers to the alternative use of two or more languages in a single conversation. It is commonly used as a communication strategy among learners. Traditional methods or formal education often discourage lingua franca among students (Setati, 2008). However, as Abad (2010) discussed, a lingua franca is a language that people use to communicate when they do not share one natively. All speakers of any language have learned how to code-switch or change their speech patterns according to the situation and environment in which they find themselves. In the academic world, lingua franca or code-switching is defined as the usage of a secondary language in addition to the primary language during speech. This can be seen most prominently in an educational setting—where students are encouraged and taught to switch between languages during class discussion.\n\nFrom a general perspective, it is known that there are many factors that determine a student’s performance and language is one of those factors. Coherently, countries, where English serves as a second language, revealed that there is no single instructional strategy or methodology that can guarantee predictable success in class (Chang et al., 2007). As a result, the focus of various research has shifted from instructional strategies and methodologies to language acquisition and utilization of English vis-à-vis native tongue in science classes (Murcia, 2011).\n\nThe science performance of students at the secondary level according to the National Achievement Test shows a retrogressive pattern where the issues underpin the comprehension and analysis of the students (Philippine Basic Education, 2019). With the aforementioned situation, the utilization of the lingua franca is presented by the Department of Education as mother tongue-based instruction to address the retrogressing science performance of the students and to cope with the demand for the UNESCO Education for All program.\n\nIn the presented idea of the Department of Education, it is believed that the official country’s language should be taught since it is a platform for learning information meaningfully to make students’ vocabulary wider in terms of understanding and comprehension. The utilization of lingua franca drives the students to have maximum performance since students actively participate during discussion and via their mother tongue, and students are able to actually learn the curricular content of instruction (Thomas and Collier, 2007).\n\nGoing with the contemporary trend in education research and considering the above-mentioned scenario, the researcher was motivated to conduct a study that looks into the outcome of lingua franca-based instruction on students’ performance as compared to the typical practice of using English language-based science education.\n\nThis study determines the influence of lingua franca-based and English-based instruction on the science performance of Grade ten students. Specifically, it examines the level of science performance in each group at the start and end of the study, the significant difference between the pre-tests of the control and experimental groups, pretests and posttests of these groups, and between the posttests of these groups, and the effect size of lingua franca-based instruction and English-based instruction on the science performance of the students.\n\nThis study was anchored on the Experiential Theory of Kolb which stressed that the educational system should give opportunity to students to acquire and apply knowledge, skills, and attitude in a way that is relevant to their experiences (Kolb, 1984; Satati, 2010). Applying the theory in this study, it can be assumed that students’ performance will be meaningful if the medium of instruction used is relevant to the language that they are using in their day-to-day lives.\n\n\nLiterature review/study site\n\nLingua franca is a multilingual association of language used as a communication strategy among learners. In a multilingual society, language choice is heavily dependent on its sociolinguistic contexts including the location, participants involved, and the topic at hand (Spolsky, 2004). Approximately, there are more or less seven thousand existing languages in the world which brings the idea of linguistic diversity (Cenoz, 2013). As elaborated by Phillipson and Skutnabb-Kangas (2017), linguistic diversity is needed as it reflects the individual’s community culture, tradition, and identity.\n\nThe Philippines’ K to 12 Basic Education Curriculum includes a mandate for mother tongue-based instruction, beginning in kindergarten and continuing through third grade. This supports the goal of having all children, regardless of background, become readers and writers by the end of third grade. Although there is an argument about the use of English as a medium of instruction versus mother tongue-based instruction in consonance with the multilingual education, the trend among teachers nowadays is teaching their students using mother tongue-based instruction. Students who were taught using mother tongue-based instruction tend to think critically and were able express themselves with greater fluency. One of the main reasons for this policy is that some people believe children learn faster when they are taught by teachers who use a form of language with which they are familiar at first. Because native and regional languages help improve children’s language skills and strengthen their social awareness, teaching materials for bilingual learners should be in such languages (Valerio, 2013).\n\nQuijano (2010) underscores that reading fluency and comprehension were lowest among those students taught to read only in English or Filipino. Hence, multidisciplinary studies have been carried out to determine the best approach to teaching students a new language: either through their native tongue or by first learning an established international language such as English before using their own. Several case studies were conducted in the Philippines over the years on mother tongue-based multilingual education and these include the Iloilo Experiments, the Rizal Experiment, and case studies of other components such as Lingua Franca Project in Ifugao Province. The aforesaid studies have shown that when teachers use students’ native language as the instructional medium, those students who read with inclination have a greater chance of developing the skills necessary to succeed in science, math and other related fields. They also tend to do better in classes where their native language is used.\n\nAs revealed by Nolasco (2009), children with a strong background in their native language can more easily pick up other languages and when children begin school with their first language as the foundation for further learning, with an understanding of the connections between these two languages—which allows for a smooth transition into learning new subject matter—they tend to emerge as more competent learners overall. Reports on the Philippine educational system tend to emphasize weak English, science, and mathematics skills among Filipino students. Mother tongue-based multilingual education seeks to address the high functional illiteracy rate in the Philippines and its consequences—the high dropout and noncompletion rates among students.\n\nEvidently, during high school education, classes are taught in Filipino and English, but teachers use their mother tongue as a way to explain concepts when necessary (Licuanan, 2007). This paves the way for the crafting and implementation of DepEd Order No. 74, series of 2009—Institutionalizing Mother Tongue-Based Multilingual Education (MLE) which replaces the Bilingual Education Policy with its assertion that research has shown that students in local communities and around the world are more likely to succeed when they learn using their mother tongue as a primary language of instruction. Further, Nolasco (2009) stresses the suitability of multilingual education in the learning process by writing an article in Education’s Primer underpinning that MLE starts from where learners are, and from what they already know: “The strategy is to develop cognitive skills of learners in their first language—and then transfer them into the medium of instruction.” It has been demonstrated in numerous surveys, proficiency tests, and in the complaints of businesses that corporate employers have not found the Bilingual Education Policy as effective as originally hoped. According to Quijano (2010), a language’s efficiency is directly related to how often it is used, for example, Cebuanos prefer to use English instead of Filipino, which negatively affects proficiency in the latter. Although today’s children are generally fluent in oral Filipino, their fluency in writing and reading the language is limited. This problem becomes even more acute when it comes to English—the country’s official national language.\n\nOn the other hand, Quijano (2010) disputes multilingual education implementation by noting that childrens’ poor command of the English language is a result of their limited exposure to it aside from the books, magazines, and other instructional materials available in schools and libraries that are written in English. As a result, language-competence problems such as poor reading comprehension have arisen among students. Licuanan (2007) points out that to combat the dwindling proficiency of Filipino English speakers, efforts have been made to improve their command of the language. However, while the push for English as a primary medium of instruction may seem like an effective strategy to solve this problem, in fact language experts have noted that it might simply be an extension of the myth that if something is good then more will be even better. The deterioration of English must be understood in the context of the overall decline in Philippine education. This is not a problem limited to English— students are also performing poorly in math and science classes.\n\nWalter (2008) noted that many of the arguments against taking language into consideration in educational policies for developing nations have little to do with effectiveness. Thus, he pointed out in his findings that people often have a number of reasons for not learning another language. One reason is that they think it will give other people an advantage over them. Another is that they think it will be easier to build a country if everyone speaks the same language. Some people also do not want their children to learn languages other than one they already know, because they think it will confuse their children. Some people believe that if you speak a certain language, you should be able to use it in school and at work, which means that it has to be developed before it can be used as a medium of instruction. Some people think that if there are not many people who speak a certain language in one place, it won’t be worth teaching it there. Some people also believe that if there are not any jobs using a particular language available right now, there would not be anytime soon either.\n\nIn bilingual classrooms, as highlighted by Baker (2009), students often switch between different languages. Teachers regularly use two languages in a bilingual classroom without official backing from policymakers. They integrate these languages to achieve their teaching tasks. On the contrary, Jacobson (2009) argues that integrating both languages in the classroom is beneficial. He says that teachers should not randomly switch from one language to another, translate, preview, or review material, or use purposeful concurrent language use. However, Baker (2009) claimed that there are many language-switching cues that cause a speaker to shift languages. These include reinforcement of concepts, reviewing material, capturing students’ attention, and changing topics. Other cues are gaining rapport with students and changing from formality to informality. Translanguaging can create opportunities for deeper learning, language enrichment and collaboration between home and school.\n\n\nMethods\n\nThis study employed a quasi-experimental research design in determining the influence of lingua franca-based and English-based instruction on the science performance of Grade ten students using the pretest-posttest non-equivalent control design. In the pretest-posttest control group design, the experimental group received treatment that was withheld from a control group. Before the introduction of the intervention, surveys and tests were conducted in both experimental and control groups last October 24, 2022. After the intervention was introduced to the experimental group, a post-test and observation of both groups took place last January 13, 2023.\n\nThe respondents of this study were identified and grouped by employing matched pairs design to address the issue of “collective similarity,” and this was employed by matching the respondents in terms of their first grading grade in Science. The participants of this study were forty-six (46) Grade ten students from Capiz State University – Roxas City Main Campus, Laboratory High School out of the total Grade ten population of fifty-three (53) students. Out of the forty-six (46), 23 students were assigned to the experimental group and were taught using lingua franca as the medium of instruction, and the remaining 23 were taught using English as the medium of instruction. The respondents together with their parents or guardian were invited for the orientation of the purpose, preliminary plans and commencement of the study last May 24, 2022. The researcher sought their approval through a parental permission form.\n\nThe data gathering instrument of this study was initially made with 100-item researcher-made tests encompassing the topics in the third quarter specified in the curriculum guide for Grade ten science. The test questions were subjected to a reliability and validity process. After the validity process, pilot testing was conducted utilizing the Grade ten students from other schools who were not included as participants in the experimentation. After the pilot testing, the researcher subjected the 100-item researcher-made test to item analysis, difficulty, and discrimination index analysis to establish whether the items will be retained, revised, or rejected. As a result, from 100 items, only 60-item were accepted. The 60 items used in the pre-test and post-test were rearranged in the post-test. To interpret the scores obtained by the participants, the researcher used the arbitrary scale of Outstanding (48.51 – 60.00), Very Satisfactory (36.51 – 48.50), Satisfactory (24.51 – 36.50), Fairly Satisfactory (12.51 – 24.50), and Did Not Meet Expectations (0.00 – 12.50).\n\nPre-experiment phase\n\nThe researcher prepared the list of topics, daily lesson plan, and researcher-made test for the pre-test and post-test before conducting the research study. Also, during this phase, pilot testing was conducted. In addition, phase validity of lesson plans and the researcher-made test were done. Further, the researcher oriented the participants about the purpose of the study. After those significant steps, the researcher was ready in employing the intervention. A copy of the pre-test and post-test can be found under Extended data (Diestro, 2023).\n\nExperiment phase\n\nAs the experimentation commenced, the experimental group was taught using lingua franca-based learning as the medium of instruction, and the control group was taught using English-based learning as the medium of instruction. At the start of each lesson, the researcher administered a pre-test and proceeded to the delivery of the lesson wherein the entire teaching was conducted using the lecture-discussion strategy and differed only on the medium of instruction used by the researcher in the respective groups. After the delivery of the lesson, the researcher administered a post-test utilizing the researcher-made test used in the pretest however; the test item placement in the pre-test was rearranged in the post-test.\n\nPost-experiment phase.\n\nAt the end of the experimentation, students in the experimental group who were taught using lingua franca-based instruction were asked about their comments or reactions with regard to the medium of instruction employed. Additionally, during this phase, the scores of the students for both groups were recorded, tabulated, and analyzed using the appropriate statistical tools.\n\nMean was used in determining the science performance of the students taught using lingua franca-based instruction and English-based instruction, t-Test for paired samples was utilized to determine the significant difference between the students’ science performance before and after teaching using lingua franca-based instruction and English-based instruction and this inferential test was set at 0.05 alpha level of significance, and Cohen’s d was used when comparing two means to determine the effect size.\n\n\nResults and discussion\n\nThe results reveal that prior to the introduction of the intervention, the control group revealed “fairly satisfactory” (m = 17.04) science performance similar to the experimental group (m = 18.96). Moreso, after the intervention, the scores obtained in the post-test of the control group revealed “satisfactory” (m = 31.57) science performance and the experimental group showed “very satisfactory” (m = 42.17) science performance. This implies that the prior knowledge of the control and experimental groups towards that subject matter and competencies to be taught was the same. Also, this indicates that there was homogeneity of the scores obtained by students in the pre-test. The full raw data can be found under Underlying data (Diestro, 2023).\n\nOn the other hand, the results of the post-test revealed that regardless of whether lingua franca or English were used as the medium of instruction, students improved their science performance. However, the increase in performance was in favor of the lingua franca-based medium of instruction which exhibits a favorable effect in improving the science performance of the students. Moreso, the favorable improvement of students in the experimental group may be due to the openness of collaboration and exchange of ideas among learners and between teacher and students using lingua franca as the medium of instruction where understating and retention were very evident since students turn active and do respond more to the topics taught. The results of this study conforms to the study of Bernardo (2005) whose findings show that multi-lingual education could be used in facilitating students’ learning and achievement since the learning process is flexible and best appropriate for code-switching. In addition, Amamio (2010) reveals that people acquire their personalities through their interactions with the world around them, including other people and language which affirms the result of the study.\n\nThe result discloses that there was no significant difference between the pre-tests of both the control and experimental groups (p-value = .053) as shown in Table 2. This implies that prior to the start of the conductance of the intervention, students assigned to both control and experimental groups possess the same level of science performance which showed that both groups were very suitable for comparing the effect of lingua franca and English as mediums of instruction. The result of this study corroborates with the study made by Marzban et al. (2013) whose result reveals that the homogeneity of the participants was obtained before introducing experimentation between experimental and control groups to attain reliable results in determining the effect of different strategies on the reading comprehension of the learners.\n\nThe result shows that there were significant differences between the pre-test and post-test of the control group (p-value = .000), and of the experimental group (p-value = .000) as reflected in Table 3. The results imply that using lingua franca or English as the medium of instruction improves the science performance of the students. Further, the results infer that students in the control group improved their performance as reflected in Table 1 where the performance in the pre-test was “fairly satisfactory” which turned to “satisfactory” in the post-test. This means that students acquired new learning during the time of experimentation using English-based medium of instruction since the teaching-learning process took place where student participation, the art of questioning, effective teaching strategy, and assessment were all adhered. The result of this study affirms Delos Santos’ (2013) findings which discloses that the pre-test and post-test of the control group exhibited significant difference which indicates that students improve their mathematical performance from moderately high to high.\n\nOn the other hand, students in the experimental group also exhibited improvement in their science performance as shown in Table 1 which entails that the pre-test resulted in “fairly satisfactory” and after the intervention using lingua Franca-based instruction resulted in “very satisfactory” and it shows greater effect in terms of science performance. The said increase can be attributed to the reason that students felt comfortable explaining and expressing their idea using a combination of language where the students may abruptly shift to give emphasis on their point. This study conforms to the study of Gardner (2001) that the use of lingua franca helps students understand what is taught and lessen the difficulties of understanding the lesson which leads to the improvement of the performance of the students. More so, Warden et al. (2004) reveal that the medium of instruction affects learners’ scholastic performance.\n\nThe results show that there was a significant difference between both groups in the post-test (p-value of.000) in favor of the experimental group as reflected in Table 4. The result infers that the students assigned to the experimental group obtained high performance in science compared to those students assigned to the control group and this can be attributed to the belongingness that the students felt during the discussion since developing one’s sense of belongingness inside the classroom makes learning meaningful and easy for the students to acquire and retrieve knowledge since the reinforcement to the learning that took place was experienced-based. Moreso, using a combination of language helps the students facilitate comprehension and deep analysis during class discussions. The result of this study affirms the findings of Libutaque (2001) which reveals that the use of primary and secondary languages of the students in teaching math and science facilitates learning well. Furthermore, the results disclosed by Clarckson (2002) show that students who were fluent in both their first and second language demonstrate outstanding cognitive processes which result in successful learning which also conforms with the results of this study.\n\nThe obtained Cohen’s d value of 2.93 as shown in Table 5 infers that there was a large size effect obtained on the science performance of the students using lingua franca as the medium of instruction. This implies that the lingua franca-based medium of instruction when used in science classes, may surely exhibit positive and desirable results in improving the performance of the students given that using lingua franca as the medium of instruction motivates students’ to actively respond and inquire, ensures rapport towards students, promotes better understanding towards word meaning and maintains active teaching-learning process atmosphere in the classroom. The results of the study corroborates the findings of Delos Santos (2013) which showed lingua franca promotes students’ understanding due to familiarity with the medium of instruction used during the discussion. Also, according to Probyn (2012), it was identified that some teachers use the home language as a tool for scaffolding since these teachers believe that first language utilization is the way to accelerate second language acquisition which corroborates the results of this study.\n\n\nConclusion\n\nIt was found that in the pre-test, both groups displayed “fairly satisfactory” science performance. Also, the scores obtained in the post-test of the control group reveals “satisfactory” science performance and the experimental group shows “very satisfactory” science performance after subjecting the participants to the intervention. Hence, the respondents in both control and experimental groups have only a satisfactory knowledge of the topics to be taught during the conduct of the intervention. After the intervention, the post-test scores reveal that there was an improvement in science performance among participants in both control and experimental groups. Thus, whether lingua franca or English as the medium of instruction is used in the classroom, students may improve their science performance. However, the increase in performance was in favor of the lingua franca medium of instruction. Lingua franca-based instruction in science facilitates understanding and stimulates active participation in the class discussion. Therefore, employing lingua franca as the medium of instruction exhibits a favorable effect in improving the science performance of the students.\n\nThere was no significant difference between groups in the pretests. Therefore, students were all in the same level of science performance which reveals that the participants have the same level of knowledge acquisition before the study.\n\nThere were significant differences between groups in the pretest and posttest. Thus, this concludes that the use of lingua franca exhibits favorable dominance compared to English-based instruction in improving the science performance of the students. Moreso, students’ interests increase since the utilization of language allows them to express their ideas freely and understand the topic well.\n\nThere was a significant difference between groups in the post-test in favor of the experimental group. Therefore, the students assigned to the experimental group were able to increase their understanding, acquisition of knowledge, and learning retention due to the language used.\n\nThere was a large size effect obtained in using the lingua franca-based medium of instruction on the science performance of the students. Therefore, the usage of lingua franca-based instruction exhibits positive and desirable results wherein students taught using lingua franca improve their performance significantly better compared to the other group.\n\n\nEthical approval\n\nThe researcher secured permissions from the University President, Campus Administrator, Dean, and Program Chairperson to carry out the study. The ethical approval letter for this study was acquired from the University President and Campus Administrator of Capiz State University dated May 20, 2022. Following the approval, participants were informed about the study’s objectives and provided their consent for the utilization of data.", "appendix": "Data availability\n\nFigshare: Exploring Students’ Performance Using Lingua Franca in Learning Science. https://doi.org/10.6084/m9.figshare.22801796.v1 (Diestro, 2023).\n\nThis project contains the following underlying data:\n\n• Raw Data Lingua Franca.xlsx (file contains the scores of the students treated as science performance of the participants)\n\n• Statistical Analysis Tables Lingua Franca.xlsx (file are the tables for interpretation presented in the results and discussion in the manuscript)\n\n• Statistical Analysis Lingua Franca.doc (file contains the statistically treated data from the SPSS)\n\nThis project contains the following extended data:\n\n• POST TEST Exploring Students’ Performance Using Lingua Franca in Learning Science.pdf\n\n• PRETEST Exploring Students’ Performance Using Lingua Franca in Learning Science.pdf\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nAbad FC: Congressional Commission on Education. Manila, Philippines: 2010.\n\nAmamio L: Attitudes of students, teachers and parents of RVM schools in metro Manila toward English and Filipino as media of instruction. (Unpublished Thesis) Presented to the UST Graduate School, Manila, Philippines.2010.\n\nBaker C: Key issues in bilingualism and bilingual education. England: Multilingual Matters, Ltd; 2009.\n\nBernardo M: Code Switching, An Alternative Resource in Science and Math. Manila, Philippines: 2005.\n\nCenoz J: Defining multilingualism. Annual Review of Appl. Linguis. 2013; 33: 3–18. Publisher Full Text\n\nChang TC, Lim HC, Lee SH: English language proficiency and academic achievement among students of English as a second language at the college level (ERIC Document Reproduction Service No. ED 074812).2007.\n\nClarckson DJ: Principles underlying English language proficiency tests and academic accountability for ELLs.Abedi J, editor. English language proficiency assessment in the nation: Current status and future practice. Davis, CA: University of California, Davis, School of Education; 2002; pp. 13–31.\n\nDelos Santos JS: Effect of Lengua Franca-Based Instruction in Mathematics Performance of High School Students. Roxas City: Filamer Christian University; 2013.\n\nDepEd issued Order No. 74 series of 2009. Institutionalizing Mother Tongue-Based Multilingual Education (MLE). https://www.deped.gov.ph/2009/07/14/do-74-s-2009-institutionalizing-mother-tongue-based-multilingual-education-mle/\n\nDiestro D: Data file for F1000. Dataset. figshare. 2023. Publisher Full Text\n\nGardner RC: Integrative Motivation: Past, Present and Future. Distinguished Lecturer Series, Temple University Japan.2001.\n\nJacobson R: Allocating two languages as key feature of a bilingual methodology.Jacobson R, Faltis C, editors. Language distribution issues in bilingual schooling. Clevedon: Multilingual Matters Ltd; 2009.\n\nKolb DA: Experiential Learning: Experience as the Source of Learning and Development. New Jersey: Prentice-Hall; 1984.\n\nLibutaque M: The Use of Filipino as Medium of Instruction as Perceived by Secondary School Teachers. Iloilo City: WVSU; 2001.\n\nLicuanan PB: Language and Learning.2007. January 24, 2007.\n\nMarzban, Marzban A, Arabahmadi S: The Effect of Written Corrective Feedback on Iranian EFL Students’ Writing.Procedia - Social and Behavioral Sciences.2013; 83. Publisher Full Text\n\nMurcia FA: Students’ concurrent performance on tests of English language proficiency and academic achievement. In The validity of administering large-scale content assessments to English language learners: An investigation from three perspectives (CSE Rep. No. 663). Los Angeles: University of California, National Center for Research on Evaluation, Standards, and Student Testing (CRESST); 2011; 47–77.\n\nNisar RD, Ahmad HJ: Predictive validity of the test of English as a foreign language for Asian graduate students in engineering, chemistry, or mathematics. Educ. Psychol. Meas. 2011; 37(2): 461–463.\n\nNolasco RM: 21 reasons why Filipino children learn better while using their mother tongue: A primer on mother tongue-based multilingual education (MLE) and other issues on language and learning in the philippines. Presented at the Guro Formation Forum, Quezon City, Philippines. 2009.\n\nPhilippine Basic Education: Why Students from the Philippines Perform Poorly in PISA 2018.2019. Reference Source\n\nPhillipson R, Skutnabb-Kangas T: Linguistics human rights, past, and present.2017.\n\nProbyn MJ: Teachers’ voices: Teachers reflections on learning and teaching through the medium of English as a second language. Int. J. Biling. Educ. Biling. 2012; 4(4): 249–266.\n\nQuijano Y: MLE in the Philippines: History and Possibilities. First National Conference onMother Tongue Based Multilingual Education. February 18-20, 2010. Capitol University, Cagayande Oro, Philippines; 2010.\n\nSatati K: Predictive validity of the test of English as a foreign language for Chinese graduate students at an American University. Educ. Psychol. Meas. 2010.\n\nSetati M: Between Languages and Discourses: Language Practices Primary Multi-language Mathematics Classroom. South Africa: 2008.\n\nSpolsky B: Language Policy. Cambridge: Cambridge University Press; 2004.\n\nThomas E, Collier W: Language proficiency and academic success: Relationships between proficiency in two languages and achievement among Mexican American students. Biling. Res. J. 2007; 21(4): 395–408.\n\nValerio MTB: Current Perspectives on Mother – Tongue Based Instruction in the Newly Implemented K to 12 Curriculum of the Philippines (Published).2013.\n\nWalter SL: Does language of instruction matter? Language and Life: Essays in Memory of Kenneth L. Pike. Wise MR, Headland T, Brend R, editors. Arlington. 13 Duncan Road, Gillingham, Kent, ME7 4LA. United Kingdom: Publications in Linguistics, No.139. Dallas: SIL International and the University of Texas; 2008.\n\nWarden L, Probyn MJ, Murray S, et al.: Taiwanese Students Attitude Made Use of the Likert Scale Combined with Open Ended Question in Teaching Mathematics.Taiwan; 2004." }	[ { "id": "229977", "date": "25 Jan 2024", "name": "Sohaib Alam", "expertise": [ "Reviewer Expertise Pedagogy and Education", "Teaching and Learning", "Pedagogy", "Learning", "Collaborative Learning", "Blended Learning", "Cooperative Learning", "ICT in Education" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nDear Author, This paper has focused on an interesting and innovative topic. However, some major issues need clarification and additional work. I have to say, if some serious problems are not properly addressed. 1. What are the findings? 2. How did these motivate the researchers to conduct this study? 3. Where are the findings? There should be concrete findings of your study. 4. The major concern is no baseline provided for further data analysis. Were there statistically significant differences in prior achievement between the two groups? If the experiment group performed better than the control group at the beginning, it is not robust to imply that an intervention was effective in promoting student achievement. The authors would like to check their data and answer this with caution. 5. Please cite related literature of last five years.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? No\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "250151", "date": "11 Mar 2024", "name": "Balachandran Vadivel", "expertise": [ "Reviewer Expertise English Language Teaching", "Second Language Acquisition", "Applied Linguistics and Educational Psychology." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nWhile the study provides valuable insights into the potential benefits of lingua franca-based instruction, there are a couple of potential limitations worth considering. Firstly, the sample size of only forty-six Grade ten students from a single school may limit the generalizability of the findings. A larger and more diverse sample across multiple schools could provide a more comprehensive understanding of the effects of instructional language on science performance. Additionally, the study could have benefited from a longer-term follow-up to assess the sustainability of the observed improvements. Without longitudinal data, it's difficult to determine whether the enhanced performance seen immediately after the intervention persists over time or if it diminishes. These limitations should be addressed in future research to ensure the validity and applicability of the findings beyond the specific context of this study.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "250138", "date": "02 Apr 2024", "name": "Salah Ben Hammou", "expertise": [ "Reviewer Expertise Applied Linguistics", "EMI", "CLIL", "Bilingual education" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nDrawing on quasi-experimental research design, the study investigated 10th grade students' science performance level in two different MOI (medium of instruction environment) environments: Lingua franca-medium instruction and English-medium instruction. The findings showed that both groups (the experimental and control groups) achieved satisfactory results in the post tests.\n\nRevisions required:\n\nAbstract:\n\nJustify the use of quantitative research design for this study in one sentence. Provide a sentence on the pedagogical implication of the study.\n\nIntroduction:\nThe reader would want to know the objective(s) of the study at this stage. A paragraph on the significance of the study and the gap the author is trying to bridge in the existing literature is also advisable at this stage.\nReview of literature\n1. This section needs consideration. A s a reader, I expected the author to report similar previous findings on the impact of Lingua franca-based instruction and EMI on students' performance in science from different context worldwide. 2. including updated existing research on the topic is advisable.\n\nMethodology: Explain the choice of the quasi-experimental research design? why not mixed methods design? Results 1. I suggest separating the section results and discussion. 2. Did the author follow any procedures to ensure the accuracy and reliability of the datasets. 3. Perhaps the author should relate the results to the research questions. Discussion\nAvoiding repeating the results presented in the previous section. The reader would want a stronger discussion which answers to the research questions by contrasting the previous studies (especially those from the local context) with the new data gathered.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? No\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1439
https://f1000research.com/articles/12-1433/v1	06 Nov 23	{ "type": "Research Article", "title": "Identification of informative genes and sub-pathways using Improved Differential Expression Analysis for Pathways (iDEAP) for cancer classification", "authors": [ "Nurul Athirah Nasarudin", "Mohd Saberi Mohamad", "Zalmiyah Zakaria", "Richard O. Sinnott", "Fatma Al Jasmi", "Noura Al Dhaheri", "Nurul Athirah Nasarudin", "Zalmiyah Zakaria", "Richard O. Sinnott", "Fatma Al Jasmi", "Noura Al Dhaheri" ], "abstract": "Background: Pathway-based analysis primarily focuses on sub-pathway-based analysis, which aids in understanding biological reactions. Several studies have found abnormalities in pathways caused by certain regions based on the etiology of diseases. The Differential Expression Analysis for Pathways (DEAP) method is one such sub-pathway-based analysis method, that identifies a local region perturbed by complex diseases based on larger pathway data. However, the method has low performance in identifying informative pathways and sub-pathways. Methods: In this paper we propose an improved DEAP (iDEAP) method for enhanced identification of perturbed sub-pathways that achieves higher performance in the detection of differentially expressed pathways. Firstly, a search algorithm adapted from the Detect Module from Seed Protein (DMSP) algorithm was implemented as part of the DEAP method to search for informative sub-pathways. Secondly, the relation among sub-pathways was taken into account by averaging the maximum absolute value for the DEAP score for the reaction among sub-pathways to support the efficient identification of informative pathways. Three gene expression data sets were applied to this research. Results: The proposed improved method shows a better performance over the previous methods. In fact, when the identified genes from the results were assessed using 10-fold cross validation to classify cancer based on accuracy, the improved method shows higher accuracy for colorectal cancer (90%) and breast cancer (94%). Conclusions: This shows that the proposed method effectively identifies informative genes related to the targeted phenotype. A biological validation was also conducted on the top five significant pathways and selected genes based on biological literature. The results from this analysis will be useful especially in the medical field for disease detection. In 10 years and beyond, computational biology will become ever more entwined with biomedical research and medicine.", "keywords": [ "Sub-pathway-based analysis", "iDEAP", "Support Vector Machine", "10-fold Cross Validation", "Cancer Classification" ], "content": "Introduction\n\nIn the life sciences, emerging high–throughput technologies such as next-generation sequencing, -omics technology and microarrays allow for creation of massive amounts of highly dimensional biological data. Such data can cover genome, transcriptome, epigenome, proteome, metabolome, molecular imaging, and molecular pathways. Many sophisticated analytic methods have been developed to broaden the biological interpretation of differentially expressed genes and pathways. The earliest approach involved gene-by-gene analysis through individual gene analysis (IGA), which produced a list of altered genes using a cut-off threshold. 1 Subsequently, systems-level methodologies have pushed forward the transition of IGA to include gene set analysis (GSA), that can identify gene sets in a more subtle way, coordinated by a single-step process. Even though GSA methods have the advantage to researchers in characterizing groups of genes, they have limitations when applied to pathway datasets. Many of GSA methods disregard the graphical structure of pathway data, resulting in the potential omission of critical information regarding the biological interactions between molecules. As a consequence, this can lead to inaccurate outcomes. Pathway topology-based analysis has been introduced to overcome the limitations of GSA methods by considering the pathway structure. This analysis has integrated the information of gene ontology and pathway structure to discover which pathways are associated with a particular phenotype. In addition, two hypothesis tests can be observed. First, entire pathways can be tested for differential expression, and secondly identified informative sub-pathways represent the entire pathway with massive information associated with the differential expression. The second hypothesis is a more recent evolution in topology-based analysis as it can improve the specificity and sensitivity of the outcomes.2 Previous studies stated that the pathway structure information can provide relevant biological insights and contribute to the understanding of higher integrative levels of biological functions that are more complex with many variations and characteristics of information.3 Recently, topology-based analysis has shifted towards sub-pathway-based analysis, which provides information on biological phenomena more precisely, and hence can identify regions of the pathway that are dysregulated by diseases.4 In addition, previous studies have proved that deformities in sub-pathway regions of the pathway might contribute to the etiology of the disease.5 An overview of sub-pathway-based analysis is illustrated in Figure 1.\n\nSeveral sub-pathway-based analysis methods have been developed which share the same target in the search pathway portion related to disease modeling, drug targeting and other objectives.6–9 However, most of the methods have constraints that need to be improved due to a range of challenges. One of the challenges is how to examine the sub-pathways.10 Most of the sub-pathway methods independently search sub-pathways without implementing any search algorithm. Moreover, some researchers assume extraction of sub-pathway strategies do not affect the results.2 In addition, the pathway structures can be complex and involve the combination of many sub-pathways and interactions. Due to this, an efficient sub-pathway-based analysis method is essential to identify the specific region that is differentially expressed by utilizing all information within a given pathway. Therefore, this paper integrates the DMSP (Module from Seed Protein) algorithm to identify informative sub-pathways from the specific nodes and expanding them to the entire pathway network. The sub-pathway with the most informative genes is used to produce the best biological insight in identifying informative pathway related to the given phenotype under investigation.\n\nNumerous biological pathway databases now exist such as Kyoto Encyclopedia of Genes and Genomes (KEGG),11 Gene Ontology (GO),12 Biocarta (https://maayanlab.cloud/Harmonizome/dataset/Biocarta+Pathways) and many more. The majority of these pathway databases are not specific to certain biological contexts such as cancer. By implementing sub-pathway-based analysis, many informative pathways can be identified and leveraged and improve biological databases. In addition, the knowledge of genes within informative sub-pathways highly related to diseases can be applied for future studies such as cancer classification.13 Previous cancer studies are clinical and have limitations in diagnostic ability.14 Usually, gene expression data that is gained from microarray experiments is used to performed cancer classification with the implementation of advanced machine learning methods. The use of gene expression data poses a challenge for cancer classification, especially when using traditional approaches.\n\n\nMethods\n\nDifferential Expression Analysis for Pathways (DEAP) method is a sub-pathway analysis method that can find informative pathways based on the maximum absolute running sum score of sub-pathways. This method calculates the sub-pathway maximum absolute summation score by considering the interaction between nodes, where catalytic/inhibitory edges are taken as positive/negative summands. There are five primary steps in the DEAP method: data pre-processing, data mapping, identification of sub-pathways, absolute value calculation and statistical calculation of pathways as shown in Figure 2. First, the gene expression data undergoes a pre-processing stage to generate a null distribution using random rotations. Then, the algorithm starts by mapping the expression data onto the pathway graph. Next, the sub-pathways in the pathway are recursively identified from root to leaves nodes. A recursive function calculates the maximum absolute running summation score for all sub-pathways within the pathway by considering whether the reactions are catalytic or inhibitory. Every reaction represents a value, which is used for calculating each sub-pathway. From the calculation, the sub-pathway with maximal differential expression is determined. The maximum absolute value represents the DEAP score, which is returned.\n\nIn order to extract a sub-pathway that is related to the targeted phenotype, many methods had been developed based around differentially expressed (DE) genes. The differential expression analysis for pathway (DEAP) method15 utilizes the information in biological pathways to identify important paths by integrating differential expression data. Despite the good performance of DEAP, the identification of sub-pathway efficiency still can be improved by integrating a searching algorithm starting from the most associated genes in the pathway. The DEAP method searches for the possible sub-pathway within the pathway without taking other information into account. It also only uses a basic search algorithm embedded in the DEAP method. Basically, the search starts from the root node to the leaves node and goes through all the nodes until the end. The efficiency of searching can be improved by implementing a search algorithm to find informative pathways. Recent research has shown that the use of search algorithms can provide better results.16 In sub-pathway-based analysis, one of the main concerns is the precision in identifying sub-pathways within a given pathway. Sub-pathways were often considered without taking into account biological relations within the pathway. Thus, non-informative sub-pathways could be erroneously identified during the analysis of data used for specific biological contexts like cancer. This is because not every gene in the pathway is involved in biological processes and diseases like cancer. This research proposes the DMSP search algorithm to the DEAP method to improve the efficiency of the identification of informative sub-pathways and genes. The proposed method is referred to as iDEAP.17 In the proposed method, the search process is improved to search for the best informative sub-pathway with an additional step introduced to average all of the DEAP scores to consider all interactions between sub-pathways. The overview of the workflow of the iDEAP method is shown in Figure 3. The improved parts that show the differences between DEAP and iDEAP can be seen in Figure 3. Where the proposed algorithm is implemented. The code from previous work is combined with a proposed searching algorithm and average calculation code. Furthermore, new codes have been developed specifically for data pre-processing, ensuring that the data are appropriately prepared.\n\nThe purpose of data pre-processing is to remove uninformative data that affect the results and to ensure the data is suitable for input. Firstly, the uninformative data are removed from the dataset, followed by a normalization process. This step is important to correct the expression data value according to different cellular inputs. Next, the geneID is converted to UniProtID as this analysis utilizes the UniProt identifier. Figure 4 shows the flowchart of the data pre-processing activity.\n\nIn this phase, the gene expression data are integrated with the pathway graphs. All cellular components within the pathway are extracted as nodes in the corresponding graph. Each node contains multiple genes which have a different unique ID. This ID is used to derive the expression values from the gene expression data. The process of mapping is illustrated in Figure 5.\n\nDMSP searching algorithm was introduced in 2007 for integrating gene expression data and protein-protein interactions (PPI) to determine functional modules. This algorithm is able to determine interactions among a set of proteins in each graph. Generally, the idea is to discover biologically relevant PPI subnetworks within a larger network, whose proteins interact significantly. This algorithm is setup to identify functional modules starting from a ‘seed’ protein (the most informative protein) in the dataset.\n\nIn this work, a search algorithm was implemented to extract sub-pathways by taking topology information into account. This algorithm is adapted from the DMSP algorithm18 where the search process starts from the most interesting node. The package edgeR19 is used to detect the most differentially expressed genes to be appointed as the most interesting nodes in the pathway. The sub-pathways were extracted based on the perturbation caused by the most interesting nodes in the pathway as shown in Figure 6(b). The most interesting nodes are selected based on the value of the most differentially expressed genes.\n\nThe search algorithm works in two phases. First, the search is conducted by selecting the internal and external nodes to form a sub-pathway. Second, the nodes in the sub-pathway are pruned to select highly connected sub-pathways as shown in Figure 6(c) and (d). To obtain a compact sub-pathway, the node is removed from the sub-pathway if it satisfies the criteria: Ninternal > Nexternal where Ninternal is the internal node nearest to the interesting node and Nexternal is the external node. This process is recursively repeated for every sub-pathway. The process is illustrated in Figure 6 and the flow of the process is shown in Figure 7.\n\nGiven an edge and all other edges in the graph together with the expression values for all genes, we consider the expression data with two conditions (health/cancer), Ex. This is defined to be the difference between the logarithm of the arithmetic mean of expression values associated with gene(s) x. Next, all edges in the sub-pathway are examined recursively, where maxrecursive represents the maximum values and minrecursive represents the minimum values of the edge. The algorithm examines all possible edges in the sub-pathway set whose reactant node is the current edge’s product node. If there are no such edges, we set maxrecursive and minrecursive as ∑Ex where y∈products refer to each gene contained in the edge’s products. Otherwise, maxrecursive and minrecursive are defined as the maximum and minimum score, respectively. The formula to calculate the maximum score and minimum scores is as follows:\n\nWhere Tedge is the multiplier associated with the edge type (-1 or 1) for inhibition or catalysis, z∈reactants refer to each protein, z, contained in the edge’s reactants. Finally, the maximum and minimum value of DEAP scores are returned for each sub-pathway. The process of establishing the DEAP score is shown in Figure 8.\n\nThe goal of this research is to include all relations or interactions within a pathway to provide a finer resolution to represent relevant biological process related to a given target phenotype. To support this, all scores of the sub-pathways are calculated together and the average of the scores taken. The average scores are calculated based on the maximum score of each sub-pathway based on the equation below:\n\nIn this equation, MaxScore represents the maximum score of every sub-pathway in the pathway. The maximum score of the i-th sub-pathway, MaxScorei is calculated based on the recursive function in DEAP. The average of the maximum score, avg is calculated by summing all MaxScores for a sub-pathway and dividing by the number of loops (n) in the recursive function for one pathway.\n\nThe average of the maximum score is used for the statistical calculation to evaluate the significance of a given pathway to the target phenotype. The statistics used here are based on maximum order statistics.15 The statistic calculation of the proposed method is shown by the following equation:\n\nSince the research focuses on pathway-based analysis, two common data sets have been used: pathway data sets and gene expression data sets. Specifically, three gene expression datasets were used in this research. Table 1 shows the gene expression dataset used here. These three datasets which are head and neck tumor,20 colorectal cancer,21 and breast cancer22 can be downloaded from the NCBI GEO database.\n\nA total of 177 graph-based pathway data sets were downloaded in Systems Biology Markup Language (SMBL) format. For the interpretation, the pathways were broken into their protein components. Every pathway represented information about protein, biochemical reaction, and other substrates. The pathway data set is used to group the gene expression data based on the sample pathways. Matching processes between the gene expression data set and the pathway data set was undertaken. All the pathways were taken from the Protein Analysis Through Evolutionary Relationships (PANTHER) database related to regulatory and metabolic pathways.23 The pathway data sets can be downloaded at http://www.pantherdb.org/downloads/.\n\n\nResults/Use cases\n\nMost sub-pathway analysis methods are assessed based on the number of differentially expressed pathways found. In order to verify the performance of the iDEAP method, this research applied three sets of biological data on the biological pathways and made comparisons with previous work. The result of the proposed method was based on comparison with previous methods, namely the DEAP,15 SubSPIA,16 MinePath,24 HiPathia,25 and PsSubpathway.26 Table 2 shows the comparison result of the iDEAP method with the previous methods based on the number of informative pathways found. As noted, significant pathways were selected based on a p-value less than 0.05 (p-value ≤ 0.05). Generally, the performance of iDEAP method was improved based on the results obtained. This proves the effectiveness of the search algorithm for identification of informative sub-pathways. In addition, the interaction and relation between sub-pathways are important as additional information to help the medical sector detect diseases. By considering all interactions, the tendency to identify informative pathways related to the targeted phenotype is increased.\n\nThe iDEAP method performed well for head and neck tumor and breast cancer but unfortunately, not for colorectal cancer. This is because the colorectal cancer data was not suitable for this research, since most of the genes were not complimentary to the pathway data resulting in sub-pathway interactions being reduced. Moreover, the huge size of colorectal cancer data could affect the results, since it might contain significant noise data that obfuscates the informative data.\n\nThe performance of the proposed method was evaluated through a 10-fold cross validation classification in terms of accuracy. These measurements were used to justify the method performance by using the identified informative genes in the sub-pathways. These genes were selected from sub-pathways with p-value less than 0.05 and underwent a classification process using support vector machine (SVM) algorithm based on a cross validation. SVM algorithm is widely used for cancer genomic classification and prediction. Therefore, the classification accuracy can be used to analyze the effectiveness of the proposed method in identifying informative genes to targeted phenotype. To get a consistent result, the classifications were run 10 times, then, the average was calculated. The comparison of the average 10-fold cross validation (CV) classification accuracy between the iDEAP method and DEAP method for all data sets is presented in Table 3.\n\nIt is important to validate the result in the biological context with literature and databases to show the relevance of the research. This validation was manually undertaken after the experiments were conducted. All the identified informative genes and pathways based on the result in the proposed method were checked through biological literatures and databases. This study used google scholar as the biological literature and Genecards as the biological database.\n\nThe pathways were analyzed individually and consequently produced corresponding p-value and informative sub-pathways. According to Figure 9, the top five pathways with the corresponding p-value and the associated list of informative genes were selected and validated based on the biological database and literature in order to show biological relevance.\n\nTable 4 presents the top five pathways selected from the proposed method for the head and neck tumor data sets. The first ranked pathway was the Notch signaling pathway that has been implicated in the regulation of self-renewal capacity, cell cycle exit, and neural stem cell survival.27 Notch signaling is often associated with cancer diseases including head and neck squamous cell carcinoma (HNSCC).28 In addition, meta-analysis reveals that this pathway plays an important role in tumor development.29 The proposed method selected three informative genes for classification where all genes found were related to the development of head and neck tumors. CSL is one of the nucleus DNA-binding factors which interacts with an intracellular fragment of NOTCH.29 The NOTCH4 gene was found to be significantly related to HEY1 gene activation in HNSCC which promotes cell proliferation, cisplatin resistance, inhibition of apoptosis and cell-cycle dysregulation.30 MAML1 regulates transcription of Notch target genes and interacts with muscle-specific genes like MEF2C as a fundamental coactivator of other cell signaling pathways.31\n\nThe second rank pathway with the lowest p-value was TGF beta signaling pathway. Transforming growth factor-β (TGF-β) is a homodimeric protein that is known as a multifunctional regulator in the target cell and plays a role in numerous types of cancer including HNSCC.32 The defective TGF-β signaling within epithelial cells promotes the growth of tumors and increases the inflammation in tumor stromal cells.33 From biological validation, seven of the 15 informative genes were identified (JUND, EP300, CITED2, JUNB, SMAD5, SKIL, SMURF1) as being related to the development of HNSCC.\n\nThe third rank pathway was the Ras signaling pathway. Ras is a family of proteins called GTPase that is commonly involved in cellular signal transduction.34 The Ras signaling pathway is rarely related to HNSCC, but this pathway had been shown to be highly significant to this cancer by a meta-analysis of differential protein expression networks.35 In this pathway, eight informative genes were selected by the proposed method and all the genes (PIK3CA, PIK3C3, PIK3CG, PIK3CB, PIK3CD, HRAS, NRAS, KRAS) were found to be involved in HNSCC.\n\nThe fourth rank pathway was the JAK STAT signaling pathway. This is involved in cell division, cell death and most importantly tumor formation.36 STAT signaling was identified by the literature to play an important role in cancer formation and progression.37 In addition, the constituents of JAK STAT activation have been recognized in many cancers including head and neck cancer.38 In this pathway, nine genes were selected for the classification and all these genes (STAT5A, STAT5B, STAT3, STAT4, STAT1, STAT6, JAK, JAK2, JAK1) were associated with HNSCC based on the literature.\n\nTable 5 shows the top five ranked pathways identified by the iDEAP method for colorectal cancer data. The first top ranked pathway was the FAS signaling pathway. FAS also known as Apo1 or CD95, are death domain-containing members of the Tumor Necrosis Factor Receptor (TNFR).55 The FAS pathway was identified to be functional in colorectal cancer and induced apoptosis.56 Based on biological validation, only one (FASL) gene was identified to be related to the growth of colorectal cancer.\n\nThe second top ranked pathway was the 5HT3 type receptor-mediated signaling pathway also known as the 5-Hydroxytryptamine3 receptor. This is a member of the cys-loop family of ligand-gated ion channels.57 Although there has been no study that has proven the direct relationship of this pathway to colorectal cancer, digestive function involvement in colorectal cancer is common.58 Furthermore, the expression of 5-HT3 subunits exist in the colon and the small intestine is related to colorectal cancer.59 From the iDEAP method, 11 genes were selected for classification and five of those genes (STX3, SNAP25, VAMP2, VAMP1, SLC6A4) were found to be involved with colorectal cancer.\n\nThe third ranked pathway was the p53 pathway which is a tumor suppressor protein in humans regulated by the TP53 gene.60 A previous study showed that the p53 tumor suppressor was frequently found in colorectal cancer.61 Consequently, six informative genes were selected by the proposed method and only five genes (TP73, RCHY1, TP53, MDM2, MDM4) were related to the development of colorectal cancer.\n\nThe fourth ranked pathway was the platelet-derived growth factor (PDGF) signaling pathway. This pathway has been studied in cancer progression as PDGF can regulate many cellular processes.62 The PDGF signaling pathway consists of four ligands and two receptors involved in colorectal cancer progression.63\n\nThe fifth ranked pathway was the JAK STAT signaling pathway, which is a chain of interactions between proteins involved in immune function, cell growth and tumor formation.64 Previous research has reported that this pathway was differentially expressed in colorectal cancer tissues.65 In this pathway, the proposed method selected nine genes for classification and all of these genes (STAT5A, STAT5B, STAT3, STAT4, STAT1, STAT6, JAK3, JAK2, JAK1) were found to be related to the development of colorectal cancer.\n\nFor the breast cancer data set, the result of the top five pathways is shown in Table 6. The first top ranked pathway was the Ras signaling pathway. As noted, Ras is a family of related proteins which belongs to the small GTPase class that is involved in cellular signal transduction. Mutation in Ras genes can cause unintended and overactive signaling inside the cell, thus Ras signaling pathways ultimately lead to cancer.79 Previous studies showed that this pathway was activated persistently in nine widely studied human breast cancer lines.80 Based on the proposed method, seven informative genes were selected for classification and all the genes (PIK3C3, PIK3CG, PIK3CB, PIK3CD, HRAS, NRAS, KRAS) were found to be involved in the development of breast cancer.\n\nThe second top ranked pathway was the Notch signaling pathway. This is involved in the development of neural tissues, blood vessels, heart, pancreas, mammary gland, T lymphocytes, hematopoietic lineages, and other cell types.81 The current study identified that the Notch signaling pathway had major participation and multiple roles during breast tumor progression.82\n\nThe third top ranked pathway was the JAK STAT signaling pathway. The Janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway significantly contributes to the transmission of signals from cell-membrane receptors to the nucleus, playing a pivotal role in this process. Moreover, the JAK-STAT pathway is indispensable for numerous cytokines and growth factors, which are responsible for crucial cellular processes such as hematopoiesis, lactation, and the development of the immune system and mammary glands.83 A previous study revealed that chemoresistance in breast cancer was associated with the activation of JAK/STAT signaling and it was suggested that JAK2 may be useful in combating chemoresistance in breast cancer.84\n\nTwo informative genes (CNR1, GNAI3) were selected for classification and both genes were identified as being involved in breast cancer progression. The fourth top ranked pathway was the Thyrotropin-releasing hormone receptor (TRHR) signaling pathway. The TRHR is a G protein-coupled receptor that binds to the tripeptide thyrotropin releasing hormone.85 Dating back to the late 18th century, the administration of thyroid extract was often used in conjunction with oophorectomy as a treatment for breast cancer.86 In this pathway, seven genes were selected by the proposed method and six genes (CACNB3, CACNA1E, CACNA1A, CACNA1B, CACNB1, CACNB4) were found to be involved in the progression of breast cancer.\n\nFinally, the fifth top ranked pathway was the Interleukin (IL) signaling pathway from the proteins of interleukins family. This pathway regulates numerous biochemical events, including cellular proliferation and long-term survival. Previous studies have shown many of the interleukin families contribute to the progression of breast cancer. For example, the IL17 family consists of six protein members, among them IL17B and its receptor. The IL17RB signaling pathway plays a key role in the development and progression of breast cancer.87 For this pathway, 12 informative genes were selected to undergo the classification process and only 11 genes (STAT5A, STAT5B, STAT2, STAT3, STAT1, STAT4, STAT6, MAPK6, MAPK1, MAPK3, MAPK7) were identified as being involved in the development of breast cancer.\n\n\nConclusions/Discussion\n\nPathway-based analysis has led to a new era in genomic studies which integrates the benefits of gene-set analysis and enhances them with prior information based on gene interaction within pathways. However, early methods of pathway-based analysis relied on enrichment-based approaches and identified differentially expressed pathways without identifying specific regions related to the target phenotype.2 The results are not entirely accurate and complete since the current methods do not consider interactions involving functional molecular pathways.113 Usually, complex diseases like cancers involve interactions between genes that causes the genes to be expressed differently compared to a single gene. In order to obtain more specific biological knowledge, pathway-based analysis needs to shift to sub-pathway-based analysis which can identify regions of pathways that are dysregulated by diseases or involved in drug-related perturbations. Therefore, investigating sub-pathways is more relevant, since it can provide finer-grained resolution representing the underlying biological processes more accurately.114\n\nOne important feature of sub-pathway-based analysis is the ability to exploit the maximum interaction between nodes in pathways. In recent years, a series of methods had been developed to find solutions for sub-pathway analysis that identify informative sub-pathways accurately. This research proposes an improved differential expression analysis for the pathway (iDEAP) method which identifies informative sub-pathways and genes in pathways by considering all the interactions involved in the pathways. The iDEAP method extends the DEAP method by implementing the DMSP search algorithm to identify the informative sub-pathway as well as through modifying the calculation algorithm based on a recursive function used to obtain the average DEAP score for all sub-pathways. This is because the DEAP score of a single sub-pathway can lead to inaccurate interpretation since the size and structure among pathways are different.15 A Support Vector Machine (SVM) classification algorithm had been implemented to measure the performance of the proposed method based on the genes selected within significant pathways. Lastly, the iDEAP method used Genecards and literatures to validate the identified pathways and genes.", "appendix": "Data availability\n\nData used in this research is available in the Gene Expression Omnibus (GEO) database:\n\nGene Expression Omnibus: Radioresistant tumor response to interferons. Accession number: GDS3126. https://identifiers.org/geo:GDS3126. 115\n\nGene Expression Omnibus: Early onset colorectal cancer: normal-appearing colonic mucosa. Accession number: GSE4107. https://identifiers.org/geo:GSE4107. 116\n\nGene Expression Omnibus: Breast cancer relapse free survival. 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Publisher Full Text\n\nManzat Saplacan RM, Balacescu L, Gherman C, et al.: The role of PDGFs and PDGFRs in colorectal cancer. Mediat. Inflamm. 2017; 2017: 1–9. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLin Q, Lai R, Chirieac LR, et al.: Constitutive activation of JAK3/STAT3 in colon carcinoma tumors and cell lines: inhibition of JAK3/STAT3 signaling induces apoptosis and cell cycle arrest of colon carcinoma cells. Am. J. Pathol. 2005; 167(4): 969–980. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGoodsell DS: The molecular perspective: the ras oncogene. Oncologist. 1999; 4(3): 263–264. Publisher Full Text\n\nEckert LB, Repasky GA, Ülkü AS, et al.: Involvement of Ras activation in human breast cancer cell signaling, invasion, and anoikis. Cancer Res. 2004; 64(13): 4585–4592. PubMed Abstract \| Publisher Full Text\n\nMiller MA, Zachary JF: Mechanisms and morphology of cellular injury, adaptation, and death. Pathologic Basis of Veterinary Disease. 6th ed.2017; pp. 2–43. Publisher Full Text\n\nKontomanolis EN, Kalagasidou S, Pouliliou S, et al.: The Notch Pathway in Breast Cancer Progression. Sci. World J. 2018; 2018: 1–11. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSeif F, Khoshmirsafa M, Aazami H, et al.: The role of JAK-STAT signaling pathway and its regulators in the fate of T helper cells. Cell Commun. Signal. 2017; 15(1): 1–13. Publisher Full Text\n\nNascimento AS, Peres LL, Fari AV, et al.: Phosphoproteome profiling reveals critical role of JAK-STAT signaling in maintaining chemoresistance in breast cancer. Oncotarget. 2017; 8(70): 114756–114768. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYamada M, Monden T, Konaka S, et al.: Assignment of human thyrotropin-releasing hormone (TRH) receptor gene to chromosome 8. Somat. Cell Mol. Genet. 1993; 19(6): 577–580. PubMed Abstract \| Publisher Full Text\n\nPage F, Bishop W: Recurrent Carcinoma Of The Female Breast Entirely Disappearing Under The Persistent Use Of Thyroid Extract Continued For Eighteen Months. Lancet. 1898; 151(3900): 1460–1461. Publisher Full Text\n\nAlinejad V, Dolati S, Motallebnezhad M, et al.: The role of IL17B-IL17RB signaling pathway in breast cancer. Biomed. Pharmacother. 2017; 88: 795–803. PubMed Abstract \| Publisher Full Text\n\nChalabi N, Satih S, Delort L, et al.: Expression profiling by whole-genome microarray hybridization reveals differential gene expression in breast cancer cell lines after lycopene exposure. Biochimica et Biophysica Acta (BBA)-Gene Structure and Expression. 2007; 1769(2): 124–130. PubMed Abstract \| Publisher Full Text\n\nZhang S, Liu J, Xu K, et al.: Notch signaling via regulation of RB and p AKT but not PIK3CG contributes to MIA PaCa 2 cell growth and migration to affect pancreatic carcinogenesis. Oncol. Lett. 2018; 15(2): 2105–2110. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nParr C, Watkins G, Jiang WG: The possible correlation of Notch-1 and Notch-2 with clinical outcome and tumor clinicopathological parameters in human breast cancer. Int. J. Mol. Med. 2004; 14(5): 779–786. PubMed Abstract\n\nDou XW, Liang YK, Lin HY, et al.: Notch3 Maintains Luminal Phenotype and Suppresses Tumorigenesis and Metastasis of Breast Cancer via Trans-Activating Estrogen Receptor-α. Theranostics. 2017; 7(16): 4041–4056. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFu YP, Edvardsen H, Kaushiva A, et al.: NOTCH2 in breast cancer: association of SNP rs11249433 with gene expression in ER-positive breast tumors without TP53 mutations. Mol. Cancer. 2010; 9(1): 113. Publisher Full Text\n\nWang JW, Wei XL, Dou XW, et al.: The association between Notch4 expression, and clinicopathological characteristics and clinical outcomes in patients with breast cancer. Oncol. Lett. 2018; 15(6): 8749–8755. 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GEO DataSet Browser.n.d.Reference Source\n\nn.d.. http\n\nn.d.. http\n\nn.d.. http" }	[ { "id": "221362", "date": "20 Dec 2023", "name": "Suhaila Zainudin", "expertise": [ "Reviewer Expertise bioinformatics", "data science" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe paper introduces an enhanced version of the Differential Expression Analysis for Pathways (DEAP) method, termed iDEAP, for improved identification of perturbed sub-pathways in biological reactions related to complex diseases. The method incorporates a search algorithm adapted from the Detect Module from Seed Protein (DMSP) algorithm and considers the relationship among sub-pathways to enhance the identification of differentially expressed pathways. The study applies three gene expression datasets and demonstrates that the proposed method outperforms previous approaches. In particular, when assessing identified genes for cancer classification through 10-fold cross-validation, the improved method shows higher accuracy for colorectal cancer (90%) and breast cancer (94%). The findings suggest that the iDEAP method effectively identifies informative genes associated with the targeted phenotype, and a biological validation of the top five significant pathways and selected genes supports its utility in disease detection. The paper concludes by highlighting the potential impact of computational biology on biomedical research and medicine in the future.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "232310", "date": "01 Feb 2024", "name": "Nora Muda", "expertise": [ "Reviewer Expertise Applied statistics", "mathematical statistics", "computational biology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nDear Authors,\nI have read your paper and would like to thank you all for your kind attention.\n\nThis manuscript is about the analysis of pathways, primarily focused on sub-pathway-based analysis. The authors proposed an extended method of DEAP by improving the identification of perturbed sub-pathways that achieves higher performance in the detection of differentially expressed pathways.\nThe comments that need to be highlighted in the manuscript are as follows: 1. Since the proposed statistics calculation is based on maximum order statistics, do you rank your maximum score before calculating it using the proposed method? 2. Is there any statistical theorem of maximum order statistics you are applying to your proposed method? It is good to provide it in your manuscript. 3. What is 'p' stand for in Equation 4? 4. Did the 'n' in Eq. 3 and Eq. 4 have the same value? 5. You are calculating twice the average of the score, one in Eq. 3 and another in Eq. 4. I am not very clear on the difference between these two methods. 6. Has any comparison been made with other previous methods? So that we can see your proposed method is better than the previous one.\nThe above comments should, at a minimum, be checked and taken into consideration.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1433
https://f1000research.com/articles/12-476/v1	09 May 23	{ "type": "Case Report", "title": "Case Report: A very rare case of a pleural effusion revealing multiple myeloma", "authors": [ "Selsabil Daboussi", "Asma Saidane", "Samira Mhamdi", "Marwa Kacem", "Samia Essbaa", "Chiraz Aichaouia", "Hela Ghedira", "Faten Gargouri", "Issam Msakni", "Zied Moatemri", "Asma Saidane", "Samira Mhamdi", "Marwa Kacem", "Samia Essbaa", "Chiraz Aichaouia", "Hela Ghedira", "Faten Gargouri", "Issam Msakni", "Zied Moatemri" ], "abstract": "Multiple myeloma is a common malignant bone-based disease. Pleural effusions reported in these patients remain rare. It is commonly due to congestive heart disease, pulmonary embolism, nephrotic syndrome or a second neoplasia. The true myelomatous pleural effusion resulting from a direct tumoral invasion of the pleural are extremely rare. We report here the case of a massive pleural effusion revealing multiple myeloma in a 71-year-old patient. The chest ultrasound showed a massive pleural effusion in the left side with a multinodular thickening of the pleura. The medical thoracoscopy showed a grape-cluster appearance. The diagnosis was made by pleural guided biopsy revealing abnormal plasma cells with an intense positive CD 138 (plasma cell marker) and MUM1 (multiple myeloma oncogene1) staining with a light kappa chain in the protein electrophoresis associated with a myeloma. Unfortunately, our patient died one month after the initial diagnosis. We present also a review of the recent literature in order to highlight the clinical presentations of the myelomatous pleural effusion, the diagnostic tools, the therapeutic strategies as well as the outcomes.", "keywords": [ "Myeloma", "pleura", "malignant", "thoracoscopy" ], "content": "Introduction\n\nMultiple myeloma is a common malignant disease due to a proliferation of an abnormal clone of plasma cells associated with a monoclonal protein or a light chain in the serum or in the urine.1 However, the myelomatous pleural invasion is very rare. We report a case of a massive left-sided myelomatous pleural effusion revealing the disease. The diagnosis was made by the identification of abnormal cells with an intense positive staining to CD138 (plasma cell marker) and a kappa light chain in the protein electrophoresis. We also present a review of the current literature in order to highlight the clinical presentation, the diagnostic tools, the therapeutic approaches and the outcomes.\n\n\nCase report\n\nA 71-year-old woman was admitted in our Department of Pneumology in November 2022 for acute respiratory failure. She complained of dyspnoea occurring at the slightest effort (III NYHA) associated with an important deterioration of her general status (weight loss, asthenia, anorexia). She has previously been treated for cardiac disease (Plavix 75 mg per day, Aspegic 100 mg/day, Statinor 80 mg per day, Sotalol 160 mg per day), atrial fibrillation (Cordarone 200 mg per day (5 days per week) with a curative dose anticoagulant treatment (Rivaroxaban 20 mg/day)) and for psychiatric depressive disorders (with treatment interruption). She is a house-wife and is Caucasian. With regards her habits, she does not smoke and does not drink alcohol. There was no significant past family medical history nor environmental exposure, especially to asbestos.\n\nPhysical examination found a deteriorated general status (performance status = 3). She was afebrile. Her pulse rate was 78 ppm. Her blood pressure was 130/70 mmHg. Her respiratory rate was 24 cpm. She did not display any sign of respiratory distress. The breath sounds were abolished in the left side. The oxygen desaturation was (89%) on room air. The electrocardiogram was normal.\n\nLab tests showed hypochromic microcytic anaemia (Hb level = 9.9 g/dl) and hypercalcemia (calcium level = 2.96 mmol/l).\n\nThe chest X-ray showed left pleural opacity with signs of compression (Figure 1). So, an exploratory and evacuating ultrasound-guided pleural puncture was immediately performed. The thoracic ultrasound showed a massive anechoic, free left pleural effusion associated with pleural nodules (Figure 2). Analysis of the pleural fluid showed a serohaematic exudative fluid with a predominantly lymphocyte formula (80%). A Gram stain fast bacilli (AFB) stain and bacterial and tuberculosis cultures for were all negative. Therefore, a malignant origin was suspected. The chest CT-scan revealed a left-sided malignant pleural effusion associated with mediastinal adenopathy, extended secondary bone lesions and subcutaneous lesions (Figure 3). Additionally, the echocardiography was normal.\n\nA medical thoracoscopy was performed 9 days after her admission in our department. It showed a multinodular pleura with a “bunch of grapes” sign (Figure 4). It allowed guided pleural biopsies as well as chemical pleurodesis in order to prevent the pleural effusion recurrence.\n\nFurthermore, the patient reported an acute neck pain associated with tetraparesis 2 weeks later, during her hospital stay. A spinal MRI showed a tumoral process infiltrating the axis resulting in a spinal cord compression. There was not any sign of spinal suffering. The MRI also revealed diffuse bone involvement with some nodular lesions. So, emergency decompressive radiotherapy was performed as a salvage therapy with immobilization of the cervical spine.\n\nThe histological examination of the pleural biopsy showed abnormal round tumoral cells (Figure 5). Immunochemistry revealed an intense and diffuse positive staining for CD138 (plasma cell marker) and MUM1 (multiple myeloma oncogene 1) with a kappa light chain (Figure 6) suggesting a myelomatous pleural effusion (MPE). The serum protein electrophoresis showed a monoclonal peak at the kappa chain gammaglobulin region. A 24-hour proteinuria was negative. Additionally, the electrophoresis and immunofixation of urinary proteins was normal.\n\nThe diagnosis of myelomatous pleural effusion was made given: the presence of abnormal plasma cells with a positive staining CD 138 and MUM1 in the pleural biopsy, the presence of a kappa-light chain in the protein electrophoresis and the presence of a multiple myeloma confirmed histologically with the presence of the CRAB criteria (hypercalcemia, anaemia, presence of several osteolytic lesions on imaging). Unfortunately, our patient died one month after the initial diagnosis because of a fatal progression of her disease.\n\n\nDiscussion\n\nMultiple myeloma is a bone-based disease, first described in Egyptian mummies. The median age of occurrence is 69 years and it occurs mainly in men.2 It can invade many organs such as the chest, the bones, and the skin. However, the pleural effusions reported in these patients remain rare (6%).3 Moreover, the true myelomatous pleural effusion (MPE) is extremely rare. It occurs in less than (1%) of cases. It is more common in IgG Myeloma types (40%).4\n\nThe diagnosis may be challenging because the symptoms are not specific. In fact, the patients can present with a wide spectrum of clinical features. They report typically hematologic symptoms due to the bone marrow invasion (anaemia, bleeding, infections) and bone lesions (hypercalcemia). They may also report symptoms due to the infiltration of the surrounding organs.5,6 Our patient complained of dyspnoea on exertion associated with an important deterioration of the general status.\n\nPleural effusion associated with multiple myeloma may be due to a congestive heart failure. It is often seen in secondary amyloidosis. It may result either from hyperviscosity or myocarditis, commonly seen in these patients especially after the treatment onset.1,7 Pulmonary embolism should be also considered because of the advanced age of the patients, the comorbidities and the current neoplasia.8 Third, the nephrotic syndrome and the chronic renal failure may be responsible of a concomitant pleural effusion.9 Pulmonary embolism was ruled out by the chest CT scan in our case. The kidney function and 24-hour proteinuria were normal. The pleural effusion may result from a second concomitant neoplasia especially a carcinoma (breast cancer in women, lung cancer in men) or a mesothelioma.1,10 This should be considered in the differential diagnosis of a malignant pleural effusion. Our patient was not a smoker and the breast exam was normal.\n\nThe diagnosis was very challenging. In fact, we initially suspected an advanced stage lung carcinoma giving the chest CT scan’s findings, or a mesothelioma due to the multinodular aspect of the pleura in the chest ultrasound and “the grape-cluster” appearance as seen in thoracoscopy. Histology helped us to assess the right diagnosis.\n\nIt is worth mentioning that true myelomatous pleural effusion is extremely rare. It may result either from: a direct extension from an adjacent skeletal or lung lesions; a direct tumoral invasion of the pleura; a lymphatic obstruction by mediastinal adenopathy; or it may be due to the presence of a multiple myeloma in the chest wall, as seen in our patient.11,12\n\nThe diagnosis requires the presence of a monoclonal protein or a light chain in the pleural fluid, abnormal plasma cells with an intense staining CD138.13–16 Medical thoracoscopy is actually recommended in the case of such malignant pleural effusion. In fact, it allows pleural guided-biopsy with a good-quality sampling, pleural fluid drainage as well as pleurodesis in order to prevent the recurrences during the follow-up.8,17,18 In our case, medical thoracoscopy showed a multinodular pleura with “a buck of grapes” like aspect. We decided to perform chemical pleurodesis given that most of the reported cases had a high rate of recurrence of the pleural effusion. Flow cytometry may be a useful tool to assess the diagnosis.19\n\nAnother interesting aspect of this case is that the pleural effusion revealed the disease. It is well known that the pleural effusion is a late manifestation during the natural history of the multiple myeloma associated with a poor prognosis.20 The median survival does not exceed four months according to literature, despite an aggressive therapeutic approach.21\n\nTreatment is based on a combination of chemotherapy (Adriamycin, vincristine, doxorubicin …) or palliative radiotherapy as a salvage therapy. A new drug has been used (Bortezomib) which is a proteasome inhibitor with good results.22 Stem cell transplantation can be also considered.23 Unfortunately, the patient’s response is often transient with a high rate of tumoral recurrence.11,24\n\n\nConclusion\n\nTo conclude, we report a very rare case of a massive pleural effusion revealing multiple myeloma. This should be considered in the differential diagnosis of a malignant pleurisy. Medical thoracoscopy is a mainstream exam in order to assess the diagnosis and to prevent the recurrence. Further studies using flow cytometry and cytogenetic analysis are required. The new therapeutic approaches using target therapy drugs seem to be very interesting pathways.\n\n\nConsent\n\nWritten informed consent for publication of their clinical details and images was obtained from the family of the patient.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\n\nReferences\n\nAl-Farsi K, Al-Haddabi I, Al-Riyami N, et al.: Myelomatous Pleural Effusion Case report and review of the literature. Sultan Qaboos Univ. Med. J. 2011; 11(2): 259–264. PubMed Abstract\n\nBabu GK, Saldanha SC, Lokesh KN, et al.: Myelomatous pleural effusion: A rare case entity reported from a tertiary care cancer center in South India. Lung India. 2017; 34(2): 176–178. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRodelo A, Joshi J: Myelomatous Pleural Effusion as a Presenting Symptom: A Case Report. Cureus. 2019; 11(7): e5153. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKim YJ, Kim SJ, Min K, et al.: Multiple Myeloma with Myelomatous Pleural Effusion: A Case Report and Review of the Literature. Acta Haematol. 2008; 120: 108–111. PubMed Abstract \| Publisher Full Text\n\nSchelle M, Schreiber C, Knolle J, et al.: Pulmonale und pleurale Manifestationen beim Plasmozytom Pulmonary and Pleural Manifestations of Multiple Myeloma. Pneumologie. 2006; 60: 743–748. Publisher Full Text\n\nShah D, Besa EC: Multiple Myeloma Clinical Presentation. Medscape. 2023. Reference Source\n\nXu X, Shen YH, Shen Q, et al.: A case of bilateral pleural effusion as the first sign of multiple myeloma. Eur. J. Med. Res. 2013; 18: 7. PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source\n\nAlshati MH, Kumar R, Kannan S: Dyspnea, massive effusion and lytic rib lesion as initial presentation of multiple myeloma in a young man. Can. Respir. J. 2013; 20(4): 253–255. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHuang TC, Chao TY: Myelomatous pleural effusion. Q. J. Med. 2010; 103: 705–706. Advance Access Publication 20 October. Publisher Full Text\n\nBen GI, Ksontini I, Houman MH, et al.: Pleurésie myélomateuse bilatérale: à propos d’une observation. Rev Méd Interne. 2001; 22: 1134–1135. Publisher Full Text\n\nKim YM, Lee KK, Oh HS, et al.: Myelomatous effusion with poor response to chemotherapy. J. Korean Med. Sci. 2000; 15(2): 243–246. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMalhotra KP, Agrawal V, Prasad N: Myelomatous pleural effusion: A diagnostic challenge. Indian J. Cancer. 2010; 47(3): 351–352. PubMed Abstract \| Publisher Full Text\n\nDeshpande AH, Munshi MM: Pleural effusion as an initial manifestation of multiple myeloma. Acta Cytol. 2000; 44: 103–104. PubMed Abstract\n\nKintzer JS Jr, Rosenow EC 3rd, Kyle RA: Thoracic and pulmonary abnormalities in multiple myeloma. A review of cases. Arch. Intern. Med. 1978; 138: 727–730. PubMed Abstract \| Publisher Full Text\n\nRodríguez JN, Pereira A, Martínez JC, et al.: Pleural effusion in multiple myeloma. Chest. 1994; 105: 622–624. Publisher Full Text\n\nKamble R, Wilson CS, Fassas A, et al.: Malignant pleural effusion of multiple myeloma: Prognostic factors and outcome. Leuk. Lymphoma. 2005; 46: 1137–1142. Publisher Full Text\n\nKuwal A, Advani M, Kumari J, et al.: Medical Thoracoscopic Assessment of Pleural Abnormalities in Patients With Malignant Pleural Effusion: A Retrospective Analysis. Int. J. Med. Rep. Prof. 2020; 6(3): 147–150. Publisher Full Text\n\nWu YB, Xu LL, Wang XJ, et al.: Diagnostic value of medical thoracoscopy in malignant pleural effusion. BMC Pulm. Med. 2017; 17: 109. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWang Z, Xia G, Lan L, et al.: Pleural Effusion in Multiple Myeloma. Intern. Med. 2016; 55: 339–345. Publisher Full Text\n\nCho YU, Chi HS, Park CJ, et al.: Myelomatous Pleural Effusion: A Case Series in a Single Institution and Literature Review. Korean J. Lab. Med. 2011; 31: 225–230. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZhong Y, Zhang J, Wang H: Myelomatous pleural effusion involvement in 23 patients with multiple myeloma: A single-center clinical analysis. Thoracic Cancer. 2015; 6: 359–362. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRamos AL, Trindade M, Pinto AS, et al.: Pleural effusion and multiple myeloma - more than meets the eye: A case report. Mol. Clin. Oncol. 2021; 15: 238. Publisher Full Text\n\nWoo WH, Ithnin A, Raffali MA, et al.: Recurrent pleural effusion in myeloma. Oxf. Med. Case Reports. 2022; 2022: 8. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSuresh A, Singh VP, Sundar S, et al.: Malignant myelomatous pleural effusion with good response to combination chemotherapy. J. Assoc. Physicians India. 2007; 55: 595–596." }	[ { "id": "202487", "date": "07 Sep 2023", "name": "Junn-Liang Chang", "expertise": [ "Reviewer Expertise Human cancers and pathology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nReviewer’s comments to the authors:\nThe authors should present laboratory assays including cell blood count, PB smear analysis, and blood chemistry values, serum electrophoresis, and serum immune electrophoresis.\n\nDoes pleural effusion cytology show abnormal proliferation of plasma cells?\n\nHas the patient had bone marrow aspiration analysis results?\n\nDoes the imaging test include skull, pelvis, and skeletal bones, and is there any abnormality?\n\nIs the background of the case’s history and progression described in sufficient detail? Partly\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Partly\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Yes\n\nIs the case presented with sufficient detail to be useful for other practitioners? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-476
https://f1000research.com/articles/11-989/v1	01 Sep 22	{ "type": "Case Study", "title": "Brain-to-brain communication during musical improvisation: a performance case study", "authors": [ "Mauricio A. Ramírez-Moreno", "Jesús G. Cruz-Garza", "Akanksha Acharya", "Girija Chatufale", "Woody Witt", "Dan Gelok", "Guillermo Reza", "José L. Contreras-Vidal", "Jesús G. Cruz-Garza", "Akanksha Acharya", "Girija Chatufale", "Woody Witt", "Dan Gelok", "Guillermo Reza", "José L. Contreras-Vidal" ], "abstract": "Understanding and predicting others' actions in ecological settings is an important research goal in social neuroscience. Here, we deployed a mobile brain-body imaging (MoBI) methodology to analyze inter-brain communication between professional musicians during a live jazz performance. Specifically, bispectral analysis was conducted to assess the synchronization of scalp electroencephalographic (EEG) signals from three expert musicians during a three-part 45 minute jazz performance, during which a new musician joined every five minutes. The bispectrum was estimated for all musician dyads, electrode combinations, and five frequency bands. The results showed higher bispectrum in the beta and gamma frequency bands (13-50 Hz) when more musicians performed together, and when they played a musical phrase synchronously. Positive bispectrum amplitude changes were found approximately three seconds prior to the identified synchronized performance events suggesting preparatory cortical activity predictive of concerted behavioral action. Moreover, a higher amount of synchronized EEG activity, across electrode regions, was observed as more musicians performed, with inter-brain synchronization between the temporal, parietal, and occipital regions the most frequent. Increased synchrony between the musicians' brain activity reflects shared multi-sensory processing and movement intention in a musical improvisation task.", "keywords": [ "Brain on arts", "hyperscanning", "brain-to-brain synchrony", "musical improvisation" ], "content": "Introduction\n\nAdvances in neuroengineering have fostered the development of mobile brain-body imaging (MoBI) technologies and denoising algorithms that allow the acquisition, interpretation, and decoding of brain activity from free-behaving individuals in real settings.1–3 These advances have led to the development of neurofeedback systems, brain-computer interfaces (BCIs) and neuroprostheses.4 These devices provide aid in the treatment of neurological disorders such as Parkinson’s disease, epilepsy and depression,5–7 motor impairments,8 and diminished brain functioning.9 Although all of these systems are extremely helpful to patients and healthy persons, they follow an individualistic, personal-use approach.10\n\nWhile an understanding of an individual’s cognitive function is of utmost importance in the development of neurological treatments, the comprehension of social interactions at a neurological level is also important, as humans are social beings by nature,11 and neurological disorders such as autism spectrum disorders (ASD) can affect social communication and interaction.12 Furthermore, many common daily human activities are carried out in groups, e.g. at school, work, sports, creative art, and leisure.11,13 Thus, research advances in social neuroscience are likely to revolutionize different fields such as entertainment, communication, education, healthcare, and social embedding, among others.14 Recently, researchers have started to explore brain activity from a collective perspective, using a contemporary approach, known as hyperscanning.15\n\nHyperscanning refers to the synchronous recording of brain activity from more than one individual simultaneously, and has been implemented to study dynamic similarities or differences between the brain signals of multiple participants engaged in interactive or social tasks.15 Such an approach holds promise in understanding the nature of cognitive traits during social interactions.15 Recent hyperscanning studies have documented traces of shared cognition, emergent during moments of social interaction, collaboration, competition, and in educational settings.16,17 The study of neural synchrony between individuals provides an insight into human connectedness and may aid in the development of treatments for social cognition disorders such as autism.18 A desired outcome of hyperscanning is the development of neural biomarkers that track in real-time the quality or strength of shared cognitive states such as brain-to-brain communication, shared attention, message conveying, and high engagement during human interactions.\n\nIndeed, recent studies on human interactions have analyzed shared brain dynamics during teamwork tasks,19 and cooperative/competitive interactions.16 It has been reported that neural synchronization increases when participants are interacting in cooperation, and it reduces when they are competing against each other. A hyperscanning study allowed the quantification of the synchronization between brain signals of infants and adults during gaze interactions, showing increased neural coupling during direct eye-contact.20 Neural coupling between humans has also been associated to the degree of mutual pro-sociality, where higher synchronization reflects stronger social relationships,21 and likeliness of interpersonal bonding.22 Considering the aforementioned studies, by analyzing inter-brain activity, hyperscanning offers a quantitative assessment of the strength and quality of different types of social interactions.23\n\nRegarding neural synchrony metrics, among the most common are coherence,17 phase coherence,16 phase locking value (PLV) and phase locking index (PLI),15 Granger causality,20 correlation,21 wavelet transform coherence (WTC),22 graph theory, and partial directed coherence (PDC).23 Bispectrum is another, more recent, metric in hyperscanning literature,19,24 and offers insights on temporal, spatial and spectral levels. The bispectrum of a signal is a high order spectra that reflects the degree of temporal synchronization and phase coupling between two time series at different frequencies.25 The bispectrum offers additional insight when compared to other neural synchrony metrics, as it provides a more complete intuition on phase coupling, resonance, temporal synchronization and non-linear interactions between any analyzed signal pair.25\n\nStudies on intra and inter neural synchrony between pairs of guitarists during musical improvisation have shown dynamical networks that connect different brain regions, depending on the situation and/or expectations, with involvement of the fronto-parietal region, as well as the somatosensory, auditory, and visual cortices.26,27 The analysis of such obtained networks can be used to study the temporal dynamics of these interactions and providing a neurophysiological interpretation of the observed behavior. Considering the rich and complex interchange of cognitive processes necessary during collaborative artistic performances, its study using the hyperscanning approach is a valid approach to explore the shared neural cognitive traces that emerge from these interactions.28,29\n\nCollaborative, free musical production (improvisation) is a complex and rich form of social interaction,30 it has also been described as a continuous process of generation and transformation of musical interaction,31 and offers an interesting object of study for hyperscanning. Similarities between music and language have been observed previously in terms of the social interaction they entail; as described in,32 a jazz improvisation can be interpreted as a conversation, and a good improvisation as a complex, meaningful conversation. Over recent years, there has been a growing interest in the study of improvisational, freely-moving, collaborative musical production in live performance settings.23 Hyperscanning in the musical context can allow the observation of neural traits of dynamic processes. For example the patterns between musicians’ brain activity when performing cooperatively or not, as it has been reported that such actions create differences in their peripersonal space,33 and in the rhythmical alignment of the overall performance.34\n\nThis process of improvised production can be perceived as a creative act of communication: one that is complex, nuanced, and technical, integrating simultaneous cognitive processes together in real time. Musical improvisation involves complex but rapid interactions of several components, including the generation and evaluation of melodic, harmonic, and rhythmic pattern ideas on a fast time-scale within a performance.35 Mobile brain-body (MoBI) imaging provides the tools for analyzing neural patterns in real-time for freely-moving participants,1,2,36 with hyperscanning techniques that provide an experimental approach to assess non-verbal communication in musical performance.28 During an improvised performance, musicians interact with each other, making use of different skills such as creativity,37 emotional expression and perception,30 self-organization,38 memory retrieval and procedural memory,39 and integration of visual and auditory stimuli with complex and precise motor coordination.40,41 Musical improvisation can also be considered as an’on the fly’ composition, one that is temporally ubiquitous, spontaneous, and is not restricted by critique.\n\nIn a theoretical model to study group jazz improvisation, Biasutti and Frezza42 identify the processes that are essential for creative musical improvisation: anticipation, use of repertoire, emotive communication, feedback, and flow. In Wopereis et al.,43 26 expert musicians provided statements about musical improvisation in two 10-min individual brainstorm sessions. The statements resulted in a 7-cluster concept map, with self-regulation as the central concept, and affect, risk-taking, ideal, basic skills, responsivity, and creation, as constituent concepts for improvisational expertise. Specifically for collaborative improvisation, monitoring, feedback, and evaluation must be performed in association with other musicians, with both generative and communicative attentional demands.44 Another study on jazz improvisation remarks that shared intentions emerge on the fly, and their presence fosters acoustic and temporal coordination, as well as improving the quality of the performance, as perceived by the performers and listeners.13\n\nRecently, the predictive coding of music (PCM) model has been introduced to model how listeners form expectations which may be fulfilled or not, through perception, action, emotion and, over time, learning.45 Under this model, musical interaction is guided by mutual reduction of prediction errors, evidenced by alpha-band intrabrain neural synchronization (phase-locking analysis) in a right-lateralized temporoparietal network, with a higher occurrence probability in mutually adapting dyads than in leader-leader dyads.46 These models of music improvisation highlight the centrality of anticipation, self-regulation, generation and evaluation, with feedback and communication in joint performance.\n\nThis article aims to contribute to the understanding of brain-to-brain communication during a creative collaboration between jazz musicians. A jazz performance incorporates each of the five elements of musical improvisation: anticipation, feedback, use of previous repertoire, emotive communication, and coordinated flow.42 Moreover, in a free jazz performance, as described in,47 a continuous process of evaluation is present, where musicians can decide to maintain or change the current theme; initiate or respond to a change; and to adopt, augment, or contrast a given idea.\n\nWhile improvising, musicians can elaborate over (but are not constrained to) a composition’s underlying chord structure48 and theme, with variations that incorporate multiple derivations from instantaneous decisions in real-world practice.49 An important aspect of jazz performance and proficiency lies in the embodied cognition and motor memory.50 However, most neuroimaging studies on musical improvisation have used functional magnetic resonance imaging (fMRI).35,44 Lying down in an fMRI scanner alters spatial and visual perception,51 and restricts body movement, which limits the capability of fMRI studies to observe realistic musical performance given the importance of embodied cognition in the task (due to the continuous retrieval and processing of spatial, auditory, visual and somatosensory information).50 Because mobile electroencephalography (EEG) does not impose movement constraints, and subsequently allows participants to naturally engage in creative production with minimal, instrumentational constraint, it may afford advantages in studying musical improvisation.52,53\n\nThe current study examines the neural correlates of brain-to-brain communication of jazz musicians during collaborative musical improvisation through hyperscanning; and addresses the limited body of knowledge on collaborative musical improvisation in an ecologically-valid production, with freely-moving expert musicians, as they interact in a jazz performance with a live audience. Here, the presence of a live audience is important, as our cohort of musicians are accustomed to them, to the point that they become a relevant part of their performance. An inter-brain synchronization analysis was implemented, by estimating the bispectrum of EEG signals between musician pairs during collaborative improvisations as they performed for a live audience. Following the concept of ecological validity, the exquisite corpse method was adopted to obtain realistic collaborative improvised art pieces.36,52 The exquisite corpse is a game played by surrealists, in which different artists integrate their contributions into a unique piece, taking turns to add their input in an iterative manner until completing a final piece with the contributions of all members.36 Under this paradigm, the complete performance is formed by a multi-participant improvisational, free jazz piece formed by the creativity from each player.\n\n\nMethods\n\nThe experimental methods were approved by the Institutional Review Board of the University of Houston, and are in accordance with the Declaration of Helsinki. All participants provided written informed consent, including agreement for publication in online open-access publication of information obtained during the experiments such as data, images, audio, and video. Three male healthy adults (P1, P2 and P3) volunteered for this study. Musicians P2 and P3 received (formal) musical instruction for 12 and 6 years, respectively, and P1 (informal) for 6 years. To the date of the experiments, P1, P2 and P3 had 31, 38, and 26 years of experience performing music, respectively. P2 and P3 were music educators at the University of Houston at the time of the experiment. The musicians performed jazz improvisation in a public event at the Student Center of the University of Houston while wearing the MoBI technology. Musicians P1 and P2 have a jazz musical background, whereas P3 had a’classical music’ education. Musician P1 played the drums, musician P2 played the saxophone, and P3 played using a soprano saxophone. P1 and P2 had performed jazz regularly together for 6 years, P2 and P3 had performed a concert together once before, and P1 and P3 had not performed together previously.\n\nHigh-density scalp EEG and electrooculography (EOG) recordings were obtained simultaneously for the three musicians during their musical performances. EEG data was wirelessly acquired using the 64 channel actiCAP (BP gel) electrodes along with the Brain Amp DC amplifer (actiCap system, Brain Products GmbH, Germany) at a sampling frequency of 1000 Hz. Electrode distribution follows the 10-20 international system. EEG data was online referenced to channel FCz on the superior region of the scalp. Four channels were used to record EOG data. Channels TP9 and TP10 were placed on the right and left temples, respectively, to record horizontal eye movement, whereas channels PO9 and PO10 were placed above and below the right eye, respectively, to record vertical eye movement. Impedance was set to less than 25 kΩ for all electrodes before starting the experiments.\n\nPerformances were recorded by three video cameras coupled with a Zoom H6 (https://zoomcorp.com/) audio recorder from a frontal, superior and lateral perspective. Audio was recorded in a single stereo file at 44100 Hz. Three Sterling ST31 FET condenser microphones (https://sterlingaudio.net/) were used to amplify the sound from each musician’s instrument during the live performance.\n\nMusicians performed three 15 minutes improvisations (trials). Each trial was divided in three 5 minutes pieces (segments). In Segment 1, one musician was performing while the other two were listening. In Segment 2, a different musician joined the first, while the remaining musician listened to the performance of the other two. In Segment 3, the third musician joined and all participants performed together until the end of the trial. In a given segment, the musicians who are performing are referred to as the “active” musicians, whereas the musicians who are not performing are the “passive” musicians. At the beginning and at the end of the experiment, three blocks comprising of an EEG impedance check, a one-minute eyes open (EO) and a one-minute eyes closed (EC) were recorded.\n\nIn each trial, the order of the musicians joining at each segment was pseudo-randomized so that each musician entered one trial as the first, second or third player. Each musician was given a visual cue to signal their start time in the piece. Between one trial and the next, there were short pauses of 3-5 minutes, in which the audience clapped and the musicians prepared for the next trial.\n\nFigure 1(a) shows the protocol for baseline measurements, and the order of musicians joining at each segment and trial. Figure 1(b) shows the locations of EEG electrodes with impedance higher than 25 kΩ at the start and at the end of the recordings, for all musicians. Figure 1(c) shows the setup of the instruments and microphones during the experiments, and the three musicians wearing the EEG caps. Figure 1(d) depicts, as an example, from top to bottom, the recorded raw EOG, EEG and audio signals obtained for the first five seconds of Segment 1 of Trial 1, when only P3 is performing.\n\nThree independent raters with training in music composition annotated the data. The annotators were not familiar with the researchers nor with the musicians involved in the performances; and were tasked to write annotations of the performance independently from each other, by watching a recording of the live performance. The annotators were asked to write a short description (e.g. “players are performing in sync”, “mirroring each other”, “performing in discord”), and the time each event happened. Table 1 shows sample descriptions from the annotators during one trial of musical improvisation.\n\nOnly synchronized performance (SP) and desynchronized performance (DP) events are presented.\n\nAt the beginning of each trial, video, audio and physiological signals were synchronized using manual event markers (i.e. pressing a button). Recordings from the three trials were obtained simultaneously using this procedure. Unfortunately, data transmission was interrupted from 4:25-5:25 of Trial 3 due to a loss in connection, which resulted in missing data. The events that happened in this period were therefore not included in the analyses.\n\nEEG signals were acquired at 1000 Hz and resampled to 250 Hz to reduce computational cost in subsequent calculations. Signals were bandpass filtered from 0.1 to 100 Hz using a 4th order Butterworth filter to remove unwanted noise. The PREP pipeline from the EEGLAB package (https://sccn.ucsd.edu/eeglab/download.php) was used as the initial step to clean the data.54 This procedure ensures the removal of power line noise, as well as a “true” average reference of the signals. EOG artifacts were removed from the EEG signals using an adaptive noise cancelling (ANC) framework, known as H∞ filter.55 Raw EOG signals were used as input in the H∞ filter with parameters γ = 1.15 and q = 1e−10 for removal of eye blinks, eye motions, amplitude drifts and recording biases simultaneously. The obtained signals were further processed using the artifact subspace reconstruction algorithm (ASR) included in the EEGLAB package.56 The ASR algorithm calculates the standard deviation of a “clean” portion of the signals in the principal component analysis (PCA) subspace, and reconstructs artifacts with standard deviations as κ times higher than in the clean portion. Here, a value of κ = 15 was chosen to remove remnants of eye movement and muscle artifacts. According to,57 κ values between 10-20 are recommended to reduce artifacts and at the same time preserve the content of EEG signals. As a final step, independent component analysis (ICA) was performed on the data and suspicious components (eye, muscle, electrode popping) were removed before projecting back the signals. A graphical representation of the pre-processing steps is presented in Figure 2, as well as feature extraction and subsequent signals analysis.\n\nEach step is described in detail in the Methods section.\n\nThe improvisational nature of the performance allowed for the examination of the musical communication between the musicians as the piece progressed. At times, they built from the theme established, returned to the main theme, or proposed new ideas. With the annotations from three independent raters, we clustered sections of the performance where the participants performed synchronously or not as synchronized performance (SP), and desynchronized performance (DP). SP included moments of in-time synchronized execution, as well as improvisation and interactions under the same underlying pulse; while DP reflected moments where musicians traded material, though not aligned to the same underlying pulse, and with no coordination (as well as deviation from the current theme). A temporal (across-time) analysis was performed to observe neural synchronization in those moments of SP and DP; and these times were evaluated across three participant conditions: passive-passive, when no musicians in a dyad were performing; passive-active, when one musician in a dyad was performing; or active-active, when the two musicians in a dyad were performing. For the sake of the ecological validity approach, rather than manipulating experimental conditions (e.g. rest vs improvisation), we observed brain synchrony across SP and DP, and the participant conditions posed by the exquisite corpse approach.\n\nThe quantitative measurement of brain-to-brain communication was achieved by calculating the bispectrum between musician dyads of EEG data obtained during improvised musical performance at different stages of interactive performance. As referred to in previous works, higher bispectrum magnitudes at given pairs of frequencies reflect non-random interactions, phase coupling,19 and non-linear multi-frequency interactions,58 which have been observed as traces of inter-brain synchrony during teamwork interactions.19,24\n\nThe denoised EEG signals were used to estimate the bispectrum between all possible channel combinations, for all participant pairs, trials and segments. Bispectrum was estimated across the EEG recordings using four-second windows with 75% (one-second) overlap. The bispectrum at each time window was estimated using Equation 1:\n\nBispectrum was estimated at 602 EEG channel combinations between pairs of participants for all segments, and trials. A bispectral representation of a segment was obtained averaging all four-second windows in each segment, for each frequency bin (1-50 Hz). Bispectral representations were normalized to the bispectral representations of the same channel combinations during pre-trial EO task using Equation 2. Pre-trial EO was treated as rest condition, where participants did not communicate with each other.\n\nBispectral values for five frequency bands were obtained as the average of the normalized bispectral representation in the following frequency ranges: delta (1-4 Hz), theta (4-7 Hz), alpha (8-12 Hz), beta (13-29 Hz) and gamma (30-50 Hz).\n\nA temporal bispectrum series was also estimated, using sliding overlapping windows of four-seconds (75%). At each window, the temporal bispectrum values were obtained as the average normalized bispectral representation (Equation 2) at each frequency band, thus obtaining a temporal representation of the EEG signals’ synchronization between musicians. These temporal bispectrum values were estimated for all windows using the channel combinations which were found to be significant in the implemented statistical analysis. The analysis is described in detail in the statistical analysis subsection.\n\nRight-tailed Wilcoxon signed rank tests were used to evaluate statistically significant differences between the average bispectrum at different frequency bands for all channel combinations. Average bispectral representations during rest and specific segments were compared.\n\nThis procedure ensures the discovery of only those channel combinations with significantly higher bispectrum at a specific segment and for a given frequency band as compared to rest. At each Segment, 602 tests were performed (p < 0.05, corrected for multiple comparisons via Bonferroni correction). Statistical tests were performed for all trials (3), segments (3), participant pairs (3) and frequency bands (5), for a total of 486, 000 tests. A different amount of samples was used for each frequency band, due to bandwidth difference; 16 for delta and theta, 20 for alpha, 68 for beta and 82 for gamma.\n\nThrough this procedure, the identified channel combinations were used as representative traces of brain-to-brain communication during musical improvisation. To further explore the behaviour of such traces, temporal and spatial analyses were implemented.\n\nThe temporal analysis of bispectrum was implemented in representative bispectrum traces to observe its dynamics under different conditions during the performance. The bispectrum traces used in this analysis were those of the most significant channel combination (in the gamma band as described in the Results section) at the third segment of each trial, for each participant pair.\n\nThe bispectrum analysis was divided into two groups of events of naturally occurring experimental conditions: SP and DP, as labelled by the annotators in the audience. For each event, a two-minutes representative bispectrum trace in a time period (-60 to 60 s) was obtained per dyad. For each specific event, the time of the annotation was considered as the 0 s mark. To observe relative differences between SP and DP, the bispectrum traces were baseline corrected at each event for both groups. To obtain baseline corrected traces, average bispectrum in the (-60 to 0 s) period was obtained and substracted from each two-minute bispectrum trace.\n\nBaseline corrected bispectrum traces were obtained for all events, trials and participant pairs, and were grouped and compared between the two groups. Wilcoxon signed rank tests were used to find statistically significant differences (p < 0.05) at every (-60 to 60 s) time point, between SP and DP at each performance condition. This analysis was implemented independently for events in the passive-passive, passive-active and active-active performance.\n\nSpatial analysis was implemented to identify regions of interest (ROIs) involved in musical performance. The selected ROIs group spatially close electrodes in 13 regions: anterior frontal (AF), left fronto-central (LFC), midline fronto-central (MFC), right fronto-central (RFC), left centro-parietal (LCP), midline centro-parietal (MCP), right centro-parietal (RCP), left parieto-occipital (LPO), middle parieto-occipital (MPO), right parieto-occipital (RPO), left temporal (LT), right temporal (RT) and occipital (O).60 Figure 7 shows the location for the 13 ROIs within the scalp map. The significant channel combinations identified through the statistical analysis at every segment and trial were grouped for the different performance conditions: passive-passive, passive-active and active-active.\n\n\nResults\n\nA general representation of the bispectral dynamics between pairs of participants during Trial 1 in shown in Figure 3. Here, normalized bispectrum is presented for the three participant pairs (P12, P13 and P23) in separate insets. In each inset, four plots are shown: a bispectrogram (top left), the average bispectrum at each time window in the gamma band (bottom left), the average bispectrum at each frequency bin (top right) and the most significant channel combination found at gamma band, between each dyad (bottom right).\n\nFor each participant pair, four insets are provided: (1) Average bispectrum in gamma band across 15 minutes of musical improvisation (bottom left); (2) Bispectrogram (1-50 Hz) across 15 minutes at (3) a representative significant channel combination in the gamma band (bottom right); (4) Average bispectrum (1-50 Hz) across the 15 minutes of performance (top right).\n\nThe bispectrogram shows positive values from 0-0.3, which means that bispectrum values were up to 30% higher during musical performance than during rest.86 From the bispectrum representation in frequency it can be observed that in average, higher frequencies show the highest values. This particular behaviour is more evident for P12 and P23. The temporal dynamics of the bispectrum shows oscillations at different moments of the Trial, which correspond to fluctuations in EEG signals synchronization between participants. Across all participant pairs, the average bispectrum in the gamma band tends to increase from the initial segments to the latter ones, where more musicians are performing together. The regions where the highest significant synchronization was found include temporo-occipital (P12), occipito-occipital (P13) and temporo-frontal (P23) connections.\n\nAs Figure 1 shows, at Trial 1, P3 starts performing. At Segment 1, participants P1 and P2 were listening to P3 perform; and the bispectrum of the dyad P12 is lowest. The bispectrum trace of dyad P13 also seems low at Segment 1. The stronger bispectrum is observed for dyad P23. At Segment 2, P1 joins the play and an increase in bispectrum is observed for P13, as both participants are performing together. The bispectral trace of P12 and P23 show slightly higher values towards the end of Segment 2. By Segment 3, P2 joins the other two participants and all are improvising together. Bispectrum increases at all participant pairs are observed during this final Segment, reflecting higher EEG synchronization, when compared to segments where participants are not actively interacting in the performance. These observations were tested statistically for all channel combinations in Figure 4.\n\nLines represent specific channel combinations with significantly higher bispectrum during improvisation than in rest condition (p < 0.05). White and gray heads show the passive and active musicians, respectively. Bars insets show the total significant channel combinations for all participant pairs for alpha, beta and gamma bands at each segment. Statistical tests were corrected for multiple comparisons via Bonferroni correction.\n\nThe procedure of the statistical tests presented in the Statistical analysis subsection was implemented for all Segments, Trials, frequency bands and participant pairs. No significant channel combinations were found for the delta and theta band for any segment. Most statistically significant channel combinations were found for the beta and gamma bands, and a few in the alpha band.\n\nFigure 4(a) shows the total significant channel combinations (between all dyads) when different musicians were performing together, and Figure 4(b) shows a topographical representation of the statistical analyses.\n\nIn Figure 4(a), the dyads P12 and P23 show consistently more significant inter-brain synchronized channels that for the P13 dyad. The three dyads showed few synchronized channels when musician P3 performed alone, and when P1 performed together with P3.\n\nThe most significant channel combinations (visualizing the top 5 channel pairs) for the alpha, beta and gamma bands are shown for each participant pair. Bar graphs show the amount of significant channel combinations at each frequency band, and dyad. At each specific segment, passive and active musicians are shown as white and gray heads, respectively. Topographical representations are presented for all trials and segments, therefore the first row of Figure 4 corresponds to the data shown in Figure 3.\n\nIn Trial 1 (top row of Figure 4(b)) and Segment 1, P3 starts performing, and only a few significant channel combinations were found for dyad P12 in the gamma band. In Segment 2, P1 joins and more channel combinations are shown in the beta and gamma bands for dyad P12 and P13, who are performing. In Segment 3, when all musicians are performing, an increase in the amount of significant channel combinations is observed for all participant pairs. In this last Segment, a few channel combinations were observed in the alpha band.\n\nIn Trial 2 (middle row of Figure 4(b)), in Segment 1, P1 starts performing. A few channel combinations were found to be significant for all participants in the beta and gamma bands. In Segment 2, P2 joins and an increase in the amount of significant channel combinations is observed for dyads P12 and P23). In the Segment 3, P3 joins and a decrease in the amount of significant channel combinations is observed across all participants).\n\nIn Trial 3 (bottom row of Figure 4(b)), P2 starts performing, and significant channel combinations for P12 and P23 are observed for beta and gamma, and only one for P13. In Segment 2, P3 joins and a similar connection pattern is observed between participants at Segment 1. In Segment 3, P1 joins and a considerable increase in significant channel combinations is observed for both P12 and P23, while those for P23 remain similar.\n\nSome general patterns were observed through this analysis. It was observed that the amount of significant channel combinations increased as more musicians joined the performance, which can be observed in Figure 4(a). Dyad P13 showed less amount of significant channel combinations throughout the experiment, at different segments and trials 4 (a). Also, in all segments, the amount of significant channel combinations was higher for the gamma band than for beta or alpha bands. Finally, the most common interconnected regions across segments and trials are those involving the temporal, occipital and parietal regions.\n\nFigure 5 shows the normalized bispectrum trace for the 15 minutes of Trial 1, using the significant channel combinations described in the temporal analysis subsection. The vertical dashed lines mark the division between segments, and bars are used to visualize the moments during the performance when the experts identified either an SP or DP event. The volume of the recorded audio file from the performance is shown below the bispectrum traces. The individual events shown in Figure 5 are described in Table 1. The corresponding Figures and Tables for Trials 2 and 3 are presented in the Extended data, in Figures S1 and S2, and Tables S1 and S2, respectively. Representative SP and DP events from Trials 1-3 are presented in Videoclips S1-S3 in the Extended data.86\n\nVertical dashed lines represent the times when a new musician joined the performance. Vertical bars represent time of synchronized performance (SP) or desynchronized performance (DP) events, as labelled by experts. A representation of the selected channels for each dyad is shown in the bottom right corner. The annotations from the events are shown in Table 1.\n\nFigure 6(a) and (b) show the average bispectrum change across participant pairs for the passive-active and active-active conditions, respectively, for both SP and DP. The 0 seconds vertical dotted line in Figure 6 indicates the start of the event: either SP or DP. The amount of averaged traces for each condition were, 24 and 31 for SP; and 14 and 26 for DP; for the conditions passive-active and active-active, respectively. Figures c) and d) show every individual trace analyzed in a) and b), respectively.\n\nBS change (%) was obtained applying baseline correction on each trace by using the 60s previous to each event. Average BS changes (%), and histograms (distribution of all traces) are shown for passive-active (a) and active-active (b) conditions. The amount of averaged traces at a) and b), respectively, are: 24 and 31 during SP (nSP); 14 and 26 during DP (nDP). Individual BS (%) changes for all nSP and nDP events are presented in c) and d) for the passive-active and active-active conditions, respectively.\n\nNo statistical significance was observed between SP and DP in the passive-active condition. Bispectrum change was significantly higher during SP than DP in the active-active condition, slightly before the onset of annotated events (− 3 s), as well as 40 s after the onset.\n\nA topographical visualization of the most significant scalp ROIs for participant pair conditions (summarizing the findings of Figure 4), is shown in Figure 7. Visualization maps were plotted to represent the degree of synchronization between and within the evaluated ROIs for all conditions (passive-passive, passive-active and active-active), dyads (3) and trials (3). Figure 7 show this representation in the gamma and beta bands. It can be observed that the most synchronized ROIs are in the active-active condition, while less are observed in the Passive-Active condition and the lesser during passive-passive condition.\n\nShading represents the degree of inter-synchronization within the same (circles) and different (lines) ROIs.\n\n\nDiscussion\n\nThe bispectrum analysis allowed us to obtain a quantitative representation of brain-to-brain communication, by analyzing the temporal synchronization strength (i.e. bispectrum) of EEG signals between musicians during a free jazz improvisation performance. In such performances, musicians continuously engage in a dynamic communication formed by perception, evaluation and action. The presented methods were applied to observe the synchronized interactions of neural activity at different stages of the performance, different recording sites and under five frequency bands.\n\nFollowing our proposed methods, a bispectral representation in the time and frequency domain were obtained for all pairs of possible combinations of the assessed variables (segment, trial, frequency band, and channel). Statistical analysis revealed that the most significant synchronization between EEG signals of paired musicians were found in high frequency bands: beta and gamma, as shown in Figure 4. Also, in the same analysis, it was noted that most of the significant neural synchronization links were formed between the occipital, parietal and left temporal regions. Such results were used to assess the most frequent connections between ROIs across pairs of musicians at different performance conditions (See Figure 7).\n\nAlthough musicians exhibit differences in their brain activity due to individual preferences, domain-specific memory for previously encountered auditory-motor patterns,35,61 as well as the nature of their instruments (e.g. drummers use more spatial and visual processing), common patterns were observed. In this study, the most synchronized ROIs between musicians were found at left temporal, and bilateral parietal and occipital sites (LT, LPO, O, RPO and RT), with increased synchronization in the beta and gamma bands.\n\nThese results have further implications for cross-modal plasticity due to musical training, between individuals. The posterior coupling between musicians can be strengthened through extensive training.62,63 Such processes are present when musicians rhythmically engage in a collaborative, creative work. Two processes give rise to this dynamical sensorimotor integration: motor commands, and sensory predictions that provide feedback regarding the given command.63–65 This feedback loop often informs individuals of errors or mismatches between predicted and real sensory feedback, which results in the reconfiguration of this perception-action cycle.65,66 However, this cycle is not restricted to self-generated action. An increasing body of research suggests that in musical contexts, musicians are able to form multiple action representations, performing real-time integration of perceptual stimuli, motor commands, and outcome predictions for one-self and others.63 This complex, moment-to-moment processing within the perception-action cycle, informed by internal forward models, may be the foundation of inter-personal synchrony in creative, musical contexts.63\n\nNeural synchronization fMRI studies of resting state in musicians have found increased functional connectivity between the auditory and motor cortices within an individual’s brain67 and in default mode network and executive control network.68 fMRI studies have shown long term induced plasticity69 in trained musicians when compared to non-musicians. Improvising jazz musicians experience weaker connectivity in pre-frontal areas during musical improvisation, compared to performing pre-learned segments.70 Studies in the literature resembling our findings regarding gamma band activity in music related processes have been reported; expert musicians exhibit neural synchronization between multiple cortical areas in the gamma band71 and left hemispheric gamma synchrony while listening to music72 while such patterns are not observed for non-musicians. Inter-brain synchronization in the theta/alpha amplitudes between temporal and lateral-parietal regions has also been described during speech-rhythm coordination in.73 The results obtained from the statistical and the ROI analysis suggest that beta and gamma synchronization is present during the performance of higher cognitive tasks that need a dynamic binding of information, such as an improvised collaborative musical performance. In our case study, the presence of higher synchronization between temporal, parietal and occipital sites during improvised musical performance suggests the establishment of functional inter-connections between musicians which reflect shared multi-sensory (visual, auditory, and spatial, respectively) processing, integration, and communication.27 Auditory and visual cues from co-performers have been reported to relate to the strength of inter-musician coordination during musical improvisation.74\n\nThese results show evidence for an inter-musician perception-action cycle, where there is a circular, feedback-based, hierarchical method of information-processing conducted by the interplay between both posterior (i.e. sensory input) and anterior (i.e. motor, executive output) regions of the cortex.66 In this experiment, inter-brain bispectrum analysis showed synchornicity in sensory areas. Cross-modal plasticity, and reinforcement of intra-brain coupling of posterior and anterior areas, has been shown to be enhanced by musical training.63 Experience in joint performance leads to fine-tuning of the internal forward model representation that allows for the prediction of observed or listened actions from fellow musicians with high temporal resolution. Our results suggest that coupling in posterior and temporal regions is associated with such predictions of the actions from other members of the performing group. The musicians generate predictions both about when and what their peers’ new musical idea will occur. Musicians with experience performing together may in fact learn which succession of tones are likely to occur, stemming from regularity from previous performances. This complex, moment-to-moment processing within the perception-action cycle, informed by internal forward models, may be the foundation of inter-personal synchrony in creative, musical contexts.\n\nMusician P1 and P3 performed together for the first time in this experiment, while musician P1 and P2 performed regularly together prior to this study. Thus, it is likely that P1 and P2 had developed strong internal forward models of each other that enabled them to predict and respond to recognized sequences between them, as shown in Figure 4(a). Musician P3 had the lesser prior musical collaboration with the other musicians. This difference in familiarity background supports the finding of a smaller number of synchronized channels between P3 and the other musicians throughout the three trials of the performance.\n\nAcross all trials of the present study, significant bispectrum synchronization was found in posterior (e.g., parietal, temporal and occipital) regions that are involved in the processing of sensory input and are important in interpreting sensory feedback from the external environment. Because musical improvisation is founded in nuanced, interpersonal exchange of motor commands that are generated based on constantly evolving sensory input, these findings support the notion that this musical, creative synchrony between participants is highly dependent on the sensory, perceptual inputs they are receiving from each other, and their surroundings. The output in this cycle (i.e. action) would be represented by activation of anterior (e.g. frontal) regions. In Figure 7, connections involving anterior regions are more present during active-active interactions, when both musicians are performing (producing an action) together, while a predictive component can also be observed in Figure 6, where a significant positive bispectrum change was observed across all analyzed SP events approximately three seconds before the onset of the labelled events. Both components were not present during passive-active and passive-passive interactions, in which action and anticipation are not as needed due to the nature of such conditions.\n\nAnother key variable to address in our study is the temporal dynamics of the bispectrum. In Figure 5, the temporal bispectrum dynamics show a consistent increasing trend, as more musicians joined the performance. Towards the final segments of the performance, there were more’musical voices’ interacting, increasing the complexity of the piece, as well as the stimulation, perception, and engagement. This increase in bispectrum was also observed in Trial 3 for P12 and P13, but not for P23, which presented a decreasing trend (See Figures S1 and S2, Extended data86). Also, in Trial 2, a general bispectrum decrease was observed for all dyads, with a sudden increase at the end of the first segment, where a new musician joined the performance. As mentioned in the Experimental design subsection, P3 is a classical trained musician, while P1 and P2 are professional jazz musicians. It is also important to mention that P1 and P2 often perform together, while P3 is not an acquaintance of them. Brain-to-brain synchrony has been studied between dyads under different social contexts, such as between romantic couples and strangers21,22 and it has been reported that higher neural coupling relate to the degree of social connectedness and mutual pro-sociality. It has also been noted from recent musical improvisation studies that the familiarity between musicians predicts stronger coordination of intentions during the performance.75 Increased neural synchrony between two participating individuals may indicate mutual, efficient, and effective social interaction76 and can be modulated by the degree to which the participating individuals feel socially connected, the activity they are engaging in, and the interaction setting.21,76–78 An interpretation of the aforementioned temporal bispectrum changes is that during bispectrum decreases, participants were not communicating effectively, and the “closeness” between each other had an important role in this communication. It can be observed from the bars in Figure 4 that a lower amount of significant channel combinations were found at the dyads where the unacquainted P3 is present, whereas a higher amount of significant channel combinations are found between the more acquainted musicians P1 and P2. This is also evident in Figure 4(a), where the highest amount of significant channels is presented between P1 and P2, and lower combinations are significant when P3 is involved.\n\nBy analyzing the fluctuations of bispectrum change relative to the stimuli type and onset, differences in bispectrum dynamics were observed whether musicians were performing in synchronization or not. This analysis revealed on average higher bispectrum during synchronized performance when compared to desynchronized performance. Higher inter-brain synchrony has been reported in participants performing cooperative tasks and lower synchrony when performing competitive tasks.15 Based on the results of this study, higher bispectrum was observed between participants while performing in synchrony, which might be a reflection of cooperative intention. On the other hand, lower bispectrum values during desynchronized performance might be indicative of a competitive behaviour (e.g. changing the current theme or proposing a new idea). A review on this topic is presented in,16 however it is noted that most papers in this regard are based on an experimental design under controlled laboratory settings. In our study, a real-world scenario is presented, therefore it is best suited to study these types of interactions. An interesting note on this regard is that in a musical improvisation, alignment and misalignment between musicians are both needed to contribute a new perspective on an established theme,49 and continuously propose new musical paths in the piece. Moreover, the observed brain synchrony dynamics are associated to the spontaneous decisions and interactions of the musicians during an unconstrained free jazz improvisation, which does not facilitate the temporal prediction that a steady pulse would cause in the musicians.29\n\nOur results suggest that bispectrum was able to detect relevant temporal and spatial information about musician’s interactions during the performance. Therefore, the proposed method could be used to track the degree of synchronized interactions and can be applied to different contexts. Some of the applications and desired outcomes of research in this field is the development of neural biomarkers that measure in real-time the quality or the strength of shared cognitive states such as: brain-to-brain communication, shared attention, message conveying, and high engagement during human interactions. Two possible applications are the use of such methods to track changes in social interactions in patients suffering from communication disorders, and to enhance learning in educational settings.\n\nWhile the social nature of individuals has been recognized and acknowledged as foundational to human interaction, research regarding the neural inter-brain basis of these interactions naturalistic social settings has only just begun in its investigation.78 Hyperscanning applied to social interactions opens the possibilities to study and enhance social exchanges.15 In a recent study, large groups of museum-goers participating in face-to-face pairs in an artistic neurofeedback installation were found to exhibit higher levels of inter-brain synchronization in low alpha and beta band frequencies, correlated with the pair’s empathy, social closeness, engagement, joint action and eye-contact.79 Observing brain-to-brain synchronization during naturalistic exchanges could not only aide in developing a more comprehensive understanding of its neural underpinnings, but also could shed light on various communication disabilities.76\n\nA bispectral analysis to observe brain-to-brain synchronization during social interactions could be used in the educational field to increase teacher-learner synchronization to enhance learning outcomes and experiences.80 A study examining the effects of brain-to-brain synchronization within a classroom setting was performed on a group of four science students and a teacher. In such study, alpha-band synchrony between students significantly predicted subsequent performance (i.e., memory retention) on both immediate and delayed post-tests.81 Inter-brain synchronization in prefrontal and superior temporal cortices between instructor-learner dyads has been shown to increase when the instructor engages with the learner through guiding questions and hints.82 Such results are also consistent with previous research on the synchrony between speakers and listeners.81,83,84 Brain-to-brain synchronization in a naturalistic musical performance provides a window to assess the perception-action and communicative cognitive processes required during musical improvisation42; and coupled with instructor-learner interactions, inter-brain synchronization metrics can inform effective pedagogical techniques.\n\nThis study faced some limitations given the logistical challenges of integrating performance and MoBI research in a public setting. First, this study has a small sample size (three professional musicians). However, this drawback can be justified by the ecological validity of our experiments, which were intended to capture the interactions of musicians during real-world improvised performance,28,29 and the experimental design that included counterbalancing to allow for testing different participants in different orders. The authors believe the ecological approach and experimental methodology used herein represent a milestone in the acquisition and understanding of brain data “in action and in context”, and the development of brain-to-brain communication metrics. It is of interest of the authors to implement the presented methods in different experimental designs oriented to unveil the shared neural traces of human interactions under a variety of contexts (e.g., dance, theater, teaming, education, etc.).\n\nAnother limitation of this study is the potential lingering effects of artifacts associated to the movements needed to perform music through the (percussion and wind) instruments involved in the experiments. Such artifacts are likely related to body and facial movements, blowing, head swaying, among others, and can contaminate the EEG signals. Although we deployed well known pre-processing and de-noising methods found in the literature1,2,85 and performed visual inspection of the raw and cleaned data, it is still possible that residual motion and muscle-related artifacts may still remain in the processed EEG signals, and thus these results may be taken with caution. As additional information on this note, a comparison of EEG signals’ independent components (ICs) before and after the de-noising framework implemented in this study is presented in Figures S5-S10 (Extended data86).\n\nNevertheless, this issue will be present in any ecologically valid study on musical improvisation due to the freedom of movement of the musicians.28 Additionally, this performance case study offers a novel way to investigate inter-brain synchronization, in “action and in context”, during a free jazz improvisation in real-world scenarios by expert musicians.\n\n\nConclusion\n\nIn this study, temporal synchronization of EEG signals between musicians interacting during a jazz performance was observed through a bispectral analysis. The most significant interactions were found between left temporal, bilateral occipital and parietal regions at the gamma band, which reveals a shared dynamic and synchronized processing of auditory, visual and spatial information needed during a cooperative improvised performance. The inter-brain interaction between electrodes in sensory integration areas among musicians provides evidence towards the centrality of sensory processing,44 feedback,42,61 and communication42,43 during a collaborative musical improvisation.\n\nA temporal analysis of the bispectrum dynamics for both synchronized and desynchronized performing allowed to observe higher bispectrum when musicians were performing in a synchronized manner, when compared to desynchronized performing. In this study, bispectrum was useful to identify differences in competitive and collaborative performance in a real world scenario such as musicians improvising a collaborative piece. Based on the presented results, the implemented bispectral analysis method is proposed to study social interactions and brain-brain communication in hyperscanning measurements.\n\n\nData availability\n\nOSF: MOBILE EEG RECORDINGS OF MUSICAL (JAZZ) IMPROVISATION.\n\nhttps://doi.org/10.17605/OSF.IO/YUEQK86\n\nThis project contains the following underlying data:\n\n• Block1_P1.mat (EEG data - Recording Block1, Participant 1).\n\n• Block1_P2.mat (EEG data - Recording Block1, Participant 2).\n\n• Block1_P3.mat (EEG data - Recording Block1, Participant 3).\n\n• Block2_P1.mat (EEG data - Recording Block2, Participant 1).\n\n• Block2_P2.mat (EEG data - Recording Block2, Participant 2).\n\n• Block2_P3.mat (EEG data - Recording Block2, Participant 3).\n\n• Impedances.xlsx (Impedance values of EEG electrodes from all participants, at start and end of recordings).\n\n• Performance Notes.xlsx (Notes with times of trials, segments and relevant events during the performance).\n\n• ZOOM0001.mp3 (Audio recording of the complete performance).\n\n• Blaffer_Floor_1210.mp4 (Video Recording1 from the performance).\n\n• Blaffer_Floor_1221.mp4 (Video Recording2 from the performance).\n\nOSF: MOBILE EEG RECORDINGS OF MUSICAL (JAZZ) IMPROVISATION.\n\nhttps://doi.org/10.17605/OSF.IO/YUEQK86\n\nThis project contains the following extended data:\n\n• Extended Data.pptx (An extended data file containing additional figures and tables from this work).\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).", "appendix": "References\n\nCruz-Garza JG, Ravindran AS, Kopteva AE, et al.: Characterization of the stages of creative writing with mobile eeg using generalized partial directed coherence. 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Publisher Full Text\n\nWopereis IGJH, Stoyanov S, Kirschner PA, et al.: What makes a good musical improviser? an expert view on improvisational expertise. Psychomusicology: Music, mind, and brain. 2013; 23(4): 222–235. Publisher Full Text\n\nBeaty R: The neuroscience of musical improvisation. Neurosci. Biobehav. Rev. 01 2015; 51: 108–117. Publisher Full Text\n\nVuust P, Heggli OA, Friston KJ, et al.: Music in the brain. Nat. Rev. Neurosci. 2022; 23: 287–305. 1471-0048. Publisher Full Text\n\nHeggli OA, Konvalinka I, Cabral J, et al.: Transient brain networks underlying interpersonal strategies during synchronized action. Soc. Cogn. Affect. Neurosci. 2021; 16(1-2): 19–30. PubMed Abstract \| Publisher Full Text\n\nWilson GB, MacDonald RAR: Musical choices during group free improvisation: A qualitative psychological investigation. Psychol. Music. 2016; 44 (5): 1029–1043. 17413087. 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Publisher Full Text\n\nContreras-Vidal JL, Cruz-Garza J, Kopteva A: Towards a whole body brain-machine interface system for decoding expressive movement intent Challenges and Opportunities. 5th International Winter Conference on Brain-Computer Interface, BCI 2017. 2017; pages 1–4. Publisher Full Text\n\nRosen DS, Yongtaek O, Erickson B, et al.: Dual-process contributions to creativity in jazz improvisations: An SPM-EEG study. NeuroImage. 2020; 213 (September 2019): 116632. 10959572. PubMed Abstract \| Publisher Full Text\n\nBigdely-Shamlo N, Mullen T, Kothe C, et al.: The PREP pipeline: Standardized preprocessing for large-scale EEG analysis. Front. Neuroinform. 2015; 9 (JUNE): 1–19. 16625196. PubMed Abstract \| Publisher Full Text\n\nKilicarslan A, Grossman RG, Contreras-Vidal JL: A robust adaptive denoising framework for real-time artifact removal in scalp EEG measurements. J. Neural Eng. feb 2016; 13(2): 026013. 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Publisher Full Text\n\nZhang Y, Prasad S, Kilicarslan A, et al.: Multiple kernel based region importance learning for neural classification of gait states from EEG signals. Front. Neurosci. 2017; 11 (APR): 1–11. 1662453X. PubMed Abstract \| Publisher Full Text\n\nPressing J: Improvisation: methods and models. John A. Sloboda (Hg.): Generative processes in music, Oxford.1988; pages 129–178.\n\nHerholz SC, Zatorre RJ: Musical training as a framework for brain plasticity: behavior, function, and structure. Neuron. 2012; 76(3): 486–502. PubMed Abstract \| Publisher Full Text\n\nNovembre G, Keller PE: A conceptual review on action-perception coupling in the musicians’ brain: What is it good for?. Front. Hum. Neurosci. 2014; 8 (AUG): 1–11. 16625161. PubMed Abstract \| Publisher Full Text\n\nCisek P: Image Schemata. Encyclopedia of Neuroscience. 2009. Publisher Full Text\n\nWolpert D, Ghahramani Z, Jordan M: An internal model for sensorimotor integration. Science (New York, N.Y.). 10 1995; 269: 1880–1882. Publisher Full Text\n\nFuster JM: Upper processing stages of the perception - action cycle. Trends Cogn. Sci. 2004; 8(4): 143–145. PubMed Abstract \| Publisher Full Text\n\nPalomar-García MÁ, Zatorre RJ, Ventura-Campos N, et al.: Modulation of Functional Connectivity in Auditory-Motor Networks in Musicians Compared with Nonmusicians. Cereb. Cortex. 2017; 27 (5): 2768–2778. 14602199. PubMed Abstract \| Publisher Full Text\n\nBelden A, Zeng T, Przysinda E, et al.: Improvising at rest: Differentiating jazz and classical music training with resting state functional connectivity. NeuroImage. 2020; 207: 116384. PubMed Abstract \| Publisher Full Text\n\nCheng L, Guo ZW, Lai YX, et al.: Musical training induces functional plasticity in perceptual and motor networks: Insights from resting-state fMRI. PLoS One. 2012; 7 (5): 1–10. 19326203. PubMed Abstract \| Publisher Full Text\n\nVergara VM, Norgaard M, Miller R, et al.: Functional network connectivity during jazz improvisation. Sci. Rep. 2021; 11(1): 1–12. Publisher Full Text\n\nBhattacharya J, Petsche H, Pereda E: Long-range synchrony in the γ band: Role in music perception. J. Neurosci. 2001; 21 (16): 6329–6337. 02706474. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBhattacharya J, Petsche H: Phase synchrony analysis of EEG during music perception reveals changes in functional connectivity due to musical expertise. Signal Process. 2005; 85 (11): 2161–2177. 01651684. Publisher Full Text\n\nKawasaki M, Yamada Y, Ushiku Y, et al.: Inter-brain synchronization during coordination of speech rhythm in human-to-human social interaction. Sci. Rep. 2013; 3: 1–8. 20452322. PubMed Abstract \| Publisher Full Text\n\nWalton AE, Richardson MJ, Langland-Hassan P, et al.: Improvisation and the self-organization of multiple musical bodies. Front. Psychol. 2015b; 06 (MAR): 1–9. 16641078. PubMed Abstract \| Publisher Full Text\n\nGoupil L, Saint-Germier P, Rouvier G, et al.: Musical coordination in a large group without plans nor leaders. Sci. Rep. 2020; 10 (1): 20377–14. 20452322. PubMed Abstract \| Publisher Full Text\n\nKruppa JA, Reindl V, Gerloff C, et al.: Interpersonal Synchrony Special Issue Brain and motor synchrony in children and adolescents with ASD–a fNIRS hyperscanning study. Soc. Cogn. Affect. Neurosci. 2020; 16: (January): 103–116. 1749-5016. PubMed Abstract \| Publisher Full Text\n\nGvirts HZ, Perlmutter R: What Guides Us to Neurally and Behaviorally Align With Anyone Specific? A Neurobiological Model Based on fNIRS Hyperscanning Studies. Neuroscientist. 2020; 26 (2): 108–116. 10894098. PubMed Abstract \| Publisher Full Text\n\nBabiloni F, Astolfi L: Social neuroscience and hyperscanning techniques: Past, present and future. Neurosci. Biobehav. Rev. 2014; 44: 76–93. 18737528. PubMed Abstract \| Publisher Full Text\n\nDikker S, Michalareas G, Oostrik M, et al.: Crowdsourcing neuroscience: inter-brain coupling during face-to-face interactions outside the laboratory. NeuroImage. 2021; 227: 117436. PubMed Abstract \| Publisher Full Text\n\nBevilacqua D, Ido Davidesco L, Wan KC, et al.: Brain-to-brain synchrony and learning outcomes vary by student-teacher dynamics: Evidence from a real-world classroom electroencephalography study. J. Cogn. Neurosci. 2019; 31(3): 401–411. PubMed Abstract \| Publisher Full Text\n\nDavidesco I, Laurent E, Valk H, et al.: Brain-to-brain synchrony between students and teachers predicts learning outcomes (preprint).2019. Publisher Full Text\n\nPan Y, Dikker S, Goldstein P, et al.: Instructor-learner brain coupling discriminates between instructional approaches and predicts learning. NeuroImage. 2020; 211: 116657. PubMed Abstract \| Publisher Full Text\n\nStephens GJ, Silbert LJ, Hasson U: Speaker-listener neural coupling underlies successful communication. Proc. Natl. Acad. Sci. U. S. A. 2010; 107 (32): 14425–14430. 00278424. PubMed Abstract \| Publisher Full Text\n\nLiu T, Saito G, Lin C, et al.: Inter-brain network underlying turn-based cooperation and competition: A hyperscanning study using near-infrared spectroscopy. Sci. Rep. 2017; 7 (1): 8684–12. 20452322. PubMed Abstract \| Publisher Full Text\n\nKontson KL, Megjhani M, Brantley JA, et al.: Your brain on art: Emergent cortical dynamics during aesthetic experiences. Front. Hum. Neurosci. 2015; 9 (NOVEMBER): 1–17. 16625161. PubMed Abstract \| Publisher Full Text\n\nRamírez-Moreno MA, Cruz-Garza JG, Paek A, et al.: MOBILE EEG RECORDINGS OF MUSICAL (JAZZ) IMPROVISATION. OSF. [Dataset].2022. Publisher Full Text" }	[ { "id": "158229", "date": "20 Jan 2023", "name": "Stephane Perrey", "expertise": [ "Reviewer Expertise Neuroscience and Neuroimaging on the field." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis study aimed to assess temporal synchronization of EEG signals between three professional musicians interacting during a jazz performance. Some relevant interactions were found through a bispectral analysis. A reflective cooperative intention was observed during the task. This original and pilot study was carried out during a free jazz improvisation tasks of long duration in real-world scenario by expert musicians. This case study is well presented and proposes very interesting thoughts and new perspectives in social neuroscience.\nAuthors were able to deal nicely with possible artifacts associated to the movements during such long plays by performing suitable analyses with EEG signals.\nBelow you will find some comments for your convenience.\nIntroduction:\nConcerning the following sentence, additional insight offered by the bispectrum, when compared to other neural synchrony metrics, needs to be clarified more. First, what does mean here \"a more complete intuition on phase coupling, resonance, temporal synchronization and non-linear interactions between any analyzed signal pair\". Second, Please highlight the added-value of bispectrum if any.\n\nThe following sentence without reference does not bring more into the rationale of this study and could be removed: \"Musical improvisation can also be considered as an’on the fly’ composition, one that is temporally ubiquitous, spontaneous, and is not restricted by critique.\"\n\nJazz performance needs to be defined/characterized in the introduction; different stages do exist.\n\n\"Brain-to-brain communication\", as an important metric proposed by authors is cited as a shared cognitive state. Please confirm is this is right and how this metric is associated to hyperscanning methods.\n\nOverall, introduction is delivering many terms around hyperscanning techniques, which makes reading difficult.\n\nPlease explain/inform on what is a \"creative collaboration\" task, not really defined in the introduction. Thereafter you wrote \"collaborative musical improvisation\". Are you saying the same?\n\nIn the following sentence (end of introduction), since the previous underlined/questioned terms are present, they have to be sufficiently clear. \"The current study examines the neural correlates of brain-to-brain communication of jazz musicians during collaborative musical improvisation through hyperscanning\".\n\nMobile EEG can impose movement constraints. This is not so straightforward. Please adjust accordingly.\nMethods\nParticipants. In terms of experiences (years performing jazz together, musical background) for collaborative tasks, P1 and P2 have likely more chance to \"interact » better during collaborative musical improvisation; P3 should present some different responses. Does this heterogeneity was voluntary targeted?\n\nImpedance was set to less than 25 kOhms. It appears relatively high as compared to classic/regular use in the field. Can you provide some methodological reference/guidelines?\n\nAnalysis methods (EEG features, temporal and spatial) are well detailed and are suitable for the data set. Assessment of brain synchrony (maybe state \"brain-to brain communication\") across two ecological sections (synchronized or not) and three participant conditions (active vs passive in a dyad) of jazz performance is relevant and within the scope of Neuroergonomics. Statistical analysis are complete.\n\nResults\nThere are many results, some of them being very observational due to the exploratory feature of this case study. However, can authors select the main findings regarding the brain-to brain communication of Jazz musicians during the collaborative musical improvisation sequences?\nDiscussion\nThis section presents some first evidence and underlines well some specific intra- inter-brain coupling modulated by musical training and previous experiences. Perspectives and methodological limitations are provided. Body movements as main sources of EEG artefacts should be captured in a further study in order to get synchronisation at the behavioral level associated to their neural correlates.\n\nEmotion is likely playing a role in the proposed collaborative musical improvisation task (see doi: 10.3389/fnhum.2022.944241) between the musicians but also with audience, as musicians interact in a jazz performance with a live audience. Please advice.\n\nThis study included freely-moving expert musicians. Do you have collected/recorded the body motion of the musicians? Have you any ideas on the kind of movements over the two sequences defined (SP and DP)?Also, it might be surprising to not observe brain synchronization with the motor regions. The ability to synchronize movement with auditory rhythms is relying on motor networks, such as cortical areas, basal ganglia and the cerebellum, which also participate in rhythm perception and movement production (doi: 10.1016/j.neubiorev.2019.12.024). Have you look at these cortical regions?\n\nMinor: correct \"synchornicity in sensory areas\"\n\nIs the background of the case’s history and progression described in sufficient detail? Yes\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate? Yes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes\n\nIs the case presented with sufficient detail to be useful for teaching or other practitioners? Yes", "responses": [ { "c_id": "9600", "date": "26 May 2023", "name": "Mauricio Ramirez", "role": "Author Response", "response": "We thank the reviewer for the time invested in reading and making such useful comments for our work. Below is a response to each of the comments (in bold letters), as well as the locations of the corrections made to the manuscript. All changes are carried out in the next version of the manuscript. Concerning the following sentence, additional insight offered by the bispectrum, when compared to other neural synchrony metrics, needs to be clarified more. First, what does mean here \"a more complete intuition on phase coupling, resonance, temporal synchronization and non-linear interactions between any analyzed signal pair\". Second, Please highlight the added-value of bispectrum if any. Introduction, Page 2, Lines 111-115. The mentioned paragraph was extended to be clearer. References were included as well as some advantages of using the bispectrum metric. The following sentence without reference does not bring more into the rationale of this study and could be removed: \"Musical improvisation can also be considered as an’on the fly’ composition, one that is temporally ubiquitous, spontaneous, and is not restricted by critique.\" Introduction, Page 2, Lines 169-173. The sentence was rephrased, and a new reference from a music composition book was added. Jazz performance needs to be defined/characterized in the introduction; different stages do exist. Introduction, Page 3, Lines 176-186. A paragraph adding more detail about jazz and free jazz performance was added with a reference. \"Brain-to-brain communication\", as an important metric proposed by authors is cited as a shared cognitive state. Please confirm is this is right and how this metric is associated to hyperscanning methods. Introduction, Page 3, Lines 231-238. A paragraph and a reference were included to add more detail about the use of hyperscanning for assessment of brain-to-brain communication. Overall, introduction is delivering many terms around hyperscanning techniques, which makes reading difficult. Introduction. Page 1 – 3. Subsections were added to the Introduction to guide the reader throughout the text. No text was removed since in further comments, reviewer suggests that some terms are explained clearly. Please explain/inform on what is a \"creative collaboration\" task, not really defined in the introduction. Thereafter you wrote \"collaborative musical improvisation\". Are you saying the same? Introduction, Page 3, Line 230 Yes, they imply the same idea, however the term “collaborative musical improvisation” was used instead, to avoid confusion. In the following sentence (end of introduction), since the previous underlined/questioned terms are present, they have to be sufficiently clear. \"The current study examines the neural correlates of brain-to-brain communication of jazz musicians during collaborative musical improvisation through hyperscanning\". Thank you for the comment, we made sure previous terms are clear and present, as suggested. Mobile EEG can impose movement constraints. This is not so straightforward. Please adjust accordingly. Introduction, Page 3, Lines 263-264. A clarification on this was added, movement constraints imposed by mobile EEG are less than for other brain activity measuring techniques, for example fMRI scans. Methods Participants. In terms of experiences (years performing jazz together, musical background) for collaborative tasks, P1 and P2 have likely more chance to \"interact » better during collaborative musical improvisation; P3 should present some different responses. Does this heterogeneity was voluntary targeted? Methods, Page 4, Lines 316-319. It was not targeted on purpose, those were the volunteers that agreed to participate in the study. An observation on this regard was included. Impedance was set to less than 25 kOhms. It appears relatively high as compared to classic/regular use in the field. Can you provide some methodological reference/guidelines? Methods, Page 4, Lines 335-339. A reference was added to how this value was selected. Analysis methods (EEG features, temporal and spatial) are well detailed and are suitable for the data set. Assessment of brain synchrony (maybe state \"brain-to brain communication\") across two ecological sections (synchronized or not) and three participant conditions (active vs passive in a dyad) of jazz performance is relevant and within the scope of Neuroergonomics. Statistical analysis are complete. Thank you, we appreciate your comments. Results There are many results, some of them being very observational due to the exploratory feature of this case study. However, can authors select the main findings regarding the brain-to brain communication of Jazz musicians during the collaborative musical improvisation sequences? Results, Page 10, Lines 730-744. Main findings were added and highlighted by the end of the results section, in a new subsection. Discussion This section presents some first evidence and underlines well some specific intra- inter-brain coupling modulated by musical training and previous experiences. Perspectives and methodological limitations are provided. Body movements as main sources of EEG artefacts should be captured in a further study in order to get synchronisation at the behavioral level associated to their neural correlates. Discussion, Page 15, Lines 1049-1051.Thank you, this is a great comment, which indeed can be explored in a future study. A note on this regard was added to the discussion section. Emotion is likely playing a role in the proposed collaborative musical improvisation task (see doi: 10.3389/fnhum.2022.944241) between the musicians but also with audience, as musicians interact in a jazz performance with a live audience. Please advice. Discussion, Page 15. Lines 1052-1059. A comment including the emotional context during musical improvisation was included as suggested. This study included freely-moving expert musicians. Do you have collected/recorded the body motion of the musicians? Have you any ideas on the kind of movements over the two sequences defined (SP and DP)?Also, it might be surprising to not observe brain synchronization with the motor regions. The ability to synchronize movement with auditory rhythms is relying on motor networks, such as cortical areas, basal ganglia and the cerebellum, which also participate in rhythm perception and movement production (doi: 10.1016/j.neubiorev.2019.12.024). Have you look at these cortical regions? Only head motion was collected, but not analyzed in this study. Indeed will be included as future steps. The auditory-motor synchronization in the references talk about such functional network within one person. However, when analyzing our data, synchronization between motor areas and motor-auditory were not observed. We believe this was due to the individuality of movement in the production of sounds (motor areas not in synch), however the perceptual information and processes were shared among musicians, therefore the synchrony in temporal, parietal, and occipital regions (perceptual areas in synch). Minor: correct \"synchornicity in sensory areas\" This error was fixed. Discussion, Page 12, Lines 838." } ] } ]	1	https://f1000research.com/articles/11-989
https://f1000research.com/articles/12-1431/v1	02 Nov 23	{ "type": "Software Tool Article", "title": "parazitCUB: An R package to streamline the process of investigating the adaptations of parasites' codon usage bias", "authors": [ "Ali Mostafa Anwar", "Salma Bayoumi", "Sagy Elzalabany", "Sameh Magdeldin", "Amr E. Ahmed", "Ali Mostafa Anwar", "Salma Bayoumi", "Sagy Elzalabany", "Sameh Magdeldin" ], "abstract": "Examining the intricate association between parasites and their hosts, particularly at the codon level, assumes paramount importance in comprehending evolutionary processes and forecasting the characteristics of novel parasites. While diverse metrics and statistical analyses are available to explore codon usage bias (CUB), there presently exists no dedicated tool for examining the co-adaptation of codon usage between parasites and hosts. Therefore, we introduce the parazitCUB R package to address this challenge in a scalable and efficient manner, as it is capable of handling extensive datasets and simultaneously analyzing of multiple parasites with optimized performance. parazitCUB enables the elucidation of parasite-host interactions and the evolutionary patterns of parasites through the implementation of various indices, cluster analysis, multivariate analysis, and data visualization techniques. The tool can be accessed at the following location: https://github.com/AliYoussef96/parazitCUB", "keywords": [ "Molecular evolution", "Natural selection", "Adaptation", "parasites", "Codon Usage Bias", "R", "RStudio" ], "content": "Introduction\n\nThe transfer of genetic information from messenger RNAs (mRNAs) to proteins occurs through codons, which are sequences of three nucleotides representing amino acids. With the exception of methionine (Met) and tryptophan (Trp), most amino acids can be encoded by multiple codons, resulting in codon degeneracy. Based on studies conducted on multiple organisms, synonymous codons, which encode the same amino acid, are not uniformly utilized within genes or across different genes in the same genome, leading to codon usage bias (CUB) phenomenon.1 In every organism, specific preferred (optimal) codons exist, which are utilized more frequently in highly expressed genes compared to genes with lower expression levels.2 The codon usage of an organism is influenced by two major forces: mutation pressure and natural selection. Nucleotide composition, synonymous substitution rate, tRNA abundance, codon hydropathy, DNA replication initiation sites, gene length, and expression level are all known to impact the CUB.1\n\nIntracellular parasites can be categorized as facultative or obligate. Facultative parasites can reproduce both inside and outside host cells, whereas obligate parasites are unable to replicate outside their host cells and solely depend on the host cell’s resources for reproduction.3 Previous studies have shown that translational selection and/or directed mutational pressure shape the codon usage of intracellular parasite genomes to optimize or deoptimize it towards the codon usage of their hosts.3,4 Previous investigations have emphasized the significance of examining the interplay between parasites and the codon usage of their hosts. For instance, research conducted on the Influenza A virus (IAV) has demonstrated that understanding the patterns of codon usage in viruses might aid in the development of novel vaccines through the use of Synthetic Attenuated Virus Engineering (SAVE), which involves weakening a virus by deoptimizing its viral codons.5 Similarly, another study demonstrated that the replacement of natural codons with synonymous triplets possessing higher CpG frequencies can effectively deactivate poliovirus infectivity.6 To understand how parasites interact with their hosts and how they evolve, it is crucial to investigate the composition of parasite genes at the codon or nucleotide level. This analysis could assist in uncovering the mechanisms underlying parasite-host interactions and help in predicting the characteristics of newly discovered parasites.\n\nA variety of metrics have been established to evaluate Codon Usage Bias (CUB), including the effective number of codons (ENc), codon adaptation index (CAI), relative synonymous codon usage (RSCU), and translational selection index (P2-index).7 Statistical analyses, such as correspondence analysis and the Neutrality Plot, have been employed to explore the influence of selection and mutation on molding CUB.7 Various tools and packages, such as coRdon,9 CodonW (http://codonw.sourceforge.net), and BCAWT.8 are available for assessing and measuring CUB. However, there is currently a lack of specialized software specifically designed to examine the co-adaptation of codon usage between parasites and their hosts. The only available package developed for studying the interaction of codon usage between viruses and hosts was created in 2019 by the same first author of this research, known as vhcub R package.\n\nThe infection of multiple organisms by various parasites is a widespread phenomenon, exemplified by the existence of 1424 known viruses that can affect humans, as documented in the virus-host database.9 Investigating the co-evolution of codon usage between parasites and their respective hosts presents a challenging task in the field of bioinformatics. However, thanks to modern techniques and software advancements, this endeavor has become feasible. To address this challenge in a scalable and efficient manner, the parazitCUB tool was developed. The ParazitCUB, in contrast to its predecessor vhcub package, offers an expanded scope that goes beyond virus-host interactions. It encompasses the co-evolution of codon usage between parasites and hosts, providing a more comprehensive analysis. Notably, ParazitCUB allows for the examination of larger and more extensive datasets, making it well-suited for handling substantial amounts of data. Additionally, it enables the concurrent study of multiple parasites, a feature lacking in vhcub, which only permits the analysis of a single organism with its host. Moreover, ParazitCUB has undergone significant optimization of its functions, resulting in improved speed and performance. A notable advantage of ParazitCUB lies in its user-friendly interface, facilitating effortless utilization even for users with limited proficiency in R programming.\n\n\nMethods\n\nParazitCUB employs several packages, such as Biostrings,10 seqinr,11 and stringr,12 to handle FASTA formate files and perform DNA sequence modifications. For CUB and multivariate analysis, the package utilizes coRdon and factoextra,13 as well as new functions implementation. To visualize the data effectively, ParazitCUB utilizes, ggplot2,14 pheatmap,15 and RColorBrewer.16 ParazitCUB efficiently extracts DNA sequences in FASTA format for each organism under study. These sequences are then combined into a comprehensive list. The package encompasses various indices for investigating CUB, as well as cluster analysis, multivariate analysis, and data visualization. A comprehensive list of the package’s functions, along with their corresponding results, can be found in Table 1. As well as, the package workflow has also been summarized in (Figure 1).\n\nparazitCUB was developed in R, and the source code can be found on GitHub and archived with Zenodo.20 It works with Windows and most Linux operating systems.\n\nThe parazitCUB package consists of six main branches, each serving a distinct purpose: nucleotide content analysis, CUB analysis at the gene level, CUB analysis at the codon level, cluster analysis, multivariate analysis, and data visualization. Within each branch, a range of methods is available for conducting CUB studies. The complete workflow of ParazitCUB is illustrated in Figure 1, providing an overview of the entire process. For comprehensive information on using ParazitCUB, detailed documentation is readily available https://github.com/AliYoussef96/parazitCUB.\n\n\nUse cases\n\nThe utilization of parazitCUB for investigating codon usage bias (CUB) in viruses (or any type of parasites), their respective hosts, and the co-adaptation between them offers a straightforward and highly customizable approach. To exemplify the capabilities of the package, the coding sequences of seven viruses, namely Influenza A, Influenza B, Influenza C, Influenza D, MERS, SARS-CoV, and SARS-CoV-2, were obtained from NCBI virus gateway.17 To showcase the package’s ability to handle larger datasets, two variants of each virus were downloaded, with one variant isolated from a non-human host and the other from a human host (except for Influenza D).\n\nTo begin, all the fasta files for the viruses should be located in a single directory, such as a folder named “virus fasta” To read all the files simultaneously for parazitCUB analysis, the following straightforward approach can be employed:\n\nAfter importing the host FASTA file, we focused exclusively on the human host for this particular analysis (only one host selection is permitted). In this instance, the genes exhibiting the highest expression levels in human lung tissues were collected from the Human Protein Atlas project database.\n\nA reasonable quality control step is to examine the coding sequence length in the virus datasets, to remove any bias (very long sequences, or very short ones) that could negatively affect the result. parazitCUB provides an easy straightforward function to do that.\n\nThis function will create a boxplot (Figure 2A) which illustrates the distribution of coding sequence lengths across the study. Through the examination of outliers in the boxplot, the QC.cutoff() function can be employed to exclude extremely long and short sequences from subsequent analyses, thereby enhancing the integrity of the data.\n\nA) A box plot for the lengths of all coding sequences in the study, serving as a quality control measure. With outliers displayed as red dots. B) A box plot illustrates the GC content of all organisms in the study for each codon position and provides an overall view of the GC content. C) A heatmap, combined with cluster analysis, utilizes the statistical representation of dinucleotide over- and underrepresentation to visually depict patterns and similarities among the data. D) Conducting cluster analysis on a Principal Component Analysis (PCA) using the RSCU values for each organism in the study enables the identification of clusters and relationships based on codon usage.\n\nTo exemplify the provided CUB workflow; we will show a case study involving the utilization of various functions from each of the six branches of the package.\n\nTo compute the GC content at every position across all viruses included in the study:\n\nGC.boxplot() function, which produces a graphical representation of the GC content distribution (Figure 2B).\n\nAll visualizations within the parazitCUB package are created using ggplot2, which allows for convenient customization of the default plot parameters.\n\nFurthermore, the package provides the capability to calculate the statistical over- and underrepresentation of dinucleotides using different models such as base, codon, and synonymous codons. The calculation can be performed as follows:\n\nAs part of this section, numerous indices can be calculated to assess Codon Usage Bias (CUB). For example, the effective number of codons (ENc) can be determined using the ENc.values.new() function, which utilizes a modified version.18 Also, MILC.values(), B.values(), and MCB.values() functions could be used to calculate the MILC, B, and MCB, respectively. All of these functions can work on the virus coding sequence without the need for a host coding sequence as a reference set.\n\nAdditionally, the MILC.values(), B.values(), and MCB.values() functions can be employed to calculate the MILC, B, and MCB indices, respectively. It is important to note that these functions can operate on the virus coding sequence alone, without requiring a host coding sequence as a reference set.\n\nSome indices within the ParazitCUB package require a reference gene set to ensure their accurate computation and cannot be executed without it. One such example is:\n\nCertain indices rely on a reference genes set for their proper functioning and cannot operate without it. For instance;\n\nMoreover, various matrices are provided within ParazitCUB to facilitate the examination of Codon Usage Bias (CUB) at the codon level. For instance:\n\nCluster analysis and multivariate analysis have been widely utilized in numerous research studies to explore codon usage patterns. Within the framework of parazitCUB, three essential functions have been integrated for this purpose. One of these functions, cub.heatmap(), facilitates the generation of a heatmap using either Relative Synonymous Codon Usage (RSCU) values or statistical representations of dinucleotide over- and underrepresentation. Additionally, cub.heatmap() supports the utilization of various clustering methods implemented through the R stats function hclust()19 (Figure 2C).\n\nPrincipal Component Analysis (PCA) is a commonly employed technique to reduce the dimensionality of RSCU values and identify the primary sources of variation and factors influencing codon usage within an organism. This task can be easily performed using the parazitCUB package (Figure 2D).\n\nSubsequently, cluster analysis can be conducted based on the PCA results as follows:\n\nThe forces that impact codon usage bias (CUB), such as mutational pressure and natural selection, have been extensively explored using various plots including the ENc-GC3 plot, PR2 plot, and Neutrality plot. In parazitCUB, two versions of the ENc-GC3 plot are available: the first version displays the ENc-GC3 of a specific virus analyzed (Figure 3A), while the second version presents the average ENc-GC3 for all the organisms studied in a single figure (Figure 3B). The same applies to the PR2 plot (Figure 4A and B). The Neutrality plot, can only be used for one organism at a time (Figure 5).\n\nA) ENc-GC3 plot displays the ENc values plotted against the GC3 content for the virus Influenza A (human CDS as reference) CDS. In this plot, the solid red line represents the expected ENc values when the codon bias is solely influenced by GC3s. B) The plot represents the average effect of ENc-GC3 for all organisms included in the study.\n\nA) A PR2-plot illustrates the coding sequences (CDS) of the Influenza A (human CDS as reference) CDS, depicting their GC bias (ratio of G3 to G3 + C3) and AT bias (ratio of A3 to A3 + T3) in the third position of each codon. The two solid red lines on the graph indicate the point where both the vertical and horizontal coordinates are 0.5, representing the condition where A is equal to T and G is equal to C. B) The plot represents the average effect of PR2 values for all organisms included in the study.\n\nOn the plot, the y-axis represents the average GC frequency at the first and second codon positions (GC12), while the x-axis represents the GC frequency at the third codon position (GC3). The equation for the slope, along with the coefficient of determination (R) and its associated p-value, are shown.\n\n\nConclusions\n\nparazitCUB streamlines the process of investigating the adaptations of parasites’ Codon Usage Bias (CUB) within the R environment, ensuring scalability and efficiency. By allowing the study of several parasites concurrently in connection to a specific host, ParazitCUB facilitates a thorough understanding of the factors influencing parasite evolution and offers potential insights for establishing effective treatment strategies", "appendix": "Data availability\n\nZenodo: AliYoussef96/parazitCUB: V1.0.0. https://doi.org/10.5281/zenodo.8393578. 20\n\nThis project contains the following underlying data:\n\n• Fasta Files: A folder containing all the fasta files used in the case study https://github.com/AliYoussef96/parazitCUB/tree/main/flu%20fasta\n\n\nReferences\n\nPlotkin JB, Kudla G: Synonymous but not the same: the causes and consequences of codon bias. Nat. Rev. Genet. January 2011; 12(1): 32–42. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHershberg R, Petrov DA: General rules for optimal codon choice. PLoS Genet. July 2009; 5(7): e1000556. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHanes A, Raymer M, Doom T, et al.: A comparision of codon usage trends in prokaryotes. Bioinformatics, 2009 Ohio Collaborative Conference. 06 2009; 83–86. Publisher Full Text\n\nChandan J, Gupta S, Babu V, et al.: Comprehensive analysis of codon usage pattern in withania somnifera and its associated pathogens: Meloidogyne incognita and alternaria alternata. Genetica. April 2022; 150(2): 129–144. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDiamantopoulos PT, Michael M, Benopoulou O, et al.: Antiretroviral activity of 5-azacytidine during treatment of a HTLV-1 positive myelodysplastic syndrome with autoimmune manifestations. Virol. J. January 2012; 9: 1. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBurns CC, Campagnoli R, Shaw J, et al.: Genetic inactivation of poliovirus infectivity by increasing the frequencies of CpG and UpA dinucleotides within and across synonymous capsid region codons. J. Virol. July 2009; 83(19): 9957–9969. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nParvathy ST, Udayasuriyan V, Bhadana V: Codon usage bias. Mol. Biol. Rep. November 2021; 49(1): 539–565. PubMed Abstract \| Publisher Full Text\n\nAnwar AM: Bcawt: Automated tool for codon usage bias analysis for molecular evolution. J. Open Source Softw. 2019; 4(42): 1500. Publisher Full Text\n\nMihara T, Nishimura Y, Shimizu Y, et al.: Linking virus genomes with host taxonomy. Viruses. March 2016; 8(3): 66. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPagès H, Aboyoun P, Gentleman R, et al.: Biostrings: Efficient manipulation of biological strings.2022. R package version 2.66.0. Reference Source\n\nCharif D, Lobry JR: SeqinR 1.0-2: a contributed package to the R project for statistical computing devoted to biological sequences retrieval and analysis.Bastolla U, Porto M, Roman HE, et al., editors. Structural approaches to sequence evolution: Molecules, networks, populations, Biological and Medical Physics, Biomedical Engineering. New York: Springer Verlag; 2007; pp. 207–232. 978-3-540-35305-8.\n\nWickham H: stringr: Simple, Consistent Wrappers for Common String Operations.2022. R package version 1.5.0. Reference Source\n\nKassambara A, Mundt F: factoextra: Extract and Visualize the Results of Multivariate Data Analyses.2020. R package version 1.0.7. Reference Source\n\nWickham H: ggplot2: Elegant Graphics for Data Analysis. New York: Springer-Verlag; 2016. 978-3-319-24277-4. Reference Source\n\nKolde R: pheatmap: Pretty Heatmaps.2019. R package version 1.0.12. Reference Source\n\nNeuwirth E: RColorBrewer: ColorBrewer Palettes.2022. R package version 1.1-3. Reference Source\n\nHatcher EL, Zhdanov SA, Bao Y, et al.: Virus variation resource - improved response to emergent viral outbreaks. Nucleic Acids Res. November 2016; 45(D1): D482–D490. PubMed Abstract \| Publisher Full Text\n\nNovembre JA: Accounting for background nucleotide composition when measuring codon usage bias. Mol. Biol. Evol. August 2002; 19(8): 1390–1394. PubMed Abstract \| Publisher Full Text\n\nR Core Team: R: A Language and Environment for Statistical Computing. Vienna, Austria: R Foundation for Statistical Computing; 2022. Reference Source\n\nYoussef A: AliYoussef96/parazitCUB: V1.0.0 (V1.0.0). Zenodo. 2023. Publisher Full Text" }	[ { "id": "220456", "date": "24 Nov 2023", "name": "Ahmed Abdelmonem Hemedan", "expertise": [ "Reviewer Expertise Systems medicine", "Disease dynamic modelling", "Biostatistics", "Mathematics", "Bioinformatics", "Molecular Pathology." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\n# parazitCUB manuscript review\nThe article introducing parazitCUB, an R package for analysing codon usage bias in parasites and hosts - It is a valuable addition to the field, addressing a unique research need.\nThe introduction would require incorporating specific examples where codon usage bias analysis has impacted our understanding of parasitic diseases and treatment strategies.\nA thorough comparative analysis with existing tools like CodonW or coRdon is required - perhaps you would like to focus on unique features, user interface, data handling capabilities, and specific functionalities, would provide valuable insights.\nExpanding on the computational methodologies and algorithms used in parazitCUB is crucial for scientific rigor and reproducibility.\nIncluding a robust performance assessment section, exploring aspects like accuracy, computational efficiency, and scalability through comparative analyses, is essential.\nDetailed case studies in the use case section, demonstrating the tool's utility with specific data and analytical processes, would illustrate its practical value.\nEnhanced documentation, including a comprehensive user guide, example datasets, troubleshooting tips, and FAQs, is necessary to make the tool approachable to a broader audience.\nEnriching the paper with more effective visuals and data visualizations would aid in conveying complex data and analyses in an engaging manner.\nThe discussion section should contextualise parazitCUB within the broader bioinformatics field and its potential impact on future research, including possible expansions or updates of the tool.\n\nIs the rationale for developing the new software tool clearly explained? Partly\n\nIs the description of the software tool technically sound? Yes\n\nAre sufficient details of the code, methods and analysis (if applicable) provided to allow replication of the software development and its use by others? Yes\n\nIs sufficient information provided to allow interpretation of the expected output datasets and any results generated using the tool? Yes\n\nAre the conclusions about the tool and its performance adequately supported by the findings presented in the article? Yes", "responses": [] }, { "id": "220457", "date": "27 Nov 2023", "name": "Xianzhao Kan", "expertise": [ "Reviewer Expertise Molecular evolution", "bioinformatics", "molecular biology." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nGeneral comments:\nparazitCUB is a useful R package for investigating codon usage co-adaptation between parasites and their hosts. Unlike its predecessor vhcub which focuses solely on virus-host interactions, ParazitCUB provides an enhanced scope. The package provides comprehensive statistical analyses, including Neutrality plot, Maximum likelihood codon bias, CU Heatmap using RSCU, and so on. Overall, I believe that ParazitCUB provides convenience for in-depth research on parasite-host co-evolution.\nImproved:\n“Host (e.g., virus) Fasta File” in the upper right corner of Figure 1 should be “Host (e.g., human) Fasta File”.\n\nTo avoid repeating the word \"previous,\" the sentence (Second paragraph of Introduction) \"Previous investigations have emphasized the significance of examining the interplay between parasites and the codon usage of their hosts\" should be rephrased as \"This has emphasized the significance of examining the interplay between parasites and the codon usage of their hosts.\"\n\nIs the rationale for developing the new software tool clearly explained? Yes\n\nIs the description of the software tool technically sound? Yes\n\nAre sufficient details of the code, methods and analysis (if applicable) provided to allow replication of the software development and its use by others? Yes\n\nIs sufficient information provided to allow interpretation of the expected output datasets and any results generated using the tool? Yes\n\nAre the conclusions about the tool and its performance adequately supported by the findings presented in the article? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1431
https://f1000research.com/articles/12-1040/v2	02 Nov 23	{ "type": "Research Article", "title": "Causal relationship model of marketing innovation and competitiveness of small and medium enterprises (SMEs) with digital technologies in Thailand", "authors": [ "Soonthonsmai Vuttichat", "Piluk Patchara", "Soonthonsmai Vuttichat" ], "abstract": "Background: This study investigates the causal model of marketing innovation and competitiveness of small and medium enterprises (SMEs) with digital technologies in Thailand. This study aimed to investigate market orientation, digital marketing strategies, competitiveness, innovative marketing behavior, and digital marketing efficiency of SMEs.\nMethods: We validated the causal model of digital marketing efficiency using empirical evidence to investigate the direct and indirect effects of market orientation, digital marketing strategies, and innovative marketing behavior on digital marketing efficiency. The study was conducted using a questionnaire.\nResults: A total of 361 samples were collected using purposive sampling. A Structural Equation Model was developed and tested using LISREL software. The study found that the causal relationship model was consistent with the empirical evidence.\nConclusions: Digital marketing strategy and competitiveness directly positively affected innovative marketing behavior and explained 89% of the variance in innovative marketing behavior. Innovative marketing behavior directly positively affects digital marketing efficiency, and competitiveness indirectly positively affects digital marketing efficiency through innovative marketing behavior and explains the variance in digital marketing efficiency by 18%.", "keywords": [ "Market orientation", "Digital Marketing Strategies", "Competitiveness", "Marketing Innovative Behavior", "SMEs" ], "content": "Introduction\n\nIt is crucial for entrepreneurs to continually develop potential knowledge to tackle the challenges and opportunities that come their way effectively. This means developing new products and services, improving processes and operations, and expanding the market reach. Investing in knowledge and skill development will assist in promoting small and medium enterprises (SMEs) in Thailand, especially in the agri-food industry, which is growing rapidly in many countries. As entrepreneurs, it is also important to focus on developing abilities and competitiveness. This means creating a supply system that can serve the global market with a focus on quality over price. Unfortunately, many SMEs in the agri-food industry struggle with insufficient competitiveness. However, SMEs are the backbone of the economy in many developing countries that rely on agri-food production (World Bank, 2018; Organisation for Economic Co-operation and Development (OECD), 2019; European Commission, 2020).\n\nAccording to Bonanno, Golgeci, and Ioppolo (2020), Radicic and Escribano (2015), and Faraaz and Mishra (2019), SMEs in the agricultural food processed products industry are one of the most substantial and fastest-growing enterprises in an agri-food supply chain with the ability to enhance a wealthy industry and sustainable profits in the long term. They are often called the core foundation of the economy because they create jobs, generate income, and contribute to economic growth. The industry is a crucial sector of the economy for many food producers. In terms of products, agri-food-processed SMEs are involved in producing a wide range of food products, including canned fruits and vegetables, dried fruits, fruit juices, and frozen foods. These products are often made using traditional recipes and methods and are known for their high quality and unique flavors.\n\nWith the emergence of digital technology, SMEs must embrace market-oriented strategies, innovate their marketing approaches, and develop effective digital marketing strategies to maintain competitiveness in the market. According to the European Commission’s “Small Business Act” (2020), most SMEs often lack digitalization and innovation, which can hinder their ability to compete in a global marketplace. This is particularly important in the context of the COVID-19 pandemic, as many SMEs have had to adapt to new digital ways of working to stay competitive. However, many SMEs need more abilities or competitiveness, which can hinder their success and growth. Common challenges SMEs face include limited access to financing, insufficient technological capabilities, inadequate marketing strategies, and difficulties in finding and retaining skilled employees. These enterprises have a high potential for competition and seek opportunities from food industry growth trends through innovation and digital technologies (Tóth et al., 2019). To increase competitiveness, productivity must be increased to sustainably build wealth for the country (Porter 1990, 2010).\n\nPorter (1990) found that if SMEs could increase their productivity to a higher level, it would result in an advantage over competitors. If enterprises in many industrial sectors increased productivity, it would also result in a higher level of competitiveness. If numerous industries in the country increased productivity, it would lead to a higher level of competitiveness in the country eventually. To achieve this, SMEs seek opportunities from agri-food industry growth trends through marketing innovation and digital technologies (Hossain & Rahman, 2017). Marketing innovation behavior is a crucial and innovative action for SMEs to enhance their competitiveness with digital technology. Digital marketing allows SMEs to improve their marketing strategies, reach more consumers, and increase their market share. Consequently, marketing innovation behavior in agri-food processing SMEs involves various management strategies, such as creating unique product features, packaging, labeling, and developing new product lines with creative communication channels and content that appeal to consumers’ changing tastes and preferences. It also involves utilizing new marketing channels such as social media, mobile marketing, and online marketplaces to reach consumers and engage with them effectively through the mutual share of innovative learning.\n\nLearning by sharing knowledge and innovation in the organization and between organizations in the industry is essential today. SME manufacturers and entrepreneurs have tried to adapt to increase their competitiveness by joining groups for knowledge exchange and mutual sharing to create innovations through marketing innovative behavior or MIB to serve the needs of consumers and society as a key (Flavián et al., 2014). However, increasing market orientation focuses on the needs of consumers and seeks competitive advantages from the three business factors. It includes customers to meet their demands and build relationships with them, competition management to establish differentiation, and resource capability or organizational management (Jaworski & Kohli, 1996; Narver et al., 2000), which may lead to competitiveness in SMEs is still questionable.\n\nBuilding competitive advantages for SMEs in the agri-food processing industry enables them to develop their market orientation continually. Promoting knowledge is essential to develop and promote entrepreneurs, particularly SMEs, in the agri-food processing industry in Thailand. These are considered the foundation and have a high proportion of supply chain system mechanisms for 70% of the country’s economic system. SMEs have been an essential source of employment in Thailand. The sector employs approximately 20 million people, 30% of whom are engaged in manufacturing SMEs (Office of SMEs Promotion 2020). In addition, Thai SMEs could develop a local market system to an international level with product quality and marketing innovation, regardless of price (Tanapoompichet & Swasdpeera, 2018).\n\nHowever, one of the challenges in the competitive enhancement of entrepreneurs is still a lack of cooperation in the supply chain system to strengthen and develop the value chain, leading to the development of marketing innovations and enhancing the ability to compete at the macro and micro levels through digital technology. It requires systematic and professional work and intermediaries with international capabilities to develop a wide range of tasks and great support from the government and regulations (Somboonsuke, 2013, pp. 87-90). The easing of rules, regulations, and promotions to build a creative economic system through innovation, according to the World Economic Forum in 2022 (World Economic Forum, 2022), encompasses strategies to promote policy and regulatory reforms to create an enabling environment for SME development and support the creation and strengthening of formal institutions that provide business development and financial services to SMEs on a sustainable basis through innovation with digital technology-based development, resulting in integrative innovations and digital technology for the economy and society in the industry. Although the government had plans and policies to develop and promote such systems, this generally led to the main problem, which was the readiness of SMEs to respond to innovation development, especially in the digital era (Schenker, 2022).\n\nMoreover, the enhancement of market innovation and competitiveness of SMEs in Thailand with digital technology is necessary to study the potential of SMEs for particular products in terms of market innovation behavior and competitiveness. Innovation in marketing has become essential for SMEs to gain competitiveness in the agricultural processed food product industry. SMEs in this sector face many challenges owing to their limited resources, such as finance and technology, and the increasingly competitive market. To compete with larger firms, they must embrace marketing innovation and use digital marketing tools to promote their products effectively. This study developed the following Research Question: Research Question 1 highlights the potential of SME entrepreneurs in the agri-food products industry in terms of market orientation, digital marketing strategy, and marketing innovation behavior. Research Question 2 dealt with current competitiveness. This will enable SMEs to develop effectively owing to marketing innovation behavior and competitiveness driven by digital technology. Research Question 3 discusses digital technology that is acceptable to SMEs. Research Question 4 deals with the type of digital technology that should be applied and how to enhance market orientation, marketing innovation behavior, and competitiveness to achieve digital marketing efficiency.\n\nTo overcome the challenges faced by small- and medium-sized enterprises (SMEs), they require more support and investment from the government and other stakeholders. Some measures that could be taken include improving access to technical expertise and knowledge transfer, investing in infrastructure, and streamlining the supply chain with digital marketing strategies. Promoting processed Thai agri-food products and increasing their visibility in overseas markets is also important, which will help boost the competitiveness of SMEs in this sector. With the help of digital technologies, this can be sustainably achieved.\n\n\nLiterature review\n\nThe study of documents, studies, and literature review is related to five variables: market orientation, digital marketing strategy, competitiveness, marketing innovative behavior, and digital marketing efficiency.\n\nMarket orientation is a classic philosophical marketing concept that focuses on the needs of consumers and society. However, market orientation centers on consumers’ demands and competitive advantages based on three determinants in business factors (Jaworski & Kohli, 1996): customer-focused orientation, competitors and competition capability, and resource-based management or internal management. According to Chinlumprasert and Tawabutr (2019), there is a relationship between market orientation and digital marketing among SMEs in Thailand. The digital marketing strategy in this study applied a digital marketing mix consisting of seven factors: product, price, place, promotion, personnel, process, and physical evidence of digital technology (Kotler & Armstrong, 2020). Additionally, marketing innovative behavior (MIB), according to Scott and Bruce (1994), refers to the following three systematic thinking processes: The first step is idea generation to identify new concepts or ideas to solve problems or find solutions. The first step is idea generation to identify new concepts or ideas to solve problems or find the solution. The second step is idea promotion, which builds a network of cooperation and supports new ideas. The last step is idea realization, which applies new ideas to practice and action.\n\nKaplan proposed a model of enterprise performance, the so-called Balance Score Card (BSC), which consists of modifying a BSC concept as well as a digital marketing concept for the digital marketing efficiency of SMEs as digital enterprise performance. Three factors are involved: learning development, sales, and technology engagement (Kotler & Armstrong, 2020). Porter (1990) finds that competitive strategy and competitiveness are related to the marketing efficiency of business entrepreneurs in a particular country. The key theoretical concept is based on competitive advantage theory, which is a modified diamond model to consider and assess the current situation of significant business environment factors (Porter, 2010) of SMEs in the agri-food processed product sector, consisting of demand conditions, production factor conditions, firm strategy, structure, and rivalry, and related and supporting industries. Another external factor also plays a role in the industry’s competitive advantage, namely, government and agency support.\n\nStudies on market orientation were likely conducted by considering behavior and processes rather than thinking about the relationship between them (Jaworski & Kohli, 1996). Jones (1996) found that the market orientation of small businesses is positively related to customer loyalty, returns on sales, and sales growth in the long term. A relationship was found between market orientation and digital marketing efficiency (Ruekert, 1992; Jaworski & Kohli, 1993; Pelham, 2000; El Mandour & Lahlou, 2021) by measuring digital marketing efficacy in terms of sales growth and profits, market share, organizational engagement, new product development success, and product quality. However, SMEs may have a strong market orientation, and many do not invest sufficient resources in digital marketing, which can limit their growth potential (Chinlumprasert & Tawabutr, 2019).\n\nIn addition, Day and Wenslay (1998) clearly indicated that formulating a marketing strategy based on market orientation requires a balance between customer market orientation, emphasis on competition, and internal activity coordination as the core of meeting customer needs. This study measured digital marketing efficacy in terms of employee learning development, sales, and technology engagement. The results of the above study for research hypothesis 1 are as follows.\n\nMarket orientation positively and directly influenced digital marketing efficiency.\n\nDay (1994) conducted a study on the capabilities of market-driven organizations and found that marketing in the organization was driven by market surveys and the ability to connect with customers. It is supported by Weerawardena, J. and A. O’Cass (2004), who conducted a study on exploring the characteristics of the market-driven firms and antecedents to sustained competitive advantage and found that entrepreneurship played an essential role in leading to market capacity as a mechanism to drive or encourage the organization to seek new markets or marketing capabilities as well as introduce new products based on entrepreneurial conditions. Senior executives would play a role in generating creative ideas and taking a risk in new business operations. The five aspects of comprehensive and reinforced digital marketing competitiveness and capabilities are as follows. First, we investigated an organization’s current market capacity. The second is future capacity demand. Third, bottom-up design of the process; fourth, involved top-down direction and commitment—lastly, creative use of information technology and continuous progress inspection.\n\nThis study also compiled theoretical ideas to explain marketing direction, and Drucker (1985) found that successful entrepreneurs had everything in common: a commitment to systematically managing innovation. For innovation sources, success is the result of attention-seeking innovative opportunities that can be found in a few scenarios. Four sources of opportunity in a firm or industry may come from unexpected events: incompatibility, process requirements, changes in the industry and market, and opportunity sources outside the firm, such as society, government and agency, related and supporting industries, and the environment. These opportunities were business and management challenges such as demographic changes. Changes in perception and the new knowledge and sources mentioned above, both inside and outside the organization, can create opportunities for innovation when the enterprise is at risk. There may be conflicts, and the innovation potential may be in more than one place at the same time, creating innovative opportunities and enhancing the benefits through competitive digital marketing technologies (Wu & Chen, 2021; Wang & Ku, 2020; Zmudzinski & Zaborek, 2018). The research hypotheses can be formulated as follows:\n\nCompetitiveness positively and directly influenced digital marketing efficacy.\n\nStudies on innovative marketing behavior and digital marketing efficiency are as follows: A study by Zahra, Ahmad, and Waheed (2017) found that with marketing innovative behavior in enterprise, employee learning happens through the ethical behavior of organizational leaders, making subordinates or followers perceive and feel the freedom to think creatively in work and to play a role that encourages proactive work and controls the outcome of the work. For SMEs, Haddad, Williams, Hammoud, and Dwyer (2019) found that innovation initiative behavior started from an important element, which is an idea, from both knowledge sharing among employees and communication with customers by relying on the support of senior executives about brainstorming from many parties to test and put in practice and action. Hsu and Wang (2017) examined the impact of digital marketing on innovation in SMEs. The study found that digital marketing strategies such as social media marketing and search engine optimization have a positive impact on marketing innovation in SMEs. They also found that SMEs that implemented digital marketing strategies were more innovative in their marketing approaches and achieved higher levels of business growth. Another study by Ratchford (2019) investigated the impact of digital marketing on innovation in SMEs. The study found that SMEs that implemented digital marketing strategies, such as social media and mobile marketing, were more likely to develop new and innovative marketing approaches. They also found that SMEs that adopted digital marketing strategies had higher levels of business growth.\n\nMarketing innovative behavior is always supported by an innovative culture in the organization as a critical component of the survival and growth of SMEs in terms of competitiveness, offering different value to customers than competitors, resulting in reduced costs and improved workflow efficiency, leading to organizational success (Noefer, Stegmaier, Molter, & Sonntag, 2009; Kong & Li, 2018). On the other hand, some studies confirmed that when they perceived an increased task demand with time pressure, they became stressed leading to work-life imbalances (Attiq, Wahid, Javaid, & Kanwal, 2017; Ren & Zhang, 2015) with employees (Aryee, Walumbwa, Zhou, & Hartnell, 2012; Kong & Li, 2018; Janssen, 2000). The findings of these studies support the idea that there is a strong relationship between digital marketing strategy and marketing innovation of SMEs in agricultural food processed products in Thailand and other countries. By adopting digital marketing strategies, such as social media and mobile marketing, SMEs can be more innovative in their marketing approaches and achieve higher levels of business growth. In turn, this can lead to increased long-term market success. From the above, research hypothesis 3 is established as follows:\n\nMarketing innovative behavior positively and directly influenced digital marketing efficiency.\n\nMarket orientation includes modifying the current customer, current competitor, future customer, future competitor, and marketing technology directions of the business (Apirungruengsakul & Pasunon, 2020). It is indirectly related to marketing achievements through innovative marketing behavior. Henard and Szymanski (2001) find that innovation focuses on the efficiency of creating new products and competitiveness. The organization should be managed to set objectives and strategies and work as a team before introducing new products. Opinions should be in the same direction throughout the organization, as well as participation from various departments to produce ideas and create new products together for a variety of specialties and efficiency. The results of the study are consistent with those of many scholars who found that innovation affects market efficiency and competitiveness (Tsai, 2001; Hult, Snow, & Kandemir, 2003; Hult, Hurley, & Knight, 2004; Taiminen & Karjaluoto, 2015). Indrawati, Caska, and Suarman (2020) found that there are five digital marketing supportive factors of SMEs with priority from highest to lowest: funding, support from the government, business partners, the potential of human resources, and the economic conditions conducive to the efficiency of digital marketing from the integration of SMEs. Carpinetti and Lima (2009) further conclude that there should be a regulatory agency for the integration of SMEs to facilitate collaboration between enterprises in the development of innovation to increase competitiveness, leading to mutual marketing efficiency.\n\nKonjane and Soonthonsmai (2016) found that most SMEs in the agricultural and organic food industry were competitive overall at a high level. Most learned through production innovation exchanges from the public sector. Moreover, there has been a transfer of learning by exchanging production innovation with other entrepreneurs in terms of production technology to SMEs owners, while exchanging marketing innovations was found to have never been learned and never had knowledge exchange on marketing innovations with other entrepreneurs. SME entrepreneurs with different learning through exchanges on marketing innovation had different competitiveness in terms of demand, competitive context, company strategy, government, and opportunities. Also, SMEs entrepreneurship was found to be positively related at a moderate level to the competitiveness of entrepreneurs engaged in the production and distribution of agri-food processed and organic products throughout Thailand. This positive relationship was observed in the exchange of knowledge on production innovation and the transfer of learning through the exchange of marketing innovations.\n\nFor the relationships of innovation and competitiveness with marketing efficiency, the researcher applied Scott and Bruce’s (1994) concept of marketing innovative behavior and Henard and Szymanski’s (2001) study on new product development. The findings revealed that innovation focused on the efficient creation of new products, while competitiveness referred to the organization’s ability to set objectives, develop strategies, and collaborate as a team before introducing new products—what is known as marketing innovative behavior. Opinion-related innovation should be in the same direction throughout the organization, as well as participation from various departments to produce ideas and create new products together for a variety of specialties and efficiency, resulting in the competitiveness of the organization. This is consistent with many scholars who found that innovation is interrelated to competitiveness (Tsai, 2001; Hult, Snow & Kandemir, 2003; Hult et al., 2004).\n\nThe results of the study were also consistent with those of Lee and Hsieh (2010), who found that innovation capability, sustainable competitive advantage, and entrepreneurship indirectly affect sustainable advantages through marketing capability and innovation capability. Market capability did not directly affect sustainable competitive advantage but was indirectly affected by innovation capability. Innovation capability also directly affects competitive advantage (Wongsansukcharoen and Thaweepaiboonwong, 2023).\n\nTherefore, marketing innovation behavior can be a key driver of digital marketing performance for SMEs in agri-food processed products in Thailand (Kaewmong & Soytong, 2020). By developing creative and effective digital marketing campaigns that leverage new digital marketing channels, adapt to changing market conditions, improve product offerings, and enhance customer experience, SMEs can achieve greater engagement, conversions, and long-term success through digital marketing strategies. Digital marketing efficiency or performance through innovative marketing behavior and digital marketing strategies results from a reflection of the efficiency of long-term successful digital marketing to develop potential and competitive advantages and the exchange of knowledge and innovation among the group (Kotler & Armstrong, 2020). Based on the above results, research hypotheses 4, 5, and 6 were established as follows:\n\nMarket orientation positively and indirectly influenced digital marketing efficacy through marketing innovative behavior.\n\nDigital marketing strategy positively and indirectly influenced digital marketing efficacy through marketing innovative behavior.\n\nCompetitiveness positively and indirectly influenced digital marketing efficiency of SMEs through marketing innovative behavior.\n\n\nMethods\n\nAfter conducting a comprehensive literature review on the topic of digital marketing for SMEs in different contexts, the researcher summarized the conceptual framework of the causal relationship model of marketing innovation and the competitiveness of SMEs with digital technologies in Thailand (as illustrated in Figure 1). The framework highlights the interdependence between marketing innovation and digital technologies, and how they contribute to the overall competitiveness of SMEs in the Thai market. This study provides valuable insights for SMEs seeking to enhance their digital marketing strategies and gain a competitive edge in the industry.\n\nThe study focused on entrepreneurs and decision-makers in SMEs as well as leaders and members of community enterprises in agricultural food processed products in Thailand. To ensure statistical accuracy, the sample size for the structural equation modeling (SEM) was 10-15 times the observed variable (Hair, Black, Babin, Anderson & Tatham, 2014), resulting in a sample size of 345. Additionally, 55 samples, or 20% of the samples, were spared in case of data loss, resulting in 361 samples collected. Using 23 variables in the model, this study aimed to provide valuable insights into the factors that influence success in these types of enterprises.\n\nThe samples were selected by purposive sampling using an online questionnaire. The researcher sent a letter to SME entrepreneurs in agri-food processed products to ask for assistance in collecting research data, along with a link to the questionnaire in a Google form format to inform the sample to know the details according to the requirements by using screening questions.\n\nThis is quantitative research as survey research. The data collection instrument was a Likert-scale questionnaire divided into five sections, verified by experts for advice and improvements. The revised questionnaire was then taken to five experts to examine the suitability in terms of content, the language used, and content validity by calculating the index of item objective congruence (IOC), and the IOC obtained passed the criteria, not less than .80 on all items (Hair, Black, Babin, Anderson, & Tatham, 2014). The questionnaire was then used to try out 20 sets of non-sample agricultural product entrepreneurs to check the reliability of the questionnaire by calculating Cronbach’s alpha coefficient for the following five factors. These were market orientation, competitiveness, marketing strategy, marketing innovative behavior, and digital marketing efficiency, resulting in .912, .879, .833, .854, and .861, respectively, which were considered to have standardized reliability coefficients not less than .7 (Cronbach, 1951) and can be used as a tool for data collection. The validity of the factor analysis was corroborated through the KMO, with a value of 0.828 and a highly significant Baltrett sphericity test. The alpha coefficients obtained, which are approximately equal to or higher than .70 are considered acceptable for measuring the variables in this study (Nunnally, 1978).\n\nThe data was collected online to cover consumers and to reduce the limitation of space. Google form was used, through social media platforms in certain groups of SMEs in agri-food processed industry in Thailand.\n\nDescriptive statistics were used to analyze data, such as frequency, percentage, mean, standard deviation, and skew and kurtosis. Inferential statistics included confirmatory factor analysis and structural equation model to examine the consistency of the research model with the empirical data by chi-square (χ2), relative chi-square (χ2/df), goodness-of-fit index (GFI), adjusted goodness-of-fit (AGFI), comparative fit index (CFI), and root mean square residuals (RMSEA). For structural equation analysis, the direct and indirect influences were analyzed according to the conceptual framework by path analysis.\n\nBefore conducting this study, a written request was made to the BUU Ethics Committee for Human Research, Burapha University Institutional Review Board. After reviewing our request, the project was approved (IRB2 – 037/2565) by the ethics committee on April 18, 2022. Participants were given the purpose of the research and were asked to participate in the study; they welcomed it. Written consent was obtained from all participants, and they were informed that their anonymized data would be published.\n\n\nResults\n\nThe analysis results of digital marketing efficacy level with market orientation, digital marketing strategy, competitiveness, and marketing innovative behavior of SMEs were shown based on research objectives as follows (Vuttichat and Patchara, 2023):\n\nThe results of the study on market orientation, digital marketing strategy, competitiveness, marketing innovative behavior, and digital marketing efficiency of SMEs were presented as shown in Table 1.\n\n1 The number of items has been checked for construct validity by measuring relationship model.\n\n2 The mean assessment are as follows: 4.21 – 5.00 = Highest, 3.41 – 4.20 = High, 2.61 – 3.40 = Moderate, 1.81 – 2.60 = Low, 1.00 – 1.80 = Lowest.\n\nThe data presented in Table 1 show that SMEs have a high level of market orientation, digital marketing strategy, competitiveness, innovative marketing behavior, and digital marketing efficiency. The highest mean was found for digital marketing efficiency (X¯ = 4.00), indicating that SMEs effectively utilize digital marketing tools to reach their target audience. Competitiveness also scored high on the mean scale (X¯ = 3.97), suggesting that SMEs can compete well in their respective markets.\n\nHowever, digital marketing strategy has the lowest mean (X¯ = 3.42), which may indicate that SMEs need to improve their overall approach to digital marketing. The standard deviation for the data was between .61-.75, with digital marketing strategy having the highest deviation (S.D. = .75). This suggests a significant amount of variability in the data for digital marketing strategies, which may indicate that SMEs have varying levels of success with their digital marketing strategies.\n\nMarketing innovation behavior also has a high standard deviation (X¯ = .70), indicating that SMEs may have different approaches to innovative marketing techniques. Finally, digital marketing efficacy has the lowest standard deviation (X¯ = .69), suggesting that SMEs are consistently efficient in their digital marketing efforts. Overall, the data suggest that SMEs are generally successful in their digital marketing endeavors but may benefit from a more focused and strategic approach.\n\nThe results of the consistency of the causal relationship model of marketing innovation and competitiveness of SMEs with digital technologies in Thailand based on hypothesis and empirical data according to objective 2 can be shown in Table 2.\n\nTable 2 shows the results of the consistency of the causal relationship model of marketing innovation and competitiveness of SMEs with digital technologies in Thailand based on hypotheses and empirical data. The study found that factor measurement models in the causal relationship model were consistent with empirical data as shown in Figure 2 with χ2/df less than 3, CFI and GFI greater than.95, and RMSEA and SRMR less than 0.05 (Wanichabuncha, 2013).\n\nFigure 2 shows a factor measurement model for the causal relationship model of marketing innovation and competitiveness of SMEs with digital technologies with the empirical data having good consistency. The statistics for measurement were all acceptable (χ2 = 81.14, df = 62, p=.05, χ2/df = 1.31, CFI = 1.00, GFI = .98, AGFI = .93, RMSEA = .03, SRMR = .04) (Wanichabuncha, 2013).\n\nThe analysis results of direct and indirect effects of factors related to digital marketing efficiency of SMEs in agri-food processed products based on objective 3 were shown in Table 3.\n\n** Statistical significance of .01.\n\n* Statistical significance of .05.\n\nThe results of the statistical initial investigation of the data showed that all variables had skewness less than ±3, kurtosis less than ±10, and Kolmogorov-Smirnov is greater than the significance level of .05, indicating that data distribution was normal according to the preliminary agreement of the structural equation model.\n\nTable 3 shows the effect of market orientation (MARPO), digital marketing strategy (STRATEGI), competitiveness (COMPETE), and innovative marketing behaviors (INNOV) on the digital marketing efficiency of SMEs in agri-food processed products as standard values. The hypothesis testing results are as follows:\n\n(H1): Market orientation does not positively or directly influence digital marketing, with an effective coefficient of.07.\n\n(H2): Competitiveness does not positively or directly influence digital marketing efficacy, with an effective coefficient of -.16.\n\n(H3): Innovative marketing behavior positively and directly influences digital marketing, with an effective coefficient of .52.\n\n(H4): Market orientation does not positively or indirectly influence digital marketing efficacy through marketing innovative behavior, with an effective coefficient of -.04. Moreover, market orientation does not positively or directly influence innovative marketing behavior, with an effective coefficient of -.08.\n\n(H5): A digital marketing strategy does not indirectly influence digital marketing efficacy through marketing innovative behavior, with an effective coefficient of .15. However, marketing strategy positively and directly influences innovative marketing behavior, with an effective coefficient of .28.\n\n(H6): Competitiveness indirectly influences digital marketing efficacy through innovative marketing behavior, with an effective coefficient of .39. Competitiveness positively and directly influences innovative marketing behavior, with an effective coefficient of .76.\n\nThe causal relationship model of marketing innovation and the competitiveness of SMEs with digital technologies in Thailand is consistent with the empirical data. For the prediction coefficient (R2), market orientation, digital marketing strategy, and competitiveness all explained the variance in innovative marketing behavior by 89% and the variance in digital marketing efficiency both directly and indirectly through innovative marketing behavior by 18%.\n\n\nDiscussion\n\nThe results of the causal relationship model of marketing innovation and competitiveness of SMEs with digital technologies in Thailand can be discussed based on the following research objectives.\n\nThe results showed that digital marketing efficiency of small and medium enterprises (SMEs) was at a high level. This was consistent with Konjane and Soonthonsmai (2016), who found that most Thai SMEs entrepreneurs had competitiveness overall at a high level with exchange and transfer production innovation and exchange of marketing innovation, which was in line with innovative marketing behavior and digital marketing strategy. This is consistent with Jones (Jones, 1996) who found that when a small enterprise’s market orientation increases, it affects the return on sales and business sales growth. In addition, a study by Amofah, Gyamfi, and Tutu (2016) examined the relationship between marketing mix and marketing efficiency and found that product, price, promotion, personnel, and physical evidence influenced marketing efficiency in terms of customer repurchase decisions. This was supported by Zahra et al. (2017), who found that increased innovative marketing behavior also resulted in increased business performance and market efficiency. Moreover, Haddad et al. (2019) explored strategies for implementing innovation and found that it could reduce costs and optimize processes and management in various areas, including marketing, and affect the survival and growth small and medium enterprises (SMEs) business.\n\nThe causal relationship model of marketing innovation and competitiveness of small and medium enterprises (SMEs) in agri-food products industry with digital technologies in Thailand based on hypothesis and empirical data had a good consistency, consisting of market orientation, digital marketing strategy, marketing innovative behavior, and digital marketing efficacy, indicating market orientation in customer and customer management, competitor and competition management, and resources or organizational management (Jaworski & Kohli, 1996) was related to digital marketing efficiency. This is consistent with studies by several scholars with consistent results (Narver & Slater, 1990; Ruekert, 1992; Jaworski & Kohli, 1993; Pelham, 2000; Narver, Slater, & MacLachlan, 2000).\n\nThis model has been confirmed by studies on marketing strategies in the digital era, where consumption methods have been changing. Causal relationship model of marketing innovation and competitiveness of small and medium-sized enterprises (SMEs) with digital technologies in Thailand. This supports the results of Apirungruengsakul and Pasunon (2020), who found that effective digital marketing should be responsive to consumers’ personal needs by offering promotions on various channels, especially contact and increased convenience for purchasing through an automated program. Marketing innovation behavior is positively related to digital marketing efficiency. This was consistent with a study by Zahra et al. (2017), who found that employees’ learning in the organization often occurs through the ethical behavior of leaders who play a role in motivating proactive work. In addition, Haddad et al. (2019) found that for small and medium enterprises (SMEs), innovations that were key process components included initiatives that required management support to brainstorm ideas from multiple parties and then put them to the test and into practice. This creates an innovative culture in an enterprise that is different from its competitors. This can reduce costs and increase efficiency in work processes, leading to organizational success. However, Attiq et al. (2017), Ren and Zhang (2015), and Kong and Li (2018) found that the relationship may occur in the opposite direction in some organizations and in some highly competitive situations. In other words, market innovative behavior with increasing demands for efficiency may result in time pressures, stress, and work-life imbalances for employees, as the hypothesis testing result of H1, which found that market orientation did not positively and directly influence digital marketing efficiency.\n\nThere are several potential reasons for the lack of a relationship between market orientation and the digital marketing efficacy of SMEs in agri-food processed products in Thailand, such as limited digital marketing expertise, insufficient investment in digital marketing, and lack of differentiation in their products. SMEs in Thailand may need more expertise and knowledge to leverage digital marketing strategies effectively. Even if an enterprise has a strong market orientation, it may need more technical knowledge or resources to implement effective digital marketing campaigns. Most small and medium enterprises (SMEs) in Thailand may need to invest more resources in digital marketing despite having a strong market orientation (Chinlumprasert & Tawabutr, 2019). This may be due to a need for more understanding of the potential benefits of digital marketing or limited financial resources allocated to this area. Additionally, there needed to be more differentiation in the products. Agri-food processed products may be seen as commodities with little differentiation, making it difficult for small and medium enterprises (SMEs) to stand out in the digital space. In such a scenario, digital marketing campaigns may not effectively drive sales or build brand awareness, even if a company has a strong market orientation. Furthermore, market dynamics are a major obstacle (Chinakidzwa & Phiri, 2020). Broader market dynamics may influence the relationship between market orientation and digital marketing performance in Thailand’s agri-food processed products industry. For example, the level of competition or the size of the market may impact the effectiveness of digital marketing campaigns regardless of a company’s market orientation. Similarly, the business performance of small and medium enterprises (SMEs) can be adapted by focusing on the promotion of integrative new idea generation and market leadership orientation (Sriboonlue & Puangpronpitag, 2019).\n\nFurther research is needed to understand why there may be no relationship between market orientation and the digital marketing performance of small and medium-sized enterprises (SMEs) in agricultural food processed products in Thailand. However, a range of factors, including technical expertise, investment, and market dynamics, can influence the effectiveness of digital marketing campaigns in this sector.\n\nHowever, the result of H2 found that competitiveness negatively and directly influenced digital marketing efficiency with an effective coefficient of -.16. This contradicts a study by Weerawardena and O’Cass (2004), who conducted a survey on the characteristics of market-driven businesses in sustainable competitive advantage and found that running conditions played a vital role in leading to marketing capability as a mechanism to drive or promote marketing capability. According to a study by Taiminen and Karjaluoto (2015), SMEs need to gain knowledge and understanding of digital marketing. They can utilize the optimized advantage of the potential of new digital tools and effectively fulfill the benefits of digital developments. This was consistent with Indrawati, Caska, and Suarman (2020), who found five digital marketing supportive factors in order of priority: funding, followed by government support, business partner, human resource potential, and economic conditions. The primary constraint of this negative relationship resulted from the need for more official and continuous essential support from the government in terms of policies to support technological innovation in practical outcomes and network expansion promotion through partnerships in the supply chain. More funding was explicitly needed for technological innovation and its promotion in small and medium enterprises (SMEs). Thus, findings on the causal relationship between competitiveness and digital marketing efficiency need to be clarified and controversial. Therefore, further studies are warranted.\n\nMarketing innovative behavior positively and directly influences digital marketing, according to H3. This was consistent with Prajogo and Ahmed (2006), who examined the relationship between business efficiency and innovation efficiency in manufacturing plants and services in Australia. The study found that innovative efficiency can be achieved through behavioral and cultural development and the organizational environment. Research and development of technology have resulted in innovative efficiency. Process innovation is more closely related to business efficiency than product innovation in manufacturing facilities. Moreover, Konjane and Soonthonsmai (2016) reported that marketing innovation was positively related to competitiveness in the production and distribution of products, leading to good firm performance.\n\nIn addition, Henard and Szymanski (2001) studied new product developments. They found that innovative behavior in management, setting objectives, strategy, and teamwork before introducing a new product was able to build mutual understanding and identify a clear direction, impacting the competitiveness and performance of innovations. This was consistent with many scholars who found that innovative management affects the competitiveness and efficiency of management and marketing performance (Tsai, 2001; Hult et al., 2003; Hult et al., 2004; Tanapoompichet & Swasdpeera, 2018). This result was also consistent with Indrawati et al. (2020), who found that funding, government support, business partners, human resource potential, and economic conditions resulted in the efficiency of digital marketing from the clusters of community enterprise entrepreneurs. Carpinetti and Lima (2009) found that regulatory agencies for clusters of small and medium enterprise (SMEs) entrepreneurs would facilitate collaboration between businesses to increase competitiveness and market efficiency. This is consistent with many scholars who find that innovation is interrelated to market efficiency and competitiveness (Tsai, 2001; Hult et al., 2003; Hult et al., 2004; Taiminen & Karjaluoto, 2015).\n\nThe study’s results found that market orientation did not indirectly influence digital marketing efficiency through innovative marketing behavior based on hypothesis H4, which is inconsistent with Jaworski and Kohli’s study (1996). This may be because small and medium enterprises (SMEs) in agricultural food processed products in Thailand are overreliant on traditional marketing methods. Even if an enterprise has a strong market orientation, proactive and reactive market orientations positively impact digital innovation (Zhao et al., 2023). They may need to be more reliant on traditional marketing methods in highly dynamic markets, such as print advertising or in-store promotions. This may limit their ability to leverage digital marketing channels effectively, even if they have a strong understanding of their target markets. It is possible that broader market dynamics influence the relationship between market orientation and digital marketing performance in the agricultural food processed products industry in Thailand or that the power of social capital positively moderates proactive market orientation and digital innovation. In addition, consumer responsibility has a partial mediating effect (Zhao et al., 2023). For example, the level of competition or the size of the market may impact the effectiveness of digital marketing campaigns, regardless of a company’s market orientation.\n\nThe findings indicate that digital marketing strategies do not indirectly influence digital marketing efficacy through innovative marketing behavior. However, marketing strategy positively and directly influences innovative marketing behavior, according to H5. In general, digital marketing strategy and marketing innovation of SMEs in agri-food processed products in Thailand are interdependent and critical for the business’s success (Kaewmong & Soytong, 2020). There are several reasons for the strong positive relationship between digital marketing strategies and marketing innovation (Gokalp & Cetindamar, 2021; Kaewmong & Soytong, 2020). A clear digital marketing strategy provides direction and guidance for SMEs to develop marketing innovation. By setting specific goals and objectives, a digital marketing strategy helps small and medium enterprises (SMEs) to identify areas of improvement and opportunities for innovation. However, some researchers have found that digital and environmental orientations positively affect product and process innovation performance; digital orientation enhances SMEs’ resources by drawing attention to existing digital resources (Ardito et al., 2021). With a digital marketing strategy, marketing innovation leads to new and creative ways of reaching the target audience, which can improve the digital marketing strategy. By incorporating new marketing tactics and channels, SMEs can enhance their digital marketing strategies and stay ahead of the competition (Wang & Ku, 2020). A digital marketing strategy helps SMEs prioritize marketing innovation initiatives based on their goals and objectives to ensure that the innovation efforts are aligned with the overall digital marketing strategy and differentiate themselves from competitors. This differentiation can help increase brand awareness, customer loyalty, and overall digital marketing performance, which develops long-term and business performance effectiveness that increases competitive advantage (Wongsansukcharoen & Thaweepaiboonwong, 2023) and the likelihood of success.\n\nHowever, competitiveness indirectly influenced digital marketing efficiency through innovative marketing behavior, based on hypothesis H6. Competitiveness in SMEs also creates uniqueness in the product of the business that is different from its competitors (Wongsansukcharoen & Thaweepaiboonwong, 2023). The ability and skills of entrepreneurs to manage resources in the organization benefit through the innovative marketing behaviors of individuals in the enterprise, resulting in business profitability and stability, especially for those who use technology in marketing. This was consistent with the findings of Apirungruengsakul and Pasunon (2020), who found that business marketing technology utilization through competitiveness was indirectly related to marketing success through firms’ innovative marketing behavior.\n\nFor the digital marketing efficiency of SMEs, the results were consistent with Indrawati, Caska, and Suarman (2020), who found that funding, government support, business partners, human resource potential, and economic conditions resulted in the efficiency of digital marketing from the clusters of community enterprise entrepreneurs. Carpinetti and Lima (2009) supported the results that regulatory agencies for clusters of SME entrepreneurs would facilitate collaboration between businesses to increase competitiveness and market efficiency. This was consistent with many scholars who found that competitiveness affected market efficiency with innovation through digital marketing technologies (Hult et al., 2004; Taiminen & Karjaluoto, 2015; Wang & Ku, 2020; Zmudzinski & Zaborek, 2018; Ardito et al., 2021; Wongsansukcharoen & Thaweepaiboonwong, 2023; Zhao et al., 2023).\n\nAccording to the results, the following academic suggestions can be made.\n\nThailand’s agri-food processed product industry is a significant contributor to the country’s economy. SMEs should be aware of the importance of components that contribute to digital marketing efficacy, such as market orientation and digital marketing strategies. These are internal factors, as well as competitiveness, resulting in a combination of internal and external factors to integrating with the innovative marketing behavior of existing enterprises to create digital marketing efficiency. Entrepreneurs should adjust approaches in conducting marketing activities to deal with the digital era by focusing on meeting consumers’ needs by offering promotions on various channels, especially contact, and increasing convenience for purchasing through an automated program, including shopping, payment, and delivery.\n\nSME owners should focus on internal factors, including product marketing strategies, distribution channels, personnel, and marketing communications, to drive digital marketing efficiency. Continually developing skills and applying digital marketing with the use of digital or semi-digital marketing strategies, together with innovative marketing behavior, to invent, improve, and introduce marketing innovations to the market with digital marketing materials. This will result in digital marketing efficacy, including sales, customer satisfaction, loyalty, market share, and the development of a back-office marketing support system for SMEs based on changing situations.\n\nFuture research should be conducted as follows.\n\n1. The relationship between marketing strategy and competitiveness affecting the digital marketing efficiency of small and medium enterprises (SMEs) in manufacturing and other services in the industry’s supply chain should be studied to understand the relational model in the supply chain according to different business contexts.\n\n2. Digital marketing is an essential aspect of modern business activities. To understand the impact of digital marketing on business success, we need to conduct action research among companies that have continuously implemented various forms of digital marketing. This research will help identify the variables that influence the efficacy of digital marketing and how they impact the performance and success of businesses. It will also encourage entrepreneurs to explore their potential and adopt digital marketing models and applications, especially SMEs, which may still need to gain knowledge and understanding of digital marketing strategies.\n\n3. Conducting a policy research study on how to increase the competitiveness and efficiency of SMEs in the sustainable agri-food processed product industry would greatly benefit business owners and stakeholders. By formulating overall strategies, policies, and guidelines for SMEs, we can foster market orientation and develop digital marketing strategies that encourage innovative marketing behavior. This undoubtedly leads to increased marketing efficacy, ultimately resulting in the growth and success of the industry.\n\nSome limitations of this study should be addressed in future research. First, the measurement of variables was based on self-reports and was conducted at one point in time, which might have influenced the research results owing to common method bias. Additionally, it should be noted that this was a cross-sectional study of SMEs in Thailand, so the results may need to be more generalizable to SMEs in other regions. Furthermore, the characteristics of specific industries (such as the agri-food processed products industry) may contain contextual factors that differ from those of other industries. Therefore, to use the results of this research as a reference for further studies, it is important to acknowledge these differences and the limitations they may pose for generalization in different contexts.", "appendix": "Data availability\n\nFigshare: Causal Relationship Model of Marketing Innovation and Competitiveness of SMEs with Digital Technologies in Thailand. https://doi.org/10.6084/m9.figshare.21916899.v1 (Vuttichat and Patchara, 2023).\n\nThis project contains the following underlying data:\n\n• Raw Data.xlsx\n\nFigshare: Causal Relationship Model of Marketing Innovation and Competitiveness of SMEs with Digital Technologies in Thailand. https://doi.org/10.6084/m9.figshare.21916899.v1 (Vuttichat and Patchara, 2023).\n\nThis project contains the following extended data:\n\n• Questionnaire - English.pdf\n\n• Questionnaire - Thai.pdf\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nThe authors would like to acknowledge QUVAE Research and Publications for their invaluable assistance in submitting the underlying data and the extended data to the standard online repository system.\n\n\nReferences\n\nAmofah O, Gyamfi I, Tutu CO: The influence of service marketing mix on customer choice of repeat purchase of restaurant in Kumasi, Ghana. 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Res. 2018; 85: 104–113." }	[ { "id": "330204", "date": "31 Oct 2024", "name": "Vincent Didiek Wiet Aryanto", "expertise": [ "Reviewer Expertise Digital Marketing & Strategic Marketing" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIntroduction Clarity: The introductory section delineates the imperative for small and medium-sized enterprises (SMEs) in Thailand, notably within the agri-food sector, to embrace digital marketing methodologies and enhance their competitive edge. It effectively establishes the pertinence of this investigation by addressing both global and local challenges encountered by SMEs in the realm of digital adaptation. Nonetheless, the focus could be refined by explicitly articulating Thailand-specific obstacles related to digitalization for SMEs. This enhancement would anchor the study more robustly within the local milieu. Structure and Flow: The introduction adheres to a coherent progression, commencing with overarching challenges faced by SMEs and subsequently converging on those pertinent to the agri-food sector in Thailand. The incorporation of a definitive statement of objectives immediately preceding the research inquiries could augment the overall readability. Research Questions: Four research inquiries are posited, each addressing a distinct facet of SME competitiveness and digitalization. These inquiries are pertinent and significant; however, they could be more effectively aligned with particular digital challenges (e.g., the impact of market orientation on digital efficiency within the agri-food sector). Highlighting these subtleties would reinforce the study’s focus.\nLiterature Review Depth and Comprehensiveness: This segment encompasses five pivotal domains: market orientation, digital marketing strategy, competitiveness, marketing innovative behavior, and digital marketing efficiency. Each construct is substantiated by a substantial body of literature, which spans foundational theoretical frameworks to contemporary empirical investigations. The incorporation of established paradigms (e.g., Porter’s competitive advantage and the Balanced Scorecard model) is advantageous, as it situates the research within a well-established theoretical context. Structure and Organization: The literature review would benefit from the introduction of additional subsections, each with distinct headings for the respective concepts, thereby augmenting overall readability. For instance, \"Market Orientation\" might be delineated from \"Digital Marketing Strategies,\" thereby allowing each section to be more prominently featured, which would facilitate readers' comprehension of the argumentative structure. Linking Literature to Research Gap: Although the literature review encompasses pertinent studies, it would be advantageous to more explicitly delineate the research gap. For instance, subsequent to the discourse on market orientation, the narrative could underscore how previous investigations have neglected the convergence of digital marketing and competitive behavior within the Thai agri-food sector. This would accentuate the critical importance of this specific study. Recent Studies and Practical Examples: The inclusion of more contemporary studies—particularly those conducted within the last three years—pertaining to digital marketing and SME competitiveness could render the literature review more contemporaneous. Moreover, citing specific instances of how digital marketing has influenced SMEs in emerging markets would significantly enhance the practical relevance of the review.\nHypotheses Clarity and Justification: The six hypotheses are articulated with precision, where each proposed relationship logically emerges from the comprehensive literature review. Each hypothesis encapsulates the variables under investigation and incorporates an appropriate justification grounded in prior empirical findings. Detailed Justification: Although each hypothesis is substantiated, the manuscript could elaborate on the justification for specific relationships. For example, what underpins the expectation that competitiveness will exert an indirect influence on digital marketing efficiency via marketing innovative behaviour? Offering a more in-depth elucidation (for instance, by referencing studies that demonstrate a link between competitiveness and innovation within comparable SME environments) could enhance the persuasiveness of the hypotheses. Hypotheses Relevance: Hypotheses such as H3 (the impact of marketing innovative behaviour on digital marketing efficiency) hold significant relevance in light of the study’s emphasis on digital efficiency. Including additional commentary regarding the particular significance of these hypotheses for Thailand’s agri-food SMEs would provide further contextualization for this distinctive environment.\nMethodology Research Design: The research methodology is meticulously structured, utilizing a structural equation modelling (SEM) framework, which is particularly advantageous for analyzing intricate relationships among variables. Nonetheless, a more comprehensive rationale for the selection of SEM in preference to alternative statistical methodologies would enhance the clarity of the approach. Sampling and Data Collection: The research employed purposive sampling, specifically targeting representatives from small and medium-sized enterprises (SMEs) via an online survey instrument. Although this method is deemed appropriate, it may potentially introduce sampling bias. A succinct explication of the purposive sampling technique, including a discussion of its limitations, would fortify the methodology section. Furthermore, delineating the inclusion criteria for participants would elucidate the representativeness of the sample. Instrument Validation: The methodology elaborates on the processes of questionnaire development, content validation, and reliability assessment utilizing Cronbach’s alpha coefficient. This comprehensive strategy bolsters the credibility of the research; however, it could benefit from increased transparency regarding the measures undertaken to ensure content validity (for instance, by specifying the areas of expertise of the five consulted experts). Analysis Procedures: The manuscript delineates the statistical tools employed (such as chi-square, GFI, CFI) and provides fit indices for the model. However, supplementary commentary regarding the rationale behind the selection of each index and the criteria for determining an acceptable threshold (for example, the reasoning for selecting a GFI value exceeding 0.95) would render this section more comprehensible for readers who may not be well-versed in SEM methodologies.\nResearch Findings Data Presentation: The findings are delineated within tables, encompassing means, standard deviations, and fit indices corresponding to various variables. This segment is extensive; however, the incorporation of graphical representations (such as bar graphs or line charts) could render the trends and interrelationships more visually comprehensible.\nHypothesis Testing and Interpretation: The outcomes of each hypothesis are encapsulated, with tables illustrating both direct and indirect effects. Nevertheless, for hypotheses that lack support (such as H1 and H2), the manuscript ought to furnish a more comprehensive interpretation. For instance, what could be the underlying reasons for market orientation not exerting a positive impact on digital marketing efficiency? Potential explanations, such as resource constraints or deficiencies in digital competencies, could be briefly conjectured here to provide readers with a deeper understanding of the findings.\nImpact of Findings: The model employed in the study elucidated a significant proportion of the variance in innovative marketing behaviour (89%), which is remarkable. However, an elaboration on the implications of this high percentage for practitioners (e.g., the criticality of nurturing marketing innovation) would augment the practical significance of the findings.\nConclusion Summary of Findings: The conclusion presents a succinct overview of the principal findings, highlighting the beneficial influence of digital marketing strategies on innovative marketing behaviour. Incorporating pertinent statistics or effect sizes in this section (for instance, “digital marketing strategy accounted for 18% of the variance in digital marketing efficiency”) would enhance the significance of the findings.\nImplications: The discourse includes actionable recommendations, advocating for small and medium-sized enterprises (SMEs) to embrace digital strategies while prioritizing innovation. Nevertheless, providing precise recommendations regarding the practical implementation of these strategies by SMEs (such as the utilization of specific digital tools or government incentives aimed at fostering innovation) would augment the document's utility. Furthermore, the text could directly engage policymakers by proposing methods to bolster SMEs through initiatives such as training programs or grant opportunities.\nLimitations and Suggestions for Future Research: The investigation recognizes certain limitations, including the cross-sectional design and the industry-specific emphasis, which restrict its generalizability. However, articulating how subsequent research could remedy these limitations—such as employing longitudinal designs or conducting comparative studies across various industries—would offer more substantial guidance.\nAdditional Recommendations Clarity and Accessibility: The document is replete with information that holds significant academic merit; however, it would greatly benefit from a refinement of language, particularly in intricating segments such as the description of the SEM model. The simplification of certain statistical terminology or the inclusion of explanatory footnotes could render the research more comprehensible to a broader audience.\nConsistency in Terminology: Concepts such as “digital marketing efficiency” and “marketing innovative behaviour” are pivotal yet are employed interchangeably with minor variations. The establishment of consistent terminology would substantially improve clarity.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "330203", "date": "07 Nov 2024", "name": "Johannes Baptista Halik", "expertise": [ "Reviewer Expertise Digital Marketing", "SMEs Performance", "Marketing Management" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nClarity: This section would be helpful to explain why you chose SMEs in the agri-food sector in Thailand. What is the urgency and why this research is important to conduct. Does the impact of SMEs in the agri-food sector in Thailand have a significant impact on the economy in Thailand? Perhaps you should provide relevant data to support your research. This section effectively establishes the relevance of this research by discussing the global and local challenges faced by SMEs in the realm of digital adaptation. However, the focus could be refined by explicitly articulating the Thailand-specific barriers related to SME digitalization. By doing so, it is hoped that the impact of this research will be of interest to agri-food SMEs in Thailand, as well as the Thai government more broadly.\nStructure and Flow: The introduction follows a coherent progression, starting with the overarching challenges faced by SMEs and then moving on to the challenges relevant to the agri-food sector in Thailand. The inclusion of a definitive statement of purpose before the research inquiry would have enhanced the usefulness of this research more comprehensively.\nResearch Questions: Four research questions are proposed, each addressing a different aspect of SME competitiveness and digitalization. These questions are relevant and significant; however, they could be more effectively aligned to specific digital challenges (e.g., the impact of market orientation on digital efficiency in the agri-food sector). Highlighting these complexities would strengthen the research focus.\nRelationship between Literature and Research Gaps: Although the literature review includes relevant studies, it would be better if the research gaps were more explicitly explained. For example, after the discourse on market orientation, the narrative could highlight how previous studies have neglected the convergence of digital marketing and competitive behavior in the Thai agriculture and food sector. This would highlight the importance of this study.\nRecent Studies and Practical Examples: Inclusion of more contemporary studies—especially those conducted within the last five years or more—related to digital marketing and SME competitiveness. This would make the literature review more innovative and contemporary. In addition, citing specific examples of how digital marketing has affected SMEs in emerging markets would significantly increase the practical relevance of the review. You can also include an empirical review based on your observations of the actual conditions in Thailand regarding the use and impact of Digital Marketing on your research object.\nResearch Results Data Presentation: The findings are presented in tables, which include means, standard deviations, and fit indices corresponding to the various variables. This segment is broad; however, incorporating graphical representations (such as bar charts or line graphs) can make trends and interrelationships more visually understandable.\nImpact of Findings: The model used in this study explains most of the variance in innovative marketing behavior (89%), which is remarkable. However, elaborating on the implications of this high percentage for practitioners (e.g., the importance of nurturing marketing innovation) would add practical significance to the findings.\nClarity and Accessibility: The document is full of information of significant academic value; however, it would benefit from some refinement of the language, especially in complex sections such as the description of the SEM models. Simplifying certain statistical terminology or including explanatory footnotes would make the study more accessible to a wider audience.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	2	https://f1000research.com/articles/12-1040
https://f1000research.com/articles/12-281/v1	14 Mar 23	{ "type": "Systematic Review", "title": "General versus spinal anesthesia in percutaneous nephrolithotomy: A systematic review and meta-analysis", "authors": [ "Rinaldo Indra Rachman", "Ponco Birowo", "Ghifari Nurullah", "Prof. Sung Yong Cho", "Widi Atmoko", "Indah Suci Widyahening", "Nur Rasyid", "Rinaldo Indra Rachman", "Ghifari Nurullah", "Prof. Sung Yong Cho", "Widi Atmoko", "Indah Suci Widyahening", "Nur Rasyid" ], "abstract": "Background: Percutaneous nephrolithotomy (PCNL) is the preferred treatment for the removal of large kidney stones, sized >20 mm. However, there is still an ongoing debate concerning the best anesthesia for PCNL. This study aimed to compare the efficacy and safety between general and spinal anesthesia for PCNL. Methods: A systematic review and meta-analysis study. A systematic, electronic literature search was performed in several databases, including PubMed, Scopus, and Google Scholar until July 1st, 2022. The quality of the articles was examined using Crombie's Items (for non-randomized controlled trials (RCTs)) and Jadad Scale (for RCTs). The outcomes assessed were operation time, fluoroscopy time, length of stay, stone-free rate, overall complication rate, specific postoperative complications, cost, pain score, and postoperative analgesic requirement. The article selection was reported based on the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines. We assessed four RCTs and eight retrospective studies. Meta-analysis of selected studies was performed using the Review Manager 5.3. Results: General anesthesia resulted in fewer Clavien–Dindo grade II (OR: 0.68; 95% CI: 0.49 – 0.94; p=0.02), major complications (OR: 0.65; 95% CI: 0.45 – 0.94; p=0.02, and lower transfusion rates (OR: 0.70; 95% CI: 0.53 – 0.94; p=0.02). Whereas spinal anesthesia resulted in faster operation time (Mean Difference: -12.98; 95% CI: -20.56 – -5.41; p<0.001, fluoroscopy time (MD: -26.15; 95% CI: -42.79 – -9.50; p=0.002), reduced length of stay (MD: -0.47; 95% CI: -0.75 – 0.20; p<0.001), and lower postoperative analgesic requirement and cost. No significant difference in stone-free rate (OR: 1.08; 95% CI: 0.92 – 1.26; p=0.37). PCNL performed using either general anesthesia or spinal anesthesia is equally safe and effective. Conclusions: Each method of anesthesia has its own advantages and disadvantages. The final choice between general and spinal anesthesia should be based on the patient's condition and surgical team preference.", "keywords": [ "PCNL", "Spinal Anesthesia", "General Anesthesia", "Complication", "Stone-free Rate" ], "content": "Introduction\n\nNephrolithiasis remains a common health problem around the globe. Its prevalence is 7–13% in North America, 5–9% in Europe, and 1–5% in Asia. According to the European Association of Urology (EAU) and American Urological Association (AUA), percutaneous nephrolithotomy (PCNL) is the first line treatment modality for renal calculi sized >20 mm.1,2 PCNL is also useful for treating multiple stones, staghorn stones, and stones that are resistant to extracorporeal shockwave lithotripsy.1,2 There are variations to PCNL, including position, imaging modality, dilation method, and anesthesia method.3,4\n\nThere is conflicting evidence between the appropriate use of general anesthesia (GA) versus spinal anesthesia (SA) for PCNL. GA was associated with a longer duration of surgery and postoperative length of hospital stay in most studies.3,5–8 GA allows greater flexibility for the anesthesiologist to extend the duration of anesthesia, whereas in SA, this would be more problematic. SA is associated with better postoperative pain control, thereby reducing the need for analgesic drugs.5,8 Some studies have also shown that GA costs more than SA and has a higher rate of complications. The complications usually occur when the patient's position is altered from supine to prone. These complications include brachial plexus injury, spinal cord injury, and lung injury.8,9\n\nThe aim of this study was to evaluate the safety and efficacy profile, in terms of stone free rate, of GA and SA in PCNL.\n\n\nMethods\n\nThis is a systematic review and meta-analysis study evaluating the efficacy and safety of SA compared to GA in PCNL. We included studies that involved patients with renal calculi sized >20 mm who underwent PCNL. The intervention group included patients who were administered SA, whereas the control group were administered GA. The safety profile was determined using the complication rate (classified by the Clavien–Dindo scoring system) and degree of specific complications (headache, urinary tract infection (UTI), urosepsis, and transfusion rate). The efficacy profile was determined using values of stone free rate, operation duration, and length of stay. Any studies that included patients with any renal anomaly, such as horseshoe kidney, malrotated kidney, or ectopic kidney were excluded. Furthermore, this study also excluded studies involving patients with contraindications for SA and GA, such as spinal deformity, severe cardiac and respiratory failure, or severe renal failure.\n\nA systematic search of the literature was performed until July 1st, 2022, using PubMed (RRID:SCR_004846), Scopus (RRID:SCR_022559), and Google Scholar (RRID:SCR_008878) databases. The keywords used were “Spinal, General, Percutaneous Nephrolithotomy, PCNL, PNL” in PubMed, “spinal, general, anesthesia, percutaneous nephrolithotomy” in Scopus, and “spinal, spine, general, PCNL, PNL, percutaneous, nephrolithotomy” in Google Scholar. All keywords were combined using the Boolean logic. The articles identified were then screened for duplicates, which were then removed. The article selection was reported based on the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines.29\n\nTwo reviewers (RIR and PB) examined the articles independently. In case of any disagreement, a discussion was conducted to resolve the matter. The articles were screened for their relevance through the titles and abstracts. The inclusion criteria were comparative studies or randomized controlled trials (RCTs) concerning the use of SA and GA in PCNL. All included articles were written in English. The exclusion criteria were non-comparative studies; studies that combined SA with another method of anesthesia, such as epidural anesthesia; studies with irrelevant outcomes; and studies that included patients with renal anomalies, such as malrotated kidney, horseshoe kidney, or ectopic kidney. The quality of the articles was examined using the Crombie's Items scale (for non-RCTs) and Jadad Scale (for RCTs).10,11\n\nThe process of data extraction of the articles was independently conducted by two reviewers (GN and PB), and any disagreement was resolved by consensus. The variables extracted from the articles were article title, author's name, year of publication, stone free rate, length of stay, operation duration, fluoroscopy time, complication rate (classified using Clavien–Dindo scores), and specific complication rate (headache, UTI, urosepsis, and transfusion rate). Major complication rate was defined by a Clavien–Dindo score 3A or higher. There was no missing data in the data extraction process.\n\nTwo reviewers (RIR and GN) performed data analysis independently. Meta-analysis of selected studies was performed using the Review Manager (RevMan) (RRID:SCR_003581) 5.3 application. Alternatively, meta-analysis of selected studies can be performed using STATA (RRID:SCR_012763). For dichotomous variables, the results were presented as odds ratio (OR) with 95% confidence interval (CI). Whereas for continuous variables, the results were presented as the mean difference with 95% CI. Heterogenicity was analyzed using the chi-squared and I2 test, as appropriate. The data were analyzed using the random-effect model when I2>50% and fixed-effect model when I2<50%. Statistical significance was set at p<0.05. Missing data were analyzed in the outcome. For studies that provided minimum and maximum values instead of standard deviation (SD) for the mean difference analysis, estimated SD was then calculated by the formula provided by Walter and Yao (2007).10 Additionally, for studies that provided 95% CI values instead of SD, SD was then calculated using the formula described in the Cochrane Handbook.12\n\n\nResults\n\nAfter screening the articles and applying the inclusion and exclusion criteria, we found 127 articles from three databases. After removing duplicates, we included 113 studies. Among these, we excluded 90 studies as they were irrelevant based on their titles and abstracts. Subsequently, after assessing the full text, we included 11 studies in the final qualitative and quantitative (meta-analysis) analyses (Figure 1).\n\nA total of four RCTs and eight retrospective studies were assessed. The retrospective studies were assessed using Crombie's items (Table 1) and RCTs were assessed using the Jadad Scale (Table 2).12,13 There were three grade B and four grade C retrospective studies. All the RCTs had a score of less than three. Anesthesia and PCNL methods for each study are presented in Table 3. Furthermore, study characteristics, such as number of patients, stone burden, mean age, stone free rate, follow-up period, and complication rate are presented in Table 4.\n\n* Grade A (6.0-7.0), Grade B (4.0-5.5), Grade C (<4).\n\n* Score below 3 considered as poor.\n\nA total of 4,072 patients were quantitatively analyzed for operation time in the included studies, with 2,393 patients in the SA and 1,679 in the GA group (Figure 2A). High heterogenicity was detected in these studies (I2=94%), and therefore a random-effects model analysis was performed. Pooled data showed that SA had a significantly faster operation time, as compared to GA, using the random-effect model analysis, and the mean difference for SA versus GA was -12.98 minutes (95% CI, 20.56 to -5.41; p=0.0008).\n\n(A) Pooled estimate of operation time using random-effect model; (B) pooled estimate of fluoroscopy time using fixed-effect model; (C) pooled estimate of length of stay using random-effect model; (D) pooled estimate of stone-free rate using fixed-effect model.\n\nThere were two studies that compared fluoroscopy time between the two groups (Figure 2B). A total of 1,120 subjects were included, with 503 in the SA group and 617 in the GA group. The studies were homogenous (I2=0%). The SA group had a significantly faster fluoroscopy time as compared to the GA group, with a fixed effect mean difference of -26.15 minutes (95% CI, -42.79 to -9.50; p=0.002).\n\nThere were eight studies that assessed the length of hospital stay of the patients. A total of 3,843 patients were included, with 2,275 in the SA group and 1,568 in the GA group (Figure 2C). These studies were heterogenous with I2=94%. Patients in the SA group were discharged sooner as compared to those in the GA group. The result of the random-effects model was statistically significant with a mean difference of -0.47 day (95% CI, -0.75 to -0.20; p=0.0008).\n\nThere was a total of nine studies that reported the stone-free rate of patients in both groups. There were 3,953 patients in total, and 2,339 patients belonged to the SA group and 1,614 to the GA group (Figure 2D). Heterogenicity was not found in these studies with I2=0%. Hence, a fixed-effect analysis was performed. There was no significant difference in the stone-free rate between the two groups, with a fixed-effect odds ratio of 1.08 (95% CI, 0.92 to 1.26; p=0.37).\n\nThere were nine studies that reported the overall complication rate (Figure 3A).5,18,20 A total of 3,953 patients were included, of which 2,339 patients were in the SA group and 1,614 patients in the GA group. The studies were homogenous (I2=0%). There was no statistically significant difference in the overall complication rates between the two groups. The fixed-effect model’s odds ratio was 0.93 (95% CI, 0.79 to 1.10; p=0.43).\n\n(A) Pooled estimate of overall complication rate using fixed-effect model; (B) pooled estimate of every Clavien–Dindo classification complication.\n\nFurther subgroup analysis was performed in relation to every Clavien–Dindo classification (Figure 3B). There were 9,090 events of complications noted, of which 5,136 were in the SA group and 3,954 in the GA group. In these studies, one patient could experience more than one complication, resulting in a higher number of events compared to the total number of patients with complications (Figure 3A). Heterogenicity was noted in patients with Clavien–Dindo grade I complications (I2=67%) and major complication rate (I2=70%). There were notably more patients with Clavien–Dindo grade II and major complications in the SA group as compared to the GA group. The result was statistically significant with a fixed-effect model odds ratio of 0.68 (95% CI, 0.49 to 0.94; p=0.02) and random-effect model odds ratio of 0.65 (95% CI, 0.45 to 0.94; p=0.29). However, there were no differences in Clavien–Dindo grade I complication rate with a random-effect model odds ratio of 1.48 (95% CI, 0.66 to 3.33; p=0.34).\n\nFurther analysis of postoperative complications showed that patients in the SA group had higher transfusion rates (Figure 4A). The odds ratio of the fixed-effects model was 0.70 (95% CI, 0.53 to 0.94; p=0.02). There were 11 studies that included transfusion rate as a parameter, with a total of 4,072 subjects, of which 2,398 were in the SA group and 1,674 in the GA group. The studies were not heterogenous (I2=40%).\n\n(A) Pooled estimate of transfusion rate using fixed-effect model; (B) pooled estimate of UTI using fixed-effect model; (C) pooled estimate of urosepsis using fixed-effect model; (D) pooled estimate of postoperative headache using random-effect model. UTI, urinary tract infection.\n\nThere was no significant difference in UTI and urosepsis between the two groups (Figure 4B and C). Meanwhile, patients in the GA group experienced more postoperative headaches (Figure 4D). The odds ratio of the random-effect model was 6.34 (95% CI, 1.76 to 22.81; p=0.005). There were six studies that reported postoperative headache as a complication, with a total of 2,977 patients, of which, 1,680 patients were in the SA group and 1,297 patients were in the GA group. Heterogenicity was noted with I2=61%.\n\nOnly one study discussed the cost difference between the GA and SA groups. Mehrabi et al., stated that GA was notably more expensive than SA. The cost of drugs and materials were USD 5.4±3.1 and USD 23±7.3 for SA and GA, respectively.8\n\nOnly one study discussed the visual analog score (VAS) between both the groups. Karatag et al., showed that there was no significant difference (p=0.365) in VAS in both groups. The VAS of the SA and GA group were 3.0±1.3 and 2.9±1.7, respectively.6\n\nPostoperative analgesic requirement was described in three studies. Overall, these studies stated that patients in the SA group required significantly less postoperative analgesic drugs, as compared to those in the GA group. Gonen et al., found that patients administered SA (53.8±39.8 mg) require significantly less postoperative intravenous tramadol, as compared to those administered GA (111.5±46.3 mg; p<0.001).5 In addition, Mehrabi et al., also stated that patients in the SA group require significantly less postoperative intravenous opioid drugs (unspecified), as compared to those in the GA group on the first (SA: 7.8±2.3 mg, GA: 12.4±3.1 mg; p=0.03) and second (SA: 11.1±2.1 mg, GA: 13.2±2.1 mg; p=0.06) postoperative days.8 However, the difference in opioid drug requirement on the second postoperative day was not significant. Moreover, Bhattarai et al., stated that patients in the SA group (8.2±1.2 mg) require significantly less postoperative analgesic drugs (unspecified) compared to those in the GA group (14.6±2.4 mg; p=0.0001).19\n\n\nDiscussion\n\nThis systematic review and meta-analysis investigated the efficacy and safety profile of SA compared to GA in PCNL. The best study design to evaluate such type of study is RCT. This study included four RCTs. This study finds that SA and GA are both equally safe and effective for PCNL. Key differences are that GA resulted in fewer Clavien-Dindo grade II complications, major complications, and lower transfusion rates. SA resulted in faster operation time, fluoroscopy time, reduced length of stay, and lower postoperative analgesic requirement and cost.\n\nPCNL is the first-line treatment for large nephrolithiasis, sized >20 mm.1,2 This procedure is traditionally performed after administering GA.20 GA may result in fluid absorption and electrolyte imbalance, therefore performing GA in patients with comorbidities could be challenging. For patients with chronic cardio-pulmonary conditions, such as chronic obstructive pulmonary disease or chronic heart failure, SA may be the method of choice for anesthesia.8,21 Moreover, GA has potential adverse effects such as allergic drug reactions, cardiopulmonary compromise, and aspiration of gastric contents. There are several studies in which SA was performed for PCNL candidates who were critically-ill and morbidly-obese to avoid cardiorespiratory compromise during the procedure.9,22,23\n\nTo date, there is no consensus concerning the best mode of anesthesia for PCNL. To address this matter, in 2015, Pu et al., published a meta-analysis comparing GA to regional anesthesia (SA, epidural anesthesia, and spinal-epidural anesthesia).24 To the best of our knowledge, this is the first meta-analysis directly comparing the efficacy and safety of GA and SA in patients who underwent PCNL.\n\nIn a majority of the studies in this review, PCNL was performed in the prone position5–7,13,14,16–18; except for one study by Mehrabi et al., in which PCNL was performed in the supine position.8 PCNL performed in the prone position results in a higher stone-free rate than that performed in the supine position. In regard to the safety profile, performing PCNL in the supine position yields superior results than in the prone position.4 The supine position also makes it easier for anesthesiologists to handle cardiorespiratory emergencies intraoperatively, as compared to the prone position.4\n\nThe authors chose stone-free rate as the study's primary outcome to compare both the methods of anesthesia from a urologist’s perspective; the secondary outcomes are operation time, fluoroscopy time, length of stay, and complications. In this study, we found that GA is superior in terms of lower Clavien–Dindo grade >II complications, and lower transfusion rate and UTIs, whereas SA is superior in terms of shorter operation time, fluoroscopy time, length of stay, and significantly fewer cases of postoperative headache. Both methods are similar in terms of stone-free rate and overall complication rate. Therefore, surgeons can freely choose between GA or SA for PCNL without having to worry about the efficacy of the anesthesia method.\n\nEvery study included in this review reported faster operation time in the SA group as compared to the GA group. The operation and fluoroscopy time was faster in the SA group. In most studies included in this review PCNL was performed in the prone position. When administering GA for PCNL, the patient must be positioned twice. The patient initially must lie in the supine position to be intubated. Thereafter, the patient is pronated for PCNL. This two-stage nature of the procedure may attribute to a longer operation and fluoroscopy time in the GA group.3\n\nLength of hospital stay is shorter in the SA group. This may be attributed to the reduced risk of systemic complications with SA as compared to GA. The method of anesthesia can affect early postoperative recovery for patients. SA is typically performed by administering bupivacaine. A study has demonstrated sensory and motor blockade for 133.16 ± 42.21 minutes after the use of bupivacaine.25 These findings correlate well with the reduced requirement of postoperative analgesics for patients in the SA group as compared to those in the GA group because SA has a lingering effect that lasts post-surgery. A study by Mehrabi et al., showed that on the first postoperative day, patients in the SA group required significantly lower doses of postoperative intravenous opioid drugs, as compared to the GA group.8\n\nOverall complications did not differ between the two groups. In terms of specific complications, patients in the GA group experienced more postoperative headaches as compared to those in the SA group. In patients who were administered GA, postoperative headache, nausea, and vomiting were a common finding.26 A prospective study assessing the postoperative complications of GA in oral and maxillofacial surgery reported that 41% of the subjects experienced postoperative headaches. On the contrary, SA has fewer systemic effects, resulting in less frequently reported postoperative headaches.26\n\nBleeding is a well-known complication in PCNL. In our meta-analysis we found that patients in the SA group had a higher transfusion rate. This result contradicts previous meta-analyses, where SA is associated with a significant decrease in blood loss compared to GA in surgeries within the pelvic, abdominal, and thoracic cavities and lower extremities.27\n\nMehrabi et al., reported that GA is more expensive than SA. The cost of drugs and materials are USD 5.4 ± 3.1 (SA) and USD 23 ± 7.3 (GA). Previous studies on orthopedic surgeries have also demonstrated that SA is relatively less expensive as compared to GA.28 Possible heterogeneity in this study may be caused by differences in sample size, anesthesia drug, and stone fragmentation method.\n\nThe limitation of this study is that there are few articles that have reported fluoroscopy time in both the groups. Postoperative analgesic consumption was also difficult to compare between the studies because there was no uniform term for the definition of postoperative analgesic consumption. Few studies have reported major complication rates using the Clavien–Dindo classification, including the incidence of urosepsis. There are only four RCTs included, and the rest of the studies are retrospective studies. Therefore, there could be an overestimation of the result due to selection bias. The RCTs assessed had low scores due to lack of blinding. However, double blinding is no applicable in the studies. Patient must provide consent to be included in the study and anesthesiologists performing anesthesia must be prepared for the upcoming procedure to ensure patient safety. Therefore, the anesthesia modality must be specified prior to the procedure. In this case, a third observer must be involved to assess the outcomes of the subjects without knowing what group they are in to ensure objectivity. There is a limitation in the review process, which is that there were only two reviewers of the articles in this study.\n\nHowever, this study addresses the advantages and disadvantages of both GA and SA in PCNL, with an aim to provide valuable insights for surgeons to choose the most appropriate method of anesthesia for PCNL.\n\n\nConclusions\n\nIn terms of efficacy marked by stone-free rate, there are no significant differences between GA and SA. GA is superior in terms of lower Clavien–Dindo grade II complications, transfusion rate, and UTI occurrence, whereas SA is superior in terms of shorter operation time, fluoroscopy time, length of stay, and a significantly lower frequency of postoperative headaches. Both methods are similar in terms of the overall complication rate. Therefore, the decision to choose between GA and SA should be based on the patient's clinical parameters and the surgical team’s preferences.\n\n\nData availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\nOpen Science Framework: PRISMA checklist for article 'General versus spinal anesthesia in percutaneous nephrolithotomy: A systematic review and meta-analysis', https://doi.org/10.17605/OSF.IO/7KR58.29\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).", "appendix": "References\n\nTürk C, Neisius A, Petrik A, et al.: EAU Guidelines on Urolithiasis. Eur. Assoc. Urol. 2018; 2018: 1–87.Reference Source\n\nNelson CP, Pace KT Jr, Pearle MS, et al.: Surgical Management of stones. Am. Urol. Assoc. 2016; (April): 1–50.\n\nKaraolides T, Moraitis K, Bach C, et al.: Positions for percutaneous nephrolithotomy: Thirty-five years of evolution. Arab. J. Urol. 2012; 10(3): 307–316. PubMed Abstract \| Publisher Full Text\n\nBirowo P, Tendi W, Widyahening IS, et al.: Supine versus prone position in percutaneous nephrolithotomy: A systematic review and meta-analysis. F1000Res. 2020; 9: 1–21. Publisher Full Text\n\nGonen M, Basaran B: Tubeless percutaneous nephrolithotomy: Spinal versus general anesthesia. Urol. J. 2014; 11(1): 1211–1215. PubMed Abstract\n\nKaratag T, Tepeler A, Buldu I, et al.: Is micro-percutaneous nephrolithotomy surgery technically feasible and efficient under spinal anesthesia? Urolithiasis. 2015; 43(3): 249–254. PubMed Abstract \| Publisher Full Text\n\nShah R, Thapa AS, Lamichhane N, et al.: Safety and efficacy of spinal anaesthesia in percutaneous nephrolithotomy. J. Nepal Med. Assoc. 2016; 55(204): 61–66. PubMed Abstract \| Publisher Full Text\n\nMehrabi S, Zadeh AM, Toori MA, et al.: General versus spinal anesthesia in percutaneous nephrolithotomy. Urol. J. 2013; 10(1): 756–761. PubMed Abstract\n\nAravantinos E, Karatzas A, Gravas S, et al.: Feasibility of Percutaneous Nephrolithotomy under Assisted Local Anaesthesia: A Prospective Study on Selected Patients with Upper Urinary Tract Obstruction. Eur. Urol. 2007; 51(1): 224–228. PubMed Abstract \| Publisher Full Text\n\nCrombie I: The pocket guide to critical appraisal: a handbook for health care professionals. London: BMJ. 1996.\n\nHalpern SH, Douglas MJ: Appendix: Jadad Scale for Reporting Randomized Controlled Trials. Evidence-based Obstet. Anesth. 2007; 237–238.\n\nHiggins J, Green S: Cochrane Handbook for Systematic Reviews of Interventions. The Cochrane Collaboration.2011.\n\nCicek T, Gonulalan U, Dogan R, et al.: Spinal anesthesia is an efficient and safe anesthetic method for percutaneous nephrolithotomy. Urology. 2014; 83(1): 50–55. PubMed Abstract \| Publisher Full Text\n\nSolakhan M, Bulut E, Ga D, et al.: Comparison of Two Different Anesthesia Methods in Patients Undergoing Percutaneous Nephrolithotomy. Endourol. Stone Dis. 2019; 16(03): 246–250.\n\nAstram A, Rasyid N, Birowo P, et al.: Success of Percutaneous Nephrolithotomy: Comparing Spinal Anesthesia with General Anesthesia. Indones. J. Urol. 2015; 22(2).\n\nBuldu I, Tepeler A, Kaynar M, et al.: Comparison of anesthesia methods in treatment of staghorn kidney stones with percutaneous nephrolithotomy. Urol. J. 2016; 13(1): 2479–2483. PubMed Abstract\n\nNouralizadeh A, Ziaee SAM, Hosseini Sharifi SH, et al.: Comparison of percutaneous nephrolithotomy under spinal versus general anesthesia: A randomized clinical trial. J. Endourol. 2013; 27(8): 974–978. Publisher Full Text\n\nMovasseghi G, Hassani V, Mohaghegh MR, et al.: Comparison between spinal and general anesthesia in percutaneous nephrolithotomy. Anesthesiol. Pain Med. 2014; 4(1): 1–6.\n\nBhattarai R, Das C, Paudel B, et al.: Comparison of General versus Spinal Anesthesia in Patients undergoing Percutaneous Nephrolithotomy: A Prospective Randomized Study. J. Nobel Med. Coll. 2016; 5(1): 37–42. Publisher Full Text\n\nLingeman JE, Matlaga BREA:Surgical management of upper urinary tract calculy.Wein AJ, Kacoussi LRNA, editors. Campbell-Walsh Urology. 9th ed.Philadelphia:Saunders Elsevier;2007; p. 1431–507.\n\nKuzgunbay B, Turunc T, Akin S, et al.: Percutaneous nephrolithotomy under general versus combined spinal-epidural anesthesia. J. Endourol. 2009; 23(11): 1835–1838. Publisher Full Text\n\nMehrabi S, Shirazi KK: Results and complications of spinal anesthesia in percutaneous nephrolithotomy. Urol. J. 2010; 7(1): 22–25. PubMed Abstract\n\nAndroniki K, Hassan R: Percutaneous nephrolithotomy under conscious sedation in morbidly obese patients. Can. J. Urol. 2006; 13(3): 3153–3155.\n\nPu C, Wang J, Tang Y, et al.: The efficacy and safety of percutaneous nephrolithotomy under general versus regional anesthesia: a systematic review and meta-analysis. Urolithiasis. 2015; 43(5): 455–466. PubMed Abstract \| Publisher Full Text\n\nMonika M, Kiwi M, Madhu S: Dhawan Sonali, Sethia Sangeeta MA. General versus spinal anesthesia in percutaneous nephrolithotomy: A comparative study. IAIM. 2017; 4(9): 59–66.\n\nAkhter LP, Wani NA, Ain Q, et al.: Common postoperative complications after general anesthesia in oral and maxillofacial surgery. Natl. J. Maxillofac. Surg. 2021; 12(2): 206–210.\n\nRichman JM, Rowlingson AJ, Maine DN, et al.: Does neuraxial anesthesia reduce intraoperative blood loss? A meta-analysis.2006; 427–35.\n\nGonano C, Leitgeb U, Sitzwohl C, et al.: Spinal versus general anesthesia for orthopedic surgery: Anesthesia drug and supply costs. Anesth. Analg. 2006; 102(2): 524–529. PubMed Abstract \| Publisher Full Text\n\nRachman RI: General versus spinal anesthesia in percutaneous nephrolithotomy: A systematic review and meta-analysis. [Dataset].2022, August 20. Publisher Full Text" }	[ { "id": "167852", "date": "02 May 2023", "name": "Noor Buchholz", "expertise": [], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nSome criticisms:\nIntroduction:\n\nYou write that some studies have also shown that GA costs more than SA. In Results it is written that only one study discussed the cost difference between the GA and SA groups. That does not fit together.\nMethods:\nThe age and gender of the subjects in the individual studies were not specified.\n\nNo indication of group sizes under methods.\n\nAccording to the size of the groups in Table 4, there are clear size differences in some retrospective studies between GA and spinal anesthesia. Why? Could that be a bias?\n\nThe objective/ the target points are described in more detail in the abstract than in the actual review.\n\nOne of the targets, the Clavien-Dindo scoring system, could possibly be briefly explained.\nResults:\nUnfortunately, the results section is divided and presented in a very confusing manner. In addition, a very confusing distribution of the results across Table 4 and Figures 2, 3 and 4. The results should perhaps be presented consistently either as in Table 4 or in Figures 2, 3 and 4. Presentation better not in the text, but possibly at the end.\n\nLiterature search and study characteristics should perhaps be listed under Methods.\n\nNumber of studies varies again and again in varying numbers; sometimes 11, then again 12.\n\nThe mean age of the subjects is mentioned under study characteristics, but no information was actually given in Table 4.\n\nIn some cases only individual result points are evaluated from the individual studies. Why were these studies not excluded, contrary to statements under Literature search. And why were items, such as costs (discussed only in one study), not removed from the review?\n\nHomogeneity or heterogeneity is mentioned again and again among the various result points. What kind of homogeneity/heterogeneity of the study in relation to what is meant by it?\n\nUnfortunately, only one or three studies seem to be somewhat incomplete to provide a comprehensive overview of the pain score and the postoperative analgesic requirement.\n\nUnlike the other parts of the results, there are no figures or tables for the pain score and the postoperative analgesic requirement.\n\nWith regard to the postoperative analgesic requirement, only opioid administration is mentioned. But not from possible other painkillers, which could falsify the statement.\n\nAre the rationale for, and objectives of, the Systematic Review clearly stated? Yes\n\nAre sufficient details of the methods and analysis provided to allow replication by others? No\n\nIs the statistical analysis and its interpretation appropriate? Not applicable\n\nAre the conclusions drawn adequately supported by the results presented in the review? Partly", "responses": [ { "c_id": "10474", "date": "29 Nov 2023", "name": "Rinaldo Indra Rachman", "role": "Author Response", "response": "Introduction: Reviewer's comment: You write that some studies have also shown that GA costs more than SA. In Results it is written that only one study discussed the cost difference between the GA and SA groups. That does not fit together. Author's response: Cost and Visual analogue score was an additional important finding of this study. However forest plot was unable to be generated because only one study addressed each of this matter. Therefore cost and visual analogue score is better suited to be included in the discussion rather than result section (already revised) Methods: Reviewer's comment: The age and gender of the subjects in the individual studies were not specified. Author's response: Has been added to table 4 Reviewer's comment: According to the size of the groups in Table 4, there are clear size differences in some retrospective studies between GA and spinal anesthesia. Why? Could that be a bias? Author's response: In table 4, the study by Astram, et al included 540 patients with spinal anestesia and 220 patients with general anesthesia. There are more than twice patients in general anesthesia group because this study included all patients who underwent PCNL during 2000 - 2011 period. The same gues for the study by Cicek et al which divided samples into two groups GA (n=564) and SA (n=440). This study included all patients who underwent PCNL between 2004-2011. Both studies mentioned the decision of using GA or SA is based on each patient's characteristics, however they did not specify the characteristics. The difference of sample size between the two group may result in bias Reviewer's comment: The objective/ the target points are described in more detail in the abstract than in the actual review. Author's response: The objective of the study is now clearly mentioned in the methods section Reviewer's comment: One of the targets, the Clavien-Dindo scoring system, could possibly be briefly explained Author's response: has been added to the methods section Results: Reviewer's comment: Unfortunately, the results section is divided and presented in a very confusing manner. In addition, a very confusing distribution of the results across Table 4 and Figures 2, 3 and 4. The results should perhaps be presented consistently either as in Table 4 or in Figures 2, 3 and 4. Presentation better not in the text, but possibly at the end. Author's response: All tables and figures in the study has it's own purpose. Table 4 serves to describe the baseline characteristics of the studies, wherease Figure 2-4 serves to explain the finding of the Meta-Analysis Reviewer's comment: Literature search and study characteristics should perhaps be listed under Methods Author's response: literature search has been listed under methods. Study characteristics is listed under results, as this is the finding of this article Reviewer's comment:Number of studies varies again and again in varying numbers; sometimes 11, then again 12 Author's response: the correct number of study is 11 Reviewer's comment: The mean age of the subjects is mentioned under study characteristics, but no information was actually given in Table 4 Author's response: mean age of the subjects has been added to table 4 Reviewer's comment: In some cases only individual result points are evaluated from the individual studies. Why were these studies not excluded, contrary to statements under Literature search. And why were items, such as costs (discussed only in one study), not removed from the review? Author's response: Cost and Visual analogue score was an additional important finding of this study. However forest plot was unable to be generated because only one study addressed each of this matter. Therefore cost and visual analogue score is better suited to be included in the discussion rather than result section (already revised) Reviewer's comment: Homogeneity or heterogeneity is mentioned again and again among the various result points. What kind of homogeneity/heterogeneity of the study in relation to what is meant by it? Author's response: Homogeinity and heterogeinity was mention to address the appropriate analysis using random-effect model for variables with I2 >50. Whereas variables with I2<50 was analyzed using fixed-effect model Reviewer's comment: Unfortunately, only one or three studies seem to be somewhat incomplete to provide a comprehensive overview of the pain score and the postoperative analgesic requirement. Reviewer's comment: Unlike the other parts of the results, there are no figures or tables for the pain score and the postoperative analgesic requirement Author's response: Visual analogue score was an additional important finding of this study. However forest plot was unable to be generated because only one study addressed each of this matter. Therefore cost and visual analogue score is better suited to be included in the discussion rather than result section (already revised) Reviewer's comment: With regard to the postoperative analgesic requirement, only opioid administration is mentioned. But not from possible other painkillers, which could falsify the statement Author's response: Bhattarai, et al gives the information of 2 drugs used, which are paracetamol 1 gram & morphine injection. Mehrabi et al only specified the need for opioid drugs postoperatively without mentioning other analgesics. Gonen et al stated that they chose tramadol to be given intravenously for postoperative pain management. This is perhaps because of opioid is a marker for significant" } ] }, { "id": "171948", "date": "13 Jul 2023", "name": "M. Hammad Ather", "expertise": [ "Reviewer Expertise Endourology and Uro oncology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nAuthors have performed a meta analysis comparing spinal anesthesia with the standard GA in performing PCNL for >20mm kidney stones. They noted that there is no clear winner and both can be safely used with matching efficacy. Authors have noted that the quality of studies and limited number of RCTs rendered the conclusions rather weak. In the discussion section authors could not provide a compelling evidence of better complication rate in GA patients and shorter fluoroscopy and operative time in regional anesthesia. They also failed to identify which type is suited when, which perhaps is the most important question for the clinicians. It is also important for the future investigators to assess operator experience (both urologist and anesthetist) with regional anesthesia and also potential difficulties and monitoring of the patient undergoing prone PCNL under regional anesthesia. In a recently reported RCT on 90 patients authors noted no major difference in the two types (GA and combined spinal epidural).\n\nAre the rationale for, and objectives of, the Systematic Review clearly stated? No\n\nAre sufficient details of the methods and analysis provided to allow replication by others? Yes\n\nIs the statistical analysis and its interpretation appropriate? Yes\n\nAre the conclusions drawn adequately supported by the results presented in the review? Partly", "responses": [ { "c_id": "10473", "date": "29 Nov 2023", "name": "Rinaldo Indra Rachman", "role": "Author Response", "response": "Spinal anesthesia is generally preferred than general anesthesia, as SA yields minimal systemic effect compared to GA. Using GA increases the risk of anaphylaxis because of the drug used in GA. However GA eases the procedure by controlling the breathing and renal movement of the patient." } ] } ]	1	https://f1000research.com/articles/12-281
https://f1000research.com/articles/12-1430/v1	01 Nov 23	{ "type": "Method Article", "title": "Efforts to enhance reproducibility in a human performance research project", "authors": [ "Jeffrey A. Drocco", "Kyle Halliday", "Benjamin J. Stewart", "Sarah H. Sandholtz", "Michael D. Morrison", "James B. Thissen", "Nicholas A. Be", "Christopher E. Zwilling", "Ramsey R. Wilcox", "Steven A. Culpepper", "Aron K. Barbey", "Crystal J. Jaing", "Kyle Halliday", "Benjamin J. Stewart", "Sarah H. Sandholtz", "Michael D. Morrison", "James B. Thissen", "Nicholas A. Be", "Christopher E. Zwilling", "Ramsey R. Wilcox", "Steven A. Culpepper", "Aron K. Barbey", "Crystal J. Jaing" ], "abstract": "Background: Ensuring the validity of results from funded programs is a critical concern for agencies that sponsor biological research. In recent years, the open science movement has sought to promote reproducibility by encouraging sharing not only of finished manuscripts but also of data and code supporting their findings. While these innovations have lent support to third-party efforts to replicate calculations underlying key results in the scientific literature, fields of inquiry where privacy considerations or other sensitivities preclude the broad distribution of raw data or analysis may require a more targeted approach to promote the quality of research output.\nMethods: We describe efforts oriented toward this goal that were implemented in one human performance research program, Measuring Biological Aptitude, organized by the Defense Advanced Research Project Agency's Biological Technologies Office. Our team implemented a four-pronged independent verification and validation (IV&V) strategy including 1) a centralized data storage and exchange platform, 2) quality assurance and quality control (QA/QC) of data collection, 3) test and evaluation of performer models, and 4) an archival software and data repository.\nResults: Our IV&V plan was carried out with assistance from both the funding agency and participating teams of researchers. QA/QC of data acquisition aided in process improvement and the flagging of experimental errors. Holdout validation set tests provided an independent gauge of model performance.\nConclusions: In circumstances that do not support a fully open approach to scientific criticism, standing up independent teams to cross-check and validate the results generated by primary investigators can be an important tool to promote reproducibility of results.", "keywords": [ "reproducibility of results", "validation studies", "evaluation methodology", "data quality" ], "content": "Introduction\n\nReproducibility of findings is a fundamental requirement of any scientific research endeavor. Nevertheless, for a variety of reasons, reproducibility remains a challenge in many areas of the life sciences.1 Batch effects, hidden variables, and low signal-to-noise ratios may interfere with researchers’ ability to draw broad conclusions based on small quantities of data.2 Similarly, data snooping may, even unintentionally, lead to the selection of models that have poor generalizability outside an original dataset.3 These difficulties can be exacerbated in exploratory studies that are by design not limited to the consideration of only one or a small number of pre-specified hypotheses, but rather constructed for the purpose of testing a very large number of possible explanatory variables using a statistical approach.\n\nTransparency in data collection and analysis has been suggested as a potential means of bringing to light methodological or other flaws that may impair the reproducibility of results in various areas of biomedical research. For example, authors who distribute notebooks integrating data, code, and text directly enable others to replicate some or all of the analysis supporting their stated conclusions.4 While such measures do not exclude all possible errors that might call into question the reliability of published findings, scientists adhering to these practices considerably reduce the ambiguity associated with the steps in their workflow subsequent to data acquisition.5,6\n\nOrganizations that fund research must take into account these considerations and others in planning new research and development (R&D) programs. The time and money available to obtain answers to the scientific questions of stakeholder interest are generally limited. Reachback tasking that would allow re-analysis of data or models following an original period of performance is not always possible, as studies frequently rely on teams assembled in an ad hoc manner to respond to the requirements of a specific project call. Moreover, publication of results is not a guarantee that the artifacts of a research program will be fully preserved. While many journals have adopted standards for sharing of data and code, compliance with these policies is imperfect.7,8 Indeed, selective publication practices have themselves been implicated as potential sources of bias in the scientific literature.9,10 Finally, the aspirations of open science may conflict with project constraints when supporting data are not suitable for release into the public domain.\n\nHere we describe the efforts of one research program, Measuring Biological Aptitude (MBA), a four-year effort sponsored by the Biological Technologies Office of the Defense Advanced Research Projects Agency (DARPA), to improve the reproducibility of studies performed with the goal of optimizing human performance in a variety of athletic and cognitive military skills tests. As the MBA program involved data encumbered by restrictions related to personal privacy, medical confidentiality, and national defense, a fully open approach to promoting reproducibility was not practical. Instead, the program sponsored the authors of this manuscript to conduct a comprehensive independent verification and validation (IV&V) program to test and evaluate the results generated by the primary program contractors and modeling teams.\n\nDARPA defines IV&V as “the verification and validation of a system or software product by an organization that is technically, managerially, and financially independent from the organization responsible for developing the product” (DARPA Instruction 70). In recent years, IV&V has become a key component of various DARPA research programs both inside and outside the life sciences domain.11 In contrast to the standard in open science, which generally relies on the free and voluntary participation of members of the scientific community to verify the results of third party studies, DARPA’s policy suggests that independent efforts to support the integrity of scientific results are of sufficient importance to merit direct funding, using teams selected for their expertise in the relevant technical areas.\n\nAccording to the MBA Broad Agency Announcement (BAA), primary performers were charged to “identify, understand, and measure the expression circuits (e.g., genetic, epigenetic, metabolomic, etc.) that shape a warfighter’s cognitive, behavioral, and physical traits, or phenotypes, related to performance across a set of career specializations.” The IV&V team, by contrast, was directed to “verify and validate whether the expression circuits, as measured by the molecular targets identified, directly correlate to dynamic changes in performance traits in the individual and independently confirm…that those circuits correlate to selection success or failure.” From this followed a corresponding but distinct schedule of tasks for each group (Figure 1).\n\nIn 2019, DARPA selected Lawrence Livermore National Laboratory (LLNL) and the University of Illinois Urbana-Champaign (UIUC) to lead the IV&V component of the MBA program. According to the IV&V plan developed by LLNL, the effort comprised four core focus areas: 1) a centralized secure data storage and exchange platform, 2) quality assurance and quality control checklists applied to data acquisition, 3) test and evaluation of performer modeling products, and 4) an archival software repository and data store.\n\n\nMethods\n\nWhile the unit costs of bioinformatic data collection have declined in recent years, the acquisition and processing of large-scale omics data remain both financially and computationally expensive.12 For reasons of reliability and cost savings, research sponsors may desire a centralized user facility for data storage and pre-processing, even in projects that involve multiple competing investigators and modeling teams. Moreover, if program managers wish to obtain a comparison of performance across several predictive models, centralized data services may help to maximize the time that data scientists and statisticians are able to devote to model selection while minimizing the risk that ambiguities in outcome labels or other metadata may lead different groups to substantially varying interpretations of the same modeling problem.13,14\n\nA separate consideration in research involving human subjects involves data security and privacy. In the U.S., federal regulations require institutional review boards (IRBs) to evaluate each proposed project’s provisions for protecting the privacy and confidentiality of human subjects information, regardless of whether a study explicitly plans to include data that is covered by other medical privacy laws.15 In addition, considerations such as the possible re-identification of putatively de-identified health data may warrant additional data protection precautions even when not required by statute or regulation.16\n\nTo ensure both data security and data consistency for all teams working on the project, LLNL built a computing enclave for storing and analyzing MBA program data (Figure 2). Following best practices employed by other centralized biomedical data repositories, the enclave implemented cyber security controls at the FISMA Moderate policy level with enhanced controls from the NIST 800-53 and 800-66 information security guidelines for privacy and HIPAA compliance.17,18,19 All accredited users of the enclave were required to complete cyber security training and a human subjects protection course prior to receiving computing accounts.20 Access to the enclave was established through multifactor authentication.\n\nTo minimize risks of intentional and/or accidental duplication of human subjects data, the enclave featured a Virtual Network Computing (VNC) portal through which external collaborators could interact with program data. As the enclave excluded other networking protocols for ordinary users, the visual interface allowed modelers to perform analyses in a standard Linux computing environment while imposing a soft barrier against the bulk download of sensitive data. Modelers were allowed to upload new data or software dependencies to the enclave via the data transfer node. However, while outbound transfers of finished analysis were supported via the same pathway, these required the additional step of administrator review and approval.\n\nTo permit utilization of high-performance computing (HPC) systems, the LLNL secure enclave was extended to include the Livermore Computing Collaboration Zone (CZ; https://hpc.llnl.gov/hardware/zones-k-enclave). This enabled analysis of multiple omics datasets using leadership-class compute platforms such as Mammoth, a 8,800 core cluster acquired via the National Nuclear Security Administration’s Advanced Simulation and Computing (ASC) Program.\n\nIn recent years, various scientific disciplines and consortia have developed minimum standards for the inclusion of data in both centralized repositories and published meta-analyses.21 These guidelines have encompassed a range of data acquisition formats, including genetic, proteomic, and other biochemical data.22,23 For example, the Human Proteome Organization’s Proteomics Standards Initiative developed the Minimum Information About a Proteomics Experiment (MIAPE) standard for mass spectrometry experiments involving protein and peptide identification.24 Similarly, in the human subjects field, the STrengthening the Reporting of OBservational studies in Epidemiology (STROBE) statement set minimum metadata standards for collection, archiving, and reporting of epidemiological research data.25\n\nOther standards-setting organizations go beyond simple metadata reporting requirements and seek to detail comprehensive processes and systems that can help ensure data quality (quality assurance, or QA) as well as specific tests and benchmarks that can flag errors during and after data collection (quality control, or QC).26 These types of procedural checks have been adopted, for example, by the Metabolomics Quality Assurance and Quality Control Consortium (mQACC) and the Encyclopedia of DNA Elements (ENCODE) Consortium.27,28 The MBA program used this type of standard as a model for its own QA/QC efforts.\n\nLLNL experimentalists generated a scoring rubric for each of the molecular and omics data collection modalities employed in the various human trials throughout MBA. Example rubrics are shown in Table 1 and Extended Data Tables 1-3. A pass/fail checklist was used to determine whether each dataset met the minimum quality standards for use by the modeling teams. Some criteria involved best practices in sample handling and study design, while others were specific to the instrumentation used and, in general, followed manufacturer recommendations. For some types of data collection, including genome sequencing data, open source tools such as multiQC were utilized as components of the scoring framework.29 Following the IV&V team’s evaluation of each dataset against the rubrics, a scorecard was transmitted to the performer team or subcontractor responsible for the data collection, and the program office was consulted for a final determination on the inclusion of the dataset in the modeling corpus.\n\nReference ranges are derived from ’CytoFLEX Platform Instructions for Use,’ Beckman Coulter, rev. 12/11/2019. In addition to the pass/fail ranges, some rubrics included a ‘warn’ range for borderline data.\n\nAdditionally, UIUC statisticians developed separate sets of metrics for the phenotypic and behavioral data collected during the course of the program. These rubrics were crafted to flag outliers and diagnose other potential data quality issues. The analyses encompassed five general domains: cognition, demographics, human performance, personality, and wearable sensors (see Table 2). Some metrics applied to only a single domain, whereas others (e.g., missing data) were relevant for multiple domains. When there is a quality assurance failure, the Potential Issue column of Table 2 provides plausible mechanisms that may underlie the faulty data collection process. The team also developed customized R software scripts that read in the data and automatically generated tables, figures, and reports.\n\nDomains Assessed: Cog=Cognition; Demo=Demographics; HP=Human Performance; Per=Personality; WS=Wearable Sensors.\n\nThe primary deliverable from the MBA IV&V effort was the test and evaluation of performer expression circuit models used to predict achievement on military skills tests. While performers trained statistical models to predict pass/fail outcomes for different candidates on a battery of human performance and cognitive tests, the IV&V team was responsible for certifying to the military cadre that the selected molecular observables were in fact predictive of the chosen outcomes.\n\nSeveral factors complicated the evaluation of performer models according to these criteria, among them: 1) small sample sizes for program cohorts, 2) the potential for subjective evaluation criteria in certain skills tests, and 3) incomplete coverage in the acquisition of omics data relative to phenotypic data, for which several years of historical data collection were already available.\n\nTo mitigate these complications, we implemented a two-pronged model evaluation strategy consisting of both a qualitative component, based on pre-registration of the key mechanistic hypotheses each performer planned to investigate, and a quantitative component, based on an evaluation of the predictions of each performer model against a held-out validation set of true outcome labels.\n\nHypothesis pre-registration is a technique used in some disciplines to avoid using the same set of data for both hypothesis generation and hypothesis testing.30 Hypotheses proposed by MBA performers at the outset of the modeling effort included a variety of potential biological mechanisms underlying task performance, such as sleep quality, metabolism, muscle tone recovery, and several proposed cognitive/psychological mechanisms. These pre-registration documents were retained by the IV&V team for later determination if the identified predictive biomarkers might plausibly correspond to the pre-specified categories.\n\nFor quantitative validation, the IV&V team held back 20-30% of the candidate outcome labels from the primary modeling teams during each year of the MBA program. The outcomes for this validation set were kept fully blinded from performer team members to prevent data snooping.31 Modelers were given all other data from each annual cohort and then asked to submit predictions of the outcomes of the blinded candidates for scoring by the IV&V team. Results were announced at each program review meeting.\n\nPreserving the ability to apply predictive models to new cohorts of individuals following the conclusion of MBA was a key goal of the program. Given the small sizes of individual cohorts, prospective model testing on future data collection was considered a significant component of the overall validation strategy. Furthermore, the IV&V team desired to ensure that, to the extent possible, the models would be independent of the choice of laboratory for omics data processing to avoid vendor lock-in.\n\nTo facilitate a single storage location for program data, LLNL data scientists generated a MariaDB database schema to contain all multi-omic, phenotypic, and outcome data collected over the course of the MBA program. As some omics data was too large to practically store within the database itself, the database contained links to the original and processed data files stored in a master data archive. It also contained metadata to track QA/QC results associated with different datasets, as well as to reconcile individual research subjects with their anonymized identifiers.\n\nTo support continued usefulness of the predictive models, the IV&V team requested that performers package their analysis and models for future use as research compendia, according to the method of Marwick et al.32 We chose this format as the R language was the preferred coding environment of the majority of modeling teams. To support long-term portability of the modeling pipelines, containerization of the computing environment using Docker or Singularity was also recommended for each team.\n\n\nResults\n\nThe primary investigator-led teams funded to perform work for MBA offered a high level of cooperation with our IV&V efforts. We were aided by support for our IV&V plan from the research sponsor, particularly when elements of the plan necessitated extra effort by the performer teams, such as in the case of hypothesis pre-registration or periodic data holdbacks.\n\nData QA/QC added a modest amount of time between the return of results from experimenters and the availability of processed data for use by modelers. However, on several occasions involving both sequencing and mass spectrometry experiments, issues flagged during the QA/QC process spurred additional consultation with the data collection teams and led to process improvement that was incorporated into subsequent data re-analysis.\n\nRegular holdout validation set tests of performer predictive models provided an unbiased, apples-to-apples comparison of model performance that assisted the sponsor in measuring progress against program goals. Unfortunately, program constraints made it difficult to test counterfactual predictions made by the modelers, i.e., predictions that certain individuals would have progressed further in the selection process than they actually did. As a result, measuring improvement over state-of-the-art in quantities such as recall, as envisioned at the outset of MBA, was not possible. Instead, the IV&V team defaulted to the use of prediction accuracy and F-score as the primary endpoints for model evaluation.33\n\n\nDiscussion and lessons learned\n\nIn recent years, studies have demonstrated that diverse types of omics data are predictive of biological phenotypes supporting human performance characteristics.34 Nevertheless, this field of research comes with a unique set of challenges that separate it from the much larger pool of clinical research seeking to drive progress in the medical domain. “Success” in the human performance context may be a more multifactorial entity than in the medical context, where it may simply entail the cessation of an identified disease process. Additionally, healthy and, in particular, athletically adept individuals may be more reluctant to participate in invasive specimen collection procedures than individuals already engaged with the medical system.\n\nIn working with cohorts that significantly depart from broader population baselines, reference data from publicly available databases may turn out to be of lesser value than modelers initially hope. For example, studies of the metabolic impact of various dietary regimens in aging or pre-diabetic populations may not have high transfer value in the warfighter population. To the extent possible, omics data collection for single individuals over long periods of time may mitigate this issue and limit the need for transfer learning from weakly representative populations.\n\nAlternatively, research sponsors may wish to consider funding short-term but larger multiomic studies that are composed of participants more closely representative of the target population. Phenotypic outcomes could be collected passively and unobtrusively using wearables technology. Cadre members could be polled to determine surrogate endpoints, measurable in this more high-throughput context, that they believe most likely related to their more holistic judgments in the selection process of interest. Additionally, if the surrogate endpoint markers are continuously valued, this type of outcome variable may allow for superior statistical power than dichotomized pass/fail outcome labels.35\n\nTo participate in blinded prediction contests, modelers may be reluctant to give up scarce training data samples as a validation holdout when the total number of observations in the dataset is small. Statistical techniques that require checking certain prerequisite assumptions, such as the normality of predictor distributions, may become tedious to implement when small amounts of data are released sequentially. The modeler experience might be subjectively improved if there is enough data to constitute multiple test sets, even if some of those are only partially blinded. For example, Kaggle, the competitive data science website, frequently splits datasets into training, “public leaderboard,” and “private leaderboard” components, with the first category being fully accessible to modelers, the second providing a basis for competitors to obtain a preliminary score during the competition, and the final category remaining fully blinded until all models have been submitted.36 Even though the “public leaderboard” data is not truly blinded, since competitors can iteratively query it throughout the model building process, modelers may nevertheless elect to use it judiciously to gauge their performance and to debug basic generalization errors.\n\nFinally, program managers wanting to drive improvements over state-of-the-art outcome prediction, particularly for quantities such as recall, should engage early with program stakeholders to develop means of testing counterfactual predictions made by data scientists. For example, modelers may predict that certain candidates “would have passed” later rounds of a tournament selection process had they been given the opportunity to compete at the higher level. While this information may be of high value from the standpoint of program goals, these predictions are impervious to validation if those individuals are lost to follow-up.\n\n\nConclusions\n\nCommunity-based efforts to promote reproducibility through open sharing of data and code have played an important role in advancing the methodological rigor of many scientific disciplines. We have demonstrated a paradigm for adapting several aspects of this approach to achieve independent verification and validation of results in the context of a research program where unlimited data exchange is not feasible.\n\nUsing holdout prediction tests and hypothesis pre-registration, our team was able to certify that predictive modeling benchmarks were achieved in the absence of data snooping. Additionally, a centralized data infrastructure and integrated QA/QC system promoted data integrity and helped to facilitate the preservation of data and algorithms generated in the course of the project for follow-on research efforts.\n\nWhile our IV&V strategy was developed for projects at the intersection of human performance and defense, we anticipate that similar protocols may prove useful in other research contexts involving multiomic data analysis and sensitive human subjects data. Though the data and trained models from this project are encumbered by distribution restrictions, other artifacts from the study, such as QA/QC rubrics, have been made available to support future work in this area.", "appendix": "Data availability\n\nNo data associated with this article.\n\nFigshare: Measuring Biological Aptitude Omics QA/QC Rubrics. https://doi.org/10.6084/m9.figshare.23802606.v1. 37 This project contains the following extended data:\n\n• ExtendedDataTables.pdf (Sequencing, Proteomics, and Metabolomics QA/QC Scoring Rubrics)\n\nData is available under the terms of the CC-BY 4.0 license.\n\n\nReferences\n\nBegley CG, Ioannidis JPA: Reproducibility in science improving the standard for basic and preclinical research. Circ. Res. 2015; 116(1): 116–126. Publisher Full Text\n\nRobson B: The dragon on the gold: Myths and realities for data mining in biomedicine and biotechnology using digital and molecular libraries. J. Proteome Res. 2004; 3(6): 1113–1119. 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Publisher Full Text\n\nBenjamin J, Morrison Michael D, Thissen James B, et al.: Measuring biological aptitude omics qa/qc rubrics.Publisher Full Text" }	[ { "id": "220464", "date": "22 Nov 2023", "name": "Michael Fitzsimons", "expertise": [ "Reviewer Expertise Genomics", "data sharing solutions", "software" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors share their approach to performing an Independent Verification and Validation (IV&V) process for the Measuring Biological Aptitude (MBA) project, which is funded by the Biological Technologies Office of the Defense Advanced Research Projects Agency (DARPA). They discuss IV&V as a general use case for supporting research reproducibility in a field where data cannot be directly shared. They also provide the specifics of their implementation including QA/QC and qualitative and quantitive evaluation of model performance. I found the article interesting and informative, but I think it is lacking some context that would help readers to understand more about IV&V. In particular, I would like to see considerably more detail about how their approach differs from or improves up other IV&V examples. A cursory review of the literature shows many papers on the topic and it would be helpful to understand how the authors' approach differs and compares.\nI would also love to see any more details about their actual model evaluation techniques. I presume the sponsors constrains what they are allowed to share, but at present it is somewhat difficult to assess the rigor of their verification and validation process.\nAdditional small and targeted comments are included below:\nIntroduction\nData snooping - I don't think this term is understood by all. It might be helpful to add another sentence or phrase about data snooping.\nReachback tasking is not a standard term as far as I know and should be modified or explained.\nIndependent verification and validation (IV&V) - I would like to see a lot more discussion of what this typically looks like and how your implementation compares. A casual review of the literature shows lots of citations about \"independent verification and validation (IV&V)\" - how does your paper add to this literature?. I cannot tell if your implementation is good, bad, or typical. Given we cannot see any actual modeling results (I assume this is a restriction by the sponsor), I think it is important to include a broader comparison to the existing IV&V literature.\nMethods\nStandards discussion is good.\nCould you really ascertain the answer to some of the QA/QC questions such as \"Were collection tubes labeled with unique, traceable codes?\" and \"Were samples thoroughly mixed before loading?\"\nCan you include any more details about the actual models or their evaluation? This is for the quantitative evaluation. Or maybe at least say explicitly what the limitations of what you are allowed to provide.\nResults\nAre there any results, or at least the format for results, that you can share?\n\nOther suggestions\nMention privacy preserving federated learning as another option for incorporating data that cannot be shared directly.\n\nIs the rationale for developing the new method (or application) clearly explained? Yes\n\nIs the description of the method technically sound? Yes\n\nAre sufficient details provided to allow replication of the method development and its use by others? Yes\n\nIf any results are presented, are all the source data underlying the results available to ensure full reproducibility? No source data required\n\nAre the conclusions about the method and its performance adequately supported by the findings presented in the article? Partly", "responses": [] }, { "id": "233971", "date": "19 Jan 2024", "name": "Sheeba Samuel", "expertise": [ "Reviewer Expertise My research focuses on reproducibility", "FAIR principles", "knowledge engineering", "provenance", "and research data management." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIn the context of human performance research, the authors present their study focusing on the Measuring Biological Aptitude (MBA) research program, organized by the Defense Advanced Research Project Agency's (DARPA) Biological Technologies Office. To address reproducibility, the authors implemented a four-pronged independent verification and validation (IV&V) strategy, including centralized data storage, quality assurance and control of data collection, evaluation of performer models, and an archival software and data repository. The IV&V plan, carried out with support from the funding agency and research teams, involved processes like quality control, error flagging, and holdout validation set tests. The paper addresses a topic which underscores the significance of reproducibility, particularly in situations where a fully open approach is unfeasible. In such scenarios, the paper advocates for the establishment of independent teams to cross-check and validate results as a pivotal step in promoting research reproducibility.\nTo offer the necessary motivation in the introduction, the paper could explore the challenges associated with ensuring reproducibility in sensitive fields where privacy concerns limit the open sharing of raw data. It would be beneficial to highlight the obstacles to reproducibility, particularly when faced with limitations related to personal privacy, medical confidentiality, and national defense. The authors could also discuss how other state-of-the-art approaches address these challenges and explore the potential applicability of this approach in similar programs within and outside DARPA research initiatives. Additionally, the authors could provide explanations for introduced terms which are not known to every readers such as data snooping, Reachback tasking, performer expression circuit models, and Holdout validation within the manuscript. Before going into the IV&V methods, a brief introduction and background on the complete test program, and data acquisition w.r.t the large scale omics data would enhance the reader's understanding. The discussion section introduces various new terms and statements that were not previously mentioned, creating a disconnect from the previous sections. It is imperative to establish a clear connection between the concept of reproducibility mentioned in the introduction section and the insights gained in the discussion section. This connection should explicitly define reproducibility in the given context and elaborate on how it is achieved, especially considering the constraint of not being able to share the data online. The paper could also discuss the limitations and the problems occurred in their context, along with proposing potential enhancements that the funding agency could implement to foster reproducibility in similar situations.\n\nIs the rationale for developing the new method (or application) clearly explained? Yes\n\nIs the description of the method technically sound? Partly\n\nAre sufficient details provided to allow replication of the method development and its use by others? Partly\n\nIf any results are presented, are all the source data underlying the results available to ensure full reproducibility? No source data required\n\nAre the conclusions about the method and its performance adequately supported by the findings presented in the article? Partly", "responses": [] }, { "id": "233980", "date": "06 Mar 2024", "name": "Timothy M. Errington", "expertise": [ "Reviewer Expertise open science", "reproducibility", "metascience", "molecular and cell biology", "preregistration" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe article describes the approach the authors implemented as an independent verification and validation (IV&V) effort within in the DARPA funded Measuring Biological Aptitude (MBA) program. This includes implementing four areas to improve and test the reproducibility of the program performers, particularly within the context of a program where not all data can be openly shared.\nOverall, this is a very nice example of how open science practices can be implemented within a program like this. My suggestions are directed on clarifications and expansions to help broaden the readership of this article.\nAdditional context about how IV&V is typically implemented in related programs and why these specific four features were developed. The benefits (and costs) of implementing open science and reproducibility steps in a program like this. That is, IV&V is running ‘alongside’ the performers. However, outside large programs it is more typical researchers (i.e., performers) disseminate their research and then other researchers (i.e., IV&V) look at the paper and outputs (e.g., data, code). In addition to discussing the steps that were implemented (and expanding on the rational), how did the specific improve the program? That is, in the authors opinions, did the effort to implement these steps increase the overall model evaluations? At the end of the results the pivot the IV&V team took to evaluate performance is indicated, but the impact of these measures is absent – this doesn’t have to be disclosing the specific F-scores, more how the four areas that the IV&V team implemented lead to improvements (however that is defined) of the performers and program at large. I think if possible, reworking Figure 1 to highlight not only the two tracks and primary responsibilities of the performers and IV&V, but how this maps onto a more general workflow of research (particularly at the intersection of human performance and defense) would be helpful. I think the abstraction beyond the specific program would help strengthen the conclusion the authors are making about this approach being useful for other projects.\nMinor comments:\nWas pre-registration implemented by performers and used by the IV&V team (which is how I read it from the text), or was it implemented by the IV&V team (which is what Figure 1 indicates. Related, are any pre-registrations openly available or is this also ‘encumbered by distribution restrictions’? Within the Independent verification and validation section of the Introduction there are extra periods in one of the sentences (“confirm…that those circuits”)\n\nIs the rationale for developing the new method (or application) clearly explained? Yes\n\nIs the description of the method technically sound? Yes\n\nAre sufficient details provided to allow replication of the method development and its use by others? Yes\n\nIf any results are presented, are all the source data underlying the results available to ensure full reproducibility? No source data required\n\nAre the conclusions about the method and its performance adequately supported by the findings presented in the article? Partly", "responses": [] }, { "id": "233981", "date": "19 Mar 2024", "name": "Rahuman S. Malik‐Sheriff", "expertise": [ "Reviewer Expertise I am a computational biologist with interest in standards and reproducibility of computational models." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors present a methodology they used to ensure the reproducibility and validation of model predictions in the MBA program, where it was not possible to publicly share the raw data due to privacy concerns or the sensitivity of the information. The authors propose and perform independent verification and validation (IV&V) of markers identified using multi-omics data analysis to predict human performance; using holdout validation sets that are not shared with the project contractors to allow unbiased evaluation of model performance. This approach is quite reasonable, the authors present the limitation in assessing the counterfactual prediction of what would have happened in scenarios where the data was not acquired.\n\nOverall, it is a very good piece of work, which fairly discusses the merits and limitations of these approaches. Holdout validation seems like an excellent strategy for the validation of models developed by diverse groups using the same set of data. Adding some quantitative table or figure showing, how many of the hypotheses/findings were independently verified and validated and how many were reproducible/not reproducible/non-verifiable, could strengthen the manuscript. If possible, some high-level categorization of the models/predictions will provide a bit more context.\n\nThe IV&V strategy includes implementation of 4 steps: 1) a centralized data storage and exchange platform, 2) quality assurance and quality control (QA/QC) of data collection, 3) test and evaluation of performer models, and 4) an archival software and data repository. These four steps are elaborately discussed in the manuscript, however, there are a few questions raised below. If the authors find it reasonable, it would be useful to discuss / clarify them in the manuscript.\nIt would be worthwhile discussing how the platform developed in step 1 differs from federated learning platforms and existing strategies, including their merits and demerits, if any.\nBaker 2016 reported that over 70% of the researchers failed to reproduce other scientists’ experiments and 50% failed to reproduce their own. In step 2, concerning QA/QC, is it sufficient to review the experimental protocols and setups? is it not required to independently reproduce a part of these experiments? Can the authors discuss this?\n\nQA/QC scoring rubric could act as a good guideline for data generation and analysis. However, some of the questions in the QA/QC scoring rubric are a little ambiguous, for example, in question “Was the sample properly recorded?” what does ‘properly’ mean?\nAre questions such as “Were samples thoroughly mixed before loading?” useful? has anyone ever answered no to these questions?\nIn step 3, the authors suggest registration of the hypothesis, however, in most cases where multi-omics data is used, it is often data-driven hypothesis generation, can the authors discuss this? How often are the hypotheses registered are successfully verified by both parties?\n\nTiwari et al. 2020 showed that about half of the computational model cannot be reproduced using the information provided in the manuscript. In step 3 for validation, did LLNL use the same codes originally developed / used by the researchers? were they verified and validated to ensure that any issues in the code were appropriately addressed? Any changes to the steps in processing data, including data normalization will impact the prediction, so where any standards and best practice guidelines are recommended to researchers to harmonize the analysis.\nIn Step 4, though the data cannot be publicly shared, can some of the software / codes / models? be shared publicly in accordance with FAIR data-sharing principles. It would be critical to ensure these are reusable if the plan is to apply these models on new cohorts.\nAs a side note, with some rewriting, the abstract and overall article could be well enhanced to appeal to a broader audience. Here are some of the points. These are optional recommendations; the authors can consider these and similar others in the article and address them as they feel appropriate.\nIn abstract, if “one human performance” is the name of the research program, using punctuation / capitalization appropriately will be helpful, otherwise please rephrase the text accordingly. In abstract: methods could be improved to increase the clarity. The authors could clarify that they present the method they developed and performed a third party independent verification and validation of outcomes of the DRDO-funded projects. In abstract: Holdout validation set, could be clarified. Clarifying in the abstract that human performance prediction models based on multi-omics data will give a bit more context. If space allows, it would be useful to mention that performance of war-warriors performances are predicted. In general, providing some definition for terms such as ‘data snooping’, and ‘expression circuits’ would be helpful.\n\nIs the rationale for developing the new method (or application) clearly explained? Partly\n\nIs the description of the method technically sound? Partly\n\nAre sufficient details provided to allow replication of the method development and its use by others? Partly\n\nIf any results are presented, are all the source data underlying the results available to ensure full reproducibility? No source data required\n\nAre the conclusions about the method and its performance adequately supported by the findings presented in the article? Partly", "responses": [] }, { "id": "220462", "date": "16 Sep 2024", "name": "Elizabeth Dhummakupt", "expertise": [ "Reviewer Expertise Multi-omic biomarker discovery - proteins", "lipids", "metabolites" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis article discusses a method to assist in reproducibility of data collected and generated by studies in which broad questions are asked and large pools of participants are not always available. Additionally, the methods proposed here are useful when full and open data sharing is not always possible, as in the case with research involving national security implications.\n\nThe method discussed herein is easily implemented in that adherence to rubrics is required, which can be done without adding software packages or investing in high performance computing.\n\nIs the rationale for developing the new method (or application) clearly explained? Yes\n\nIs the description of the method technically sound? Yes\n\nAre sufficient details provided to allow replication of the method development and its use by others? Yes\n\nIf any results are presented, are all the source data underlying the results available to ensure full reproducibility? No source data required\n\nAre the conclusions about the method and its performance adequately supported by the findings presented in the article? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1430
https://f1000research.com/articles/12-1429/v1	31 Oct 23	{ "type": "Research Article", "title": "Assessment of weed species composition and diversity in tomato (Solanum lycopersicum L.) farms in Ethiopia", "authors": [ "Dereje Geremew", "Nagassa Dechassa", "Shashitu Bedada", "Getachew Bekele", "Niguse Hundessa", "Tesfaye Abdisa", "Desalegn Gella", "Mosisa Duguma", "Nagassa Dechassa", "Shashitu Bedada", "Getachew Bekele", "Niguse Hundessa", "Tesfaye Abdisa", "Desalegn Gella", "Mosisa Duguma" ], "abstract": "Background: Weeds compete for vital nutrients and water with the tomato plants and serve as an alternative host for other pests. Although there is limited information on the weed species composition in major tomato-growing areas in Ethiopia, this study was conducted to identify the economically important weed species in tomato fields in the East Wollega, Shewa, West Shewa, Arsi, and Sidama zones. Methods: The weed flora assessment was conducted in major tomato growing zones of the country, such as East Shewa, Wollega, West Shewa, Arsi, and Sidama zones, in the 2018–2019 cropping season during the off-season. In each tomato field a 1 m x 1 m quadrat was thrown following an inverted X pattern, and weed species were recorded. Weed species composition was calculated using standard formulas. Results: The results revealed that 63 weed species under 23 families were identified at the surveyed locations. Portulaca oleracea was the most abundant and dominant, with 50% of the dominance value. Weed frequency values ranged from 0.06 to 72% in the studied areas. The weed similarity index was also different among the surveyed locations. The results of farmers perceptions on weed management in tomato fields showed that 61% of respondents practiced hoeing followed by hand weeding, and Tuta absoluta was the most significant constraint to tomato production in the surveyed areas.\nConclusions: This assessment showed that there are differences in the composition and distribution of weed species in tomato fields in Ethiopia, which aid in predicting the population dynamics of weed flora. Farmers perceptions of weed management practices in tomato fields were also different. Therefore, different environmentally friendly weed management options such as good crop husbandry, biological control, and integrated management should be designed to increase tomato production and productivity.", "keywords": [ "Abundance", "Dominance", "Portulaca oleracea", "Weed species" ], "content": "Introduction\n\nThe tomato is one of the most extensively grown vegetable crops in Ethiopia, ranking 8th in terms of annual national production (Tasisa et al., 2012). Nevertheless, in Ethiopia, the yield of tomato is 8 tons per hectare, which is lower than other tomato-producing countries, which produce 34 tons per hectare (FAOSTAT, 2012).\n\nThe increase in agricultural pests is one of the major constraints on vegetable production and productivity in Ethiopia, particularly in the mid-Rift Valley. Weeds compete with the host plant for vital nutrients and available space, resulting in decreased yields in both quantity and quality (Reddy and Reddy, 2019; Kebede et al., 2017). Even though most farmers give less attention to the impact of weeds, a study shows that 45% of annual losses of agricultural products are caused by weeds (Terfa, 2018).\n\nWeeds are currently complicating pest issues to a great extent. The increasing prevalence of numerous viral diseases in tomatoes further highlights the significance of weed control because they may serve as alternative hosts and raise production costs (Hanzlik and Gerowitt, 2016).\n\nThe success of plant protection initiatives depends on the capacity to identify pests (Kalaris et al., 2014). Crop weed flora varies throughout habitats and fields based on the weather, irrigation, fertilizer use, soil type, weed management methods, and cropping patterns (Anderson and Beck, 2007; Kebede et al., 2017).\n\nIn a particular agro-ecology, weed flora assessment aids in the prediction of changes in weed populations brought on by climate change, agricultural practices, and pesticide resistance. In Ethiopia, there is limited information and no well-documented weed species composition and distribution that explain the reduction in tomato production and productivity. The proper identification and prioritization of the most economically significant weed species is essential for solving this issue and aids in the development of effective weed control measures. This supports the claim that accurate information on the presence, composition, importance, and ranking of weed species is required in order to develop effective weed management approaches (Migwi et al., 2017). Similar research has demonstrated that it is possible to determine from survey data the local, regional, and national relevance of weed species and the functions they perform (Hanzlik and Gerowitt, 2016).\n\nOn the other side, weed flora identification aids in maintaining the natural enemies of crop pests and maintaining the balance between crop and non-crop vegetation (Terfa, 2018). This study was carried out to evaluate, catalog, and rank the most economically significant weed species in Ethiopia’s main tomato-growing regions.\n\n\nMethods\n\nThis study was approved by the review committee of the Ethiopian Institute of Agricultural Research, Ambo Agricultural Research Center, Ambo, Ethiopia. Ethical approval was obtained from review committee on January 10, 2018, before the implementation of the study. Verbal consent was obtained from participants through interviewing because all respondents were illiterate.\n\nThe weed survey was conducted at major tomato-growing areas of the country (Table 1), such as East Shewa, Wollega, West Shewa, Arsi, and Sidama zones, in the 2018–2019 cropping season during the off-season (Figure 1).\n\nWeed assessment was done at 5–10 km intervals on the main roadside and gravel roads of major tomato growing areas. In each field, a 1 m × 1 m quadrat was used, following an inverted X pattern.\n\nA quadrat was randomly thrown along transects in each field, and the weed types and extent in a quadrat were recorded according to the methodology (Thomas, 1985; Kevine McCully et al., 1991). The first quadrat sample was taken following where the surveyor walks 50 paces along the edge of the field, turns right and walks 50 paces into the field, throws the quadrat, and starts taking the sample. A new specimen that was difficult to distinguish was brought to the Ambo Agricultural Research Center (Ambo ARC) and identified by experienced researchers using identification manuals. The Global Positioning System (GPS) coordinates of the sampling points were also collected. About 39 tomato fields and 195 weed samples were taken in the studied areas.\n\nThe nomenclature of weed flora was done by using the flora of Ethiopia and Eritrea as a weed identification guide (Stroud and Parker, 1989). Information on cropping systems, crop varieties used, crop management practices, adopted weed control methods, and herbicide use history was collected simultaneously during weed flora assessment from farmers and other tomato growers through interviews (Dereje, 2023b). At each surveyed site, 2–3 voluntary respondents were randomly selected and interviewed.\n\nField data on weed species composition and similarity index (SI) were collected at every sampling point. At each site of data collection, weed species were recorded on a data sheet and later weed species composition (frequency, abundance and dominance) and similarity index was calculated using standard formulas.\n\nData on weed species was summarized using the formula described by Tessema et al. (1999) and Thomas (1985).\n\nFrequency: is the percentage of sampling plots in which a particular weed species is found in a field. It explains how often a weed species occurs in the survey area (Marshall, 1988).\n\nWhere, F = Frequency of particular weed species, X = number of samples in which a particular weed species occurs, N = total number of samples.\n\nAbundance: is the population density of a weed species expressed as the number of individuals of weed plants per unit area (Tessema et al., 1999).\n\nWhere, A = Abundance, ∑W = Sum of individuals of a particular weed species across all samples, N = total number of samples in a field.\n\nDominance: abundance of an individual weed species in relation to total weed abundance in a field (Thomas, 1985; Tessema et al., 1999).\n\nWhere, D = Dominance of a particular species, A = Abundance of the same species, ∑W = Total abundance of all weed species.\n\nSimilarity index (community index): is the similarity of weed communities between different locations or crops. If the index of similarity is >60%, it is assumed that the two locations are similar in species composition and hence the same control method can be applied (Tessema et al., 1999).\n\nWhere, SI = Similarity index; Epg = number of species found in both locations; Epa = number of species found in location I; Epb = number of species found in locations II.\n\nData on the farmers perceptions of weed management practices in tomato fields were calculated through descriptive statistics using Statistical Package for Social Sciences (SPSS) (RRID:SCR_002865) version 26.\n\n\nResults and discussion\n\nA field survey on weed species in major tomato growing areas showed that 63 weed species from 23 families were identified in the surveyed areas (Dereje, 2023a). Based on number of taxa, the Asteraceae family contained 13 weed species, followed by the Poaceae family with 11 weed species (Table 2). Among weed species, 50 of them were categorized as broad-leaved, whereas 13 were classified as grasses and sedges.\n\nPortulaca oleracea and Cyperus rotundus were the most dominant weed species in East Shewa and West Arsi, with 50 and 47.25%, respectively (Tables 3 and 4). The lowest weed species dominance was obtained by Sonchus asper at East Shewa, Bideance pilosa at East and West Shewa (Table 3), Oplismenus hirtellus, Sonchus oleraceus, and Setaria pumila at Sidama Zones.\n\nThe highest weed frequency was also recorded by Portulaca oleracea (95%) and Cyperus rotendus (70%) in the West Arsi Zone (Table 4). A similar finding reported that Cyperus species, Portulaca oleracea, and Amaranth species were the most frequent weed species at Rift Valley areas in the country (Firehun and Tamado, 2006).\n\nThe result revealed that the highest weed community index was obtained between East Shewa and Wollega zones, with 58.62% similarity (Table 5). Whereas the lowest weed species similarity index value (28.57) was registered between West Shewa and Arsi zones.\n\nThis diversity and distributions in weed species composition may be due to variation in climatic conditions, soil types, and farmer practices (Saavedra et al., 1990; Mennan and Isik, 2003). Karar et al. (2005) also stated that crop husbandry, the use of the same herbicide, and the entrance of new invasive weed species into the areas may cause the diversification and distribution of weed flora in a given habitat.\n\nIn this study, 61% of the respondents used hoeing, followed by hand weeding (23%), and hoeing plus hand weeding (15%) for the management of weeds in the tomato fields (Figure 2). About 48% of tomato growers rotated maize after tomato, and 25% of them planted onion after tomato to reduce the weed seed bank in the soil by interrupting germination and suppressing weed growth.\n\nOn the other hand, amongst tomato growers, 48% used an improved tomato variety named “Galilea,” followed by Koshoro (17%), whereas the remaining used local and unknown varieties (Table 6). Furthermore, a significant proportion of growers (69%) used recommended fertilizer rates, whereas the others used above and below recommendation rates.\n\nAbout 46% of respondents stated that Tuta absoluta was one of the biggest obstacles to tomato production in the surveyed locations, along with weeds. This could be as a result of the insect’s resistance to numerous insecticide classes and a lack of knowledge about integrated pest management.\n\n\nConclusions\n\nA total of 63 weed species under 23 families were identified, and the composition, distribution, and diversity of the weed flora were determined. Portulaca oleracea and Cyperus rotundus were the most abundant and dominant weed species. About 61% of respondents in the surveyed areas adopted hoeing for the management of weeds in tomato fields. The weed community index was also different among the studied areas. Therefore, various eco-friendly weed management strategies, such as good crop husbandry, biological control, and integrated management, should be designed to increase tomato yield in Ethiopia.", "appendix": "Data availability\n\nDANS-EASY: DATA OF WEED ASSESSMENT IN TOMATO FIELDS, https://doi.org/10.17026/dans-z9v-c53d (Dereje, 2023a).\n\nThis project contains the following underlying data:\n\n• Arsi.xlsx (weed species composition in tomato farm in West Arsi Zone)\n\n• East Wollega.ods (weed species composition in tomato farm in East Wollega Zone)\n\n• East shewa.ods (weed species composition in tomato farm East Shewa Zone)\n\n• West shewa.xlsx (weed species composition in tomato farm in West Shewa)\n\n• Sidama.xlsx (weed species composition in tomato farm in Sidama Zone)\n\nFARMERS PERCEPTIONS ON THE WEED MANAGEMENT IN TOMATO FIELDS, https://doi.org/10.17026/dans-xwe-ufj6 (Dereje G., 2023b).\n\nThis project contains the following underlying data:\n\n• farmers perception.xlsx (farmers perception and practices on weed management in tomato fields)\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nAcknowledgment\n\nWe acknowledged the Ethiopian Institute of Agricultural Research (EIAR) for the funding. The authors are thankful to the Ambo Agricultural Research Center for providing vehicles for the survey.\n\n\nReferences\n\nAnderson RL, Beck DL: Characterizing weed communities among various rotations in central South Dakota. Weed Technol. 2007; 21(1): 76–79. Publisher Full Text\n\nDereje G: data of weed assessment in tomato fields. [Dataset]. DANS. 2023a. Publisher Full Text\n\nDereje G: farmers perceptions on the weed management in tomato fields. [Dataset]. DANS. 2023b. 10.\n\nFAOSTAT data: Agricultural data. Provisional 2012 Production Indices Data. Crop Primary.2012. Reference Source\n\nFirehun Y, Tamado T: Weed flora in the Rift Valley sugarcane plantations of Ethiopia as influenced by soil types and agronomic practices. Weed Biol. Manag. 2006; 6(3): 139–150. Publisher Full Text\n\nHanzlik K, Gerowitt B: Methods to conduct and analyse weed surveys in arable farming: a review. Agron. Sustain. Dev. 2016; 36(1): 1–18. Publisher Full Text\n\nKalaris T, Fieselmann D, Magarey R, et al.: The role of surveillance methods and technologies in plant biosecurity. The handbook of plant biosecurity. Dordrecht: Springer; 2014; pp. 309–337.\n\nKarar RO, Mohamed BF, Marrs RH: Factors influencing the weed flora in the Gezira Scheme, Sudan. Weed Res. 2005; 45(2): 121–129. Publisher Full Text\n\nKebede M, Gerama G, Mamo K: Qualitative and quantitative assessment of weed in the major wheat growing areas of Western Oromia Region, Ethiopia. J. Nat. Sci. Res. 2017; 7(19): 15–21.\n\nMarshall EJP: Field-scale estimates of grass weed populations in arable land. Weed Res. 1988; 28(3): 191–198. Publisher Full Text\n\nMcCully KV, Sampson MG, Watson AK: Weed survey of Nova Scotia lowbush blueberry (Vaccinium angustifolium) fields. Weed Sci. 1991; 39(2): 180–185. Publisher Full Text\n\nMennan H, Isik D: Invasive weed species in onion production systems during the last 25 years in Amasya, Turkey. Pak. J. Bot. 2003; 35(2): 155–160.\n\nMigwi GG, Ariga ES, Michieka RW: A Survey on Weed Diversity in Coffee Estates with Prolonged Use of Glyphosate in Kiambu County, Kenya. Int. J. Innov. Sci. Res. Technol. 2017; 4(2): 82–94.\n\nReddy TY, Reddy GH: Principles of agronomy. Kalyani Publishers; 2019.\n\nSaavedra L, Torres G, Hernan G, et al.: Influence of environmental factors on weed flora of field crops. Quiver Valley; 1990.\n\nStroud A, Parker C: A weed identification guide for Ethiopia. A weed identification guide for Ethiopia. 1989.\n\nTasisa J, Belew D, Bantte K: Genetic associations analysis among some traits of tomato (Lycopersicon esculentum Mill.) genotypes in West Showa, Ethiopia. Int. J. Plant Breed. Genet. 2012; 6(3): 129–139. Publisher Full Text\n\nTerfa AE: Weed species diversity, distribution and infestation trend in small scale irrigated vegetable production area of mid-rift-valley of Ethiopia. Biodiversity Int. J. 2018; 2(1): 75–81. Publisher Full Text\n\nTessema T, Lema Y, Admasu B: Qualitative and quantitative determination of weeds in tef in West Shewa Zone. Arem (Ethiopia). 1999.\n\nThomas AG: Weed survey system used in Saskatchewan for cereal and oilseed crops. Weed Sci. 1985; 33(1): 34–43. Publisher Full Text" }	[ { "id": "233143", "date": "15 Apr 2024", "name": "Abdellatif Boutagayout", "expertise": [ "Reviewer Expertise Agroecological Weed Management" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nGeneral Comments: The study titled \"Assessment of weed species composition and diversity in tomato (Solanum lycopersicum L.) farms in Ethiopia\" presents crucial insights into prevalent weed species in major Ethiopian tomato-growing regions. While the methodology involving quadrats and field surveys appears suitable, enhancing the study's credibility would benefit from a more comprehensive exposition of sampling procedures and statistical analyses. The study's wide-ranging coverage across multiple tomato-growing zones and identification of 63 weed species across diverse regions constitute a noteworthy contribution. However, a more profound investigation into the ecological impact and potential management strategies targeting dominant species like Portulaca oleracea and Cyperus rotundus would augment the study's depth. Furthermore, there are significant areas in need of additional elucidation and clarification.\nSpecific Comments: Weed Species Diversity: Although the identification of dominant weed species is insightful for forecasting population dynamics, the article could amplify its depth by discussing the ecological impact or potential management strategies specifically aimed at these dominant species. Weed Management Practices: Commendably, the study incorporates farmers' perceptions and practices regarding weed management. However, reinforcing the analysis of these practices' effectiveness in controlling identified weed species would substantiate the study's findings. Weed Similarity Index: While the discussion on weed community index variation among different regions is informative, a more comprehensive exploration of contributing factors such as climate, soil types, or agricultural practices would enrich the analysis. Figure 1 Revision: Improving the visualization of the studied areas is recommended. Presenting an isolated map of Ethiopia within a frame, accompanied by an inset box highlighting the different study areas in distinct colors, would provide a clearer representation. Quadrats in Field Studies: The absence of information regarding the number of quadrats utilized within each field studied diminishes the methodology section's clarity. Including this detail would strengthen the methodology by offering insight into sampling intensity. Materials and Methods Rewrite: Enhancing the methodology section with subheadings and recent citations would enhance clarity and comprehensibility regarding the study's procedures. Missing Information on Farmer Surveys: Despite referencing surveys conducted near farmers, essential details about survey methodology, sampling techniques, respondent numbers, and survey questions are missing. The inclusion of this information is crucial for understanding farmers' perspectives on weed management practices. Herbicide Use Data: While Table 6 presents fertilizer rates, the absence of herbicide use information is notable. Considering the study's focus on weed species, including data on herbicide frequency and types used would align with the primary objectives. Inadequate Results Presentation: The article lacks the presentation of crucial data obtained during interviews with farmers, including information on cropping systems, crop varieties, management practices, and herbicide history. Advanced Statistical Tests: While the article outlines frequency, abundance, dominance, and similarity index calculations, further detailed explanations or statistical validations of these indices would bolster methodological rigor. Integrating more advanced statistical analyses like ANOVA, multivariate tests, or Multiple Correspondence Analysis (MCA) would provide comprehensive insights into weed species variation across different zones. Discussion and Conclusion: The discussion section requires enhancement, particularly in relating the obtained results and the study's impact. Moreover, the conclusion could benefit from specific, actionable recommendations for improving weed management strategies based on identified weed species and prevalent practices.\nIn summary, the study significantly contributes to understanding weed species in Ethiopian tomato fields. Strengthening the methodology and discussion by providing a deeper analysis of dominant species, assessing the effectiveness of current weed management practices, and offering specific recommendations would elevate the article's impact in addressing challenges related to weed management in tomato production.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1429
https://f1000research.com/articles/12-759/v1	28 Jun 23	{ "type": "Review", "title": "A literature survey on healthcare supply chain management", "authors": [ "Amit Mittal", "Archana Mantri", "Archana Mantri" ], "abstract": "Supply Chain Management (SCM) is a practice that has rapidly spread across industries. SCM may boost output while simultaneously satisfying customers. Despite SCM's recognition as a key factor in enhancing healthcare efficiency, widespread adoption remains in its infancy. Hospitals, a crucial element of the healthcare supply chain (HSC), have failed to fulfill the primary goals of lowering costs and providing high-quality treatment due to their inadequate knowledge of supply chain management (SCM). This research was conducted to fill in the blanks in the current HSC literature. Achieving the healthcare supply chain's goal of reducing costs will be greatly aided by the thorough literature study completed for this report. This review of healthcare supply chain management can quantify the benefits of supply chain initiatives and identify opportunities for improvement. Healthcare institutions can make informed decisions on optimizing their supply chains by understanding customer and supplier needs. This includes making strategic decisions on how to improve inventory management, streamline processes and reduce costs. The focus of this study is on the relationship between supply chain practices, the efficiency of supply chain performance, and the financial outcomes for healthcare organizations. By highlighting certain key research issues that are shared by supply chain management and healthcare management, this article contributes to the literature in both areas.", "keywords": [ "Supply Chain Management (SCM)", "healthcare", "Healthcare Supply Chain (HSC)", "supply chain performance (SCP)", "organizational performance (OP)", "healthcare organization", "hospital supply chain", "safety practices" ], "content": "I. Introduction\n\nMost businesses nowadays employ some form of “Supply Chain Management (SCM)”. The “healthcare industry” has struggled with rising prices for decades. Researchers and practitioners alike have been giving more attention to different supply chain techniques for SCM to adapt to the ever-shifting nature, context, and requirements of the industry.1 Supply chain strategies allow healthcare systems to strategically manage and continually control their targets, making them a crucial management tool and the engine for success. In addition to lowering the overall quantity of resources needed to deliver the requisite level of customer service, efficient SCM may boost customer satisfaction by lowering prices and increasing product availability, and decreasing the time it takes to place an order. The growing importance given to creating and preserving beneficial connections with channel partners—who might include “suppliers, middlemen, service providers, and even customers”—can be credited with SCM's rise to prominence. The supply chain of a hospital is very different from a regular manufacturing supply chain. It's a convoluted system that relies on a constant stream of goods and services to meet the demands of its service providers.2 High operational efficiency in the healthcare industry is impossible without the adoption of “Supply Chain Management (SCM)” procedures due to the industry's vast product diversity, fluctuating product life cycles, growing outsourcing, emerging IT trends, and widespread corporate globalization.3 Implementing “healthcare supply chain management (HSCM)” is more challenging since it is a matter of someone's life, just as healthcare organizations and hospitals must carry out incredibly precise activities. However, it was noticed in the literature that the supply perspective is not thoroughly investigated and explored when it comes to the implementation challenges of HSCM procedures. Healthcare service redesign and healthcare program implementation are common practices in this setting with the goals of maximizing the efficiency of available resources and improving the quality of treatment provided.4 In the past decade, numerous impressive literature reviews have emerged in fields related to healthcare to better utilize resources and improve healthcare service. These disciplines include, among others, “decision-making, cold chain, internal medication distribution, operations, management roles, operation research, and outsourcing, agility, and facility maintenance management”.4,5 However, each study was conducted independently. Healthcare supply chain (SC), pharmaceutical supply chain (SC), hospital supply chain (SC), drug supply chain (SC), lean supply chain management (SC) in healthcare, and agile supply chain management (SC) in healthcare are all terms that have been used by researchers in their studies of SC in the healthcare sector.6\n\nThis research follows the following structure: Section II contains the healthcare supply chain. The challenges facing healthcare supply chains are described in Section III. Section IV contains the vital role of supply chain management in the healthcare industry.\n\n\nII. Healthcare supply chain\n\nImprove therapeutic results and keep costs down by using the HCSC, which was defined as data, tools, and cash flow across the supply chain from manufacturer to customer The Healthcare Supply Chain Consortium (HCSC) is an innovative approach to improving therapeutic results and controlling costs across the healthcare supply chain from manufacturer to customer. By leveraging data, tools, and cash flow, the HCSC provides the necessary resources to ensure that healthcare products are delivered effectively and efficiently.5 The focus of HCSC is hospitals. Whether an organization is for-profit or nonprofit, it can be categorized as a service provider. The healthcare value chain comprises five primary players. They function as providers, buyers, sellers, product intermediates, and financial mediators.7 Numerous kinds of research evaluated the effectiveness of the HCSC in terms of costs, and supply chain management is suggested in the literature as a strategy for lowering costs.8 Healthcare provides performance measures for operations such as ordering and inventory management, receiving, storage, and replenishment because it acknowledges the value of “supply chain performance” assessment in the “healthcare industry”.9 It emphasizes how crucial internal client satisfaction is to the healthcare supply chain. They point out that techniques like internal customer satisfaction surveys can result in changes in healthcare delivery, which eventually enhance the entire performance of the supply chain.10 We used five healthcare-inspired metrics to evaluate the performance of the supply chain. These include i) adaptability, ii) integration, iii) patient response, iv) physician performance, and v) partnership quality, and via these aspects, an integrated model was developed.11 HCSC is viewed as a networked constellation made up of medical professionals, advisors, experts, hospitals, clinics, pharmacies, and health-related plans.12 This constellation's overarching goal is to contribute value via cooperation. Customer value in the healthcare industry is produced by suppliers, clients, hospital staff, strategic partners, and others via mutual collaboration, coordination, and teamwork.13 Each care bundle in the healthcare supply chain is made up of resources that are supplied and used by actors including suppliers, clients, hospital staff members, business partners, and others that collaborate to add value for the client.14 Due to interdependencies across several business partners in the supply chain, there is an increase in the creation of customer value. The healthcare supply chain is highly interconnected, with multiple business partners providing essential services, supplies, and support to ensure the successful delivery of patient care. This interdependency has resulted in an increased focus on customer value and experience. As businesses increasingly rely on each other for the delivery of services, products, and support, there is a greater emphasis on creating a seamless customer experience that maximizes the value of the services and products delivered. To create customer value, healthcare supply chain partners focus on improving customer experience through improved customer service, better product quality, and increased product availability. By optimizing the customer experience, healthcare supply chain partners can ensure that they can provide the best possible care to patients and maximize the value of each transaction. Additionally, by creating a positive customer experience, healthcare supply chain partners can drive customer loyalty and build strong long-term relationships with their customers.7,9,11\n\n\nIII. Challenges facing healthcare supply chains\n\nThere was a lot of progress made in the supply chain industry throughout the 1990s. Given the importance of logistics in modern society, this invention was based on that field. Up until that point, it was thought that managing, storing, and transporting things was the organization's agenda.15 The majority of practitioners, consultants, and academics believe that the ideas of supply chain management and logistics management are similar.16 The notion of Supply Chain Management (SCM) has recently been reframed from the integration of logistics throughout the supply chain to the integration and management of essential business activities throughout the supply chain. The new SCM concept focuses on the use of effective management practices to reduce production costs, maximize customer service levels, and increase market share in a competitive global marketplace. SCM has been reframing since the early 2000s, as organizations began to recognize the need for a more comprehensive approach to the management of their supply chains. Consequently, the SCM concept has become a critical component of an organization's strategic planning process. Furthermore, the focus on SCM has allowed organizations to access new markets, expand their competitive advantage, and reduce costs.14,15\n\nWieland examined hospital-supplier cooperation in the healthcare supply chain to determine common goals and strategies for reaching them.17 In that trial, some businesses pooled resources to pay for full-time professionals to oversee logistics.18 The results of the study suggested that improving patient care may be achieved by collaborating with other organizations. The two companies weighed the merits of sharing information and learning about one another's operations.19 The authors highlight the difficulties and constraints of blockchain technology implementation while outlining the potential advantages in supply chain operations, such as better transparency, improved efficiency, and lower prices. The article further discusses the need for additional studies to fill in knowledge gaps and establish best practices for the supply chain implementation of blockchain technology.20 By reducing the time and effort spent searching for a suitable alternative to an out-of-stock item, these delay management strategies can enhance patient care. Their focus was on pharmacists' roles in both primary and secondary care settings. According to Yu et al., and Queiroz et al., to achieve patient outcomes at the lowest feasible cost, medication management should be a collaborative effort between patients and clinicians that considers all aspects of patient usage.20,21 To aid the new service in identifying areas that may be improved as part of the ongoing process of hospital quality improvement,22 they propose an integrated management of pharmaceuticals within the hospital that seeks to cover the main components and analyze the effect of their administration. Researchers considered the whole course of a patient's hospitalization; from the moment they were admitted to the moment they were discharged.\n\nSeveral factors, including the nature of the injury, the patient's age, and the patient's financial situation, among others, contribute to an inaccuracy in the demand forecast for materials and medications, as stated by Venkatesh et al.23 This is especially true in the case of medical supplies, where different types of supplies are used on different schedules depending on the individual's condition. For this reason, people frequently rely only on their expertise while shopping for medical products. Due to the ambiguity in how demand is defined, Rowan et al., found that there are occasionally surpluses and other times there are shortages of medical supplies.24 To address this issue, they first suggested using the Activity Based Costing (ABC) curve, a technique that divides items into three groups and emphasizes 80% of the company's most crucial products. A hospital's inability to accurately estimate demand is further complicated by the absence of quantifiable metrics that would enable smart buying, according to Govindan et al.25 According to the authors, a common cause of medication supply inconsistencies is the lack of reliable logistic data to support the hospital's scheduled procurement plan.26 They recommended obtaining accurate and quantifiable information from three sources: (1) a study of “internal and external processing” times of prescription drug instructions that ensure safe stock levels; (2) a list of standardized medications with “descriptions, units of supply, and spreadsheets to track monthly” samples to utilize each piece of equipment in the hospital and (3) an ABC classification of the meds.26,27 The hospital can enhance its inventory control and management procedures by obtaining precise and quantifiable information from the sources mentioned, such as the internal and external processing times of prescription drug instructions, a list of standardized medications with descriptions and units of supply, and an ABC classification of medications. This can have several advantages, including lowering waste and stockouts, increasing inventory efficiency, enhancing patient safety, and lowering expenditures related to outdated or unused pharmaceuticals. Ultimately, these advancements may help HSC perform and operate more effectively.25\n\nThe public sector purchases medications through a competitive public auction procedure, which was the subject of research.27 According to the Tönnissen & Teuteberg, overstocking frequently resulted from a lack of integrated systems to manage supply, distribution, and consumption.28 The environment is not suitable for public service, as noted by.28 There are a few things they recommend doing to work better with vendors: (1) gathering out pertinent information about the vendor and compiling a shortlist of credible candidates based on such research “information system that addresses legal capacity, suitability, financial standing, and technical suitability”; and (2) learning about the product itself. Medicines are an example of a product category with a lot of technical details. The logistics process requires the participation of qualified specialists, including pharmacists and trained technological workers. Additional indications of the quality of the inputs should be employed, such as the verified “observations of individuals who use the product in the organizations”. The healthcare supply chains in Sri Lanka are managed by several different organizations, all of which report directly to the Ministry of Health.29 These organizations are responsible for providing the necessary medical supplies to healthcare facilities, as well as ensuring the quality and safety of the services they provide. Registration with the Cosmetics, Devices, and Drugs Regulatory Authority (DRA) is required for all pharmaceutical companies wishing to submit bids to the State Pharmaceutical Corporation (SPC).\n\n\nIV. Supply chain management vital role in the healthcare industry\n\nProviding patients with high-quality, cost-effective care requires a well-oiled supply chain in the healthcare industry.30 According to their roles in the healthcare supply chain, the various participants may be divided into four broad groups: manufacturers, buyers, distributors, and providers (Figure 1).\n\nLogistics is concerned with a wide range of tasks, including but not limited to: managing demand and supply, controlling manufacturing, operating, stocking inventories, distributing goods, and transporting people and goods.31 The first function of logistics is resource management, which includes things like wheelchairs, stretchers, and ambulance availability, as well as the storage and distribution of medical supplies and pharmaceuticals (patient, wheelchair, stretcher, ambulance). Logistics duties, such as managing resources, are essential to healthcare stakeholders, such as wheelchairs, stretchers, ambulance accessibility, and medical supplies. Healthcare supply chain stakeholders are essential in managing these resources. For instance, to meet patient needs, hospitals, and clinics must ensure that they have sufficient medical supplies and medications. To ensure they are used successfully and efficiently, they must control the distribution and availability of other resources, including staff, beds, and medical equipment. Moreover, the transportation of patients and medical supplies between facilities such as hospitals, clinics, and warehouses may involve coordination between healthcare stakeholders. Rigorous planning and coordination are required to guarantee that resources are deployed effectively and that they reach their intended destination on time.25–27\n\nThe management of resources and cost optimization in healthcare nowadays revolves around maintaining patient health. The hospital tasks are carried out via a variety of subsystems. Electronic Health Record (EHR) systems–Data about patients, such as medical history, lab results, and medication records, are managed through these systems. Picture Archiving and Communication System (PACS) - this system is used to manage medical images. Hospital Information Systems (HIS) - hospital operations are managed by this system, including admissions, scheduling, and billing. Pharmacy Information Systems - it oversees medication orders, inventory, and dispensing. Clinical Decision Support Systems (CDSS) - the purpose of these systems is to assist healthcare professionals in making decisions based on patient information.32 Most healthcare organizations are moving away from unit procedures and toward supply chains to save money and make better use of available resources.33 This study examined many hospital procedures to illustrate the significance of supply chain management, including medical strategy and service excellence, patient admission and reception, diagnosis and treatment, medical record maintenance, patient release, and rehabilitation services (Figure 2).\n\nThe administrative pillars of a hospital include check-in details (critical patient information), inventory management, billing and collections, patient medical records, staff information systems, and patient information security (Table 1).\n\nThe hospital's ultimate focus is to provide patients with high-quality medical care. What's needed is a robust stock of high-quality pharmaceuticals available in local drugstores. Supply chain management is essential for the hospital pharmacy to ensure the timely availability of drugs at the most cost-effective purchasing price. Different Suppliers,34 “Vendor Agreements,14 Floating of Tenders, rounds of negotiations, and freezing on processes of product delivery”25 are required in the supply chain since some medications can only be carried at specific temperatures. It is challenging to foresee the precise demand for medications. To determine the trend of drug usage, it is crucial to collect correct data. Due to the lack of expertise in supply chain management among today's hospital storekeepers, the supply chain is sometimes plagued by situations of “High Demand and Low Availability”21 or “Reverse, with Low Demand and High Availability”11 for some medicines, thereby increasing the likelihood of medication expiration and extending shelf life.35\n\nThe crucial issue in healthcare is the control of the blood supply. Dynamically managing the blood supply chain is the hospital's aim.36 According to a study,4 donor blood supply is erratic; hence the places chosen for blood collecting sites need to be considered. This study stated that dynamically managing the blood supply chain is a crucial issue in healthcare, and hospitals aim to balance the demand and supply of blood to achieve the goal of providing blood as needed. This study further demonstrated that the number and location of blood banks should be chosen carefully to ensure a balanced supply and demand for blood, and delivery systems should be tightly coupled to satisfy runtime requirements. The authors also suggest that blood banks should be opened around the clock to ensure a timely response to emergencies in hospitals or nearby hospitals. Depending on the transfusion services supply that is needed, the number of local blood banks, that demand and supply should be balanced to achieve the goal, transporting blood as needed, Delivery systems must be coupled tightly to satisfy runtime requirements. For any emergency in hospitals or nearby hospitals, blood banks should be open around-the-clock.4,37\n\nAccording to research, 440,000 patients each year pass away due to medical mistakes that could have been avoided and unsafe work environments.\n\nFigure 3 depicts the drug flow supply chain. The vital life and smooth operation of the company are crucially dependent on the healthcare supply chain. Better healthcare supply chains increase patient safety and improve the standard of service.38 The management of expired medications by automating the monitoring and identification of medications and products, as well as taking appropriate action, has been connected by many hospitals to patient safety and other operations in the correct manner.39\n\nReducing drug-search times, human error, and unnecessary processes can be achieved by streamlining the whole supply chain. To cut down on repetition and human error, the whole data sheet filled out by the doctor needs to be electronically input utilizing RFID technology.40 To increase patient satisfaction and take into account the value of human life, all procedures must adhere to supply chain transparency. A supply chain function and patient safety practices can be found in Figure 4.\n\n\nV. Conclusion\n\nHealthcare supply chain management was the primary focus of this literature survey. The healthcare sector is under a lot of strain right now because of factors such as heightened competition, governmental limitations, rising costs, and consumer expectations for higher-quality services. Due to the industry's expanding worldwide reach, evolving organizational structures, mergers, workforce, and growth of information technology, managing healthcare as a business becomes exceedingly difficult. It is solely used to distribute medicines and vaccines, and our review does not include the delivery of devices that support life. SC implementation in healthcare requires a highly specialized, nation-specific methodology. Healthcare organizations should monitor the performance of their supply chains to make sure they are adding value at each level. Modern RFID technology, SUM, and centralized virtual supply chain management lay the groundwork for the future of supply chain management. Radio Frequency Identification (RFID) technology, Self-organizing Maps (SUM), and centralized virtual supply chain management are three key technologies that are paving the way for future supply chain management. RFID technology provides superior visibility to supply chain data, making it easier to track and monitor inventory. SUM provides an intuitive way to visualize and analyze data, allowing for better decision making. Centralized virtual supply chain management systems enable greater collaboration between suppliers and customers, helping streamline processes and reduce costs.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nReferences\n\nGovindan K, Mina H, Alavi B: A decision support system for demand management in healthcare supply chains considering the epidemic outbreaks: A case study of coronavirus disease 2019 (COVID-19). Transp. Res. E Logist. Transp. Rev. 2020; 138: 101967. 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Publisher Full Text\n\nSazvar Z, Zokaee M, Tavakkoli-Moghaddam R, et al.: Designing a sustainable closed-loop pharmaceutical supply chain in a competitive market considering demand uncertainty, manufacturer's brand, and waste management. Ann. Oper. Res. 2022; 315(2): 2057–2088. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSharma R, Shishodia A, Kamble S, et al.: Agriculture supply chain risks and COVID-19: mitigation strategies and implications for the practitioners. Int. J. Log. Res. Appl. 2020; 1–27. Publisher Full Text\n\nLi Y, Chen K, Collignon S, et al.: Ripple effect in the supply chain network: Forward and backward disruption propagation, network health and firm vulnerability. Eur. J. Oper. Res. 2021; 291(3): 1117–1131. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSingh S, Kumar R, Panchal R, et al.: Impact of COVID-19 on logistics systems and disruptions in food supply chain. Int. J. Prod. Res. 2021; 59(7): 1993–2008. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDutta P, Choi TM, Somani S, et al.: Blockchain technology in supply chain operations: Applications, challenges, and research opportunities. Transp. Res. E Logist. Transp. Rev. 2020; 142: 102067. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYu Z, Razzaq A, Rehman A, et al.: Disruption in global supply chain and socio-economic shocks: a lesson from COVID-19 for sustainable production and consumption. Oper. Manag. Res. 2022; 15(1): 233–248. Publisher Full Text\n\nQueiroz MM, Ivanov D, Dolgui A, et al.: Impacts of epidemic outbreaks on supply chains: mapping a research agenda amid the COVID-19 pandemic through a structured literature review. Ann. Oper. Res. 2022; 319(1): 1159–1196. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMajumdar A, Sinha SK: Analyzing the barriers of green textile supply chain management in Southeast Asia using interpretive structural modeling. Sustain. Prod. Consum. 2019; 17: 176–187. Publisher Full Text\n\nVenkatesh VG, Kang K, Wang B, et al.: System architecture for blockchain-based transparency of supply chain social sustainability. Robot. Comput. Integr. Manuf. 2020; 63: 101896.\n\nRowan NJ, Laffey JG: Challenges and solutions for addressing the critical shortage of supply chain for personal and protective equipment (PPE) arising from Coronavirus disease (COVID-19) pandemic–A case study from the Republic of Ireland. Sci. Total Environ. 2020; 725: 138532. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGovindan K, Shaw M, Majumdar A: Social sustainability tensions in multi-tier supply chain: A systematic literature review towards conceptual framework development. J. Clean. Prod. 2021; 279: 123075. Publisher Full Text\n\nMardani A, Kannan D, Hooker RE, et al.: Evaluation of green and sustainable supply chain management using structural equation modeling: A systematic review of the state of the art literature and recommendations for future research. J. Clean. Prod. 2020; 249: 119383. Publisher Full Text\n\nSharma R, Kamble SS, Gunasekaran A, et al.: A systematic literature review on machine learning applications for sustainable agriculture supply chain performance. Comput. Oper. Res. 2020; 119: 104926. Publisher Full Text\n\nTönnissen S, Teuteberg F: Analyzing the impact of blockchain-technology for operations and supply chain management: An explanatory model drawn from multiple case studies. Int. J. Inf. Manag. 2020; 52: 101953. Publisher Full Text\n\nAbu-Elezz I, Hassan A, Nazeemudeen A, et al.: The benefits and threats of blockchain technology in healthcare: A scoping review. Int. J. Med. Inform. 2020; 142: 104246. PubMed Abstract \| Publisher Full Text\n\nBadi S, Murtagh N: Green supply chain management in construction: A systematic literature review and future research agenda. J. Clean. Prod. 2019; 223: 312–322. Publisher Full Text\n\nTseng ML, Islam MS, Karia N, et al.: A literature review on green supply chain management: Trends and future challenges. Resour. Conserv. Recycl. 2019; 141: 145–162. Publisher Full Text\n\nCentobelli P, Cerchione R, Del Vecchio P, et al.: Blockchain technology for bridging trust, traceability, and transparency in circular supply chains. Inf. Manag. 2022; 59(7): 103508. Publisher Full Text\n\nChang Y, Iakovou E, Shi W: Blockchain in global supply chains and cross border trade: a critical synthesis of the state-of-the-art, challenges and opportunities. Int. J. Prod. Res. 2020; 58(7): 2082–2099. Publisher Full Text\n\nWong LW, Leong LY, Hew JJ, et al.: Time to seize the digital evolution: Adoption of blockchain in operations and supply chain management among Malaysian SMEs. Int. J. Inf. Manag. 2020; 52: 101997. Publisher Full Text\n\nNasiri M, Ukko J, Saunila M, et al.: Managing the digital supply chain: The role of smart technologies. Technovation. 2020; 96: 102121.\n\nSchmidt CG, Wagner SM: Blockchain and supply chain relations: A transaction cost theory perspective. J. Purch. Supply Manag. 2019; 25(4): 100552. Publisher Full Text\n\nSharma A, Adhikary A, Borah SB: Covid-19's impact on supply chain decisions: Strategic insights from NASDAQ 100 firms using Twitter data. J. Bus. Res. 2020; 117: 443–449. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYadav G, Luthra S, Jakhar SK, et al.: A framework to overcome sustainable supply chain challenges through solution measures of industry 4.0 and circular economy: An automotive case. J. Clean. Prod. 2020; 254: 120112. Publisher Full Text\n\nKhan SAR, Yu Z, Golpira H, et al.: A state-of-the-art review and meta-analysis on sustainable supply chain management: Future research directions. J. Clean. Prod. 2021; 278: 123357. Publisher Full Text\n\nMosteanu NR, Faccia A, Ansari A, et al.: Sustainability integration in supply chain management through systematic literature review. Calitatea. 2020; 21(176): 117–123." }	[ { "id": "195717", "date": "19 Apr 2024", "name": "Ramakrishna Yanamandra", "expertise": [ "Reviewer Expertise Supply Chain Management", "Healthcare Supply Chain Management", "SCM in SMEs", "Digital Supply Chain", "Smart Supply Chain", "Supply Chain Resilience", "Circular Supply Chain" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe article partially meets the requirements to get accepted for indexing. I suggest the following modifications for improving its value and recommend resubmission after making those modifications.\n\nIn the Introduction section, the purpose of this research should be clearly stated Section-II should be renamed as Background and Literature Review or just Literature Review. 3. Section-II and III should be part of this Literature Review. A separate section titled 'Research Methodology' should be included. This section should explain how the literature review was done, which journals were considered, period of articles considered, how the articles were selected, gaps in the past studies and then the purpose of this study once again highlighting the role of this research in plugging those gaps. The title of Section-III is 'Challenges facing healthcare supply chains'. However, only a description of some aspects of SCM are written in this section WITHOUT any mention of challenges! The author should highlight the challenges based on literature review. Section-IV should begin with 'In view of the above challenges, this section proposes how SCM can resolve some of those challenges etc.' The Conclusion section should clearly be linked to the 'gaps' identified in the literature review, it should address the conclusion for each challenge. The last conclusion should be about 'why this research is a unique one and how it contributes to the existing body of knowledge in the area of HSCM.\n\nMy Best Wishes!\n\nIs the topic of the review discussed comprehensively in the context of the current literature? Yes\n\nAre all factual statements correct and adequately supported by citations? Yes\n\nIs the review written in accessible language? Yes\n\nAre the conclusions drawn appropriate in the context of the current research literature? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-759
https://f1000research.com/articles/12-411/v1	17 Apr 23	{ "type": "Research Article", "title": "A blended model of interior design studio practice due to COVID-19", "authors": [ "Imad Assali" ], "abstract": "Background: The coronavirus disease 2019 (COVID-19) pandemic upended the educational system around the globe. During this challenging period, universities and colleges looked for other effective alternative methods of learning, such as Virtual Learning Environments (VL). Besides, Ahlia University has implemented E-learning in response to COVID-19. There needs to be more attention given to the challenges associated with technology adoption facing interior design and architecture programs, where over 60% of courses are practical, especially design studios, which form the core of the curriculum. According to a review of the relevant literature, there needs to be more research on blended learning in interior design and architecture. In order to enhance the teaching and learning of interior design and architecture, further research is required to combine cutting-edge techniques and technology. The aim of this study was to review the classroom materials for Ahlia University's interior design studio. Methods: After completing the INTD 212, INTD 216, INTD 311, and INTD 404 studios in mid-March 2022, a short Qualtrics poll was done to assess the difficulties of e-learning and offer potential consequences. Results: Though students were conveniently attending courses online, there was not much discussion and interaction like in the face-to-face model. Blended teaching in design studio courses offered many benefits. The results showed that blended design studios achieved pedagogical results as students developed their knowledge.\n\nConclusions: Based on the findings, this research concludes that teaching and learning should be shifted from face-to-face and online learning to the best practice of a blended format.", "keywords": [ "Virtual Design Studio (VDS)", "COVID-19", "E-learning", "Blended learning", "Virtual learning environments (VLSs)", "Moodle", "MS Teams", "Face-to-Face learning" ], "content": "Introduction\n\n\n\n“A flexible learning approach aims to provide learners with excellent choice, convenience, and personalization to suit their needs. With flexible learning, learners can choose where, when, and how they learn through a variety of technologies that support teaching and learning.p.02”.1\n\nThe environment for interior design education is developing and reaching new heights due to cutting-edge technologies, innovative teaching strategies, and raised standards for design education.\n\nHowever, worldwide interior design education needs to rapidly prepare for extreme situations, given the ongoing coronavirus disease 2019 (COVID-19) pandemic. However, to finish the academic year without squandering the current semester, colleges worldwide are switching to online and e-learning methods. Within a week of the first COVID-10 lockdown, online classes at Ahlia University started. However, neither the teachers nor the students anticipated how it would affect every course, academic or practical.2,3\n\nAll educational institutions worldwide have shut down as the world fights COVID-19, which impacts learning. It is essential to offer learners a face to face learning method.4,5 However, this conclusion highlights the significance of Virtual Learning Environments (VLSs) as a replacement for the conventional pedagogical strategies of lectures, tutorials, and exam to guarantee the effectiveness of education. The rise of information communication technology and the internet-savvy youth, which increased learning options during the COVID-19 epidemic, are prerequisites for these new learning techniques.6,7 It emphasizes the necessity for faculty members knowledgeable about and skilled in online pedagogies and emphasizes student-centered practices more.1 Like many other campuses, Ahlia University shut down its physical structure in reaction to COVID-19 to stop the virus’s spread. It set up various Microsoft Teams-based virtual professional development sessions for its faculty members who were taking their first e-learning courses to give them the assistance they needed to transition to virtual learning and get students ready for this change in environment. Students could access resources and submit their assignments after faculty members presented live lectures using the Microsoft Teams (MS Teams) platform, post-learning activities, and related resources on Moodle. Along with the traditional design studio (face-to-face learning), the mainstay of interior and architectural design education has been dealing with real-world problem-solving and developing students’ conceptual grasp of design since the 19th century.8 Today’s design studio education is experiencing unheard-of difficulties due to new technology and COVID-19 constraints. It is necessary to incorporate technology tools, create online instruction, and evaluate students’ levels of understanding through synchronous and asynchronous interactions to design blended learning that uses face-to-face and virtual learning in a hurry. This will help to create a productive learning environment.9–11 As cited by Brown et al.10 from Hulon [Ref. 8: p.6], the studio is “a virtual world representing the real world of practice, but relatively free of its pressures and risks”. Consequently, design studios prepare students for professional practice. Since the middle of the 1990s, the Virtual Design Studio (VDS) has grown significantly, urging attention to the potential of digital technologies and their influence on instructional strategies.12–14 In order to better understand the preferences of the students in Ahlia University’s interior design program for blended, online, and face-to-face learning, this research examines the engagement of blended learning, faculty members’ roles, and students’ experiences in blended learning in interior and architecture design education. Investigate how blended learning can be used effectively in the interior design curriculum.\n\n\nLiterature review\n\nThe design studio is the guiding principle of interior design education and other design disciplines. It serves as a model for educational innovation. It is devoted to teaching students how to construct products that develop their technical and social abilities by combining knowledge from the classroom with real-world experience.7,15,16 The design studios have provided the development of students’ creativity, imagination, professional expertise, and post-graduate practical experience with the utmost relevance and legitimacy.16,17 At all design education levels, a design studio entails one-on-one interaction between students and the studio educator, group discussions, and teamwork in creating design ideas that are ultimately presented to the studio educator and other invited experts’ audiences.18 As a result of the coronavirus epidemic, design studios’ pedagogy has changed from face-to-face classes to online classrooms, posing a new challenge for students and faculty members who must provide the highest education as specified by the Council of Interior Design Accreditation. New technologies, social networking sites, and the emergence of artificial intelligence have revolutionized teaching and learning methods and facilitated distance learning that allows student autonomy through online learning models. The current and ongoing coronavirus situation is also a result of these changes.18 Despite the pandemic affecting all design schools in late 2019, e-learning has always been part of design studios and has proliferated in all design schools, particularly virtual design studios (VDSs). Faculty and students use MOODLE, Microsoft Teams, MOODLE, and other platforms to present online, distribute class materials, and submit final work.\n\nA hybrid or mixed-mode course that combines traditional in-person instruction with online learning is known as blended learning. In addition, blended learning (also known as hybrid or mixed-mode courses) is a competency-based paradigm that combines traditional face-to-face instruction with online learning.\n\nAccording to Schnabel et al.,11 blended learning refers to a hybrid course that substitutes technology-supported activities for a portion of the standard course’s time. According to Troha,19 blended learning is the term for online and in-person activities. They continued by defining course objectives before course activities, such as assignments and assessments, as they determined the delivery method between in-class and online activities. Blended learning, they said, refers to the combination of on-campus class meetings and online activities. Here, the question of how much in-person instruction must be replaced with online coursework arises. McGee and Reis18 estimates that 30% and 79% of blended learning lessons are provided online. It is challenging for studio-based learning to have a large student enrollment since a traditional studio environment has a small student-to-studio educator ratio where students and their tutors interact in a creative and reflective process. Therefore, blended learning can offer a comprehensive enrollment studio course to address this difficulty. It also overcomes the limitation of physical classroom space in some universities and courses that may not lend themselves to online delivery, such as labs.11 According to McGee and Reis,18 modern technology that eliminates physical barriers between students from different cultures encourages collaboration. It allows collaboration on design projects with other design schools worldwide. Besides the physical distance, tutorials, and desk crits, studio instructors’ roles in both VDSs and traditional design are similar. However, despite employing various mediums, students must present design solutions. As cited in the BlendKit Course,5 “Students in online conditions performed modestly better, on average than those learning the same material through traditional face-to-face instruction” and, “Instruction combining online, and face-to-face elements had a larger advantage relative to purely face-to-face instruction than did purely online instruction. The University of Central Florida (UCF) has been using blended learning for over 17 years and has offered 10,941 blended courses since the end of the 2015-2016 academic year. Despite these benefits, the virtual studio procedure is more complex than it seems because it is almost certainly more complicated and time-consuming than face-to-face studios. According to Schön,16 students construct ideas in a virtual design studio that respond to the issue in the design brief and offer solutions, typically by utilizing different software like AutoCAD to make digital drafts, which takes more time than creating hand-drawn sketches. Students must therefore have extensive training in interior and architectural design and comprehensive education in ICT.12 In order to help their students, faculty staff must also be knowledgeable in ICT and online instruction.20 According to Schön,16 social media and digital tools help people develop their talents, making it simple to implement virtual design studios (VDS) into design studio education. Students in interior design and architecture schools employed software tools like AutoCAD, 3DMax, Building Information Modeling, and Adobe Photoshop in their design projects at various design studio levels.12\n\n\nProblem statement\n\nThe idea of virtual and online design studio classrooms for interior design raises several issues, focusing on the communicative and practical components of learning. The COVID-19 global lockdown has significantly impacted the practical, communicative, and team-building scenarios that are part of interior design studio classes. Universities worldwide have a sense of losing credibility in the design studio when they fall short of the ideal quality of global design education. Maher and Simoff14 emphasized that the studio needs to be an appropriate size to ensure a direct connection between students and their studio educators and each other. It is therefore vital to understand and adapt to the extensive extent to which design studios will change in the future.\n\n\nObjectives\n\nThis study aims to understand the crucial changes and measures that virtual design studios will have to adapt to due to the current COVID-19 pandemic in interior design and architecture education. By assessing the results of the analysis presented to the faculty and students, this study also aims to understand their perspectives on online and e-learning education. Thus, the main summary of the research objectives is:\n\n• Integrating physical and online education in interior design courses in order to determine the success rate.\n\n• Developing comprehensive solutions to make online and e-learning education a global norm.\n\n• To meet the education standards set by the Council of the Interior Design Accreditation (CIDA) during this challenging time of virtual and e-learning methods being the only feasible and safe options available.\n\n• Emphasizing and demonstrating the importance of how virtual design studios (VDSs) can enhance internships and post-graduate jobs for students.\n\nFor academic research to be credible and accurate, bias must be avoided. In this study, efforts were made to address potential sources of bias, including discussing the results with the responders.\n\n\nMethods\n\nBased on the abovementioned research objectives, this study’s primary goal is to evaluate the effectiveness and standard of blended learning in design studio procedures as a result of COVID-19. Thus, this pilot study used a blended learning environment that combines a face-to-face studio environment with an online studio environment for design studio courses. The learning environment includes INTD 212, INTD 216, INTD 311, and INTD 404 of interior design students from different levels, faculty members, and interior and architecture professionals, along with appropriate journals and literature assessments over five months. It was conducted over ten weeks in the spring semester of 2020.\n\nParticipants were selected in all design courses (probability sampling).\n\nThe goal was to compile and watch the details of participants’ behaviors in the blended learning setting. The author of this work did the observation by visiting some classes at Ahlia University face-to-face and through MS TEAMS. Students’ confidence level, access to the Internet during online education, convenience in using MOODLE, Microsoft Teams, computer-aided software or developing projects, students’ listening abilities, students’ progress on projects, students’ response to the online environment, and increased opportunities for interaction and collaboration with peers are the factors considered for this study.22,23 Further, the factors observed in this study were the impact of online education allowing students to communicate with other students better than a conventional face-to-face teaching model. It is overall monitoring between the instructor and the students through discussion, feedback, and filling of the survey questions.\n\nParticipants were selected from all design courses: INTD 212, INTD 216, INTD 311, and INTD 404 (probability sampling). Students were positive and satisfied with taking part in the study, both on the implementation of blended learning. Students were also permitted to reopen the lesson without any pressure. A total of Three hundred and fifty-seven undergraduate students (283 females and 74 men) enrolled in interior design and architecture courses at six institutions in Bahrain responded to the survey.\n\nIn addition, in-depth interviews with eighteen (18) teaching staff (11 women and 7 men) about the students’ awareness and cognition of the difference between online and blended design studio courses were carried out. A survey was also conducted which examined students’ software literacy in a design project and their preparedness for online learning. Moodle and LMS make it easy for students to look back on earlier materials and to move through coursework at their own pace. It also examines studio lecturers and students’ satisfaction with Moodle and MS Teams in communication, feedback, and project progress using Likert scales.\n\nAfter preparing and refereeing the tools for data collection, a written request was made to the Deanship of Graduate Studies and Research (DGSR), Scientific and Ethical Committee, Ahila University, who approved the study on 22nd March 2020. Students, the core of the study, were given the purpose of the research and asked to participate in the study. A form containing details of the project, such as the objective and duration of the study, was given to the students from the design courses: INTD 212, INTD 216, INTD 311, and INTD 404. A total of Three hundred and fifty-seven undergraduate students agreed to participate in this study. After the participants filled a written statement of willingness to participate in the study. We held a zoom meeting with students who agreed to participate in the study. We explained the objectives of the survey and informed them that they were requested to provide feedback on blended learning. Similarly, a detailed explanation of the project was provided to the 18 staff members willing to participate in the study, and written consent was obtained from them. The researcher told them that all their information would remain anonymous and would not be used for other purposes.\n\nDesign work was done on MOODLE and Microsoft Teams throughout the design studio courses, using both synchronous and asynchronous interaction platforms, such as lectures, discussions, announcements, presentations, project briefs, research studies, and sharing images, drawings, and evaluation criteria.\n\nThe questionnaire via Google Online Surveys includes closed and open questions for all students in these four design studio classes.23 It uses qualitative and quantitative approaches to examine student opinions on the methodology and tools employed. Closed questions were used to gather information on demographics and e-learning preferences.8 In order to understand more about how students, interact with the various e-learning platforms, open-ended questions on learning and teaching in higher education were posed to the participants.\n\nThe study considers adding some questions that was earlier presented by Hulon et al.8 This is our initial attempt to focus on studying the face-to-face and online learning and blended format. Therefore, prior to implementation of the pilot study of our survey, a panel of experts from our university were provided with questions and they were asked to provide feedback on how well each question measure the construct in question. Since this is our preliminary pilot work, we have developed the questions and the experts reviewed it. In analyzing the data, we ensured that the questions all reliably measure the same latent variable. We employed SPSS’s Version 27.0.1.0 reliability command to test for internal consistency using Cronbach’s alpha. Our interpretation of the output was based on George and Mallery’s (2003) rule.\n\nOf the Three hundred and fifty-seven undergraduate students who agreed to participate in the study, it has continued for over ten weeks of the project duration, and all the students who participated filled out the survey questions. None of the patients withdrew or dropped out of the study. Interview details from 18 staff members and data from their responses were collected for analysis. The student’s t-test was used to analyze the data. Student’s t-test was used to compare the effectiveness of online, face-to-face, and blended teaching. When performing a student’s t-test for online and face-to-face teaching comparisons, the significance level was set at 0.05. This means that the results of the t-test must be statistically significant to a level of 95% confidence. SPSS Version 27.0.1.0 Statistical Software Package for analyzing the data.\n\nThe use of Moodle and Microsoft Teams for supporting learning and teaching that provide chat, virtual meetings, shared files, and the ability to organize live sessions synchronously or asynchronously is advised in response to COVID-19 restrictions forbade face-to-face instruction when the lockdown started in March 2020. The Ahlia University IT staff quickly delivered training and thorough instructions on using the new Microsoft Teams. As a result, the instructors in the interior design program at Ahlia University (AU) used Microsoft Stream and Moodle as communication tools for both theoretical and design studio courses like INTD 212, INTD 216, INTD 311, and INTD 404 in the spring semester (2020). They discovered its effectiveness in the delivery of the courses.\n\n\nResults\n\nOnline teaching has become increasingly popular in higher education. Online teaching is becoming increasingly popular in higher education and can be a challenge for staff that have not previously taught in this format. Our staff reported when conducting interviews that teaching a design studio course online requires demonstrating both technical and creative skills. While there are many positive aspects of online learning, such as increased accessibility, they encountered difficulty in transferring the design studio course from an in-person setting to an online one. Staff reported that they had struggled to create an engaging online environment for students and to ensure that the course was just as enriching as if it were taught in person. Additionally, the lack of face-to-face interaction in an online setting makes it difficult to build relationships with students. They reported that they faced difficulty in understanding how well each student was performing and in providing the necessary feedback. Another concern is that Staff they had some issues when technical difficulties arise during an online lesson which included on learning how to use the various software programs, troubleshooting any connectivity issues. All these elements were challenging for the staff to effectively teach an online design studio course.\n\nBased on probability sampling, the students were selected from the following design courses: INTD 212, INTD 216, INTD 311, and INTD 404. A total of 357 students participated in this study.22 The participants were informed of the study objectives. All the students who showed interest in participating in the study gave their feedback, and none of the students dropped out during the course of this study. As an initial step in validating the survey is to establish face validity. The experts related to our study read through the questionnaire completely and evaluated whether the questions effectively capture the objective. Our interpretation of the output was based on George and Mallery’s (2003) rule. Cronbach’s alpha coefficient increases as the number of variables increased, and the average inter-item correlations increased.\n\nUsing quantitative analysis for 357 students, 79.2% (283) were women, and 74 were men, from different design departments in Bahrain. The questionnaire included information about the students’ degree in digital literacy, showing that 98% of them had access to a computer at home for academic purposes. The fact that 73% of students used social media daily, 71 used it sometimes, 14 typically used it once per week, and 11 said they did not use it at all demonstrated their suitability for design studio E-learning education. E-learning environments were suitable for design education by 72% of the respondents. In contrast, blended learning environments—including instructor and technical support from the university—were deemed the most effective and suitable in 78.5% of the responses. The results demonstrated a significant impact on the quality of online design studios. Besides, more than 73.8% of the respondents affirm that they have become more proficient in computer software, which helped them in their design projects as online learning provides them with the flexibility of time to develop their skills. Overall, 82 of the respondents reported that face-to-face teaching is crucial in design education. On the contrary, first-year students in all the universities in Bahrain were less satisfied with online learning (Figure 1A), especially in courses like graphic design and basic design. Also, findings about MS Teams and Moodle revealed that 94% of the respondents were satisfied with these platforms (Figure 1B). They provide satisfactory results because they are easy to use and effective for online design studios for communication, discussion, and the exchange of design information, enabling studio educators to monitor student work (Figure 1D).\n\nAccording to the survey, students share their design creations online to receive feedback and criticism from studio educators. The findings showed that 96.7% of students choose not to post their design work on Moodle or MS Teams (Figure 1E), keeping it instead for in-person meetings with their studio instructor or uploading it as soon as it is finished out of concern for intellectual theft. Furthermore, online learning saves students’ travel time to the campus in developing their technical skills for design projects. The survey results revealed that 11 participants said they had trouble downloading and uploading files and connecting to the internet. Smaller class sizes in design courses would be preferable, according to 32 of those surveyed, while 64 students noted e-learning and blended learning as more novel and interactive. In this way, the questionnaire results provided insightful information about how students had used Moodle and Microsoft Teams to study effectively in a hybrid environment (Figure 1F). In addition, students believe that in-person instruction is a crucial component of design education.\n\nOnline learning could lead to a more flexible, student-centered, and creative educational process. The role of faculty members will be transformed from the traditional teacher-centric model to a student-centric model with the implementation of e-learning. Implementing online learning has increased globally recently, and some higher education institutes in developing countries are embracing this trend. This study aimed to investigate the perception, challenges, and predictors of online and face-to-face learning acceptance during the COVID-19 pandemic among university students. We found that most of our participants agreed that the online course offered greater flexibility in teaching times. In contrast, many participants reported that online learning could take time and lead to listening difficulties and experienced network and technological problems affecting learning progress.\n\nDuring online education, most participants were satisfied with the effort involved in developing a project using computer-aided software. According to 72% of respondents, e-learning environments are suitable for design education. In contrast, first-year students were less satisfied with online learning, particularly in graphic design and basic design courses. Despite this, 96.7% of students chose not to post their design work on Moodle or MS Teams, instead choosing to present it in person with their studio instructors or upload it as soon as possible to avoid intellectual theft. According to the study, 89.1% of participants strongly agreed that instructors could teach at their own pace through an online course. Although they are geographically separated from the instructor, they feel connected to the course and the instructor.\n\nBy combining face-to-face and online learning, blended learning offers learners the benefit of both learning methods. Studying asynchronously enables participants to study at their own pace, but questions are generally posted on an online forum so instructors can answer them. As a result, interactions are delayed. During face-to-face teaching, instructors and students are in constant communication, allowing for instant discussions and clarifications of questions. Online learning has numerous benefits, but many learners still prefer face-to-face learning due to the discipline and familiarity it offers. Most study participants reported that online education prevented them from communicating effectively with other students. Most participants felt they could communicate with other students in a conventional face-to-face approach.\n\nThe outcomes of this study revealed that online learning is more flexible than traditional learning, and face-to-face learning is generally more practical for learners. The majority of our study participants preferred blended learning (both online and face-to-face) for developing knowledge and skills and better outcomes.\n\nAccording to 86.7% of the students, blended learning gives them flexible time and lets them study when it is convenient. Additionally, it allows them to communicate with their studio educator for guidance and feedback more frequently than in a face-to-face setting, which enables them to advance their design project more quickly. As cited by some students, “it is such a help to know the next studio whether there is something to improve on, or just to ask simple questions that would not be answered until the next studio. It also allows you to express your ideas more in e-learning to make sure you fully get your point across. You can forget things easily in the studio”.\n\nA quick review of the teaching staff revealed that all of them had access to computers at home, used online learning environments like Moodle and Microsoft Teams for educational reasons, and took part in various virtual workshops and conferences utilizing the Zoom platform. The survey also showed that 93% of respondents used social media sites like Facebook, Twitter, and Instagram, which Ahlia University offers, to communicate. During the pandemic epidemic, all interior design and architecture professionals’ work was moved to the virtual world by employing computer-mediated collaboration platforms to discuss design projects with their clients and contractors. Therefore, interior design and architecture programs use the same approach to prepare their graduates for the market needs.\n\n\nDiscussion\n\nIn this study, both the students and staff reported that both online teaching and face-to-face teaching have advantages and limitations when used in a design studio course. Online teaching can provide more flexibility and convenience as students can access resources, such as video tutorials and lectures, from wherever they are, and can complete their work at their own pace. However, the lack of direct contact with instructors and peers can be a major limitation of online teaching in a design studio course, as it can be difficult to replicate the traditional in-person learning experience. On the other hand, face-to-face teaching provides a more immersive learning environment, as students are able to interact and collaborate directly with their instructors and peers. However, it was difficult to implement if the staff were unable to meet in person due to travel restrictions or social distancing measures. Additionally, face-to-face teaching is more time-consuming, as it requires instructors and students to be physically present during the course. Overall, both online and face-to-face teaching have advantages and limitations when used in a design studio course. Ultimately, it is important for instructors to consider their specific situations and available resources when deciding which teaching method is best for their course.\n\nIn this study, blended teaching in design studio courses offered many benefits to the students and staff. By combining traditional face-to-face instruction with online components, blended teaching allows students to benefit from both. For example, students can access online resources, such as lectures and demonstrations, from anywhere, allowing them to review as needed to gain a thorough understanding of the concepts presented. Additionally, blended teaching can provide students with the opportunity to work at their own pace and on their own schedule, increasing the likelihood of successful course completion. The benefits of blended teaching also extend to the staff. Additionally, the use of online components allows staff to have more control over the pacing of the course and to be able to cover more material in the allotted time. Overall, blended teaching offers many benefits to both the students and staff. Students are able to access online resources, work at their own pace, and access up-to-date information and materials.\n\nPreviously, the new technologies were limited to specific sectors, but nowadays, their low cost makes them affordable to promote learning outcomes in education. This research highlights the need to apply changes in online learning by developing the design program’s pedagogy. This incorporation of new technology and digital media into design pedagogy has the potential to support online design studio learning environments.21 Furthermore, blended learning is not a novel learning system for universities that need a different mode of design pedagogy. Although students use the new technology, they still need more support and guidance, especially first-year students, to improve the quality and outcome of their design projects.\n\nThe results of this study showed that blended design studios achieved pedagogical results as students developed their knowledge and skills, especially in computer software. Students in various design studio courses now more widely appreciate blended learning in design studios due to in-depth study, literature reviews, and data discoveries. The benefits of combining in-person and online interactions create a much more inclusive learning environment than one-on-one instruction, as noted by Sopher.6 The data collection shows that Ahlia University’s design studio’s use of blended learning has a high success rate, and most participants believe it is flexible. The comprehensive criteria of accessibility, preference, practicability, and student needs are all things blended learning attempts to address. The study concentrated on the many levels of design studio education’s legitimacy, influencing the next generation of architects and designers through technical and online education.\n\nThe alternative to blended learning is changing how architecture and interior design are taught, and this will likely be the following significant change in higher education that must be prepared. For students and professors, the ideal balancing act of physical and virtual learning opens up new ideas and opportunities to investigate the philosophy of distant learning and preserve practicality by attending physical design studio workshops. As previously mentioned, Ahlia University performed virtual learning using a variety of communication channels, allowing faculty members and students to experiment with various forms of engagement, including emails, text messages, audio calls, and video calls. The global COVID-19 pandemic necessitates the search for potential replacements where the educational framework of design studio classes is not jeopardized by a lack of communication, method learning, and practical experience. As a result, blended learning is crucial at this point in the pandemic to be acknowledged as a viable alternative to our everyday physical learning, which is impractical given the state of the world’s health. When it comes to meeting the requirements of traditional design studio sessions, blended learning is the ideal fit for the new reality of design education. Additionally, the results demonstrated that throughout the COVID-19 epidemic, all partners contributed to ensuring the caliber of online design studio instruction.", "appendix": "Data availability\n\nA BLENDED MODEL OF INTERIOR DESIGN STUDIO PRACTICE DUE TO COVID-19 – Underlying Data. https://doi.org/10.6084/m9.figshare.21543417.v2. 22\n\nThis project contains the following underlying data:\n\n- Underlying data.xlsx (responses to the survey)\n\n- Interview – Underlying data.xlsx (interview responses)\n\nFigshare: Blended Learning - Questionaire.pdf. https://doi.org/10.6084/m9.figshare.21940130.v1. 23\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nHuang RH, Liu DJ, Tlili A, et al.: Handbook on Facilitating Flexible Learning During Educational Disruption: The Chinese Experience in Maintaining Undisrupted Learning in COVID-19 Outbreak. Beijing: Smart Learning Institute of Beijing Normal University, UNESCO International Research and Training Centre for Rural Education Version 1.2; March 2020. Reference Source\n\nReza Saghafi M, Franz J, Crowther P: Perceptions of Physical Versus Virtual Design Studio Education. [ebook] Archnet-IJAR, MIT- Massachusetts Institute of Technology; 2012; 6(1): pp. 6–22. Publisher Full Text Reference Source\n\nBender M, Vredevoogd J: Using Online Education Technologies to Support Studio Instruction. International Forum of Educational Technology & Society. 2006; 9(4): 114–122. 9th ed. Reference Source\n\nBozkurt A, Jung I, Xiao J, et al.: A global outlook to the interruption of education due to COVID-19 Pandemic: Navigating in a time of uncertainty and crisis. Asian J. Distance Educ. 2020; 15(1): 1–126. Publisher Full Text\n\nBlendKit Course: BlendKit Reader: Chapter 1, “Understanding Blended learning”.Futch L, Chen B, editors. 3rd ed.Publisher Full Text Reference Source\n\nSopher H: Use of Immersive Virtual Environment in the Design Studio: An Assessment Model. VR, AR & VISUALISATION\|Explorations - Volume 2 - eCAADe 36. 2018. Reference Source\n\nSchön DA: The reflective practitioner: How professionals think in action. 1st ed.London: Routledge; 1992. Publisher Full Text\n\nHulon S, Tucker MH, Green AM: Virtual Professional Learning for In-service Teachers to Support Teaching and Learning in Online Environments. Teaching, Technology, and Teacher Education During the COVID-19 Pandemic: Stories from the Field. Published by AACE-Association for the Advancement of Computing in Education. NACOL Research Committee, Online Conference. October 26-28, 2020. Reference Source\n\nPektaş ST, Gürel MÖ: Blended learning in design education: An analysis of students’ experiences within the disciplinary differences framework. Australas. J. Educ. Technol. 2014; 30(1): 31–44. Publisher Full Text Reference Source\n\nBrown SE, Karle ST, Kelly B: An Evaluation of Applying Blended Practices to Employ Studio-Based Learning in a Large-Enrollment Design Thinking Course. Contemp. Educ. Technol. 2015; 6(4): 260–280. Publisher Full Text Reference Source\n\nSchnabel MA, Kvan T, Kruijff E, et al.: The First Virtual Environment Design Studio. eCAADe proceedings. 2001. Publisher Full Text\n\nShao YJ, Daley L, Vaughan L: Exploring Web 2.0 for virtual design studio teaching. ascilite Singapore 2007 Conference. 2 – 5 December 2007. Reference Source\n\nZehner R, Forsyth G, de la Harpe B , et al.: Optimising studio outcomes: Guidelines for curriculum development from the Australian studio teaching project. ConnectED2010–2nd International Conference on Design Education. Sydney: University of New South Wales; June 28th-July 1st. Retrieved on 25 April 2015. Reference Source\n\nMaher ML, Simoff S: Variations on the Virtual Design Studio. Proceedings of Computer-Supported Collaborative Work in Design. 1999. Reference Source\n\nFotaris P, Mavrommati I, Leinfellner R, et al.: Teaching Design from A distance: A case Study of Virtual Design Studio Teaching via A social Network. Proceedings of the 7th International Conference on Education and New Learning Technologies EDULEARN15. 2015. Reference Source\n\nSchön D: The Design Studio: an Exploration of its Traditions and Potentials. J. Archit. Educ. London: RIBA; 1985; 43(1): 53–55. Publisher Full Text\n\nIoannou O: Opening up design studio education using blended and networked formats. Int. J. Educ. Technol. High. Educ. 2018; 15: 47. Publisher Full Text\n\nMcGee P, Reis A: Blended course design: A synthesis of best practices. JALN. Jun 2012; v16(n4): p7–22. Publisher Full Text Reference Source\n\nTroha F: Bulletproof Blended Learning Design: Process, Principles, and Tips. Author house; October, 2003.\n\nAl Najadah AS: Challenges of E-learning for Interior Design and Art Education students studying at the College of Basic Education in Kuwait. J. Art Des. Music. 1(1): 1–8. Publisher Full Text\n\nIavarone AH: An evaluation of internet-based design studios in the context of learning styles. Yıldız Journal of Art and Design. June 2021; 8(1): 33–42. Publisher Full Text\n\nAssali I: A BLENDED MODEL OF INTERIOR DESIGN STUDIO PRACTICE DUE TO COVID-19. [Dataset]. figshare. 2022. Publisher Full Text\n\nAssali I: Blended Learning - Questionaire.pdf. Dataset. figshare. 2023. Publisher Full Text" }	[ { "id": "170043", "date": "25 Apr 2023", "name": "Noor Dheyaa Azeez", "expertise": [ "Reviewer Expertise Information technology", "information systems", "e-learning", "mobile government", "e-commerce", "decision-making systems." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe topic of study is vital and important.\nHowever, there are a set of observations that the researcher must follow:\nThe article did not mention the previous studies from which it was based. It is preferable to make a table that includes the previous studies, including the name of the study, the year, the title of the study, and the aim of the study. Then discuss these studies.\n\nIn the discussion section, the researcher did not compare the results of the article with the previous studies.\n\nAt the end of his article, the researcher did not provide a section for conclusion.\n\nThe researcher did not present the limitations of the article.\n\nThe article needs to enhance the statistical analysis method through the use of more advanced analytics.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "9653", "date": "20 Jun 2023", "name": "Imad Assali", "role": "Author Response", "response": "The article did not mention the previous studies from which it was based. It is preferable to make a table that includes the previous studies, including the name of the study, the year, the title of the study, and the aim of the study. Then discuss these studies. Answer: Thank you for the comments. Based on the comments, the details have been added in the revised article. 20, 21Several studies have been conducted on the impact of COVID-19 on various aspects of the design industry, including interior design. Some of the key aims of these studies have been to understand the challenges faced by designers during the pandemic and identify strategies for adapting to the new realities of remote work and social distancing. For example, 24a study published in 2020 explored the challenges faced by design educators in transitioning to online teaching during a pandemic. This study identified several key challenges, including technological barriers, difficulties in adapting to new teaching methods, and reduced opportunities for hands-on learning. 25A recent study examined the impact of the pandemic on interior design students' learning experience. The study found that students faced significant challenges in adapting to online learning but also identified several strategies that could be used to enhance their learning experiences in the future. Overall, these studies highlight the importance of adapting to new technologies and teaching methods to continue delivering high-quality interior design education during the pandemic. By adopting a blended approach to studio practice and combining both online and in-person learning opportunities, educators and students can continue to develop their skills and expertise despite the challenges posed by COVID-19. References: DeCoito I, Estaiteyeh M. Transitioning to Online Teaching During the COVID-19 Pandemic: an Exploration of STEM Teachers' Views, Successes, and Challenges. J Sci Educ Technol. 2022;31(3):340-356. Ulusoy, B. (2023). Effects of COVID-19 Pandemic on Students’ Written Outcomes: An Interior Architecture Research/Theory Module Case Study in the UK. Education Sciences, 13(1), 71. In the discussion section, the researcher did not compare the results of the article with the previous studies. Answer: Thank you for the comments. Based on the suggestions given, the results of this study were compared with previous studies. The COVID-19 pandemic has led to significant changes in the way businesses operate; interior design studios are no exception. As social distancing and remote work protocols have become the new norms, traditional studio-based practices have had to adapt to accommodate the restrictions imposed by the pandemic. A blended model of interior design studio practices has emerged as a way to balance the demands of client work with the need for safety and flexibility. This model combines in-person and remote work using technologies, such as video conferencing, file sharing, and collaborative software, to maintain team communication and project management. By allowing for both remote and in-person work, designers can also schedule client consultations in a way that balances the need for safety with the need for face-to-face interaction. A recent study showed that this blended model has proven to be effective, with interior design studios successfully completing projects on time and within budget, while maintaining the safety of both employees and clients. Compared to previous studies, this approach to interior design studio practice has been shown to be resilient and adapted to the current pandemic landscape. As the circumstances continue to evolve, it is likely that the blended model will remain a viable option for interior designers looking to navigate the post-pandemic landscape. 21Recent studies have explored the efficacy of a blended model in interior design studio practices in light of the COVID-19 pandemic. This model combines both in-person and online instruction and collaboration, allowing for social distancing and safer working conditions, while maintaining the benefits of traditional studio practice. 17One study found that students successfully engaged in design collaboration and critique through online platforms, suggesting that virtual communication tools could become an integral part of the design process even after the pandemic. However, the same study also highlighted the challenges of monitoring student progress in a virtual environment and maintaining a studio culture of mentorship and collaboration. 15Another study examined the use of augmented reality and virtual reality tools in the design process, which could enhance the collaborative experience of students and create a more immersive design experience. 18, 24While technology is still developing, it holds promise for supporting the blended model and even expanding the possibilities of interior design education. At the end of his article, the researcher did not provide a section for conclusion. Answer: Thank you for the comments. Based on the comments, conclusion section is added. The results of this study showed that blended design studios achieved pedagogical results as students developed their knowledge and skills, especially in computer software. The blended model of interior design studio practice has proven to be a successful approach for adapting to the challenges brought about by COVID-19. By combining virtual and in-person experiences, students and professionals can continue their work in a safe and efficient manner. The virtual aspect of the blended model allows for greater flexibility and accessibility, whereas the in-person component provides hands-on learning and collaboration opportunities. This hybrid approach also encourages the use of technology and problem-solving skills, which are essential in the current industrial landscape. The blended model is a valuable tool for interior design education and practice, and has the potential to continue beyond the pandemic. As we navigate this new normal, it is important to continue exploring innovative solutions that prioritize safety while maintaining the quality and effectiveness of our work. The researcher did not present the limitations of the article. Answer: Thank you for the comments. Based on the comments, limitations of the study are added. While this article presents a compelling argument for this blended model, it is important to note the limitations of the study. First, the study may not be generalizable to all interior design programs because it is based on the experience of one course. Different programs may have different teaching approaches and face different challenges. However, this study did not address the long-term effectiveness of the blended model. While it may have been effective in the short term because of the pandemic, it is unclear whether this model will continue to be effective in the future or if it will need to be modified as circumstances change. Overall, while the proposed blended model of interior design studio practice is promising, further research is required to fully understand its effectiveness and feasibility in various contexts. The article needs to enhance the statistical analysis method through the use of more advanced analytics Answer: Than you for the comments. Based on the comments, the details have been updated in the revised article. Enhancing statistical analysis methods through advanced analytics is essential to provide designers with the data-driven insights they need to create more valuable and customer-centric interior design solutions. It is a powerful tool that provides the ability to easily perform intricate analyses. Moreover, it includes a comprehensive range of statistical procedures that are useful for analyzing complex datasets. With the onset of the COVID-19 pandemic, interior design studio practices have been significantly impacted. The adoption of a blended model can help in mitigating the effects by enabling online collaboration. Using SPSS version 27.0.1.0 to analyzing data from the blended model can enable practitioners to derive meaningful insights from the collected data." } ] } ]	1	https://f1000research.com/articles/12-411
https://f1000research.com/articles/12-1428/v1	31 Oct 23	{ "type": "Research Article", "title": "Butterfly eyespots exhibit unique patterns of open chromatin", "authors": [ "Suriya Narayanan Murugesan", "Antónia Monteiro" ], "abstract": "Background: How the precise spatial regulation of genes is correlated with spatial variation in chromatin accessibilities is not yet clear. Previous studies that analysed chromatin from homogenates of whole-body parts of insects found little variation in chromatin accessibility across those parts, but single-cell studies of Drosophila brains showed extensive spatial variation in chromatin accessibility across that organ. In this work we studied the chromatin accessibility of butterfly wing tissue fated to differentiate distinct colors and patterns in pupal wings of Bicyclus anynana. Methods: We dissected small eyespot and adjacent control tissues from 3h pupae and performed ATAC-Seq to identify the chromatin accessibility differences between different sections of the wings. Results: We observed that three dissected wing regions showed unique chromatin accessibilities. Open chromatin regions specific to eyespot color patterns were highly enriched for binding motifs recognized by Suppressor of Hairless (Su(H)), Krüppel (Kr), Buttonhead (Btd) and Nubbin (Nub) transcription factors. Genes in the vicinity of the eyespot-specific open chromatin regions included those involved in wound healing and SMAD signal transduction pathways, previously proposed to be involved in eyespot development. Conclusions: We conclude that eyespot and non-eyespot tissue samples taken from the same wing have distinct patterns of chromatin accessibility, possibly driven by the eyespot-restricted expression of potential pioneer factors, such as Kr.", "keywords": [ "Chromatin accessibility", "ATAC-Seq", "Butterfly eyespots", "Notch", "wound healing" ], "content": "Introduction\n\nDynamic changes in chromatin opening and closing in a cell nucleus facilitates the binding of transcription factors and the expression of nearby genes. Chromatin is often opened via the binding of pioneer transcription factors, that start the process of unwinding DNA from its highly packed configuration around histones. This unwinding then exposes potential cis-regulatory elements (CREs) to other transcription factors (TFs). If these TFs are present in the cell, and if the newly opened regulatory sequences contain their respective target binding sequences, this second set of TFs, with the help of the pioneer factors, co-operatively bind DNA and begin the process of either activating or repressing nearby genes (Balsalobre and Drouin 2022; Iwafuchi-Doi et al. 2016).\n\nStudies examining the opening and closing dynamics of chromatin obtained from different tissues showed that most genes share the same chromatin profile at specific points in development, but have a different profile at different stages of development. For example, chromatin extracted at the same stage of development from legs, halteres, and wings of Drosophila, had the same chromatin profile, except for chromatin around tissue-specific selector genes (e.g. Ultrabithorax (Ubx), vestigial), that was uniquely open in that tissue (McKay, Estella, and Mann 2009; McKay and Lieb 2013). Chromatin profiles, however, were distinct between larval and pupal stages (van der Burg et al. 2019), suggesting that global developmental progression, driven by hormonal signals, rather than by tissue-specific patterns of gene expression, produced these distinct chromatin profiles.\n\nSimilarly, work on Heliconius and Junonia butterflies’ forewings and hindwings showed that except for the hindwing selector gene (Ubx), all other genes shared the same chromatin profile between the two wing types (van der Burg et al. 2019; Lewis et al. 2019; Lewis and Reed 2019). This was the case even for genes that were expressed only in a specific region of the Heliconius forewing, like Optix in the red band area, which showed no difference in its chromatin profile between fore and hindwings (Lewis et al. 2019; Lewis and Reed 2019). These results suggested, again, that all tissues sampled at the same time in development had the same chromatin profile.\n\nRecent work performed at higher resolution, however, contradicted these initial results. Chromatin profiles from small sections of Drosophila embryos had differences in chromatin accessibility, which were not identified when using whole embryos (Bozek et al. 2019). Similarly, there were extensive differences in chromatin accessibilities of single cells in developing brains of Drosophila (Janssens et al. 2022). These studies suggest, instead, that the similar chromatin profiles observed across tissues in earlier studies may be a result of averaging out distinct signals across many cells (McKay and Lieb 2013).\n\nHere we explored whether variation in chromatin accessibility can be associated with gene expression differences in a highly patterned tissue. We examined chromatin accessibility across the forewings of Bicyclus anynana butterflies, focusing on a region with eyespot patterns and two control regions next door. Eyespots consist of multiple concentric rings of color, and the central cells of these pattern elements appear to be reusing part of a limb gene regulatory network (GRN) for their differentiation (Murugesan et al. 2022). Hundreds of genes are differentially expressed (DE) in eyespot centers compared to adjacent regions in the wings (Murugesan et al. 2022; Özsu and Monteiro 2017). It is currently unclear, however, whether the eyespot DE genes display unique patterns of chromatin accessibility, compared to other genes. It is also unclear to what extent chromatin from eyespot regions, in general, differs in accessibility from chromatin from other wing regions. Pioneer factors, evenly expressed across the wing, might create the same chromatin profile across all wing cells, but the expression of unique TFs in the eyespot regions could lead to hundreds of DE genes in those regions. Alternatively, pioneer factors, expressed specifically in the eyespots, could produce eyespot-specific open chromatin regions that contribute to the differential regulation of genes in that region alone.\n\nTo investigate these alternate hypotheses, we performed fine dissections of eyespot and two flanking control wing tissues from 3 h old pupal forewings and conducted assays for transposase-accessible chromatin with Sequencing (ATAC-Seq) separately on each tissue type, and also for a complete wing, to identify genomic areas with open chromatin. We then identified differences in chromatin accessibility across these wing regions. Having found eyespot-specific regions of open-chromatin, we conducted a motif enrichment analysis for these regions to identify potential pioneer factors and/or specific TFs associated with eyespot development. Finally, we examined the ontology of genes that mapped to the vicinity of the uniquely open chromatin of eyespot tissues to identify their general identities.\n\n\nResults\n\nTo identify region-specific open chromatin, we first identified ATAC-Seq peaks that were consistently present across biological replicates of each wing region, and then added them across wing regions. A total of 166,062 peaks were identified across the three wing regions. Differentially open (DO) peaks between eyespot and control regions were identified by pairwise comparisons, as described in Murugesan et al. (2022). A comparison of eyespots with a non-eyespot sector region (NES1) in a more proximal part of the wing identified 562 peaks (397 + 165) that were DO in NES1, and these 562 peaks mapped to the vicinity of 506 genes (Figure 1B, Data S1). Similarly, a comparison of eyespots with a second non-eyespot sector region (NES2), in a more anterior part of the wing, identified 785 peaks (619 + 1 + 165) that were DO in NES2, and these peaks mapped to 647 genes (Figure 1B, Data S1). A comparison of eyespots with the two non-eyespot sector regions (NES1 and NES2) identified 82 eyespot-specific DO peaks that mapped to 80 genes (Figure 1B, 1C, Figure S1: Murugesan and Monteiro, 2023, Data S1). These data indicate that multiple distinct patterns of open chromatin are present across B. anynana 3 h old pupal forewings. Furthermore, only some of these patterns resemble the pattern of peaks obtained from ATAC-Seq performed on whole-wings of the same age (Figure 1C, Data S1).\n\n(A) Tissues chosen for the study were the eyespot region and two flanking control tissues (NES1 and NES2). (B) Number of peaks differentially open between different comparisons (differentially open- peaks are open in eyespot compared to NES1 or NES2 sector. Differentially closed – closed in eyespot and open in NES1 or NES2). (C) Example of an ATAC-peak next to the gene Notch that was DO in the eyespot region. Note how this peak was shown as open across the whole wing as well.\n\nOpen areas of chromatin often have specific genomic signatures/binding sites for pioneer TFs and for secondary TFs that have bound or will bind these regions in the future (Galupa et al. 2023). To examine whether specific DNA motifs were enriched in DO peaks we performed motif enrichment analysis. We identified nine motifs that were enriched in the 82 eyespot-specific ATAC-Seq peaks. Binding site motifs for the transcriptional regulator for the Notch signalling pathway Su(H) (present in 14.63% of the motifs identified), Adh transcription factor 1 (Adf1) (3.66%), pair-rule gene Runt (Run) (23.17%), Nubbin (Nub) (6.10%), gap genes Giant (Gt) (34.15%), Buttonhead (Btd) (19.51%), and Krüppel (Kr) (3.66%), Smad TF Medea (Med) (14.63%) and POL009.1_DCE_S_II (2.44%) were among the top enriched motifs (p-value < 0.01) in eyespot-specific peaks (Figure 2, Figure S2: Murugesan and Monteiro, 2023). In the more proximal NES1 wing region, we found 22 motifs that were enriched in the 562 DO peaks of this wing region (Figure S3: Murugesan and Monteiro, 2023). These motifs included binding sites for Pioneer factor Grainy-head (Grh), and TFs involved in the ecdysone signalling pathways such as Ultraspiracle (Usp), Broad (Br), and Hormone receptor-like in 46 (Hr46) (Figure S3: Murugesan and Monteiro, 2023). Similarly, 24 TF binding motifs were enriched in the 785 DO peaks of the more anterior NES2 wing region, with motifs for Hr46, Nubbin (Nub), Odd-skipped (odd), paired (Prd), Toll (Tll) being the most enriched (Figure S4: Murugesan and Monteiro, 2023). 10 TFs motifs were enriched in both NES1 and NES2 DO peaks, relative to eyespots, including Br, Hr46, Pangolin (Pan), and Dorsal (Dl) (Figures S3, S4: Murugesan and Monteiro, 2023).\n\nSuppressor of Hairless (Su(H)), Buttonhead (Btd), Adh transcription factor 1 (Adf1), Giant (Gt), Krüppel (Kr), Nubbin (Nub), Medea (Med), DCE_S_II predicted from JASPAR database, Runt (Run).\n\nIn order to identify a potential functional role for the genes in the vicinity of all the identified open chromatin regions, we performed an ontology analysis with those genes. The 166,062 peaks were first annotated for their position in the genome as part of promoters, distal intergenic sequences, or UTRs to nearby genes. More than 50% of the peaks were in distal intergenic regions and around 25% of the regions fell in promoter regions (Figure S5: Murugesan and Monteiro, 2023). Molecular pathways involved in wound-healing, cell differentiation, cell-cell signalling, and SMAD signal transduction were over-represented in the 80 genes found in the vicinity of the 82 eyespot-specific DO peaks (Figure 3B).\n\n(A) Pie chart and upset plot showing the percentage of the 82 eyespot-specific DO peaks that fall in distinct regions of the genome relative to nearby genes. This includes promoters, distal intergenic regions and UTRs. The maximum number of peaks falls in distal-intergenic and intergenic regions (36.59%). (B) Gene ontology for the 80 genes in the vicinity of the 82 eyespot-specific DO peaks.\n\n\nDiscussion\n\nIn this work, we examined the chromatin organization of butterfly eyespots and adjacent wing segments to test if the vast gene expression differences found across these different wing regions was also associated with differences in chromatin regulation. Previous transcriptomic experiments showed that ~800 genes were differentially expressed in eyespots relative to adjacent wing sectors, similar in size and location to the ones tested in the current study with ATAC-Seq (Murugesan et al. 2022; Özsu and Monteiro 2017). The large number of DE genes in the eyespot region is believed to be due, in part, to the co-option of a limb gene-regulatory subnetwork to the wing (Murugesan et al. 2022). Here we tested whether the same chromatin profile was present across all three wing segments, regardless of gene expression complexity, or whether pioneer factors, expressed specifically in the eyespots, were producing eyespot-specific open chromatin regions. We found that each of the three sectors of the wing investigated here had a unique chromatin profile, suggesting that DNA from distinct areas of the wing is being opened by sector-specific pioneer factors (Larson et al. 2021; Pihlajamaa et al. 2014). This contradicts the findings from McKay and Lieb, (2013) that showed that the only DO peaks between wings, halteres, legs cell homogenates were next to selector genes, such as Ubx and vestigial.\n\nThe large majority of the DE genes in eyespots (~800) had no variation in their chromatin accessibility in eyespot and control tissues. However, there were 82 chromatin regions that were differentially open in eyespots and closed in the other control wing regions. These chromatin regions had an over-representation of DNA motifs recognized by eight transcription factors (Figure 2) that may represent eyespot-specific pioneer factors or TF that act cooperatively with pioneer factors to regulate eyespot-specific genes. We discuss these genes below.\n\nEyespots DO peaks were enriched for Su(H) motifs. Su(H) is a transcription cofactor of the Notch signalling pathway (Baonza and Garcia-Bellido 2000; Bray and Furriols 2001), which can act as a repressor in the absence of Notch signalling (Bray and Furriols 2001; Larson et al. 2021). Notch signalling is activated by the presence of the ligand (Notch) (Baonza and Garcia-Bellido 2000; Bray and Furriols 2001). This pathway is likely active in eyespots as N shows an eyespot-specific DO peak (Figure 1C, Data S1) and is expressed in the eyespot centers of B. anynana as well as other nymphalid butterfly species (Oliver et al. 2012; Reed and Serfas 2004). N is also one of the four genes whose novel expression in the eyespot centers coincides with the origin of butterfly eyespots at the base of the nymphalids (Oliver et al. 2012; Monteiro 2015). The relative temporal expression of N and Distal-less (Dll) proteins in larval wings, indicated that Notch signalling precedes Dll expression in the eyespot centers (Reed and Serfas 2004). Dll is an essential gene in eyespot development (Connahs et al. 2019), but the role of N in eyespot development is not yet known. We propose that eyespot origins might be connected with a region-specific modification of chromatin accessibilities involving Su(H), essential for Notch signalling, in the eyespot regions of the wing.\n\nMotifs for the gap gene Btd, which belongs to the family of Sp genes, were also enriched in the eyespot-specific open peaks. Ectopic expression of btd in dorsal imaginal discs of Drosophila (wings, haltere and eyes) led to the formation of ventral structures (antenna and legs) through local activation of segment polarity genes engrailed (en) and wingless (wg) as well as decapentaplegic (dpp) (Estella et al. 2003). Eyespots may be reusing this ancestral limb network, because they also express en, wg and dpp at their centers (Keys et al. 1999; Monteiro et al. 2006; Özsu et al. 2017; Banerjee et al. 2022). As we haven’t observed any differential expression of btd in eyespots compared to NSE1 and NSE2 wing tissues (Murugesan et al. 2022; Özsu and Monteiro 2017), further expression analysis with in situ probes is required to test the expression of this gene in eyespots at earlier stages of development. Functional analysis should also test its potential role in eyespot development.\n\nTwo other gap genes, Kr and Gt, showed enrichment for eyespot-specific open chromatin regions. Kr acts as a downstream target gene in Notch-mediated cell differentiation in Malpighian tubules (MT) in Drosophila (Hoch and Jäckle 1998). Homologs of Kr in humans are called Krüppel-like factors (KLFs), and are involved in various cellular mechanisms along with Sp1-like proteins. KLF4 in humans is one of the pioneer factors that regulates cellular division, proliferation and differentiation (Borisova et al. 2022; Farrugia et al. 2016; Kaczynski, Cook, and Urrutia 2003). KLF4 along with Sox2, Oct4 and Nanog are essential for maintaining stem cell properties and their activation in somatic cells enables induced pluripotency (Liu et al. 2020). The enrichment of Kr in eyespot-specific peaks suggests that Kr could likely act as a potential pioneer factor under the influence of Notch signalling to open the eyespot peaks.\n\nGt is a gap gene that when disrupted shows broad regions of the body missing as well as segmentation defects (Bucher and Klingler 2004; Brent et al. 2007). Gt and Kr are expressed in a complementary non-overlapping manner in early Drosophila embryos, mutually repressing each other (Kraut and Levine 1991). It would be interesting to investigate the expression pattern of both genes in and around the eyespots in future.\n\nIn addition, TFs Nub and Med motifs were also enriched in eyespot-specific open peaks. Nub belongs to a POU-family of TFs. It is expressed in early wing primordium and is required for proximal-distal patterning of wings in Drosophila (Ng, Diaz-benjumea, and Cohen 1995; Zirin and Mann 2007). Med is a genetic modifier of Dpp signalling. It regulates wing patterning along the anterior-posterior axis through binding with the selector gene scalloped (sd) in Drosophila (Guss et al. 2001). The function of these TFs in eyespot development should be investigated in future.\n\nGiven the importance of ecdysone signalling and the presence of its receptor Ecdysone receptor (EcR) in eyespot centers, we looked for enrichment of EcR and other ecdysone signalling transcription factors that may play important roles in shaping the chromatin dynamics in the eyespot centers, compared to control tissues. However, we didn’t see any motif enrichment for EcR or other ecdysone TFs. For a long time, EcR has been thought to be a pioneer factor responding to ecdysone signalling and changing chromatin architecture (Uyehara and McKay 2019). Recent research in Drosophila wings and salivary glands, however, showed that EcR binds opportunistically to chromatin previously opened by tissue-specific pioneer factors, in particular Forkhead and Grainyhead (Uyehara et al. 2022). Also, in Junonia coenia butterflies, Spineless binding sites were enriched in chromatin peaks that were dynamically opening during wing metamorphosis, whereas EcR binding sites were only enriched in persistently open sites throughout development (van der Burg et al. 2019). These data point to EcR not being a pioneer factor for wing development. However, this needs to be further tested, as here we picked a single developmental stage, 3 h after pupation, before a large surge in 20E titers. Analysis of chromatin peaks right after this surge, not examined here, could have distinct motif enrichments, including those for EcR.\n\nHormone receptor-like 39 (Hr39) also showed eyespot-specific chromatin accessibilities (Figure S1: Murugesan and Monteiro, 2023). Hr39 is an orphan nuclear hormone receptor considered as a master regulator with a conserved role in female reproductive glands and secretive tissues in animals (Cattenoz et al. 2016; Praggastis et al. 2021). Hr39 is also a downstream canonical ecdysone response gene likely regulated through the EcR in wings (Uyehara and McKay 2019). Many butterfly eyespots express EcR at their center, and in B. anynana the expression of EcR in eyespot centers is tightly controlled by the ecdysone signalling pulse (Monteiro et al. 2015). The evolution of eyespots in nymphalid butterflies is also coincident with the novel expression of EcR in the eyespot centers (Bhardwaj et al. 2020). The enrichment of Hr39 binding sites in the open chromatin peaks specific to eyespot tissue could have facilitated ecdysone signalling in the eyespot centers and eyespot origins.\n\nIn summary, eyespot regions of the wing had areas of open chromatin that had an enrichment of DNA motifs recognized by a potential pioneer factor like Kr, as well as several other TFs. Some of these TFs have previously been suggested to be involved in eyespot development. Others should be examined in the future, including Kr.\n\nTo test if the genes next to the eyespot-specific open peaks were involved in any of the previously proposed signalling or molecular pathways involved in eyespot development, we performed gene ontology analysis. We found that wound healing is one of the significantly enriched gene ontologies along with SMAD signal transduction, cell-cell signalling and cell differentiation (Figure 3).\n\nPrevious work on eyespot DE genes using RNA-Seq highlighted many genes involved in wound-healing mechanisms (Özsu and Monteiro 2017), including Notch signalling (Vizcaya-Molina et al. 2018). Piercing pupal wings produces ectopic eyespots and activates genes involved in eyespot development in the wounded region (Monteiro et al. 2006), suggesting that eyespot and wound healing GRNs share common genes and regulatory elements. In our previous work, we identified two ATAC-Seq peaks next to Distal-less and spalt that were open in eyespots, whole wings, as well as wounded wings (Murugesan et al. 2022). The extent that additional genes and regulatory elements are shared between eyespots and wound healing needs to be further explored to understand how these two GRNs are related.\n\nSMAD signal transduction was also enriched in our gene ontology analysis. SMAD proteins are the intracellular mediators of both Dpp and Wnt signalling (Wrana and Attisano 2000), and both Smad2 and Smad6 are differentially expressed in the eyespot centers (Murugesan et al. 2022). Furthermore, both Dpp and Wingless are potential morphogens involved in eyespot development, and CRISPR knockout of both genes led to the loss of eyespots (Banerjee et al. 2022).\n\nWe propose that co-option of pre-existing GRNs, such as the limb GRN (Murugesan et al. 2022) and/or the wound healing GRN (Özsu and Monteiro 2017) to the novel location where eyespots develop was potentially mediated via the generation of a novel eyespot-specific open chromatin region next to a top-regulator of the network. This would allow that top regulator to have a novel activity in a novel domain of the body of the butterfly. Any of the eighty genes with DO chromatin identified in this study might be such a top regulator. Furthermore, we propose that Kr, the only confirmed pioneer factor whose binding sites were enriched in the eyespot-specific open chromatin, is a good candidate for mediating these eyespot-specific patterns of open chromatin.\n\nOverall, our study shows that eyespots show differential chromatin accessibility compared to two other nearby wing regions. Eyespot-specific accessibility peaks are enriched for motifs and gene ontology corresponding to many of the genes and pathways previously proposed to be involved in eyespot development. This clearly demonstrates that chromatin accessibility varies across the wing, and with specific wing patterns in B. anynana butterflies.\n\nThe main limitation of this study lies in the tissue collection method adopted to distinguish eyespot from non-eyespot cells. Though we were able to capture differences in chromatin accessibilities between the different sections of the wings, dissections of eyespot tissues contain non-eyespot cells which might hinder the precision in identifying the ATAC peaks corresponding to eyespot cells alone. One way to overcome this limitation in future is to use single-cell sequencing approaches, and eyespot-specific marker genes, to study ATAC-peaks only on those marker-expressing candidate cells.\n\n\nMethods\n\nBicyclus anynana larvae and adults were reared in 27°C and 60% humidity inside a climate room with 12:12 h of dark and light cycle. Adults were fed with bananas and larvae were fed on young corn plants.\n\nATAC-Seq was performed for the tissues as described in Murugesan et al. (2022). Two replicates were made for the eyespot-specific sector and three replicates for each control sector. Eyespot dataset was taken from Murugesan et al. (2022). The control tissues NES1 and NES2 were from the same forewings as eyespot tissues dissected around 3 h to 6 h after pupation. We dissected as many tissues as possible within 15 minutes, and then snap-froze them using liquid nitrogen. Tissues were stored at -80°C before nuclei extraction. We pooled 50 wing bits from the same wing area to produce a single biological replicate, which contained approximately 80,000 nuclei. Prepared libraries were sequenced using Novoseq 6000, with an average read depth of 30 million reads per library and 2×50 bp paired-end at Novogene, Singapore. We also used 3 h pupal whole wing data from Murugesan et al. (2022) as a control dataset for the peaks identified from micro dissections.\n\nATAC-Seq analysis was carried out as described in Murugesan et al. (2022) with few modifications. Reads obtained from the sequencing were processed using bbduk scripts from bbmap tools (Bushnell 2014) to remove any adapters. Reads were mapped to Bicyclus anynana genome, version BaGv2 (Murugesan et al. 2022) using bowtie. Reads mapped to the mitochondrial genome and marked for duplicates were removed using samtools idxstats and GATK MarkDuplicates respectively. Peak calling was done using Fseq (Boyle et al. 2008). Consensus peaks for all samples were obtained using bedtools intersect, followed by bedtools merge with peaks within 50 bp distance from each other resulting in a total of 166,062 peaks. The consensus peaks were used for all downstream analyses. FRiP score was used to measure the fraction of reads mapped to the genome falling into peaks. Our ATAC-Seq data showed a median FRiP score of 0.41 which is greater than the ENCODE standard (>0.3) (Table 1). Deeptools (Ramírez et al. 2014) were used to measure the correlation between replicates and quality of the ATAC-Seq data. A read count matrix was produced corresponding to the consensus peaks for the libraries using Featurecount from Subread tools (Liao, Smyth, and Shi 2014). Differential peak analysis was performed using DESeq2 (Love, Huber, and Anders 2014) and the peaks were considered differentially accessible with a p-value < 0.05. Peaks were called differentially open (DO) if their logFC > 0 and p-value ≤ 0.05. Quality check for the ATAC data was done using Deeptools (Figure S6: Murugesan and Monteiro, 2023).\n\nRead depth and FRiP score.\n\nConsensus ATAC peaks and subset of peaks obtained from the differential peaks were all annotated using the R package ChiPseeker (Bushnell 2014). The gene annotation gff file for BaGv2 genome version was loaded into the pipeline using GenomicFeatures (Lawrence et al. 2013) library and was used to locate the position of peaks in the genome with respect to the gene structures. Promoters were defined as peaks 3000 bp upstream and downstream of the gene’s transcription start site. The distal-intergenic peaks are mapped to genes which are within 300 Kb from the transcription start site (Data S1)\n\nGenes corresponding to the differential peaks were obtained from the ChiPseeker annotation. Gene ontology was carried out as described in Murugesan et al. (2022). Gene ontology for the BaGv2 genome was obtained from Interproscan (Jones et al. 2014). ClusterProfiler package (Wu et al. 2021) in R was used to identify the enriched gene ontology for the given gene lists.\n\nHomer findMotifsGenome.pl (Heinz et al. 2010) was used to identify enriched motifs in differentially accessible regions identified from DESEq2. Bed files for the peaks of interest were produced. BaGv2 genome was incorporated into Homer to run locally. Transcription factor binding motifs were searched just within the peaks’ exact sizes, with GC normalization for the background sequences and in insect databases in Homer.\n\n\nAuthors contribution\n\nSNM and AM designed the study. SNM carried out the experiment and analysed the data. SNM and AM wrote the manuscript. All authors read and approve the manuscript.", "appendix": "Data availability\n\nNCBI BioProject: Bicyclus anynana (squinting bush brown). Accession number: PRJNA685019; ATAC-Seq reads for eyespot and control tissues are submitted in NCBI under Bioproject, https://identifiers.org/bioproject:PRJNA685019 (Murugesan et al. 2023).\n\nBiosamples: SAMN17214324, SAMN17214325, SAMN33923641, SAMN33923642, SAMN33923643, SAMN33923644, SAMN33923645, SAMN33923646.\n\nDRYAD: Data for: Butterfly eyespots exhibit unique patterns of open chromatin, https://doi.org/10.5061/dryad.q573n5tp9 (Murugesan and Monteiro 2023).\n\nThis project contains the following extended data:\n\n- Data for: Butterfly eyespots exhibit unique patterns of open chromatin\n\n- Figure S1\n\n- Figure S2\n\n- Figure S3\n\n- Figure S4\n\n- Figure S5\n\n- Figure S6\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nReferences\n\nBalsalobre A, Drouin J: Pioneer Factors as Master Regulators of the Epigenome and Cell Fate. Nat. Rev. Mol. Cell Biol. 2022; 23(7): 449–464. 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PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "267378", "date": "19 Apr 2024", "name": "Richard ffrench-Constant", "expertise": [ "Reviewer Expertise butterfly wing pattern formation" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis is a really nice paper that begins to fill the gap between gene and phenotype in butterfly wing pattern formation. By carefully staging and then carefully dissecting developing eyespot tissue the authors have shown how chromatin structure differs with wing scale development. This now allows us to identify potential transcription factors in an unbiased manner (in other words not simply looking for those for which we already have probes or antibodies). This sort of approach should pay dividends if used across other butterfly wing pattern elements. Congratulations to the authors.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "232273", "date": "11 May 2024", "name": "Craig Brunetti", "expertise": [ "Reviewer Expertise virology", "immunology", "butterfly wing patterning." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nPrevious studies on insect whole-body parts have demonstrated little variation in chromatin accessibility. More recently, studies have shown that in fact variation may exist if smaller regions or single cells are tested. The rationale is that the average of chromatin accessibility over a body part may mask variations within subsets of the tissue. The manuscript “Butterfly eyespots exhibit unique patterns of open chromatin” explores chromatin accessibility in various pattern elements on the wings of Bicyclus anynana. ATAC-Seq was performed on 3hr pupal wing discs from either the eyespot or adjacent non-eyespot tissue. The authors demonstrated that there were distinct open chromatin regions in the eyespot-specific tissue versus control tissue. Areas of open chromatin corresponded to pathways previously implicated in eyespot formation. These data suggest that there is variation in open chromatin across tissues and provides further insight into the patterning of wings of butterflies.\nSome comments related to the manuscript:\nIn the 5th paragraph of the Introduction, second sentence, change “control regions next door” to “adjacent control regions” Figure 3B shows a large % of the genes in the eyespot DO set are involved in wound healing. There should be a mention in the discussion about why this isn’t the result of wound healing induced during surgical removal of the tissue. Highlighting the fact that the tissue is removed quickly and fast frozen (within 15 minutes) along with the fact that this is differential expression and induction of wound healing would happen in all tissue (eyespot and control).\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1428
https://f1000research.com/articles/12-1427/v1	31 Oct 23	{ "type": "Study Protocol", "title": "Effectiveness of sensory stimulation and early mobility on consciousness, mental state, RLA stage, and hospital stay in patients with Traumatic brain injury: A research protocol", "authors": [ "Devyani Purushe", "Moh’d Irshad Qureshi", "Raghumahanti Raghuveer", "Moh’d Irshad Qureshi", "Raghumahanti Raghuveer" ], "abstract": "TBI is the most difficult to treat and has the highest mortality and morbidity rates in ICU. The most common cause of neurological abnormalities and physical disabilities in people is traumatic brain injury (TBI). Brain injury occurs when an external force causes a change in brain function or other signs of brain pathology. TBI is a leading cause of epilepsy and is linked to a high morbidity and mortality rate. Over the last few decades, India has seen an increase in traumatic brain injury (TBI) as a result of increased transportation development, a rapidly growing construction industry, and urbanization. A total of 30 patients with traumatic brain injuries will be included in this experimental study, and they will be evenly divided into two groups. Sensory stimulation with early mobility will be delivered to Group A, while traditional physiotherapy will be given to Group B for two weeks at 40 minutes per session, two sessions per day, 5 days a week. The Coma Recovery scale, Mini-mental scale examination (MMSE), Glasgow Coma Scale (GCS), Ranchos Los Amigos scale for cognitive functioning, and length of stay in ICU will be used as outcome measures. The goal of this study is to see how early mobility exercises and sensory stimulation affect consciousness, mental state, RLA, and hospital stay when used in conjunction with physiotherapy.\nCTRI Reference Number CTRI/2022/07/044127", "keywords": [ "Sensory stimulation", "Consciousness", "Mental state", "RLA", "Traumatic Brain Injury", "Research Protocol." ], "content": "Introduction\n\nThe most common causes of neurological anomaly and physical disabilities in persons is traumatic brain injury (TBI). It encompasses both sensory and motor capabilities that have an impact on one's overall quality of life.1 TBI is the most difficult to treat and has the greatest morbidity and fatality rates in intensive care units.1 TBI is thought to affect 200 people per 100,000 in wealthy countries each year.2 The country's rapid development in transportation, construction, and urbanization in recent decades has resulted in a high risk of traumatic brain injury.3\n\nThe most vulnerable to TBI are adolescent/young adults, children (under 25 years old), and the elderly. A severe TBI may occur in one-third of patients, resulting in long-term cognitive and behavioural deficits. In 20% of cases, brain traumas cause a multisystem condition due to related injuries, which might involve complicated neurological deficits, neuroendocrine, and neuro medical issues.3\n\nTransportation network safety improvements, alcohol and drug awareness programs, and regulations requiring the use of safety belts and helmets have all been important prevention strategies.4\n\nTBI has been linked to a large number of neuromuscular, cognitive, and behavioural disorders, all of which can impair activity, social involvement, and overall quality of life.5 Motor function is often affected in people who have suffered a TBI. Paresis in the upper and lower extremities, as well as decreased coordination, postural control, aberrant tone, and irregular gait, can all be life-long deficits.6 Tremor and chorea are examples of abnormal, involuntary movements, but dystonic motions are less prevalent. Patients may also have compromised somatosensory function, depending on where the lesion is located.7\n\nThe mental process of knowing and using knowledge is referred to as cognition. Arousal, attention, focus, memory, learning, and executive functions are among the many complicated brain processes that make up cognition. People with altered states of awareness are common. Arousal states such as vegetative state, coma, and minimally conscious state are common after severe brain trauma.5 Cognitive impairment was the primary contributor to disability in most brain-injured subjects who scored moderate to severe on the GOS.8\n\nThe Glasgow Coma Scale is frequently used to determine whether a traumatic brain injury is severe, moderate, or mild (GCS). Teasdale and Jennett developed the GCS, which is the most widely used clinical scale for measuring consciousness and the severity of an injury. The GCS is made up of three different types of responses: motor, verbal, and eye-opening. When the individual response scores are added together, the total score ranges from 3 to 15. A chronic injury is defined as one with a score of 8 or less, a serious brain injury as one with a score of 9 to 12, and a light brain injury as one with a score of 13 to 15.5\n\nPhysical therapy's main purpose is to minimize secondary consequences from the TBI and prolonged bedrest/immobilization, start early mobilization after receiving medical clearance, and start patient and family education. Monitoring vitals and other comorbidities is used to manage respiratory treatment, coma stimulation, and passive/active mobility for patients.1 So, the goal of the study will be conducted to see how early mobility exercises and sensory stimulation affect awareness, mental state, and RLA staging in traumatic brain injury patients. To identify the impact of early mobility exercises and sensory stimulation on consciousness using physiotherapy intervention and the goal of this study will be to see how helpful physiotherapy maybe for early mobility exercises and sensory stimulation during an ICU stay.\n\nThe goal of the study is to see how early mobility exercises and sensory stimulation affect awareness, mental state, and RLA staging in traumatic brain injury patients.\n\nThe following are the study's goals:\n\n1) To determine the impact of early mobility exercises and sensory stimulation on consciousness using physiotherapy intervention.\n\n2) This research aims to see how effective physiotherapy intervention on early mobility exercises and sensory stimulation is for mental health.\n\n3) To determine the efficacy of physiotherapy intervention on early mobility exercises and sensory stimulation in patients with RLA stage.\n\n4) The goal of this study is to see how helpful physiotherapy maybe for early mobility exercises and sensory stimulation during an ICU stay.\n\nStudy setting\n\nFollowing ethical approval by the Institutional Ethical Committee of the Datta Meghe Institute of Medical Science the Reference number is DMIMS (DU)/IEC/2022/896 and date is 11/04/2022, the study will be carried out in the Intensive Care Unit (ICU) of the Acharya Vinobha Bhave Rural Hospital (AVBRH) in the Sawangi Meghe Wardha.\n\nStudy design\n\nIt is a two arm parallel interventional study. The total amount of participants in Group A will be 15, while Group B will have 15 (n=30).\n\nParticipants\n\nInclusion criteria: Patients aged 35 to 60 years old with mild to moderate traumatic brain injury with or without craniotomy and a Glasgow Coma Scale (GCS) score of 13.\n\nExclusion criteria: The patient with a history of visual and Auditory dysfunction (blindness, color blindness, deafness, hearing loss). The Patient with alcohol and drug in toxification with he/she is on anticonvulsants were excluded.\n\nRecruitment procedure\n\nPatients admitted to the Intensive Care Unit with traumatic brain injuries (ICU) at Acharya Vinoba Bhave Rural Hospital and meet the criteria for inclusion will be included in the study. Patients will be randomly allocated into two groups using simple random sampling and will be assigned through sequentially numbered opaque sealed envelopes. The experimental group will receive sensory stimulation and early mobility exercises whereas the control group will receive traditional physiotherapy.\n\nParticipant timeline\n\nThe study will last one year, and the physiotherapy intervention will last 30 minutes per session, two times per day, five days a week for two weeks.\n\nImplementation\n\nThe research coordinator and principal investigator will be in charge of randomization. For recruitment into either group, participants will be asked to choose a sealed group allocation from an envelope.\n\nBlinding\n\nTo assign the subjects to the groups, the tester(s) will be blinded. Subjects will be required not to reveal any details about their treatment to the tester in order to ensure binding.\n\nFollowing the ethical committee's approval, the participants will be given a thorough explanation of the technique, and their informed consent will be obtained. The study will include all those who are willing to participate.\n\nThe research will be implemented in the ICU setting. The patients will be chosen based on the inclusion criteria listed above. After obtaining signed informed consent from a caregiver or family member, patients who meet the inclusion criteria will be enrolled in this study. Informed consent will be taken by caregiver as the patient will be unaware about the situation. Simple random sampling will be used to assign patients to one of two groups, which will be assigned through sequentially numbered opaque sealed envelopes. The patients’ blood relatives or primary caregiver is the point of contact for randomization and allocation. The experimental group will be receiving Sensory stimulation and early mobility exercises whereas the control group will receive conventional physiotherapy. For two weeks, both groups will receive 30 minutes of instruction per session, two times per day.\n\n40 min a session, two sessions per day, 5 days a week for 2 weeks.\n\nThe program of sensory stimulation was divided into five sections:\n\n1. Awakening for 5 Minutes\n\nThe position of Therapist and caregiver will be at the side of the patient's bed. The therapist and caregiver will call out the patient's name and give commands for an eye-opening. The patient's response will be noted for the response. If in case, the patient does not open the eyes, wet gauze will be applied to the patient's eyes.\n\n2. Auditory Stimulation for 10 Minutes\n\nThe therapist and caregiver first clean the earplugs of the earphones with a spirit swab and then connect the earplugs to the mobile, will put the earplugs in the patient's ears and turn on the Carnatic instrumental theme (Hare Krishna Hare Krishna, Krishna Krishna Hare Hare). It will be given for 10 minutes. If patient will be applied the same musical note repeatedly to get accustomed to the music.\n\n3. Visual Stimulation for 10 Minutes\n\nThe physiotherapist will open the patient's right eye with his/her left hand and stimulated it with bright light on/off for 1 second and will repeat it for 10 seconds. The source of light will be a torch held in the therapist’s right hand. After 10 seconds, the same will be repeated for the left eye.\n\n4. Tactile Stimulation for 5 Minutes\n\nTactile stimulation will be aimed to provide several elements of tactile sense such as touch, pressure, and temperature. The tactile sense of touch will be given with the help of a wisp of cotton, the pressure was given with the use of a comb, and the temperature will be provided with the help of hot and cold packs. This stimulation shall be given to all four limbs of the patient. Each stimulus will be given every 5 seconds. The response will be varied from patient to patient. If patients have difficulty recognizing different types of touch, such as light touch versus deep pressure then repeated tactile stimulation will be given.\n\n5. Olfactory Stimulation for 10 Seconds\n\nThe therapist or caregiver, first, will clean each nostril with the earbuds. Then one nostril will be closed with the thumb, and the coffee or camphor or spices will be kept near the opposite nostril of the patient, so that the patient could smell and the same will be repeated in the other nostril. Herbs such as cloves will be used with the help of cotton for 10 seconds. The stimulus which will be used placed 3 cm from the nostrils. It will be given for 10 seconds after every 15 seconds. If the patient will be having difficulty perceiving the distinct aroma of coffee and the strong scent of camphor or the fragrance of clove, then repeated olfactory stimulation will be given.9\n\nEarly mobility: Early mobility is described as starting the mobility program when the patient can participate in therapy minimally, has a stable hemodynamic status, and is obtaining adequate oxygen levels.\n\nBecause it meets parts of therapy goals during the early stages of recovery, upright sitting is particularly significant. The patient should be transported to a sitting posture and out of bed onto a wheelchair as soon as they are medically stable. All safety precautions should be followed. The head should be properly supported because the patient is unlikely to have adequate neck and head control to maintain an upright posture without assistance. Co-treatment with an occupational therapist provides two skilled professionals to assist the patient because maximal assistance is often required. The use of a tilt table is also advantageous because it allows early weight-bearing through the LEs. The upright position, both on a tilt table and in a wheelchair, may improve the overall level of alertness.10\n\nGroup B- subjects in the control group will undergo traditional physiotherapy, which will involve passive limb exercises as well as an in-bed posture to avoid pressure sores. The treatment would last 40 minutes.\n\nPre and post outcome measures will be taken by utilizing the Coma Recovery Scale (CRS), Mini-mental scale examination (MMSE), Glasgow Coma Scale (GCS), and Ranchos Los amigos scale parameters by the assessor who will be aware of the outcome measures and having a similar experience that of the Physiotherapy resident doing the study. The assessor will be blinded for the treatment.\n\nComa Recovery Scale (CRS)\n\nThe Revised JFK Coma Recovery Scale (CRS-R) is a tool for assessing patients who will be in a level of awareness such as coma. It can help distinguish the vegetative state (VS) from the minimally conscious state (MCS). It will also be used to track the transition from a semi-conscious to conscious state.\n\nReliability: 0.94\n\nValidity: 0.9711\n\nMini-mental scale examination (MMSE)\n\nThe Mini-Mental State Examination (MMSE) is a reliable and effective tool for determining whether or not an older person has cognitive impairment. By including detailed administration and scoring criteria, the original instrument's reliability was improved (the Standardized MMSE [SMMSE]). The SMMSE is a thorough assessment tool for seniors. It generates a global score of cognitive capacity that is linked to daily task performance.\n\nReliability: 0.80-0.95\n\nGlasgow Coma Scale (GCS)\n\nThe Glasgow Coma Scale is used to categorize traumatic brain damage as severe, moderate, or mild (GCS). The GCS was created by Teasdale and Jennett and is the most extensively used clinical scale for evaluating consciousness and helps in the characterization and classification of injury severity. The GCS is composed of three reaction scores: verbal response, motor response, and eye-opening. The sum of the individual activity scores yields a total score ranging from 3 to 15. A score of 8 or less indicates severe brain injury, a score of 9 to 12 indicates serious brain injury, and a score of 13 to 15 indicates mild brain injury.12\n\nReliability- 0.86\n\nAcute care professionals also use the Rancho Los Amigos Scale (RLA) (Hagen, 1984) to classify a patient's condition as well as decide discharge placement. It is divided into ten degrees of cognitive functioning:\n\nNo response, generalized response, localized response, confused agitated, confused-inappropriate, confused-appropriate, automatic-appropriate, purposeful-appropriate, Purposeful, Appropriate: Stand by Assistance and Purposeful, Appropriate: Stand by Assistance on request.5\n\nReliability- 0.87\n\nValidity- 0.92\n\nThe subject shall be examined immediately after the coma stimulation and the scoring shall be done for GCS, CRS, MMSE, and RLA scales. The scoring shall be done on the 0th day and after 15 days.\n\nSample size Calculation: Daniel formula for sample size (1999).\n\nZα/2 is the level of significance at 5% i.e., 95% Confidence interval= 1.96\n\nP = Prevalence of TBI= 0.97% = 0.0097\n\nD = Desired error of margin= 5% = 0.05\n\n= 15 patients needed in each group\n\nAs per this article G. Gururaj et alEpidemiology of traumatic brain injuries: Indian scenario. The prevalence of TBI is p value = 0.0097.13\n\nAnalysis\n\nResults over the outcome variables will be tabulated and described using descriptive statistics; data over the outcome variables will be tested for normal distribution for the mean and standard deviation (SD) mean statistics will be positioned for finding skewed distributions and interquartile range (IQR). Frequency and percentages for binary and categorical variables will be tabulated for descriptive statistics. SPSS 27.0 software will be used for all statistical analysis.\n\nWe are planning to publish in an Indexed journal.\n\nThe Recruitment of participants has been done.\n\n\nDiscussion\n\nThe goal of the protocol will be conducted to see how sensory stimulation and early mobility affect patients with traumatic brain injury's consciousness, mental state, RLA stage, and hospital stay. This sensory stimulation is broken down into five sections: Five minutes of awakening, ten minutes of visual stimulation ten minutes of auditory stimulation, and five minutes of tactile stimulation and olfactory stimulation for ten seconds, with early mobility exercises in between. This will be given to one group and for other group we will give passive limb exercises as well as an in-bed posture to avoid pressure sores.\n\n\nConclusion\n\nThis study will show effectiveness of early mobility exercises and sensory stimulation on the level of consciousness, mental state, RLA stage and ICU stay in individuals with traumatic brain injury.", "appendix": "Acknowledgements\n\nI would like to acknowledge Mr. Laxmikant Umate Sir, who has helped me in sample size calculation and data analysis planning.\n\n\nReferences\n\nThomas A, Shravani B, Poulose L, et al.: Early Mobilisation Improves Conscious Levels, Cognitive Levels and Reduction in Number of Days in Intensive Care Unit Stay in Moderate to Severe Traumatic Brain Injury Patients-A Hospital Based Study.2018 Mar 1.\n\nBruns J, Hauser WA: The Epidemiology of Traumatic Brain Injury: A Review: EPIDEMIOLOGY OF TRAUMATIC BRAIN INJURY. Epilepsia. 2003 Sep 26; 44: 2–10. Publisher Full Text\n\nPanwar N, Purohit D, Deo Sinha V, et al.: Evaluation of extent and pattern of neurocognitive functions in mild and moderate traumatic brain injury patients by using Montreal Cognitive Assessment (MoCA) score as a screening tool: An observational study from India. Asian J. Psychiatr. 2019 Mar; 41: 60–65. PubMed Abstract \| Publisher Full Text\n\nBorg J, Röe C, Nordenbo A, et al.: Trends and Challenges in the Early Rehabilitation of Patients with Traumatic Brain Injury: A Scandinavian Perspective. Am. J. Phys. Med. Rehabil. 2011 Jan; 90(1): 65–73. PubMed Abstract \| Publisher Full Text\n\nChesnut RM, Carney N, Maynard H, et al.: Evidence Report on Rehabilitation of Persons with Traumatic Brain Injury.287.\n\nHellweg S, Johannes S: Physiotherapy after traumatic brain injury: A systematic review of the literature. Brain Inj. 2008 Jan; 22(5): 365–373. Publisher Full Text\n\nChua KS, Ng YS, Yap SG, et al.: A Brief Review of Traumatic Brain Injury Rehabilitation. A Brief Review of Traumatic Brain Injury Rehabilitation. 2007; 36(1): 31–42. Publisher Full Text\n\nHellweg S: Effectiveness of Physiotherapy and Occupational Therapy after Traumatic Brain Injury in the Intensive Care Unit. Crit. Care Res. Prac. 2012; 2012: 1–5. Publisher Full Text\n\nPassler MA, Riggs RV: Positive outcomes in traumatic brain injury–vegetative state: Patients treated with bromocriptine. Arch. Phys. Med. Rehabil. 2001 Mar; 82(3): 311–315. PubMed Abstract \| Publisher Full Text\n\nZaninotto AL, El-Hagrassy MM, Green JR, et al.: Transcranial direct current stimulation (tDCS) effects on traumatic brain injury (TBI) recovery: A systematic review. Dement Neuropsychol. 2019 Jun; 13(2): 172–179. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLippert-GrÜner M, Wedekind C, Klug N: Outcome of prolonged coma following severe traumatic brain injury. Brain Inj. 2003 Jan; 17(1): 49–54. Publisher Full Text\n\nPhysiopedia: [cited 2022 Feb 12]. Glasgow Coma Scale.Reference Source\n\nGururaj G: Epidemiology of traumatic brain injuries: Indian scenario. Neurol. Res. 2002 Jan; 24(1): 24–28. PubMed Abstract \| Publisher Full Text" }	[ { "id": "264987", "date": "30 Apr 2024", "name": "Domenico Intiso", "expertise": [ "Reviewer Expertise severe acquired brain injury", "rehabilitation", "spasticity", "stroke", "critical illness polyneuropathy and myopathy" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors proposed a study protocol to investigate the effects of sensory stimulation and early mobilization on the recovery of consciousness, mental state and long of stay of adult traumatic brain injury subjects admitted in ICU. Although the study is interesting severe criticisms arise about the protocol and method design.\nBackground is bare and based on outdated references. Some of them are incomplete (ref. 1 and 5). The choice of early mobilization and sensory stimulation to facilitate the outcome of TBI subjects and particularly the recovery of consciousness is poorly motivated and explained. The reasons to investigate these rehabilitative intervention are poorly addressed and the introduction is predominantly conceived and organized in relation to one reference. The authors should highlight the role and the effect of mobilization on the recovery and focus the aim of study based on recent literature data. Indeed, a lot of studies about early mobilization in critically ill patients has been published in particular to prevent muscular weakness (1), but several questions remain unsolved about early mobilization in neurological ICU patients(Bartolo M, et al., 2017[Ref 1]), (Maia TFLD, et al., 2024[Ref 2]). The authors affirmed that inclusion criteria were: patients aged 35 to 60 years old with mild to moderate traumatic brain injury with or without craniotomy and a Glasgow Coma Scale (GCS) score of 13. In this respect is not clear the reason to employ CRS as measure. Indeed, this scale is generally used in subjects unresponsive or in coma, but inclusion criteria considered only subjects with moderate or mild TBI or subjects graded to GCS 13. This score generally is observed in awake, responsive patients. The choice of CRS is conflicting and unclear. Likewise, the use of sensory stimulation in responsive subjects is unclear given that those are awake (GCS 13) and might be conscious, even if confusion and agitation might be observed. However, the authors described the delivery of sensory stimulation (auditory, visual, tactile, etc.) as if subject was unresponsive. Furthermore is unclear the timing of start the early mobilization. The authors affirmed that the mobility program start when the patient can participate in therapy minimally, has a stable hemodynamic status, and is obtaining adequate oxygen levels. The patient should be transported to a sitting posture and out of bed onto a wheelchair as soon as they are medically stable. However, this condition might occur late during ICU stay opposing the early mobilization. Early or late mobilization is debated. Recent literature show different modality to early mobilization (Huebner L, et al., 2024[Ref 3]). No clinical and neurological variable commonly occurring during ICU stay that can act on the recovery is considered. Likewise, no previous clinical or neurological disease that can bias the finding has been considered in exclusion criteria.\n\nIs the rationale for, and objectives of, the study clearly described? No\n\nIs the study design appropriate for the research question? No\n\nAre sufficient details of the methods provided to allow replication by others? Partly\n\nAre the datasets clearly presented in a useable and accessible format? Not applicable", "responses": [] } ]	1	https://f1000research.com/articles/12-1427
https://f1000research.com/articles/12-1424/v1	31 Oct 23	{ "type": "Research Article", "title": "Endometriosis and sexual disorders: the effect of surgical and medical treatment, a multicentre cross-sectional study", "authors": [ "Tommaso Capezzuoli", "Elisa Maseroli", "Fabio Barra", "Silvia Vannuccini", "Linda Vignozzi", "Paola De Mitri", "Silvia Baggio", "Marcello Ceccaroni", "Felice Petraglia", "Tommaso Capezzuoli", "Elisa Maseroli", "Fabio Barra", "Silvia Vannuccini", "Linda Vignozzi", "Paola De Mitri", "Silvia Baggio", "Marcello Ceccaroni" ], "abstract": "Background Sexual health is a major concern in women with endometriosis, however only a few controlled studies have examined this with validated instruments. The effect of hormonal treatments on sexual function in endometriosis is also an underrated topic. The aim of this study was to investigate sexual function of patients with endometriosis by a specific tool to better evaluate their sexual function (including different domains), and the influence of hormonal treatment or surgery on these parameters.\n\nMethods An observational, cross-sectional, multicentre study was conducted in a group (n=194) of sexually active, women aged 25–45 years old, with surgical or ultrasonographic diagnosis of endometriosis, referred to the Endometriosis Center of Careggi University Hospital or Negrar di Valpolicella. Sexual function was assessed by administering the Female Sexual Function Index (FSFI), which assesses the domains of desire, arousal, lubrication, orgasm, satisfaction and pain. FSFI scores were compared to those of a control group (n=58) and according to the treatment received by patients with endometriosis.\n\nResults Ovarian endometriosis was present in 50 patients (25.8%), deep infiltrating endometriosis in 65 patients (33.5%) and both in 79 patients (40.7%). Adenomyosis coexisted in 102 patients (52.6%). Women with endometriosis reported a mean total FSFI score of 18.3 [4.2-25.8] (< 26.55), indicating female sexual dysfunction (FSD) in all patients. At multivariate analysis, after adjusting for confounders (BMI and hormonal therapy), women with endometriosis presented significantly lower scores than controls in all the FSFI (p<0.001). Patients with endometriosis under hormonal treatments (n=124; 64%), regardless of the type, had significantly lower scores in all FSFI subscales and total score, even after adjusting for confounders—age, BMI and history of surgery.\n\nConclusions Patients with endometriosis are at risk for FSD, encompassing not only dyspareunia, but all domains of sexual function. Hormonal treatments do not result in improvement in sexual symptoms.", "keywords": [ "Endometriosis", "sexual health", "hormonal treatment", "surgical treatment", "female sexual dysfunction" ], "content": "Introduction\n\nEndometriosis is a benign chronic inflammatory disease defined as the presence of endometrium outside of the uterine cavity, and the most recognized mechanisms that explain the ectopic location of the endometrial cells are retrograde menstruation and stem cell differentiation.1 The disease affects 10% of women at reproductive age and it is characterized by painful symptoms and infertility, affecting quality of life (QoL).2–5\n\nConsidering that patients with endometriosis are young and sexually active, sexual health is a major concern and should be carefully evaluated.6 In fact, endometriosis is associated with reduced psychological well-being, and impaired sexual life and relationships. Patients often complain about dyspareunia, decreased sexual satisfaction, painful sex and overall altered sexual functioning,7 driving to an impaired quality of sexual life with a significant negative effect on their relationships.8–11 Besides, the coexistence of adenomyosis is associated with a worse quality of sexual life than only endometriosis.12,13\n\nMoreover, multiple surgery and long-term hormonal treatments may represent additional contributing factors to negative sexual function in endometriosis. Induced transient menopausal state associated with some hormonal treatments, such as gonadotropin releasing hormone analogues (GnRHa), affects brain areas involved in sexual response. Also combined oral contraceptives (COCs)14 and progestin treatment may induce a change in sexual functioning.15 In fact, despite medical treatment having a role in endometriosis management given its efficacy on painful symptoms (e.g., dyspareunia) and prevention of disease recurrence and progression, it may influence the sexual well-being.7\n\nThe aim of the present study was to investigate sexual function of patients with endometriosis by a specific tool to better evaluate their sexual function (including different domains, i.e., desire, arousal, lubrification, orgasm, satisfaction and pain), and the influence of hormonal treatment or surgery on these parameters.\n\n\nMethods\n\nThe study was approved by the local Institutional Review Board (Comitato Etico Regione Toscana-Area Vasta Centro-CEAVC, n. 14558_oss approved on 28 May 2019), and all participants provided written informed consent to be included in the series.\n\nAn observational cross-sectional multicentre study was conducted in a group (n=194) of fertile age women (25–45 years old) with endometriosis, consecutively referred to two different hospitals, both reference Centres for Endometriosis (Careggi University Hospital, Florence and Negrar di Valpolicella, Verona, Italy) and recruited between January 2022 and February 2023.\n\nThe study group was constituted of patients with surgical or imaging diagnosis of endometriosis (mean age was 35.45 ± 7.44 years). Ovarian endometriosis (OMA) was present in 50 patients (25.8%), deep infiltrating endometriosis (DIE) in 65 patients (33.5%) and both in 79 patients (40.7%). Adenomyosis coexisted also in 102 patients (52.6%). Previous surgery for endometriosis was performed by 24.2% (n=47) patients: 37 (78.7%) underwent one previous surgery, whereas 21.3% more than two surgeries for endometriosis. Regarding medical treatments, 64% (n=124) were under a hormonal therapy. The majority of patients were treated with dienogest (59.7%, n=74), followed by those treated with COCs (21%, n=26), desogestrel (7.3%, n=9), GnRHa (4%, n=5), norethisterone acetate (NETA) (3.2%, n=4), drospirenone (2.4%, n=2) and levonorgestrel intrauterine system (LNG-IUS) (2.4%, n=3).\n\nA group of women without endometriosis attending the two Women’s Endocrinology outpatient clinics were enrolled as the control group (n= 58) (mean age was 34.55 ± 8.76 years). Controls were consulting for contraceptive needs, or for follow-up for thyroid or metabolic disorders. The abovementioned endocrinologic conditions were clinically stable since at least six months, respondent to treatment and presenting laboratory or imaging parameters within the normal ranges. These were the criteria to be eligible as controls in the study. Data were collected by an extensive review of clinical records of patients in the follow-up of these outpatient clinics.\n\nExclusion criteria were: menopausal status, pregnancy, desire of pregnancy when the survey was conducted or previously attempts to conceive, both naturally or through assisted reproductive technologies, breastfeeding, systemic diseases—including previous or active cancer, polycystic ovary syndrome, hyperandrogenism, hyperprolactinemia, uncontrolled psychiatric diseases, alcohol or drug abuse, and use of medications with a possible influence on sexual function except for hormonal contraception (i.e., antidepressant and anxiolytic drugs). The ability to provide written informed consent and having engaged in sexual activity in the previous month were considered as inclusion criteria.\n\nDuring the follow-up visit, patients were interviewed through:\n\n(i) a structured questionnaire containing all clinical information regarding the history of the patient (in particular age, body mass index (BMI), parity and current use of hormonal treatment). The hormonal treatments used were: progestins, GnRHa or continuous COCs, for a minimum of 12 months;\n\n(ii) a structured questionnaire containing all clinical information about female sexual function. Sexual symptoms were investigated by using the gold standard tool for the screening of Female Sexual Dysfunction (FSD), the Female Sexual Function Index (FSFI). This self-administered questionnaire analyses overall levels of sexual function and its primary components: sexual desire, arousal, lubrication, orgasm, pain, and satisfaction.16 The FSFI is composed by items with answers codified on a 5-point Likert scale ranging from 1 to 5, with higher scores indicating greater levels of sexual functioning for each item. The total score, resulting from the sum of the five domains, ranges from 2 to 36; a total score of 26.55 has been found to provide an excellent cut-off to distinguish women with and without FSD.17\n\nAll participants filled out the answers to the FSFI questionnaire themselves, whereas baseline and medical data were asked by a healthcare professional.\n\nData were reported as mean ± SD when normally distributed, as median (quartiles) when non-normally distributed and as percentage and number when categorical. The unpaired 2-sided Student’s t-test and the Mann-Whitney U test were applied for the assessment of between-group differences, whenever appropriate. Multivariate models, with adjustment for relevant clinical confounders, were conducted by means of analysis of covariance. Statistical analyses were performed in SPSS 26.0 IBM SPSS Statistics (RRID:SCR_016479) for Windows (SPSS Inc, Chicago, IL, USA).\n\n\nResults\n\nCases and controls were similar for age, whereas those with endometriosis showed a lower BMI than controls (22.55 ± 3.81 vs. 24.74 ± 6.93, p=0.002) and were more likely to use hormonal therapy (64% vs. 19%, p<0.001). In terms of sexual function, women with endometriosis reported a mean total FSFI score of 18.3 [4.2-25.8] (< 26.55), indicating FSD in all patients. Conversely, those in the control group obtained a mean total score of 32.0 [28.8-33.4], thus excluding FSD. Comparing each FSFI domain, at univariate analysis, women with endometriosis presented significantly lower scores than controls, in desire, arousal, lubrication, orgasm, satisfaction and pain (p<0.001) (Table 1), indicating sexual functioning impairment.\n\nMultivariate analysis was adjusted for age, BMI and use of hormonal therapy. The symbol * indicates statistically significant difference. BMI = body mass index. FSFI = Female Sexual Function Index. HT = hormonal therapy. Data were reported as mean ± SD when normally distributed, as median (quartiles) when non-normally distributed and as percentage and number when categorical. Multivariate models, with adjustment for relevant clinical confounders, were conducted by means of analysis of covariance.\n\nAt multivariate analysis, after adjusting for confounding factors (BMI and use of hormonal therapy), all the reported differences between the two groups retained statistical significance (p<0.001 for all the scores; Table 1).\n\nIn a second step, sexual function in patients with endometriosis was further investigated, exploring the differences between those taking (64%, n=124) and not taking (36%, n=70) hormonal treatment (GnRHa, progestins, oral contraceptives) and the potential influence of previous surgery for endometriosis. Hormonal therapies, regardless of the type, were associated with significantly lower scores in all FSFI subscales and in total score, indicating worse sexual functioning. After adjusting for confounders—age, BMI and history of surgery—all the differences retained statistical significance (Table 2).\n\nMultivariate analysis was adjusted for age, body mass index, and a history of previous surgery for endometriosis. The symbol * indicates statistically significant difference. indicates statistically significant difference. FSFI = Female Sexual Function Index. Multivariate models, with adjustment for relevant clinical confounders, were conducted by means of analysis of covariance.\n\n\nDiscussion\n\nThe present study confirmed that patients with endometriosis have worse sexual function compared to healthy controls in all FSFI domains (desire, arousal, lubrification, orgasm, satisfaction and pain) and FSFI total score.\n\nOur results confirmed those of a recent meta-analysis18 showing that patients with endometriosis have lower scores of FSFI total score with poor sexual function. Considering that endometriosis is a disease that affects sexually active young women, the evaluation of sexual function in the context of patient’s global management is crucial in order to improve the global QoL.7 In fact, endometriosis is not only characterized by sexual pain, but they also present an important impairment in several domains of sexuality (desire, arousal, lubrification, orgasm, satisfaction). A recent study showed patients with endometriosis have a worse score in the short-form of McGill Pain Questionnaire (SF-MPQ), pain subscale of FSFI, and Sexual Distress Scale (FSDS).19 Furthermore, they reported more negative emotions toward sexuality and seem to be characterized by an impairment in body image,20 depressive symptoms,21 worse health related QoL (HRQoL) and unemployed work status.22\n\nSeveral mechanisms play a role in increasing the risk of FSD in women with endometriosis. First, in women with dyspareunia, fear and anticipation of pain strongly affect the global sexual response, thus increasing sexual inhibition and reducing spontaneous desire and sexual fantasy. In addition, psychological and interpersonal correlates of endometriosis, including fertility issues, low self-esteem, and body image concerns, contribute to negatively affect sexual function in its different areas.7 In recent years, evidence is accumulating that indicates a relevant overlap between endometriosis and superficial dyspareunia, and a high prevalence of Genito-pelvic pain and penetration disorder (GPPPD) in endometriosis.23 These comorbidities are also likely to play a role in compromising the sexual experience of affected women.\n\nConsidering that endometriosis is a chronic condition, medical treatment is the primary choice for improving symptoms, preventing or treating recurrences and planning surgery or ART. GnRH analogues or antagonists, progestins, combined oral contraceptives block cyclic menstruation and reduce endometriosis-related pain.1,5 The present study showed that hormonal treatment, regardless of the type, is not associated with an improvement in sexual function, showing lower scores in all FSFI subscales and total score. After adjusting for confounders—age, BMI and history of surgery—all the differences retained statistical significance.\n\nThe strong hypoestrogenic effect of GnRH agonists seems to be associated with a significant decline in libido and vaginal lubrication.6 Despite the use oral contraceptives and progestins in healthy women was previously described to be associated with negative sexual side effects (sexual activity, arousal, pleasure and orgasm and more difficulty with lubrication),6,14 a recent observational study detected an improvement of sexual quality of life in patients with DIE with or without adenomyosis after 12 months of treatment with a combined oral contraceptive (2 mg dienogest/30 μg ethinyl oestradiol).13\n\nConsidering the effect of surgical treatment, it is an option that works on pain relief and improvement of quality of sex life in symptomatic women with endometriosis.24,25 On the other hand, persistent or recurrent endometriosis after unsuccessful first-line conservative surgery is associated with severe deep dyspareunia and low FSFI score, below the cut-off for normal sexual function.6,26 Furthermore, the comparison between surgical treatment versus a low-dose progestin therapy among patients with deep dyspareunia shows an immediate significant improvement of pain after surgery, but recurrent over time; on the contrary, those on low dose of NETA have a slight decrease in dyspareunia, though progressively declining.26 Medical and surgical treatment should be carefully evaluated because they often do not consistently allow for the global improvement of sexual function, despite their strong effect on painful symptoms of affected patients.\n\nFinally, endometriosis is frequently associated with gynaecological and systemic comorbidities that may cause sexual dysfunction and impair the Qol.11,27–29 Adenomyosis determines further a high rate of altered sexual function in patients with endometriosis.12,13 Autoimmune, inflammatory, psychiatric and neurological disorders are commonly described in patients with endometriosis30,31 and have a strong effect on global QoL and also sexuality. Therefore, the evaluation of eventual gynaecological and systemic comorbidities is mandatory in patients with endometriosis.\n\nThe present study has some limitations. First, sexual-related distress was not assessed, and this is a relevant aspect of sexual dysfunction. Second, all women were sexually active, but the relational component (i.e., the presence of a stable relationship, couple conflicts, sexual dysfunction in the male partner) was not evaluated. Furthermore, our study population is a selected sample of patients with severe endometriosis referred to highly specialized centres and most likely include cases with recurrent symptoms after either surgical or hormonal treatment.\n\nIn conclusion, sexual dysfunction is a common finding in patients with endometriosis and a multidisciplinary approach, including a psychological support and the contribution of other specialists for systemic comorbidities, is warranted.7 In fact, the traditional hormonal or surgical management do not significantly improve such as an important aspect as sexual function, and a multimodal approach is required.", "appendix": "Data availability\n\nTo protect the patients’ privacy the present study data access was restricted. The anonymous data about patients’ sexual function can be shared with readers and reviewers. To apply for access to the data, readers or reviewers can contact Dr. Tommaso Capezzuoli ( tommasocapezzuoli@unifi.it ). While applying for access, reader or reviewer should give a signed letter mentioning that they will not share the data with a third party and it will used only for academic purpose.\n\n\nReferences\n\nCapezzuoli T, Rossi M, La Torre F, et al.: Hormonal drugs for the treatment of endometriosis. Curr. Opin. Pharmacol. 2022 Dec; 67: 102311. Publisher Full Text\n\nChapron C, Marcellin L, Borghese B, et al.: Rethinking mechanisms, diagnosis and management of endometriosis. Nat. Rev. Endocrinol. 2019 Nov; 15(11): 666–682. PubMed Abstract \| Publisher Full Text\n\nZondervan KT, Becker CM, Missmer SA: Endometriosis. N. Engl. J. Med. 2020 Mar 26; 382(13): 1244–1256. Publisher Full Text\n\nSaunders PTK, Horne AW: Endometriosis: Etiology, pathobiology, and therapeutic prospects. Cell. 2021 May 27; 184(11): 2807–2824. PubMed Abstract \| Publisher Full Text\n\nVannuccini S, Clemenza S, Rossi M, et al.: Hormonal treatments for endometriosis: The endocrine background. Rev. Endocr. Metab. Disord. 2022 Jun; 23(3): 333–355. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPluchino N, Wenger JM, Petignat P, et al.: Sexual function in endometriosis patients and their partners: effect of the disease and consequences of treatment. Hum. Reprod. Update. 2016 Nov; 22(6): 762–774. PubMed Abstract \| Publisher Full Text\n\nVannuccini S, Maseroli E, Vignozzi L, et al.: The challenge of endometriosis for female sexual health. J. Sex. Med. 2023 Feb 27; 20(3): 240–246. PubMed Abstract \| Publisher Full Text\n\nDenny E, Mann CH: Endometriosis-associated dyspareunia: the impact on women’s lives. J. Fam. Plann. Reprod. Health Care. 2007; 33: 189–193. PubMed Abstract \| Publisher Full Text\n\nFlynn KE, Lin L, Bruner DW, et al.: Sexual Satisfaction and the Importance of Sexual Health to Quality of Life Throughout the Life Course of U.S. Adults. J. Sex. Med. 2016 Nov; 13(11): 1642–1650. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLukic A, Di Properzio M, De Carlo S, et al.: Quality of sex life in endometriosis patients with deep dyspareunia before and after laparoscopic treatment. Arch. Gynecol. Obstet. 2016 Mar; 293(3): 583–590. PubMed Abstract \| Publisher Full Text\n\nMissmer SA, Tu FF, Agarwal SK, et al.: Impact of Endometriosis on Life-Course Potential: A Narrative Review. Int. J. Gen. Med. 2021 Jan 7; 14: 9–25. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAlcalde AM, Martínez-Zamora MÁ, Gracia M, et al.: Assessment of Sexual Quality of Life and Satisfaction in Couple Relationships Among Women With Deep Infiltrating Endometriosis and Adenomyosis. J. Sex Marital. Ther. 2022; 48(3): 263–272. Publisher Full Text\n\nAlcalde AM, Martínez-Zamora MÁ, Gracia M, et al.: Assessment of Quality of Life, Sexual Quality of Life, and Pain Symptoms in Deep Infiltrating Endometriosis Patients With or Without Associated Adenomyosis and the Influence of a Flexible Extended Combined Oral Contraceptive Regimen: Results of a Prospective, Observational Study. J. Sex. Med. 2022 Feb; 19(2): 311–318. PubMed Abstract \| Publisher Full Text\n\nBoth S, Lew-Starowicz M, Luria M, et al.: Hormonal Contraception and Female Sexuality: Position Statements from the European Society of Sexual Medicine (ESSM). J. Sex. Med. 2019 Nov; 16(11): 1681–1695. PubMed Abstract \| Publisher Full Text\n\nOppenheimer A, Verdun S, Perot M, et al.: Do high-dose progestins impair sexual function in women treated for endometriosis? A prospective observational longitudinal study. Acta. Obstet. Gynecol. Scand. 2021 May; 100(5): 850–859. PubMed Abstract \| Publisher Full Text\n\nRosen R, Brown C, Heiman J, et al.: The Female Sexual Function Index (FSFI): a multidimensional self-report instrument for the assessment of female sexual function. J. Sex. Marital. Ther. 2000 Apr-Jun; 26(2): 191–208. PubMed Abstract \| Publisher Full Text\n\nWiegel M, Meston C, Rosen R: The Female Sexual Function Index (FSFI): Cross-validation and development of clinical cutoff scores. J. Sex. Marital. Ther. 2005; 31: 1–20. PubMed Abstract \| Publisher Full Text\n\nShi C, Xu H, Zhang T, et al.: Endometriosis decreases female sexual function and increases pain severity: a meta-analysis. Arch. Gynecol. Obstet. 2023 Jan; 307(1): 195–204. PubMed Abstract \| Publisher Full Text\n\nRossi V, Galizia R, Tripodi F, et al.: Endometriosis and Sexual Functioning: How Much Do Cognitive and Psycho-Emotional Factors Matter? Int. J. Environ. Res. Public Health. 2022 Apr 27; 19(9): 5319. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMelis I, Litta P, Nappi L, et al.: Sexual Function in Women with Deep Endometriosis: Correlation with Quality of Life, Intensity of Pain, Depression, Anxiety, and Body Image. Int. J. Sex. Health. 2015; 27: 175–185. Publisher Full Text\n\nDe Graaff AA, Van Lankveld J, Smits LJ, et al.: Dyspareunia and depressive symptoms are associated with impaired sexual functioning in women with endometriosis, whereas sexual functioning in their male partners is not affected. Hum. Reprod. 2016 Nov; 31(11): 2577–2586. PubMed Abstract \| Publisher Full Text\n\nvan Poll M , van Barneveld E , Aerts L, et al.: Endometriosis and Sexual Quality of Life. Sex. Med. 2020 Sep; 8(3): 532–544. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWahl KJ, Orr NL, Lisonek M, et al.: Deep Dyspareunia, Superficial Dyspareunia, and Infertility Concerns Among Women With Endometriosis: A Cross-Sectional Study. Sex. Med. 2020; 8(2): 274–281. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDubuisson J, Pont M, Roy P, et al.: Sexualité féminine après chirurgie pour endométriose pelvienne profonde [Female sexuality after surgical treatment of symptomatic deep pelvic endometriosis]. Gynecol. Obstet. Fertil. 2013 Jan; 41(1): 38–44. PubMed Abstract \| Publisher Full Text\n\nFritzer N, Hudelist G: Love is a pain? Quality of sex life after surgical resection of endometriosis: a review. Eur. J. Obstet. Gynecol. Reprod. Biol. 2017 Feb; 209: 72–76. PubMed Abstract \| Publisher Full Text\n\nVercellini P, Frattaruolo MP, Somigliana E, et al.: Surgical versus low-dose progestin treatment for endometriosis-associated severe deep dyspareunia II: effect on sexual functioning, psychological status and health-related quality of life. Hum. Reprod. 2013; 28: 1221–1230. PubMed Abstract \| Publisher Full Text\n\nVannuccini S, Lazzeri L, Orlandini C, et al.: Mental health, pain symptoms and systemic comorbidities in women with endometriosis: a cross-sectional study. J. Psychosom. Obstet. Gynaecol. 2018 Dec; 39(4): 315–320. PubMed Abstract \| Publisher Full Text\n\nCapezzuoli T, Vannuccini S, Fantappiè G, et al.: Ultrasound findings in infertile women with endometriosis: evidence of concomitant uterine disorders. Gynecol. Endocrinol. 2020 Sep; 36(9): 808–812. Publisher Full Text\n\nCapezzuoli T, Orlandi G, Clemenza S, et al.: Gynaecologic and Systemic Comorbidities in Patients with Endometriosis: Impact on Quality of Life and Global Health. Clin. Exp. Obstet. Gynecol. 2022; 49(7): 157. Publisher Full Text\n\nShigesi N, Kvaskoff M, Kirtley S, et al.: The association between endometriosis and autoimmune diseases: a systematic review and meta-analysis. Hum. Reprod. Update. 2019 Jul 1; 25(4): 486–503. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nShafrir AL, Farland LV, Shah DK, et al.: Risk for and consequences of endometriosis: A critical epidemiologic review. Best Pract. Res. Clin. Obstet. Gynaecol. 2018 Aug; 51: 1–15. PubMed Abstract \| Publisher Full Text" }	[ { "id": "231806", "date": "21 Feb 2024", "name": "Maria Grazia Porpora", "expertise": [ "Reviewer Expertise endometriosis", "adenomyosis", "pelvic pain", "minimally invasive surgery" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nI have carefully read the manuscript entitled Endometriosis and sexual disorders: the effect of surgical and medical treatment, a multicentre cross-sectional study, by Tommaso Capezzuoli, Elisa Maseroli, Fabio Barra, Silvia Vannuccini, Linda Vignozzi, Paola De Mitri, Silvia Baggio, Marcello Ceccaroni and Felice Petraglia. The study aimed to analyze del sexual functioning in women with endometriosis and /or adenomyosis. In this multicentric study, all women with surgical or sonographic diagnosis of endometriosis or adenomyosis answered to a structured questionnaire containing all clinical information on their medical history and to the FSFI questionnaire which is validated for the evaluation of the sexual function. Results were compared with those of a control group. The authors found that all affected women, independently from the type of lesions, had a lower FSFI score than controls, indicating FSD in all patients. Women taking hormonal therapies had worse results in terms of FSD.\nThe paper is interesting and well written, but I have a few comments for the Authors:\n1. Surgery, particularly when involving the bowel and its function, can worsen the self- body image perception and the presence of bowel problems significantly affects sexual functioning (Boyd T, et al 2022.) [Ref 1]; did you find any difference between those who had undergone surgery and those who received only medical treatments? In addition, did you find differences in sexual function according to the type of surgery?\n2. Surprisingly, in this study, all kinds of medical therapy worsened the sexual functioning of patients. These results can be easily justified in women taking GnRh analogues or even oral progestins but the detrimental effects on sexual functions are less clear in women taking oral contraceptives. Do you think that the lack of difference between treatments could be related to the small number of patients in each group? Could it be possible that a long history of disease may have affected the quality of life and have determined psychological problems, thus influencing the results?\n\n3. Psychological factors and mood disorders were not evaluated in this paper, but they may have influenced the results. Please make a comment about this.\n\n4. It is also possible that marital status and a stable affective relationship can improve the patient’s sexual life as observed by Giuliani et al. in 2016 (Giuliani et al. 2016)[Ref2]. Could you please make a comment on this aspect?”\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "285939", "date": "11 Jul 2024", "name": "Giussy Barbara", "expertise": [ "Reviewer Expertise gynecology", "endometriosis", "sexual health" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nI read with interest the article by Capezzuoli et al. entitled \"Endometriosis and sexual disorders: the effect of surgical and medical treatment, a multicenter cross-sectional study\". The authors conducted a multicenter cross-sectional study to evaluate sexual function in women with and without endometriosis using the FSFI. Several studies have already investigated the impact of endometriosis on sexual function, and it is well known that endometriosis has a negative impact on female sexuality. However, in this study, the authors found that patients with endometriosis under hormonal treatment (n=124; 64%), regardless of type, had significantly lower scores on all FSFI subscales and total score, even after adjusting for confounders - age, BMI, and history of surgery. This is a somewhat novel finding and warrants further investigation. However, so far studies have suggested that medical treatment of endometriosis can reduce pain during intercourse and consequently improve sexuality. However, female sexuality is a very complex dimension, influenced not only by pain but also by several other potential factors, both psychological and social and relational. My suggestion to the authors is to discuss this point in the paper as a limitation of the results, since they did not have an investigation of psychological, social and relational factors, and this could have had an impact on the results.\nOverall, the paper is well written and the discussion is logically supported by the results. I recommend indexing.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1424
https://f1000research.com/articles/12-503/v1	16 May 23	{ "type": "Research Article", "title": "Study of significance of bone marrow microvessel density in myeloproliferative neoplasms in correlation with CD34 blasts, mast cell count and fibrosis", "authors": [ "Kesiya Thomas", "Ranjitha Rao", "Chaithra G V", "Sharada Rai", "Sneha Rao A R", "Kudurugundi Basavaraju Vatsala", "Kesiya Thomas", "Chaithra G V", "Sharada Rai", "Sneha Rao A R", "Kudurugundi Basavaraju Vatsala" ], "abstract": "Background: Myeloproliferative neoplasms (MPN) are clonal hematopoietic stem cell diseases characterised by myeloid cell growth from one or more lineages. Angiogenesis, in contrast to other subtypes, plays a substantial role in the pathophysiology of primary myelofibrosis (PMF). Research expressing the correlation of microvessel density (MVD), blasts, fibrosis and mast cell count in MPN cases are rarely conducted. We aimed to study the significance of MVD in correlation with CD34 blasts, mast cells and fibrosis in bone marrow biopsies of MPN patients. Methods: The current research was a cross sectional study conducted on 66 cases diagnosed as MPN during a six-year period. This comprised of 32 chronic myeloid leukemia (CML), 31 PMF and three essential thrombocythemia (ET) cases. Routine staining along with reticulin stain to look for fibrosis and immunohistochemistry (IHC) using CD34 and mast cell tryptase (MCT) were performed. Results: We found increased MVD in PMF, when compared to CML and ET (p = 0.042). Further, mean MVD was observed to be increased with high blast counts (p = 0.036). On follow up, raised mean MVD was seen in those cases with relapse/deceased as compared to disease-free patients, which was highly significant (p = 0.000). Conclusions: Increased MVD score was mostly associated with PMF subtype among all the MPNs. Further, higher MVD was observed to be associated with increased blast count and poor prognosis. With angiogenesis playing a critical role in disease outcome, we now have drugs to regulate angiogenesis that are supported by contemporary research. However, further studies with larger cohorts to establish the theranostic role of MVD in MPNs is recommended.", "keywords": [ "myeloproliferative neoplasms", "microvessel density", "CD34", "mast cell tryptase" ], "content": "Introduction\n\nMyeloproliferative neoplasms (MPN) are a category of disorders characterised by enhanced erythroid, megakaryocytic, or granulocytic cell proliferation.1 The global incidence of MPNs ranges from 1.15 to 4.99 instances per 100,000 individuals.2 In a review article by Titmarsh and others, the highest incidences of chronic myeloid leukaemia (CML) and essential thrombocythemia (ET) were found to be in Europe, but primary myelofibrosis (PMF) was more prevalent in Australia.3 Varma and colleagues found that in India, 231 out of 18,14,298 patients (0.0127%) were diagnosed as MPN, out of which, 207 (89.6%) were CML with BCR-ABL mutation. The remaining MPN cases (n = 24/231, 10.4%) were BCR-ABL negative, the majority of whom were polycythemia vera (PV), which was identified in 11 cases (4.7%), followed by PMF (n = 7, 3%) and ET (n = 6, 2.6%) cases.4\n\nSeveral studies have demonstrated the significance of neoangiogenesis in tumour growth and progression, but only a few have focused on malignant haematological diseases.5 Angiogenesis is important in the pathophysiology of PMF but less so in PV and ET. In myelofibrosis, transforming growth factor-β (TGFB) and vascular endothelial growth factor (VEGF) are two cytokines that help to enhance the stromal response that entails angiogenesis. The principal focus of angiogenesis is in the bone marrow, which is typically measured as vessel concentration and expressed as MVD.6\n\nPatients with MPN also have higher numbers of bone marrow mast cells. Mast cells originate from multipotent hematopoietic progenitor cells in the bone marrow, but they often do not mature there. Instead, they circulate as immature progenitors via the circulatory system to complete their development peripherally in connective or mucosal tissues.7,8 Many inflammatory cells surround tumour cells, including mast cells, which can secrete many such proangiogenic factors, leading to endothelial cell proliferation and angiogenesis.8\n\nStudies expressing the significance of MVD in specific MPNs like CML and PMF are done in the past; however, studies to explore the relationship between mast cells, blasts, bone marrow fibrosis and MVD are seldom available in literature. Hence, this study is undertaken to understand the significance of MVD in various MPNs, and to ascertain its relation with blast count, mast cell count and fibrosis along with its role in recovery/prognosis.\n\n\nMethods\n\nThe current research is a cross sectional study of cases diagnosed over six years duration spanning from January 2016 to December 2022 (five years two months retrospective and 10 months prospective).\n\nThis study follows ‘The Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) statement guidelines. A completed STROBE checklist can be found in the reporting guidelines.9\n\nWe included 66 cases diagnosed as MPN during a six-year period. This comprised of 32 chronic myeloid leukemia (CML), 31 PMF and three essential thrombocythemia (ET) cases.\n\nThe Institutional Ethics Committee (Reg. No. ECR/541/Inst/KA/2014/RR20; DHR Reg. No. EC/NEW/INST/2020/742), provided ethical clearance on 17th February 2022 with reference no. IEC KMC MLR 12-2020/444 after submitting the proposal letter beforehand. The ethics committee also waived the informed consent as the study is performed on archived tissue blocks.\n\nAll bone marrow biopsies diagnosed as MPNs in our laboratory during the study duration were included. Cases in which the blocks and/or slides were not available or the tissue was inadequate for immunohistochemistry (IHC) were excluded.\n\nAll previously histomorphologically diagnosed MPNs were retrieved from the Pathology department archives and clinical characteristics, including age, gender, symptoms, signs, laboratory investigations, genetic mutations, treatment and follow up details were gathered from the medical records and patient case record files. A total 66 cases of MPN were studied, out of which 32 cases were CML, 31 cases were PMF and three cases were ET. Bone marrow biopsies were received in 10% neutral buffered formalin fixative. Paraffin embedded tissue blocks were made. Sections of four-micron thickness were prepared using a microtome (Leica RM2255) and stained using haematoxylin and eosin (H&E).\n\nPreparation of Mayer’s hematoxylin was done by dissolving one gram of hematoxylin in one litre of water in a bottle. Aluminium sulphate was then dissolved in a small amount of water in a separate beaker, followed by addition of 0.2 g of sodium iodate to this solution and mixed properly. This was then added to the hematoxylin solution bottle and mixed properly. Finally, 0.2 g of citric acid crystals were added and mixed.\n\nEosin stain was prepared using Eosin Y powder (10 g) dissolved in distilled water (1000 ml) and 1 ml of glacial acetic acid. The mixture was filtered to get a total of 1000 ml of eosin.\n\nFollowing section cutting, deparaffinization was caried out first, by incubating the slides in an oven for 40 minutes. They were then cooled and dipped in xylene two times, first for 15 minutes and the second time for 10 minutes. The slides were air dried after which they were dipped in absolute alcohol twice for one minute each. They were then air-dried following which they were dipped in water for five minutes. Following deparaffinisation, sections were washed in tap water. They were dipped for 15 minutes in Mayer’s hematoxylin and kept for 5–10 minutes in running tap water to ensure ‘bluing’. Next, they were dipped for 30 seconds in eosin stain and then washed in tap water, dried, and mounted.\n\nMicroscopic examination of the cases was based on morphology according to the World Health Organisation classification as per the data proforma9 by using an Olympus CX 21i microscope. MPNs were typed according to the WHO 2016 classification. This is comprised of chronic myeloid leukemia (CML), BCR-ABL1 positive, chronic neutrophilic leukemia (CNL), polycythemia vera (PV), primary myelofibrosis (PMF) i) Prefibrotic/early stage ii) Overt fibrotic stage, essential thrombocythemia (ET), chronic eosinophilic leukemia, not otherwise specified (NOS), myeloproliferative neoplasm, unclassifiable.10\n\nImmunohistochemical evaluation with CD34 and mast cell tryptase were done on the archived blocks. IHC staining was performed with CD34 (RTU mouse monoclonal; clone: QBEND/10; PDM050) (BD Biosciences Cat# 550390; RRID:AB_393656) and mast cell tryptase (RTU mouse monoclonal; clone: JC/70A; PDM020) (IMGENEX Cat# IMG-80250; RRID:AB_1152624).\n\nThe tissue taken for IHC was performed on paraffin embedded, formalin fixed samples. A representative block was selected in every case. IHC staining was performed with CD34 (RTU mouse monoclonal; clone: QBEND/10; PDM050) and mast cell tryptase (RTU mouse monoclonal; clone: JC/70A; PDM020) according to the instructions provided by the manufacturer. The secondary antibody used was of Diagnostic Biosystem (Ref: UMR1000PD) against mouse primary antibodies. The staining for IHC was done according to standard procedures.\n\nPreparation of Tris (tris hydroxymethyl aminomethane) EDTA buffer: Tris buffer, 1.21 g; EDTA disodium salt 0.372 g was dissolved in 1L of distilled water. This was used for antigen retrieval.\n\nPreparation of stock solution of wash buffer/tris buffer: Tris buffer and sodium chloride (NaCl) solution were used for preparing the working solution. Tris buffer was prepared by mixing 121.1 g of buffer powder with 1000 ml of distilled water and adjusting the pH to 7.4 by adding hydrochloric acid (HCl) drop by drop. NaCl solution was prepared in a separate beaker by adding 83.33 g of NaCl powder to 1000 ml of distilled water. The working solution was prepared by mixing 30 ml of Tris buffer and 70 ml of NaCl solution in 700 ml of distilled water.\n\nThe IHC staining was carried out on 4 μm thick sections of tissue. The tissue sections were incubated on poly L-lysine coated glass slides overnight at 37°C. De-paraffinization was performed by keeping the sections at 60°C for 20 minutes followed by two changes of xylene for 15 minutes each. Slides were dried and alcohol changes for 2 times for 15 minutes each was carried out. The slides were immersed in a solution of 99% methanol and 1% hydrogen peroxide to quench the endogenous peroxidase, for 20 minutes. They were washed in running tap water for 5 minutes.\n\nAntigen retrieval: The EDTA buffer was pre-heated for 5 minutes at high power in the microwave oven. After 5 minutes, washed slides were dipped in pre-heated solution for 5 minutes, 10 minutes, and another 5 minutes and allowed to cool. They were washed under running tap water for 5–10 minutes. Slides were dipped in distilled water, then working solution/wash buffer (pH 7.4) for 10 minutes. Next, the slides were incubated for 10 minutes in buffered casein solution with sodium azide to suppress the non-specific binding (power block/peroxidase blocking solution). The slides were incubated with the primary antibody CD34 (RTU mouse monoclonal; clone: QBEND/10; PDM050) and mast cell tryptase (RTU mouse monoclonal; clone: JC/70A; PDM020) for 30 minutes at room temperature. After incubation, they were washed in the working solution for duration of 10 minutes. Next, the slides were treated with Diagnostic Biosystem rabbit/mouse secondary antibody (catalogue number K8002, monoclonal antibody) and incubation was carried out at room temperature. Slides were washed in working solution for 10 minutes and then treated with a freshly prepared solution of 20 μL of 3,3’- Diaminobenzidine (DAB) chromogen and 1000 μl of substrate buffer for 5 minutes to obtain a brown colour. Next, the slides were washed with working solution for 5 minutes. Counterstaining was performed for 2–3 minutes with Meyer’s hematoxylin and slides were rinsed in tap water thereafter. Finally, the slides were dehydrated, cleared and mounted.\n\nControls used for IHC: Positive controls used for respective markers were as follows: CD34, Kidney; Mast cell tryptase, Tonsil\n\nThe average number of thin-walled vessels with or without lumen and sinusoids that were detectable by CD34+ endothelial cells were considered to assess MVD. These were then counted in ten hot spot fields and an average was obtained per high power field (HPF). Average MVD in 10 hotspots was scored as follows; 0–7 mvd/10 hotspots – score 1, 8–15 mvd/10 hotspots – score 2, 16–25 mvd/10 hotspots – score 3.8,11\n\nCD34 is expressed in the cytoplasm of blast cells. For determination of blast percentage on CD34 stained bone marrow biopsy, five representative fields were selected and the number CD34+ cells were counted in a total of five high power fields (40x).12,13 The scoring was performed as follows; 1–5 blasts/5HPF – score 1, >5–10 blasts/5HPF – score 2, >10–19 blasts/5HPF – score 3. MCT shows cytoplasmic staining in mast cells. Average mast cell numbers were counted using tryptase in 10 oil immersion fields. The grading of mast cells using tryptase was as follows; 3–5 mast cells – 1+, >5–10 mast cells – 2+, >10 mast cells – 3+.14\n\nReticulin fibrosis was graded according to WHO guidelines, where loose network of intersecting reticulin fibres around the perivascular region was considered grade 1, diffuse and dense reticulin fibres with extensive intersections along with focal bundles of thick collagen fibres as grade 2 and diffuse and dense increase in reticulin fibres with extensive intersections and coarse bundles consistent with thick collagen fibres as grade 3 fibrosis.10\n\nFor statistical analysis, SPSS software version 25 (IBM SPSS Statistics (RRID:SCR_016479)) was used. Chi-square test, Fishers exact test, mean MVD and standard deviation were calculated. P-value was used to determine significance of the study. Statistical significance was defined as p-value ≤0.05.\n\n\nResults\n\nIn a total of 66 cases of MPN studied, 32 (48.48%) were CML, 31 (46.96%) were PMF and three (4.54%) cases were ET. The age range of patients were from 14 to 78 years with the mean age being 51.53 years. The present study found that elderly males were primarily affected and the male to female ratio of 1.9:1.\n\nOn morphological assessment of these 66 bone marrow biopsies, the majority of them (n = 59, 89.39%) were hypercellular for age. Increased myelopoiesis was observed in 54 cases (81.81%), out of which left shift was seen in 39 (59%) of them. Erythropoiesis was found to be decreased in 36 (54.54%) cases. Megakaryopoiesis was markedly increased in 62 cases (93.93%), in which 17 (26%) and 8 (12.12%) cases showed MVD scores 2 and 3 respectively. Cases with megakaryocytes in clusters, sheets and diffuse patterns were found to have increased mean MVD (8.85, 8.38 and 11.50 respectively), while the presence of dyspoietic megakaryocytes did not alter the mean MVD significantly. These observations, however, were not statistically significant (Table 1).\n\n1 MVD – Microvessel density.\n\n2 MPN – Myeloproliferative neoplasms.\n\nIn CML, MVD score 2 and 3 were found only in six (18.75%) and five cases (15.62%) respectively. Out of 31 cases of PMF, 12 (38.7%) cases expressed MVD of score 2, while score 3 was seen in three (9.67%) cases only. In ET, two cases (n = 2/3, 66.66%) expressed MVD of score 3 (p = 0.042). Highest mean MVD was observed in ET (Mean MVD -12.67) compared to CML and PMF (Mean MVD -7.34 and 8.29 respectively) as depicted in Table 2.\n\n1 MVD – Microvessel density.\n\n2 MPN – Myeloproliferative neoplasms.\n\n3 CML – Chronic myeloid leukemia.\n\n4 ET – Essential thrombocythemia.\n\n5 PMF – Primary myelofibrosis.\n\nOut of 66 cases, 9 (13.63%), 26 (39.39%) and 31 (46.96%) cases were showing grade 1, 2 and 3 fibrosis respectively. In cases with MVD score 2, 66.7% cases were found to have grade 2 fibrosis and in cases with score 3, 50% of them were showing grade 2 fibrosis (p = 0.332) (Table 3, Figure 1).\n\n1 MVD – Microvessel density.\n\n2 MPN – Myeloproliferative neoplasms.\n\n1PMF – Primary Myelofibrosis, 2H&E - Haematoxylin and Eosin stain, 3WHO - World Health Organisation, 4MVD - Microvessel Density, 5IHC – Immunohistochemistry, 6MCT – Mast cell tryptase.\n\nWhen we correlated CD34 blast count score with mean MVD, there was a positive correlation between CD34 blast count and mean MVD (Table 4). Increased mean MVD was observed with score 2 and 3 of CD34 blasts, which was statistically significant (mean MVD = 8.37 and 10.67 respectively, p = 0.036) (Figure 2).\n\n1 MVD – Microvessel density.\n\n2 MPN – Myeloproliferative neoplasms.\n\n3 HPF – High power field.\n\n1CML – Chronic myeloid leukemia, 2H&E - Haematoxylin and Eosin stain, 3WHO - World Health Organisation, 4MVD - Microvessel Density, 5IHC – Immunohistochemistry, 6MCT – Mast cell tryptase.\n\nOn analysis, highest mean MVD was seen in cases with mast cell positivity of 3+ and there was a positive correlation between mast cell score and mean MVD. However, we did not get statistical significance with MVD score and mean MVD as seen in Table 5 (p = 0.447).\n\n1 MVD – Microvessel density.\n\n2 MPN – Myeloproliferative neoplasms.\n\n3 MCT – Mast cell tryptase.\n\nBCR-ABL1 mutation was seen in 32 (48.48%) cases, out of which six (33.3%) and five (50%) cases showed MVD scores 2 and 3 respectively. JAK2 mutation was seen in a total of 32 (48.48%) cases, in which MVD scores of 2 and 3 were found in 11 and five cases respectively. Only five cases showed CALR mutation, out of which only one case showed score 2 MVD as depicted in Table 6. Mean MVD of 8.72 and 6.60 was seen in JAK2 and CALR positive cases respectively (p = 0.053 and 0.059 respectively).\n\n1 MVD – Microvessel density.\n\n2 MPN – Myeloproliferative neoplasms.\n\n3 BCR-ABL1 – Breakpoint cluster region – Abelson gene.\n\n4 JAK2 – Janus kinase 2.\n\n5 CALR – Calreticulin.\n\nIn the present study, all 66 patients were followed up for one year with regular checkups at three-month intervals. Among which, 37 (68.18%) and 17 (15.15%) subjects were rendered disease-free with MVD score of 1 and 2 respectively. Disease relapse was seen in eight cases (12.12%), with the majority (n = 6/8, 75%) expressing MVD score of 3 (p = 0.000) as depicted in Table 7. Thus, mean MVD was higher in those cases with relapse/deceased as compared to disease-free patients.\n\n1 MVD – Microvessel density.\n\n2 MPN – Myeloproliferative neoplasms.\n\n\nDiscussion\n\nThe present study analysed 66 cases of MPNs, out of which 32 (48.48%) cases of CML, 31 (46.96%) cases of PMF and three (4.54%) cases of ET were considered. Megakaryopoiesis was markedly increased in 62 cases (93.93%), in which 17 (26%) and eight (12.12%) cases showed MVD scores 2 and 3 respectively. Cases with megakaryocytes in sheets and diffuse patterns were found to have increased mean MVD (8.38 and 11.50 respectively), while mean MVD was not majorly altered with respect to dyspoietic megakaryocytes. These observations were not statistically significant. Mesa R. et al. observed that increased MVD was associated with clustering of megakaryocytes (p <0.01) in a study conducted in 114 MPN subjects.15 Ponce et al. also studied that increased MVD is associated with hypercellularity and megakaryocytic clustering in 56 patients.16\n\nIn CML, MVD score 2 and 3 were found only in six (18.75%) and five (15.62%) cases respectively. Out of 31 cases of PMF, 12 (38.7%) cases expressed MVD of score 2, while score 3 was seen in three (9.67%) cases only. In ET, two cases (n = 2/3, 66.66%) expressed MVD of score 3. The highest MVD was observed in ET (mean = 12.67) compared to CML and PMF (mean -7.34 and 8.29 respectively) (p = 0.042). Lundberg L. et al. found that MVD was significantly increased by a two-fold degree in CML when compared to normal bone marrow (p <0.01)5 Gianelli U. and colleagues studied 98 cases of Philadelphia negative MPNs and concluded that there were significantly more MVD “hot spots” in PMF (mean ± SD, 25.6 ± 6.3) cases when compared to ET cases (10.1 ± 4.5) and normal control samples (7.5 ± 3.6) (P <0.05).17 These observations were concordant with the findings in our study. Wrobel T. et al. found that PMF is the disease with the most evident angiogenesis (increased MVD) and the expression of VEGF positive MVD in bone marrow in PMF patients was significantly higher than in other myeloproliferative disorders.18\n\nOut of 66 cases, nine (13.63%), 26 (39.39%) and 31 (46.96%) cases were showing grade 1, 2 and 3 fibrosis respectively. In cases with MVD score 2, 66.7% cases were found to have grade 2 fibrosis and in cases with score 3, 50% of them were showing grade 2 fibrosis (p = 0.332). Increased MVD at the prefibrotic stage of PMF suggested angiogenesis as an early event signalling the development of fibrosis, according to Boveri et al. The study found that the grade of fibrosis increased with increasing MVD.6 Lekovic et al. found that CD34-MVD and CD105-MVD strongly (p = 0.001 and p = 0.001, respectively) linked with the grade of bone marrow fibrosis in the entire cohort of MPN patients.19 These observations were in line with the findings of the current study.\n\nWhen we correlated CD34 blast count score with mean MVD, there was positive correlation between CD34 blast count and mean MVD. Increased mean MVD was observed with score 2 and 3 of CD34 blasts, which was statistically significant (mean MVD = 8.37 and 10.67 respectively, p = 0.036). Padró et al. found that increased angiogenesis was seen in 62 patients with acute myeloid leukemia (AML).20 Kuzu et al. and Pruneri et al. also concluded that increase in marrow micro-vascularity has adverse prognosis in a study carried out on AML patients.21,22\n\nOn analysis, highest mean MVD was seen in cases with mast cell count score of 3 and there was positive correlation between mast cell score and mean MVD. However, we did not get statistical significance (p = 0.447). Xu P. et al. found that MVD was increased with increase in mast cells in CML patients, which was in concordance with the present study, but statistically significant.8 Studies on MVD with mast cell association is seldom seen in hematological malignancies.\n\nIn the current study, BCR-ABL1 mutation was seen in 32 (48.48%) cases, out of which six (33.3%) and five (50%) cases showed MVD scores 2 and 3 respectively. JAK2 mutation was seen in a total of 32 (48.48%) cases, in which MVD scores of 2 and 3 were found in 11 and five cases respectively. Only five cases showed CALR mutation, out of which only one case showed score 2 MVD. Mean MVD of 8.72 and 6.60 was seen in JAK2 and CALR positive cases respectively (p = 0.053 and 0.059 respectively). Boveri et al. and Medinger et al, in their study stated that JAK2 mutation was strongly associated with increasing numbers of MVD in all subjects with MPNs.6,13 Lekovic D. et al. also found that MVD was notably raised in patients with JAK2 mutation (CD34-MVD: p = 0.491, p <0.001) but not with CALR mutation.19\n\nIn the present study, all 66 patients were followed up for one year with regular check-up at three-month intervals. Among which, 37 (68.18%) and 17 (15.15%) subjects were rendered disease-free with MVD score of 1 and 2 respectively. Disease relapse was seen in eight cases (12.12%), with majority (n = 6/8, 75%) expressing MVD score of 3 (p = 0.000). Thus, mean MVD score was higher in those cases with relapse/deceased as compared to disease-free patients (p = 0.000). Koopmans S. et al. in a study conducted on 106 MPN patients found that increased MVD and development of marrow fibrosis correlated with worse prognosis.23 Ponzoni M. et al. studied MVD using CD105 and CD34 in 55 MPN patients and observed that MVD was associated with poor prognosis, especially in PMF.24 Pizzi M. et al. and Lundberg L. et al. also stated the above in their study.5,25 The above-mentioned study results were concordant with our study.\n\nThe present study evaluates bone marrow angiogenesis using MVD score and mean MVD. The p-value obtained using both data have been analysed to draw conclusions between various parameters and we observed that raised MVD score was significantly associated with PMF, while raised mean MVD was found in association with increased CD34 blast counts. Both, MVD score and mean MVD was significant statistically when correlated with disease outcome. Angiogenesis is crucial in the growth and progression of hematopoietic malignancies.6,13,18 Anti-angiogenic therapy is primarily an anti-vascular endothelial growth factor (VEGF) or anti-VEGF-receptor (VEGFR) therapy. Several anti-angiogenic drugs, either by themselves or in combination with other anti-tumor therapies, have been licensed for the treatment of these disorders.26\n\n\nConclusions\n\nIn the present study, raised MVD is mostly associated with PMF subtype than in other MPNs. In MPNs, cases with increased blast count, mast cell count and fibrosis are also associated with increased MVD. We also found that increased angiogenesis was observed in cases with relapse/deceased, implying the prognostic significance of studying MVD in MPN. With angiogenesis playing a critical role in disease outcome, we now have drugs (bevacizumab) to regulate angiogenesis that are supported by contemporary research.\n\nOut of 66 MPN cases, only three cases of ET and no case of polycythemia vera were evaluated. Hence, further studies with larger cohorts comprising a greater number of each subtype of MPN would be recommended to establish the prognostic/theranostic role of MVD in MPNs.", "appendix": "Data availability\n\nOpen Science Framework: Underlying data for ‘Study of significance of bone marrow microvessel density in myeloproliferative neoplasms in correlation with CD34 blasts, mast cell count and fibrosis. https://doi.org/10.17605/OSF.IO/E739P. 9\n\nThis project contains the following underlying data:\n\n• Data.xlsx (Anonymized responses in Excel sheet)\n\n• Unedited micrographs: Figures 1a-f and 2a-f.jpg\n\nOpen Science Framework: Extended data for ‘Study of significance of bone marrow microvessel density in myeloproliferative neoplasms in correlation with CD34 blasts, mast cell count and fibrosis’. https://doi.org/10.17605/OSF.IO/E739P. 9\n\nThis project contains the following extended data:\n\n• Coding.xlsx (Codes for responses)\n\n• Proforma.pdf\n\nOpen Science Framework: STROBE checklist for ‘Study of significance of bone marrow microvessel density in myeloproliferative neoplasms in correlation with CD34 blasts, mast cell count and fibrosis.’ https://doi.org/10.17605/OSF.IO/E739P. 9\n\nData are available under the terms of the Creative Commons Attribution 4.0 International (CC-BY 4.0)\n\n\nAcknowledgements\n\nI would like to thank our technical staff and co-postgraduates for their research assistance.\n\n\nReferences\n\nSkoda RC, Duek A, Grisouard J: Pathogenesis of myeloproliferative neoplasms. Exp. Hematol. 2015; 43(8): 599–608. Publisher Full Text\n\nPatterson-Fortin J, Moliterno AR: Molecular pathogenesis of myeloproliferative neoplasms: Influence of age and gender. Curr. Hematol. Mal. Rep. 2017 Oct; 12(5): 424–431. PubMed Abstract \| Publisher Full Text\n\nTitmarsh GJ, Duncombe AS, Mcmullin MF, et al.: How common are myeloproliferative neoplasms? A systematic review and meta-analysis. Am. J. Hematol. 2014; 89(6): 581–587. PubMed Abstract \| Publisher Full Text\n\nVarma S, Naseem S, Malhotra P, et al.: Incidence Rates of Myeloproliferative Neoplasms in India-A Hospital Based Study. Int. J. Epidemiol. 2015 Oct 1; 44(suppl_1): i198–i199. Publisher Full Text\n\nLundberg LG, Lerner R, Sundelin P, et al.: Bone marrow in polycythemia vera, chronic myelocytic leukemia, and myelofibrosis has an increased vascularity. Am. J. Pathol. 2000 Jul 1; 157(1): 15–19. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBoveri E, Passamonti F, Rumi E, et al.: Bone marrow microvessel density in chronic myeloproliferative disorders: A study of 115 patients with clinicopathological and molecular correlations. Br. J. Haematol. 2008 Jan; 140(2): 162–168. PubMed Abstract \| Publisher Full Text\n\nKeski H: Association of mast cells and bone marrow reticulin fibrosis in patients with bcr-abl negative chronic myeloproliferative neoplasms. Blood Cells Mol. Dis. 2020 May 1; 82: 102420. PubMed Abstract \| Publisher Full Text\n\nXu P, Zhang C, Wang Y, et al.: Increased number of mast cells in the bone marrow of chronic myeloid leukemia may herald the pending myeloid transformation-the mast cell is an indicator of myeloid transformation. Transl. Cancer Res. 2019 Sep 1; 8: 2121–2129. 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Hematol. 2000 Apr 15; 95(8): 2637–2644. Publisher Full Text\n\nKuzu I, Beksac M, Arat M, et al.: Bone marrow microvessel density (MVD) in adult acute myeloid leukemia (AML): Therapy induced changes and effects on survival. Leuk Lymphoma. 2004 Jun; 45(6): 1185–1190. PubMed Abstract \| Publisher Full Text\n\nPruneri G, Bertolini F, Soligo D, et al.: Angiogenesis in myelodysplastic syndromes. Br. J. Cancer. 1999 Dec; 81(8): 1398–1401. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKoopmans SM, Bot FJ, Schouten HC, et al.: The involvement of Galectins in the modulation of the JAK/STAT pathway in myeloproliferative neoplasia. Am. J. Blood Res. 2012; 2(2): 119–127. PubMed Abstract\n\nPonzoni M, Savage DG, Ferreri AJM, et al.: Chronic idiopathic myelofibrosis: Independent prognostic importance of bone marrow microvascular density evaluated by CD105 (endoglin) immunostaining. Mod Pathol. 2004 Dec; 17(12): 1513–1520. 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PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "197587", "date": "29 Aug 2023", "name": "Anna Rita Migliaccio", "expertise": [ "Reviewer Expertise Experimental Hematology including Myeloproliferative neoplasms" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nNeoangiogenesis is one of the features of myelofibrosis, the most severe of the Phyladelphia negative myeloproliferative neoplasms and is the subject of extensive investigation. A pubmed search for “angiogenesis” and “myelofibrosis” provided 43 hits. Methods to quantify angiogenesis in chronic myeloproliferative disorders were published by Thiele (Histil Histopathol 2004, 1245-1260. Angiogenesis was correlated with the expression of vascular endothelial growth factor (likely produced by megakaryocytes) and of its receptor by Medinger et al Br J Haemat 2009, 146:150-157, Boiocchi et al (J Clin Pathol 2011:64, 226 and others. A robust assessment on neoangiogenesis in chronic myeloproliferative disorders (115 patients) was provided by Boveri et al (Br J Haematol 2008, 140: 162 and it is one of the diagnostic criteria for patients with Ph-negative chronic myeloproliferative diseases (Michielis et al Acta Haematol 2015:133:36). Of these papers only that from Boveri is cited in the manuscript.\nGiven this extensive background information, the study presented in the paper currently under review correlates the extent of microvessel density with the frequency of CD34+ blasts, mast cells and fibrosis in a cohort of 66 patients with myeloproliferative neoplasms (32 patients with chronic myeloid leukemia, 31 primary myelofibrosis and three essential thrombocythemia) from diagnosis over a period of six years (almost five years retrospectively and 10 months prospectively). The end points were assessed by immune-histomorphology of bone marrow sections with antibodies against CD34 (blasts and endothelial cells) and mast cell tryptase. Based on the data presented the authors conclude that microvessel density is present in patients with Ph-negative myelofibrosis patients and not in patients with BCR-ABL chronic myeloid leukemia and that it frequency correlated with adverse prognosis in myelofibrosis patients.\nImages of the bone marrow two representative patients are presented in Figure 1 (primary myeloprolibrosis) and Figure 2 (chronic myeloproliferative neoplasm). Correlation studies with the data from more patients per group is presented in a table format.\nThe Figures are the main problem of the paper: the claimed presence of hypolobulated micromegakaryocytes in Figure 1B and of dwarf megakaryocytes in Figure 2B is not evident. Also not evident are the presence of CD34+ microvessels in 1D and 2D and of CD34+ blasts in 1E and 2E. The quality of the immunohistochemistry is overall disappointing. In addition, normal bone marrow processed in parallel as baseline control is missing.\nAnother concern is that it is not clear whether the number of deceased patients is sufficiently high for powerful assessments of adverse factors.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? No\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10389", "date": "16 Nov 2023", "name": "Ranjitha Rao", "role": "Author Response", "response": "The figures are edited as suggested by the reviewer. The hypolobulated micromegakaryocytes in figure 1B, dwarf megakaryocytes in figure 2B are indicated by black arrow.CD34 staining micro vessels and CD34 staining blasts are indicated by black arrow in figure 1D,2D and 1E,2E respectively. The statistical tests have been performed by a qualified statistician. The results try to compare the mean vessel density between those who are deceased of myeloproliferative disease and those who have survived the disease at the time of study. Yes, the number of deceased cases are low and studies with large number of cases are required to validate this finding." } ] }, { "id": "197588", "date": "04 Sep 2023", "name": "Danijela Lekovic", "expertise": [ "Reviewer Expertise Myeloproliferative neoplasms" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe limitations of the study should be noted, that there are no patients with PV and that there is an uneven distribution of patients in the analyzed groups. I think that statistical analysis cannot be done on three patients with ET. I also did not see comparisons with healthy controls. It is also necessary to have the work checked by a statistician because I am not competent to say that it is possible to correlate how it was done.\nAfter supplementing the work with suggestions, it can be accepted\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nI cannot comment. A qualified statistician is required.\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10388", "date": "16 Nov 2023", "name": "Ranjitha Rao", "role": "Author Response", "response": "All the statistical tests in this study were conducted by a qualified statistician. The number of essential thrombocythemia cases were very few and no polycythemia vera cases were identified during the study and this limitation has been stated after conclusion in the article." } ] } ]	1	https://f1000research.com/articles/12-503
https://f1000research.com/articles/12-1423/v1	31 Oct 23	{ "type": "Brief Report", "title": "Enhancement of private schools for collaboration to address gender equity gap of STEM education: Case study in Mbeya city of Tanzania", "authors": [ "Isack Ephraim Kibona" ], "abstract": "Investment in education to guarantee gender equity in most developing countries has focused on modern teaching pedagogy, teaching and learning facilities, cultural issues and the likes. However, performance in private schools do set learnable examples of which if not ignored, a nation in question may take lessons and improve gender equity in science, technology, engineering and mathematics (STEM) education. Private secondary schools may serve as on way to enrich girls' enrolment in STEM for future career opportunities by scoring qualifiable grades in STEM subjects, provided that education investment environment not only favors public schools but also be in favor of private schools. In the study conducted in Mbeya city of Tanzania, the author investigated 58 secondary schools of which 32 were private schools. From these schools 7936 candidates sat for Certificate of Secondary Education Examination (CSEE) of 2022 in Mbeya city. 2232 candidates were from private schools. The analysis indicated that more girls with a potential to enter into STEM education came from private schools, and it is about twice the number of girls from public schools regardless of more girls candidates from public schools. Chi-square testing of pass in STEM subjects between boys and girls verified that performance in private schools had no gender equity gap between boys and girls. On the other hand, boys outperformed girls in public schools. Moreover, an estimation of 2202 candidates of which 999 were girls had at least a minimum pass in basic mathematics. Private schools contributed 626 (63%) of passes in basic mathematics. Thus, appropriate collaboration with private schools might revamp not only girls’ enrolment but also clear gender equity gap in performance of STEM subjects if policy makers in education investment can do more improvement in the environment for education investment through private schools.", "keywords": [ "STEM education", "gender equity", "Mbeya city of Tanzania", "Private schools" ], "content": "Introduction\n\nEstablishment of Sustainable Development Goals (SDGs) by the United Nations (UN) in September 2015 placed the world to ensure gender equity in science, technology, engineering and mathematics (STEM) education (Koehler, 2016; Leal Filho et al., 2022; Zorzano, 2020). Gender is commonly representing female or male, however, in this study, added to that describes the socio-cultural characters of masculinity and femininity according to practices by individual based on their culture, while sex describes the biological characteristics of women and men (Unicef & others, 2020). In this regard, gender may change with time and place depending on roles taken by women and men, whereas sex never change. Before 2016, sub-Saharan Africa had a substantial number of secondary schools whose quality of education was questionable according to United Nations, this being one of the reasons to launch SDG4 (Unterhalter, 2019).\n\nSome regions of the world are closing gender equity gaps in STEM education like USA, and Europe (Kamberidou & Pascall, 2019), but gender inequity in STEM education is substantial in sub-Saharan Africa, the Arab states, and south and west Asia (Ismail, 2018; Loyalka et al., 2021). Emphasis to enhance STEM education in literature has centered resolution on pedagogical skills, low teacher student ratio, incompetent teachers, and education facility availability (Allen et al., 2016; Huang et al., 2022; Teo & Ke, 2014). Moreover, girls' poor participation in STEM subjects in secondary schools especially in sub-Saharan Africa are more associated with cultural practices and other reasons associated with masculinity (Adams & Baddianaah, 2023; Lewin, 2009).\n\nEducation policy makers in Tanzania engaged private sectors to run private schools along with public schools (Komba, 2017). This lead to contribution in containing not only enrolment issues in STEM education but also great improvement of gender equity (Weaver, 2011). Evaluation of private schools’ contribution is indispensable, this fact holds following the reality that graduates from private education sectors become part of the nation decent and future human resource capital (Achoui, 2009; Budhwar & Sparrow, 2002; Sebola, 2023). Table 1 presents the performance of students from 32 private schools in Mbeya city. Majority of students in privates’ schools passed in first and second division. Unlike in private schools, candidates in public schools' majority were in the fourth division Table 2.\n\nIn Tanzania, efforts to solve gender issues in STEM education is historical (Samoff, 1987). Several bodies like United States Agency for International Development (USAID) have supported to ease the tension of gender inequity in STEM education (Stromquist, 2006; Swainson, 2000).\n\nThis report unleashes aspects of private schools’ contributions in gender equity in STEM education following credible educational policies by the Tanzanian government and promotes improvement in education investment for harmonization of public and private schools. This study illustrated by concrete examples the contribution of private schools in STEM education. Contribution is not only an increase in number of students specializing in STEM subjects but also significant improvement in diminishing of gender equity gap of STEM education.\n\nFigure 1 depicts performance in STEM subjects of biology, chemistry, physics and basic mathematics of 2232 candidates from 32 private secondary schools in Mbeya city. Comparable performance of 5704 candidates from 26 public schools Figure 1 indicated deprived performance in STEM subjects. In parallel to more girls’ enrolment over boys, more girls passed every STEM subject in private schools except for physics. However, regardless of more girl’s enrolment in public schools, boys outperformed girls in every STEM subject except for biology.\n\nThe study is for enhancing education investment policies along with engaging private school investments for collaboration to address challenges of gender equity in STEM education.\n\nSpecific objectives\n\ni. Stimulate harmony and more collaboration between education policy makers and private schools investors to enrich gender equity in STEM education.\n\nii. Compare private schools and public schools’ performance of girls and boys in STEM subjects.\n\n\nMethods\n\nMethodologies in this study undertook over form four secondary school examination results for Mbeya city, Tanzania. In this regard, this study dealt with secondary data, already processed for student placement to the next education level, in this situation, analysis of sex or gender was not applicable from the raw data. That is study design, data collection and data analysis carried out free of considerations due to sex or gender. Thus, this study is about what is observed in the examination results and not how teachers reach these results. On the other hand, this study calls for more insights as to how these results are reached, which is not the scope of this study. Therefore, the scope of study is to examine whether gender has influence in the examination performance.\n\nThe author collected secondary schools’ performance data from National Examination Council of Tanzania. The data involved 58 secondary schools of which 32 were private and 26 public schools. The data tabulated in separation of private and public schools as in Table 3 and Table 4 respectively, indicating number of girls and boys in every STEM subject. The researcher processed the data, analyzed by employing chi-square test for interpretation, and eventually reported the finding.\n\nThe Author conducted data collection by reading and recording from the report by NECTA of CSEE for the year 2022, particularly, selecting all Mbeya city secondary schools participated in the form four national examinations of year 2022. In addition to division-wise performance as organized by NECTA, the author went further by listing overall performance of every STEM subject and eventually bar charts plotted as in Figure 1 and Figure 2.\n\nThe author used the chi-square approach to determine whether there was a significant difference in the passing of STEM subjects between boys and girls. The focus of the STEM subjects was biology, chemistry, physics and basic mathematics. The degree of freedom evaluated leads to a critical value (p) from statistical tables, a test statistic computed for each group of students separately, and the decision made based 95% confidence interval. Records of number of passes in STEM subject are as listed in Table 3 and Table 4. No software were involved and all computations conducted manually, aided by Casio scientific calculator.\n\nCalculation of Chi – square, χ2:\n\nNumbers in parentheses of Table 3 are theoretical expectations of gender equity. The author went through five steps to justify whether gender has an effect in the performance of STEM subjects.\n\nStep 1: Define Null (H0) and Alternative Hypotheses (H1):\n\nH0: For the students in Mbeya city from private secondary schools, gender has no effect to pass in STEM subjects.\n\nH1: For the students in Mbeya city private secondary schools, gender has effect to pass in STEM subjects.\n\nStep 2: State the confidence interval: α=0.05\n\nStep 3: Calculate degree of freedom (df) and state the critical value (p):\n\ndf=rows−1columns−1=2−14−1=3, so critical value, p = 7.81473.\n\nThat is if chi-square (χ2) is greater than 7.81473, rejectH0.\n\nStep 4: Calculation of test statistic χ2=∑fo−fe2fe, where fe=fcfrn and f0=observed frequence, fe=expected frequence, fc=frequence of the column,fr=frequence of therow, n=total number of subjects.\n\nStep 5: Calculation of theoretical pass expectations: Girls expected to pass biology in first cell of Table 3,\n\nχpr2=4.28<7.81473=p. Therefore, the null hypothesis is true.\n\nThe author applied a similar testing to candidates of public schools.\n\nWith similar calculations as Table 3 of private schools, χp2=∑fo−fe2fe=318.33425 for Table 4. However,χp2=318.334>7.81473=p. Therefore, we reject the null hypothesis.\n\nThe author went further to ascertain every student with a pass of at least D grade (minimum pass) in both physics and basic mathematics and classified this group as minimum pass in STEM. Moreover, students with at least two C grades and one D pass grade in any of three subjects: physics, chemistry and biology (PCB) or same passes in any of three: physics, chemistry and mathematics (PCM) classified as potential PCM or PCB candidates. A collection of students with minimum passes in STEM and/or potential PCM or PCB candidates grouped as potential STEM candidates.\n\n\nResults\n\nThe degree of freedom for each group is three with α =0.05 of which the collected critical value, p =7.81473. Computed chi-squares were χpr2=4.28 for private schools χp2=318.334 for public schools. In addition to chi-square testing of the data, student performance in STEM subjects were listed in tabular (Table 3, Table 4) form along with plotting the bar charts (Figure 1, Figure 2).\n\nPerformance in STEM subjects determines number of candidate placements in high school PCB or PCM combinations and prospects of STEM career candidates in higher education institutions. Out of 2232 candidates, private schools contributed 783 (35%) candidates of which 380 were girls able to further studies in STEM education (Table 5). On the other hand, out of 5704 candidates, public schools contributed 699 (12%) candidates of which 206 were girls with potential to advance in STEM careers in higher education (Table 6).\n\nPCM: physics, chemistry and mathematics; PCB: physics, chemistry and biology; STEM: science, technology, engineering and mathematics.\n\nPCM: physics, chemistry and mathematics; PCB: physics, chemistry and biology; STEM: science, technology, engineering and mathematics.\n\n\nDiscussion\n\nBased on the data analysis, we calculated critical value, p = 7.81473 and a test statistic computed χpr2=4.209 for all students’ performance in STEM subjects for private schools and χp2=318.33425 for public schools. It is clear that χpr2=4.28<7.81473=p, and χp2=318.33425>p.Recall the null and alternative hypothesis:\n\nNull (H0) and Alternative Hypotheses (H1):\n\nH0: For the students in Mbeya city from private/public secondary schools, gender has no influence to pass in STEM subjects.\n\nH1: For the students in Mbeya city from private/public secondary schools, gender has influence to pass in STEM subjects.\n\nIn this regard, we do accept the null hypothesis for private schools and reject the null hypothesis for public schools. Therefore, private schools in Mbeya city resolved the gender equity gap in performance of STEM subjects. On the other hand, gender equity gaps are unresolved between girls and boys in STEM subjects’ performance for public schools. That implies that the gender equity gap shall extend to the future STEM careers of graduates from public schools, and therefore, spread to the whole nation and worldwide at large.\n\nWe suppose education policy had had recognized private sectors contribution and further promote teamwork along with private schools then more output that is stunning would surface. Thus, in so speaking, this study saves to encourage education policy makers to enrich policy mechanism for collaboration and possible embolden of private schools for more enrolment in STEM education. This is in parallel with the implementation of SDG4 realization in 2030. It is also clear that passing in both physics and basic mathematics determines the potential of STEM career prospects from secondary schools (Table 5, Table 6).\n\n\nConclusion and recommendations\n\nPrivate schools’ reflection of balancing gender in STEM education in the case of Mbeya city offers a great step toward achieving SDG4 of the United Nations. The author recommends policy makers to engage private schools to address the challenges of gender equity in STEM education in a collaborative manner rather than existing in with public schools. STEM education stakeholders need to stress not only STEM careers position in the 21st century of job placements but also the importance of passing both basic mathematics and physics as determinants of STEM career for secondary school students. Basic mathematics alone is not enough.\n\nThis study is calling for further investigation as to why in the same nation with likely the same environment, private schools outperformed public schools to such great extent. The researcher will explore both parties separately to acquire reliable information to share experiences to avert the situation in public schools.", "appendix": "Data availability\n\nData used in this study are available and accessible for reproducibility. Information about public and private schools involved are all associated to Mbeya city CSEE results of 2022. Specifically, the Author used the data published by the National Examination Council of Tanzania (NECTA) from all test centers of the year 2022 (NECTA of CSEE for the year 2022).\n\n\nAcknowledgements\n\nFirst, I do appreciate Mbeya University of Science and Technology leadership for their strength and determination. Secondly, I do acknowledge the support in ideas by all colleagues in the Department of mathematics and statistics, to mention a few are; Mr. Justin Kisakali, Mr. Paulo Ngayekamwe and Ms. Tatu S. Irunde, who as well offered their time to work on behalf for some of the office routine activities. That gave a room for me to concentrate on writing. The whole members of the Department were willing to teach more classes, this boosted ending in small teaching load and therefore ability to write. Finally, my family support is indispensable in this task. They have been patient to all my late coming at home to ensure I am okay with this task. I must point out my youngest son, David.\n\n\nReferences\n\nAchoui MM: Human resource development in Gulf countries: An analysis of the trends and challenges facing Saudi Arabia. Hum. Resour. Dev. Int. 2009; 12(1): 35–46. Publisher Full Text\n\nAdams A-M, Baddianaah I: Factors affecting female enrolment in technical and vocational education and training institutions in sub-Saharan Africa: Insights from north-western Ghana. International Journal of Training Research. 2023; 1–24. Publisher Full Text\n\nAllen M, Webb AW, Matthews CE: Adaptive teaching in STEM: Characteristics for effectiveness. Theory Pract. 2016; 55(3): 217–224. Publisher Full Text\n\nBudhwar PS, Sparrow PR: An integrative framework for understanding cross-national human resource management practices. Hum. Resour. Manag. Rev. 2002; 12(3): 377–403. Publisher Full Text\n\nHuang X, Erduran S, Zhang P, et al.: Enhancing teachers’ STEM understanding through observation, discussion and reflection. J. Educ. Teach. 2022; 48(5): 576–591. Publisher Full Text\n\nIsmail Z: Benefits of STEM education.2018.\n\nKamberidou I, Pascall N: The digital skills crisis: Engendering technology–empowering women in cyberspace. European Journal of Social Sciences Studies. 2019.\n\nKoehler G: Tapping the Sustainable Development Goals for progressive gender equity and equality policy? Gend. Dev. 2016; 24(1): 53–68. Publisher Full Text\n\nKomba AA: Educational accountability relationships and students’ learning outcomes in Tanzania’s public schools. SAGE Open. 2017; 7(3): 215824401772579. Publisher Full Text\n\nLeal Filho W, Kovaleva M, Tsani S, et al.: Promoting gender equality across the sustainable development goals. Environ. Dev. Sustain. 2022; 1–22.\n\nLewin KM: Access to education in sub-Saharan Africa: Patterns, problems and possibilities. Comp. Educ. 2009; 45(2): 151–174. Publisher Full Text\n\nLoyalka P, Liu OL, Li G, et al.: Skill levels and gains in university STEM education in China, India, Russia and the United States. Nat. Hum. Behav. 2021; 5(7): 892–904. PubMed Abstract \| Publisher Full Text\n\nSamoff J: School expansion in Tanzania: Private initiatives and public policy. Comp. Educ. Rev. 1987; 31(3): 333–360. Publisher Full Text\n\nSebola MP: South Africa’s public higher education institutions, university research outputs, and contribution to national human capital. Hum. Resour. Dev. Int. 2023; 26(2): 217–231. Publisher Full Text\n\nStromquist NP: Gender, education and the possibility of transformative knowledge. Compare: A Journal of Comparative and International Education. 2006; 36(2): 145–161. Publisher Full Text\n\nSwainson N: Knowledge and power: The design and implementation of gender policies in education in Malawi, Tanzania and Zimbabwe. Int. J. Educ. Dev. 2000; 20(1): 49–64. Publisher Full Text\n\nTeo TW, Ke KJ: Challenges in STEM teaching: Implication for preservice and in service teacher education program. Theory Pract. 2014; 53(1): 18–24. Publisher Full Text\n\nUnicef & others: A rigorous review of global research evidence on policy and practice on school-related gender-based violence.2020.\n\nUnterhalter E: The many meanings of quality education: Politics of targets and indicators in SDG 4. Global Pol. 2019; 10: 39–51. Publisher Full Text\n\nWeaver NE: Educational policy in Tanzania from independence to the present: Continuity and transformation. University of Pittsburgh; 2011. [PhD Thesis].\n\nZorzano M-P: Gender balance in Mars exploration: Lessons learned from the Mars Science Laboratory. Sustainability. 2020; 12(24): 10658. Publisher Full Text" }	[ { "id": "242769", "date": "22 Feb 2024", "name": "María Goretti Alonso de Castro", "expertise": [ "Reviewer Expertise Education", "Technology", "ICT", "Educational European projects", "Educational Inspection" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nWhile the study's objectives are clearly delineated and the methodologies are thoroughly explained, there remains a need for a clearer rationale behind the chosen analysis method. Furthermore, the study lacks an explicit discussion of its limitations and fails to address the variables that could potentially impact the observed differences between public and private educational centers. It is essential to delve into the analysis of contextual variables such as methodologies utilized, available resources, family support systems, socioeconomic status, and cultural influences to better comprehend the disparities in outcomes between these educational settings. A more comprehensive examination of the study's limitations and the myriad factors influencing the differences outlined in the article is warranted. This would significantly enhance the depth and rigor of the research, providing a more nuanced understanding of the observed discrepancies. Regarding replicability, the text references the state test database; however, it lacks clarity on the selection process, sample criteria, and the variables considered in the presented tables. A more detailed explanation of the methodology behind selecting the state test database, the criteria for sample inclusion, and the specific variables incorporated into the tables would significantly enhance the transparency and replicability of the study.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate? Partly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "11162", "date": "13 Apr 2024", "name": "Isack Ephraim Kibona", "role": "Author Response", "response": "The Author has responded by including a section informing limitations of the study, see an attachment. Moreover, the data involved (CSEE 2022) in the study enabled the study to reveal unfair prevailing situation in secondary schools but on the other hand situation about contextual variables such as methodologies utilized, available resources, family support systems, socioeconomic status, and cultural influences shall need more data pertaining to tools such as questionnaire and interview. On selection criteria, all schools in the city in the given year of study went into participation in this study, therefore, the study involved city school population. Thus, for reproducibility of the study, one shall need to study all secondary schools in the city that participated in the CSEE 2022." } ] } ]	1	https://f1000research.com/articles/12-1423
https://f1000research.com/articles/12-364/v1	04 Apr 23	{ "type": "Research Article", "title": "Comparative evaluation of pH and Ca+ ion release from MTA on interaction with platelet-rich fibrin and blood clot: an in vitro study", "authors": [ "Sonia Khatri", "Sylvia Mathew", "Shruthi Nagaraja", "Swaroop Hegde", "Soumyadeep Ghosh", "Kavimalar Ravichandran", "Sonia Khatri", "Sylvia Mathew", "Shruthi Nagaraja", "Swaroop Hegde", "Kavimalar Ravichandran" ], "abstract": "Background: ‘Regenerative endodontics’ using host-derived scaffolds and biomaterials (MTA) is popular in the management of teeth with open apex. Alkaline pH and bioactivity contribute to tissue healing and remineralization. We assessed the influence of PRF and Blood Clot on the pH and Ca+ ion release from MTA. Methods: A total of 15 single-rooted human extracted teeth were sectioned at the level of the cementoenamel junction. Based on the type of scaffolds used, samples were divided into three groups. Group 1 (MTA+ PRF), Group 2 (MTA + Blood Clot), Group 3 (control MTA). The prepared specimens were transferred to a fresh falcon tube containing 10mL of distilled water and the collected solutions were analysed for pH and Ca+ ion release at 3h, seventh day and 14th day. Results: It was observed that the mean pH and Ca+ ion release were significantly lower in the experimental groups as compared to the control group. Though there was an increase in the pH recorded in Group 1 and 2 at all time periods, the difference was not significant. Ca+ ion release peaked at Day 7 (Group3 > Group2 > Group1) and reduced significantly on the 14th day for all groups. Conclusions: Within the limitations of the study, it can be concluded that PRF and blood clot influence the pH and Ca+ ion release from MTA.", "keywords": [ "Regenerative Endodontics", "Mineral Trioxide Aggregate", "Platelet Rich Fibrin" ], "content": "Introduction\n\n‘Regenerative endodontics’ using biomaterials, stem cells, biomimetic scaffold and bioactive growth factors have contributed immensely to the clinical management of teeth with pulp necrosis and underdeveloped roots. Success in the form of regression of apical lesion, continued root maturation in length and thickness may return the tooth to vitality.1 Pulp revascularization techniques have gathered much consideration due to their feasibility, cost-effectiveness and reasonable success rate.\n\nCurrent regenerative endodontic procedures utilize various host-derived scaffolds such as intracanal blood clots (BC) or platelet substitutes like platelet-rich fibrin (PRF),2 platelet-rich plasma (PRP), which supply necessary signalling molecules with growth factors for tissue regeneration. The alkaline irrigants (sodium hypochlorite), medicaments (triple antibiotic paste and calcium hydroxide) and biomaterials like mineral trioxide aggregate (MTA) or biodentine with different biological properties can also impact the outcome.\n\nOn hydration calcium silicates present in MTA produce calcium silicate hydrate and calcium hydroxide. Some of the hydration products such as calcium hydroxide dissociate into Ca+ and OH ions, increasing the pH, contributing to antibacterial activity, osteogenic differentiation and bone formation.3 It has been observed that acidic environments increase the solubility of materials, inhibiting the setting reaction and the sealing ability.4\n\nCalcium ions released from the MTA have been shown to pass through the cell membrane by depolarization or activation of membrane-bound calcium channels stimulating the expression of bone-associated proteins which may contribute to the repair process.5 It can further activate ATP leading to osteoblast and cementoblast differentiation and hard tissue mineralization.6\n\nThere is limited literature on the influence of scaffolds like PRF and BC on the pH and Ca+ ion release of MTA. Interaction of PRF and BC with MTA may positively or negatively affect the pH and Ca+ ion release. This knowledge may contribute to the understanding of the regenerative process and help modify regenerative treatment strategies. Therefore, this study evaluated the influence of PRF and BC on the pH and Ca+ ion release from MTA.\n\n\nMethods\n\nThe study protocol was approved by the Human Research Ethics Committee of M S Ramaiah University of Applied Sciences, Bangalore, Karnataka. Whole fresh blood was collected from a healthy volunteer after obtaining written informed consent.\n\nFifteen previously extracted, single-rooted human teeth were selected. The teeth were sectioned at the level of the cementoenamel junction (CEJ) using a slow-speed diamond disc.7 Only roots with a single canal were selected. The apical end of the sectioned root was further trimmed to obtain a uniform root length of 10mm for all the specimens. The root canal was then prepared using Peeso reamers Number 1–6 (Kerr, Kerr Corporation, Orange, CA) followed by the use of long straight diamond point to achieve a final canal diameter of 4mm. After mechanical instrumentation, the canals were irrigated with 1.5 % sodium hypochlorite (20mL, 5 min) 5mL sterile physiological saline followed by a final rinse with 20 mL of 17% EDTA and dried using sterile paper points.\n\nA 5 mL blood volume was drawn by venipuncture of the antecubital vein from a healthy volunteer and transferred to a 5mL sterile PRF tube (BD vacutainer serum tubes 5mL) without anticoagulant and centrifuged immediately at 3000 revolutions/min (rpm) for 10 min. The resultant PRF clot between the acellular platelet poor plasma and red blood cells was separated out, squeezed on a piece of sterile gauze to obtain PRF membrane.8\n\nCoronal end of the root was sealed with wax. MTA was mixed (one scoop powder was mixed with 0.33mL of liquid) and condensed into the prepared canal to obtain a thickness of 4mm and confirmed by a graduated probe. Fifteen samples thus prepared were divided into three groups.\n\nPRF membrane cut into fragments was placed into the root canal and gently compacted over the MTA using a hand plugger to achieve a thickness of 5mm.\n\nBlood drawn by venipuncture of the antecubital vein of a healthy volunteer was introduced into the root canal and left to form a clot.9\n\nMTA was mixed according to the manufacturer’s instructions (one scoop of powder was mixed with 0.33 mL of liquid). Cement was then placed into the prepared canal to obtain a thickness of 4 mm and confirmed by a graduated probe.\n\nThe prepared specimens were transferred to sterile falcon tubes containing 10 ml of distilled water7 and stored at 37oC and 100% relative air humidity throughout the testing period. At the end of every experimental period (three hours, seven days and 14 days), the specimens were transferred to a fresh falcon tube containing 10mL of distilled water and the collected solutions were analysed for pH and Ca+ ion release.\n\nThe digital pH meter (HI5221 Hanna Instruments Cal Check, United States) was calibrated to pH 7 with standard buffer solution before use. The refillable calomel electrode was placed into the falcon tube containing 10mL of solution to record the pH. The electrode was washed with distilled water and wiped dry between readings.\n\nTo determine the release of Ca+ ions, ICP-OES Inductively Coupled Plasma- Optical Emission Spectrometry (Thermo Fisher ICAP 7400 ICP-OES, Radial N. American, USA). equipment was used, owing to its high sensitivity and stability for inorganic analysis. It provides rapid and multi-element analysis of the solutions.\n\nThe pH and Ca+ ion release were presented as mean values. One-way ANOVA followed by post hoc Bonferroni test were done to determine the significant differences between groups using IBM SPSS software version 22.0 (IBM Corp., Armonk, NY, USA). The results were considered statistically significant if the p-value was less than 0.05.\n\n\nResults\n\nThe mean pH values of the individual groups at different time periods are presented in Table 1. The control group (MTA) exhibited a mean value of 10.76 at 3h, 11.54 on the seventh day, and 11.16 on the 14th day, which was significantly higher than the other groups (p=0.000). pH readings for Group 1 (MTA +PRF) were 7.56 at 3h, 7.78 at seven days which increased to 9.10 on the 14th day. Group 2 (BC) presented pH of 6.68 at 3h which increased to 8.64 on the seventh day and 9.28 on the 14th day. There was a significant difference in pH between Groups 1 and 2 at 3h and seven days (p < 0.001) but on the 14th day there was no significant difference between the groups (p > 0.973).\n\nThe mean Ca+ ion release of the individual groups at different time periods is given in Table 2. The mean values of Ca+ ions at 3hin the control group (Group 3) was 1.70, MTA +PRF (Group 1) was 0.77 and MTA+ BC (Group2) was 0.72. Group 3 exhibited significantly higher Ca+ ion release as compared to Group 1 and Group 2 at 3h. On the seventh day, MTA+PRF (Group 1) had a Ca+ ion value of 18.39, MTA+ BC (Group 2) recorded a value of 39.20. The control group (Group 3) recorded a maximum value of 203.08, which was significantly higher than both experimental groups. On the 14th day Ca+ion release from all groups reduced significantly. The recorded Ca+ ion release of MTA+PRF (Group 1) was 2.01, MTA+ BC (Group 2) was 2.75 and control group (Group 3) had a value of 4.73. The Ca+ ion release from Group 1 and Group 2 was not significantly different for all the experimental time periods. The control group exhibited the maximum Ca+ ion release for all the time periods as compared to the experimental groups.\n\n\nDiscussion\n\nThe availability of calcium and hydroxyl ion depends on the dissociation of calcium hydroxide, which affects the mineralization process, and surrounding pH. The definitive objective of regenerative endodontic treatment modality for immature root apex with pulpal necrosis is continued root development.10 Immature teeth with open apex with a diameter of 1.1 mm or more respond favorably to REPs11 as it permits the migration of mesenchymal stem cells into the canal space. The revascularization probability in such cases increases by approximately 18-34%.12\n\nThe blood collected from inside the root canal was found to have up to a 600-fold increase of CD73 and CD105 markers of mesenchymal stem cells compared to systemic blood.13 Mechanically irritating the periapical tissues may cause discomfort to the patient and in some cases apical bleeding may not always be possible. Various scaffolds have been used as an alternative to (BC) which is rich in platelets, specifically PRP and PRF.\n\nPRF contains physiological thrombin which creates symmetrical intersections in polymerized fibrin, which helps to release growth factors for up to 28 days. Additionally, their flexible fibrin network facilitates cell migration. Platelet derived growth factors (PDGF) and cytokines are abundant in platelets which play an important role in cellular differentiation. Thus, PRF scaffold has emerged as an effective biological tool in REPs.14 Blood clot makes a weak fibrin mesh as compared to PRF. The use of biomaterials along with platelet concentrates containing fibrin and growth factors could lead to a paradigm shift in how endodontic regeneration can be achieved. MTA provides signalling molecules for the maturation of stem cells.15 Ca+ ions play a crucial role in the formation of mineralized hard tissues. Therefore, the study evaluated the release of Ca+, a key element for regenerating pulp dentine complex.\n\nAn alkaline environment promotes osteogenic differentiation and bone formation. Mineralisation enzymes such as alkaline phosphatase peaks at pH 7.37 and significantly diminished under physiologic level. In vitro and in vivo research has shown that a pH above 8.0 inhibits the mineralization process. Therefore, various biological and molecular responses that influence repair and regeneration may depend on the local pH.\n\nIt is important to create an antibacterial environment at the tooth restoration interface or to control the residual bacteria in the canal space to reduce the risk of reinfection.16 Hydroxyl ions released from biomaterials thus create a hostile environment for bacterial survival and proliferation reducing this reinfection.\n\nWhite MTA (WMTA) when in contact with tissue fluid, MTA dissolves and releases hydroxyl ions (OH-) increasing the pH to 11-12 and contributing to reparative dentin formation.17 Therefore the present study evaluated the influence of two different scaffolds along with MTA on the pH.\n\nAll experimental groups showed an increase in pH over period of time. The highest alkaline pH was recorded in the control group, MTA with distilled water at seven days (11.54) and the lowest in PRF/MTA group (7.78). The difference in pH between PRF and blood clot at 3h and seven days (p < 0.001) was significant, but on the 14th day these groups recorded similar values. Group 3 recorded a mean pH value of 10.76 at 3h, 11.54 on the seventh day and 11.14 on the 14th day which was significantly different from the experimental groups. This is in accordance with the study.9 In the same study, WMTA with blood recorded higher pH values in contrast to our study where the BC with MTA group recorded lower pH values. This could be because the experimental set up was different in that study: in that study, the MTA cylinder was prepared and immersed in blood whereas in our study, PRF/BC was placed over MTA and the pH recorded.\n\nCalcium ions are known to act on osteoblasts and cementoblast cells, causing their differentiation and hard tissue mineralization. Hunter et al. (2018) reported that calcium ions released from biomaterials may influence the repair process, as they pass through the cell membranes by depolarization or activation of membrane-bound calcium channels.18 Although Ca+ ions are one of the major components released by MTA, the role of Ca ions in regenerative endodontics is largely underexplored. In the present study, the mean values for Ca+ ion release from Group1 (PRF/MTA) and Group 2 (BC/MTA) was 0.77 mg L-1 and 0.72 mg L-1 respectively at 3h, which were not significantly different from each other. At seven days, Group 1 recorded a value of 18.39 mg L-1 and Group 2 recorded a value of 39.20 mg L-1. Though the mean Ca+ ion release values of Group 2 were higher than for Group 1, it was not statistically significant at seven days (p ≤ 0.001). At day 14, there was significant reduction in the Ca+ ion release where Group 1 recorded a value of 2.01 mg L-1 and Group 2 recorded a value of 2.75 mg L-1, which were again not significantly different from each other. Overall Group1 (PRF/MTA) showed the lowest release of calcium ions compared to Group 2 (BC/MTA) in14 days. At all experimental periods, Group 3 recorded the maximum Ca+ ion release, which was statistically significant for both experimental groups. A previous study9 evaluated Ca+ ion release from WMTA with and without blood respectively, and reported that the group with WMTA and blood showed greater calcium ion release, which is contrary to what was recorded in our study. This could be because the experimental set up was different in that study, where a MTA cylinder was prepared and immersed in blood; in contrast, in our study, blood clot was placed over MTA and the ion release was recorded. To measure the release of Ca+ ions, ICP-OES equipment was used, owing to its high sensitivity and stability for inorganic analysis. It provides rapid and multi-element analysis of the solutions. In this in vitro study, both experimental groups showed diffusion of Ca+ ions from MTA through scaffolds at three hours, seven days and 14 days, though PRF/MTA allowed lower diffusion of Ca+ ion as compared to the blood clot. However, various clinical studies have suggested that PRFs induce regeneration of pulp dentine complex better than blood clot as it is a rich source of platelets and growth factors.15\n\nThere have been a limited number of studies evaluating the effect of PRF/BC with MTA on the pH and calcium ion release. In the present study, we observed that PRF and BC influenced the pH and Ca+ ion release from MTA.\n\n\nConclusions\n\nWithin the limitations of this in vitro study, which evaluated the influence of two scaffolds on the pH and calcium ion release from MTA over different time periods, it can be concluded that:\n\n1. PRF and BC influenced the pH and Ca+ ion release from MTA. The values were significantly lower in the experimental groups as compared to the control group for all experimental durations.\n\n2. Ca+ ion release in the PRF and BC group increased up to the seventh day, which reduced significantly by the fourteenth day with no significant difference between both groups.", "appendix": "Data availability\n\nMendeley Data: pH and Calcium ion release from MTA when interacted with various substances, https://doi.org/10.17632/jr9sm49gj9.2. 19\n\nThis project contains the following underlying data:\n\n‐ Mendley Dataset Values.docx\n\n‐ Raw data.csv\n\n‐ Results.csv\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nNakashima M, Akamine A: The application of tissue engineering to regeneration of pulp and dentin in endodontics. J. Endod. 2005 Oct 1; 31(10): 711–718. Publisher Full Text\n\nChrepa V, Pitcher B, Henry MA, et al.: Survival of the apical papilla and its resident stem cells in a case of advanced pulpal necrosis and apical periodontitis. J. Endod. 2017 Apr 1; 43(4): 561–567. PubMed Abstract \| Publisher Full Text\n\nSong M, Yu B, Kim S, et al.: Clinical and molecular perspectives of reparative dentin formation: lessons learned from pulp-capping materials and the emerging roles of calcium. Dent. Clin. 2017 Jan 1; 61(1): 93–110. PubMed Abstract \| Publisher Full Text\n\nMirhadi H, Moazzami F, Safarzade S: The Effect of Acidic pH on Microleakage of Mineral Trioxide Aggregate and Calcium-Enriched Mixture Apical Plugs. Iran. Endod. J. 2014; 9(4): 257–260. PubMed Abstract\n\nJung GY, Park YJ, Han JS: Effects of HA released calcium ion on osteoblast differentiation. J. Mater. Sci.: Mater. Med. 2010 May; 21(5): 1649–1654. PubMed Abstract \| Publisher Full Text\n\nShawky MO, Said Badr AE, El-Monem Tawfik MA, et al.: Evaluation of Calcium Ion Release from Two Calcium Silicate-Based Endodontic Materials. SCIOL Biotechnol. 2018; 1: 34–38.\n\nSáez MD, López GL, Atlas D, et al.: Evaluation of pH and calcium ion diffusion from calcium hydroxide pastes and MTA. Acta odontológica latinoamericana: AOL. 2017 Apr 1; 30(1): 26–32. PubMed Abstract\n\nDohan DM, Choukroun J, Diss A, et al.: Platelet-rich fibrin (PRF): a second generation platelet concentrate. Part I: technological concepts and evolution. Oral Surg. Oral Med. Oral Pathol. Oral Radiol. Endod. 2006; 101: e37–e44. Publisher Full Text\n\nTaweewattanapaisan P, Jantarat J, Ounjai P, et al.: The effects of EDTA on blood clot in regenerative endodontic procedures. J. Endod. 2019 Mar 1; 45(3): 281–286. PubMed Abstract \| Publisher Full Text\n\nChaniotis A: The use of MTA/blood mixture to induce hard tissue healing in a root fractured maxillary central incisor. Case report and treatment considerations. Int. Endod. J. 2014 Oct; 47(10): 989–999. PubMed Abstract \| Publisher Full Text\n\nGarcia-Godoy F, Murray PE: Recommendations for using regenerative endodontic procedures in permanent immature traumatized teeth. Dent. Traumatol. 2012 Feb; 28(1): 33–41. PubMed Abstract \| Publisher Full Text\n\nLee BN, Moon JW, Chang HS, et al.: A review of the regenerative endodontic treatment procedure. Restor. Dent. Endod. 2015 Aug 1; 40(3): 179–187. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLovelace TW, Henry MA, Hargreaves KM, et al.: Evaluation of the delivery of mesenchymal stem cells into the root canal space of necrotic immature teeth after clinical regenerative endodontic procedure. J. Endod. 2011 Feb 1; 37(2): 133–138. PubMed Abstract \| Publisher Full Text\n\nPreeja C, Arun S: Platelet-rich fibrin: Its role in periodontal regeneration. Saudi J. Dent. Res. 2014 Jul 1; 5(2): 117–122. Publisher Full Text\n\nPrabhakar AR, Rani NS, Yavagal C: Revascularization of immature necrotic teeth with platelet-rich fibrin and blood clot. Int. J. Oral Health Sci. 2016 Jan 1; 6(1): 4. Publisher Full Text\n\nGandolfi MG, Siboni F, Prati C: Chemical–physical properties of TheraCal, a novel light-curable MTA-like material for pulp capping. Int. Endod. J. 2012 Jun; 45(6): 571–579. PubMed Abstract \| Publisher Full Text\n\nSong M, Yu B, Kim S, et al.: Clinical and molecular perspectives of reparative dentin formation: lessons learned from pulp-capping materials and the emerging roles of calcium. Dent. Clin. 2017 Jan 1; 61(1): 93–110. PubMed Abstract \| Publisher Full Text\n\nHunter AR, Kirk EE, Robinson DH, et al.: A slow release calcium delivery system for the study of reparative dentine formation. Dent. Traumatol. 1998 Jun; 14(3): 112–118. Publisher Full Text\n\nKhatri S, Mathew S, Nagaraja S, et al.: pH and Calcium ion release from MTA when interacted with various substances. Mendeley Data. 2023; V2. Publisher Full Text" }	[ { "id": "191859", "date": "24 Aug 2023", "name": "Janet N Kirilova", "expertise": [ "Reviewer Expertise Conservative dentistry –glass-ionomer cements", "regenerative endodontics-indirect and direct pulp cupping", "digital dentistry-inlay", "onlay", "overlay" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe proposed research is interesting. The results reflect changes in pH and emission of Ca ions alone with MTA administration or in combination with PRF and blood clots in the case of teeth with an open apex.\nTo improve the work, I would recommend some changes such as:\nIntroduction. To formulate more precisely the purpose of the study.\nThe methodology does not specify the manufacturers of the different drugs. The methodology can be described in more detail for the individual groups. The PRF protocol differs from that of Shoukorun, whether the resulting product has the same properties – rich in cytokines and growth factors. The Shoukroun study is cited as number 8.\nThe coronal end of the root was sealed with wax. What kind and what is the purpose of sealing tightly or not?\nIn the discussion. To discuss the importance of 17% EDTA for tertiary dentin formation.\nThe sources of the last 5 years are relatively few.\nThere is a need to improve English.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10447", "date": "16 Nov 2023", "name": "Soumyadeep Ghosh", "role": "Author Response", "response": "Thank you for your valuable contribution. 1. Purpose of study - The regenerative technique should create an environment that favours disinfection and tissue regeneration. Inducing bleeding or use of PRF for facilitating root closure followed by placing a biomaterial may positively or negatively affect the pH and Ca + release. Knowing the release kinematics can improve our understanding of the regenerative process and help modify treatment strategies. This study thus explored the influence of PRF and BC on the pH and Ca + release from MTA. 2. The methodology - Specification of manufacturers of different drugs - After mechanical instrumentation, the canals were irrigated with 1.5 % sodium hypochlorite (20mL, 5 min) (PERCAN N, SEPTODONT) ,5mL sterile physiological saline(0.9% NaCl) followed by a final rinse with 20 mL of 17% EDTA liquid (PRO Endo EDTA liquid, SS WHITE) and dried using sterile paper points PRF protocol - Same as mentioned by Shoukron, 10 ml blood centrifuged immediately at 3000 revolutions/min (rpm) for 10 min, without anticoagulant. Resulting product has the same properties. Coronal end of the root was sealed with wax (Modelling wax, PYRAX). The purpose of sealing it tightly was to form a matrix against which MTA could be condensedinto the prepared canal to obtain a thickness of 4mm. 3. Discussion Importance of 17% EDTA- Root conditioning with 17% EDTA may neutralize the cytotoxicity provoked by NaOCl, enhancing cellular spreading and the liberation of bioactive molecules from the conditioned dentine. It enabled the release of growth factors i-e TGF- β present in dentin. TGF- β shows the ability to induce odontoblastic differentiation and contribute to dentinogenesis. Sources of last 5 year are incorporated. Shawky MO, Said Badr AE, El-Monem Tawfik MA, et al.: Evaluation of Calcium Ion Release from Two Calcium Silicate-Based Endodontic Materials. SCIOL Biotechnol. 2018;1:34–38. Taweewattanapaisan P, Jantarat J, Ounjai P, et al.: The effects of EDTA on blood clot in regenerative endodontic procedures. J. Endod. 2019 Mar 1;45(3):281–286. 30803535 10.1016/j.joen.2018.10.010 Kucukkaya Eren S, Bahador Zırh E, Zeybek ND, Askerbeyli Örs S, Aksel H, Parashos P. Effect of benzalkonium chloride addition to EDTA on attachment and proliferation of dental pulp stem cells on dentin and on transforming growth factor-β1 release. Odontology. 2021 Apr;109:313-20. dos Reis‐Prado AH, Abreu LG, Fagundes RR, Oliveira SD, Bottino MC, Ribeiro‐Sobrinho AP, Benetti F. Influence of ethylenediaminetetraacetic acid on regenerative endodontics: A systematic review. International Endodontic Journal. 2022 Jun;55(6):579-612. Smoczer C, Yuth KR, Askar MA, Young LA, Paurazas SB. Growth Factors Released from Advanced Platelet-Rich Fibrin in the Presence of Calcium-Based Silicate Materials and Their Impact on the Viability and Migration of Stem Cells of Apical Papilla. Dentistry Journal. 2023 Sep 19;11(9):220 Improvement in English - Rectified." } ] }, { "id": "199278", "date": "07 Sep 2023", "name": "Amjad Abu Hasna", "expertise": [ "Reviewer Expertise Endodontics" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nI would like to extend my congratulations to the author for their dedication and hard work in conducting this study. Certainly, I will try to help the authors to improve their article in some comments\nIt is advisable to include the study's hypothesis towards the end of the introduction section for better contextualization.\n\nCould you please provide the protocol number for the Human Research Ethics Committee of M S Ramaiah University of Applied Sciences in Bangalore, Karnataka?\n\nA more concise conclusion is strongly recommended.\n\nThe authors have referenced a total of 19 sources, with merely 2 of them originating from the past five years. To enhance the scholarly foundation of this study, it is advisable to incorporate more up-to-date references into the bibliography. These recent sources can subsequently be leveraged to bolster the discussion, with a particular focus on highlighting the study's limitations.\nThank you for your attention to these matters.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10446", "date": "16 Nov 2023", "name": "Soumyadeep Ghosh", "role": "Author Response", "response": "Thank you for your valuable comments 1. Hypothesis is included at the end of introduction and it is stated as :- The regenerative technique should create an environment that favours disinfection and tissue regeneration. Inducing bleeding or use of PRF for facilitating root closure followed by placing a biomaterial may positively or negatively affect the pH and Ca + release. Knowing the release kinematics can improve our understanding of the regenerative process and help modify treatment strategies. This study thus explored the influence of PRF and BC on the pH and Ca + ion release from MTA. 2. Protocol Number for the Human Research Ethics Commitee of M S Ramaiah University of Applied Sciences, Bngalore, Karnataka is (EC-2018/PG/25). 3. Conclusion :- While this study provides valuable insights into the interactions between MTA, PRF, and BC, further research, including in vivo studies and clinical trials, is needed. Additionally, it is crucial to evaluate various other interactions between PRF and different biomaterials. It can be concluded that PRF and BC adversely influenced the pH and Ca + release from the biomaterial MTA, and this effect decreased with time. Any interference with the release of calcium and hydroxy ions might impact the regenerative potential of MTA. 4. Recent references have been added : Kucukkaya Eren S, Bahador Zırh E, Zeybek ND, Askerbeyli Örs S, Aksel H, Parashos P. Effect of benzalkonium chloride addition to EDTA on attachment and proliferation of dental pulp stem cells on dentin and on transforming growth factor-β1 release. Odontology. 2021 Apr;109:313-20. dos Reis‐Prado AH, Abreu LG, Fagundes RR, Oliveira SD, Bottino MC, Ribeiro‐Sobrinho AP, Benetti F. Influence of ethylenediaminetetraacetic acid on regenerative endodontics: A systematic review. International Endodontic Journal. 2022 Jun;55(6):579-612. Smoczer C, Yuth KR, Askar MA, Young LA, Paurazas SB. Growth Factors Released from Advanced Platelet-Rich Fibrin in the Presence of Calcium-Based Silicate Materials and Their Impact on the Viability and Migration of Stem Cells of Apical Papilla. Dentistry Journal. 2023 Sep 19;11(9):220" } ] } ]	1	https://f1000research.com/articles/12-364
https://f1000research.com/articles/12-224/v1	28 Feb 23	{ "type": "Study Protocol", "title": "Implementing patient-centred outcome measures in palliative care clinical practice for adults (IMPCOM): Protocol for an update systematic review of facilitators and barriers", "authors": [ "Bárbara Antunes", "Stephen Barclay", "Isla Kuhn", "Kathy Eagar", "Claudia Bausewein", "Fliss Murtagh", "Simon Etkind", "Ben Bowers", "Sarah Dixon", "Roberta Lovick", "Richard Harding", "Irene Higginson", "Farhad Shokraneh", "Bárbara Antunes", "Stephen Barclay", "Isla Kuhn", "Kathy Eagar", "Claudia Bausewein", "Fliss Murtagh", "Simon Etkind", "Ben Bowers", "Sarah Dixon", "Roberta Lovick", "Richard Harding", "Irene Higginson" ], "abstract": "Background: Despite the development of patient-centred or patient-reported outcome measures (PCOMs or PROMs) in palliative and end-of-life care over recent years, their routine use in practice faces continuing challenges. Objective: To update a highly cited literature review, identify and synthesise new evidence on facilitators, barriers, lessons learned, PCOMs used, models of implementation, implementation outcomes, costs, and consequences of implementing PCOMs in palliative care clinical practice. Methods: We will search MEDLINE, PsycINFO, CINAHL, Embase, Emcare, SCI-Expanded, SSCI, ESCI, and BNI. The database search will be supplemented by a list of studies from the expert advisory committee, hand-searching of reference lists for included articles, and citations of the original review. We will include primary studies using a PCOM during clinical care of adult patients with advanced disease in palliative care settings and extract data on reported models of implementation, PCOMs, facilitators, barriers, lessons learned, costs, and implementation outcomes. Gough’s Weight of Evidence Framework will be used to assess the robustness and relevance of the studies. We will narratively synthesise and tabulate the findings. This review will follow PRISMA, PRISMA-Abstract, PRISMA-P, and PRISMA-Search as the reporting guidelines. Source of funding: Marie Curie. The funder is not involved in designing or conducting this study. Protocol registration: CRD42023398653 (13/02/2023)", "keywords": [ "Patient-centred outcome measurement", "patient-reported outcome measurement", "palliative care", "systematic reviews", "implementation", "barriers", "facilitators" ], "content": "Introduction\n\nThe importance of using outcome measures to understand the effect and effectiveness of health interventions has been growing as they are increasingly credited as an essential component of evidence-based clinical practice, specifically, outcome measures that the patient completes - so-called patient-reported outcome measures or PROMs - because of their specificity to patients’ individual needs. Data collected at the individual patient level can be used immediately for patient-centred care, as it allows healthcare professionals to act on any distressing symptoms or palliative needs they might have. It can also inform shared decision-making and advance care planning (Antunes et al. 2014, 2022; Antunes and Ferreira 2020). Additionally, PROMs data can be aggregated for audit, research, quality improvement and benchmarking (Currow et al. 2015). However important the PROMs are, measuring can be challenging when the patient is becoming too ill or approaching the end of their life.\n\nIn palliative and end-of-life care, it is equally important to learn about physical effects and psychological, existential, emotional and practical outcomes. However, implementing PROMs as a routine part of palliative care clinical practice has been slow and challenging in a number of countries. Compared to other settings and conditions, measuring outcomes in palliative care faces unique challenges. One reason is that patients’ health is expected to deteriorate, and symptoms may worsen; deterioration will make the outcome measurement challenging, as well as changes in cognitive abilities towards the end of life. PROMs are impossible to use closer to the time of death once the patient becomes unable to communicate (Bausewein et al. 2011). This raises ethical and practical challenges when proxy outcome measurements are used for patients with cognitive impairments (Martins Pereira and Hernández-Marrero 2018).\n\nOur previous review summarised barriers, facilitators, and lessons learned from the published palliative care literature and provided recommendations for implementing outcome measures at the patient, healthcare professional, and management and policy-makers levels for three timepoints: preparation, implementation, and assessment/improvement (Antunes et al. 2014).\n\nOur new systematic review will update and expand the previous review conducted in 2013 on facilitators, barriers, and lessons learned in implementing PROMs in palliative care clinical practice. In addition, we will seek to identify the implementation models used and the costs of implementing those measures. We will also review the literature concerning patient-centred outcome measures reported by family members and healthcare professionals when the patient is not able to do so themselves. For this review, we will use the term ‘patient-centred outcome measures’ (PCOMs) to include both patient-reported and proxy-reported measures that focus on the concerns important to patients.\n\nThe original systematic review (cited 307 times on 16 January 2023) was published in Palliative Medicine and included the literature published between 1985 and 2013. In the last decade, there has been a plethora of publications demonstrating the exponential growth in palliative care outcome measurement. An update on these developments is warranted.\n\n\n\n1. To identify the patient-centred outcome measures implemented in palliative care clinical practice;\n\n2. To identify the facilitators of implementing patient-centred outcome measures in palliative care clinical practice;\n\n3. To identify the barriers to implementing patient-centred outcome measures in palliative care clinical practice;\n\n4. To identify lessons learned on implementing patient-centred outcome measures in palliative care clinical practice;\n\n5. To identify the implementation models used when implementing patient-centred outcome measures in palliative care clinical practice;\n\n6. To identify what implementation outcomes were measured and how, when implementing patient-centred outcome measures in palliative care clinical practice;\n\n7. To assess the financial costs of implementing patient-centred outcome measures in palliative care clinical practice;\n\n8. To update the recommendations from our previous literature to inform the implementation process in palliative care clinical practice for stakeholders at different levels.\n\n\nMethods\n\nThis systematic literature review and narrative synthesis will be reported based on PRISMA-P (Moher et al. 2015), PRISMA including PRISMA Abstract (Page et al. 2021), and PRISMA-Search (Rethlefsen et al. 2021). This protocol has been registered on PROSPERO (CRD42023398653) on 13th February 2023.\n\nTypes of studies\n\nPrimary research studies reporting information related to one of the eight objectives of this review outlined above. All types of evidence synthesis will be considered research studies and included.\n\nTypes of participants\n\nAdult patients (18 years old and over) with advanced diseases and receiving palliative and end-of-life care, their informal carers, and healthcare professionals.\n\nTypes of setting\n\nAll settings in which palliative care is provided. As in the original review, we will include studies conducted both in specialist and non-specialist palliative care settings, provided that in the non-specialist settings, a palliative care measure was implemented.\n\nImplementation of outcome measures as the intervention\n\nIncluded articles will report on the implementation of PROMs. PROMs are a form of outcome measure and comprise standardised validated questionnaires that patients complete to measure their perceptions of their own health status and well-being (Etkind et al. 2015). Since focusing on PROMs alone runs the risk of excluding less well or unwell patients who may not be able to complete those measures themselves, proxy outcome measures completed by families and professionals - PCOMs - are useful and will be considered in this review. Previous authors have highlighted patient-centeredness as key to outcome measurement in palliative care (Etkind et al. 2015; Anhang Price and Elliott 2018).\n\nTypes of studies\n\nNarrative reviews, editorials, commentaries, and case reports will be excluded. If we identify relevant narrative reviews, we will check their references for eligible studies.\n\nTypes of participants\n\nStudies with a paediatric population or a mixed population where fewer than 50% of participants are aged 18 years old or over.\n\nTypes of setting\n\nStudies with non-palliative care settings or mixed settings when fewer than 50% of participants are from palliative care settings.\n\nImplementation of outcome measures as the intervention\n\nStudies reporting data from routine outcome measurement without information on the implementation process as they relate to settings in which implementation has already taken place.\n\nStudies reporting exclusively on the development, testing and feasibility phases of new measures or validation of the linguistic translation of measures, as they do not relate to implementing measures in clinical practice.\n\nElectronic searches\n\nAn information specialist will systematically search MEDLINE, Embase, and Emcare via Ovid SP, CINAHL and PsycINFO via EBSCOhost, British Nursing Index via ProQuest, Science Citation Index-Expanded, Social Science Citation Index, and Emerging Sources Citation Index via Web of Science without any limitations to language, date, document type, or publication status.\n\nThe searches were designed by an expert information scientist and peer-reviewed by another information scientist in collaboration with two clinical experts. The search strategies were tailored based on the included studies in the original review and tested against a new set of relevant studies supplied by the experts in MEDLINE before the agreement on the final search strategy.\n\nThe search strategy for MEDLINE via Ovid SP can be found as Extended data (Shokraneh et al. 2023a).\n\nSearching other resources\n\nCitations of the original review in Web of Science, Google Scholar, and Scopus will be collected and screened. In addition, we will hand-search the reference lists of all included articles and relevant review articles.\n\nThe results will be imported into EndNote and de-duplicated. We expect to screen in the region of 20,000 records after removing duplicate records. The de-duplicated search results will be exported from EndNote and imported into Rayyan (Ouzzani et al. 2016). Two reviewers will screen 20% of the results independently using the “blinding mode”. If there is more than 90% agreement, a single reviewer will screen each result after this point. Any discrepancies between the two reviewers will be resolved by discussion. All full reports will be independently checked by two reviewers. If a disagreement between the two reviewers continues, a senior topic expert will make the final decision on the eligibility of the studies.\n\nAs in the initial systematic review, we anticipate considerable heterogeneity of papers and that eligibility criteria would need to be supplemented with decision rules. These will be formalised and applied consistently; disagreements between research team members will be resolved by discussion.\n\nThe data extraction form will be designed, piloted and revised based on the following data points:\n\n- Patient-centred outcome measures used (name, frequency, format, length)\n\n- Outcome reporter (patient, nurse, family member, carer, general practitioner)\n\n- Factors (facilitators, barriers) related to the implementation\n\n- Implementation models used\n\n- Outcomes of implementation (Peters et al. 2013)\n\n- Costs associated with implementation\n\n- Type of implementation research objective (Peters et al. 2013)\n\n- Implications, proposal, or suggestions (lessons learned)\n\n- Country\n\n- Date of research and publication\n\n- Age groups\n\n- Healthcare conditions\n\n- Setting (home, hospice, hospital, palliative care unit, care home, etc.)\n\n- Scale of the study (pilot, local or national studies of implementation)\n\n- Study design\n\n- Objectives of the study\n\n- Limitations and strengths of the study\n\n- Sample size\n\n- Notes (including relevant information not matching other data points)\n\n- Potentially relevant references\n\n- Lead author's name and email address\n\nWe will initially use the form in Google Docs document format for piloting, and when finalised, we will use Google Sheets to extract data.\n\nQualitative data will be extracted and analysed following the Guidance on the Conduct of Narrative Synthesis in Systematic Reviews (Popay et al. 2006): Element 1 is the role of theory in evidence synthesis; Element 2 is the development of a preliminary synthesis; Element 3 is exploring relationships within and between studies; Element 4 is the assessment of the robustness of the synthesis.\n\nIn this update, we aim to extract frameworks from the studies. Frameworks are graphical or narrative representations of a phenomenon’s factors, concepts, or variables. Implementation frameworks provide a structure for: (a) describing/guiding the process of translating effective interventions and research evidence into practice (process frameworks); (b) analysing what influences implementation outcomes (determinant frameworks); and (c) evaluating implementation efforts (outcome frameworks) (Moullin et al. 2020; Nelson et al. 2020; Van Der Wees et al. 2019). The application of conceptual and theoretical frameworks is key to advancing implementation knowledge. These studies will guide clinical teams on how best to implement patient-centred outcome measures in their respective contexts by providing a foundation on generalisable evidence. The main strength of frameworks is that they organise, explain, or describe information and the range and relationships between concepts. They delineate processes and hence are useful for communication (Bradshaw et al. 2021a, 2021b). We will determine which implementation frameworks are used (if any) and how useful they are. Suboptimal use of frameworks is known to impact the viability and success of implementation efforts, resulting in wasted resources, erroneous conclusions, and specification errors in implementation methods and data analyses (Stover et al. 2021).\n\nWe will consider and, where feasible, extract the PROGRESS plus (O'Neill et al. 2014) characteristics: the place of residence, race/ethnicity/culture/language, occupation, gender/sex, religion, education, socioeconomic status and social capital.\n\nWe will adopt a well-tested multilevel framework for health program evaluation to describe and analyse implementation with six domains: delivery; impact; sustainability; capacity building; generalizability; and dissemination (Masso et al. 2017). The extent to which this will be possible will depend on the degree of implementation undertaken and reported, including the complexity, description of constructs, visual representation and organisation levels.\n\nWe will update the recommendations on implementing patient-centred outcome measures in palliative care clinical practice generated in the original systematic review.\n\nAssessment of robustness and relevance of included studies\n\nThe previous review used the Modified Harden criteria (Barnett-Page and Thomas 2009; Slort et al. 2011) to appraise the quality of included studies. For this update, we will use Gough’s Weight of Evidence Framework (Gough 2007) to assess the robustness and relevance of studies in line with our objectives; the framework applies to quantitative, qualitative and mixed-methods research at the study level. The weight of evidence refers to the preponderance of evidence used to inform decision-making. It considers judgements on different generic judgement (Weight of Evidence A), review-specific judgement on research design (Weight of Evidence B), and a review-specific criterion of evidence focus (Weight of Evidence C), and then a combination of them for an overall judgement (Weight of Evidence D) (Gough 2004):\n\nWeight of Evidence A (WoE A): A generic and non-review-specific judgement about the coherence and integrity of the evidence in its own terms, i.e. the generally accepted criteria for evaluating the quality of this type of evidence.\n\nWeight of Evidence B (WoE B): A review-specific judgement about the appropriateness of that form of evidence for answering the review questions, i.e. the fitness for the purpose of that form of evidence.\n\nWeight of Evidence C (WoE C): A review-specific judgement about the relevance of the focus of the evidence for the review question, e.g. the relevance of the type of sample, evidence gathering or analysis in relation to the review question.\n\nThese three sets of judgements can then be combined to form an overall assessment Weight of Evidence D (WoE D) of the extent that a study contributes evidence to answer a review question.\n\nTwo reviewers will independently make the assessments, with any disagreement resolved by discussion and, if needed, by a third assessor.\n\n\nChanges from the original review\n\nWe have replaced the terminology patient-reported outcome measures (PROMs) with patient-centred outcome measures (PCOMs).\n\nFour new objectives in this update were not part of the previous review: implementation outcomes, costs, implementation models, and updating the recommendations.\n\nWe will not have any date limitations in this update.\n\nSince more studies today use the term “implementation”, we added implementation and related terms to the search strategy after consultation and testing the search.\n\n\nDissemination\n\nThe findings will be published in a peer-reviewed MEDLINE-indexed journal. An abstract from the current work has been accepted for EAPC 2023 (European Association for Palliative Care's 18th World Congress) (Shokraneh et al. 2023b).\n\nThe search stage has been completed and the reviewers are screening the search results.\n\nOne limitation of this update is that we will focus on the settings in which palliative and end-of-life care occur. We will not be able to include studies focusing on particular conditions, such as advanced cancer, outside a palliative care context.", "appendix": "Data availability\n\nNo underlying data are associated with this article.\n\nOpen Science Framework: IMPCOM: Implementing Patient-Centred Outcome Measures in Palliative Care Clinical Practice for Adults: An Update Systematic Review of Facilitators and Barriers. https://doi.org/10.17605/OSF.IO/M4F6A (Shokraneh et al. 2023a).\n\nThis project contains the following extended data:\n\n‐ Appendix 1. Search Strategy for MEDLINE via Ovid SP.docx\n\nOpen Science Framework: PRISMA-P checklist for ‘Implementing patient-centred outcome measures in palliative care clinical practice for adults: (IMPCOM): Protocol for an update systematic review of facilitators and barriers’. https://doi.org/10.17605/OSF.IO/M4F6A (Shokraneh et al. 2023a).\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nAcknowledgements\n\nAn abstract from this research has been accepted for presentation in EAPC 2023 (Shokraneh et al. 2023b).\n\n\nReferences\n\nAnhang Price R, Elliott MN: Measuring Patient-Centeredness of Care for Seriously Ill Individuals: Challenges and Opportunities for Accountability Initiatives. J. Palliat. Med. 2018; 21(S2): S-28–S-35. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAntunes B, Harding R, Higginson IJ, et al.: Implementing patient-reported outcome measures in palliative care clinical practice: a systematic review of facilitators and barriers. Palliat. Med. 2014; 28(2): 158–175. PubMed Abstract \| Publisher Full Text\n\nAntunes B, Ferreira PL: Validation and cultural adaptation of the Integrated Palliative care Outcome Scale (IPOS) for the Portuguese population. BMC Palliat. Care. 2020; 19(1): 178. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAntunes B, Pereira Rodrigues P, Higginson IJ, et al.: Process Evaluation of a Mixed Methods Feasibility Study to Identify Hospital Patients with Palliative Care Needs in Portugal. Acta Medica Port. 2022; 35(2): 94–104. PubMed Abstract \| Publisher Full Text\n\nBarnett-Page E, Thomas J: Methods for the synthesis of qualitative research: a critical review. BMC Med. Res. Methodol. 2009; 9: 59. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBausewein C, Daveson B, Benalia H, et al.: Outcome measurement in palliative care: the essentials. PRISMA. 2011 Mar 23; 1–48.\n\nBradshaw A, Santarelli M, Khamis AM, et al.: Implementing person-centred outcome measures (PCOMs) into routine palliative care: A protocol for a mixed-methods process evaluation of The RESOLVE PCOM Implementation Strategy. BMJ Open. 2021a; 11: e051904. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBradshaw A, Santarelli M, Mulderrig M, et al.: Implementing person-centred outcome measures in palliative care: An exploratory qualitative study using Normalisation Process Theory to understand processes and context. Palliat. Med. 2021b; 35(2): 397–407. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCurrow DC, Allingham S, Yates P, et al.: Improving national hospice/palliative care service symptom outcomes systematically through point-of-care data collection, structured feedback and benchmarking. Support. Care Cancer. 2015 Feb; 23(2): 307–315. Publisher Full Text\n\nEtkind SN, Daveson BA, Kwok W, et al.: Capture, transfer, and feedback of patient-centered outcomes data in palliative care populations: does it make a difference? A systematic review. J. Pain Symptom Manag. 2015; 49(3): 611–624. PubMed Abstract \| Publisher Full Text\n\nGough DA: Systematic research synthesis.Thomas G, Pring R, editors. Evidence-based Practice in Education. Buckingham: Open University Press; 2004; 44–62.\n\nGough D: Weight of evidence: a framework for the appraisal of the quality and relevance of evidence.Furlong J, Oancea A, editors. Applied and Practice-based Research. Special Edition of Research Papers in Education. 2007; 22(2): 213–228.\n\nMartins Pereira S, Hernández-Marrero P: Ethical challenges of outcome measurement in palliative care clinical practice: a systematic review of systematic reviews. Ann. Palliat. Med. 2018; 7(Suppl 3): S207–S218. PubMed Abstract \| Publisher Full Text\n\nMasso M, Quinsey K, Fildes D: Evolution of a multilevel framework for health program evaluation. Aust. Health Rev. 2017; 41(3): 239–245. PubMed Abstract \| Publisher Full Text\n\nMoher D, Shamseer L, Clarke M, et al.: Preferred reporting items for systematic review and meta-analysis protocols (PRISMA-P) 2015 statement. Syst. Rev. 2015; 4(1): 1. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMoullin JC, Dickson KS, Stadnick NA, et al.: Ten recommendations for using implementation frameworks in research and practice. Implement. Sci. Commun. 2020; 1: 42. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNelson TA, Anderson B, Bian J, et al.: Planning for patient-reported outcome implementation: Development of decision tools and practical experience across four clinics. J. Clin. Transl. Sci. 2020; 4(6): 498–507. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nO'Neill J, Tabish H, Welch V, et al.: Applying an equity lens to interventions: using PROGRESS ensures consideration of socially stratifying factors to illuminate inequities in health. J. Clin. Epidemiol. 2014; 67(1): 56–64. PubMed Abstract \| Publisher Full Text\n\nOuzzani M, Hammady H, Fedorowicz Z, et al.: Rayyan-a web and mobile app for systematic reviews. Syst. Rev. 2016; 5(1): 210. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPage MJ, McKenzie JE, Bossuyt PM, et al.: The PRISMA 2020 statement: an updated guideline for reporting systematic reviews. BMJ. 2021; 372: n71. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPeters DH, Adam T, Alonge O, et al.: Implementation research: what it is and how to do it. BMJ. 2013; 347: f6753. PubMed Abstract \| Publisher Full Text\n\nPopay J, Roberts H, Sowden A, et al.: Guidance on the conduct of narrative synthesis in systematic reviews. A product from the ESRC methods programme Version.2006; 1(1): b92.\n\nRethlefsen ML, Kirtley S, Waffenschmidt S, et al.: PRISMA-S: an extension to the PRISMA Statement for Reporting Literature Searches in Systematic Reviews. Syst. Rev. 2021; 10(1): 39. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nShokraneh F, Antunes B, Barclay S, et al.: IMPCOM: Implementing Patient-Centred Outcome Measures in Palliative Care Clinical Practice for Adults: An Update Systematic Review of Facilitators and Barriers.2023a, February 16. Publisher Full Text\n\nShokraneh F, Barclay S, Kuhn I, et al.: Implementing patient-reported outcome measures in palliative care clinical practice: A systematic review update of facilitators and barriers. Rotterdam, The Netherlands: EAPC 2023 (European Association of Palliative Care); 15-17 June 2023b.\n\nSlort W, Schweitzer BP, Blankenstein AH, et al.: Perceived barriers and facilitators for general practitioner-patient communication in palliative care: a systematic review. Palliat. Med. 2011; 25(6): 613–629. PubMed Abstract \| Publisher Full Text\n\nStover AM, Haverman L, van Oers HA , et al.: ISOQOL PROMs/PREMs in Clinical Practice Implementation Science Work Group. Using an implementation science approach to implement and evaluate patient-reported outcome measures (PROM) initiatives in routine care settings. Qual. Life Res. 2021; 30(11): 3015–3033. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nvan der Wees PJ , Verkerk EW, Verbiest MEA, et al.: Development of a framework with tools to support the selection and implementation of patient-reported outcome measures. J. Patient Rep. Outcomes. 2019; 3(1): 75. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "211225", "date": "06 Oct 2023", "name": "Madeleine Hinwood", "expertise": [ "Reviewer Expertise Epidemiology", "biostatistics" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis is a protocol describing a planned update of a systematic review. The justification for updating the review is sound, and the objectives are clear.\nIn the Design section of the methods, four PRISMA reporting guidelines are referenced, including PRISMA-P (for systematic review protocols). I would imagine that this protocol should be reported according to PRISMA-P, with the review itself reported according to PRISMA and PRISMA-S.\nOther changes from the previous review were reported, however not consistently. I would like to understand which things changed, and also which remained the same. For example, in 'Data synthesis', you are using an implementation framework published after the original review. What did you previously use to guide data synthesis? The contrast would be helpful here.\nOverall, this will be an interesting update, which has been well-justified and well described.\n\nIs the rationale for, and objectives of, the study clearly described? Yes\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Not applicable", "responses": [ { "c_id": "10461", "date": "09 Nov 2023", "name": "Farhad Shokraneh", "role": "Author Response", "response": "Dear Dr Hinwood, Thank you for spending your time studying and commenting on our protocol. We very much appreciate it. We have accepted both your comments and made changes to this version accordingly. Comment: Four PRISMA reporting guidelines are referenced in the Design section of the methods, including PRISMA-P (for systematic review protocols). I imagine this protocol should be reported according to PRISMA-P, with the review reported according to PRISMA and PRISMA-S. Response: Thank you for this comment. You are right. We followed PRISMA-P for the protocol, and the review will be reported following PRISMA, PRISMA-Abstract and PRISMA-S. We have now clarified it in the text. Comment: Other changes from the previous review were reported, however, not consistently. I would like to understand which things changed and which remained the same. For example, in 'Data synthesis', you use an implementation framework published after the original review. What did you previously use to guide data synthesis? The contrast would be helpful here. Response: Thank you for this comment. Adding implementation processes was not a change in methods but an addition to the existing methods. While conducting the original review, very few studies reported on implementation. We hope this may have changed, and this time, we may have enough information to report on implementation processes using the Masso 2017 multilevel framework for health program evaluation. Using this framework will be specific and sensible for the implementation process and not the other objectives. For data synthesis, we used \"Guidance on the Conduct of Narrative Synthesis in Systematic Reviews\", as cited in both versions, regardless of the addition to our objectives/collected data. We have now added the \"Data synthesis for implementation models and processes\" headline and updated \"Changes from the original review\"." } ] }, { "id": "215104", "date": "25 Oct 2023", "name": "Mitra Tewes", "expertise": [ "Reviewer Expertise Palliative Care", "patient reported outcome measurement" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors of the presented manuscript describe a study protocol for a systematic review of implementing patient-centered (or patient-reported) outcome measures in palliative and end-of-life care. The study objective is the identification of new evidence on facilitators, barriers, lessons learned, used instruments, models of implementation, implementation outcomes, costs, and conseqences of implementing PCOMs. Therfore, the authors describe the search tools, types of studies, participants and settings. Proxy outcome measures will be considered in the presented review to include patients in bad conditions. The study protocol describes the exclusion criteria and search methods in detail.\nThe description of the underlying disease (e.g. cancer vs non-cancer) would be helpful in the \"Data extraction\" section.\nThe changes from the original systematic review are clearly described and relevant.\nIn sum, the authors deal with an important update of the systamatic review for PROMs in palliative care. Therefore, I would accept this manuscript for indexing with only the described little minor revision.\n\nIs the rationale for, and objectives of, the study clearly described? Yes\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [ { "c_id": "10462", "date": "09 Nov 2023", "name": "Farhad Shokraneh", "role": "Author Response", "response": "Dear Dr Tewes, Thank you for spending your time studying and commenting on our protocol. We very much appreciate it. We have accepted your comment and made changes to this version accordingly. Comment: The description of the underlying disease (e.g. cancer vs non-cancer) would be helpful in the \"Data extraction\" section. Response: Thank you. Yes, collecting underlying diseases under Healthcare conditions is one of the data points. We have now added (Cancer, non-cancer, etc.) for clarification." } ] } ]	1	https://f1000research.com/articles/12-224
https://f1000research.com/articles/11-1208/v1	24 Oct 22	{ "type": "Opinion Article", "title": "Dignity in the care of older adults living in nursing homes and long-term care facilities", "authors": [ "Patrick Wachholz", "Karla Giacomin", "Karla Giacomin" ], "abstract": "Depending on the fields and actors involved, dignity may involve, signify, and encompass different meanings. This fundamental right can be subjectively experienced and rooted in a person's perception of being treated and cared for. Care refers to a set of specific activities combined in a complex life-sustaining network, including long-term Care, which involves various services designed to meet a person's health or personal care needs. However, older residents' human rights have been disrespected and widened the gaps between theory and practice regarding the precarious protection of their rights and dignity inside long-term facilities and nursing homes. This paper aims to discuss threats to dignity and elucidate some strategies to promote and conserve dignity in care, including the person-centered practice in long-term care. Some barriers to the dignity of older residents involve the organizational culture, restraints of time, heavy workload, burnout, and lack of partnership between the residents, their families, and the long-term care homes' staff. Person-centered integrated care quality frameworks are core components of a good quality of care in these spaces in high-income countries. Unfortunately, the COVID-19 pandemic highlighted how weak long-term care policies were and demonstrated that much progress in the dignity of care in long-term care facilities and nursing homes is needed. In low- and middle-income countries, long-term care policies do not accompany the accelerated and intense aging process, and there are other threats, like their invisibility to the public sector and the prejudices about this service model. It's urgent to create strategies for designing and implementing sustainable and equitable long- term care systems based on a person-centered service with dignity to everyone who needs it.", "keywords": [ "long-term care", "older adults", "dignity", "aging" ], "content": "Introduction\n\n\n\n“All human beings are born free and equal in dignity and rights. They are endowed with reason and conscience and must act towards one another in a spirit of brotherhood.\n\n(Universal Declaration of Human Rights, 1948).\n\nFlagship value, dignity may involve, signify, and encompass different meanings depending on the fields and actors involved and expected reciprocity between them. For example, the perception of dignity for professionals and healthcare providers can sound different when compared to the perceptions of healthcare users, their families and policy makers.1,2\n\nDespite some controversial criticism on defining it,3 dignity is a core concept that must be guaranteed to any human being, whatever their condition is. Some authors argue that dignity may be a link that explains the interchange between the promotion and protection of human rights and an individual’s health status.4,5 Recently, a substantial body of literature reviewing and analyzing the concepts of ‘dignity’, ‘care with dignity’, and ‘dignified care’ have been published,6,7 reinforcing that dignity is considered to be a fundamental right, subjectively experienced, and rooted in a person’s perception of being treated and regarded as essential and valuable to others.8\n\nCare refers to the provision of a set of specific activities combined to provide help, protection, or supervision in a complex life-sustaining network. It may involve distinct actors and actions, including self-care, caring for others, the caregiver, and the care recipient.9 Of note, the increased demand for care by the older population occurs concomitantly with changes in its provision: a declining number of carers challenges the informal sector due to trends toward reduced family size, a higher proportion of single households, more opportunities for women in the labor market, and increasing migration rates and geographic separation between parents and children.10\n\nLong term care (LTC) involves services designed to provide what is necessary to reach personal care needs, or to maintain their daily lives during a short or long period, particularly for those with limited capacity for self-care because of a chronic illness, injury, physical, cognitive, or mental disabilities.11 LTC includes assistance with activities of daily living (ADLs) and instrumental activities of daily living (IADLs), including dispensing and correct administration of medications, engaging in household chores and hobbies and self-care tasks.\n\nIn most of the countries, LTC provision involves a mix of formal (state, local authority and nongovernment) and informal (primarily unpaid family) provision – though there are wide differences in the balance between these sectors in the context of national cultures and welfare traditions.12 In low- and middle-income countries, LTC is mostly provided informally (typically by family and friends), even for the most vulnerable and functionally-impaired older adults, despite the growth of public and private LTC services in many of these nations.\n\nLTC services help people improve or maintain an optimal quality of life and physical functioning, including but not limited to help from third parties or assistive devices in various settings: in the community (adult daily service center); at home, from a home health agency, hospice, or family and friends; in facilities (nursing home or specialized infirmary); or other residential settings.13 According to the World Health Organization, “LTC homes are living spaces for adults with significant health challenges” in accessing 24-hour nursing and personal care.14\n\nDespite global and regional initiatives in favor of strengthening strategies to promote respect and dignity of older adults, such as the Decade of Healthy Aging,15 LTC facility residents do not seem to have the same priorities and guarantees as their counterparts. The disrespect for older residents’ human rights during the recent period of the COVID-19 pandemic, for example, widened the gaps between theory and practice regarding the precarious protection of their rights and dignity.16\n\nThis paper aims to discuss threats to dignity in LTC facilities (LTCF) and nursing homes (NH) and elucidate some strategies to promote and conserve dignity in care, including the person-centered practice in LTC.\n\nEven though care is provided proficiently or technically competent, residents and family members may perceive it as lacking in dignity. The concept of dignity for the older people living in LTC homes relates to feelings of comfort, autonomy, meaning, interpersonal connection, hope, physical and spiritual state, and belonging, and is influenced by their social interactions and positively or negatively affected by others.1,17\n\nThe Nordenfelt’s theoretical dignity model, developed within the Dignity and Older Europeans Project,18 provides a comprehensive definition of dignity that is very useful to understand how fostering a culture of dignity may have an impact on older residents. It distinguishes between intrinsic and contingent value in four concepts as follows: “Dignity of merit: related to a person’s formal or informal status in society; Dignity as moral stature: linked to self-respect and dependent on the conduct of the individual; Dignity of identity: attached to the person’s identity as a human being, which can be altered by others or external events; Dignity of Menschenwürde: a German word meaning innate or inner dignity that is afforded all humans.”18\n\nThe first three concepts of dignity described by Nordenfelt can vary and often depend on individuals’ conduct, autonomy, integrity, and the people they interact. In the context of aging or illness, dignity of identity is probably the most important of the previous concepts.1 In contrast, Menschenwürde’s dignity deals with innate dignity, which we all possess equally.19\n\nDespite some differences in the causes of admission to NH and LTCF in low- and high-income countries, it is common for a large proportion of residents to have a significant reduction in their cognitive and functional abilities, depending on third parties to perform ADLs and IADLs. According to some authors, dependency affects their dignity (of identity), because it can reduce their control and choice.17,19\n\nRigid or inflexible technical and organizational routines depersonalize care in LTC homes, depriving residents of expressing their opinions and desires. Due to time constraints, resources, and caregivers’ propensity for task-oriented care, the depersonalization of care often compromises the resident’s dignity, who is forced to “obey” mealtimes, hygiene standards, and continence, participation in social activities, and sometimes even control over one’s belongings.1,17,19 Even when residents have their cognition and desire for autonomy preserved, a tension may emerge when organizations decide to maintain a ‘risk-free environment’ by forcing staff and residents to obey rules that limits autonomy and control.1,19\n\nCaregivers’ communication with older residents (or other workers) about themselves or their peers can also threaten the dignity of LTC, even when a resident has impaired communication skills. Using potentially stigmatizing or ageist labels, diminutives or nicknames when referring to a resident is highly undesirable, as well as publicly exposing personal information due to hearing impairment in collective settings.1,17,18\n\nDignity in LTC must always be linked to values of personhood and unique identity and disaggregated from the use of any form of physical or chemical restraints. The “zero tolerance” culture of abuse against the older resident must be an organizational dogma understood and practiced by all staff, including volunteers.\n\nThe right to privacy includes concepts of respect for the dignity of identity also in the promotion of assistance during the control and rise of continence, respecting the resident’s desire for service provided by caregivers of the same sex, for example. The right to privacy includes reducing exposure to the body or assistive devices (such as prostheses or urinary catheters).\n\nEven in environments where economic deprivation can substantially impact access to inputs and food, ensuring frequent, healthy, and palatable meals must be essential. Disregarding food consumption preferences, especially during the approximation periods after entering an LTC home, can significantly impact the perception of dignity and outcomes related to weight loss, sarcopenia, and, consequently, worsening of functional abilities.\n\nSome authors previously suggested best practices for compliance related to resident dignity,20 focusing on requirements that include respecting care needs, maximizing the dining experience, living in a secure facility, participating in activities, and respecting residents’ personal space.21 Best practices may include, for instance, assuring residents preferences related to personal appearance are consistently honored, developing a policy for selective use of clothing protectors during meals and an environment to ensure that direct staff can comfortably assist with feeding, besides addressing residents by their names and providing meaningful activities considering the residents’ abilities and past interests.\n\nIt is important to highlight that dignity of older residents cannot be promoted without reciprocal partnership between them, their family and the LTC homes’ staff.1 Despite previous studies found that organizational culture, restraints of time, heavy workload and burnout have been cited as barriers to a dignified care,6,22,23 providers must make sure that the care and treatment they provide ensure people’s dignity, including having privacy when they need and want it, treating them as equals and providing the support they might need, including involving them in the local community activities.24\n\nPerson-centered integrated care (PC-IC) quality frameworks are core components of a good quality of care in LTC. It is possible to build a 4-stage goal-oriented PC-IC process,17 including (a) personalizing goal settings, (b) care planning aligned with goals, (c) care delivery according to plan, and (d) evaluation of goal attainment.25 A theoretical framework for person-centered practice in long-term care (PeoPLe) is another example of providing a comprehensive guide to empirical inquiry, education, and practice development in LTC homes, serving as a low-threshold starting point for practice development.26\n\nIn addition to constructs in the framework of person-centered practice, previous authors have found significant associations between self-rated health, mobility, and dementia and perceptions of dignity and well-being.27–29\n\nUsing a modified Delphi process to prioritize essential ‘dignity-conserving care markers’, Thompson and colleagues17 found the following practices to be good markers: staff make residents feel valued as a person; staff are compassionate in providing care; residents can trust staff; staff do not make residents feel like a burden to others; residents are able to make choices in their everyday life;17 assistance with hygiene and personal matters is adequate and sensitive; there is freedom to complain without fear of repercussions; staff does not talk about residents in front of other residents; the personal space of the residents and the need for privacy are respected; efforts are made to make residents feel safe.\n\nThe COVID-19 pandemic highlighted long-known weaknesses and shortcomings, which were continually postponed in terms of public and social relevance in its resolutions. It demonstrated that we need to make much progress in the dignity of care in LTC. The challenge of caring for older residents is particularly felt in low- and middle-income countries, where the development of LTC policies does not accompany the accelerated and intense aging process in a context of marked social and gender inequality.13\n\nHowever, given the lack of a national LTC policy, this gap’s most explicit practical expression is the reduced and heterogeneous offer of institutional care in these countries.\n\nWhen considering the provision of comprehensive and person-centered care in LTCF, fundamental aspects must be considered by the workers, family, and managers. Among these aspects, the life project (i.e., the direction that the individual wants to take according to their beliefs) stands out; preferences and their scale of values; the story of life, with which we can get to know the person more deeply and pay close attention. Finally, individualized service plans facilitate the detection of needs, turning LTCFs into units of conviviality closer to a domestic environment in terms of organization, schedules, and spaces.\n\nAccording to the person-centered practice framework, significant associations were found between attitudes of staff, thriving in the indoor-outdoor-mealtime environment, and perceptions of dignity and well-being. This approach targets the attitudes of staff and the care environment, which could be used when designing interventions to promote dignity and well-being.27\n\nIn conclusion, dignity in the LTC goes through the recognition of its need and the support of public policies that, in addition to monitoring, promote more significant knowledge about the reality of the care offered. This also means the confrontation of prejudice about this service model27 and the urgent creation of strategies for designing and implementing sustainable and equitable LTC systems28 that ensure a person-centered service with dignity to everyone who needs it.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nReferences\n\nŠaňáková Š, Čáp J: Dignity from the nurses’ and older patients’ perspective: A qualitative literature review. Nurs. Ethics. agosto de 2019; 26(5): 1292–1309. PubMed Abstract \| Publisher Full Text\n\nHarasym P, Brisbin S, Afzaal M, et al.: Barriers and facilitators to optimal supportive end-of-life palliative care in long-term care facilities: a qualitative descriptive study of community-based and specialist palliative care physicians’ experiences, perceptions and perspectives. BMJ Open. 5 de agosto de 2020; 10(8): e037466. PubMed Abstract \| Publisher Full Text\n\nGallagher A: Editorial: What do we know about dignity in care? Nurs. Ethics. 1o de julho de 2011; 18(4): 471–473. Publisher Full Text\n\nMann null.: Dignity and Health: The UDHR’s Revolutionary First Article. Health Hum. Rights. 1998; 3(2): 30–38. PubMed Abstract \| Publisher Full Text\n\nTangcharoensathien V, Mills A, Patcharanarumol W, et al.: Universal health coverage: time to deliver on political promises. Bull. World Health Organ. 1o de fevereiro de 2020; 98(2): 78–78A. PubMed Abstract \| Publisher Full Text\n\nOosterveld-Vlug MG, Pasman HRW, van Gennip IE , et al.: Changes in the Personal Dignity of Nursing Home Residents: A Longitudinal Qualitative Interview Study. PLoS One. 12 de setembro de 2013; 8(9): e73822. PubMed Abstract \| Publisher Full Text\n\nKinnear D, Williams V, Victor C: The meaning of dignified care: an exploration of health and social care professionals’ perspectives working with older people. BMC. Res. Notes. 27 de novembro de 2014; 7(1): 854. PubMed Abstract \| Publisher Full Text\n\nSimões Â, Sapeta P: Conceito de dignidade na enfermagem: análise teórica da ética do cuidado. Rev Bioét. junho de 2019; 27(2): 244–252. Publisher Full Text\n\nGiacomin KC, Duarte YAO, Camarano AA, et al.: Care and functional disabilities in daily activities – ELSI-Brazil. Rev. Saude Publica. 2018; 52(Suppl 2): 9s–9s. Publisher Full Text\n\nSchulz R, Eden J; Adults C on FC for O et al.: Family Caregiving Roles and Impacts. Families Caring for an Aging America. National Academies Press (US); 2016 [citado 1o de setembro de 2022].Reference Source\n\nNational Institute on Aging: What Is Long-Term Care? National Institute on Aging;2017 [citado 1o de setembro de 2022].Reference Source\n\nBarbieri D, Ghibelli P: FORMAL vs INFORMAL LONG-TERM CARE: ECONOMIC & SOCIAL IMPACTS.[citado 20 de junho de 2022]. (Social Protection Investment in Long-Term Care).Reference Source\n\nLong-term Care Facilities\|CDC:2021 [citado 30 de setembro de 2022].Reference Source\n\nWorld Health Organization: Integrated continuum of long-term care.2020 [citado 1o de setembro de 2022].Reference Source\n\nWorld Health Organization: Decade of Healthy Ageing 2020–2030 [Internet]. Geneva:World Health Organization;2020 [citado 7 de janeiro de 2021].Reference Source\n\nDe Albuquerque E, Green C: The human rights of people living in care homes: never again an afterthought. Nat. Aging. 30 de agosto de 2022; 2: 767–769. Publisher Full Text\n\nThompson GN, McArthur J, Doupe M: Identifying Markers of Dignity-Conserving Care in Long-Term Care: A Modified Delphi Study. Thompson Coon J, organizador. PLoS One. 15 de junho de 2016; 11(6): e0156816. PubMed Abstract \| Publisher Full Text\n\nNordenfelt L, Edgar A: The four notions of dignity. Qual. Ageing Older Adults. 1o de janeiro de 2005; 6(1): 17–21. Publisher Full Text\n\nKane J, de Vries K : Dignity in long-term care: An application of Nordenfelt’s work. Nurs. Ethics. setembro de 2017; 24(6): 744–751. Publisher Full Text\n\nAgency for Health Care Administration: Best Practices For Compliance Related To Resident Dignity.2018 [citado 2 de setembro de 2022].Reference Source\n\nAmerican Health Care Association: Best Practices For Compliance Related To Resident Dignity In Skilled Nursing Facilities.2003 [citado 20 de junho de 2022].Reference Source\n\nJakobsen R, Sørlie V: Dignity of older people in a nursing home: narratives of care providers. Nurs. Ethics. maio de 2010; 17(3): 289–300. PubMed Abstract \| Publisher Full Text\n\nDwyer LL, Andershed B, Nordenfelt L, et al.: Dignity as experienced by nursing home staff. Int. J. Older People Nursing. 2009; 4(3): 185–193. PubMed Abstract \| Publisher Full Text\n\nCare Quality Comission: Regulation 10: Dignity and respect - Care Quality Commission.2022 [citado 3 de setembro de 2022].Reference Source\n\nBerntsen G, Høyem A, Lettrem I, et al.: A person-centered integrated care quality framework, based on a qualitative study of patients’ evaluation of care in light of chronic care ideals. BMC Health Serv. Res. 20 de junho de 2018; 18(1): 479. PubMed Abstract \| Publisher Full Text\n\nMayer H, McCormack B, Hildebrandt C, et al.: Knowing the person of the resident – a theoretical framework for Person-centred Practice in Long-term Care (PeoPLe). International Practice Development Journal. 18 de novembro de 2020; 10: 1–16. Publisher Full Text\n\nRoos C, Alam M, Swall A, et al.: Factors associated with perceptions of dignity and well-being among older people living in residential care facilities in Sweden. A national cross-sectional study. Health Soc. Care Community. 2022; 30(5): e2350–e2364. PubMed Abstract \| Publisher Full Text\n\nKelly F, Reidy M, Denieffe S, et al.: Older adults’ views on their person-centred care needs in a long-term care setting in Ireland. Br. J. Nurs. 9 de maio de 2019; 28(9): 552–557. PubMed Abstract \| Publisher Full Text\n\nBéland F, Hollander MJ: Integrated models of care delivery for the frail elderly: international perspectives. Gac. Sanit. dezembro de 2011; 25(Suppl 2): 138–146. PubMed Abstract \| Publisher Full Text" }	[ { "id": "154087", "date": "15 Nov 2022", "name": "Neil H. Chadborn", "expertise": [ "Reviewer Expertise Public health research and health and social care services research." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis article discusses dignity, an important topic in health and social care, which can be overlooked in health research literature. This is partly because it is difficult to quantify but also because it may not be seen as a 'health outcome'. However, similar to person-centred care, dignity is a crucial element of care as expressed by residents and family.\n\nThe introduction, following the abstract, starts from an ambiguous point, with dignity potentially being different to different people and between different groups of people. This appears at odds with the quote about dignity and rights as being universal concepts. While, clearly there may be different attitudes or priorities with respect to dignity. The next paragraph develops the approach that there is a ‘core concept’ (at least) for the meaning of dignity, so I wonder whether this could be incorporated into the first paragraph so the article does not start with such an open definition. “Of note, the increased demand for care by the older population occurs concomitantly with changes in its provision” – clarity could be improved by stating what scale of change is being discussed – it appears that demographic shift is being discussed, however it could be read as the care of a cohort of people changes as they age. Further, this demographic change may relate to USA and is different other countries? Also, I question the term ‘informal sector’ – if this mainly means family members, I would argue that this group is not a ‘sector’ in an economic sense. My point is that if someone is not receiving a service, then they may be supported by family members; I don’t believe they would be described as ‘receiving long-term care provided by the informal sector’.\nAlso, I have same concerns with this para “In most of the countries, LTC provision involves a mix of formal (state, local authority and nongovernment) and informal (primarily unpaid family) provision – though there are wide differences in the balance between these sectors in the context of national cultures and welfare traditions.” I would argue that family provided care should not be included in the category of ‘receiving care service’. I would define the different sectors, then, as: voluntary (third sector – informal or formal), for-profit, and state. The reason that I labour this point is that individuals (particularly in ‘Western’ cultures) do express different views when they are receiving care from family or friends compared to receiving ‘a service’; particularly for intimate care, individuals may feel that this compromises their dignity, whereas receiving care from a careworker or volunteer from an NGO is likely to be seen as protecting their dignity. This point may be Western-centric and may not apply in different cultural contexts, but I believe the distinction remains important. In fact maybe dignity cannot be described independently of familial duty.\n“Rigid or inflexible technical and organizational routines depersonalize care in LTC homes, depriving residents of expressing their opinions and desires. Due to time constraints, resources, and caregivers’ propensity for task-oriented care, the depersonalization of care often compromises the resident’s dignity, who is forced to “obey” mealtimes, hygiene standards, and continence, participation in social activities, and sometimes even control over one’s belongings” For this paragraph, I think it would be helpful addition to quote the work of Tom Kitwood on malignant social environments (Dementia Reconsidered, 1997).\nI think it would help clarity of the article to state that discussion is focused on long term care facilities – rather than care in own home (also included in the term long-term care).\n\nWhile the authors emphasise protecting, promoting and conserving dignity, it may be worth touching on where there may be exceptions or trade-offs; particularly for safety of the individual, care staff or co-residents (including protection from outbreak of COVID-19).\n\nSpecific comments:\nAbstract: “long-term facilities” should be long-term care facilities\n\nIntro “Flagship value, dignity may involve, signify, and encompass different meanings” for readability amend to: Dignity, as a flagship value, may involve…\n\nCheck formatting of reference 4 – should be Mann, J. “Dignity and Health: The UDHR’s Revolutionary First Article.” Health and Human Rights, vol. 3, no. 2, 1998, pp. 30–38. JSTOR, https://doi.org/10.2307/4065297. Accessed 14 Nov. 2022.\n\nIs the topic of the opinion article discussed accurately in the context of the current literature? Yes\n\nAre all factual statements correct and adequately supported by citations? Yes\n\nAre arguments sufficiently supported by evidence from the published literature? Yes\n\nAre the conclusions drawn balanced and justified on the basis of the presented arguments? Yes", "responses": [ { "c_id": "9315", "date": "08 Feb 2023", "name": "Patrick Wachholz", "role": "Author Response", "response": "We very much appreciate the reviewer's comments and recommendations. We have included your suggestions in this new version, corrected minor errors, and revised the bibliographic references. We agreed with the reviewer that it was necessary to adapt the first paragraph to start the manuscript with a less open definition of dignity. We agree that the term \"informal sector\" may convey the wrong message to the audience and have amended this paragraph accordingly. We have included a mention of Tom Kitwood's work on malign social psychology and its impact on the attitudes and practices of caregivers in the formal sector. Likewise, we corrected the title so that the manuscript focuses on long-term care facilities. We agree that during the COVID-19 pandemic, adaptations and exceptions to practice were necessary to reduce the odds of outbreaks and ensure the safety of care homes, their workers, and residents." } ] } ]	1	https://f1000research.com/articles/11-1208
https://f1000research.com/articles/12-21/v1	09 Jan 23	{ "type": "Study Protocol", "title": "Protocol for a scoping review of work system design in health care", "authors": [ "Oladunni Sarah Okunade", "Victor O. Oladokun", "Chinwe Juliana Iwu-Jaja", "Anelisa Jaca", "Charles Shey Wiysonge", "Victor O. Oladokun", "Chinwe Juliana Iwu-Jaja", "Anelisa Jaca", "Charles Shey Wiysonge" ], "abstract": "Background: Delivery of safe and reliable healthcare to patients and the healthcare workforce shortage amidst growing demand has been major challenge to the healthcare system. Addressing this challenge calls for designing or redesigning of healthcare work system. Work system design which is usually associated with productivity in manufacturing offers a wide spectrum of applicability in addressing this challenge of healthcare system. Despite the availability of primary studies on work system design in healthcare, there are sparse published reviews in specific contexts. This scoping review explores the existing evidence to understand the state of the art of work system design in healthcare. Methods: The scoping review adopts the methodology of Joanna Briggs Institute for scoping review which is based on the methodological framework of Arksey and O’Malley. The search will be done on PubMed, Scopus, and Web of Science for the identification of eligible studies. A grey literature search will also be performed. A two-phase screening and extraction of data will be done by two independent reviewers. Data extraction will be done on a pre-piloted data extraction form. The findings will be presented in tables, figures, and a narrative summary. The scoping review will highlight the state of the art, gaps in knowledge and provide directions for future research. Ethics and dissemination: This is a scoping review of primary studies and therefore ethical approval is not required. The report of the findings will be presented in line with the PRISMA reporting guidelines for scoping reviews (PRISMA-ScR). The results will be submitted to a peer-reviewed scientific journal for publication and presented at relevant conferences.", "keywords": [ "Health care", "Work system design", "Work design", "Work place design", "Macroergonomics", "Time and motion study", "System engineering", "Human factors" ], "content": "Introduction\n\nThe provision of safe and reliable healthcare for patients with an adequate and accessible health workforce is the bedrock of an efficient and effective healthcare work system. The ability of the healthcare system to deliver safe and reliable healthcare to patients is vital to earning public trust.1 Delivery of safe and reliable healthcare to the patient has been a challenge due to ineffectiveness and inefficiencies in the performance of the healthcare work system thereby resulting in patient safety problems.\n\nAdequate and accessible health workforce is fundamental to an integrated and effective health system and provision of care, however, there is a global projection of a shortfall of 15 million health workers by 2030, with a major proportion in the low and middle-income countries.1 Poor design of the healthcare work environment also contributes to the shortage of health workers. The shortage of nurses has been linked to the inability to attract and retain nurses because of poor working condition.2,3 Carayon et al. (2012)4 argue that the working conditions of healthcare professionals and workers are sources of stress, burn out and dissatisfaction. Poor infrastructure, unsafe environment, and unfair distribution of incentives among other factors are accountable for poor working conditions of health care personnel.5\n\nAddressing these challenges of the inability of the healthcare system to deliver safe and reliable health care to patients and workforce shortage effectively amidst growing demand for healthcare services calls for designing or redesigning of healthcare work system. Work system design which is usually associated with productivity improvement in the manufacturing sector offers a wide spectrum of applicability in addressing the challenges of healthcare systems.6–12 The knowledge and application of work system design characteristics and principles provide the basis for healthcare organizations to engage in work improvements that can ultimately result in a variety of positive outcomes such as reduced turnover for organizations, increased job satisfaction for the workers and improved quality and safety of care for patients and their caregivers.4,13–16 The term “system” as used in healthcare literature refers to an entity that aids the improvement of quality of care.17–20 But the concept of “system” from Industrial and Systems Engineering point of view, also positioned by Carayon et al.4 refers to “various elements of work that a healthcare person uses, encounters or experiences in order to perform his/her job”. The system approach considers all elements of the system and their interconnections as well as the layout of the system in ensuring the goal(s) of the system is achieved.21\n\nWork system design (WSD) involves a systematic approach that considers various components that make up the work system. It aims at minimizing or eliminating the negative aspects of work which contribute to the poor performance of the work system.4 Work system design ensures that the work environment is designed for workers to have optimal workload, human safety, health and well-being ensured and the overall system performance is optimised.22 The design of work systems has benefited greatly from the principles, theories, tools and techniques of Human Factors, Systems and Method Engineering. Human Factors Engineering has been identified by the National Academy of Engineering and the Institute of Medicine as an important tool for designing better healthcare systems.4,23 This suggests work system design is a robust tool that has potential in improving the healthcare system. However, there is a need to investigate work system design approaches that have been used in health care.\n\nDespite the availability of primary research studies on work system design in healthcare, there are sparse published reviews. Also, published reviews considered specific context in the application of work system design to healthcare. For instance, reviews have explored the use of human factors and ergonomics approach to work system design in healthcare.24,25 Also, a review has demonstrated the use of the Systems Engineering Initiative for Patient Safety (SEIPS) model to improve patient work.26 To the best of our knowledge, a review with focus on work system design in healthcare in a broader context has not been conducted. A comprehensive understanding of work system design in healthcare is vital to exploring its approaches to addressing the challenges of the healthcare system. Consistent with scoping review which is known for mapping out the body of literature on a topic area,27 this scoping review aims to explore the existing evidence to understand the state of the art of work system design in healthcare. To achieve this aim, the review objectives are:\n\n1. to report work system design approaches in healthcare\n\n2. to identify areas where it has been applied in healthcare\n\n3. identify gaps for further research and make recommendations for future research.\n\n\nMethods\n\nThe scoping review adopts Joanna Briggs Institute (JBI) scoping review methodology which is based on the methodological framework of Arksey and O’Malley.28,29 The methodology has 6 stages which are as follows:\n\nStage 1. identification of the research question;\n\nStage 2. identification of relevant studies;\n\nStage 3. selection of eligible studies;\n\nStage 4. charting the data;\n\nStage 5. collating, summarising and reporting the results;\n\nStage 6. consultation with stakeholders,\n\nStage 6 of the methodology, although optional, is valuable if it can be explored. This scoping review will not consider it because of constraints of budget and time. Also, the Preferred Reporting Items for Systematic reviews and Meta-Analyses extension for Scoping reviews (PRISMA-ScR) checklist was followed in preparing the protocol.\n\nThe research questions were developed using the iterative process of Arksey and O’Malley through consultation with the team and the team came up with the following questions: (1) What are the work system design approaches used in healthcare? (2) Which areas have work system designs been applied in healthcare? (3) What are the gaps and recommendations for future research?\n\nPopulation, Concept, and Context (PPC) framework was used to determine relevant terms and studies.28 The search will identify all studies without any restriction to date and geographical location. The search will be implemented across three electronic databases: PubMed, Scopus, and Web of Science. These databases were selected for comprehensiveness and coverage of a broad range of disciplines. The search strategy consists of keywords generated from related studies and are approved by all authors to describe the scoping review and its methodology: work system design, work system, system engineering, human factors, work design, workplace design, job design, organisation design, macroergonomics, work organisation, time and motion study, healthcare, health system, health care. The preliminary search on PubMed (see Table 1) combines keywords with Boolean operators. This will be adapted for search in other databases in consultation with a librarian.\n\nThe same search strategy will be used for a web search to be conducted on Google Scholar to identify grey literature. In consideration of screening time for each hit and the likelihood of not yielding many more relevant articles, the decision to screen only the first 50 hits was made. Other identified websites such as National Academy of Engineering, Ergonomics and Human factors organization will be manually searched. The studies to be included in this scoping review must meet the following inclusion and exclusion criteria as depicted in Table 2.\n\nThe Population/Concept/Context (PCC) framework is outlined to guide in the identification and screening of relevant studies. The population which identifies important characteristics of participants is not applicable to this study. The concept is focused on exploring the existing evidence in understanding work system design approaches as applied to healthcare. Primary studies on work system design in healthcare will be included. All reviews whether literature or systematic will be excluded. The context for the scoping review is global and studies in the English language only will be included while all studies in other languages will be excluded. There is no date limit to this study.\n\nThe screening process is in two phases; screening of titles and abstracts and screening of full texts. Two independent reviewers will carry out screening of titles and abstracts, to select studies that are in line with PPC framework as given in the inclusion and exclusion criteria. Full-text screening is the second phase to be done by two independent reviewers to select studies that met the inclusion criteria for this scoping review. At this phase, full-text studies will be excluded if they do not meet inclusion requirements and the reason for excluding the studies will be provided in the final report. Data generated by two independent reviewers will be extracted, assessed and discussed. The report of the final results of the search will be presented following the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) flow diagram. Disagreements in the selection process by the two independent reviews will be resolved through dialogue or a third-party reviewer. Consensus will be used in resolving disagreement but if it is not achieved, a third reviewer will be consulted.\n\nData extraction gives a descriptive summary of study results to address scoping review objectives and research questions. Excel spreadsheet was used in the development of a template for data extraction. The validity of the template will be piloted and tested prior to the commencement of data charting. Key information to be extracted includes authors, year, title, journal, study objectives, study setting, and study site, to mention a few. Table 3 gives the data charting framework for this scoping review. A data extraction form will be used for the collection of general information from relevant studies and data relevant to answering the research questions. Data extraction will be done by independent reviewers and consensus will be reached by team members on resolving discrepancies. Revision and update will be done on the data extraction form during the review process to accommodate necessary changes.\n\nThe collation of findings from the data extraction form will be analysed using themes relating to the review objectives. Descriptive analysis to summarize the characteristics of the included studies such as publication year, study setting and site and also the evidence on the approaches of work system design in healthcare, the area of its applications in healthcare and the limitations of the approaches and application of work system design in healthcare. On the other hand, areas that are not well-researched and may require further investigation will also be presented. Results will be presented in form of tables and figures where appropriate. Also, a narrative summary of the findings will be presented.\n\nThis study will explore the existing evidence to understand the state of the art of work system design in healthcare by providing breadth and depth of work system design knowledge as an important tool for designing better healthcare systems. It will also identify gaps in knowledge and provide directions for future research.\n\nCurrently developing and piloting data extraction form, and the next step will be running of the search and screening at a later date.\n\nThe report of the findings will be presented in line with the PRISMA reporting guidelines for scoping reviews (PRISMA-ScR). The results will be submitted to a peer-reviewed scientific journal for publication and presented at relevant conferences and events.", "appendix": "Data availability\n\nNo data are associated with this article.\n\nFigshare: PRISMA-ScR-Fillable-Checklist-ProtocolSubmission.docx. https://doi.org/10.6084/m9.figshare.21618258.v1. 30\n\n- PRISMA-ScR-Fillable-Checklist-Protocol Submission.docx\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nAcknowledgements\n\nThe authors acknowledge the South African Medical Research Council for supporting CJI, AJ, and CSW during the preparation of this manuscript. In addition, the article publication cost for this article is paid for by the South African Medical Research Council, through Cochrane South Africa (Project code 43500).\n\n\nReferences\n\nWorld Health Organization: World health assembly resolution on human resources for health. Seventy-Fifth World Health Assemby. 2022. Publisher Full Text\n\nBuchan J, Aiken L: Solving nursing shortages: A common priority. J. Clin. Nurs. 2008; 17: 3262–3268. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAiken LH, Sloane DM, Bruyneel L, et al.: Nurses’ reports of working conditions and hospital quality of care in 12 countries in Europe. Int. J. Nurs. Stud. 2013; 50: 143–153. PubMed Abstract \| Publisher Full Text\n\nCarayon P, Alvarado CJ, Hundt A:Work System Design in Health Care. Handbook of human factors and ergonomics in health care and patient safety. Carayon P, editor.CRC Press Taylor & Francis Group;2012; 65–79.\n\nManyisa ZM: The Current Status of Working Conditions in Public Hospitals at a Selected Province, South Africa: Part 1. J. Hum. Ecol. 2016; 56: 210–219. Publisher Full Text\n\nAsadzadeh SM, Azadeh A, Negahban A, et al.: Assessment and improvement of integrated HSE and macroergonomics factors by fuzzy cognitive maps: The case of a large gas refinery. J. Loss Prev. Process Ind. 2013; 26: 1015–1026. Publisher Full Text\n\nPerez J, de Looze MP , Bosch T, et al.: Discrete event simulation as an ergonomic tool to predict workload exposures during systems design. Int. J. Ind. Ergon. 2014; 44: 298–306. Publisher Full Text\n\nRealyvásquez-Vargas A, Maldonado-Macías AA, García-Alcaraz JL, et al.: A macroergonomic compatibility index for manufacturing systems. Int. J. Ind. Ergon. 2018; 68: 149–164. Publisher Full Text\n\nMurphy LA, Huang YH, Robertson MM, et al.: A sociotechnical systems approach to enhance safety climate in the trucking industry: Results of an in-depth investigation. Appl. Ergon. 2018; 66: 70–81. Publisher Full Text\n\nSaurin TA, Patriarca R: A taxonomy of interactions in socio-technical systems: A functional perspective. Appl. Ergon. 2020; 82: 102980. PubMed Abstract \| Publisher Full Text\n\nEl Mouayni I, Etienne A, Lux A, et al.: A simulation-based approach for time allowances assessment during production system design with consideration of worker’s fatigue, learning and reliability. Comput. Ind. Eng. 2020; 139: 105650. Publisher Full Text\n\nWahyuni D, Budiman I, Nasution H, et al.: Improving Work System Design using Macro- Ergonomics Approach in Rubber Processing Plant ant. IOP Conf. Series: Materials Science and Engineering 288 012077. 2018. Publisher Full Text\n\nSainfort F, Karsh B, Booske BC, et al.: Applying quality improvement principles to achieve healthy work organizations. J. Qual. Improv. 2001; 27: 469–483. Publisher Full Text\n\nCarayon P, et al.:Work system and patient safety. In Human Factors in Organizational Design and Management-VII. Luczak H, Zink KJ, editors.IEA Press; 2003.\n\nCarayon P, Alvarado C, Hsieh Y, et al.: A macroergonomic approach to patient process analysis: application in outpatient surgery. Proceedings of the XVth Triennial Congress of the International Ergonomics Association. 2003.\n\nPickup L, Nugent B, Bowie P: A preliminary ergonomic analysis of the MRI work system environment: Implications and recommendations for safety and design. Radiography. 2019; 25: 339–345. Publisher Full Text\n\nLeatherman S, Ferris TG, Berwick D, et al.: The role of quality improvement in strengthening health systems in developing countries. Int. J. Qual. Health Care. 2010; 22: 237–243. PubMed Abstract \| Publisher Full Text\n\nElayan H, Aloqaily M, Guizani M: Digital Twin for Intelligent Context-Aware IoT Healthcare Systems. IEEE Internet Things J. 2021; 8: 16749–16757. Publisher Full Text\n\nArah OA, Klazinga NS, Delnoij DMJ, et al.: Conceptual frameworks for health systems performance: A quest for effectiveness, quality, and improvement. Int. J. Qual. Health Care. 2003; 15: 377–398. PubMed Abstract \| Publisher Full Text\n\nMacNeill AJ, McGain F, Sherman JD: Planetary health care: a framework for sustainable health systems. Lancet Planet. Heal. 2021; 5: e66–e68. PubMed Abstract \| Publisher Full Text\n\nBadiru AB:2014; Handbook of industrial and systems engineering :CRC Press.\n\nCharles-Owaba OE: Organisational Design: a quantitative approach. Oputoru Books;2002.\n\nKhunlertkit A(N), et al.: Human Factors in the Wild. Proc. Hum. Factors Ergon. Soc. Annu. Meet. 2016; 60: 652–656.\n\nXie A, Carayon P: A systematic review of human factors and ergonomics (HFE)-based healthcare system redesign for quality of care and patient safety. Ergonomics. 2015; 58: 33–49. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCarayon P, et al.: A systematic review of mixed methods research on human factors and ergonomics in health care. Appl. Ergon. 2015; 51: 291–321. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWerner NE, Ponnala S, Doutcheva N, et al.: Human factors/ergonomics work system analysis of patient work: State of the science and future directions. Int. J. Qual. Health Care. 2021; 33: 60–71. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHanneke R, Asada Y, Lieberman L, et al.: The Scoping Review Method: Mapping the Literature in “Structural Change” Public Health Interventions. Department of Public Health Scholarship and Creative Works.2017. Publisher Full Text\n\nPeters MDJ, et al.: Updated methodological guidance for the conduct of scoping reviews. JBI Evid. Synth. 2020; 18: 2119–2126. PubMed Abstract \| Publisher Full Text\n\nArksey H, O’Malley L: Scoping studies: towards a methodological Framework. Int. J. Soc. Res. Methodol. 2005; 8: 19–32. Publisher Full Text\n\nOkunade O, Oladokun V, Jaja C-JI, et al.:PRISMA-ScR-Fillable-Checklist-Protocol Submission.docx. figshare. Dataset.2022. Publisher Full Text" }	[ { "id": "179129", "date": "04 Jul 2023", "name": "Cecilia Berlin", "expertise": [ "Reviewer Expertise Production ergonomics", "social sustainability", "work system design", "human factors in manufacturing", "macroergonomics", "cognitive ergonomics" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis protocol for a scoping review aims to broadly map out the scope of available work (system) design literature that is applicable to healthcare settings. Its intention, aim, proposed methodology and intended further steps are detailed and described in a sensible and mostly satisfactory way. I think the forthcoming paper sounds promising and I would be interested to read it.\n\nThe protocol (and its subsequent paper) will benefit from a thorough language check. The way this is written, I have a feeling that the authors are competent writers, but there are some incorrect singular/plural noun choices, and sentence structure errors here and there that could be edited for clarity, mostly by reducing length and complexity. One example is the first sentence of the 3rd paragraph (\"Addressing these...\").\n\nResearch question 2 should grammatically be written, \"Which areas have work system designs been applied _to_ in healthcare?\". I would however suggest that \"areas\" is rather vague in this sense, so I would like to suggest \"How have work system design interventions been applied in healthcare?\". In this way, the focus can be placed on reporting intervention studies and empirical findings in RQ2. However, this is just a suggestion to increase clarity.\n\nIn your search terms, remember that there may be some variation in whether it is called \"system engineering\" or \"systems engineering\". I might also recommend searching \"sociotechnical systems\" and \"socio-technical systems\".\n\nIs the rationale for, and objectives of, the study clearly described? Yes\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Not applicable", "responses": [ { "c_id": "10443", "date": "28 Nov 2023", "name": "Oladunni Okunade", "role": "Author Response", "response": "Thank you for your comments. The article has been copyedited. RQ2 has been reviewed as suggested to increase clarity." } ] }, { "id": "185072", "date": "01 Aug 2023", "name": "Carlo Fabricatore", "expertise": [ "Reviewer Expertise Human factors engineering in healthcare and education." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nOverall, the core structure and contents of this protocol seem sound.\nThe article requires some further proofreading to improve readability and amend some sparse typos and/or improve the editing. I believe that there is need and scope to improve the protocol by considering the following points:\nThe introduction proposes a valid emphasis on “work system design”. However, I would recommend strengthening the background, by introducing an overview of the work system theory, in order to better contextualize the reader by explaining what a work system is, what the involved elements and relationships are, and why their perspective is important in healthcare practice.\n\nBy focussing on “application areas” RQ2 seems to me overly vague, and it can lead to difficulties in analyzing data from the review. It seems to me that this scoping review warrants a systematic examinations of applications of work system design (WSD) in healthcare, and that “application areas” can be one of the multiple analyses to be done based on the “application data” extracted from the reviewed studies. Other analyses should then ensue, such as, for example, how many studies have empirically corroborated the effects of WSD, types of effects ascertained, etc. A deeper and better structured analysis along these lines would then facilitate the identification of trends and gaps in current research, and the subsequent proposal of orientation for future research.\n\nI would recommend reporting (and further detailing) the systematic literature search protocol based on the PRISMA-S statement (Rethlefsen et al., 20211), in order to improve the replicability of the search.\n\nThe authors indicate that data extraction will be based on a pre-developed template, and that “The validity of the template will be piloted and tested prior to the commencement of data charting”. This is somewhat confusing to me:\n(a) Firstly, what do the authors mean by “charting”? If this is about identifying and classifying excerpts in the original texts based on predefined categories, then this would sound to me like a data coding procedure appropriate to extract data prior to analysis (e.g., identifying all the excerpts describing which elements of a work system have been re-designed, to later on propose a synthesis highlighting which elements have been more frequently addressed, by how many studies, etc.) If “charting” means something else to the authors, then it should be clarified and justified.\n(b) Secondly, the authors should clarify how they intend to “pilot and test” the template prior to the commencement of the data extraction process. I would also consider that data extraction templates do not need to be “fixed” a-priori. They may be modified through iterative processes of data extraction and coding, e.g., as suggested by the template analysis technique (King, 20122).\n\nConsider discussing the broader relevance of the protocol: does it represent a contribution in and of itself? Can it be useful to guide other reviews?\n\nIs the rationale for, and objectives of, the study clearly described? Partly\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Partly\n\nAre the datasets clearly presented in a useable and accessible format? Not applicable", "responses": [ { "c_id": "10444", "date": "28 Nov 2023", "name": "Oladunni Okunade", "role": "Author Response", "response": "Thank you for your comments. The background of this study has been strengthened as suggested. RQ2 has been reviewed. The PRISMA-S as given by Rethlefsen et al. (2021) may not be applicable to the study because it is meant for reporting systematic review. PRISMA-ScR as stated in the protocol is appropriate PRISMA reporting guidelines for scoping review. The charting refers to data extraction, one of the six stages of the scoping review methodology. “pilot and test” template prior to the commencement of data extraction process is to ensure the validity of the process. This will be carried out on a sample of included studies in order to ensure consistency in the use of the framework for data extraction and if need be, the data extraction framework can be modified" } ] } ]	1	https://f1000research.com/articles/12-21
https://f1000research.com/articles/12-603/v1	05 Jun 23	{ "type": "Case Report", "title": "Case Report: A prenatal diagnosis of osteogenesis imperfecta in a patient with a novel pathogenic variant in COL1A2", "authors": [ "Melissa Sindy Peláez Chomba", "Guillermo Raúl Vásquez Gómez", "Yasser Ciro Sullcahuaman Allende", "Julio Cesar Mendoza Fernández", "Nelson David Purizaca Rosillo", "Alejandra Zevallos", "Vicente Leandro Cruzate Cabrejos", "Melissa Sindy Peláez Chomba", "Guillermo Raúl Vásquez Gómez", "Yasser Ciro Sullcahuaman Allende", "Julio Cesar Mendoza Fernández", "Nelson David Purizaca Rosillo", "Vicente Leandro Cruzate Cabrejos" ], "abstract": "Osteogenesis imperfecta is considered a rare genetic condition which is characterized by bone fragility. In 85% of cases, it is caused by mutations in COL1A1 and COL1A2 genes which are essential to produce type I collagen. We report the case of a female neonate delivered to a 27-year-old women at San Bartolomé Teaching Hospital with a family history of clavicle fracture. A prenatal control with ultrasound was performed to the mother at 29 weeks. A fetus with altered morphology and multiple fractures was found. Therefore, a prenatal diagnosis of osteogenesis imperfecta was performed. The neonate was born with a respiratory distress syndrome and an acyanotic congenital heart disease. Therefore, she remained in NICU until her death. We highlight the importance of prenatal diagnosis, genetic counseling and a multidisciplinary evaluation in this type of pathologies and report a new probably pathogenic variant in the COL1A2 gene detected by exomic sequencing in amniotic fluid.", "keywords": [ "Osteogenesis imperfecta", "newborn", "prenatal diagnosis" ], "content": "Introduction\n\nSkeletal disorders (SD) are a genetically diverse group of 461 different diseases classified into 42 groups.1 This group of rare bone diseases accounts for 5% of all birth defects.2 Among the most common SD in the prenatal stage are achondroplasia, thanatophoric dysplasia, osteogenesis imperfecta (OI), and achondrogenesis.3\n\nOI is a connective tissue genetic disorder with an incidence of 1 in every 15 to 20,000 births and comprises a heterogeneous group of diseases characterized by susceptibility to bone fractures of varying severity.4 The clinical picture presents a wide intra- and interfamily variability, which ranges from individuals with occasional fractures and regular height, to phenotypes characterized by severe skeletal fragility, bone deformities, and significant growth deficiency, to neonatal lethality. Other signs include blue sclera, hearing loss, dentinogenesis imperfecta, heart defects, and joint hypermobility.4,5\n\nThe classification of OI has undergone significant changes with the discovery of new genes, and there is great genetic heterogeneity, including forms of autosomal dominant, autosomal recessive, and X-linked inheritance. Currently, five (05) types of OI are considered, of which around 90% of cases are associated with heterozygous pathogenic/probable pathogenic (PV/PVP) variants in the COL1A1 or COL1A2 genes.1,6\n\nThe objective of the present case was to report the prenatal diagnosis of osteogenesis imperfecta associated with a new probably pathogenic variant in the COL1A2 gene detected by exomic sequencing in amniotic fluid.\n\n\nMethods\n\nFor the extraction of fetal DNA, amniocentesis was performed at 29 weeks and 4 days of gestation, collecting 3 ml of amniotic fluid in an EDTA tube. The sample was labeled and processed by centrifugation at 3000 rpm for 10 minutes to obtain the cell pellet. After washing with Phosphate Buffered Saline (PBS) pH 7.4, the cells resuspended in the same buffer were aliquoted. DNA extraction was performed with the ROCHE® High Pure PCR Template Preparation Kit according to the manufacturer's instructions. The eluate (30uL) was quantified using an Invitrogen Qubit 4 fluorometer and stored at -20°C until use.\n\nFor complete exome sequencing using massive sequencing by synthesis (NGS-Exome) technology, the sample was sent by air to Macrogen according to the requested requirements: Sample [58.348ng/uL] with genomic DNA integrity test by electrophoresis in agarose. The platform used was the following. Sequencing on Novaseq 6000, 150bp PE. Library preparation: Exome capture (SureSelect V6 post capture kit).\n\n\nCase report\n\nThe proband was the first child of a 27-year-old Peruvian mother who works as a call center agent. The mother had a personal and family history of clavicle fracture (Figure 1). An ultrasound was performed in another institution at 22 weeks, which suggested a clinical suspicion of achondroplasia. Therefore, she was referred to the Saint Bartolome National Hospital for specialized treatment.\n\nThe mother was admitted to our institution at 23 weeks and 2 days, an ultrasound evaluation was performed, finding a podalic fetus with reduced mobility, long bones below the fifth percentile (corresponding to biometry at 16 weeks), altered morphology (Figure 2A), and continuity solution, in addition to femur length/abdominal circumference=0.11. Fractures were also found at the level of the femur, tibia, fibula, ulna, and ribs (Figure 2B, C) and at the cephalic level there was a skull deformity on pressure echocardiography and cranial fractures (Figure 2D).\n\nA) Ulna with altered morphology. Arrow indicates a point fracture, B) Ribs that exceed the middle of the thorax, C) multiple rib fractures. D) Deformation of fetal head and cranial fractures can also be observed.\n\nIn the thoracic evaluation, the ribs comprise two thirds of the fetal thorax, suggesting a low risk of pulmonary hypoplasia. Finally, it was concluded that she was pregnant at 21.3 weeks by fetal biometry, with a probable skeletal disorder, for which an evaluation by the Genetics Office was requested.\n\nAfter providing genetic counseling and a multidisciplinary evaluation, a complete exome sequencing study was proposed in amniotic fluid and with parental consent before amniocentesis was performed at 29 weeks and 4 days.\n\nThe variant c.1612G>T (p.Gly538Cys) was identified in the COL1A2 gene (NM_000089.4) in a heterozygous state. This variant was evaluated according to the criteria recommended by the American College of Medical Genetics and Genomics (ACMG) and classified as a likely pathogenic variant.7 The clinical findings of the patient and the presence of the probably pathogenic variant in the COL1A2 gene confirmed the diagnosis of Osteogenesis Imperfecta type II.\n\nAfter 9 prenatal controls in the institution, an elective cesarean section was scheduled at 38 weeks and 6 days, delivering a female newborn with a weighi of 3.330 kg, height 43cm, head circumference 34.5cm, chest circumference 34cm, APGAR 5 at minute, 6 at 5 minutes, and 7 at 10 minutes, associated with respiratory distress, which required endotracheal intubation and hospitalization in the neonatal intensive care unit (NICU) (Figure 3). She was admitted with diagnoses of a 39-week-old newborn, osteogenesis imperfecta type II, and respiratory distress syndrome to rule out acyanotic congenital heart disease.\n\nDoppler echocardiography reported a bicuspid aortic valve, severe aortic stenosis, plus hypoplasia of the aortic ring without a significant obstructive gradient; as well as a cardiothoracic index of 0.70, which is considered pulmonary hypoplasia.\n\nAdvanced life support was provided to the newborn for cardiogenic shock with vasoactive drugs to improve pre- and post-loading. The drugs used were norepinephrine and dobutamine in continuous infusion at low doses. Additionally, a drug treatment with furosemide and captopril at low doses were administrated to control congestive heart failure.\n\nOn the other hand, Intratracheal intubation was performed following initial sedoanalgesia protocols with 0.2mg/kg midazolam. Subsequently, she was maintained with sedoanalgesia and 0.1mg/kg morphine for pain management and dyspnea. She remained hospitalized for 17 days in the NICU with an invasive ventilation support to improve gas exchange and mechanical ventilation. An Enteral nutrition was given with calcium and vitamin D and a prophylactic iron dose of 2mg/kg/day. Furthermore, Anasarca was reduced with diuretic and colloid therapy.\n\nOn mechanical ventilation, the newborn developed pneumonia and was treated with antimicrobial agents for 7 days based on the culture results (Ampicillin: 50 mg/kg every 8 h and Gentamicin: 3 mg/kg once a day). Finally, despite intermittent episodes of dyspnea, the patient's progression was stable, therefore, she was referred to the neonatal unit for follow-up care. However, six days after being discharged from NICU, she was readmitted for a sudden worsening and died from a cardiopulmonary arrest.\n\n\nDiscussion\n\nOI is a genetic disorder of connective tissue with an incidence of 1 in every 15 to 20,000 births. It comprises a heterogeneous group of diseases, with autosomal dominant, autosomal recessive, and X-linked inheritance patterns, whose clinical picture is primarily characterised by susceptibility to bone fractures to a variable degree, as well as bone deformities, significant growth deficiency, blue sclera, hearing loss, dentinogenesis imperfecta, heart defects, and joint hypermobility.4,5,8\n\nClassically, the classification of OI included up to 18 types,9 which have undergone significant changes with the discovery of new genes. Currently, IAO is part of the 25 ED groups, called the Osteogenesis Imperfecta and Decreased Bone Density Group, and five types of IAO are considered,1,6 of which approximately 90% of cases are associated with heterozygous PV/PVP in the COL1A1 or COL1A2 genes. VP/VPP in the COL1A1 or COL1A2 genes alter the alpha-1 and alpha-2 chains of type I collagen, respectively, which is the main component of the bone matrix, generating alterations in the bone architecture that explain the clinical manifestations of OI.8,9\n\nRegarding the prenatal parameters found in the patient, chest circumference <p5 for gestational age, chest circumference <0.75 and relative cardiomegaly have a sensitivity of 55-80%, a specificity of 90-100%, a positive predictive value of 80-100%, and a negative predictive value of 87-91% for the association with pulmonary hypoplasia. The characteristics found in the prenatal ultrasound evaluation, such as altered morphology of the long bones, foot malposition, rib, and skull fractures, support the clinical diagnosis of Osteogenesis Imperfecta.4,5,8\n\nPostnatal physical examination revealed height below the third percentile, blue sclera, short nose, and micromelia. Both thighs were abducted and in external rotation. Neonatal radiography confirmed the presence of curvature and humeral and femur fractures, as well as rib fractures and generalized hypomineralization. Finally, the patient died at 23 days. These findings coincide with the phenotype of OI type II and the reports that 90% of these patients die at 4 weeks of age.4,5,8\n\nThere are several ways to diagnose type II OI. The mainstay is prenatal ultrasound, which can reveal bone fractures during pregnancy. Skeletal abnormalities can be detected as early as 14 weeks, regardless of whether there is a family history.10 It is important to note that misdiagnosis of the foetus during a prenatal ultrasound is common, as ultrasound details might be unreliable.11,12 Therefore, biochemical analysis, such as analysis of collagen production in fibroblast cultures or DNA extraction from blood to verified mutations, is important.13,14 However, technologies such as next-generation sequencing have led to a better personalization of medical care in pathologies such as OI, as they enable a faster diagnosis and a more accurate identification of the polymorphisms.15 Moreover, an accurate diagnosis has a direct impact on the selection of patient treatment. On the other hand, a prenatal diagnosis improves the genetic counseling offered to parents, since it is common that two healthy parents have a child with OI and determine whether it is a sporadic mutation or inherited could be crucial for effective case-to-case surveillance.16,17\n\nIn the present case, the patient's prenatal clinical diagnosis was made through an exome sequencing study with amniotic fluid. We identified the variant c.1612G>T (p.Gly538Cys) in the COL1A2 gene (NM_000089.4) in a heterozygous state. Most cases of type II-IV OI are associated with heterozygous variants in the COL1A1 and COL1A2 genes that generate glycine substitutions. About 80% of these variants have been reported to be of missense type, and, in general, glycine substitutions near the carboxyl-terminus are associated with a more severe phenotype.8,18 We must highlight that this variant is not registered with gnomAD or CLINVAR. Likewise, it is found in exon 28, one base away from the splicing site; furthermore, different computational predictors (REVEL, SIFT, PROVEAN, MutPred, DEOGEN2, MutationTaster) report it as harmful or pathogenic. Based on the criteria recommended by ACMG, this variant was classified as probably pathogenic.7 With these clinical and molecular prenatal findings, the diagnosis of osteogenesis Imperfecta type II was confirmed as associated with a probable pathogenic variant in the COL1A2 gene, also known as the lethal perinatal form.1\n\nCurrently, exome sequencing is a tool that allows for better clinical management of complex cases. According to a study by Jenny Lord et al.19 reported that the application of exome sequencing facilitates the genetic diagnosis of ultrasound-detected fetal structural abnormalities, allowing the prediction of the prognosis and the risk of recurrence in future pregnancies, but the use of exome sequencing should be carefully considered. case selection to maximize clinical utility. Similarly, Zhao et al.20 report that the clinical application of exome sequencing allows detecting the monogenic etiology of pregnancy loss and provides more information for genetic counseling of the risk of recurrence and the management of subsequent pregnancies.\n\nIn this case report, the genetic characterization of the parents was not possible, since genetic tests are not offered free of charge in the public health system. Furthermore, we must emphasize that the mutation in the COL1A2 gene described in this case had incomplete penetration, which causes a phenotypic variation between individuals and highlights the importance of screening the parents. On the other hand, with the sequencing of exomic regions, we missed important regions in the introns, such as microRNA regions or long noncoding RNA regions.21,22\n\nIn conclusion, the diagnosis of osteogenesis imperfecta is quite complex, mainly in the prenatal stage, due to the diversity of types that exist. Therefore, the clinical application of exome sequencing during this stage is essential, since it will allow us to confirm the clinical diagnosis, determine the genetic cause, inheritance patterns, and evaluate multidisciplinary therapeutic options for adequate care, as well as carry out the correct genetic counseling. parents about the prognosis and risk of recurrence in future pregnancies. This is the first prenatal diagnosis of osteogenesis Imperfecta type II confirmed by an amniotic fluid exome sequencing study carried out in our country, thus, as a hospital, we seek to implement measures that ensure that genetic studies are carried out in pregnancies with clinical suspicion of monogenic disease.\n\n\nConsent\n\nWritten informed consent for publication of their clinical details and/or clinical images was obtained from the patient parents.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\n\nReferences\n\nMortier GR, Cohn DH, Cormier-Daire V, et al.: Nosology and classification of genetic skeletal disorders: 2019 revision. Am. J. Med. Genet. A. 2019 Dec; 179(12): 2393–2419. PubMed Abstract \| Publisher Full Text\n\nSabir AH, Cole T: The evolving therapeutic landscape of genetic skeletal disorders. Orphanet J. Rare Dis. 2019; 14(1): 300. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVan Dijk FS, Sillence DO: Osteogenesis imperfecta: clinical diagnosis, nomenclature and severity assessment [published correction appears in Am J Med Genet A. 2015 May; 167A(5):1178]. Am. J. Med. Genet. A. 2014; 164A(6): 1470–1481. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNijhuis WH, Eastwood DM, Allgrove J, et al.: Current concepts in osteogenesis imperfecta: bone structure, biomechanics and medical management. J. Child. Orthop. 2019; 13(1): 1–11. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMarini JC, Cabral WA, Barnes AM, et al.: Components of the collagen prolyl 3-hydroxylation complex are crucial for normal bone development. Cell Cycle. 2007a; 6: 1675–1681. PubMed Abstract \| Publisher Full Text\n\nVan Dijk FS, Cobben JM, Kariminejad A, et al.: Osteogenesis Imperfecta: A Review with Clinical Examples. Mol Syndromol. 2011 Dec; 2(1): 1–20. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJohn BM, Patnaik SK, Thergaonkar RW: Multiple fractures in neonates and osteogenesis imperfecta. Med. J. Armed Forces India. 2006; 62(1): 73–74. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCole WG: Advances in osteogenesis imperfecta. Clin. Orthop. 2002; 401: 6–16. Publisher Full Text\n\nEdelu B, Ndu I, Asinobi I, et al.: Osteogenesis imperfecta: a case report and review of literature. Ann. Med. Health Sci. Res. 2014; 4(7): 1–S5. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVon Bubnoff A: Next-generation sequencing: the race is on. Cell. 2008; 132: 721–723. Publisher Full Text\n\nSteiner RD, Basel D: COL1A1/2 Osteogenesis Imperfecta. 2005 Jan 28 [Updated 2021 May 6].Adam MP, Mirzaa GM, Pagon RA, et al., editors. GeneReviews®. Seattle (WA): University of Washington, Seattle; 1993-2023. Reference Source\n\nÁrvai K, Horváth P, Balla B, et al.: Next-generation sequencing of common osteogenesis imperfecta-related genes in clinical practice. Sci. Rep. 2016; 6: 28417. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMarini JC, Blissett AR: New Genes in Bone Development: What's New in Osteogenesis Imperfecta. J. Clin. Endocrinol. Metabol. 1 August 2013; 98(8): 3095–3103. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLord J, McMullan DJ, Eberhardt RY, et al.: Prenatal Assessment of Genomes and Exomes Consortium. Prenatal exome sequencing analysis in fetal structural anomalies detected by ultrasonography (PAGE): a cohort study. Lancet. 2019 Feb 23; 393(10173): 747–757. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZhao C, Chai H, Zhou Q, et al.: Exome sequencing analysis on products of conception: a cohort study to evaluate clinical utility and genetic etiology for pregnancy loss. Genet. Med. 2021 Mar; 23(3): 435–442. PubMed Abstract \| Publisher Full Text\n\nLiu Y, Wang L, Yang YK, et al.: Prenatal diagnosis of fetal skeletal dysplasia using targeted next-generation sequencing: an analysis of 30 cases. Diagn. Pathol. 2019 Jul 13; 14(1): 76. Publisher Full Text\n\nMucaki EJ, Shirley BC, Rogan PK: Expression Changes Confirm Genomic Variants Predicted to Result in Allele-Specific, Alternative mRNA Splicing. Front. Genet. 2020 Mar 5; 11: 109. Publisher Full Text" }	[ { "id": "195013", "date": "21 Sep 2023", "name": "Xiuli Zhao", "expertise": [ "Reviewer Expertise In the recent ten years", "my research focus on the precise medicine of monogenic diseases", "especially osteogenesis imperfecta. We perform the genetic research in Chinese population with osteogenesis imperfecta." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis paper performed a prenatal diagnosis on a fetus suspected of type II Osteogenesis Imperfecta (OI), a rare hereditary bone disease. It provides a detailed background of the case’s history and progression, including the case’s examination, diagnosis, treatment plan, and final outcomes of the fetus.\nThe genotype-phenotype correlation in OI and similar diseases is still under-researched, and there is significant heterogeneity in patient phenotypes, with a persistent risk of lethal OI in fetuses. The most effective approach currently is prenatal screening for OI-related gene mutations in combination with a pregnancy diagnosis.\nThe potentially pathogenic mutation COL1A2: c.1612G>T (p. Gly538Cys) identified in this paper is reported for the first time, expanding the spectrum of OI pathogenic mutations. Unfortunately, no genetic mutation testing or co-segregation analysis was performed on family members, which are crucial for assessing the pathogenicity of the mutation. In summary, this report presents a very typical case of lethal Type II OI, providing an important diagnostic treatment case and a significant pathogenic mutation.\n\nIs the background of the case’s history and progression described in sufficient detail? Yes\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Yes\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Yes\n\nIs the case presented with sufficient detail to be useful for other practitioners? Yes", "responses": [] }, { "id": "201010", "date": "22 Sep 2023", "name": "Roy Morello", "expertise": [ "Reviewer Expertise Connective tissue diseases" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors report on a case of osteogenesis imperfecta type II that was identified in utero, showing multiple abnormalities by ultrasound, and then molecularly diagnosed by exome sequencing using DNA obtained by amniocentesis. They identified a new heterozygous Gly538Cys substitution in the COL1A2 which is predicted to be pathogenic by multiple computational programs.\nI have the following comments/suggestions:\nAbstract: “…a 27-year-old women…” should read woman.\n\nAbstract: “..a prenatal diagnosis of osteogenesis imperfecta was performed.” Probably better to say ‘issued or requested or suspected’.\n\nAbstract: “…detected by exomic sequencing in amniotic fluid CELLS”\n\nFigure 1: what is the meaning of 26 weeks next to the proband? Arrows should be singular (there is only 1 arrow)\n\n“…drug treatment with furosemide and captopril at low doses were administrated…”, please change to administered.\n\nDiscussion: “Currently, IAO is part of the 25 ED groups….”. Please define what IAO stands for.\n\nDiscussion: “..but the use of exome sequencing should be carefully considered. case selection to maximize clinical utility.” Broken sentence, please fix.\n\nDiscussion: “Furthermore, we must emphasize that the mutation in the COL1A2 gene described in this case had incomplete penetration, which causes a phenotypic variation between individuals and highlights the importance of screening the parents. On the other hand, with the sequencing of exomic regions, we missed important regions in the introns, such as microRNA regions or long noncoding RNA regions.” Please provide some more explanation and/or context about these two sentences which are not easy to follow.\n\nDiscussion: “..the correct genetic counseling. parents about the prognosis”. There is a period in the middle of the sentence. Please fix.\n\nIs the background of the case’s history and progression described in sufficient detail? Yes\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Yes\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Yes\n\nIs the case presented with sufficient detail to be useful for other practitioners? Yes", "responses": [ { "c_id": "10337", "date": "16 Nov 2023", "name": "Alejandra Zevallos", "role": "Author Response", "response": "We highly appreciate the reviewer’s insightful and helpful comments on our manuscript. Many sentences of the manuscript have been carefully rewritten or reorganized to enhance the logic flow and make the statements stricter in a proper tone. Comments/suggestions: 1. Abstract: “…a 27-year-old women…” should read woman. We agree with the reviewer’s observations. We have made the change requested. 2. Abstract: “..a prenatal diagnosis of osteogenesis imperfecta was performed.” Probably better to say ‘issued or requested or suspected’. We thank the suggestion from this reviewer. We have carefully edited the manuscript according to the inputs from the reviewer. 3. Abstract: “…detected by exomic sequencing in amniotic fluid CELLS” We thank the suggestion from this reviewer. We have carefully edited the manuscript according to the inputs from the reviewer. 4. Figure 1: what is the meaning of 26 weeks next to the proband? Arrows should be singular (there is only 1 arrow) We agree that we were unclear in our Figure 1 text. We have added more details to clear what we meant with the words “26 week”. 5. “…drug treatment with furosemide and captopril at low doses were administrated…”, please change to administered. We thank the suggestion from this reviewer. We have carefully edited the manuscript according to the inputs from the reviewer. 6. Discussion: “Currently, IAO is part of the 25 ED groups….”. Please define what IAO stands for. We apologize that our original statement was unclear. We have rewritten this paragraph as follow “Currently, SD are classified into 42 groups with OI been part of the 25th group, called the Osteogenesis Imperfecta and Decreased Bone Density Group. 1 Furthermore, there are five types of OI, with approximately 90% of cases being heterozygous PV/PVP in the COL1A1 or COL1A2 genes. 6” 7. Discussion: “..but the use of exome sequencing should be carefully considered. case selection to maximize clinical utility.” Broken sentence, please fix. We thank the suggestion from this reviewer. We have carefully edited the manuscript according to the inputs from the reviewer. 8. Discussion: “Furthermore, we must emphasize that the mutation in the COL1A2 gene described in this case had incomplete penetration, which causes a phenotypic variation between individuals and highlights the importance of screening the parents. On the other hand, with the sequencing of exomic regions, we missed important regions in the introns, such as microRNA regions or long noncoding RNA regions.” Please provide some more explanation and/or context about these two sentences which are not easy to follow. We admit that the logic may be jumping at some places. Thus, the sentence has been rewritten as “In this case report there were some limitations. First, variant c.1612G>T in the COL1A2 gene has incomplete penetration, leading to variations in phenotypic characteristics among individuals and highlighting the necessity of screening parents. However, genetic testing was not available for free in the public health system, so genetic characterization of the parents was not possible. In addition, we might have missed microRNA regions, long noncoding RNAs, and other important regions in introns, since we sequenced just exomic regions. 21 , 22 On the other hand, it is important to highlight that this is the first prenatal diagnosis of osteogenesis imperfecta type II confirmed by an amniotic fluid exome sequencing study carried out in Peru. In this context, this case report emphasizes the importance of conducting genetic analyses when monogenic diseases are suspected during pregnancy.” 9. Discussion: “..the correct genetic counseling. parents about the prognosis”. There is a period in the middle of the sentence. Please fix. We thank the suggestion from this reviewer. We have reorganized this paragraph to enhance the logic flow and make the statements clearer: “In conclusion, the diagnosis of OI is quite complex, mainly in the prenatal stage, due to the diversity of types that exist. Therefore, exome sequencing during this stage is essential, since it allows professionals to confirm the clinical diagnosis, determine the genetic cause, and inheritance patterns. Additionally, we can provide parents with genetic counseling about the prognosis and risk of recurrence in future pregnancy, along with evaluating multidisciplinary therapeutic options for adequate care.”" } ] } ]	1	https://f1000research.com/articles/12-603
https://f1000research.com/articles/12-919/v1	01 Aug 23	{ "type": "Research Article", "title": "Association between the risk perception of contracting COVID-19 and sociodemographic characteristics in a Peruvian population", "authors": [ "Jhon Alex Zeladita-Huaman", "David Esteban-Espinoza", "Michelle Lozada-Urbano", "Eduardo Franco Chalco", "Marcelo Fernandes Costa", "Henry Castillo-Parra", "Jhon Alex Zeladita-Huaman", "David Esteban-Espinoza", "Michelle Lozada-Urbano", "Marcelo Fernandes Costa", "Henry Castillo-Parra" ], "abstract": "Background: The perception of risk regarding coronavirus disease 2019 (COVID-19) has been widely researched due to its association with the adoption of preventive measures. In addition, since the onset of vaccination, it has been reported that the population perceives a lower risk of getting infected. However, few studies have analyzed the factors associated with risk perception in low- and middle-income countries. The aim of this study was to determine the association between the risk perception of contracting COVID-19 and sociodemographic characteristics in Peruvian population. Methods: An analytical and cross-sectional study was conducted in four cities in Peru from October to December, 2021. The sample consisted of 821 individuals aged 18 years and older. A virtual questionnaire was used to collect sociodemographic data and assess the risk perception of contracting coronavirus based on the Health Belief Model. The process of back-translation, expert judgment, and reliability analysis using split-half correlation was conducted. Student's t-tests, analysis of variance with post hoc Tukey's test, and Spearman's correlation were employed. Results: Of the participants, 53.71% were women and 73.3% had a higher education level, 45.55% are self-employed, and 40.44% did not have a family member infected with COVID-19. The risk perception of COVID-19 infection was associated with participants' family antecedent of COVID-19 (p < 0.05). Regarding the factors analyzed, perceived susceptibility to COVID-19 was associated with age (p=0.002), occupation (p<0.05), and a history of COVID-19 (p<0.05), while the perceived benefits of adopting preventive measures against this disease were associated with educational level (p < 0.001). Conclusions: The risk perception of contracting COVID-19 was higher among those who had multiple infected relatives. Furthermore, the perception of susceptibility and the perceived benefits of using preventive measures were associated with sociodemographic characteristics.", "keywords": [ "Covid-19", "Perception", "Disease Susceptibility", "Health Belief Model" ], "content": "Introduction\n\nEmerging viral infections pose a threat to public health due to their rapid transmission and the increased morbidity and mortality rates in the population. At the end of the coronavirus disease 2019 (COVID-19) pandemic worldwide, over 192 million people were reported to have been infected, with 9.2 million deaths resulting from this viral disease.1\n\nTo prevent the spread of COVID-19, the World Health Organization (WHO) recommended pharmacological measures (vaccines and antiretroviral drugs) and non-pharmacological measures such as hand hygiene, mask usage, and physical distancing. At the beginning of the pandemic, studies conducted in different countries reported that the population had a high level of knowledge, favorable attitudes, and appropriate practices regarding these measures.2 However, subsequent research reported that although the population had a good level of knowledge, adherence to preventive measures was low.3\n\nBased on the theory of common-sense model of self-regulation, the decrease in the adoption of prevention measures could be attributed to a change in risk perception.4 Additionally, at the onset of the pandemic, media and government messages increased the population’s perception of risk, leading individuals to adopt preventive measures. However, after this initial stage and with the prolonged presence of the virus, people gradually ceased to fear it and began to perceive it as part of their daily lives. This can be explained by the “mere exposure effect” proposed by Zajonc,5 who suggested that repeated exposure to a stimulus is sufficient to develop a more positive attitude towards that stimulus. Zajonc5 proposed that the relationship between exposure and liking follows a positive deceleration curve, where the initial exposures have a greater impact on the attitude towards the stimulus compared to subsequent exposures.\n\nThe strength of the mere exposure effect depends on the individual’s pre-existing attitude towards the stimulus and tends to be stronger when the individual is unaware of the stimulus presented. To support this hypothesis, Zajonc discussed three types of supporting studies: word frequency and word evaluation, interpersonal contact and interpersonal attraction, and familiarity with musical selections and other stimuli for which individuals express their liking.6 In other words, when the brain encounters a new or unfamiliar stimulus, it activates the amygdala (fear response). However, with repeated exposure to that stimulus, familiarity is generated, leading to increased confidence. In other words, living with a risk factor makes the brain perceive it as more familiar, gradually decreasing the perception of risk.\n\nAlthough several studies highlight the association between adherence to COVID-19 prevention measures and risk perception,7,8 few studies analyze the risk perception of coronavirus infection. A study conducted in Brazil that developed a scale on risk perception based on the Health Belief Model (HBM) indicates that factors such as the type of transportation used, low-income individuals, and those with autoimmune diseases significantly affect risk perception.9 Furthermore, in addition to stay-at-home measures, different non-uniform patterns of behavior in the population have been demonstrated. Specifically, demographic, socioeconomic, and environmental density characteristics were associated with health outcomes related to exposure risk.10\n\nAnalyzing the factors associated with the perception of contagion risk is an important psychological contribution to understanding population behavior during this and future pandemics. In addition, having studies that explore risk perception in the Peruvian population will allow the design of proposals to reduce risky behaviors and promote the adoption of preventive measures in future pandemics. Therefore, this research aimed to determine the association between the risk perception of contracting COVID-19 and sociodemographic characteristics among Peruvian population.\n\n\nMethods\n\nThe population consisted of residents from four cities in Peru (Callao, La Libertad, Lambayeque, and Lima Metropolitana). The study included residents of the selected cities who had access to an internet-connected device. Participants under 18 years old and those who were unavailable to complete the questionnaire were excluded. The sample size was determined in 492 participants considering that it was expected to find moderate to small size effects (f = 0.15) and a statistical power of 80%. However, data was collected from 1044 participants due to the fact possibility of having significant losses in data due to non-complete answers and also to increase the statistical power. Participants taking part in the online survey were chosen using a convenience sampling method.\n\nThis was an analytical and cross-sectional study in which participants were surveyed using a self-administered questionnaire developed in Google Forms.31 The questionnaire was distributed via email and social media from October to December, 2021.\n\nThe online form consisted of three sections. The first section included the informed consent. The second section collected information on sociodemographic characteristics (sex, age in terms of the complete years from your birthday, marital status, highest level of education, occupation, personal and family history of COVID-19 in the household). The third section had the scale of risk perception of contracting the COVID-19 infection, which was designed based on the Health Belief Model (HBM) and validated in Brazil.9 The scale had 24 questions that assessed five factors: a) perceived susceptibility, corresponding to knowledge and belief about the possibility of getting coronavirus; b) perceived severity, asking about personal beliefs about how the individual would experience the disease process and the intensity of symptoms; c) perceived benefits, corresponding to the effectiveness of adopted behavioral mechanisms to prevent infection; d) perceived barriers, aiming to understand the difficulties in adhering to protective and preventive measures for coronavirus transmission, and e) health motivation, seeking to improve overall health. Responses were rated on an analog scale from 0 to 100, where zero represented “not at all” and 100 represented “extremely high.”\n\nBefore the validation process, a back-translation was performed. Firstly, the questionnaire was translated from Portuguese to Spanish by a certified translator, and then vice versa. Subsequently, a pilot test was conducted with 30 participants to evaluate the comprehension of the questions and response options. In this activity, it was identified that participants had difficulty rating the probability of risk (response option) with a value greater than 100. This is why the range of response options was modified from the original version, which ranged from 0 to 120, to a range from 0 to 100.\n\nThe content of the instrument went through a process of expert judgment validation. In this process, health professionals who are methodologists, members of the COVID-19 command, and epidemiologists.\n\nIn order to explore the reliability of the instrument, a split-half correlation procedure was conducted, correlating all possible halves of items. Based on the obtained correlations, a data distribution was obtained for each subscale of the instrument, and the 2.5th, 50th, and 97.5th percentiles are reported. Split-half correlations with a score of 0.70 or higher are considered adequate.\n\nTable 1 shows the results of the reliability analysis conducted for the scales of perception of COVID-19 contagion risk and its factors. Specifically, we can observe that the median split-half correlations for the general risk perception scale (0.91), susceptibility scale (0.83), and severity scale (0.88) are considerably higher than 0.70, indicating that these scales have very good levels of reliability. On the other hand, the severity, benefit, and barrier scales have low levels of reliability as the median split-half correlations are below 0.70 (0.67, 0.53, 0.57, respectively). Regarding the descriptive statistics of the subscales, it can be observed that the highest average perception of risk is found in the barrier subscale (M = 42.34, SD = 18.03), while the lowest average perception of risk is observed in the benefit perception (M = 28.90, SD = 15.68), followed by susceptibility perception (M = 28.94, SD = 19.50).\n\nIn order to determine the differences in susceptibility to COVID-19 based on sex, marital status, education level, occupation, and family history, a series of Student’s t-tests and Analysis of Variance (ANOVA) were conducted, depending on the number of categories in the contrasting variable. In cases where statistically significant differences were found in the ANOVA, post hoc Tukey analysis was performed to determine the nature of these differences. In all cases, the assumptions of parametric analyses were verified. Finally, to examine the relationship between age and susceptibility to COVID-19, a Spearman correlation coefficient was estimated, as age did not follow a normal distribution. All analyses were carried out using R software v4.2.1.11\n\nDue to the absence of an available sampling frame and the utilization of convenience sampling, the sample obtained in this study was not representative of the population across the four Peruvian cities, limiting the ability to generalize the findings. Nonetheless, the number of participants in this study, nearly twice the minimum sample size, allows for statistically robust results. On another note, as the survey was conducted virtually, the results may be subjected to biases, such as social desirability bias—the tendency for respondents to provide socially acceptable responses rather than responding truthfully.\n\nLastly, given that the participants had to respond numerically to the risk perception questionnaire, potential difficulty in providing responses may have arisen, as some individuals have limitations in quantitatively evaluating their perceptions, or confusion surrounding the interpretation of the questions. For this reason, clear instructions were provided in the questionnaire, and each question reiterated the numerical scale that could be used for responses. In addition, rigorous quality control was conducted on the responses to select the questionnaires that were considered for statistical analysis.\n\nThe study was approved on May 24, 2021 by the Institutional Ethics Committee of Norbert Wiener University (file number 560-2021). All participants provided informed consent before responding to the questionnaire. The data collected through the survey in each city were coded, excluding identifying information, ensuring the confidentiality of the information and exclusive access to the data by the researchers.\n\n\nResults\n\nA total of 1044 participants answered, of which 223 were excluded for quality control purposes because they provided letters or words instead of indicating a number between 0 and 100 in their responses.30 Out of the 821 participants, 53.71% (n = 441) were women, with an average age of 28.29 years (SD = 11.56). Regarding marital status, 76.49% (n = 628) reported being single, 20.95% (n = 172) reported being married, while 2.56% (n = 21) reported being separated or divorced. In terms of participants’ education level, it was observed that 73.33% (n = 602) had a higher education level, while 26.31% (n = 216) had completed secondary education. In regard to the occupation of the participants, 45.55% (n = 374) reported being self-employed, 42.39% (n = 348) reported being employed, 11.33% (n = 93) reported being homemakers, and the remaining 0.73% (n = 6) indicated being retired. Finally, with regard to the history of COVID-19 among family members or individuals living in the same household, at the time of the survey, 40.44% (n = 332) of the participants reported that no family member or household member had contracted COVID-19, 28.01% (n = 230) reported that one family member had been infected, while the remaining 31.55% (n = 259) reported that several members of their family and household had been infected with COVID-19.\n\nIn Table 2, the Pearson correlations for the different scales of risk perception can be observed. In this table, it can be seen that the general scale of risk perception maintains a high and significant correlation with all the subscales (0.72 < r’s < 0.80). On the other hand, the different subscales show moderate to strong correlations if (0.32 < r’s < 0.60).\n\n* p < 0.001.\n\nIn Table 3, the Student’s t-tests for the means of risk perception of contracting the COVID-19 infection can be observed according to the reported sex of the participants. Specifically, it can be indicated that none of the subscales showed statistically significant differences (p’s > 0.5). These results indicate that the risk perception of contracting the COVID-19 infection is the same for men and women.\n\nIn Table 4, the Analysis of Variance (ANOVA) comparisons of the means of risk perception of contracting the COVID-19 infection can be observed according to the marital status reported by the participants. Particularly, it can be observed that none of the subscales showed statistically significant differences (p’s > 0.5). These results indicate that the risk perception of contracting the COVID-19 infection is the same for married, single, and separated participants.\n\nIn Table 5, we can observe the results of the Analysis of Variance (ANOVA) for the means of risk perception of contracting the COVID-19 infection according to the participants’ educational level. For this sociodemographic variable, significant differences were found in the benefit subscale based on educational level (F = 5.75, df = 2,818, p = 0.003). According to the post-hoc Tukey analysis, participants with a secondary education level have a significantly higher benefit score than those with a higher education level (p = 0.004). No other significant differences were observed in any of the other subscales of risk perception of contracting the COVID-19 infection.\n\n* p < 0.05.\n\n p < 0.01.\n\n* p < 0.001.\n\nIn Table 6, we can see the results of the Analysis of Variance (ANOVA) for the mean scores of risk perception of contracting the COVID-19 infection according to participants’ occupation. For this variable, significant differences were observed in the susceptibility subscale (F = 7.47, df = 3,817, p < 0.001). According to the post-hoc Tukey analysis, participants who are employed reported higher levels of susceptibility compared to homemakers (p = 0.008). Similarly, participants who are employed reported higher levels of susceptibility compared to self-employed participants (p < 0.001). No significant differences were found in the other subscales of the instrument.\n\n* p < 0.05.\n\n p < 0.01.\n\n* p < 0.001.\n\nTable 7 shows the results of the ANOVA for the mean scores of risk perception of contracting the COVID-19 infection according to participants’ family history of COVID-19. Significant differences were observed in the general perceived risk scale (F = 3.06, df = 2,818, p = 0.047) and the susceptibility scale (F = 9.66, df = 2,818, p < 0.001) for this sociodemographic variable. Following post-hoc Tukey analysis, it was found that participants with multiple infected family members had higher levels of perceived risk on the general perceived risk scale compared to participants with no infected family members (p = 0.04). In terms of the susceptibility scale, participants with no infected family members reported lower levels of susceptibility compared to those with at least one infected family member (p = 0.03), and those with multiple infected family members (p < 0.001). No other significant differences were observed in any of the other subscales of risk perception of contracting the COVID-19 infection.\n\n* p < 0.05.\n\n p < 0.01.\n\n* p < 0.001.\n\nFinally, regarding the correlation between age and perceived risk, Table 8 reveals that only the susceptibility scale shows a positive, weak, and significant relationship with age (Rho = 0.11, p = 0.002). This indicates that older participants in the sample reported higher susceptibility scores compared to younger participants. None of the other subscales showed significant associations.\n\n\nDiscussion\n\nIn this study, after validating a scale of risk perception of contracting the COVID-19 infection perceived in the Peruvian population, which was constructed based on the HBM, differences in perception (global scale and two factors) were described according to certain sociodemographic characteristics. This finding contributes to the growing literature on addressing risk perception of contracting the COVID-19 infection based on the HBM to understand its variability in the population.\n\nThe direct and positive correlation between age and perceived susceptibility to COVID-19 found in this study suggests that as individuals get older, they have more favorable beliefs regarding the likelihood of contracting coronavirus. Considering that studies comparing different age groups have found that the adoption of preventive behaviors is generally associated primarily with susceptibility and to a lesser extent with the perception of the impact of COVID-19, this finding is particularly interesting. It implies that older adults, with a higher perception of susceptibility, are more likely to adopt preventive measures. On the other hand, younger people having less favorable perceptions regarding the probability of getting infected could represent a risk group in terms of adopting preventive behaviors against COVID-19. Therefore, preventive interventions should be targeted towards these population groups.\n\nSimilarly, a study conducted during the first wave of infections in Argentina found that perceived severity was positively associated with age.12 Additionally, another study conducted in Italy found that the perception of susceptibility among older adults was higher compared to younger individuals.13 Conversely, a study that used the HBM in Canada and analyzed differences in the perception of personal impact of COVID-19 across age groups found that older adults, compared to younger individuals, had greater concerns about being hospitalized or dying (severity), but not about the risk of infection despite having higher susceptibility.14 Furthermore, another study conducted in the United States showed that older individuals perceive themselves as less susceptible to getting sick, but are more likely to experience severe consequences if they do contract the disease.15 Even though the literature has reported an association between age and different perceptions of risk related to COVID-19,16–18 the contribution of this study confirms that susceptibility to the possibility of contracting COVID-19 increases with age.\n\nAdditionally, differences in perceived susceptibility to COVID-19 were found based on occupation. Specifically, employed individuals had higher mean scores compared to retirees and homemakers. This finding is consistent with the results of a study conducted in Iran, where they reported that the mean score of risk perception of contracting the COVID-19 infection among employed individuals was up to seven times lower than that of homemakers and retirees.19\n\nAs observed in all the presented tables, the mean values related to perceived barriers are higher than those of the other dimensions. This result can be interpreted as a general difficulty for study participants in finding behaviors and actions in the face of the possibility of contamination. Possible income and salary reductions, fear of job loss, and limited transportation alternatives without crowds could be examples of events contributing to greater resistance to adopting behaviors that reduce exposure to the virus. Furthermore, the weak to moderate correlation between perceived barriers and perceptions of benefits and susceptibility indicates that even when individuals are aware of the benefits and conditions that make them more susceptible, the lack of options to modify their work and living conditions is evident in the population.\n\nAdditionally, it was found that participants who had multiple previous COVID-19 infections had a higher perceived susceptibility score for COVID-19 compared to those with a single infection or those who reported not being infected with the virus. Similarly, the risk perception of university students in China whose family members or friends had been exposed to confirmed or suspected COVID-19 patients was higher than those who had not been exposed.20 Furthermore, another study conducted in ten countries reported that direct experience is a predictor of COVID-19 risk perception.8 One possible reason could be that awareness of the ineffectiveness of the immune system in preventing the disease and the experience of being diagnosed with COVID-19 lead to an increased susceptibility perception of COVID-19.\n\nOn the other hand, regarding the perception of benefits, the sociodemographic characteristic in which differences were found was the level of education. Specifically, individuals with primary education perceive greater efficacy of the mechanisms adopted to prevent COVID-19 infection compared to those who completed secondary or higher education. This could be explained by research conducted in Peru, which reported that a higher level of education among household heads was associated with higher scores of myths and inappropriate beliefs. It is worth noting that in that study, the most frequent myths were “spraying alcohol or chlorine kills the virus” and “home remedies can cure or prevent coronavirus”.21 However, in a study conducted in four cities in Latin America, older adults with higher academic degrees showed better adherence to self-care measures.22\n\nRegarding the perception of risky behaviors for COVID-19 transmission, it was found that participants who had multiple infected family members had higher scores than those who had no infected family members. One explanation for this finding could be that people have a higher perception of risk for COVID-19 when they perceive themselves as more personally vulnerable to infection or when they perceive the pandemic as more severe.23\n\nWhen comparing the mean perception scores in each of the dimensions obtained in this study conducted in the Peruvian population, it can be observed that in four out of the five dimensions, the score is lower compared to the study conducted in Brazil at the beginning of the pandemic.9 This indicates that as the population becomes more familiar with the coronavirus, due to the mere exposure effect, their perception of risk decreases.24 Additionally, the start of COVID-19 vaccination generated rejection and uncertainty25 because the population perceived a lower susceptibility risk and therefore had less motivation to adopt other preventive measures.\n\nOverall, the findings of this study, in line with the literature, highlight the impact of sociodemographic variables on susceptibility perception and perception of benefits. This aspect can be useful in targeting public health interventions aimed at calibrating risk perception in the population to promote compliance with preventive measures, as preventive behavior is only evident when the event is perceived as highly contagious or dangerous.23 While risk perception influences the adoption of preventive behaviors during a pandemic,20,26,27 especially the affective dimension, this construct alone is not sufficient to promote the adoption of these behaviors.24 Therefore, to reduce the incidence of the disease, it is necessary to consider the sociodemographic characteristics of the population, complemented by the issuance of prevention guidelines, and thus ensure adequate vaccination coverage.28,29\n\nAnother implication of this study stems from the psychometric analysis conducted, which demonstrated that the scale developed in Brazil to measure the perception of risk behaviors for COVID-19 contagion based on the health belief model9 exhibits adequate psychometric properties in the Peruvian population. This suggests that this scale could be used in other Latin American countries with a previous a validation process.\n\nThe first limitation is that data collection was based on self-reporting, which could increase the likelihood of common method variance. However, these results represent an initial approach to the evaluated phenomenon and should be verified using complementary methodologies to control this effect. The second limitation is related to the convenience sampling method employed, which allowed quick data collection and access to remote areas of Peru during the pandemic but diminished the representativeness of the sample. The third limitation is the unequal composition of the sample according to sociodemographic characteristics, which could impact the detection of differences. Nevertheless, the statistical tests employed enabled the identification of statistically significant differences. Despite these limitations, this study reports findings from residents of different cities in Peru.\n\n\nConclusion\n\nThis study found that perceived susceptibility to COVID-19 correlates with age, occupation, and having a history of COVID-19 infection. The perceived benefits of adopting preventive measures for this disease are associated with educational level. Furthermore, the perception of risk of coronavirus contagion is linked to the history of infection with this viral disease. Additionally, the psychometric properties of a scale measuring the perception of risk behaviors related to COVID-19 in the Peruvian population, developed under the HBM, have been confirmed.", "appendix": "Data availability\n\nFigshare: Risk perception dataset English.xlsx. https://doi.org/10.6084/m9.figshare.23703939.v1. 30\n\nFigshare: Questionnaire on risk perception of contracting COVID-19 in a Peruvian population. https://doi.org/10.6084/m9.figshare.23669154.v1. 31\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nWe would like to thank Prof. Juana Cuba, Prof. José Chiroque, and Rubén Burga for their collaboration in data collection.\n\n\nReferences\n\nEpidemic Diseases - Cumulative suspected and confirmed COVID-19 cases reported by countries and territories in the Americas.[cited May 22th 2023]. Reference Source\n\nPuspitasari IM, Yusuf L, Sinuraya RK, et al.: Knowledge, Attitude, and Practice During the COVID-19 Pandemic: A Review. J. Multidiscip. Healthc. 2020; 13: 727–733. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOmotoso OE, Omotoso EF, Paimo KO, et al.: Knowledge and Adherence to COVID-19 Preventive Measures: A Continental Review. Sudan J. Med Sci. 2021; 16(3): 371–385. Publisher Full Text\n\nCameron LD, Fleszar-Pavlović S, Khachikian T: Changing Behavior Using the Common-Sense Model of Self-Regulation.Hamilton K, Cameron LD, Hagger MS, et al., editors. The Handbook of Behavior Change. Cambridge: Cambridge University Press; 2020 [cited May 22th 2023]; pp. 60–76. (Cambridge Handbooks in Psychology). Publisher Full Text Reference Source\n\nZajonc RB: Attitudinal effects of mere exposure. J. Pers. Soc. Psychol. 1968; 9: 1–27. Publisher Full Text\n\nBerkowitz L, editor. Advances in Experimental Social Psychology. Academic Press; 1977 [cited May 22th 2023]; pp. 39–83. Reference Source\n\nAlegria KE, Fleszar-Pavlović SE, Ngo DD, et al.: The Role of Risk Perceptions and Affective Consequences in COVID-19 Protective Behaviors. Int. J. Behav. Med. 2021; 28(6): 801–807. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDryhurst S, Schneider CR, Kerr J, et al.: Risk perceptions of COVID-19 around the world. J. Risk Res. 2020; 23(7-8): 994–1006. Publisher Full Text\n\nCosta MF: Modelo de crença em saúde para determinantes de risco para contaminação por coronavírus. Rev Saúde Pública. 2020; 54.\n\nHong B, Bonczak BJ, Gupta A, et al.: Exposure density and neighborhood disparities in COVID-19 infection risk. Proc. Natl. Acad. Sci. 2021; 118(13): e2021258118. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nR Core Team: R: The R Project for Statistical Computing.2021 [cited February 9th 2022]. Reference Source\n\nCuesta LS, Tumas N, Berra S: Percepción de riesgo ante el coronavirus en la primera fase de la pandemia en Argentina. Hacia la Promoción de la Salud. 2021; 26(1): 163–178. Publisher Full Text\n\nCommodari E, La Rosa VL, Coniglio MA: Health risk perceptions in the era of the new coronavirus: are the Italian people ready for a novel virus? A cross-sectional study on perceived personal and comparative susceptibility for infectious diseases. Public Health. 2020; 187: 8–14. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBechard LE, Bergelt M, Neudorf B, et al.: Using the Health Belief Model to Understand Age Differences in Perceptions and Responses to the COVID-19 Pandemic. Front. Psychol. 2021; 12: 609893. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGarfin DR, Fischhoff B, Holman EA, et al.: Risk perceptions and health behaviors as COVID-19 emerged in the United States: Results from a probability-based nationally representative sample. J. Exp. Psychol. Appl. 2021; 27(4): 584–598. Publisher Full Text\n\nFernández-Castillo E, Fernández-Fleites Z, Broche-Pérez Y, et al.: The Risk Perception COVID-19 Scale (RP-COVID19-S): Initial Validation and Its Relationship with Gender and Age in a Cuban Population Sample. Int. J. Ment. Heal. Addict. 2021; 21: 1466–1486. Publisher Full Text\n\nBruine de Bruin W: Age Differences in COVID-19 Risk Perceptions and Mental Health: Evidence From a National U.S. Survey Conducted in March 2020. J. Gerontol. B Psychol. Sci. Soc. Sci. 2021; 76(2): e24–e29. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIorfa SK, Ottu IFA, Oguntayo R, et al.: COVID-19 Knowledge, Risk Perception, and Precautionary Behavior Among Nigerians: A Moderated Mediation Approach. Front. Psychol. 2020; 11: 566773. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nArefi MF, Babaei AP, Barzanouni S, et al.: Risk Perception in the COVID-19 pandemic; a health promotion approach. J. Educ. Health Promot. 2022; 11: 118.\n\nDing Y, Du X, Li Q, et al.: Risk perception of coronavirus disease 2019 (COVID-19) and its related factors among college students in China during quarantine. PLoS One. 2020; 15 (8): e0237626. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCano-Gómez LC, Castillo-Tejada RD, Mena-Ordoñez SS, et al.: Percepción de riesgo, automedicación, mitos y creencias relacionados con COVID-19 entre jefes de hogar peruanos. Revista de la Universidad Industrial de Santander Salud. 2022 [cited May 22th 2023]; 54. Publisher Full Text Reference Source\n\nLiberona DF: Comunicación del riesgo en Latinoamérica: una evaluación de su impacto frente a la pandemia del COVID-19.2021 [cited May 22th 2023]. Reference Source\n\nPérez de Celis-Herrero M d l C, Cavazos Arroyo J: Percepción del riesgo de COVID-19 y medidas preventivas en México. Rev. Med. Inst. Mex. Seguro Soc. 2021 [cited May 22th 2023]; 59(5). Reference Source\n\nSavadori L, Lauriola M: Risk perceptions and COVID-19 protective behaviors: A two-wave longitudinal study of epidemic and post-epidemic periods. Soc. Sci. Med. 2022; 301: 114949. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSchaffer DeRoo S, Pudalov NJ, Fu LY: Planning for a COVID-19 Vaccination Program. JAMA. 2020; 323(24): 2458–2459. Publisher Full Text\n\nZeladita Huaman JA, Arcaya Moncada MJ, Zegarra Chapoñan R, et al.: Factors associated with the use of the N95 respirator in university students in the daily life of COVID-19. Rev. Bras. Enferm. 2022; 75: e20210412–e20210412. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nQin H, Sanders C, Prasetyo Y, et al.: Exploring the dynamic relationships between risk perception and behavior in response to the Coronavirus Disease 2019 (COVID-19) outbreak. Soc. Sci. Med. 2021; 285: 114267. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDönges P, Wagner J, Contreras S, et al.: Interplay between risk perception, behaviour, and COVID-19 spread. Front. Phys. 2022; 10: 842180. Publisher Full Text\n\nZeladita-Huaman JA, Franco-Chalco E, Zegarra-Chapoñan R, et al.: Development and validation of a COVID-19 risk perception scale in Peru. Rev. Peru. Med. Exp. Salud Publica. 2023; 40(2): 1–9.\n\nFranco E, Lozada-Urbano M, Zeladita-Huaman JA, et al.: Risk perception dataset English.xlsx. [Dataset]. figshare. 2023. Publisher Full Text\n\nZeladita-Huaman JA, Esteban Espinoza D, Lozada-Urbano M: Questionnaire on risk perception of contracting COVID-19 in a Peruvian population. figshare. Preprint.2023. Publisher Full Text" }	[ { "id": "193526", "date": "22 Aug 2023", "name": "María de la Concepción Pérez de Celis-Herrero", "expertise": [ "Reviewer Expertise Public Health and Computer Science" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis article analyzes the perception of the risk of contracting COVID-19 in Peru taking into account the socioeconomic characteristics of the respondents. For the authors, the analysis of the factors associated with the perception of contagion risk is an important psychological contribution to understanding the behavior of the population during this and future pandemics and will allow the design of proposals to reduce risk behaviors and promote the adoption of preventive measures in the Peruvian population.\nTo carry out the study, an online questionnaire of our own design with 24 questions based on the Health Belief Model (HBM) was used, but this questionnaire is not included in the article, only the five factors that were evaluated are mentioned: a) perceived susceptibility, corresponding to knowledge and belief about the possibility of contracting coronavirus; b) perceived severity, asking about personal beliefs about how the individual would experience the disease process and the intensity of the symptoms; c) perceived benefits, corresponding to the effectiveness of the behavioral mechanisms adopted to prevent infection; d) perceived barriers, with the aim of understanding the difficulties in adhering to coronavirus transmission protection and prevention measures, and e) health motivation, seeking to improve overall health. Reason why I consider that this study cannot be reproduced by other working groups. The questionnaire was applied in four cities in Peru from October to December 2021. The sample consisted of 821 people aged 18 and over.\nThe authors conclude that perceived susceptibility to COVID-19 is correlated with age, occupation, and having a history of COVID-19 infection. The perceived benefits of adopting preventive measures for this disease are associated with the educational level. In addition, the perception of risk of contagion from coronavirus is linked to a history of infection by this viral disease.\nIt should be noted that at the time the study was carried out in Peru, vaccination against covid19 had already begun (start date February 9, 2021), starting with health personnel, older and vulnerable groups, continuing with the rest of the groups. of age to reach most of the target population of 18 years and over during the year 2021[1]. And if we add to this that in October 2021 PAHO[2] reported that the rates of infection of COVID-19 in the region of the Americas was at the lowest levels in 2021, we can assume that the results found could be influenced by both facts, which is why I would assume that the conclusions are partially supported by the results.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nI cannot comment. A qualified statistician is required.\n\nAre all the source data underlying the results available to ensure full reproducibility? No source data required\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "193525", "date": "08 Sep 2023", "name": "Evelyn Fernández-Castillo", "expertise": [ "Reviewer Expertise Psychology", "addiction prevention", "mental health", "disease prevention", "health promotion", "validation of psychological evaluation instruments" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe manuscript approaches a current topic. The results presented contribute to refining health interventions for future emergency situations.\n\nIt is considered that the introduction should:\n\nDeepen the definition of risk perception addressed in the research.\n\nHighlight the relationship between the definition assumed and the Health Belief Model and its importance for this research.\nIn the methodological section and analysis of results:\n\nIt is stated that its uses a non-probabilistic sampling by convenience. And in the description of the statistical analyses performed, it is stated that \"In all cases, the assumptions of parametric analyses were verified\", however, in the results section, the results that allow verifying the fulfillment of these assumptions are not made explicit. I suggest incorporating this into the manuscript.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-919
https://f1000research.com/articles/11-964/v1	19 Aug 22	{ "type": "Brief Report", "title": "Building a portfolio for not-for-profit activities rather than maximizing social return on investment ratio", "authors": [ "Fuminobu Mizutani" ], "abstract": "Background: An influential piece of literature on effective altruism insists that not-for-profit organizations (NFPs) should concentrate their investments on a few activities to maximize their social return on investment (SROI) ratio. However, this creates greater risk for an NFP than building a portfolio of investments in activities. This study investigates whether it is desirable for executives and contributors of NFPs to build a portfolio rather than maximize the expected SROI ratio, and if so, how to build one. Solving these questions will help the chief financial officers (CFOs) of NFPs, who serve as their agents, fulfill their obligations to contributors, who are their principals, and will help advisors provide better services for their contributors, their clients. Methods: Data were collected from a ranking of NFPs, then non-parametric tests were performed on this ranking and the Herfindahl-Hirschman Index (HHI). Results: The HHI are between 2013 and 2688. The results of non-parametric tests do not deny that rank and HHI are independent of each other. Most of the NFPs’ investments in activities were in accord with their core competencies. Conclusions: It was found that successful executives build portfolios. The findings of this study should be sufficiently practical in helping NFP executives and contributors decide whether to build portfolios, and if so, how.", "keywords": [ "Portfolio", "NFP", "SROI", "HHI", "Non-parametric test", "Core competency", "Financial Engineering", "Accounting" ], "content": "Introduction\n\nSocial return on investment (SROI) is a hot topic in accounting; in practice, a not-for-profit organization (NFP) can calculate its SROI ratio annually, the way a profit-oriented entity calculates return on equity (ROE).\n\nThe Wall Street Journal (Feb. 2, 2022) stated that Melinda French Gates will not concentrate her donations within the Bill & Melinda Gates Foundation, but instead spread her donations across various philanthropic endeavors. We know from Tuan (2008) that this foundation has been interested in measuring SROI and other forms of social impact.\n\nThe frequently quoted MacAskill (2015) insists that NFPs should concentrate on a few activities and suggests that Oxfam and WorldVision are involved in too wide a variety of activities. Moreover, some scholars use fictional examples in which contributors concentrate their donations within one NFP with the highest SROI ratio, which would maximize the expected SROI ratio; however, some NFPs unfortunately fail to achieve their goals, and some even become embroiled in scandal, meaning that each NFP is accompanied with risk for its executives and contributors.\n\nBuilding a portfolio, an often-discussed topic in finance, may fulfill the needs of NFP executives and contributors who are risk-averse. The Markowitz model, also called the Modern Portfolio Theory (MPT), advises investors of profit-oriented entities against concentrating investments within only one profit-oriented entity, and this may provide insight on how contributors to NFPs should donate.\n\nMany finance textbooks, including Brealey et al. (2020), discuss agency theory. The executives of an NFP are the agents of its contributors, who are its principals. If governance of an NFP is effective, its agents fulfill its principals’ needs. If building a portfolio were desirable for principals, agents would also build a portfolio. If decisions to concentrate are desirable for contributors, they would also be desirable for NFP executives.\n\nThis paper intends to help investment professionals in decision-making, including chief financial officers (CFOs) of NFPs and advisors for contributors to NFPs, who are considering building a portfolio. A review of the literature is provided in the next section, then the data and adopted methodology, which are non-parametric tests, are described in the section after that. This is followed by results, then a discussion of the results, and finally, conclusions.\n\nGneezy et al. (2014) is a widely read paper on NFP financial ratios that discusses not only how to calculate them, but also how to use them. Gargani (2017) is an influential and well-written paper that also discusses risk, written by a scholar with a professional background. This paper found that SROI ratio can be calculated using the formula below:\n\nB means net social impact and C means costs, while T means the number of years of an NFP project and t is the particular financial year.\n\nGargani uses a model in which only the NFP with the highest SROI ratio is selected. Yates and Marra (2017) indicate the weak points of SROI and warn that SROI may lead to radical concentration of donations in the real world.\n\nMacAskill (2015) is a typical and practical example of literature on effective altruism based on rationality. The basis of SROI is utilitarianism, as Maier et al. (2015) states, and utilitarianism is a philosophical thought based on rationality. Some supporters of effective altruism are utilitarian. Singer (2015) in particular is clearly based on utilitarianism and is also widely read. MacAskill is not written from the viewpoint of an extreme risk-lover, meaning that supporters of SROI cannot neglect this paper.\n\nOn the other hand, scholars such as Pahlevani et al. (2021) have considered a portfolio of donations. Luenberger (2014) is a widely read textbook about financial engineering that contains explanations of the Markowitz model, a model created by Nobel laureate Harry Markowitz that has become the standard for scholars. The Markowitz model is suited to risk-averse investors, whose optimal portfolio is one that is not too concentrated.\n\nLukomnik and Hawley (2021) depict the Markowitz model as a bloodless model that cannot tackle social problems, implying that the Markowitz model is harmful for stakeholders. Their emphasis on social problems themselves is agreed upon by most investors and contributors, as governments around the world aim to achieve the United Nations’ Sustainable Development Goals (SDGs) with the cooperation of profit-oriented entities. However, it is doubtful that the Markowitz model is really bloodless.\n\nRegarding methodology, Clark et al. (2022) show that simple methodologies sometimes bring interesting findings on economics. Flori et al. (2019) is an influential paper stating that the Herfindahl-Hirschman index (HHI) can be used to quantify concentration of investments. Basu et al. (2009), a somewhat older study, used non-parametric tests extensively, showing that accounting researchers can adopt not only parametric tests but non-parametric tests as well.\n\nIt is difficult to foresee the results of statistical methods, because of the difference between NFPs and profit-oriented entities. Angerer et al. (2015) shows that higher risk tolerance among contributors relates to higher contributions. Some contributors may be even risk-lovers. In contrast, mainstream investors in profit-oriented entities are obviously risk-averse.\n\n\nMethods\n\nDue to a small sample size that makes postulating Gaussian distribution impossible, this study uses a simple methodology which utilizes non-parametric tests, which can be used for small sample sizes and do not require Gaussian distribution.\n\nSample NFPs were collected from NGO Advisor’s “World 200 Best SGOs” for 2021. This is a leading ranking of NFPs. NGO Advisor is a Swiss organization that provides a wealth of information on NFPs in English and French and sometimes uses the word SGO (social good organization). This abbreviation was perhaps influenced by the word association in French.\n\nWhile the characteristics of each NFP often differ, Mercy Corps, Oxfam, and Save the Children rank high on the list and have similar characteristics. Thus, this study sampled NFPs whose activities are similar to these three NFPs, are concerned with SROI, and whose headquarters are within the English-speaking world. If detailed financial information on an NFP was difficult to collect from its annual report, it was excluded from this study.\n\nNFPs ranked in the World 200 Best SGOs are excellent in the aspect of social impact, and executives who use SROI in such NFPs are assumed to carefully consider how to invest donations received into their activities.\n\nEach NFP conducts several activities, and the HHI of these activities can be calculated from financial reporting. Numbers showing each NFP’s investments into each segment were gathered. This study assumes that disclosed data on activities is similar to segment reporting by profit-oriented entities, which NFP executives use in their decision-making. The objective financial information used was as the list in the next section.\n\nData from before the coronavirus disease (COVID-19) pandemic was used in order for this study to be applicable to a wider range of situations than the special circumstances of the pandemic. The time lag between financial information and the world ranking is not an issue for this study. Whether an NFP conducted emergency relief for the pandemic does not affect how investments in activities were made in the past, and thus it is expected that the world ranking is based on investments before the pandemic.\n\nThis study calculates the HHI of each NFP in order to see how each NFP distributes the donations received among their activities. The percentage invested in each activity within the NFP was used for this calculation. If HHI was low, it can be assumed that building a portfolio rather than concentrating donations in one area is a preferable strategy for NFPs.\n\nThis study uses Spearman’s rank correlation coefficient, which is widely used among scholars, to calculate correlation between HHI and the rank of the NFP on the world ranking. A statistically significant correlation (p < 0.05) would indicate that these high-ranking NFPs have not built an optimal portfolio of donations. This study also calculates Kendall’s rank correlation coefficient, which is not widely used among scholars, as a supplement because it is known that these two statistical methods sometimes show different results.\n\nIBM SPSS Statistics 27, a reliable application widely used in social sciences, was used to calculate these two statistical methods.\n\n\nResults\n\nSix NFPs met the requirements of this study, meaning that the sample size was 6. Data were collected from the below documents:\n\n• 2019 Annual Impact Report by Mercy Corps (The left schedule of page 14.)\n\n• Annual Report 2018-19 by Oxfam (The right schedule of page 47.)\n\n• Save the Children Annual Report 2019 by Save the Children (The upper right graph of page 23.)\n\n• CARE USA 2019 Annual Report by CARE (The graph named “How We Work” of page 31.)\n\n• 2020 Annual Report by ChildFund (The right financial statement of page 11.)\n\n• Annual Report and Accounts 2018/2019 by Voluntary Service Overseas (VSO) (The upper graph of page 38.)\n\nThe name, rank, and rounded HHI of each NFP are shown in Table 1.\n\nThe HHI of the six NFPs ranged from 2013 to 2688. There were no NFPs that met the requirements of the study below rank 148 in this ranking.\n\nA scatter chart is shown in Figure 1. At a glance, there is no significant correlation between rank and HHI.\n\nSpearman’s rank correlation coefficient was ρ = -0.314, statistically not significant with not only p ≥ 0.05, but p ≥ 0.1. Kendall’s rank correlation coefficient was Tau-b = -0.2, also statistically not significant at p ≥ 0.1. Neither of these results denies that rank and HHI are independent of each other.\n\nAdditionally, most of the six NFPs’ investments in activities were in accord with their core competencies. Even the NFP that made the largest investments in activities not in accord with its core competence among the six NFPs invested more than two-thirds in activities that were in accord with its core competence.\n\n\nDiscussion\n\nFirst, all six NFPs showed low HHI, showing that executives of these NFPs have built portfolios. Second, because all six NFPs had similar HHI, it would seem at a glance that an optimal portfolio exists. Statistical analysis does not deny the existence of an optimal portfolio. Finally, executives of all six NFPs appeared to prefer an optimal portfolio that consisted of activities in accord with the NFPs’ core competencies.\n\nThis suggests that skillful and rational executives build a portfolio, and that an optimal portfolio of investment into activities and an optimal portfolio of donations exist; if NFP executives neglect their core competencies, their SROI ratio will be low.\n\n\nConclusions\n\nThese findings suggest that skillful and rational NFP executives build a portfolio of investments based on their activities. Similarly, from the statistical analysis, it can be assumed that building a portfolio of donations would benefit contributors as well. There also appears to be an optimal portfolio of donations. This study also suggests that core competencies should not be neglected, and that the Markowitz model is likely not bloodless.\n\nThis paper suggests that investment professionals should avoid overly concentrating investments, but instead build a portfolio for risk-averse clients, who make up the majority of clients. CFOs in NFPs, who serve as their agents, will be able to better fulfill their obligations to contributors, their principals, than in the past, and advisors will be able to provide better services for their clients, who are NFP contributors.\n\nOne of the limitations of this study is its small sample size. The findings of this study could be checked more rigorously if there were a method to increase the sample size. Another limitation is the widely known limits of statistical methods, which are suitable for finding differences but are not as suitable for finding similarities. The well-known statistical methods can only show that something is not denied, a slightly ambiguous indication. A method suited to finding similarities may be preferable.\n\nEven with the limitations above, the findings of this study should be sufficiently practical in helping NFP executives and contributors decide whether to build a portfolio and, if so, how.\n\n\nData availability\n\nData was collected from a webpage of a third party “NGO Advisor”. Because NGO Advisor changed its name to thedotgood in 2022, readers can access the data from the website of thedotgood. Its URL is https://thedotgood.net/. The author utilized “World 200 Best SGOs” for 2021. In order to access the full data, registration is required. The cost for purchasing a pass varies depending on the type of the pass. The latest ranking is for 2022 in July 2022. If a reader wants to view the ranking not for 2022 but for 2021, they need to send an inquiry to thedotgood.", "appendix": "Acknowledgment\n\nThe author would like to thank b-cause Inc. for its English proofreading service.\n\n\nReferences\n\nAngerer S, Glätzle-Rützler D, Lergetporer P, et al.: Donations, Risk Attitudes and Time Preferences: A Study on Altruism in Primary School Children. J. Econ. Behav. Organ. 2015; 115: 67–74. Publisher Full Text\n\nBasu S, Dickhaut J, Hecht G, et al.: Recordkeeping Alters Economic History by Promoting Reciprocity. Proc. Natl. Acad. Sci. 2009; 106(4): 1009–1014. PubMed Abstract \| Publisher Full Text\n\nBrealey RA, Myers SC, Allen F: Principles of Corporate Finance. 13th ed.McGraw-Hill Education;2020.\n\nClark AE, D’Ambrosio C, Onur I, et al.: COVID-19 Compliance Behaviors of Older People: The Role of Cognitive and Non-Cognitive Skills. Econ. Lett. 2022; 210: 110158. PubMed Abstract \| Publisher Full Text\n\nFlori A, Pammolli F, Buldyrev SV, et al.: Communities and Regularities in the Behavior of Investment Fund Managers. Proc. Natl. Acad. Sci. 2019; 116(14): 6569–6574. PubMed Abstract \| Publisher Full Text\n\nGargani J: The Leap from ROI to SROI: Farther than Expected? Eval. Program Plann. 2017; 64: 116–126. PubMed Abstract \| Publisher Full Text\n\nGneezy U, Keenan EA, Gneezy A: Avoiding Overhead Aversion in Charity. Science. 2014; 346: 632–635. Publisher Full Text\n\nLuenberger DG: Investment Science. Second ed.Oxford University Press;2014.\n\nLukomnik J, Hawley JP: Moving Beyond Modern Portfolio Theory: Investing That Matters. Routledge;2021.\n\nMacAskill W: Doing Good Better: How Effective Altruism Can Help You Make a Difference. Guardian Books;2015.\n\nMaier F, Shober C, Simsa R, et al.: SROI as a Method for Evaluation Research: Understanding Merits and Limitations. VOLUNTAS: International Journal of Voluntary and Nonprofit Organization. 2015; 26(5): 1805–1830. Publisher Full Text\n\nNGO Advisor: World 200 Best SGOs. [data set].2021.Reference Source\n\nPahlevani M, Choi T, Heydar J, et al.: Cooperative Donation Programs in Supply Chains with Non-Governmental Organizations (NGOs). IEEE Trans. Eng. Manag. 2021; 1–12. Early Access.\n\nSinger P: The Most Good You Can Do: How Effective Altruism is Changing Ideas about Living Ethically. Yale University Press;2015.\n\nTuan MT: Measuring and/or Estimating Social Value Creation: Integrated into Eight Integrated Cost Approaches. [Report].2008.Reference Source\n\nGlazer E: Melinda French Gates No Longer Pledges Bulk of her Wealth to Gates Foundation. Wall Street J. 2022, Feb. 2.\n\nYates BT, Marra M: Social Return on Investment (SROI): Problems, Solutions … and is SROI a Good Investment? Eval. Program Plann. 2017; 64: 136–144. PubMed Abstract \| Publisher Full Text" }	[ { "id": "148079", "date": "01 Sep 2022", "name": "Marco Bellucci", "expertise": [ "Reviewer Expertise Accounting" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThanks for the opportunity to read and review this study that investigates \"whether it is desirable for executives and contributors of NFPs to build a portfolio rather than maximize the expected SROI ratio\". While the topic is relevant and interesting, I believe the present version of this study would benefit from several revisions to the literature review, research design and discussion of contributions. The subject of the paper is more than relevant nowadays, but, in my view, the study also presents an important set of shortcomings that hampers, in this present version, its potential to contribute to the literature.\nFrom a constructive perspective, the authors could consider the following suggestions.\nI have several concerns about the study's premises. The calculation of an SROI ratio and the construction of a portfolio are not incompatible avenues, and they are not alternatives to each other. This is especially true if we consider the usefulness of SROI as a planning and control tool. The paper is highly focused on the calculation of the SROI ratio and the selection of projects with a higher value. Still, in this way, it oversees how an organization can use the process behind the SROI methodology to foster social impact.\n\nThe literature review is scarce and ignores recent contributions on this topic. Moreover, the SROI methodology, the Markowitz model, and the HHI are not properly presented nor framed in the context of NFPs.\n\nIt is not completely clear how the author built the sample for this study, and the author should address how the criteria behind the report on the \"200 Best SGOs\" could affect his results. Moreover, the paper uses many terms (NGOs, NFPs, SGOs, associations) that refer to different organizations in the third sector. Furthermore, the study is based on six organizations, and this may not be enough to provide significant findings.\n\nThe author sometimes refers to \"an optimal portfolio of donations\". For whom? For the donor organization? For the beneficiaries? For the society at large? I fear this utilitarian perspective may neglect the true significance of social impact. I think the author should better clarify the contribution of this study in terms of how it can improve the selection of projects to be financed and, thus, their social impact.\n\nIn this version, the author does not clearly explain how he calculated the HHI and how this financial measure of market concentration can be useful to explain the choice of an \"optimal portfolio\" for nonprofit donors.\n\nThe absence of a theoretical framework led to a very brief and underwhelming discussion of results.\n\nThe author writes that \"These findings suggest that skillful and rational NFP executives build a portfolio of investments based on their activities. Similarly, from the statistical analysis, it can be assumed that building a portfolio of donations would benefit contributors as well\". However, in the present version, I do not see how the results can support this conclusion; moreover, it is difficult to know the contribution to the literature on SROI and social impact evaluation.\n\nIn general, I believe that author would need to consider deeply revising their study, particularly concerning the theoretical and managerial contribution and the research premises. I do realize that I have been critical throughout this review, but I also truly hope this feedback can help the authors to reconsider their conceptual approach to this interesting research strand.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate? Partly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [ { "c_id": "8723", "date": "06 Sep 2022", "name": "Fuminobu Mizutani", "role": "Author Response", "response": "Thank you very much for your report. I would like to revise my manuscript after receiving other reviewers' comments." }, { "c_id": "10426", "date": "16 Nov 2023", "name": "Fuminobu Mizutani", "role": "Author Response", "response": "Thank you very much for your comments. Your comments were helpful for the revision. 1. This paper’s focus moved from SROI to social impact in general. 2. The section of literature review was revised to explain SROI, Markowitz model, and HHI in the context of NFPs. 3. The direct explanation of sampling is in paragraph 3 of “Methods” and the sampling of part two made the sampling criteria clearer. 4. If the limitation is necessary, for contributors who are principles as explained in “Introduction”. 5. HHI was calculated by a spreadsheet as written in “Methods”. Why optimal was explained through Buchak (2023). 6. Some sentences were added to “Discussion”. 7. Additional data were collected and another statistical analysis were conducted." } ] }, { "id": "178726", "date": "03 Aug 2023", "name": "Linda M. Parsons", "expertise": [ "Reviewer Expertise Nonprofit accounting research using archival reports and disclosures." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe author examines whether a common claim that concentrating investments (rather than building a portfolio of activities) leads to better social return on investment (SROI). This is an interesting question and, as is often the case in the nonprofit sector, involves considering the appropriateness of using corporate (profit-maximizing) techniques in nonprofit organizations. In this example, financial theory suggests a diversified portfolio is less risky and more profitable than a concentrated one. However, given the focus on SROI, perhaps dedicating nonprofit resources and expertise to a specific activity is more effective.\nThe primary question/concern is the measures employed. Rather than directly measuring social impact, the author uses the ranking from a reputable international source. The assumption is that ranking of the 200 best social good organizations is an appropriate proxy for (and highly correlated with) social impact. If this is the case, a sentence or two to acknowledge this assumption and the logic of doing so would be helpful. Are there other factors (such as organizational growth or governance) that significantly affect rankings?\nThe author calculates the correlation of ranking (as a proxy for impact) with a widely-accepted measure of portfolio diversity, the Herfindahl-Hirschman Index (HHI). The results show no significant association between HHI and ranking of six large nonprofits. However, later the author states that there is little variation in the HHI of the sample organizations. The lack of variation implies that both high- and lower-ranked organizations have similar investment diversity. The conclusion is that diversifying does not appear to lead to a high or low ranking, and that results “suggest that investment professionals should avoid overly concentrating investments.” I don’t think the results allow this conclusion. You show that the organizations in your sample (which are all in the top 200 internationally) do, in fact, diversify, and that this diversity is similar for those at the top of the ranking and those further down the list. What you cannot observe is whether concentrating investments would improve rankings for those at the bottom or hurt the rankings of those at the top. A more appropriate comparison (to reach your conclusion) would be a look at organizations that are not in the top 200 to see if their investments are more or less concentrated.\nI think the author can address these questions/concerns with a little more explanation of the choice of measures.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate? Yes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10030", "date": "16 Nov 2023", "name": "Fuminobu Mizutani", "role": "Author Response", "response": "Thank you very much for your valuable reviewing. I will revise this manuscript as soon as possible." }, { "c_id": "10427", "date": "16 Nov 2023", "name": "Fuminobu Mizutani", "role": "Author Response", "response": "Thank you very much for your comments. Your comments were helpful for the revision. Paragraph 1 This paper’s title was changed. This paper’s focus moved from SROI to social impact in general. Paragraph 2 More explanations of the data source were added in “methods”. Paragraph3 Additional data were collected and another statistical analysis was conducted." } ] } ]	1	https://f1000research.com/articles/11-964
https://f1000research.com/articles/11-1075/v1	21 Sep 22	{ "type": "Research Article", "title": "Identification of prognostic biomarkers of invasive ductal carcinoma by an integrated bioinformatics approach", "authors": [ "Albeiro Marrugo-Padilla", "Johana Márquez-Lázaro", "Antistio Álviz-Amador", "Johana Márquez-Lázaro", "Antistio Álviz-Amador" ], "abstract": "Background: Invasive ductal carcinoma (IDC) is the most common breast cancer worldwide. Nowadays, due to IDC heterogeneity and its high capacity for metastasis, it is necessary to discover novel diagnostic and prognostic biomarkers. Thus, this study aimed to identify new prognostic genes of IDC using an integrated bioinformatics approach. Methods: Using the Gene Expression Omnibus (GEO) database, we downloaded publicly available data of the whole-genome mRNA expression profile from the first three stages of IDC in two expression profiling datasets, GSE29044 and GSE32291; intra-group data repeatability tests were conducted using Pearson’s correlation test, and the differentially expressed genes (DEGs) were identified using the online tool GEO2R, followed by the construction of a protein‑protein interaction network (PPI-net) with the common DEGs identified in the three analyzed stages using the Search Tool for the Retrieval of Interacting Genes (STRING) database and Cytoscape software, from these PPI-net we identify the hub genes (prognostic genes). Results: We found seven genes [WW domain-containing E3 ubiquitin-protein ligase 1 (WWP1), STIP1 homology and U-box containing protein 1 (STUB1), F-box and WD repeat domain containing 7 (FBXW7), kelch like family member 13 (KLHL13), ubiquitin-conjugating enzyme E2 Q1 (UBE2Q1), tripartite motif-containing 11 (TRIM11), and the beta-transducin repeat containing E3 ubiquitin-protein ligase (BTRC)] as potential candidates for IDC prognostic biomarkers, which were mainly enriched in the Ubiquitin-specific protease activity, cytoskeletal protein binding, and ligase activity. The role of these genes in the pathophysiology of IDC is not yet well characterized, representing a way to improve our understanding of the process of tumorigenesis and the underlying molecular events of IDC. Conclusions: Genes identified may lead to the discovery of new prognostic targets and precise therapeutics for IDC.", "keywords": [ "Invasive ductal carcinoma", "prognostic biomarkers", "hub genes", "microarray technology", "differentially expressed genes", "GEO", "GEO2R" ], "content": "Introduction\n\nBreast cancer (BC) is the most prevalent diagnosed neoplasm in women worldwide and one of the most important causes of death among them.1,2 According to The Global Cancer Observatory, in 2020, there were more than 2.3 million new cases worldwide. On the other hand, BC deaths are reported more frequently in countries such as Melanesia, Western Africa, Micronesia/Polynesia, and the Caribbean.3\n\nBC has been categorized into two major histological types, invasive ductal carcinoma (IDC) and invasive lobular carcinoma (ILC),4 with IDC being the most common (80%)5; this neoplasm begins in the cells lining a milk duct in the breast, from there, the cancer cells invade the wall of the duct, and grows into nearby breast tissues. At this point, it may have the ability to spread (metastasize) to other parts of the body through the lymphatic system and bloodstream.6\n\nThe clinicopathological characteristics and differences between IDC and ILC and BC prognosis have been well described.6 Identified prognostic factors of BC have been crucial in the diagnostic markers, workup, and treatment of this pathology7; these include the hormone receptors [estrogen receptor (ER)/progesterone receptor (PR) positive], human epidermal growth factor receptor 2 (HER2/neu), and germline mutations in BReast CAncer gene 1 (BRCA1) or BReast CAncer gene 2 (BRCA2), which are associated with an increased risk of BC incidence.5 Prognostic markers are also helpful in determining the effectiveness of the established intervention (surgical or pharmacological treatment), the probability of recurrence, and the establishment of additional follow-up and treatment strategies.8\n\nDespite efforts to identify biomarkers for BC, due to its heterogeneity and its high capacity for metastasis, an increasing percentage of patients are demanding personalized treatments,9 which makes it necessary for the discovery of novel biomarkers for diagnosis and prognosis that allow for an early evaluation of the development of the pathology to formulate effective diagnosis and treatment strategies.10–12\n\nNowadays, the analysis of gene expression profiles [verification of differentially expressed genes (DEGs)] using bioinformatics tools has represented a notable advance in research in clinical oncology aimed at the identification of genes related to tumors, new molecular markers of diagnosis and prognosis, and evaluation of therapeutic effects among others.13 DEGs and protein-protein interaction network (PPI-net) analysis have been widely used to identify biomarkers and potential drug targets. Open access databases such as Gene Expression Omnibus (GEO) are broadly employed as microarray resources for this purpose.14\n\nPrevious studies have identified prognostic genes from ductal carcinoma in situ (DCIS), such as Fibroblast growth factor 2 (FGF2), Growth arrest-specific protein 1 (GAS1), and Secreted frizzled-related protein 1 (SFRP1) using GEO15; however, currently, IDC is little understood from the genomic point of view, and there are no studies from the analysis of expression of genes using bioinformatic methods. Thus, this study aimed to identify new prognostic genes of this type of BC using an integrated bioinformatics approach.\n\n\nMethods\n\nThe two expression profiling datasets from BC, GSE29044 and GSE632291, were downloaded from GEO (RRID:SCR_005012), which were based on the platforms [GPL570 (HG-U133_Plus_2 - Affymetrix Human Genome U133 Plus 2.0 Array)] and [GPL4091 (Agilent-014693 Human Genome CGH Microarray 244A (Feature number version)], respectively. GSE29044 was a collection that analyzed the whole-genome mRNA expression profile from 73 patients with tumors and 36 adjacent disease-free tissues using the Affymetrix GeneChip Human Genome U133 Plus 2.0 Arrays.16 On the other hand, GSE32291 was analyzed using whole-genome CGH arrays from Agilent 394 invasive ductal breast carcinomas and 20 normal breast biopsies.\n\nThe inclusion criteria used for the selection of the samples were that they were both ER and PR positive and a wild type of strain. The MeSH (RRID:SCR_004750) terms used for the selection of the datasets were (“carcinoma, ductal, breast” [MeSH Terms] OR invasive ductal carcinoma [All Fields]) AND “Homo sapiens”[porgn]).\n\nTo verify the intra-group data repeatability per each group of datasets, as proposed by Xu et al. (2019), we developed a Pearson’s correlation test using the R programming language, R Project for Statistical Computing (RRID:SCR_001905). The degree of correlations between all samples from the same dataset was visualized by heat maps built in R.17\n\nDEGs in the first three stages of the IDC were obtained by online analysis in GEO2R (RRID:SCR_016569), which is an interactive online tool from GEO that finds DEGs through the comparison of the original submitter-supplied processed data tables using the GEOquery (RRID:SCR_000146) and limma R packages from the Bioconductor project.18–20 Initially, the experimental groups were built from the datasets, grouping the samples as tissues with IDC and controls (adjacent disease-free tissues). A comparative analysis was carried out for each IDC stage evaluated.1–3 The cut-off criterion was P < 0.05 and a fold-change among ≥ 1.5 or ≤ 1.5. Volcano plots with the DEGs found were drawn in GEO2R. An intersection analysis between DEGs extracted from the three stages assayed was made by drawing Venn diagrams, delineated in the functional enrichment analysis tool (FunRich).17,21\n\nA PPI-net was built with the DEGs product of the intersection analysis between the three IDC stages evaluated, using the online Search Tool for the Retrieval of Interacting Genes [STRING (RRID:SCR_005223)], thus identifying the prognostic candidate genes (hub genes). Next, through the software Cytoscape (RRID:SCR_003032) (version 3.8.0),22 the PPI-net was visualized; on the other hand, the MCODE App (RRID:SCR_015828) (Molecular complex detection tool; version 1.6.1) was used to identify the most important module of the network.23 The criteria for MCODE analysis were a degree of cut-off of 2, scores >5, a maximum depth equal to 100, a node score cut-off of 0.2, and a k-score of 2. Genes with degrees ≥10 were selected as hub genes.17,19\n\nAfter the identification of the main module of the network, the top 10 central genes were evaluated through the cytoHubba (RRID:SCR_017677) application of Cytoscape,24 using the five most reported calculation methods: Degree, EcCentricity, closeness, Maximum Neighborhood Component (MNC), and Maximal Clique Centrality (MCC).25 An intersection analysis was performed between the hub genes identified for each method in a virtual tool VennDiagram image GP.26 Finally, a functional enrichment analysis of the hub genes identified in FunRich was performed, and through the Kyoto Encyclopedia of Genes and Genomes (KEGG) (RRID:SCR_012773) we analyzed the enrichment around the molecular function.27,28\n\nWe assayed the expression patterns of hub genes between different stages of IDC based on Gene Expression Profiling Interactive Analysis (GEPIA) (RRID:SCR_018294), a web server for cancer and normal gene expression profiling and interactive analyses.29\n\nAll analyses were conducted in GraphPad Prism (RRID:SCR_002798) (version 8.0.2) [free alternative, JASP (RRID:SCR_015823) (version 16.3)] and RStudio (RRID:SCR_000432). One-way analysis of variance (ANOVA) was used for comparing the mean between groups in the analyses conducted in GEPIA. P<0.05 was considered to indicate a statistically significant difference.\n\n\nResults\n\nR script for GSE29044 and GSE632291 can be found as Underlying data.62–67 Pearson’s correlation coefficient showed that both datasets (GSE29044 and GSE32291) had a strong correlation among the samples from the control group and IDC (Supplementary Figure S1, which can be found as Extended data70). Next, we classified the samples of the datasets per stage,1–3 and, through GEO2R, a volcano plot analysis was performed to identify the DEGs in the three stages assayed. Nodes that conformed to the cut-off criterion (fold-change ≥1.5 or ≤-1.5, and a P<0.05) were represented in blue or red color; the first represented downregulated DEGs and the red the upregulated DEGs in IDC samples, regarding the controls (Figure 1a).68 An intersection analysis in FunRich was made with the DEGs from each dataset per stage, and those genes were used to find the common DEGs in the three stages assayed; we found 1,085, 3,213, and 3,477 common DEGs in stages 1, 2, and 3, respectively (Supplementary Figure S2, which can be found as Extended data71). We also found 724 common DEGs in the three stages (Figure 1b) (P<0.05).\n\na) Volcano plots obtained in GEO2R show the difference in gene expression between tissues of IDC and controls. The X and Y-axis represent the fold-change and the P-value (log-scaled). Each symbol represents a different gene; red and blue symbols represent upregulated and downregulated genes. b) Venn diagram showing the shared genes per stages assayed. DEG, differentially expressed gene; IDC, invasive ductal carcinoma.\n\nFrom 724 common DEGs, a PPI network was built in STRING using the following parameters: medium confidence of > 0.4 (minimum required interaction score) and that the network will only display the query proteins; Supplementary Figure S3, which can be found as Extended data,72 described the network features. Next, we identified the most significant PPI-net module by the MCODE app from Cytoscape, which had 73 edges, 17 nodes, and a score of 9.125 (Figure 2a); from it and using the five most reported calculation methods of cytoHubba (Degree, EcCentricity, MCC, MNC, and Closeness) we identified seven hub genes by intersection analysis [WW domain-containing E3 ubiquitin-protein ligase 1 (WWP1), STIP1 homology and U-box containing protein 1 (STUB1), F-box and WD repeat domain containing 7 (FBXW7), kelch like family member 13 (KLHL13), ubiquitin-conjugating enzyme E2 Q1 (UBE2Q1), tripartite motif-containing 11 (TRIM11), and the beta-transducin repeat containing E3 ubiquitin-protein ligase (BTRC)] (Figure 2b, 2c and Table 1),69 which are described in Table 2. All of those genes were upregulated in patients with IDC regarding the controls.\n\na) Central cluster of the PPI-net built from the 724 common DEGs (MCODE score: 9.125). b) Identification of hub genes (IDC prediction) by cytoHubba algorithms (Degree, EcCentricity, MCC, MNC, and Closeness) in the central cluster of the network. c) Intersection analysis of the genes identified in each algorithm. DEG, differentially expressed gene; PPI, protein-protein interaction; IDC, invasive ductal carcinoma; MCC, Maximal Clique Centrality; MNC, Maximum Neighborhood Component.\n\nWW domain-containing E3 ubiquitin-protein ligase 1 (WWP1), STIP1 homology and U-box containing protein 1 (STUB1), F-box and WD repeat domain containing 7 (FBXW7), kelch like family member 13 (KLHL13), ubiquitin-conjugating enzyme E2 Q1 (UBE2Q1), tripartite motif-containing 11 (TRIM11), beta-transducin repeat containing E3 ubiquitin-protein ligase (BTRC), ubiquitin conjugating enzyme E2 H (UBE2H), thyroid hormone receptor interactor 12 (TRIP12), ankyrin repeat and SOCS box containing 1 (ASB1), epidermal growth factor receptor (EGFR), fibronectin 1 (FN1), insulin-like growth factor 1 (IGF1), Maximum Neighborhood Component (MNC), Maximal Clique Centrality (MCC), protein-protein interaction (PPI).\n\nEnrichment analysis of hub genes was developed in FunRich, classifying them by their ‘biological process’, ‘molecular function’, ‘cellular components’, and the ‘Catalogue of Somatic Mutations in Cancer (COSMIC)’ (RRID:SCR_002260). The results obtained showed that hub genes were mainly enriched in “protein metabolism”, “metabolism”, “cell growth” and “energy pathways”; in turn, among the main molecular functions, analyzed by KEGG pathways showed that hub genes were mainly enriched in “Ubiquitin-specific protease activity”, “cytoskeletal protein binding”, and “ligase activity”; these were associated with the main cellular components where genes are found (“nucleoplasm”, “ubiquitin ligase complex”, “SCF ubiquitin ligase complex”, “ubiquitin conjugating enzyme complex” and “nuclear inclusion body”). Finally, according to COSMIC, the “breast”, “endometrium”, “stomach”, “bone”, and “soft tissue” were the primary site of action of the hub genes (Figure 3).27,28\n\nThe figure shows the enrichment percentages in terms of ‘biological process’, ‘molecular function’, ‘cellular component’, and the ‘Catalog of Somatic Mutations in Cancer (COSMIC)’. The enrichment analysis was performed in FunRich.\n\nThe analysis developed in GEPIA is shown in Figures 4 and 5; this evidenced no statistical differences in expression patterns of hub genes in different stages of IDC. The concentrations of the genes remain constant throughout the evolutionary process of the disease, which could denote an important prognostic factor (Figure 4). On the other hand, high expression of BTRC, FBXW7, and WPP1 was related to the low percentage of survival of the patients (Figure 5).\n\nIDC, invasive ductal carcinoma; GEPIA, Gene Expression Profiling Interactive Analysis; WWP1, WW domain-containing E3 ubiquitin-protein ligase 1; STUB1, STIP1 homology and U-box containing protein 1; FBXW7, F-box and WD repeat domain containing 7; KLHL13, kelch like family member 13; UBE2Q1, ubiquitin-conjugating enzyme E2 Q1; TRIM11, tripartite motif-containing 11; BTRC, beta-transducin repeat containing E3 ubiquitin-protein ligase.\n\nIDC, invasive ductal carcinoma; GEPIA, Gene Expression Profiling Interactive Analysis; WWP1, WW domain-containing E3 ubiquitin-protein ligase 1; STUB1, STIP1 homology and U-box containing protein 1; FBXW7, F-box and WD repeat domain containing 7; KLHL13, kelch like family member 13; UBE2Q1, ubiquitin-conjugating enzyme E2 Q1; TRIM11, tripartite motif-containing 11; BTRC, beta-transducin repeat containing E3 ubiquitin-protein ligase.\n\n\nDiscussion\n\nIDC is a nonspecific invasive carcinoma that belongs to epithelial tumors. This cancer is considered extremely malignant and the main cause of death in women.30,31 Thus, the search for molecular markers that allow its detection in the early stages is necessary for the diagnosis, early treatment, and prognosis of patients. In this sense, several bioinformatics techniques were integrated into this study, whose objective was to investigate data to screen and identify hub genes related to IDC. Two datasets, GSE29044 and GSE32291, were screened for IDC. Seven gene hubs in common were discovered (WWP1, STUB1, FBXW7, KLHL13, UBE2Q1, TRIM11, and BTRC) (Table 2).\n\nThe WWP1 encodes the WW domain-containing E3 ubiquitin-protein ligase 1 protein, a HECT domain-containing E3 ligase regulating apoptosis,32 which has been associated with colorectal, osteosarcomas, oral, gastric, melanoma, prostate, hepatocellular, and BC.33 In BC, WWP1 is frequently amplified and overexpressed34,35; also, the overexpression of WWWP1 in colorectal cancer and BC has been associated with the worst prognosis and poor survival in patients.36 The expression of WWP1 in breast tumors correlated with the positive ER (ERα) status, inducing breast cell growth. On the other hand, WWP1 depletion in ERα positive BC cell lines suppressed cell proliferation and induced apoptosis.32 Despite the above, the role of WWP1 in IDC is not yet well studied. In this context, Chen et al. (2009) reported that WWP1 is overexpressed in a cell line of IDC and was associated with ER and IGF-1R proteins.37,38 On the other hand, Zhou et al. (2012) demonstrated that WWP1 depletion by small interfering (si) RNA activated the extrinsic apoptotic pathway increasing the TRAIL-induced caspase-8 recruitment.32 Although WWP1 has not been associated with any stage of BC, the results of this study show its importance in IDC.\n\nThe STUB1 encodes the carboxyl-terminus of HSP70-interacting protein (CHIP); this is a co-chaperone protein that interacts with Hsp70 and negatively regulates chaperone functions. In oral, lung, and colorectal cancer, STUB1 seems to have an important role in the progression of cancer.39,40 According to Lui et al. (2020), the lower expression of STUB1 in oral cancer can be linked to a poorer prognosis.39 Xu et al. (2011) reported that overexpression of STUB1 (CHIP expression) glioma cell lines is associated with the histological grade of the tumor.41 In BC, Kajiro et al. (2009) reported that knockdown of CHIP in breast cancer cells resulted in rapid tumor growth and metastatic phenotypes,42 while Hiyoshi et al. (2014) found that promoting the expression of CHIP can inhibit cell growth and metastatic potential of BC cells.43 Also, Wei et al. (2021) indicated that overexpression of Tripartite motif-containing protein 6 (TRIM6) promoted the degradation of STUB1, which facilitated the growth and migration of malignant cells.44 According to the aforementioned, STUB1 could participate in different ways in the progression of cancer, however, in BC, it may be acting as a tumor suppressor. Regarding the IDC, there is no available evidence associating the presence of the STUB1 with this type of BC.\n\nThe TRIM11 encodes the Tripartite motif-containing 11 protein, identified as an oncogene in colon, hepatocellular, and lung cancer. However, its role in BC cells remains unclear. In this sense, Song et al. (2019) reported that TRIM was overexpressed in BC tissues, which was linked to the metabolism of glycolysis.45 Also, Tang et al. (2020) found that the protein level of TRIM11 is highly correlated with ERα, and its depletion significantly decreases the cell proliferation and migration of BC cells.46 As was described above, TRIM6 (member TRIM family), was associated with the degradation of STUB1 on BC cells; thus, we hypothesized that in IDC, these genes could be related metabolically. However, there is no available evidence linking TRIM 11 to invasive ductal cancer.\n\nThe FBXW7 encodes the F-box protein family members and has a role important in cell cycle regulation, transcriptional regulation, apoptosis, and cell signal transduction.47,48 The FBXW7 in triple-negative BC (TNBC) has been related to the suppression of proliferation and invasion of TNBC cells. Wu et al. (2020) reported that the inhibition of the TLR4/NF-κB pathway could increase the BXW7 expression, which suppresses the proliferation and invasion of TNBC cells,49 while Singh et al. (2020) demonstrated that downregulation of FBXW7 in a mouse model increased the tumorigenesis and metastasis in TNBC cells.50 Also, Wang et al. (2022) reported that microRNA (miR)-223-3p decreases the expression of FBXW7, which promotes the invasion and metastasis of BC cells.51 According to the studies described above, FBXW7 could have an important role as a suppressor of tumors in BC. Though, its function in IDC has not yet been studied.\n\nThe KLHL13 encodes Kelch-like proteins (KLHLs), which act as substrate adaptors of Cullin3-RING ligases (CRL3). CRL3 regulates the degradation of proteins that function as tumor suppressors, which participate in tumor development.52 In this context, Xiang et al. (2021) reported that the upregulation of KLHL proteins contributes to the progression of lung cancer through binding with CRL3, which showed that KLHL13 could be considered a potential target therapeutic.53 However, the involvement of this gene in BC and IDC has not been studied.\n\nThe UBE2Q1 encodes ubiquitin-conjugating enzyme E2 Q1 (UBE2Q1), identified as upregulated in human breast and colorectal cancer.54,55 Shafiee et al. (2015) showed that UBE2Q1 in BC cell lines was overexpressed. Also, these authors found that UBE2Q1 could be interacting with p53 through a complex, which explains its involvement in the proliferation and migration of tumor cells.56 Also, Topno et al. (2021) identified UBE2Q1 as a potential prognostic marker in high-grade serous ovarian cancer, using an integrated gene expression analysis and gene co-expression network analysis (WGCNA).57 Nevertheless, the involvement of UBE2Q1 in IDC remains unstudied at present.\n\nThe BTRC encodes F-box protein, which has been associated with colorectal, glioma, esophageal, and BC. In this sense, Zheng et al. (2020) reported that the inhibition of BTRC by miR-224 in colorectal cancer promotes cell migration and invasion. The miR-224 silencing promoted the overexpression of BTRC, which decreased the cell progression,58 while Zhou et al. (2021) found that the invasion and migration cells induced by miR-193a-3p in patients with glioma could be reversed by overexpression of BTRC.59 Zhang et al. (2018) indicated that BTRC activity mediated by upregulated tetraspanin 15 (TSPAN15) in esophageal cancer promotes the degradation of phosphorylated (p-)IκBα and triggers NF-κB nuclear translocation and subsequent activation of transcription of several metastasis-related genes [intercellular adhesion molecule 1 (ICAM1, vascular cell adhesion molecule 1 (VCAM1), urokinase-type plasminogen activator (uPA), matrix metallopeptidase 9 (MMP9), tumor necrosis factor α (TNFα), and C-C motif chemokine ligand 2 (CCL2)].60 Lim et al. (2022) found that BTRC acts as an oncogene in TNBC through NF-κB activation (IκBα ubiquitination).61 As described above, the function of BTCR oncogene/gene suppressor could be dependent on cancer type. Therefore, BTRC, like the other identified genes in this study, could be considered a potential therapeutic target and biomarker in IDC.\n\n\nData availability\n\nCode with which R was fed for the preliminary analysis of the data (Intra-group reproducibility):\n\nFigshare: R-Script_GESE32291_STAGE 1. https://doi.org/10.6084/m9.figshare.2041911962\n\nFigshare: R-Script_GESE32291_STAGE 2. https://doi.org/10.6084/m9.figshare.2041916463\n\nFigshare: R-Script_GESE32291_STAGE 3. https://doi.org/10.6084/m9.figshare.2041916764\n\nFigshare: R-Script_GESE29044_STAGE 1. https://doi.org/10.6084/m9.figshare.2041917065\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\nFigshare: R-Script_GESE29044_STAGE 2. https://doi.org/10.6084/m9.figshare.2041917666\n\nFigshare: R-Script_GESE29044_STAGE 3. https://doi.org/10.6084/m9.figshare.2041917967\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nRaw data derived from differential expression analysis performed in GEO2R for each IDC stage in the datasets:\n\nFigshare: Raw data derived from differential expression analysis performed in GEO2R for each IDC stage in the datasets. https://doi.org/10.6084/m9.figshare.2041920668\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\nData derived from the analysis in Cytohubba from Cytoscape:\n\nFigshare: Data derived from the analysis in Cytohubba from Cytoscape. https://doi.org/10.6084/m9.figshare.2041921869\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nFigshare: Supplementary Figure S1. https://doi.org/10.6084/m9.figshare.2029384270\n\nFigshare: Supplementary Figure S2. https://doi.org/10.6084/m9.figshare.2029384571\n\nFigshare: Supplementary Figure S3. https://doi.org/10.6084/m9.figshare.2029384872\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).", "appendix": "References\n\nZangouri V, Akrami M, Tahmasebi S, et al.: Medullary breast carcinoma and invasive ductal carcinoma: A review study. 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PubMed Abstract \| Publisher Full Text\n\nWu X, Chen H, Wu M, et al.: Downregulation of miR-182-5p inhibits the proliferation and invasion of triple-negative breast cancer cells through regulating TLR4/NF-κB pathway activity by targeting FBXW7. Ann. Transl. Med. 2020; 8(16): 995–995. PubMed Abstract \| Publisher Full Text\n\nSingh S, Kumar S, Srivastava RK, et al.: Loss of ELF5–FBXW7 stabilizes IFNGR1 to promote the growth and metastasis of triple-negative breast cancer through interferon-γ signalling. Nat. Cell Biol. 2020; 22(5): 591–602. PubMed Abstract \| Publisher Full Text\n\nWang Y, Shi S, Wang Y, et al.: miR-223-3p targets FBXW7 to promote epithelial-mesenchymal transition and metastasis in breast cancer. Thorac. Cancer. 2022; 13(3): 474–482. PubMed Abstract \| Publisher Full Text\n\nCheng J, Guo J, Wang Z, et al.: Functional analysis of Cullin 3 E3 ligases in tumorigenesis. Biochim. Biophys. Acta Rev. Cancer 2018; 1869(1): 11–28. 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PubMed Abstract \| Publisher Full Text\n\nTopno R, Singh I, Kumar M, et al.: Integrated bioinformatic analysis identifies UBE2Q1 as a potential prognostic marker for high grade serous ovarian cancer. BMC Cancer 2021; 21(1): 220. PubMed Abstract \| Publisher Full Text\n\nZheng Q, Yu JJ, Li C, et al.: miR-224 targets BTRC and promotes cell migration and invasion in colorectal cancer. Biotech 2020; 10(11): 485.\n\nZhou DD, Li HL, Liu W, et al.: miR-193a-3p Promotes the Invasion, Migration, and Mesenchymal Transition in Glioma through Regulating BTRC. Biomed. Res. Int. 2021; 2021: 1–22. Publisher Full Text\n\nZhang B, Zhang Z, Li L, et al.: TSPAN15 interacts with BTRC to promote oesophageal squamous cell carcinoma metastasis via activating NF-κB signaling. Nat. Commun. 2018; 9(1). Publisher Full Text\n\nLim YX, Lin H, Chu T, et al.: WBP2 promotes BTRC mRNA stability to drive migration and invasion in triple-negative breast cancer via NF-κB activation. Mol. Oncol. 2022; 16(2): 422–446. PubMed Abstract \| Publisher Full Text\n\nMarrugo Padilla A: R-Script_GESE32291_STAGE 1. figshare. [Dataset].2022. Publisher Full Text\n\nMarrugo Padilla A: R-Script_GESE32291_STAGE 2. figshare. [Dataset].2022. Publisher Full Text\n\nMarrugo Padilla A: R-Script_GESE32291_STAGE 3. figshare. [Dataset].2022. Publisher Full Text\n\nMarrugo Padilla A: R-Script_GESE29044_STAGE 1. figshare. [Dataset].2022. Publisher Full Text\n\nMarrugo Padilla A: R-Script_GESE29044_STAGE 2. figshare. [Dataset].2022. Publisher Full Text\n\nMarrugo Padilla A: R-Script_GESE29044_STAGE 3. figshare. [Dataset].2022. Publisher Full Text\n\nMarrugo Padilla A: Raw data derived from differential expression analysis performed in GEO2R for each IDC stage in the datasets. figshare. [Dataset].2022. Publisher Full Text\n\nMarrugo Padilla A: Data derived from the analysis in Cytohubba from Cytoscape. figshare. [Dataset].2022. Publisher Full Text\n\nMarrugo Padilla A: Supplementary Figure S1. figshare. [Dataset].2022. Publisher Full Text\n\nMarrugo Padilla A: Supplementary Figure S2. figshare. [Dataset]2022. Publisher Full Text\n\nMarrugo Padilla A: Supplementary Figure S3. figshare. [Dataset].2022. Publisher Full Text" }	[ { "id": "153740", "date": "24 Oct 2022", "name": "Xingxin Pan", "expertise": [ "Reviewer Expertise Bioinformatics", "computational biology", "machine learning", "genome", "and genetics" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors analyzed public IDC datasets and identified differentially expressed genes between IDC and control samples. After constructing a protein-protein interaction network, they found seven genes and thought these genes may serve as prognostic targets for treating IDC.\n\nThe review of related work is not sufficiently thorough and not sufficiently specific, especially for IDC. The authors should cite the latest references if appropriate.\n\nGSE632291 is wrong, I couldn’t find it in GEO.\n\nIt’s better for the authors to summarize the clinical information for the samples they used. When identifying DEGs, clinical factors such as gender are important effectors to consider.\n\n“a fold-change among ≥ 1.5 or ≤ 1.5” is not correct.\n\nWhat is the quantification measure the author analyzed for expression analysis?\n\nCould the authors provide the lists of DEGs they identified?\n\nWhen it comes to the Venn diagram, did the authors take account of up-regulated and down-regulated respectively, or take it as a whole?\n\nThe paragraph: “WW domain-containing E3 ubiquitin-protein ligase 1 (WWP1)…protein-protein interaction (PPI).” looks odd. This paragraph makes no sense, the authors should double-check the texts.\n\nThe result of Figure 5 seems not significantly different, the conclusion the authors got from Figure 5 didn't get statistical support.\n\nIt’s better to take into account up-regulated and down-regulated DEGs in PPI-net; consider whether the DEGs are up-regulated or down-regulated when constructing PPI-net.\n\nI suggest that the Discussion section should be improved to better reflect the quality of the work. There are many limitations to this research work. The authors should reflect on the quality of their work and conclude their findings and compare them with other relevant works.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10397", "date": "30 Nov 2023", "name": "Albeiro Marrugo Padilla", "role": "Author Response", "response": "Thank you very much for your comments and notes, which were crucial to the work's development. Below are the responses to the sent queries. The review of related work is not sufficiently thorough and not sufficiently specific, especially for IDC. The authors should cite the latest references if appropriate. Thanks for the appreciation. Based on the comment, the introduction was restructured and nourished with current references. GSE632291 is wrong, I couldn’t find it in GEO. Thank you. It was a typo; the code was corrected in the document. It’s better for the authors to summarize the clinical information for the samples they used. When identifying DEGs, clinical factors such as gender are important effectors to consider. Correct. In response to the comment, the characteristics of the selected samples of the datasets are broken down in supplementary tables 1–3. “a fold-change among ≥ 1.5 or ≤ 1.5” is not correct. Thanks, the expression was corrected in the document. What is the quantification measure the author analyzed for expression analysis? That provided by the data collections consulted Could the authors provide the lists of DEGs they identified? The list of DEGs is shown in Supplementary Tables 4–7. When it comes to the Venn diagram, did the authors take account of up-regulated and down-regulated respectively, or take it as a whole? We take into account those regulated upwards and downwards; although they are analyzed separately, in the end, the network is built with the union of both. The paragraph: “WW domain-containing E3 ubiquitin-protein ligase 1 (WWP1)…protein-protein interaction (PPI).” looks odd. This paragraph makes no sense, the authors should double-check the texts. Based on the other referee's recommendations, this paragraph was eliminated. This gene was absent from the new hub gene listing when expanding the comparative analysis by adding a new collection. The result of Figure 5 seems not significantly different, the conclusion the authors got from Figure 5 didn't get statistical support. Thanks, indeed, this graph was edited. This comment was considered in accordance with the current results analysis. It’s better to take into account up-regulated and down-regulated DEGs in PPI-net; consider whether the DEGs are up-regulated or down-regulated when constructing PPI-net. Thank you very much. In fact, the design of PPI-net associates both DEGs categories. I suggest that the Discussion section should be improved to better reflect the quality of the work. There are many limitations to this research work. The authors should reflect on the quality of their work and conclude their findings and compare them with other relevant works. In response to your comment, the new discussion section had been significantly improved to provide a comparative analysis of the bibliographic reports that was more in-depth." } ] }, { "id": "153739", "date": "31 Oct 2022", "name": "Russell Hamilton", "expertise": [ "Reviewer Expertise Bioinformatics", "computational biology. Transcriptomics", "single-cell. Translational biology." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nMarrugo-Padilla et al. present a bioinformatics analysis of two previously published mRNA expression array datasets for invasive ductal carcinoma, the most common form of breast cancer worldwide. Through a differential expression analysis, followed by a protein-protein interaction network analysis, seven hub genes are identified as potential biomarkers for patient prognosis.\nMajor Points:\nThe introduction is lacking an in-depth review of the related literature.\n\nThere is very little mention of the patient metadata in the text despite being utilised in the analyses. How many patients are in each stage, age ranges, etc.? Perhaps a table could summarise this information?\n\nA combined analysis of two datasets was used to derive the seven genes, however, a more thorough approach would be to identify the genes in study 1 and see if they replicate in study 2; or, do the seven genes classify with IDC survival in a 3rd or 4th study (a quick search of GEO reveals 92 expression array datasets for IDC). There are several Bioconductor packages available to perform this type of analysis, e.g. geNetClassifer and genefu.\n\nThe discussion is limited to a paragraph for each of the seven identified hub genes. A final paragraph bringing together combined insight from all seven genes together is necessary to conclude the paper. For example, all seven genes appear to be linked to ubiquitination, but this is not discussed. This could be speculative, but then provide suggestions for how this analysis should be followed up before being utilised in the clinic.\n\nMore recent technologies such as single-cell sequencing may be able to reveal the cell type specificity of the seven hub genes in IDC patients. A discussion or ideally a query of publicly available single-cell datasets would put more weight behind the findings in this paper. Many single-cell datasets are available to search via web interfaces.\n\nMinor Points:\nCode, in the form of R-scripts, are provided via FigShare. It is commendable to provide code, however, a more appropriate resource such as GitHub or BitBucket would be more appropriate for others to easily download and replicate/extend this analysis.\n\nTypo GSE32291 is miss entered as GSE632291.\n\nA PCA plot of the intra-group variability would be preferable to a heatmap, coloured by stage and e.g. age for each of the studies.\n\nResults, first paragraph: “we classified” – are the samples not already classified in the GEO submission?\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10398", "date": "30 Nov 2023", "name": "Albeiro Marrugo Padilla", "role": "Author Response", "response": "Thank you very much for your comments and suggestions, which contributed significantly to improving the work's quality. Major Points: The introduction is lacking an in-depth review of the related literature. In response to this, the work's introduction was reorganized and expanded. There is very little mention of the patient metadata in the text despite being utilised in the analyses. How many patients are in each stage, age ranges, etc.? Perhaps a table could summarise this information? Sincere appreciation. In fact, three additional supplementary tables containing information about the experimental subjects utilized by the researchers in the development of the work were included. A combined analysis of two datasets was used to derive the seven genes, however, a more thorough approach would be to identify the genes in study 1 and see if they replicate in study 2; or, do the seven genes classify with IDC survival in a 3rd or 4th study (a quick search of GEO reveals 92 expression array datasets for IDC). There are several Bioconductor packages available to perform this type of analysis, e.g. geNetClassifer and genefu. Thank you greatly. In response to your request, the developed analysis is modified by adding a third collection, resulting in a comparison of three datasets and an important modification to the obtained results. The discussion is limited to a paragraph for each of the seven identified hub genes. A final paragraph bringing together combined insight from all seven genes together is necessary to conclude the paper. For example, all seven genes appear to be linked to ubiquitination, but this is not discussed. This could be speculative, but then provide suggestions for how this analysis should be followed up before being utilised in the clinic. Correct; The discussion surrounding the newly reported hub genes was substantially enhanced, and a comparative analysis of the signaling pathways associated with these was developed, as a result of the changes in the results obtained by modifying the study by adding a new data set. More recent technologies such as single-cell sequencing may be able to reveal the cell type specificity of the seven hub genes in IDC patients. A discussion or ideally a query of publicly available single-cell datasets would put more weight behind the findings in this paper. Many single-cell datasets are available to search via web interfaces. I express my gratitude for your assistance. In accordance with your inquiry, we have extended the scope of analysis for the novel hub genes by assessing their performance in GEPIA or by examining their overall survival in IDC using the Kaplan-Meier tracer. Subsequently, the significance of the hub genes was assessed by utilizing mRNA-seq data for BC sourced from the Cancer Genome Atlas (TCGA). Minor Points: Code, in the form of R-scripts, are provided via FigShare. It is commendable to provide code, however, a more appropriate resource such as GitHub or BitBucket would be more appropriate for others to easily download and replicate/extend this analysis. We attach the link to the GitHub repository in FigShare Typo GSE32291 is miss entered as GSE632291. Thank you. the code was corrected in the document. A PCA plot of the intra-group variability would be preferable to a heatmap, coloured by stage and e.g. age for each of the studies. A graph with the PCA for each collection was attached in the supplementary material. Results, first paragraph: “we classified” – are the samples not already classified in the GEO submission?" } ] } ]	1	https://f1000research.com/articles/11-1075
https://f1000research.com/articles/12-1418/v1	30 Oct 23	{ "type": "Case Report", "title": "Case Report: Rehabilitation of anterior esthetic region with ovate pontic", "authors": [ "Prasanna R. Sonar", "Aarati Panchbhai", "Ravina Khairkar", "Aarati Panchbhai", "Ravina Khairkar" ], "abstract": "The period of time between extraction and recovery following tooth loss in the esthetic zone can be awful to the patient's appearance and mental health. A suitable alternative to extraction and replacement of a maxillary anterior tooth is quick tooth replacement with an ovate pontic on a temporary fixed partial denture. The interdental papilla, which is preserved by ovate pontic, in turn protects the contour of gingiva that after extraction would have been lost. Utilizing an ovate pontic, an instantaneous tooth replacement removes the emotionally distressing partial edentulous stage, while also producing a considerably more aesthetically acceptable, hygienic, and natural-looking replacement tooth. Ovate pontics have the additional benefit of avoiding the dissatisfaction associated with an unattractive ridge lap pontic. The case study demonstrated an ovate pontic replacement for an upper anterior tooth that helped a patient whose mobile anterior tooth was advised for extraction and finally resulting to both improved esthetics and health.", "keywords": [ "aesthetic", "gingival contour", "ovate pontics" ], "content": "Introduction\n\nWhen an individual loses their anterior teeth, it has a profoundly negative impact on their ability to integrate into society as usual. Any patient who extracts or loses a single anterior tooth may find it difficult in social integration, but unless extensive bone and soft tissue loss is present along with the tooth, replacing the tooth with a fixed partial denture (FPD) or an implant yields an expected esthetic result.1 Even so, the outcome is typically acceptable when handled by a qualified practitioner. To succeed, many things must be taken into account. These characteristics include the pontic's size, form, color, and location, as well as its profile as it emerges from the soft tissues.2\n\nPontics that look natural and are hygienic have been the subject of many different designs over the years. We have the chance to fulfill both of the above objectives because of the ovate pontic's design. The ovate pontic, also known as the emergence profile, is a method used to give the appearance that the tooth is emerging from the gums. It can also aid to form and preserve the interdental papilla. The contour is a cleanability-enhancing design in ovate pontic.3\n\nThe ovate pontic is a method for giving the appearance that the tooth is poking through the gum.4 However, clinicians don't typically use an ovate pontic design on a regular basis. Preserving inter-proximal tissue following tooth removal is one of the difficult aspects of a dental treatment plan. In restorative dentistry, it is highly desirable to prevent alveolar bone collapse.\n\n\nCase report\n\nAn Asian female patient, 33 years old, reported to the dental hospital with complaint of a loosening of tooth in anterior region. Patient also wished to replace the mobile tooth with a firm tooth. There were not any previous medical interventions or treatments or any systemic history. On examination, right central incisor (11) was grade III mobile with supraeruption of the same tooth (Figure 1). On radiographic examination, orthopantomogram (OPG) revealed that there was severe bone loss in 11 region (Figure 1). A diagnostic impression recorded and treatment planning was done.\n\nThe appearance of the patient's maxillary anterior teeth bothered her. Patient was advised and recommended to remove the mobile tooth and replace it with an ovate pontic that, in terms of form, functionality, and appearance, exactly mimics the missing tooth.\n\nWith maxillary right lateral incisor (12) and maxillary left central incisor (21) teeth, final tooth preparation was completed [Figure 2A]. Then, the right central incisor (11) was extracted under local anesthesia with no complication [Figure 2B]. During extraction, considerable care was taken to avoid damaging the lingual and buccal plates. This process is essential to maintaining both the interdental papillae and the bone. The cast was scored around the tooth to be removed, creating a 3 mm depression to represent the extraction socket. Fabrication of a temporary restoration using luxatemp material was done [Figure 3]. On the prepared tooth, a temporary fixed partial denture (FPD) was placed with the pontic portion immersing into the extraction socket by 2-3 millimeters, resulting in a depression of 1-1.5 millimeters once the tissues receded throughout the healing phase. The required changes, including the placement of the ovate pontic, were made to enable the appropriate placing of the FPD. The tissue surface of the pontic was cleaned and polished to avoid irritating the socket's healing tissues and to stop the buildup of bacterial plaque. A temporary restoration substance was used to solidify the work [Figure 3].\n\nAfter extraction and temporary prosthesis, the patient was told to come back in 48 hours to have the temporary restoration removed and have the healing socket checked [Figure 4]. Following evaluation, the temporary restoration was restored using temporary luting material. To manipulate the soft tissues and reproduce the ovate pontic receptor location, a succession of temporary fixed partial denture was made. An elastomeric material was used to create an impression of the abutment tooth and the prepared soft tissue site after three to four months, until the soft tissues had developed and the extraction site had healed [Figure 5]. GIC was used to build and cement the ceramic-metal repair. After giving post-cementation instructions, the patient was asked to come back for routine dental follow-up. The patient was happy and satisfied with the prosthesis [Figure 6].\n\n\nDiscussion\n\nAbrams invented the ovate pontic in 1980.5 Both implant prostheses and fixed and removable partial dentures have been utilized effectively using the concept itself. Super floss can be used to keep the pontic's tissue surface clean in order to avoid any tissue inflammation. Although some have questioned its viability, the tissue surface of the pontic's convex form always allows for appropriate cleansing of the area beneath it.6 Counselling may take time, and some patients may not accept having teeth created close to missing teeth.\n\nFor a satisfactory marginal fit, careful attention to the preliminary repair is required. It is crucial to take final impressions for the FPD as soon as the temporary reconstruction is removed. If not, tissue may revert to its original state and result in an ovate pontic space on the model that is considerably smaller than the temporary pontic.7\n\nFor a good esthetic result in this case, the ovate pontic shape was intended to draw attention to the mucosa's position on the alveolar ridge by recreating a concave soft tissue outline. For directed papilla growth and stabilization, the tissues were molded.2\n\nOvate pontics have the following advantages when used in the anterior aesthetic zone:\n\n1. Maintenance of the normal gingival contour and the interdental papilla\n\n2. Gets rid of the emotionally distressing partial edentulous phase\n\n3. A substitute that is hygienic and aesthetically appealing and seems natural\n\n4. Discards the dissatisfaction brought on by an unappealing ridge lap pontic\n\n5. Gets rid of unsightly “black triangles”.2\n\nThe height of the apical pontic was chosen and selected by the tissue/bone complex already present, aesthetics, support, cleaning convenience, and prevention of food impaction. For the majority of pontic forms, passive ridge contact was recommended; however, the ovate pontic requires close contact in order to support, shape, and preserve tissue.8,9\n\n\nConclusion\n\nOvate pontic can meet the aesthetic, functional, and hygienic specifications of an artificial tooth in FPD. For individuals with a high smile line or for the replacement of missing teeth in the cosmetic zone, ovate pontics are advised. The pontic minimizes the black triangles while creating the impression of a free interdental papilla and gingival border. The patient's oral hygiene routine will determine if the prosthesis is ultimately successful.\n\n\nConsent\n\nThe patient was provided with an informed consent form and provided a thumb print signature to consent for the publication of their clinical details and images.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\n\nReferences\n\nSpear FM: The use of implants and ovate pontics in the esthetic zone. Compend Contin. Educ. Dent. 2008; 29(72-4): 76–80.\n\nEdelhoff D, Spiekermann H, Yildirim M: A review of esthetic pontic design options. Quintessence Int. 2002; 33: 736–746. PubMed Abstract\n\nGuruprasada LC: Creating natural gingival profiles of missing anterior teeth using ovate pontic. Medical Journal Armed Forces India. 2015; 71: S124–S126. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNoriega EB: Ovate Pontic: Natural Look. Active Member of the American Academy of Cosmetic Dentistry No. 001568.\n\nAbrams L: Augmentation of the deformed edentulous residual ridge for fixed prosthesis. Compend Contin Educ. Gen. Dent. 1980 MayeJun; 1(3): 205e213.\n\nJohnson GK, Leary JM: Pontic design and localized ridge augmentation in fixed partial denture design. Dent. Clin. North Am. 1992 Jul; 36(3): 591–605. Publisher Full Text\n\nDylina TJ, Merced.: Contour determination for ovate pontics calif. J. Prosthet. Dent. 1999; 82: 136–142. Publisher Full Text\n\nTarantola GJ, Becker IM: Definitive diagnostic waxing with light-cured composite resin. J. Prosthet. Dent. 1993; 70: 315–319. PubMed Abstract \| Publisher Full Text\n\nMhatre S, Gala A, Ram SM, et al.: Modified ovate pontic design for immediate anterior tooth replacement. J. Contemp. Dent. 2012 May; 2(2): 64–68. Publisher Full Text" }	[ { "id": "279412", "date": "06 Jun 2024", "name": "Matheel AL-Rawas", "expertise": [ "Reviewer Expertise Prosthodontics", "Dental materials", "Maxillofacial prosthodontics", "tooth wear and oral rehabilitation" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe case study demonstrated an ovate pontic replacement for an upper anterior tooth. The language used is not clear and needs a lot of improvements. The abstract, intro and discussion is weak and need more information and citations. The report quality is unsatisfactory in various aspects. The value of this one case report is thus unclear to me.\nAs anterior esthetic case with low smile line which don't show the ovate pontic part when the patient smile, so, I am not sure why this was made from the beginning. There was a possibility for more conservative option such as zirconia resin bonded bridge. Did the author provide treatment options to the patient?.\nThere was no full examination details of this esthetic case such smile line, smile design and as well pocket depth of the abutments and dynamic occlusion info such protrusion, excursion details.\nIn conclusion, the case report is very simple to be indexed.\n\nIs the background of the case’s history and progression described in sufficient detail? Partly\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Partly\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? No\n\nIs the case presented with sufficient detail to be useful for other practitioners? No", "responses": [] } ]	1	https://f1000research.com/articles/12-1418
https://f1000research.com/articles/11-1186/v1	17 Oct 22	{ "type": "Research Article", "title": "Potential impact of artificial intelligence on the emerging world order", "authors": [ "Anupama Vijayakumar" ], "abstract": "The fast-paced diffusion of technologies broadly falling under the umbrella of artificial intelligence (AI) is said to be shaping the emerging world order in international relations (IR). It is expected that the global AI race will pave the way for another rise and fall of great powers in the international system, similar to the impact caused by the three industrial revolutions of the past. The literature in IR identifies three major powers – namely, the United States of America (USA), China, and Russia, as the leading contenders in this AI race. The ongoing AI-enabled fourth industrial revolution is all the more unique due to the markedly different approaches these three powers have adopted for integrating AI into their military, political, and socio-economic spheres. The AI strategies of these countries further reflect their capabilities and intentions towards how they plan on employing the technology to elevate their prestige and power status in the international system. This paper draws from a historiography of the First, Second, and Third Industrial Revolutions to study how technological innovations have altered relative power capabilities of nations, triggering a re-ordering of power hierarchies at a systemic level. Drawing from this understanding, it analyses the nature of AI as an emerging technology and assesses whether it can cause systemic alterations. It critically examines and compares the AI strategies of the USA, China, and Russia as leading contenders in the global AI race and outlines their strengths and weaknesses. It further draws from the Adoption Capacity Theory to argue that the AI race may well be determined by the relative capacity of the major institutions in each of these countries to manage and adapt to the disruptions this technology is bound to bring to the fore.", "keywords": [ "AI", "Technological Diffusion", "World Order", "Great Power Competition", "Fourth Industrial Revolution", "Emerging Technologies", "Technological Innovation", "Great Power Status" ], "content": "Introduction\n\nThe world is currently witnessing the Fourth Industrial Revolution (Schwaab, 2015; Rotatori, Lee, & Sleeva, 2021; Amaresh, 2022). The advent of technologies including artificial intelligence (AI), robotics, big data analytics, and the Internet of Things (IoT) is ushering in a wave of changes across military, political, economic, and societal spectra and forcing a re-think into how activities from individual to global levels have so far been conducted (Rotatori, Lee, & Sleeva, 2021). These technologies offer tremendous potential to solve problems plaguing the status quo. However, the nature of disruption that they could bring comes with novel challenges whose nature is still unfolding at this stage. These debates have arguably been highlighted the most around AI which has been touted to be the ultimate gamechanger amongst the Fourth Industrial Revolution technologies (Ayyar, 2016; Del Castillo, 2018; Girasa, 2020).\n\nAI has already crept into activities at various levels to heavily influence human behaviour in tangible or intangible ways. For instance, user behaviour on the internet within platforms ranging from social media to online shopping is heavily conditioned by manipulation through AI algorithms (Petropoulos, 2022; Rathenau Instituut, 2022). Meanwhile, the technology also learns or evolves through the various patterns it picks up from a person’s online preferences. Law enforcement is another sector where use of AI, particularly through predictive policing is revolutionising traditional methods of preventing and apprehending criminal or terrorist threats (Verma, 2022; Hunt, 2022). Ethical concerns including those pertaining to the violation of individual privacy and systemic bias have been raised against the practices accompanying AI integration in several instances (Jenkins & Purves, 2020; Heaven, 2020).\n\nThe transformative effects of AI are particularly relevant to understand in the context of geopolitics and international relations (IR). Through its integration into various sectors of the society, the technology is slowly shaping major geopolitical trends ranging from the United States of America (USA)-China technological rivalry to triggering an interest in intelligent warfare among major powers around the world (Kapetas, 2020). Moreover, these powers seem to view AI as a means to improve their relative position in the international system in economic and military terms (Alexandre & Miailhe, 2017). The interest in AI, particularly from an international relations point of view, can be seen to be springing primarily from three factors:\n\n♦ The nature and purpose of the technology.\n\n♦ Rapid advances in the field of AI over the past decade\n\n♦ Historical experience from the past industrial revolutions.\n\nSimply put, AI looks to simulate human thought process and functioning on machines. The idea has been intriguing, yet fear-inducing in several ways to humans who fear that their existence would not amount to much if a machine could serve their purpose. Further, the technology has been advancing at a fast pace since 2012, a year that witnessed several breakthrough events in deep learning (House, 2019). The years that followed had witnessed the technology rapidly integrated into aspects ranging from national security to day-today lives of individuals with its disruptive effects receiving wide coverage in all kinds of media. The final factor that drives the interest in AI is rooted in historical experiences. The coming of new technologies in the past have often turned existing power structures on their heads, a phenomenon that particularly stands true while speaking of industrial revolutions of the past. AI is an enabling technology like the steam engine or electricity and can boost the efficiency of anything that it is applied to (Lyu, 2020). Actors who make prudent and effective use of the potential AI offers are arguably well-positioned to draw benefits to improve their relative position compared to their competitors and influence others’ behaviour to benefit themselves. In this context, a deep look is warranted into the ways in which AI can enhance the power status of a country. An examination into understanding the exact role that AI will play in shaping the outcome of major power competition in the current context is also warranted.\n\nIn pursuit of global domination or a relative improvement in their status, major powers including the USA, China, Russia, India, and the European Union (EU) have been seen to be ramping up their investments on AI research and development (R&D) to arrive at major breakthroughs. Several scholars argue that these nation-states are engaged in a ‘global AI race’ to utilise the technology to boost their economic productivity as well as military effectiveness to get ahead of the rest (Geist, 2016; Savage, 2020; Levy, 2021; Stanford University, 2021). On the commercial front, AI can trigger largescale unemployment as well as enable the rise of new commercial technology giants. AI can equip a country with formidable economic power. The potential for competition to achieve a technological breakthrough in AI is particularly said to manifest as an accelerated arms race that will create instability at national and international levels. This is because AI can potentially facilitate newer forms of warfare entailing highly advanced offensive and defensive capabilities wielded by the warring nation-states. Increased integration of autonomous or semi-autonomous weaponry is further expected to increase the speed of warfare. In a projection for a worst-case scenario, AI and its contribution to reducing the human cost of war can possibly incentivise nation-states to engage in them frequently.\n\nIn this background, it becomes pertinent to attempt to try to project whether or how AI will impact the emerging global order and the nature of the global order shaped by the technology. While exact predictions in this regard are impossible given the uncertainty surrounding the nature of evolution of the technology, this article would base its analyses on select factors drawn from historical instances namely, the First, Second, and Third Industrial Revolutions. It critically examines the concept of first-mover advantage in technological innovation and its correlation to great power status in IR. It draws from this understanding to assess how patterns of technological diffusion alter relative power capabilities of nation-states. The study shall examine the impact of the advent of nuclear and space technologies to infer how a nation-state’s technological capabilities determine its power status. The qualitative analysis on the aforementioned aspects shall be utilized as a basis to understand the extent to which AI can re-order international power hierarchies.\n\nThe research paper has been built on a foundation examining theoretical discourse on technology and its correlation to the concept of power in IR. It grounds itself on the understanding on National Power and its systemic implications understood through the lens of balance of power in the international system. The fundamental point of inquiry pertains to how nation-states translate their technological capabilities into strategic dividends, such as elevation or preservation of power status. Analyses on theoretical formulations have been correlated with historical analyses. The paper then places the concept, nature, and applications of AI within the historical experience and correlates the same to the current standing of the USA, Russia, and China with respect to their AI capabilities through analysing their relative approaches and capabilities. The three nation-states are noted to have been entangled in competitive power dynamics amid ‘the ‘New Era of Great Power Competition’ (Congressional Research Service, 2022) as China and Russia are increasingly seen to be challenging the US-led world order (Allison, 2020; Savoy, 2022; Tiezzi, 2022). The three countries have consequently been identified as countries with stated intention to emerge as victors in the global AI race (Simonite, 2017; Minevich, 2017; Lant, 2017; Garcia, 2019). The article additionally tries to identify variables pertaining to the nature of domestic polity and socio-economic organization in the USA, China, and Russia that might determine whether these nation-states could draw from AI to enhance their power status in the international system. An attempt is finally made through scenario building to demonstrate a possible interplay between these attributes within the evolving geopolitical context.\n\n\nTechnology as a catalyst of systemic change\n\nThe rise and fall of great powers in the modern era has arguably followed a techno-economic logic. Small Powers or Middle Powers might look to acquire technology to boost their National Power to rise to a Major power or Great Power status. Meanwhile, Power Transition theories in IR highlight a country’s ability to innovate as well as dominate in leading sectors as an important indicator of a country’s ability to rise to as well as preserve a superior power status. A country’s position in the international power hierarchy in this context is determined by whether a country can sustain its power through efficiently managing its technological growth. As well-acknowledged in the dominant discourse in IR, the periodic rise and fall of Great Powers in the international system has often occurred amid an overall milieu defined by the coming of new technological innovations. Countries may move up or down the power hierarchy based on how a country adopts and manages the disruption from a new technological innovation. Kennedy underlines this fact while explaining the rise and fall of powers as a consequence of change in relative military and economic capabilities as;\n\n“differentials in growth rates and technological change, leading to shifts in the global economic balances, which in turn gradually impinge upon the political and military balances (Kennedy, 1989).”\n\nThe expectations on how AI will impact international relations in the coming years are largely based on these patterns as seen from time immemorial. For instance, the coming of chariots in the 1700 BC altered the power structures and changed the character of warfare in the ancient civilisations of Mesopotamia, India, and China and is said to have facilitated the movement of Aryans from Central Asia into Northern India (World Supporter, 2014). Those in possession of the technology enjoyed a superior social standing which flowed from their ability to use the technology as a tool to subjugate the weaker classes. The revolution in iron smelting technology that came forth around the 1200 BC had a similar effect. With the availability of iron for armour and weaponry, infantry was effectively able to quickly neutralise chariots. The invention of stirrups in medieval Europe if further said to have paved the way for a power structure that would manifest in the form of Feudalism (Derby, 2001). The use of stirrups by the Mongols is further said to have enabled their relentless, quick forward movement into invading territories (Ingliss-Arkell, 2017). This is because stirrups would allow mounted knights to emerge as a core strike element of any major armed conflict of the time. Their status that would pervade into other aspects of social life to yield an unequal relationship between peasants and nobles in the society. The major centres of power in the modern and immediate pre-modern era such as the Ottoman, Mughal, Safavid, Ming, and Tokugawa empires rose to prominence through possession of superior strategic technologies, prominently gunpowder and cannons. Nicknamed as the ‘gunpowder empires’, these powers were able to subjugate their rivals and consolidate territory by utilising this technology against their awestruck rivals, although they were greater in number in several cases (Kennedy, 1987).\n\nThis effect of technology on power dynamics between nation-states is seen to be much more profound in the Westphalian era. The three Industrial Revolutions that the world has witnessed over the past three centuries have altered the nature of the global order through causing the fall of prevailing great powers and facilitating the rise of others to occupy those positions (Ding, 2021). The First Industrial Revolution which came about in the 18th century saw technologies including the steam engine revolutionise as well as boost the means of production. The United Kingdom (UK), the country that played host to these innovations reached the peak of its hegemony in the phase that immediately followed. The UK was able to defeat resistant inhabitants of its colonial territories in Asia and Africa who unsuccessfully employed inferior technology to attempt to defeat them and thereby consolidate their power over vast swathes of territory that held abundant resources. Additionally, the spurt in manufacturing and mercantile friendly policies enabled it to become the economic epicentre of the world, its mastery over shipbuilding and naval technology bolstered its command over the seas and sea commerce, in turn allowing it to emerge as a formidable military power. In other words, the UK was successfully able to leverage its first mover advantage to emerge as a global hegemon reigning over the international system for at least two centuries that came after the First Industrial Revolution.\n\nThe era following the Second Industrial Revolution witnessed innovations including electricity and a boom in oil mining technologies occur in the USA. Meanwhile, Germany and Japan were able to advance in the chemical and iron and steel sectors. As these countries started accumulating capabilities through harnessing technological innovation, there was a minor upset over the prevailing balance of power. These countries were effectively able to utilise these technologies to improve their economic and military capabilities. The technology-power interplay is perhaps most visible in the case of Germany, following its reunification in 1871. Under the Prussian Empire, the country had forged ahead with its booming coal, iron, and steel and chemical industries (Chandler Jr., 1990, 251). Along with facilitating a rapid development of rail networks ideal for troop movement, Germany would successfully translate its technological strength into rendering its military into a force that could defeat any other formidable power in the world. The massive arms build-up by other European powers in response to Germany’s amassing of techno-military might would eventually culminate in World War I. While Germany failed to translate its superior technological capabilities into a military victory, Germany’s technological rise and its bid to challenge the status quo would leave the UK significantly weakened. Moreover, this would further allow the USA and Japan to gain international influence as British hegemony was experiencing a relative decline.\n\nBritish hegemony would ultimately come to an end by the conclusion of World War II with the USA donning the mantle of superpower in the decades that followed (Lozada, 2005). The destructive might it demonstrated through using the atomic bomb in Hiroshima and Nagasaki in 1945 would go on to establish its status as an indisputable great power. By being the first to acquire this new weapon, the USA signalled the start of the decline of the UK’s supremacy, and also the beginning of a new power-balancing structure in world affairs. The USA’s acquisition of German blueprints for strategic technologies, prominently the V1 and V2, the earliest prototypes of the ballistic and cruise missile respectively, also significantly allowed the country to amass capabilities in a major way (Jacobsen, 2014). It can be argued that the fundamentals gained from German technology is what enables the USA’s global force projection through various land, sea, air and space assets in the current context.\n\nIn spite of all the myriad political, economic, and structural changes that have occurred since 1945, the USA has continued to draw from its first-mover advantage in nuclear weapon technology to preserve its position as a world leader, giving it the ability to set international norms in its interests. In addition to catapulting the USA to the superpower status, nuclear technology acted as one of the forces driving bipolar politics during the Cold War era, yielding both cooperative and competitive dynamics between the two blocs spearheaded by the USA and the Soviet Union. The five de facto Nuclear Weapon States (NWS) - the USA, Russia (the former Soviet Union), the UK, France, and China identified as so under the Nuclear Non-Proliferation Treaty (NPT), 1970, have managed to bolster their exceptional status through preserving their roles as sole holders of permanent membership at the United Nations (UN) Security Council. Their first mover advantage has further been preserved through setting and solidifying norms including the acceptability of peaceful uses of nuclear technology, non-proliferation, and the nuclear taboo which reinforces the notion that use of nuclear weapons is akin to the destruction of mankind itself (Ying, 2019). Their interests continue to be protected by a series, institutions such as export control regimes and treaty arrangements including the NPT and the United Nations Conference on Disarmament.\n\nNewer dimensions of the technology-power interplay can be identified from the experiences of the Third Industrial Revolution. The advent of technologies including advanced electronics and microprocessors would effectively render Information and Communication Technologies (ICT) indispensable to how nation-states carry out governance along with strategic and economic activities in the modern day. The array of technologies falling under the umbrella term ICTs, particularly those connected to rapid advances in computers and semiconductors, ushered in an ‘epochal shift’ from mechanised systems of the industrial era to information-based systems (Galambos, 2013, 2-4).\n\nThrough serving as a medium fostering high level of interdependence among nation-states, ICTs effectively ushered in globalisation and paved way for the multipolar global order that prevails today. The USA which pioneered these technologies since the 1950s and 60s sought to benefit from the ICT revolution which came about in the 1990s. However, it would briefly feel threatened by Japan during the last quarter of the 20th Century as the country would forge ahead in key areas of high technology such as electronics and semiconductors.\n\nJapan’s lead in electronics and Information Technology encouraged several scholars to predict its rise to challenge the USA as the world’s leading industrial power (Lohr, 2011; Gilpin, 1997; Ozawa, 1974; Ingersoll, 1985-86). Fears were rife that Japan would abandon its pacifist posturing to convert its economic might into military and disrupt the prevailing world order at the time. Time would eventually douse these fears as the growth of Japan’s export-oriented economy stagnated around the 1990s. The USA would maintain its first-mover advantage in ICTs effectively adapting its ICT-enabled service industries to computerisation. Moreover, the USA’s lead would also draw from its ability to cultivate as well as attract a pool of talent that can advance its lead (Ding, 2021). The country was effectively able to capitalise on an opportune unipolar moment to integrate ICT into its military and economic activities. The Third Industrial Revolution effectively helped the USA strengthen its pre-eminent position through maintaining and improving its global force projection. Meanwhile, other countries such as India and the Association of Southeast Asian Nations (ASEAN) countries took advantage of the cheap manufacturing costs of the technology and human capital to exploit the opportunities provided by an interdependent world to emerge as powerful economies.\n\nAs highlighted in the above discussion, new technologies often interact with existing power structures to completely transform them. While it may cause the rise as well as fall of great powers, the geopolitical context as well as factors including the nature of polity or economy, the nature of leadership, and of the technology itself will determine whether the prevailing great powers can employ the technology to bolster their predominant status. In this context, the rate of diffusion of technology, access to resources, and openness to change can be identified as primary factors that are seemingly determinative of how the coming of new technologies can impact the nature of the world order.\n\nIn order to understand how AI will interact with geopolitics, a detailed examination of the technology itself is required. It is important to understand if AI will shape the global order like the industrial revolution technologies of the past or like a technology of military origin such as the nuclear weapon. The constituent components of the technology as well as its overall nature further needs to be examined.\n\n\nAI: Concept, nature, and applications\n\nAI is a technology that is still unfolding. Hence, there are inherent limitations to predicting the exact form the technology might evolve to assume in advanced stages. In the current stage of AI development, it can perform activities including recognition of patterns, statistics, and images and natural language processing. However, even in its routine tasks, such as image classification, AI in its current form is generally seen as unable to perform activities using common sense like humans, but instead relies on the ways that it perceives the inputs (Scott, Heumann, & Lorenz, 2018).\n\nThere are two broad approaches in AI development: symbolic and connectionist. In symbolic AI, the algorithms operate through deducing key behavioural pathways, while the connectionist approach trains algorithms to solve problems through calculations. In this regard, algorithms learn to perform complex tasks through two prominent types of learning as described in programming parlance namely machine learning and deep learning. While machine learning uses computational techniques such as decision trees to feed information and rules to the algorithm, deep learning relies on neural networks that function much like a human or animal brain to think and perform tasks such as image recognition (Dong, 2017).\n\nIn this background three types of AI can be identified: Artificial Narrow Intelligence (ANI), Artificial General Intelligence (AGI), and Artificial Super Intelligence (ASI) (Fourtane, 2019). Much of AI that is in use today falls within the scope of ANI. In ANI, algorithms learn from a specific data set to acquire the capability to solve single, rather straightforward problems, such as identifying certain objects from images, monitor weather, or play chess. While it may appear that narrow AI is capable of quick-thinking to understand complex scenarios, the scope of the technology is narrowly pre-determined. The next stage of evolution of technology is AGI wherein the algorithm can mimic an average human brain in performing tasks of varying levels of complexity. ASI has been said to be the ultimate form of evolution and represents the point where AI surpasses human intelligence to become super-intelligent machines, the entities that alarmists fear would bring about the elimination of the human race. According to a scientific school of thought the achievement of ASI will trigger the stage of singularity which would result in ‘unforeseeable, irreversible changes to human civilisation’. According to a version of this theory, the super-intelligent entity will keep upgrading itself hence causing an ‘intelligence explosion’ (Hvistendahl, 2019).\n\nAs previously noted, much of AI in use today is ANI with the technology being trained on a specific dataset to perform specific tasks such as data or image processing, identification of patterns, and so on. In addition to speeding up processes that humans take time to perform such as sorting through and connecting millions of data points, combinations of AI and robotics can also be used to perform tasks that are considered too risky or dangerous for humans. Such applications could become more widespread in scenarios such as disaster relief and rescue as well as in military applications such as detection of mines or Improvised Explosive Devices (IED) or guarding of borders. For instance, the USA has been using TALON, a military robot for identifying and disposing IEDs in locations such as Bosnia and Herzegovina, Afghanistan, and Iraq in the post-1990s era (TALON Tracked Military Robot, 2020). The USA’s Army is further said to have deployed the Special Weapons Observation Remote Reconnaissance Direct Action System (SWORDS), an improved variant of TALON fitted with a semi-autonomous gun in Iraq (Wired, 2007). Objections and ethical concerns have been raised against Lethal Autonomous Weapon Systems (LAWS) that can potentially strike targets without human oversight by prominent figures including Stephen Hawking and Tesla and SpaceX CEO Elon Musk (Cifford, 2017). While these weapons are perceived by critics as killer robots with their own ability to kill, nation-states including the USA and China are not looking to remove human oversight upon LAWS, although the scope of human supervision may face several challenges if and when the technology advances sufficiently for humans to comprehend its actions (Scharre, 2018).\n\nWhile AI offers lucrative options to militaries, the technology is pervading rapidly into various sectors of the economy to become an omnipresent part of how products and services are developed, manufactured, and sold. Such uses of AI-enabled predictive analytics seem to be growing in sectors ranging from healthcare to outer space. Combinations of ANI with big data analytics is already being tried out in various arenas such as in the development of models to analyse the spread of the COVID-19 pandemic, to assess the nature of the disease as well as the patterns of its spread. Use of algorithms has also become a quintessential element of modern-day marketing with AI learning from user’s data to gauge their preferences and offer suitable options for them. AI, big data analytics, and robotics is further expected to improve the quality and quantity of products and speed up as well as streamline manufacturing and movement of goods and services hence boosting the overall economic productivity of a nation-state. To get ahead in the AI race through military or economic means, the contenders may place an equal amount of emphasis on both with both the strands building into each other. Nation-state’s priorities with respect to AI may further be shaped by their unique geopolitical and security circumstances. The next section highlights the key pillars of AI plans of major powers.\n\n\nAI strategies of major powers: An overview\n\nGiven the hype surrounding AI, major powers including the USA, Russia, China, India, and the EU have started to place a large amount of importance on it. The governments of these countries have further been engaged in charting out legal and policy frameworks to facilitate faster development of AI, to reap its full potential as well as manage the disruption that it already seems to be bringing about. A glance at the national AI strategies of the USA, China, and Russia provides key insights into where the powers currently stand relatively to each other.\n\nChina recognises AI as a strategic technology that will guide international competition in the future and underlines its role in the protection of national security as well as in enhancing national competitiveness. As per China’s New Development Artificial Intelligence Plan released in July 2017 (DigiChina, 2017), it has its eyes set on becoming a world leader in AI by 2025. In doing this, it is looking to challenge the USA’s supremacy in both the civilian and military spheres. Beijing here is following the doctrine of ‘civil-military fusion’ which entails a blurring of lines between civilian and military resources in pursuing advances in science and technology (Tay, 2020). The goal here is to channel all national energies including those of academic institutions, military and private players to fasten the country’s military modernisation as well as economic growth (Pecotic, 2019). In the military sphere, AI is central to the People’s Liberation Army’s (PLA) ‘intelligentization doctrine’ (Bassler & Noon, 2022). This accords for the technology an important role in helping commanders with strategic decision-making through analysing volumes of data including satellite imagery and GPS locations of troops that they are expected to manoeuvre.\n\nThe government has further hand-picked Chinese AI giants, Tencent, Alibaba, Baidu, and iFlytek (a leading speech recognition company) as its dream team, tasking them to work on different priority areas (Jiang & Dai, 2017). While Baidu has been tasked with technologies relating to autonomous driving, Tencent has been tasked with looking at AI in healthcare and medical diagnostics. Alibaba Cloud (Aliyun) is looking into smart cities. Meanwhile, China has also been integrating AI into its domestic governance to maintain stability and exercise social control (Ding, 2018). Rampant surveillance using facial recognition cameras are used to identify those who violate traffic rules and commit crimes. It’s policy in the Xinjiang Autonomous Region using AI algorithms is specifically intended at racial profiling to track and control the Uighurs (Mozur, 2019; Wakefield, 2021).\n\nThe USA’s Department of Defense’s (DoD) Third Offset Strategy released in 2014 places a heavy emphasis on AI to be integrated into the military domain to maintain a military edge over near-peer competitors, Russia and China (Gentile, et al., 2021). Moreover, through this document, the DoD has discussed how the USA can employ technology to negate its military disadvantage vis-a-vis its competitors, placing a heavy emphasis on human-machine collaboration. This has been termed the ‘Centaur Model’ and aims to combine the efficiency of humans and machines in a way that they complement each other by addressing each other’s flaws (Horowitz, 2018). The Pentagon’s AI strategy released in February 2019 (US Department of Defense, 2019) further laid out specific priorities in this regard. It states that AI will be integrated in areas including 'improving situational awareness and decision-making, increasing the safety of operating equipment, implementing predictive maintenance and supply, and streamlining business processes' (US Department of Defense, 2019, 7). In doing all this, the overall goal is to augment the capabilities of the troops to free them of “tedious, cognitive physical tasks” (US Department of Defense, 2019, 7). The emphasis here seems to be on handing over to AI the tasks that humans may find mundane and time-consuming such as analysis of surveillance data (Cassano, 2018). For instance, the DoD’s Project Maven - which witnessed a 580% increase in funding from 2018 to 2019 - is intended to analyse feeds collected by thousands of drones (Cassano, 2018). Meanwhile, robots are deployed in the field to perform dangerous missions such as detecting IEDs. On the commercial front, leading players including Facebook, Apple, Amazon, Netflix, and Google (commonly referred to using the acronym FAANG), as well as Microsoft, have an edge over Chinese firms in access to data owing to their collective dominance over the global technology landscape (Mulrenan, 2020), which is crucial for AI’s evolution.\n\nCompared to the USA and China, Russia is often not regarded as a “frontrunner in the global AI race” (Nair, 2022) and has been termed akin to an outsider in the same (Nacetti, 2020). Russia lags significantly behind China and the USA in terms of key indicators that are employed to assess a country’s technological capability such as number of patents, publications in journals and total investment in R&D. The strength of its digital economy has been deemed weak with fewer private players involved. While Russia is said to have a strong basis in mathematics and basic sciences (Pecotic, 2019), it is said to have shortcomings in terms of talent that can be employed in developing AI (Petrella, Miller, & Cooper, 2021). It is further affected by the ‘brain drain’ phenomenon with prospective talent often migrating to the USA or Israel in search of lucrative opportunities. In a stark contrast to China and the USA, Russia further appears to rely on state-owned entities to innovate as well as implement its AI strategy (Petrella, Miller, & Cooper 2021). Much like China, Russia handed over the task of developing roadmaps for developing technology to various state-owned enterprises. While Rostec was tasked with the 5G implementation roadmap, Rosatom was assigned the task of developing a quantum computing roadmap. Other state-owned entities such as Sberbank, Rostec, Yandex, and Gazprom Neft have been tasked with developing AI systems for their own diverse purposes, such as improving bank operations, streamlining military manufacturing, creating driverless cars and managing oil production, respectively (Petrella, Miller, & Cooper 2021, 81).\n\nRussia’s 2019 AI strategy (Ministry of Digital Developments Communications and Mass Media of the Russian Federation, 2019; Bendett, 2019) sets out a goal for the country to become a leading contender in the global AI race through sharpening its existing capabilities in science, engineering and mathematics and through making coding expertise available. It also emphasises upon AI ethics and data sovereignty by calling for various kinds of data including those from surveillance systems, weather, sound, and medical sources to be stored in Russian databases. It further lays focus on legal and ethical aspects with respect to the handling of data as well as to govern the “interaction of the individual with AI” (Bendett, 2019). It further emphasises upon integrating the technology in the healthcare and education sectors. The role of the private sector here seems relatively muted in comparison to the USA and China. However, Russia is rather reticent on its intentions toward military (Bendett, 2019) applications of AI although it has been forging ahead with plans for using AI to strengthen its military. Russia’s intentions for military applications are largely unknown, although it has already been using the technology to make smarter weaponry. Russia’s strategy for AI in the military domain rests on two key pillars: the strengthening of its existing weapons and platforms through integrating AI as well as through use of AI in asymmetric tactics. Russia is believed to be integrating AI into weapon systems such as those entailed in electronic warfare, air defence, guided missile systems, and drones.\n\nIt is reported to have tested several of these systems including the Uran-9 autonomous tank in the Syrian conflict (Robitzski, 2019) where its military tactics also underwent several changes. These ambitions can also be seen reflected in Russia’s plans for modern systems including the Su-57 multi-role fifth generation fighter jet as well as the T14 Armata Main Battle Tank. Russia is also said to be working on a smart missile which can alter its course based on the incoming missile defence system (Futurism, 2017). AI also occupies a central place in Russia’s hybrid warfare strategies. The country is said to use AI algorithms to power its disinformation campaigns intending to influence the politics of other countries (Polyakova, 2018). Such tactics have been evident from its military campaigns in Ukraine in both 2014 and 2022 (Blankenship & Ordu, 2022; Baumann, 2020; Kuzio, 2019).\n\n\nAI and the emerging world order\n\nWhile AI could usher in a range of technological innovations, these need not always translate to military innovations that demand radical changes in organisational characteristics or war-fighting strategies. Since AI technology is relatively nascent at this stage and given the unpredictability surrounding its applications, it is far too early to draw comparisons with previous instances of major technological change, such as that of the development of tanks during World War I. In this context, the purpose is best served by examining the capabilities major powers while placing them within the organisational attributes that are most likely to position entities that constitute a nation-state such as the government and the military to adapt quickest to a major technological change and achieve an edge.\n\nThe Adoption Capacity Theory posits that specific organisational and financial considerations of militaries determine the rate of diffusion of technological innovations and its impact on the balance of power. While financial considerations relate to attributes including the cost per unit for hardware and other investments, organisational considerations pertain to the military mindset, specifically as to how the military would perceive the resultant changes in war-fighting, or existing bureaucratic practices that may obstruct certain actors from facilitating the adoption of technology (Gilli & Gilli, 2014). As discussed before, technological innovations by themselves do not impact the balance of power, although the way in which the technology is used may set a military apart (Posen, 1984).\n\nFor instance, the British Royal Navy invented the aircraft carrier to operate as a platform for surveillance aircraft to spot battleships. Invested with a traditional mentality, the Royal Navy saw carriers as an aide to battleships. However, rising naval powers, Japan and the USA were quick to realise that the most effective use of the aircraft carrier was as a mobile attacking platform. In this case, the UK found itself at a disadvantage in terms of the mindset, as it would have been difficult for commanders to think outside the box. The emerging navies of USA and Japan were less bogged down by such considerations. Therefore, the chances are that countries with powerful battle-hardened militaries might find themselves at a disadvantage when it comes to adopting AI, while emerging powers could hold the advantage.\n\nAs elaborated in the above discussion, the scale at which AI can shape the global balance of power depends on three factors: firstly, the rate of development and diffusion of technology, secondly, the mode of utilisation and level of institutionalisation, and thirdly, on domestic factors. Understanding the patterns of diffusion in this context is crucial as changes to the balance of power in AI’s context will depend on whether the state that gains a first-mover advantage through being the first to achieve a breakthrough application of the technology is able to sustain it. For instance, nuclear technology which had a military innovation was hardly diffused. After the five nuclear weapon states got together to enforce the hierarchical NPT, merely three states namely, India, Pakistan, and North Korea have tested nuclear weapons. The NWS were essentially able to use their exceptional status secure their collective first-mover advantage to sustain a viable strategic upper hand on international affairs. ICT in contrast represents a widely commercialised military innovation. Low costs of manufacturing and ease of imitating within a globalised geopolitical context enabled several countries to rise to become major poles of the international system. The first mover advantage is hence likely to wither away once AI applications become imitable. While it can be argued the AI applications exclusively meant for the military are difficult to mimic, private industries as drivers of AI technology innovation might change the tide. Particularly, the nature of the relationship between governments and industries will provide insights into how fast nation-states can follow the first mover.\n\nBoth the USA and China are heavily relying on private players to develop AI technology. Dual-use technologies that private players are developing, such as facial recognition, could form the basis of potential applications in national security. Existing in the world’s most technologically advanced nation, American technology leaders including Google, Microsoft, and Facebook are favourably positioned to operate in a nurturing business ecosystem, characterised by less government oversight. However, increasing voices arguing for more government regulation of technology, particularly in the cyber domain, pose a challenge to relations between the government and the private sector, and could prevent ambitious partnerships due to a lack of trust (Huddleston Jr., 2019).\n\nMeanwhile, in China’s authoritarian capitalist economy, tech giants including Alibaba and Tencent maintain close ties with the government, whereas the business ecosystem thrives despite strict regulations. China’s AI strategy seems to follow a larger governmental outline, with designated roles for various actors including the industry and academia. Russia is at a relative disadvantage given its relative weaknesses in the digital sector. Unlike China and the USA, Russia has no significant private players to join hands with. This could perhaps be the reason why Russia is focusing on using AI to better its military strengths and through asymmetric means. Instead, Russia’s defence industry, which works closely with the Kremlin, is spearheading research on AI in the country. Recent technological advancements Russia has showcased, such as the Avangard hypersonic missile, clearly shows the resurgent capabilities of this Russian military-industrial complex (Garcia, 2019).\n\nWhile the USA remains the world’s most technologically advanced nation, China is fast catching up and envisions utilising AI and robotics among other things to level the playing field with the former. For China to replace the USA as the most advanced nation in the world, it is safe to say that it needs to come out clearly on top in the AI arms race. In keeping with the predicted scenario where the ones who master AI become masters of the world, China would have used its advantages in AI to attain parity with the USA. What could make this scenario possible is the ability of China’s ruling dispensation to maintain the level of control it has over all the public and private machineries functioning in its state and ensure dissent to the introduction of AI remains low-key. Indeed, through other policies aimed at opening up the markets it has access to, China is showing an awareness of ensuring AI does not take away jobs from its large population, and instead can open more opportunities for an upwardly mobile citizenry. Moreover, the USA’s military as an older, more experienced organisation, may have to cross multiple bureaucratic obstacles to adopt AI into their organisation. In comparison, the PLA seems more willing to take chances with giving AI wider scope and responsibilities and could stand to benefit from this. While it is next to impossible to say who will achieve a definitive lead, both nations will benefit from being at the forefront of the AI race and will inspire similar research to follow in other nations.\n\nEventually, the attaining of a Sino-American parity on AI might proceed on similar lines to the evolution of nuclear technology during the middle of the Cold War. Keeping strategic gains in sight, both countries may share technology with friendly nations through trade or partnerships, with some information leaking out through channels such as espionage. Particularly if information about commercial AI remains open source, these nations aspiring for a place at the high table of global affairs will be able to follow in the footsteps of China, the USA, and Russia, which based on current indications should continue to be a leading player in the race. The three powers may also develop countermeasures to defend against and anticipate adversarial operations, leading to a multiplication of technological applications such as confusing AI through manipulating images or triggering illogical reactions. War strategies are also likely to evolve to include a range of asymmetric tactics intended to stave off an adversary’s superior AI technology. However, the possibility of attaining a broader parity in terms of military capabilities is highly unlikely or in any case, could take a long time.\n\n\nConclusion\n\nIn pursuit of global domination or a relative improvement in their status, major powers including the USA, China, Russia, India, and the EU have been ramping up their investments on AI R&D to arrive at major breakthroughs. Several scholars argue that these nation-states are engaged in a ‘global AI race’ to utilise the technology to boost their economic productivity as well as military effectiveness to get ahead of the rest. For the USA, the prevailing superpower of the international system, AI offers the means to preserve or strengthen their existing position. In other words, AI can arguably compensate for the dent in its international perception due to a perceived decline of power following its misadventures in Afghanistan and the weaning away of economic dominance to China. Meanwhile, the latter seemingly views AI as a shot in the arm that can boost its national power to either supplant the USA or at least match its might to serve its own ambition to emerge as a global superpower in the next few decades. Russia’s perspective partially reflects the Chinese view with the country looking to employ AI in combination with asymmetric cyber tactics and space weapon technologies to enhance its existing military capabilities to gain an edge over the USA. Through AI, Russia hopes to get the USA to the negotiating table through which the country hopes to regain its past glory. Countries such as India or Singapore look to tap into the strength of their thriving knowledge economies and boost their skills in AI to address their larger security challenges.\n\nThis study has drawn from historical experience to assess whether major powers in the international system today, the USA, China and Russia can harness the opportunities offered by AI to enhance their power status as well as goals envisioned under national interest. Each of these players have variations in their approaches as well as differing abilities as organisations to absorb the technology into the fundamental ways in which they operate. A large part of whether any of these powers forge ahead and the ways in which the rise or fall these powers might come about is also contingent on whether the technology evolves from ANI to AGI or even ASI. Evolution to both AGI and/or ASI is likely to yield different projections given the geopolitical circumstances of the time. If economics is to shape the contours of the competition, China and India are likely to expand as major powers with a significant influence over international affairs. The USA holds a unique advantage in terms of the wealth of technological knowledge it possesses as well as through its private ecosystem which is churning out AI innovations at an ever-increasing rate. The world system will likely continue in its multipolar configuration in this case with commercial AI diffusing fast to enable multiple poles of power to strengthen their position.\n\nIf military innovation and its impact on war decides the outcome of the AI race, the USA and China are well-positioned to emerge on top. While the USA still has a substantial lead in capabilities and fighting experience compared to the Chinese, AI might slightly change the equation in China’s favour. China’s authoritarian system of governance, access to data, and smart policies such a civil-military fusion may further put the country at an advantage. Ultimately, the winner(s) of the AI race may be those who adopt a prudent approach to the technology and make calculated moves towards using this unique AI moment to rise to the top.\n\n\nData availability\n\nNo data are associated with this article.", "appendix": "References\n\nAlexandre L, Miailhe N: The Geopolitics of AI and Robotics, Interview of Laurent Alexandre by Nicolas Miailhe.2017. Retrieved December 1, 2020.Reference Source\n\nAllison G: The New Spheres of Influence Sharing the Globe With Other Great Powers. Foreign Aff. 2020; 30–40.\n\nAmaresh P: How is Fourth Industrial Revolution changing our economy?2022, February 3. Retrieved April 10, 2022.Reference Source\n\nAyyar R: Industrial Revolution 4.0 a game changer: Ramadorai.2016, February 28. 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Retrieved from World Supporter.Reference Source\n\nYing F: A Preliminary Analysis of the Impact of AI on International Relations. Quarterly Journal of International Politics. 2019. Reference Source" }	[ { "id": "164732", "date": "02 May 2023", "name": "W Lawrence S Prabhakar", "expertise": [ "Reviewer Expertise International Relations Theories", "Technology", "Foreign Policy Analysis", "Strategic Studies" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe article \"Potential impact of artificial intelligence on the emerging world order\" by Anupama Vijayakumar is a comprehensive and meticulous study and analysis of the role and impact of Artificial Intelligence on the emerging world order that is driven by Technology, specifically Artificial Intelligence and its constituent forces provides a competition paradigm among the Great Powers notably USA, Russia and China.\nArtificial Intelligence itself is an Industrial Revolution that has brought about a Paradigm Shift to the nature of industrialization and specifically Defense Industrialization. The quest for primacy and dominance is cast in the most tenuous competition that sees elimination of rivals and with the augmentation of all dimensions and vistas of national power and comprehensive national power.\nThe research article has well documented the cumulative impacts of the First, Second and Third Industrial Revolutions and works out the trajectories of the competing AI technologies paradigms that would eventually shape the Power primacy of each power. The author has well used the Adoption Capacity Theory to argue that the AI race is determined by strengths of relative capacity of the primary institutions, processes and their strengths and the relative disruptions it is likely to cause in the overall institutional and industrial capacities of the countries.\nThe article is well annotated with facts and is salient in analysis. The critical reviews of the strengths of the AI domain capacities of the USA, Russia and China are well analysed. The references are varied and are salient by the theme and reflect the correct citations. The text of analysis is sound and is reflective of the theme. The theoretical premises are well applied and analysed and the conclusions are well drawn and meticulously placed in the correct context.\nThe article could be indexed with no alterations and is most pertinent in its topic, theme and salience.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "180402", "date": "03 Jul 2023", "name": "Guangyu Qiao-Franco", "expertise": [ "Reviewer Expertise Artificial Intelligence", "Cyber Security", "China Studies", "Norm Diffusion", "International Organisations." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis manuscript engages with the important topic of AI and geopolitics. Overall, it is a strong, well-written and researched paper. It contributes to the literature by situating AI competition in a broader historical context of technology evolution and setting out clearly the AI ambitions and development status of China, the US and Russia. I support its publication, and my recommendation for the current manuscript is for minor revisions that bring more focus and depth to the theoretical argument and empirical operationalisation. Below, I offer comments that will assist the author in improving the piece.\nFirst, I would suggest the author state her stance to the research question of whether major powers can harness the opportunities offered by AI to enhance their power status and shape the world order explicitly in the introduction. This will help clarify the contribution of the paper and the need for studying the subject matter of AI and international order. (The conclusion can also be stronger in order to highlight this important original contribution that the author has set forth.)\nRelatedly, the author can be clearer on what the specific contribution of the paper is and in what ways this research builds on previous contributions on this topic. The topic of AI and geopolitics has already been studied elsewhere, albeit to a limited extent (see e.g., Bode et al. 20231 published in Ethics and Information Technology, which similarly compares the practices of China, the US and Russia). In addition, the issue has been extensively discussed at the UNGGE on LAWS. The author can do a more thorough job of situating the study in the existing literature(s) and teasing out the innovative points it brings to the table.\nAnother issue is with the AI definition. The term AI is used a bit unproblematically throughout, as if its referents were self-evident. Since the term can be interpreted differently by readers, a clear definition of the term would be helpful.\nI would also encourage the author to consider adding some reflections in three empirical sections on how the power balance and international order will evolve in the light of these discussions. For instance, to what extent, can we expect the trajectories of technology development and power shift to continue in the Fourth Industrial Revolution in the context of AI? What do the different policy approaches to AI of China, Russia, and the US say about the IR landscape? This will be helpful in clarifying and substantiating the theoretical discussion developed from p. 9 onwards.\nThe manuscript can also better source all the ideas it is drawing upon to better engage with the literature. The following statements all need to be sourced: “AI can equip a country with formidable economic power.” (p. 3); “The potential for competition to achieve a technological breakthrough in AI is particularly said to manifest as an accelerated arms race that will create instability at national and international levels. This is because AI can potentially facilitate newer forms of warfare entailing highly advanced offensive and defensive capabilities wielded by the warring nation-states.” (p. 3); the concept of “first-mover advantage” (p. 5); and “While it may cause the rise as well as fall of great powers, the geopolitical context as well as factors including the nature of polity or economy, the nature of leadership, and of the technology itself will determine whether the prevailing great powers can employ the technology to bolster their predominant status.” (p. 6). In absence of proper in-text citations, the analysis comes off as generalized (and unsubstantiated) personal opinion.\nSome other sentences and claims made by the authors need further clarification or refinement. These include, for instance, the following:\np. 7 “While these weapons are perceived by critics as killer robots with their own ability to kill, nation-states including the USA and China are not looking to remove human oversight upon LAWS, although the scope of human supervision may face several challenges if and when the technology advances sufficiently for humans to comprehend its actions (Scharre, 2018).” This argument is largely true but can be refined. States such as China and the US agree on retaining human oversight over weapons systems but differ on what constitutes human control. The US for instance has a much broader understanding of human control (or human judgement) than many other countries, especially developing countries.\npp. 7-8 discussions on China’s AI policies are not updated. Two of my latest papers might be helpful:\nGuangyu Qiao-Franco, Ingvild Bode, Weaponised Artificial Intelligence and Chinese Practices of Human–Machine Interaction2\n\nGuangyu Qiao-Franco & Rongsheng Zhu (2022) China’s Artificial Intelligence Ethics: Policy Development in an Emergent Community of Practice3\np. 8 “Russia’s intentions for military applications are largely unknown, although it has already been using the technology to make smarter weaponry.” I do not agree with this point as Russia’s position has been fairly clearly stated at the UNGGE on LAWS meetings. A few studies have been developed in this vein. An example would be Anna Nadibaidze (2022) Great power identity in Russia’s position on autonomous weapons systems4.\np. 9 “The first mover advantage is hence likely to wither away once AI applications become imitable”. This point can be developed by drawing on literature discussing the proliferation risk of AI weapons. For instance, Denise Garcia, Lethal Artificial Intelligence and Change: The Future of International Peace and Security5.\np. 10 “In comparison, the PLA seems more willing to take chances with giving AI wider scope and responsibilities and could stand to benefit from this.” I am not convinced of this point as the PLA similarly faces political restraints and takes into account the risk of AI to human control in AI development and deployment. See my paper on weaponized AI mentioned above.\nA few other issues are related to the assumption that economic might will convert into military advantage without discussing the scenario that military advantage translates into economic strength.\nLastly, the analysis seems to be largely optimistic about the potential of AI while containing relatively limited considerations of the risk of AI (e.g., cognitive bias, automation bias, the limits of datafication, technological limitations) and the increasing awareness of the risk of AI among power contenders around the world. It would be helpful to situate these AI political implication discussions in more balanced discourse that considers both the potentials and concerns of AI.\nI hope these comments will be of help to the author, to whom I wish the best of luck with the next stages of work on this interesting piece!\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "9896", "date": "11 Jul 2023", "name": "Anupama Vijayakumar", "role": "Author Response", "response": "Dear Ma'am, Hope you are doing well. I would like to convey my heartfelt gratitude to you for your detailed and invaluable suggestions. I am also thankful to you for sharing with me some excellent references for the purposes of revising my article. However, I do not have access to the following source through my institution or otherwise. Qiao-Franco G, Zhu R: China’s Artificial Intelligence Ethics: Policy Development in an Emergent Community of Practice. Journal of Contemporary China. 2022. 1-17 I really think it could help me broaden my understanding on China's AI strategy (otherwise found lacking in Western sources) if I could refer to your work for the purposes of incorporating the minor revisions. I would appreciate it greatly if you could share the same with me. Thanking you. Yours sincerely, Anupama" }, { "c_id": "10489", "date": "02 Nov 2023", "name": "Anupama Vijayakumar", "role": "Author Response", "response": "Once again, I am extremely grateful to the reviewer for her insightful suggestions and excellent references which have significantly expanded my knowledge and thinking on this topic. I have made a modest attempt to incorporate her suggestions to the best of my ability. Point-by-point responses to the reviewer’s comments as well as the corresponding revisions implemented have been described below: I apologise for the lack of clarity with respect to situating the specific contribution of the paper within existing literature. An attempt has been made in the introduction and conclusion sections to clearly articulate the research gap the study is trying to address. I apologise for omitting to include definitions of AI. This has been incorporated in the third section of the article titled “AI: Concept, nature and applications”. Analytical reflections on how the power balance and international order will evolve in light of the discussions entailed in the three empirical sections have been added in an attempt to substantiate the theoretical discussion. I apologize for omitting to add references for central ideas highlighted in the article. Sources have been added against the following statements: “AI can equip a country with formidable economic power.” The potential for competition to achieve a technological breakthrough in AI is particularly said to manifest as an accelerated arms race that will create instability at national and international levels. This is because AI can potentially facilitate newer forms of warfare entailing highly advanced offensive and defensive capabilities wielded by the warring nation-states.” the concept of “first-mover advantage”- a definition of first mover advantage has also been added in While it may cause the rise as well as fall of great powers, the geopolitical context as well as factors including the nature of polity or economy, the nature of leadership, and of the technology itself will determine whether the prevailing great powers can employ the technology to bolster their predominant status.” I apologise for not looking deeper into how the USA and China understand meaningful human control. This discussion has been incorporated into the third section of the article titled “AI: Concept, nature and applications”. Updated information on China’s AI policies have also been added to the fourth section of the article. Respectfully, I would beg to differ with the suggestion that Russia’s domestic AI development is in compliance with its stated position at the UNGGE. A stance that a country adopts at an international level is primarily intended to safeguard its sovereignty and allow itself space to bolster its national power, through AI in this case. A number of assessments on the ongoing Russia-Ukraine war for instance have pointed to Russia employing autonomous systems including attack drones and air defence systems. I have attempted to elaborate on the argument that imitability will erode first mover advantage in AI through expanding the discussion on technology diffusion and its implications on global power dynamics. These additions have been made in paragraphs four to six in the fifth section of the article titled “AI and the emerging world order”. I apologise for suggesting that the PLA may be more willing to give a wider scope and responsibilities to AI. This was influenced by dominant western views on China and is not necessarily true as a wider reading of the literature suggested. This has been removed from the article. I apologise for not considering the scenario where military advantage translates to economic strength. A modest attempt has been made to elaborate upon this angle in the sixth paragraph of fifth section titled “AI and the emerging world order”. I apologise for not giving due consideration to the risks emanating from AI and its negative impact on national power. This fact has been alluded to in the second paragraph of the conclusion and the second section titled “Technnology as a catalyst of systemic change”." } ] } ]	1	https://f1000research.com/articles/11-1186
https://f1000research.com/articles/12-1417/v1	30 Oct 23	{ "type": "Study Protocol", "title": "Impact of drainage catheter material, size, and anti-dislodgement mechanism on percutaneous nephrostomy exchange intervals: a systematic review protocol", "authors": [ "Deepak Iyer", "Menelaos Konstantinidis", "Hanzhou Li", "Zachary Bercu", "John Moon", "Menelaos Konstantinidis", "Hanzhou Li", "Zachary Bercu" ], "abstract": "Background: Percutaneous nephrostomy (PCN) is a commonly performed procedure by interventional radiology and urology to treat urinary obstruction. In this procedure, a catheter is percutaneously placed into the renal pelvis for urinary diversion or hemorrhagic cystitis. Material type, catheter size, and catheter shape (anti-dislodgement feature) ultimately contribute to the inherent traits of longevity in drainage catheter device. Reviewing the relative strengths or weaknesses of products in the existing clinical market may help clinicians critically appraise the devices they use with evidence-based findings from this review. Furthermore, a deeper understanding of the relative strengths and weaknesses of existing devices may help inform the next generation of drainage catheter devices to prolong the interval between exchanges without detriment to patient safety. Methods: The following electronic databases will be queried: PubMed, Web of Science, Cochrane from their inception to January 2023 to identify randomized controlled trials (RCTs) and cohort studies to investigate the differences that our interventions of catheter material, size, and dislodgement mechanism will have on the exchange interval (standard of care 90 days vs. 60 days vs. 45 days vs. 30 days). The primary outcomes will be the drainage catheter exchange frequency. Ethics and dissemination: We aim to share our findings through high-impact peer reviewed journals. As drainage catheters and minimally invasive interventional radiology procedures become more popular, it is important for healthcare providers taking case of these populations to understand which variables might optimize patient care and minimize emergent exchanges. Data will be made available to readers. Registration: PROSPERO (CRD42023432788, 16 June 2023).", "keywords": [ "Drainage catheter", "exchange interval", "anti-dislodgement", "interventional radiology" ], "content": "Introduction\n\nPercutaneous nephrostomy (PCN) is a commonly performed procedure by interventional radiology and urology to treat urinary obstruction. In this procedure, a catheter is percutaneously placed into the renal pelvis for urinary diversion. Obstruction is the most common indication, accounting for 85-90% of all PCN placements.1 Obstruction can lead to infection, and in turn can lead to sepsis, which increases patient morbidity and mortality.2 PCNs are also required for access to nephrolithotomy. Urinary diversion in the setting of urinary leak, fistula, or hemorrhagic cystitis is another common indication of PCN tubes as a modality to reroute urine from areas of inflammation.2–4\n\nLike any other procedure, PCNs are associated with major and minor complications. The Society of Interventional Radiology’s published major complication rate is around 2-10% and includes sepsis, hemorrhage, pneumothorax, puncture of adjacent organs, urine leak, and mortality.5–7 The technical success rate of PCN placement is typically at least 95% in dilated calyces without stones, making it a useful procedure for the patients with the previously mentioned indications.8 Prolonged use of indwelling drainage catheters has also been associated with poor outcomes, such as sepsis and encrustation, resulting in the exchanging of these catheters.9 There is no consensus in the literature regarding the optimal interval at which these drainage catheters should be exchanged, however, leading many institutions to opt for a 90-day exchange interval for percutaneous nephrostomies. While this interval is sufficient for majority of patients, previous studies have shown that there is an increasing incidence of Emergency Department visits for acute PCN exchanges, resulting in increased economic burdens on these patients and the health system.10\n\nThe size and material of the catheter used in the procedure can have an impact on success and complications rates.\n\nWhile small sized catheters might be more beneficial for patient comfort, some drainage catheter literature suggests that larger bore catheters may be associated with lower rates of obstruction due to thrombosis or encrustation and malpositioning.11–13 Prior research in cystostomy tubes suggests that catheter flow velocity increases significantly with increasing catheter lumen size per Poiseuille’s Law.14\n\nThe type and material of catheter used in percutaneous nephrostomy can also impact the success and complications of the procedure. Two of the more commonly used tubes in PCN procedures include Malecot tubes (made of silicone) and pigtail catheters (typically made of polyurethane), each of which have different complication rates.15 For example, silicone catheters have been shown to have higher rates of obstruction and dislodgement as compared to that of polyurethane catheters.\n\nAnother potential complication of percutaneous nephrostomies that might prompt an earlier than expected exchange frequency is the accidental dislodgement of the catheter, which can lead to further sequelae of obstructive uropathy. A number of anti-dislodgment mechanisms exist including pigtail “Cope” catheters, balloon retention features, and Malecot re-entry tubes.16\n\nThere have been no studies to our knowledge that explore these variables in the context of PCN exchange procedure intervals.\n\nMaterial type, catheter size, and catheter shape (anti-dislodgement feature) ultimately contribute to the inherent traits of longevity in drainage catheter device. Reviewing the relative strengths or weaknesses of products in the existing clinical market may help clinicians critically appraise the devices they use with evidence based findings from this review. Furthermore, a deeper understanding of the relative strengths and weaknesses of existing devices may help inform the next generation of drainage catheter devices to prolong the interval between exchanges without detriment to patient safety.\n\n\nMethods\n\nThis protocol follows the Preferred Reporting Items for Systematic Review and Meta-analysis Protocols (PRISMA-P).17,30 Should any modifications to the protocol be performed, they will be updated in PROPSERO (CRD42023432788, 16 June 2023). The methods for this protocol will be modelled from previously published protocols.18\n\nTypes of studies\n\nWe will be examining randomized controlled trials (RCTs) and cohort studies, case-control trials, case series, and observational studies comparing any two interventions against one another or against a placebo. Qualitative studies will be excluded. Only studies written in English will be included; however, we will not be excluding based on date of publication or setting.\n\nTypes of participants\n\nAdults (aged 18 years or older), of any sex and ethnicity, who have received a percutaneous nephrostomy as determined by the individual study inclusion criteria.\n\nTypes of interventions\n\nThe major interventions that will be explored through this systematic review include catheter sizes, catheter material, and anti-dislodgement mechanisms in the context of percutaneous nephrostomy exchange frequency. As discussed previously, these three variables all contribute to complications with drainage procedures that result in earlier than expected exchanges. We will be exploring studies that discuss the complication rates associated with varying catheter sizes based on French size. We will also be examining various catheter material types, such as silicone and polyurethane, as it relates to PCN complications and increased exchange frequencies. Finally, we will be examining the role that anti-dislodgement mechanisms play in the context of drainage catheter complications and need for emergent exchanges.\n\nTypes of outcome measures\n\nPrimary outcomes\n\n• Exchange frequency will be the primary outcome of interest in our systematic review. Currently, the optimal exchange frequency is not well established in the literature and many institutions opt for performing exchanges in a 90 day interval. Through this systematic review, we will be investigating the differences that our interventions of catheter material, size, and dislodgement mechanism will have on the exchange interval (standard of care 90 days vs. 60 days vs. 45 days vs. 30 days).\n\nSecondary outcomes\n\n• Morbidity\n\n• Mortality\n\n• Rate of sepsis\n\n• Hospital admission and readmission rates\n\nOur search strategy aims to identify all published literature for possible inclusion in this review. We will adapt the search strategy we developed for PubMed to search the following electronic databases.\n\nThe Web of Science (WoS) and Cochrane databases will be subsequently searched.\n\nWe will restrict publications to only those published in the English language.\n\nOur institution’s Information Specialist will search MEDLINE (PubMed; 1966 to date of search), Cochrane database, Embase (Elsevier; 1947 to date of search), Scopus (Elsevier; 1788 to date of search). Grey literature sources will also be included such as ProQuest Dissertations & Theses Global, Journal of Vascular and Interventional Radiology Supplements, and Cardiovascular and Interventional Radiology Journal Supplements.\n\nMEDLINE search strategy\n\n(((((“nephrostomy, percutaneous”[MeSH Terms] OR “nephrostomy catheter”[Text Word] OR “percutaneous nephrostom”[Text Word] OR “nephrostomy tub”[Text Word] OR ((“Kidney”[MeSH Terms] OR “kidney”[Text Word] OR “nephrotom”[Text Word]) AND (“Catheters”[MeSH Terms:noexp] OR “catheter”[Text Word] OR “catheters, indwelling”[MeSH Terms] OR “tube”[Text Word] OR “tubing”[Text Word]) AND “nephrostom”[Text Word])) AND (“Postoperative Complications”[MeSH Terms] OR “post operat”[Text Word] OR “infections”[MeSH Terms] OR “infect”[Text Word] OR “adverse effects”[MeSH Subheading] OR “adverse effect”[Text Word] OR “risk factors”[MeSH Terms] OR “risk”[Text Word] OR “treatment outcome/adverse effects”[MeSH Terms] OR “injur”[Text Word] OR “complicat”[Text Word] OR “sepsis”[Text Word] OR “tube exchange”[Text Word] OR “tube replace”[Text Word] OR “fail”[Text Word] OR “failure”[Text Word])) OR (“nephrostomy, percutaneous/instrumentation”[MeSH Terms] AND (“catheters, indwelling/adverse effects”[MeSH Terms] OR “catheters/adverse effects”[MeSH Terms])) OR ((“Nephrotomy”[MeSH Terms] AND “Catheter-Related Infections”[MeSH Terms]) OR (“nephrotom”[Text Word] AND (“tube size”[Text Word] OR “tube material”[Text Word] OR “catheter”[Text Word]) AND (“infect”[Text Word] OR “complicat”[Text Word] OR “injur”[Text Word] OR “risk”[Text Word] OR “outcome”[Text Word] OR “sepsis”[Text Word] OR “adverse effect”[Text Word] OR “fail”[Text Word] OR “failure”[Text Word])))) NOT “nephrolithotomy, percutaneous”[MeSH Major Topic]) AND “English”[Language]) NOT (“systematic review”[Publication Type] OR “meta analysis”[Publication Type] OR “systematic review”[Title] OR “meta analys”[Title] OR “literature review”[Title] OR “narrative review”[Title] OR “scoping review”[Title] OR “case report”[Title] OR “case series”[Title] OR “case stud*”[Title]).\n\nSelection of studies\n\nAll results identified in the search strategy will be imported into the software Covidence. Title and abstract review will be performed independently by two authors. Next, for all selected abstracts, the full text will be referenced and reviewed by two independent authors for inclusion or exclusion based on previously mentioned criteria. Should any disagreements occur throughout this process, a third author will be used to yield final decision. A PRISMA-P flow chart will depict the final set of inclusion and exclusion criteria.19 Throughout the process, we will be collecting the study characteristics, outcomes, and any relevant effect modifiers.\n\nStudy characteristics data\n\nArticle title, corresponding author, publication date, study setting, all conflicts of interest, study design, patient demographics and comorbidities, and inclusion and exclusion criteria will be noted. All primary and secondary outcomes will be noted along with any levels of uncertainty.\n\nBias will be assessed independently for each study that will be included in our systematic review. Like previously mentioned methods, two reviewers will independently assess risk with a third reviewer to yield final decision as necessary. The Cochrane Risk of Bias tool will be utilized to evaluate randomized control trials.20 Cohort studies will be assessed using the ROBINS-II tool across their specified domains.21\n\nSummary of study characteristics\n\nWe will use descriptive statistics to give an overview of the listed studies. Information on the study population’s characteristics, such as the different comparisons and design elements, will be included in this. Additionally, for each result, we will show a network diagram with the size of the nodes based on the patient count and the thickness of the edges based on the number of studies per comparison.\n\nEstimation of relative treatment effect\n\nWe will compute the odds ratio for dichotomous outcomes, mean difference or normalized mean difference for continuous outcomes, and hazard ratios for time-to-event or survival outcomes to ascertain the relative treatment effects. The surface under the cumulative ranking curve or p-score, which offers a scalar value ranging from 0 (least effective) to 1 (most effective) for each therapy in respect to a particular outcome, will be used to rank the treatments overall for each outcome.22\n\nAssessment of statistical inconsistency\n\nThe back-calculation technique will be used to assess local inconsistencies. We will also evaluate global inconsistency using the design-by-treatment interaction model.23 The R programming language and software will be used to carry out the consistency checks.\n\nConsideration of unit of analysis issues and missing data\n\nWe will consider the correlation between effect estimates in the network and incorporate multi-arm studies. Even though we do not plan to include any cluster or crossover randomized controlled trials (RCTs), if any are, we will abide by the recommendations in the Cochrane Handbook for Systematic Review of Interventions. Every analysis will be done with the objective of treating each individual. For each included study, we will investigate the level of missing data and examine the approaches taken to deal with it in the risk of bias analysis. We will do a sensitivity analysis for the key outcomes to determine the effect of including studies with a significant quantity of missing data. Measures of uncertainty will, whenever possible, be converted into standard errors. If no certainty is provided or can be inferred, the mean of known standard errors from included studies will be calculated.24\n\nAssessment of reporting biases\n\nTo assess the possibility of reporting bias and small-study effects, we will visually inspect the funnel plots or each treatment where possible (i.e., at least 10 studies for a given treatment). Additionally, to evaluate small-study effects, we will assess comparison-adjusted funnel plots, and where possible, conduct meta-regression with study variance as the covariate.25,26\n\nWhen possible (i.e., when there are at least 10 trials for a given therapy), we will visually evaluate the funnel plots for each treatment to check for reporting bias and small-study effects.25 Additionally, we will employ comparison-adjusted funnel plots (Chaimani et al.) and, if necessary, do meta-regression using study variance as the covariate to evaluate small-study effects.26–28\n\nFor the primary outcomes, we will explore possible sources of heterogeneity, such as the indication for percutaneous nephrostomy, other comorbidities, and the method of outcome ascertainment. If enough studies are available, we will perform subgroup analyses based on these characteristics. Additionally, for the primary outcomes, we will conduct sensitivity analyses by:\n\nRestricting the analysis to larger studies, as smaller studies may introduce bias. The study size will be determined by the median of the included studies, and in the sensitivity analysis, studies below the median size will be excluded.\n\nRemoving studies with more than 20% missing data.\n\nDescriptive synthesis and summary of findings will be performed in accordance with the Synthesis without Meta-Analysis in Systematic Reviews (SWiM) guidelines.29 For each primary outcomes, two authors will independently assess the nine SWiM domains as very low, low, moderate, or high confidence.29\n\n\nConclusions\n\nWhile this is a timely and important topic to explore as a systematic review, there are some limitations to this study. First, there potentially is only scant literature on the subject. As a result, conclusions drawn from the systematic review might not fully represent the entire clinical context of the role these interventions play on our outcome. Additionally, since this is a multicomponent analysis and only a descriptive synthesis will be performed through this systematic review, no relative effectiveness data will be collected. Additionally, it is possible that we will find statistically relevant results by chance alone; however, there are no methods to account for these issues in systematic reviews. Finally, different companies of these devices might manufacture their devices differently and with proprietary technology preventing a true one-to-one comparison across catheter materials and anti-dislodgement mechanisms. Given these limitations, we still strongly believe that the corresponding results will be meaningful to clinical practice but should be interpreted with caution.\n\nThis systematic review will serve as the foundation for comparison of varying catheter tubes and sizes on the frequency of drainage catheter exchange interval. As the incidence of drainage procedures increases, there may be patients with certain comorbidities that may benefit from a shorter exchange interval.\n\nNew institutional review board approval is not required for this study since it involves a systematic review of existing published data. Our intention is to disseminate our results through reputable peer-reviewed journals and conference presentations of significant influence. Given the growing prevalence of percutaneous drainage procedures, it is crucial for healthcare practitioners involved in the management of these patients to have access to concise and synthesized educational resources on this subject. The data synthesized from our study will be made accessible to readers.\n\nThis protocol was developed without the involvement of any patients or members of the public, and their inclusion in the completion or dissemination of the corresponding review and results is not planned.\n\nWe plan to begin title/abstract review upon successful submission of this protocol.", "appendix": "Data availability\n\nNo data are associated with this article.\n\nFigshare: PRISMA-P checklist for ‘Impact of drainage catheter material, size, and anti-dislodgement mechanism on percutaneous nephrostomy exchange intervals: a systematic review protocol’, https://doi.org/10.6084/m9.figshare.23937171.v1. 30\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nRamchandani P, Cardella JF, Grassi CJ, et al.: Quality improvement guidelines for percutaneous nephrostomy. J. Vasc. Interv. Radiol. 2003; 14(9 Pt 2): S277–S281.\n\nPearle MS, Pierce HL, Miller GL, et al.: Optimal method of urgent decompression of the collecting system for obstruction and infection due to ureteral calculi. J. Urol. 1998; 160(4): 1260–1264. PubMed Abstract \| Publisher Full Text\n\nBird VG, Fallon B, Winfield HN: Practice patterns in the treatment of large renal stones. J. Endourol. 2003; 17(6): 355–363. PubMed Abstract \| Publisher Full Text\n\nZagoria RJ, Hodge RG, Dyer RB, et al.: Percutaneous nephrostomy for treatment of intractable hemorrhagic cystitis. J. Urol. 1993; 149(6): 1449–1451. PubMed Abstract \| Publisher Full Text\n\nFarrell TA, Hicks ME: A review of radiologically guided percutaneous nephrostomies in 303 patients. J. Vasc. Interv. Radiol. 1997; 8(5): 769–774. PubMed Abstract \| Publisher Full Text\n\nRadecka E, Magnusson A: Complications associated with percutaneous nephrostomies. A retrospective study. Acta Radiol. 2004; 45(2): 184–188. Publisher Full Text\n\nDegirmenci T, Gunlusoy B, Kozacioglu Z, et al.: Utilization of a modified Clavien Classification System in reporting complications after ultrasound-guided percutaneous nephrostomy tube placement: comparison to standard Society of Interventional Radiology practice guidelines. Urology. 2013; 81(6): 1161–1167. Publisher Full Text\n\nPabon-Ramos WM, Dariushnia SR, Walker TG, et al.: Quality improvement guidelines for percutaneous nephrostomy. J. Vasc. Interv. Radiol. 2016; 27(3): 410–414. PubMed Abstract \| Publisher Full Text\n\nSzvalb AD, El Haddad H, Rolston KV, et al.: Risk factors for recurrent percutaneous nephrostomy catheter-related infections. Infection. 2019; 47(2): 239–245. Publisher Full Text\n\nLu X, Zhou B, Hu D, et al.: Emergency decompression for patients with ureteral stones and SIRS: a prospective randomized clinical study. Ann. Med. 2023; 55(1): 965–972. PubMed Abstract \| Publisher Full Text\n\nAten Q, Killeffer J, Seaver C, et al.: Causes, Complications, and Costs Associated with External Ventricular Drainage Catheter Obstruction. World Neurosurg. 2020; 134: 501–506. PubMed Abstract \| Publisher Full Text\n\nRobinson J: Selecting a urinary catheter and drainage system. Br. J. Nurs. 2006; 15(19): 1045–1050. Publisher Full Text\n\nDe Sio M, Autorino R, Quattrone C, et al.: Choosing the nephrostomy size after percutaneous nephrolithotomy. World J. Urol. 2011; 29(6): 707–711. PubMed Abstract \| Publisher Full Text\n\nLee C, Gill BC, Vasavada SP, et al.: Does size matter? measured and modeled effects of suprapubic catheter size on urinary flow. Urology. 2017; 102: 266.e1–266.e5. PubMed Abstract \| Publisher Full Text\n\nDepartment of Urology, Faculty of Medicine, “Lucian Blaga” UniversitySibiu, Grigore N, et al.: Comparative Study of Silicone and Polyurethane Nephrostomy Catheters used for long-term Urinary Drainage in Malignancy. Ro. J. Med. Pract. 2017; 12(2): 79–82. Publisher Full Text\n\nBayne D, Taylor ER, Hampson L, et al.: Determinants of nephrostomy tube dislodgment after percutaneous nephrolithotomy. J. Endourol. 2015; 29(3): 289–292. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMoher D, Shamseer L, Clarke M, et al.: Preferred reporting items for systematic review and meta-analysis protocols (PRISMA-P) 2015 statement. Syst. Rev. 2015; 4(1): 1. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChockalingam A, Konstantinidis M, Koo B, et al.: Surgical resection, radiotherapy and percutaneous thermal ablation for treatment of stage 1 non-small cell lung cancer: protocol for a systematic review and network meta-analysis. BMJ Open. 2022; 12(6): e057638. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPage MJ, McKenzie JE, Bossuyt PM, et al.: The PRISMA 2020 statement: an updated guideline for reporting systematic reviews. Syst. Rev. 2021; 10(1): 89. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHiggins JPT, Thomas J, Chandler J, et al., editors. Cochrane Handbook for Systematic Reviews of Interventions. Cochrane; 2019. Version 6. Publisher Full Text\n\nSterne JAC, Hernán MA, Reeves BC, et al.: ROBINS-I: a tool for assessing risk of bias in non-randomised studies of interventions. BMJ. 2016; 355: i4919. Publisher Full Text\n\nRücker G, Schwarzer G: Ranking treatments in frequentist network meta-analysis works without resampling methods. BMC Med. Res. Methodol. 2015; 15: 58. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWhite IR, Barrett JK, Jackson D, et al.: Consistency and inconsistency in network meta-analysis: model estimation using multivariate meta-regression. Res. Synth. Methods. 2012; 3(2): 111–125. Publisher Full Text\n\nFurukawa TA, Barbui C, Cipriani A, et al.: Imputing missing standard deviations in meta-analyses can provide accurate results. J. Clin. Epidemiol. 2006; 59(1): 7–10. PubMed Abstract \| Publisher Full Text\n\nSterne JAC, Sutton AJ, Ioannidis JPA, et al.: Recommendations for examining and interpreting funnel plot asymmetry in meta-analyses of randomised controlled trials. BMJ. 2011; 343: d4002. Publisher Full Text\n\nChaimani A, Higgins JPT, Mavridis D, et al.: Graphical tools for network meta-analysis in STATA. PLoS One. 2013; 8(10): e76654. Publisher Full Text\n\nChaimani A, Salanti G: Using network meta-analysis to evaluate the existence of small-study effects in a network of interventions. Res. Synth. Methods. 2012; 3(2): 161–176. Publisher Full Text\n\nMavridis D, Efthimiou O, Leucht S, et al.: Publication bias and small-study effects magnified effectiveness of antipsychotics but their relative ranking remained invariant. J. Clin. Epidemiol. 2016; 69: 161–169. Publisher Full Text\n\nCampbell M, McKenzie JE, Sowden A, et al.: Synthesis without meta-analysis (SWiM) in systematic reviews: reporting guideline. BMJ. 2020; 368: l6890. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIyer D: PRISMA P. ‘Impact of drainage catheter material, size, and anti-dislodgement mechanism on percutaneous nephrostomy exchange intervals - a systematic review protocol’.doc. [Dataset]. figshare. 2023. Publisher Full Text" }	[ { "id": "234532", "date": "08 Feb 2024", "name": "Azza Aly", "expertise": [ "Reviewer Expertise medical surgical nursing" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nDear Researchers I appreciate your efforts in preparing this protocol However, I have some points that provide specific areas for improvement and clarification to enhance the scientific integrity of the systematic review. The protocol is well structured, but the extensive technical details would benefit from simplification for a broader understanding. regarding the Methodology Provide a clear rationale for the chosen methodologies, addressing potential biases and limitations. I think you need to engage the patients in the research process to enhance the relevance of the study and its applicability to real-world scenarios. Contain the current status of the study or any progress made since the protocol was submitted.\n\nIs the rationale for, and objectives of, the study clearly described? Yes\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [] }, { "id": "244991", "date": "20 Feb 2024", "name": "Dragos Puia", "expertise": [ "Reviewer Expertise Urology" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nDear Researchers, I think your research is of interest considering that there are many patients with PCN, and many have problems with the catheter. However, the patients' pathology is very heterogeneous, and the underlying disease could influence the outcome. I would suggest performing a subgroups' analysis according to their underlying disease (cancers, kidney stones, other disease) and also if they have received a specific treatment like chemotherapy (in these patients there is high catabolism with an increased excretion of uric acid)\n\nIs the rationale for, and objectives of, the study clearly described? Yes\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1417
https://f1000research.com/articles/12-204/v1	21 Feb 23	{ "type": "Review", "title": "Anemonefishes: A model system for evolutionary genomics", "authors": [ "Marcela Herrera", "Timothy Ravasi", "Vincent Laudet", "Marcela Herrera", "Timothy Ravasi" ], "abstract": "Anemonefishes are an iconic group of coral reef fish particularly known for their mutualistic relationship with sea anemones. This mutualism is especially intriguing as it likely prompted the rapid diversification of anemonefish. Understanding the genomic architecture underlying this process has indeed become one of the holy grails of evolutionary research in these fishes. Recently, anemonefishes have also been used as a model system to study the molecular basis of highly complex traits such as color patterning, social sex change, larval dispersal and life span. Extensive genomic resources including several high-quality reference genomes, a linkage map, and various genetic tools have indeed enabled the identification of genomic features controlling some of these fascinating attributes, but also provided insights into the molecular mechanisms underlying adaptive responses to changing environments. Here, we review the latest findings and new avenues of research that have led to this group of fish being regarded as a model for evolutionary genomics.", "keywords": [ "adaptive radiation", "Amphiprion", "chromosome-scale assembly", "clownfish", "genome", "pigmentation", "proteomics", "transcriptomics" ], "content": "1. Introduction\n\nThe increasing availability of genomic tools and resources is revolutionizing our understanding of the molecular basis of evolution.1 Rapid advancements are being made in addressing questions such as: how does speciation occur, and how do new adaptations drive this process? Which genetic changes are responsible for morphological, physiological, and behavioral traits? How do organisms cope with rapidly changing environments? For the past decade, anemonefish have been a valuable tool for ecological and evolutionary research, but the development of molecular methods in recent years has made it possible to apply it to previously intractable problems in developmental biology, adaptive evolution, and speciation (reviewed in Refs. 2, 3). Whole-genome, transcriptome, and proteome sequencing, collectively known as “omics” tools (Figure 1), have opened anemonefish research up to new possibilities, hypotheses, and information regarding their ecology and evolution.\n\nThe green circle represents metabolomics, orange -proteomics, blue -transcriptomics, and pink -genomics. Colored arrows indicate interactions between the metabolome, proteome, transcriptome, and genome and how they affect each other. Circle sizes illustrate estimated complexity (adapted from Braun et al. 2021). Number of publications using “omics” tools were retrieved from the Web of Knowledge (https://apps.webofknowledge.com/) and plotted according to year of publication. Since the early 2000’s when the first studies investigating gene expression and technological advancement of various molecular sequencing platforms, the application of “omics” tools has increased steadily and led to the achievement of milestones such as the assembly of one of the most contiguous chromosome-scale fish genomes and the successful use of CRISPR/Cas9 gene editing in a reef fish (as shown by the light areas in the plot). Keywords used to determine these studies were separated into independent variables (or) within two categories donated by (and): “gene expression or genome or transcriptome or proteome or genomics or transcriptomics or proteomics or omics” and “clownfish or anemonefish or Amphiprion”.\n\nThe year 2018 saw the publication of the first chromosome-scale genome for an anemonefish4 (Figure 1). Of all published chromosome-level fish genomes to that date, the Amphiprion percula genome stood out as one of the most contiguous fish genomes with ~98% of the assembled genome ordered into chromosomes.4,5 This impressive feat not only highlighted the power of modern genome sequencing but most importantly, it empowered an array of studies in anemonefish (it has been cited 46 times as of February 2023 according to Google Scholar). Furthermore, the possibility of whole mRNA sequencing propelled transcriptome studies in a variety of tissues from multiple species and life stages to examine gene expression changes in development and adaptive responses to environmental stressors (reviewed in Section 2.3). Though not specifically for anemonefish, a growing number of studies are applying proteomics in coral reef fish (see Section 2.4). Monroe and colleagues (2020) used a mass spectrometry data-independent acquisition method for proteome quantification in a non-model fish species for the first time,6 a stepping stone for the application of proteomics to study many anemonefish.\n\nPreviously, “omics” methods were only used in a handful of studies, but now are one of the most common tools applied in the field and thus the primary focus of this review. Here, we describe the numerous attributes that make anemonefish an exceptional model system for studying evolutionary genomics. We then present a detailed synthesis of recent research that has provided important insights into the incredible adaptive radiation anemonefish have undergone,7–11 and how their genomic architecture underlies the evolution of complex phenotypic traits such as sex change12,13 and color patterning.14–16 We further describe how researchers are using anemonefish as a model system to understand the genomic basis of symbiosis with giant sea anemones10,17–19 and environmental plasticity.20,21\n\n\n2. Anemonefish as a model system for evolutionary biology\n\nThere are 28 species of anemonefish2 (Figure 2a), yet the two clownfish A. ocellaris (Figure 2b) and A. percula are perhaps the most recognizable ones, especially following the Disney movie “Finding Nemo”.22 Within the more than 300 species in the family Pomacentridae (to which anemonefish belong), two genera have been previously described: Amphiprion and Premnas, the latter including only one species which is being now considered as part of Amphiprion (reviewed in Section 3.1)23,24 (Figure 2a). All anemonefishes are protandrous hermaphrodites (i.e., male to female transition) that live in association with sea anemones.25,26 It is a mutualistic relationship in which the sea anemone provides food and shelter from predators25,26 and, in return, the territorial fish protects its host from predation by attacking other animals that attempt to feed on the tentacles.27 Furthermore, the fish serves as a supplemental nutrition source28 and also increases oxygen uptake by modulating water flow among the tentacles.29,30 Social groups typically consist of an adult breeding pair and several smaller (immature) juveniles ranked by size.31 Basically, a large dominant female is followed by a male so that if the female is removed, the male changes sex and the largest non-breeder matures into a breeding male.12 This male is also the one providing most of the parental care to the eggs by keeping them clean and well-oxygenated, another fascinating and rare feature of anemonefishes.2\n\na) Phylogeny of anemonefishes based on the 20 most informative genes (adapted from Marcionetti et al. 2022). Geographical distributions (light blue: NWI – North-Western Indian Ocean, dark blue: WI – Western Indian Ocean, green: CIP –Central Indo-Pacific Ocean, orange: CP – Central Pacific Ocean, yellow: SWP – South-Western Pacific Ocean, red: P – Polynesian Ocean), sea anemone hosts, and phenotypes are shown for each species. Asterisk denotes a recent revision of anemonefish phylogenetic data that suggests Premnas biaculeatus should be recognized as Amphiprion. Lastly, DNA symbol is shown next to the species for which genomes have been sequenced. b) The iconic false clownfish Amphiprion ocellaris. c) The white bonnet anemonefish Amphiprion leucokranos is a naturally occurring hybrid species found in the WI and CIP regions. d) Clownfish lay hundreds of eggs on the substrate near their host anemone. Pictures taken by Pascal Kobeh.\n\nLiving in symbiosis with 10 distantly related sea anemones (Cryptodendrum spp., Entacmaea spp., Heteractis spp., Macrodactyla spp., and Stichodactyla spp.), anemonefish can be found in shallow, tropical waters of the Indo-Pacific Ocean, from Australia to the Ryukyu archipelago (Japan) and from the Red Sea and southwest coast of Africa to the Maldives and French Polynesia. There is no anemonefish in the Caribbean nor in the Eastern Pacific (e.g., Hawaii).26,32 The highest diversity is in the Coral Triangle,26,33 where up to nine species have been observed coexisting together.34 Distribution varies greatly for each species, some are widespread (e.g., A. clarkii, A. sandaracinos), while others have a limited regional distribution (e.g., A. bicinctus, A. percula) or are even restricted to a few islands (e.g., A. chagosensis, A. latezonatus). Similarly, some species can only associate with one sea anemone (i.e., specialists), whereas other may have various possible hosts (i.e., generalists) (Figure 2a). For example, the yellowtail clownfish A. clarkii, for which a chromosome-scale reference genome was recently published,35 is the only species that has been observed to inhabit all 10 species of sea anemone.26,32 As such, it also has the widest distribution32 and temperature tolerance,36 making it a robust and accessible study species. In contrast, the tomato clownfish A. frenatus can only be found in one species of sea anemone (Entacmaea quadricolor).26,32 Furthermore, hybridization in the wild has been observed and several hybrid species (such as A. leucokranos (Figure 2c)) are known,37–40 prompting the study of its role in anemonefish evolution.8,11,41 Early studies have already characterized the ecology, behavior, and diversity of many traits of anemonefish, laying the groundwork for subsequent research using a variety of genomic tools that facilitated key discoveries across many biological fields (reviewed in Ref. 42).\n\nOne of the main reasons why anemonefish have become a model organism for a broad range of biological disciplines (e.g., host-microbiome interactions,10,17 developmental20 and phenotypic plasticity,43 behavior,44,45 and larval dispersal dynamics46–48) is because they can be easily found in the field due to their symbiosis with giant sea anemones, but they can also complete their life cycle in captivity.2,49 In contrast to adults, which are easy to observe and collect in their natural environment, studying wild anemonefish larvae represents a major difficulty. This has led to the development of husbandry methods and experimental protocols for “low-volume” rearing, and rearing and hatching embryos without parental care.49,50 Furthermore, anemonefish can adapt to different system types and culture conditions (e.g., closed or open systems, filtered or natural seawater, different temperatures and salinities), and do not require the presence of a host anemone, which makes their maintenance easier.49,50 Such rearing methods and intrinsic features of anemonefish has expanded their potential as a model organism by opening new avenues for the use of molecular tools (e.g., micro-injections51,52), functional approaches, and ecotoxicological and/or pharmacological experiments (reviewed in2). Crosses can be performed in the laboratory16 and anemonefish can spawn every two to three weeks, typically laying between 100 and 500 eggs on the substrate near their host anemone (Figure 2d). Further, they have a short embryonic and larval development (which have been characterized in precise detail53,54) of 10–15 days (depending on the water temperature and the species), thus allowing large-scale studies.2 Generation times, however, can be as long as 18 months (depending on the species), which might impose practical constraints on experimental approaches. Nevertheless, the biological traits and practical features in their breeding described above make them a growing model organism for developmental biology, ecology, and evolutionary sciences.\n\nAs significant advances in sequencing technologies have been made and genome projects become more affordable,55,56 a new era of genome biology in anemonefishes has also begun. It was only until recently, in 2018, that the first draft genome assembly for an anemonefish, that of the false clownfish A. ocellaris,57 was published. Though the coverage of this genome was low (~11×), it allowed the prediction of 27,240 high-quality protein-coding genes (96.3% of which were functionally annotated) and a genome size of 880 Mb. This was soon followed by the genomes of A. frenatus58 and A. percula,4 the latter of which was, until not long ago, one of the most contiguous and complete teleost fish genome assemblies currently available.5 Constructing a high-quality chromosome-level assembly for a species with no previous genome-scale data was certainly a major achievement in a world of sticklebacks and zebrafish (Figure 3a). Since then, the genomes of at least nine other species10,35 have been sequenced and deposited in public databases (Figure 2a). While these resources can provide valuable insights into the molecular evolution and adaptation of common anemonefish traits,10,11,35,59,60 with the exception of A. percula4 and the recently published chromosome-scale assemblies of A. ocellaris60 and A. clarkii35 (Figure 3a), most of these genomes10,58 are mainly based on Illumina technology and are therefore highly fragmented. A large number of scaffolds can result in multiple gaps and mis-assemblies, which can then hinder the understanding of genomic features such as chromosome rearrangements, gene duplications, repetitive regions, and changes in regulatory sequences.61,62 Thus, even if the functional content of these genomes (26,917–29,913 genes containing 92.7–94.9% of the core set of actinopterygian orthologs) is similar to that of the chromosome-level assemblies mentioned above, their lower quality might pose a challenge for certain types of analyses.\n\na) A comparison of genome contiguity for the three anemonefish chromosome-scale genomes (the false clownfish Amphiprion ocellaris, the orange clownfish Amphiprion percula, and the yellowtail clownfish Amphiprion clarkii) and 26 other previously published chromosome-scale fish genomes assemblies until 2019 (adapted from Lehman et al. 2019 and Hotaling et al. 2019). Data points are color-coded by order. b) Chromatin contact mapping takes advantage of the inverse relationship between proximity of nuclear DNA and genomic distance thus allowing contigs to be clustered into chromosomal groups. Here, the Hi-C heatmap of interactions between pairs of chromosomal loci (chr01-chr24) throughout the A. clarkii genome is shown. Interactions were drawn based on the chromatin interaction frequencies between pairs of 100 kb genomic regions (as determine by Hi-C). Darker red cells indicate stronger and more frequent interactions, which in turn imply that the two sequences are spatially close. Close-ups on the right show an overview of features revealed by Hi-C maps. Top squares show the long-range contact pattern of a locus (left) and its nuclear subcompartments (right). Middle squares show enhanced contact frequency along the diagonal (left) which indicate the presence of small domains of condensed chromatin (right). Bottom squares show peaks in the contact map (left) and the presence of loops that lie at domain boundaries and bind CTCF (right) (adapted from Rao et al. 2014). c) Availability of high-quality chromosome scale assemblies allow for large-scale genomic comparisons. Here, a dual synteny plot between all 24 chromosomes from A. ocellaris and A. percula shows conserved sequences within chromosomes of both species. Chromosomal rearrangements such as translocations and inversions are shown as red ribbons, whereas blue ribbons represent unchanged regions.\n\nCompared to second-generation sequencing technologies, third-generation sequencing platforms such as Pacific Biosciences (also known as single-molecule, real-time (SMRT) sequencing) and Oxford Nanopore Technologies (ONT) can produce long reads (5-50 kb for PacBio sequencing and up to the current record of 2.3 Mb for Nanopore reads63), thus making it possible to assemble genomes with higher contiguity and completeness. Indeed, inclusion of Nanopore reads together with Illumina data led to a 94% decrease in the number of scaffolds and an 18-fold increase in N50, and increased the genome completeness of A. ocellaris by an additional 16%.64 Moreover, the past decade has also seen the rise of chromatin contact (Hi-C) mapping to achieve chromosome-level assemblies.65 By crosslinking and fragmenting stretches of DNA that are physically close, and using short-read technology to paired-end sequence these fragments, the frequency distribution of how often two fragments of the genome interact (i.e., how physically close they are to one another) can be known and used to cluster contigs into chromosomal groups65 (Figure 3b). The contiguity of the A. percula assembly, the first chromosome-scale reference assembly for an anemonefish, improved dramatically following this approach. Scaffold N50 increased from 1.9 to 38.1 Mb, a more than 20-fold improvement, and over 1,000 contigs were placed into 24 chromosomal scaffolds.4,5 Furthermore, the recently published chromosome-scale assembly of the yellowtail clownfish A. clarkii35 has the highest quality and completeness (protein-coding genes encompassing 97.0% of conserved actinopterygian genes) of all published anemonefish genomes to date4,10,58,60,64 and is possibly in the upper echelons of all previously published fish genomes as well66–71 (Figure 3a).\n\nThe availability of genomes for multiple species has been critical to gain insights into the evolutionary history and adaptive radiation of anemonefishes. High-quality assemblies allow for comparative genomics and molecular evolution analyses on different traits of anemonefish. For example, using the first A. ocellaris long-read assembled genome,57 gene-editing with CRISPR/Cas9 became possible for the first time.51 The recently published chromosome-scale assembly of A. ocellaris60 revealed genomic elements conserved only in A. ocellaris and its sister species A. percula (Figure 3c). Importantly, the authors found that these elements are close to genes implicated in various nervous system functions and distinct expression patterns in the brain, potentially highlighting the genetic toolkits involved in lineage-specific divergence and behaviors of the ocellaris/percula branch. Comparative analysis using the A. clarkii genome35 identified higher copy numbers of the erbb3b gene. Notably, erbb3b encodes for an epidermal growth factor receptor (EGFR)-like tyrosinase kinase linked to melanophore development, suggesting then a possible link between this gene and the natural melanism polymorphism observed in A. clarkii.72 Studying anemonefish genomes has also enabled the identification of visual opsin genes and analysis of their synteny with the A. percula genome.59 There is evidence of two tandem duplication events involving the ultraviolet-sensitive (SWS1) opsin gene, of which two functionally-coding genes (SWS1α and SWS1β) are retained in the genomes of all anemonefishes. This is an exceptionally rare finding as most teleost fishes have lost this gene altogether.73 Last but not least, the genomes published by Marcionetti and colleagues (2019) have been crucial to identify genes under positive selection.10 Particularly, this study identified 17 genes at the origin of anemonefish radiation, two of which (versican core protein and protein O-GlcNAse) have been hypothesized to remove and/or mask N-acetylated sugars present in the skin mucus that protect anemonefish from getting stung by the sea anemone (reviewed in Section 3.2). Thus, providing the first insights into the genetic mechanisms of clownfish mutualism with sea anemones.\n\nThe first molecular insights of anemonefish biology came well before any of the genomes now available were sequenced. Instead, it came when the first study using quantitative polymerase chain reaction (qPCR) to investigate the role of the aromatase cyp19a1 gene on sex differentiation of the yellowtail clownfish A. clarkii was published in 2010,74 pioneering transcriptome expression research in anemonefish. Nowadays, applications of transcriptomics have rapidly expanded, and with it our understanding of the mechanistic underpinnings of various biological processes such as development, adaptive evolution, disease progression, and stress response. Indeed, the transcriptome is dynamic compared to the genome and is useful for dissecting the relationship between genotype and phenotype.75,76 RNA-seq and qPCR are low-cost methods that can provide high-resolution data without the need for extensive genomic resources.76 In anemonefish, transcriptome sequencing and qPCR experiments have provided insights into gene expression changes during development12,15,74,77–81 and adaptive responses to variations in the environment.21,82–84 Numerous efforts have contributed to the paramount abundance of transcriptomic data that is currently available for different tissues from multiple species and developmental stages (reviewed in Ref. 85), which has enabled the study of specific genes involved in a variety of functions including pigmentation,15,86 vision,59,87 thyroid hormone regulation of metabolism,77 and sex change.12,74,78,80,81\n\nRecent analysis of gene expression patterns across tissues in A. ocellaris60 and A. clarkii35 revealed interesting findings pertaining to the total number of genes and unique number of genes expressed per tissue. For example, ~15% of all genes (in both species) were expressed in nearly all tissues without biased expression (as determined by the tau index which quantifies tissue-specificity88) and therefore considered as housekeeping genes. On the other hand, less than 5% of all genes retrieved were absolutely specific genes (i.e., only expressed in one tissue type). Interestingly, the brain had the highest number of (total and unique) expressed genes, thus highlighting the complex role of this organ as the body control center. In A. ocellaris, some of the genes identified with higher expression levels in the brain are known to be involved in synapse formation in the central nervous system, chemo- and mechano-sensing of the environment, neuroplasticity, and development of spatial memory.60 Whilst future research should delve deeper on the characterization of these genes, their expression levels, and the roles they may play in anemonefish phenotypic traits, both studies35,60 provide the most accurate and complete transcriptomic atlas for two clownfish species to date.\n\nA main focus of anemonefish research has been the study of the transcriptomic programs of development. Particularly, several studies have investigated developmental gene expression related to sex change.12,74,78,80,81,89–93 After a first examination of the role of the aromatase gene during sex change in A. clarkii,74 Casas et al. (2016) was the first transcriptome-wide study to provide insights into the genetic mechanisms governing social sex change and gonadal restructuring in clownfish.12 Differential expression analyses in A. bicinctus revealed a complex genomic response of the brain associated with sex change that is subsequently transmitted to the gonads (see more in Section 4.1). Moreover, it identified a large number of genes, some of them well-known and others novel, that facilitated further research on sex change in anemonefish78,94 as well as in other hermaphrodite fish.95 The clownfish is a conspicuously colored species (possessing a bright orange body with three iridescent white bars bordered with black), and understanding the molecular basis of pigmentation has also become a fundamental question of evolutionary biology.2,14,16,96 Studies on A. ocellaris and A. percula have provided insights on how pigmentation patterns are phylogenetically conserved but also exhibit developmental and environmental plasticity.15,86 Other applications of transcriptomics in anemonefish research have provided in-depth characterization of visual opsins59,77,87 and the rhythmic expression of internal clock genes.79 More recently, Roux et al. (2022) provided a detailed, global overview of changes in gene expression across the post-embryonic development of the false clownfish.77 Transcriptomic analysis of each of the seven developmental stages of A. ocellaris has revealed three distinct phases: larval development, a pivotal stage that marks the onset of metamorphosis, and metamorphosis per se that corresponds to the actual transformation. By identifying expression patterns of genes specifically implicated in thyroid hormone and metabolic pathways, the authors describe how the morphological and physiological changes coupled with the ecological function of A. ocellaris in an integrative and coherent manner. Seeing that the environment can have significant effects on metamorphosis, especially during early stages where larvae and/or juvenile phenotypes can carry-over to later life stages, this study lays the foundation for further research investigating the adaptive potential of anemonefish to climate change.\n\nFinally, research has also examined changes in gene expression under different environmental stressors.21,82–84 Experiments using qPCR analyses have revealed the differential expression of genes strongly correlated to oxidative stress resulting from UV radiation83 and exposure to bisphenol A (BPA).84 This last study proposed the use of cytochrome P450 1A gene (cyp1a) as a biomarker for monitoring BPA pollution. Further, and in line with previous research investigating the impacts of elevated pCO2 on the brain transcriptome of coral reef fishes,97–99 Schunter et al. (2021) identified changes in the expression of genes related to circadian rhythm regulators and hormone pathways in A. percula subjected to diel pCO2 fluctuations.21 Notably, the authors suggest that environmental pCO2 fluctuations might enable reef fishes to phase-shift their clocks and adjust more successfully to ocean acidification conditions by “anticipating” pCO2 changes. Lastly, a new study has examined the molecular responses of A. ocellaris to the UV filter benzophenone-3 (BP-3) and found profound changes in the regulation of lipid metabolism that result in lipid accumulation in the liver of fish exposed to this long-time sunscreen ingredient.82 As the importance of anemonefish in developmental, evolutionary, and ecological research continues to grow, but also the availability of transcriptomic resources, it is only fair to assume that so will the number of studies investigating the impacts that environmental change has on this iconic group of fish. Integrating gene expression data with physiological and other molecular measurements will certainly provide key information on adaptive phenotypes.\n\nTo date, most research investigating physiological and behavioral changes of coral reef fish under warming and ocean acidification conditions has focused on transcriptomic and epigenetic modifications.97,99–105 Transcriptomic expression alone, however, is not sufficient to reflect protein levels and to therefore explain genotype-phenotype relationships.106 Measuring the presence and abundance of proteins is thus indispensable for the complete understanding of biological processes and cellular phenotypes, especially since post-translational modifications inferred from proteomics have been shown to be more strongly correlated to phenotypic observations than those from transcriptomics106–108 (Figure 1). Yet, technologies for quantifying the proteome are still lagging behind other “omics” fields.6 Until recently, only a couple of studies have measured changes in protein expression of a closely related species to anemonefish, exposed to elevated CO2.6,97,109 Proteomics can then be a powerful tool for identifying specific proteins and pathways that are crucial to stress responses, but more general for studying the evolution, biodiversity, and physiological adaptations of fish living in extreme environments.110\n\nConventional methods in proteomics first focused on isolating specific proteins to study their structure and function,107 which led to a very small number of intensely studied proteins over the past decades. Though protein biomarkers have facilitated a deeper understanding of various aspects of fish,110–113 the use of protein-based analyses changed when new technological advancements made it possible to accurately and reliably quantify amino acids at a proteome-wide scale.114,115 Indeed, mass spectrometry has become a mainstream analytical tool for proteomic profiling with diverse ecological applications.107 Proteomic techniques have been classified as shotgun, the optimal method for discovering more proteins but with the drawback that has reduced quantitative accuracy and reproducibility, or targeted, which is better for reproducibility if the proteins in question are known but limited in the number of measurements and therefore the number of peptides that can be identified.108,116 Particularly, iTraq (isobaric tags for relative and absolute quantification) labeled shotgun proteomics has become popular due to its use in non-model organisms. With this method, samples are labeled and processed together thus enabling the relative comparison of protein accumulation. A big limitation to this approach is, however, that the number of samples that can be compared directly is limited to a maximum of eight. Hence, pooling biological samples within one label is commonly done to increase the number of individuals that can be analyzed but results cannot be compared across experiments.117 Nonetheless, previous studies measuring protein responses with the iTraq method have done so on pooled samples and found distinct proteomic patterns in fish exposed to elevated CO2.97,109\n\nA newer method that combines the advantages of both shotgun and targeted proteomics is SWATH (sequential window acquisition of all theoretical spectra)-MS, a label-free strategy capable of quantifying thousands of proteins in a single measurement.114,115,118 The data are acquired on a fast, high-resolution mass spectrometer by cycling through sequential isolation windows over the entire chromatographic elution range.114,118 Since it is label-free, it is relatively cheap, and it has also been shown to have high reproducibility across different labs.115,119 SWATH-MS is versatile and has been for the quantifying proteins in a number of model organisms, diseases states, and bacteria, but also characterizing different post-translational modifications (reviewed in Refs. 119, 120). A recent study laid the groundwork for using this approach on a non-model fish species.6 It evaluated the performance of SWATH-MS in detecting significant proteomic expression differences in a complex experimental design of fish exposed to multiple climate change stressors. Most of all, the authors provided a guide on the efficiency, cost-effectiveness and applicability of this method in creating future proteomics references in non-model organisms aiming to identify genome-wide and ecologically relevant differential protein expression.6\n\nCertainly, the advancement of new techniques allows for the broad application of proteomics to study many aspects of anemonefishes. A few studies have investigated the proteomic responses of the spiny chromis damselfish Acanthochromis polyacanthus to ocean acidification,6,97,109 and one provided the proteomic profile of a sea anemone species from temperate seas.121 Seeing the potential of proteomics to identify ecologically relevant molecules and mechanisms, more studies should then focus on the application of these techniques to provide new insights that we have not yet obtained from genomic and/or transcriptomic expression alone. Proteomic data could unravel the processes driving symbiosis with the host anemones, sex change, complex social behaviors, and responses of anemonefish to environmental change, for example.\n\nThe combination of “omics” technologies can provide without question a wider vision of the organism of study and indicate the direction of future research. Integrating data from as many levels of information (from gene regulatory networks, RNA and protein measurements, metabolites and cell-cell interactions, to individuals, populations and ecologies) as possible is the ultimate goal of modern systems biology approaches.122 In situ hybridization, for example, is a powerful approach for studying the temporal and spatial patterns of specific genes especially because not only it enables maximum use of tissue that is difficult to obtain but can be frozen for future use.123 Though protocols were originally established in zebrafish, in situ hybridization has also been successfully performed in anemonefish embryos124 and different tissues.15,74,78,80,125 Importantly, this technique revealed unique aspects of the embryogenesis of the tomato clownfish A. frenatus that suggest an evolutionary adaptability of the teleost developmental program.124 Similarly, fluorescent in situ hybridization (FISH) has provided a detailed understanding of the visual system of various anemonefish species by visualizing and quantifying patterns in opsin gene expression.59,87\n\nCommercial enzyme immunoassay (EIA) kits have also proven to be a useful method to detect and quantify specific molecules. It is fast, simple, and cost-effective, and it has already been validated for measuring hormone concentrations in several species of anemonefish.77,126,127 Particularly, the measurement of thyroid hormones in A. ocellaris has revealed an important link with metabolic regulation, morphological transformation, and behavioral changes during the transition of pelagic larvae and benthic reef associated juveniles.77 Functional studies are possible now due to the development of a “low-volume” rearing protocol for anemonefish larvae, which allows the use of pharmacological approaches to alter specific biological pathways.49 Experiments testing different drugs have been conducted to investigate the metamorphosis and pigmentation changes through larval development of A. ocellaris.15,77 Cell lines have assumed an importance in molecular studies as well, especially for genetic manipulation.128 Yet, so far there is only one report on cell culture from anemonefish explants.129 Patkaew and colleagues (2014) described a simple and reliable method to for culturing A. ocellaris cells using vertebrae explants. Cytogenetic studies have further contributed to different fields of fish biology by providing basic information on the number, size and morphology of chromosomes.130 Karyological analyses have been done for several anemonefish species131–134 and they have consistently revealed 24 chromosomes.\n\nAs of today, no quantitative trait locus (QTL) analysis or genome-wide association studies (GWAS) have been performed in anemonefishes, thus limiting the potential for forward genetic studies (reviewed in Ref. 85). However, following a transcriptomic analysis of the mechanisms involved in sex differentiation in the Red Sea clownfish, A. bicinctus,12 the same authors published a high-density genetic map for this species.13 Essentially, this map provides a platform to study the main gene regulatory networks governing social sex change in anemonefish and other protandrous fish as well. Finally, a gene-editing protocol for applying the CRISPR/Cas9 system was recently developed in A. ocellaris (Figure 1). Micro-injection of eggs was used to demonstrate the successful use of this approach at two separate target sites with 75–100% efficiency in producing biallelic F0 mutants.51 Specifically, CRISPR/Cas9 knockout of the tyrosinase encoding gene (tyr) involved in melanin production resulted in embryos exhibiting varying degrees of hypomelanism, thus clearly showing a loss-of-function.51 This is undoubtedly a steppingstone for reverse genetic studies with exciting prospects to study the genetic basis of various unique traits of anemonefishes.\n\n\n3. Elucidating the genomic basis of adaptation\n\nAnemonefish are an extraordinary example of adaptive radiation, a process driven, in this case, by the mutualistic relationship they maintain with sea anemones.135 Indeed, whilst ubiquitously distributed across the tropical Indo-Pacific Ocean, anemonefish have occupied different ecological niches according to the habitat preference (e.g., reef zonation, substrate, depth) of their host sea anemones. The distribution and abundance of clownfish are thus strongly dependent on the distribution and abundance of sea anemones.135,136 Coexistence of multiple anemonefish species is in fact possible because of difference in host and habitat utilization.34 The effect of mutualism on clownfish diversification was first examined using different nuclear and mitochondrial gene regions (e.g., 12S, 16S, ATP6-8, COI, cytochrome b, ND3, BMP-4, RAG1, RAG2),8,9,135,137,138 but now the availability of high-quality genomes4,10,35,57,58,60 has further clarified the phylogenetic relationships between anemonefish.\n\nWhile most phylogenetic inferences8,9,135,137,138 agree in the overall placement of six major species groups (the ocellaris/percula clade, the Australian clade, the skunk anemonefishes (known as the akallopisos group), the “ephippium” complex, the polymnus group, the clarkii, and the Indian clade), some discordance has been observed between mitochondrial and nuclear trees8 with the main difference being the positioning of the maroon clownfish Premnas biaculeatus (reviewed in Refs. 23, 139). It has been long accepted to place P. biaculeatus together with the ocellaris/percula clade and separately from the rest of anemonefish (Figure 2a),8,10,135–137 but recent analysis23,60,140 and a thorough systematic analysis of 322 damselfish species24 have suggested that Premnas is, in fact, related to A. ocellaris and A. percula and should not be separated from the genus Amphiprion. The latter study reported a level of divergence within the range of what is observed between Amphiprion species,24 which was further reinforced by Salamin and colleagues (2022), who found that gene trees estimated from 100 kb windows display an ambiguous placement for Premnas (either as a sister species to the ocellaris/percula clade or at the base of the tree).23\n\nCertainly, establishing a well-resolved phylogeny is critically important to understanding the evolution and genomic underpinning of anemonefish lifestyle. Nonetheless, despite the topological inconsistencies mentioned above, these studies have provided impressive insights into the adaptive radiation of clownfish. Litios et al. (2012) were the first to show higher rates of speciation and diversification for clownfish compared to their closest relatives without anemone mutualistic associations. Similarly, their findings also revealed a strong link between the appearance of mutualism and increased morphological evolution.7 Following this study, the same authors inferred the effect of the geographical range of species on the diversification of clownfish by implementing geographic state speciation and extinction models on phylogenetic reconstructions.9 Results of this study showed that most species originated in the Indo-Malay Archipelago, with one independent radiation event along the eastern coast of Africa (including the Red Sea, Maldives, and central Indian Ocean) that gave rise to seven species that now span the whole range of possible associations with sea anemones. This is interesting as instances of replicated ecological speciation over large geographic scales (of the marine realm) are quite rare, most examples being found on islands or lakes.141–143\n\nWhilst ecological speciation is likely to be the main driver of clownfish diversification, it is not the sole factor. A study by Tim and colleagues (2008) showed clear geographical subdivisions (up to ~19% of sequence divergence) in A. percula from Papua New Guinea and the Solomon Islands, thus suggesting that novel species might be arising by parapatric means in a region where partial isolation between subregions reinforces isolation along genetic and ecological gradients.144 Hybridization has also played an important role in the evolution of anemonefish8,11,41 and the several known hybrid species37–40 show that it is an ongoing process. Increased glaciations and low sea levels during the Pleistocene likely promoted hybridization of many coral reef associated fish species,37,144 and in the case of anemonefish this process may be further facilitated by the presence of co-existing, closely related species within the same sea anemone hosts.33,145 Unlike other fish, in which it is not known what the parent species are, how often they come into contact or whether the resulting hybrids interbreed with one or both parent species, anemonefish provide a unique opportunity to understand how patterns of hybridization and introgression can be controlled by resource use and reproductive behaviour (reviewed in Ref. 37). Parent species have specific habitat requirements and may only interbreed where they overlap and co-occur. For example, the skunk clownfish A. sandaracinos and orange-fin anemonefish A. chrysopterus, which distribution overlaps in the northwestern regions of Papua New Guinea and the Solomon Islands and can co-habit in H. crispa and S. mertensii anemones, have been described as the putative parent species of the natural hybrid A. leucokranos37 (Figure 2c). Several other Amphiprion species appear to be hybridizing as well, such as A. bicinctus and A. omanensis in Socotra Island146 and the historical hybridization that occurred between A. mccullochi and A. akindynos in southern Australia.40 Notably, the species A. thiellei (probably resulting from A. sandaracinos and A. ocellaris)3,147 is still under debate as there is no definitive genomic proof of its hybrid condition.42 Previous studies were based on limited genetic data and did not include all species, but as progress is made in this field and availability of genomic data increases, new species may be described in the years to come.\n\nIndeed, new whole-genomic data for all 28 species has confirmed multiple past hybridization events throughout the evolutionary history of anemonefishes.41 The findings of Schmid and colleagues (2022) also shed light on the functional role of introgressive hybridization during clownfish adaptive radiation. Specifically, they show distinct phylogenetic and introgression patterns in chromosome 18 compared to the rest of the genome. This is interesting as it potentially indicates that the introgression signal was removed from the rest of the genome by extensive backcrossing but persisted on chromosome 18. This was through the disruption of recombination, and genomic inversions that break recombination and are known for creating clusters of loci controlling ecologically important traits that are consequently fixed by natural selection (as it is the case of supergenes).41,139 For example, the authors noted that genes in this chromosome are associated with the nervous system and embryonic development, and DNA damage and external stressors responses, which they suggest could be linked to advantageous traits involved in local adaptation and pre-/postzygotic isolation.41 Further genomic studies are needed, however, to better characterize the various chromosomal rearrangements and the role they played in the evolution and diversification of anemonefish.\n\nSimilar to the cichlid fishes, which are famous for their large, diverse, and replicated adaptive radiations in the Great Lakes of East Africa,142 a recent study found that anemonefish genomes also show major bursts of transposable elements (TE) and accelerating coding evolution.11 Given that a large fraction (20–25%) of clownfish genomes consist of TE,4,11,35,60 and that transposition bursts are common in teleost fishes,148,149 these findings are to be expected. TE have been proposed to contribute to adaptation, speciation, and diversification processes,148,149 and they may also be associated with interspecific hybridization.150 Marcionetti and Salamin (2022) thus suggest that the high percentage of TE in clownfish genomes originated from two bursts of transpositions, which in turn might have played a key role in the adaptive radiation of anemonefishes.11 The authors also detected increased evolutionary rates and positively selected genes (~5% of the genome) including genes with functions linked to clownfish social behavior and ecology. Surprisingly, this study did not find an excess of gene duplications in anemonefish, a remarkable finding as gene duplication has been shown to be critical for genome evolution and adaptive radiation of fish like the African cichlids.142,148 Furthermore, Marcionetti and Salamin (2022) go a step further by examining the evolutionary rates and selective pressures of genes involved in the ecological divergence of clownfishes (i.e., specialist and generalist species). Altogether, the results of this study are extraordinary as they lay the foundation to understand the genomic substrate of anemonefish adaptive radiation but also open an avenue for future research investigating the genomic mechanisms governing species diversification.\n\nClownfish and sea anemones are perhaps one of nature’s most iconic duos. This mutualistic relationship has long fascinated biologists (first observations date back to 1868151) and it has become the subject of more recent studies investigating the evolutionary history of anemonefishes (reviewed in Section 3.1). Two aspects of this association make it particularly interesting to study: first, anemonefish can inhabit sea anemones without being harmed (unlike other fish that can be killed), and second, there is a complex species-specificity of this symbiotic relationship between the 28 species of clownfish and the 10 possible sea anemone hosts (probably related to the toxicity levels of the anemone).2 The mechanism(s) underlying this mutualism remains poorly understood, but two conflicting hypotheses have been proposed to explain how anemonefish are able to live safely in their host (reviewed in152,153). One hypothesis proposes that anemonefish acquire certain components (e.g., antigens) of the anemone mucus that protect them from being stung (i.e., chemical camouflage).154,155 Indeed, an early study156 found that the mucus coating of the fish changes during the behavioral process of acclimation to resemble that of the anemone. The other hypothesis suggests that clownfish produce their own protective mucus,157–159 which either prevents nematocyst discharge by the host160,161 or protects the fish from the consequences of the sting.162,163 Particularly, N-acetylneuraminic acid (Neu5Ac), a member of the sialic acids family, has been recognized to have a critical role in the chemical recognition of the host.157,161 Indeed, it has been shown that clownfish mucus lacks this sialic acid (e.g., 1.6 mg/mL in A. ocellaris compared to 50.4 and 71.89 mg/mL in other reef fish species), making them “invisible” to the anemone and thus avoiding being stung.161 Altogether, the above certainly suggests that the mucus of both the fish and host anemone is key for the success of this association (Figure 4). Studies investigating the anemonefish mutualistic relationship have brought insights into the biochemical mechanisms developed by clownfish to avoid being stung by the sea anemone nematocysts155,156,160,161,163 as well as the variable host specificity displayed by different Amphiprion species and developmental stages.159,164–166 More recently, studies are leveraging the power of next-generation sequencing technologies to better understand the genetic basis10 and potential microbial role in clownfish adaptation to sea anemones.17–19,167\n\nTwo conflicting hypotheses have been proposed to explain how anemonefish are able to live safely in their host: 1) CF acquire antigens of the SA mucus that protects them from being stung, and 2) CF produce their own protective mucus, which either prevents nematocyst discharge by the host or protects the fish from the consequences of the sting. Particularly, N-acetylneuraminic acid (Neu5Ac) might have a critical role in the chemical recognition of the host.\n\nOnly recently, candidate genes that may grant anemonefish protection from the host toxins were identified for the first time.10 Marcionetti and colleagues (2019) highlighted the genes versican core protein and protein O-GlcNAse as particularly interesting due to their functional link with N-acetylated sugars.10 Versican core protein is known to be a critical extracellular matrix regulator of immunity and inflammation168 that interacts with several matrix molecules including glycosaminoglycans such as N-acetylglucosamine (GlcNAc).169 Expression of versican core protein in clownfish skin is thought to bind to N-acetylated sugars, masking their detection by the host chemoreceptors and therefore preventing nematocyst discharge (reviewed in Ref. 10). On the other hand, protein O-GlcNAse has the potential to cleave N-acetylated sugars from different cell surface molecules170 and has also been found to be expressed in anemonefish epidermis.10 Additionally, clownfish-specific duplicated genes involved in immunity (e.g., T cell receptor alpha) and detoxification (e.g., cytochrome P450, glutathione S-transferasas) responses were also identified.10 Conclusions regarding the potential role of these genes in the protection from anemone secreted toxins cannot be drawn, however, without further experimental evidence.\n\nGiven that the skin is the first line of physical contact between clownfish and sea anemones, and that epithelial microbial communities are important drivers of symbiotic interactions,171 more studies are now investigating the microbial signatures of the clownfish-sea anemone mutualism.17–19,167 Pratte et al. (2018) were the first to show that contact with host anemones can significantly reshape the clownfish skin microbiome.17 Their findings revealed a drastic shift in the epithelial bacterial communities of A. clarkii within one week of association with their host. Interestingly, they also showed that these changes are reversible. Following this study, Roux and colleagues (2019) examined the microbiome of A. ocellaris and H. magnifica mucus simultaneously before and after initiation of the symbiotic relationship.18 The authors found distinct microbial signatures for each symbiont before initial contact (e.g., Alteromonadaceae dominated the clownfish skin bacterial taxa whereas sea anemone mucus was mainly composed of Pseudoalteromonadaceae and Endozoicomonadaceae) that were subsequently modified during the establishment of symbiosis (e.g., fish-anemone microbiota shared the families Haliangaceae, Pseudoalteromonadaceae, and Saprospiraceae). Until then, this was the only study to have tested the effect of clownfish association in the mucosal microbiota of the sea anemone and shown microbial convergence between both partners (but see Refs. 19, 167). Notably, the latter seems to substantiate the hypothesis that anemonefish cover themselves with their host mucus to avoid being stung.154,155\n\nA more recent study examined shifts on the skin microbiota in clownfish when in direct contact with its host (i.e., fish and anemone are in the same tank) but also tested the effect of “remote interaction” (i.e., fish and anemone are in separate tanks, both connected to the same water flow) on the epithelial microbiome restructuration in both partners.19 The results of this study are compelling as they provide evidence of a strong water-mediated chemical communication between both symbiotic partners (as seen by the gradual convergence in the microbial communities of the fish and its host when they are both placed in the same water system). Interestingly, increasing abundances of three sequence variants closely related to a tyrosinase-producing Cellulophaga tyrosinoxydans bacterium were observed during microbiota convergence. Noteworthy, bacterial tyrosinases (which catalyze melanin synthesis) have been shown to be immunologically active compounds, providing skin protection against radiation, viral agents, immunogens, and/or toxic compounds.86,172 Whether convergence of microbial communities might play any role in the symbiotic relationship between clownfish and sea anemones remains to be determined nonetheless. Metagenomic and metatranscriptomic approaches would be the next step to gain a deeper understanding on specific gene functions and expression patterns of the microbial communities involved in this mutualism. Finally, it is worth mentioning a study by Titus et al. (2020) as it shows for the first time the effect of host identity and symbiotic association on the functional diversity and composition of the microbiome.167 Microbiota of different anemones (C. adhaesivum, E. quadricolor, H. aurora, H. magnifica, and S. mertensii) harboring the same species of clownfish (A. nigripes or A. clarkii) were more similar to each other than to that of anemones that were hosts to different species of anemonefish. Furthermore, this is the only study examining in situ microbiomes so far. Experiments in field conditions are needed to ultimately establish the role of microbial communities in the clownfish-sea anemone symbioses.\n\nAdaptation to changing environments has long been a central question in evolution.173 Phenotypic plasticity, the ability of a species to produce multiple phenotypes (e.g., alternative morphology, physiological state, behavioral response) from a single genotype, has been shown in many terrestrial and aquatic organisms and is critically important for adaptation of populations to local environments. Environmentally induced non-genetic effects on phenotypes can alter the strength and direction of selection affecting transmitted gene frequencies by shifting the range of phenotypes expressed. In such cases, a phenotype, which initially is produced only in response to a specific environment, becomes assimilated genetically so that it is formed even in the absence of the environmental influence that had been necessary before (reviewed in Refs. 173, 174). In anemonefish, color polymorphisms within populations have received considerable attention and have been attributed to developmental plasticity (reviewed in Ref. 175). The latter referring to the ubiquitous ability to adjust phenotypic development in response to environmental cues experienced in early life stages.176\n\nPhenotypic (developmental) plasticity as a phenomenon enables the study of the link between gene expression and phenotype since it involves the production of various phenotypes without genetic changes. Species with adult individuals that can be experimentally induced to transition between distinct phenotypes are notably valuable as they make it possible to isolate phenotypic effects of gene expression by comparing the gene expression profiles of groups of individuals who differ in their phenotypes due to plasticity rather than genetic differences.177 Such is the case of the yellowtail clownfish A. clarkii, for example, a species known for showing a high degree of melanism polymorphism.72 Particularly, melanism in A. clarkii varies with social rank,36,178 local variations in habitat (e.g., temperature),179 and host anemone.72 Other species of anemonefish (A. chrysopterus, A. percula, A. polymnus) also exhibit polymorphic melanistic morphs depending on the host they associate with. Indeed, observations have noted that fish inhabiting Stichodactyla spp. are generally darker (i.e., more melanic), whereas individuals in other anemones (e.g., Heteractis spp., Entacmaea quadricolor) tend to be more orange.72 Moreover, Salis et al. (2021) recently showed that the developmental timing of white bar formation in juvenile A. percula depends on the anemone species to which they have recruited.20 Specifically, earlier formation of white bars when clownfish developed in Stichodactyla gigantea rather than Heteractis magnifica was observed (Figure 5a). Using a combined approach of transcriptomic analysis and pharmacological treatments, the authors showed that thyroid hormones are essential in modulating the timing of adult color pattern formation and which are, in turn, associated with ecological differences. This study offers great promises to understand the genomic and developmental basis of plastic phenotypes observed in wild clownfish.\n\na) Color patterns in anemonefish can vary greatly depending on their ecology, development, and evolution (adapted from Refs. 15, 175). White bars could be necessary for species recognition and could be adaptive for camouflage or even used as an aposematic signal. Pigmentation polymorphisms have also been observed in Amphiprion percula living in Heteractis or Stichodactyla anemones: 1) juveniles in Heteractis exhibit a delayed white bar formation and 2) adults in Stichodactyla show higher melanism. The three white bars arise sequentially from anterior to posterior body parts during ontogenesis whereas during evolution, bars are lost in the opposite sequence of ontogenesis (from the posterior to anterior region). b) Natural melanism polymorphism observed in Amphiprion clarkii (adapted from Ref. 35). c) Examples of Amphiprion ocellaris color mutants available from aquaculture companies: naked phenotypes (“Naked” and “Extreme Misbar”), phenotypes with extra white markings (“Gladiator/Da Vinci”) to an almost nearly complete white colored body (“Wyoming White”), melanic phenotypes (“Black/Darwin” and “Domino”), and phenotypes with irregular patterning (“Snowflake” and the Premnas biaculeatus “Lightning”) (adapted from Ref. 16).\n\n\n4. Genomic architecture underlying anemonefish phenotypes\n\nFish exhibit extraordinary sexual plasticity, changing sex naturally as part of their life cycle12,180 or because of environmental stressors.181,182 Indeed, sequential hermaphroditism has been reported for at least 27 teleost families spread across nine orders in three different forms: protogynous (i.e., female to male), protandrous (i.e., male to female), or bidirectional sex change (reviewed in Ref. 183). From these, protandry is rarer among teleosts, occurring sporadically across six families including anemonefish.183 Anemonefish are particularly interesting, as contrary to most protandrous species, which need to attain a threshold age/size to change sex, their sex change is regulated socially.94 The first molecular characterization of sex change in anemonefish dates to the late 2000s when Miura and colleagues (2008) performed immunohistochemical detection to examine the expression of the aromatase gene (cyp19a1) during gonadal development of A. clarkii.184 Soon after, numerous studies started investigating the expression patterns of different hormones involved in sex change by using qPCR.74,80,81,89–93 However, it was not until the year 2016 that Casas et al. explored the transcriptome-wide expression landscape during sex change in a clownfish.12 To this, followed a high-density map containing tens of genes involved in sex differentiation.13 Collectively, these studies identified several genes (and their location) that may be important orchestrators of sex change and gonadal transformation in anemonefish. Importantly, these studies are also shedding new light on the gene regulatory mechanisms underlying functional hermaphroditism in fish.94,95,183\n\nTranscriptomic analysis of the Red Sea clownfish A. bicinctus showed a gradual decline in male-related gene expression and up-regulation of female-pathway genes as the gonads transitioned from ovotestis to ovaries (reviewed in Refs.12, 94, 95). Active feminization of the (male) brain starts two weeks after the female is removed (i.e., social cue) so that the transcriptional response is subsequently transmitted to the gonads where differential expression and histological changes can be clearly observed after three to four weeks. Specifically, cyp19a1 (steroidogenic enzyme operating in the female pathway by converting androgens into estrogens) exhibited increasing expression in the brain of transitional males. This might be, at the same time, regulated by the expression of two other genes: sox6 (transcription factor involved in spermatogenesis) and foxp4 (transcription regulator with an important role in sexual development). A specific mechanism of action remains to be established though. Changes at the gonadal level, on the other hand, are also driven by the over-expression of cyp19a1 which then triggers the up-regulation and down-regulation of foxl2 (transcription factor involved in ovarian differentiation) and dmrt1 (gene involved in the development and regression of testis), respectively. Based on this, Casas et al. (2016) propose a feedback loop combining transcriptional regulation with steroid hormonal activity where dmrt1 and foxl2 regulate the production of cyp19a1 and thereby, gonadal restructuring during sex change in clownfish.12\n\nImportantly, the same molecular pathways have been described for another Amphiprion species.78 Wang and colleagues (2022) also conducted comparative transcriptomic analysis of gonads of the complete social group (females, males, and non-breeders). In addition to this, they also performed in situ hybridization to show expression localization of foxl2 and dmrt1. Consistent with previous findings, foxl2 could only be detected in the granulosa cells of oocytes in female gonads, whereas signals of dmrt1 were detected in spermatogonia and spermatocytes in male gonads.78 New research using exogenous steroid (i.e., estradiol and cortisol) treatments combined with qPCR have further validated the role of cyp19a1 in feminization in A. ocellaris.81 Particularly, cortisol pellet-fed fish exhibited a decline in expression of cyp19a1 and dominant behavior intensity. The authors showed that high cortisol concentrations inhibit transcription of the aromatase gene, which results in masculinization. Thus, suggesting that the interaction between cortisol and aromatase might play a pivotal role in sex change of anemonefish.\n\nColor patterns in adult fish provide a unique opportunity to study the interplay between ecology, development, and genetics that is the basis for trait diversification (reviewed in Ref. 96). Indeed, fish have the highest number of pigment cell types (chromatophores) and color diversity among vertebrates, which is not surprising considering the ample pigmentation gene repertoire they have as a result of undergoing a third (and fourth, in the case of salmonids) whole genome duplication round.185,186 Body coloration of fish varies greatly, different species and/or life stages display diverse combinations of spots, stripes, bands, eyespots, etc.96 that can also change depending on environmental cues and geographic distribution20,72,178,179,187–189 (Figure 5a). Certainly, color patterns have clear ecological and behavioral significance, with functions ranging from recognition of conspecifics, to avoidance of predators, sexual attraction, and protection against ultraviolet radiation.96 In anemonefish, pigmentation plays a key role in the complex hierarchical social system they have. Young recruits are colored distinctly different than older juveniles to potentially avoid antagonistic and aggressive behaviors from the larger individuals (this is yet to be confirmed with behavioral experiments).175 Loss of white vertical bars during ontogeny has indeed been observed in multiple Amphiprion species.14\n\nSalis and colleagues (2018) mapped the occurrence of these bars throughout the phylogenetic tree and showed that the diversification of color patterns in anemonefish is the result of successive (posterior to anterior) losses of bars during clownfish radiation. The sequential appearance/disappearance of white bars during the development of distantly related species is remarkable as it suggests a highly conserved mechanism of pigmentation pattern ontogeny across anemonefish14 (Figure 5a). Different phylogenetic approaches have also shown evolutionary pathways linking the number of bars with host specificity and host toxicity (i.e., fish with fewer bars associate with fewer and more toxic anemone species than fish with higher number of bars).190 Color polymorphisms are also known to occur frequently in anemonefish (also see Section 3.3), whether they are rare natural variants43,72 or mutants found in pet shops.16 Widely distributed species such as A. clarkii exhibit great intraspecific polymorphism for melanic pigmentation according to geographical variation and environmental conditions26,72,178,179 (Figure 5b). Interestingly, a recent study revealed higher copy numbers of the receptor protein kinase erbb3b gene (which is involved in melanocyte development) in A. clarkii compared to other anemonefish, thus implying a possible link between erbb3b and the natural melanism polymorphism observed in this species.35 Morphotypes such as albinism or individuals with no bands in species that usually have, are never or very rarely observed in the wild but can be found in the aquarium trade industry (reviewed in Ref. 16). In the wild, mutations that result in such drastic color pattern alterations have a negative effect on the survival of individuals and are therefore negatively selected against, but they can be bred for several generations in aquaculture. As global trade in ornamental fish has become a multi-billion dollar industry (reviewed in Ref. 191), many color mutations have been characterized.16 For example, mutations related to “naked” phenotypes (i.e., absence of white bars) may be specifically caused by genes such as ltk (leucocyte tyrosine kinase), sox10 (SRY-related HMG-box) and endothelin receptors edn3b and ednr3b that are responsible for iridophore specification. Klann et al. (2021) further review the mutations underlying many of the pigmentation variants known for anemonefish until now and present a global picture of their origins and crosses16 (Figure 5c).\n\nStudying pigmentation also allows further understanding of the cellular basis of adult form, as the cells that produce diverse color patterns are readily visible in the skin during development.15,54 Thus far, however, genetic and cellular mechanisms of lineage specification, differentiation, and morphogenesis during pigment pattern formation have been studied most extensively in zebrafish (reviewed in Ref. 192). It is only until recently that Salis et al. (2018, 2021) described in detail the emergence of pigmentation during embryonic development in the anemonefishes A. ocellaris and A. perideraion.54,193 In another study, the same authors also investigated the molecular basis of white barring in clownfish.15 Using transcriptomic approaches, they showed that white skin in clownfish have a transcriptomic signature of purine-containing iridophores, similar zebrafish and oppose to leucophores in medaka, for example. Particularly, four genes (fhl2a – four and a half LIM domains 2a, pnp4a – purine nucleoside phosphorylase 4a, prtfdc1 – phosphoribosyl transferase domain containing 1b, and tfec – transcription factor EC) were inferred to be essential for the development and function of iridophores. Great progress in identifying the genetic and cellular bases of pigment patterns formation in anemonefish is being made in our laboratory, nonetheless there is still much to understand. Beyond zebrafish, a classical fish model for vertebrate biology, we are only just beginning to understand how the molecular mechanisms underlying the diverse pigmentation in clownfish and other teleosts, but the emerging genomic and imaging technologies offer a promising future in this field.\n\nThe evolutionary theory of aging predicts that individuals with low extrinsic mortality will show delayed senescence (i.e., the process of physiological deterioration with age) and increased lifespan (reviewed in Ref. 194). Here, low extrinsic mortality is correlated to the low predatory pressure anemonefish face thanks to their association with sea anemones (which provides protection from predation). Indeed, the overall rate of mortality amongst anemonefish is low compared to other coral reef fish, thus leading to clownfish having slow aging and increased longevity.195 Buston and García (2007) estimated a life expectancy of 30 years for wild A. percula,195 and similarly for A. ocellaris and A. melanopus in captivity.196 Noteworthy, this estimate is two times greater than the longevity estimated for any other coral reef fish and up to six times greater than the longevity expected for a fish of that size.195 Anemonefish therefore stand out as quite unique under this criterion also.\n\nTranscriptome sequencing of several Amphiprion species196 revealed 157 positively selected genes, several of which are related to processes linked to xenobiotic and glutathione metabolism, detoxification, mitochondrial translation, inflammation, and autophagy. In particular, the authors found a positive selection of two lysosomal membrane proteins (LAMP2 and CD63) known for playing an important role in chaperone-mediated autophagy, lysosomal protein degradation, adaptive immune response, and apoptosis. These results are consistent with earlier findings that have associated lysosomal function as one of the key hallmarks of aging,194 thus implying that positive selection of lysosomal genes plays an important role in the evolution of exceptionally long life of anemonefish.196 Interestingly, this study also showed evolutionary convergence with the short-lived killifish and the long-lived mole rat. Signs of convergence were observed for genes and pathways involved in the biogenesis of mitochondrially-encoded proteins with the remarkable observation that MTERF (mitochondrial transcription termination factor 1) is under positive selection in all three taxa. This parallels previous evidence suggesting mitochondrial biogenesis as a core genetic substrate in the evolution of lifespan.194 Furthermore, the observation that the same pathway is under positive selection in both exceptionally short- and long-lived species indicates that the same genetic architecture underlies both evolution of lifespan and longevity.194,196 Altogether, this makes anemonefish the first long-living fish model for aging research.197\n\n\n5. Evolutionary genomics of complex traits in anemonefish\n\nAdding to the many traits that make these fishes fascinating, anemonefish also exhibit interesting behaviors such as social group formation and parental care. The mechanisms involved in social evolution in clownfishes, and more specifically the interspecific variation in the genetic benefits and ecological constraints of forming social groups, have been a major focus of study in anemonefish research (reviewed in Ref. 198). Parental behaviors in anemonefish have also been well described, with a growing number of studies investigating the neural pathways and brain regions regulating parental care. Particularly, there is an interest in understanding the plasticity of parental care in response to changes in ecological context (i.e., resource availability) and social roles (across sex change) (reviewed in Ref. 199). Behavioral genomics is still in its infancy; the complexity of individual and group behaviors, and the highly polygenic nature of these traits, make it challenging to study the mechanistic links between genes and behavior. Nonetheless, with the recent advent of (affordable) “omics” approaches, it is increasingly possible to identify the precise genetic contributors of a wide diversity of behaviors, allowing for new insights into how behavior evolves in the wild. More studies are now focusing on investigating the role of genetic/genomic variation (from DNA sequences to brain gene expression, to neuronal dynamics, to gene regulatory networks) to understand the genetic/genomic bases of behavior.200 The establishment of anemonefish as a model organism in different biological disciplines, and the availability of high-quality reference genomes and transcriptomic data will certainly facilitate the quest for answers on clownfish behaviors. Seeing that behavioral traits are complex, carefully designed experiments are needed to disentangle individual and group behaviors. The plastic nature of behavior and the likelihood that many genes with small effects are involved also makes quantifying the role of natural genetic variation difficult.201 Newer tools have been developed to test candidate genes with behavioral functions. James and Bell (2021) used virus-mediated transgenesis (through direct brain injection) to study how overexpression of certain genes affected aggressive behaviors in the fish model Gasterosteus aculeatus.202 Indeed, species such as the threespine stickleback and zebrafish have long been the subject of behavioral genetics,202–206 providing valuable insights that can potentially be transferred to anemonefish.\n\n\n6. Conclusions and perspectives\n\nAnemonefish have become an invaluable model system for answering some of the most fundamental and long-standing questions in evolutionary genomics. Future research will most likely be more integrative, incorporating not only the topics discussed here, but also other fields such as ecotoxicology and neuroendocrinology, as well as continued integration with ecology and behavior. Recent technological advancements have facilitated the generation of huge amounts of genomic, transcriptomic, and proteomic data that can be leveraged to answer complex questions pertaining to the many traits that make anemonefish extraordinary. Importantly, one of these traits is that, compared to most marine fish, anemonefish (almost) never abandon their host sea anemone thus making them ideal subjects for long-term monitoring studies. Indeed, the first multigenerational pedigree for a marine fish population was constructed using data from a 10-year genetic survey of A. percula from Kimbe Bay in Papua New Guinea.48 Such genealogy provides a unique opportunity to study how maternal effect, environment or philopatry can shape wild fish populations, for example. Long-term genomic monitoring will certainly become a powerful tool to assess species and ecosystem vulnerability to environmental change. Anemonefishes are becoming a mainstay to study adaptive responses of marine fish to climate change and ocean acidification, a body of work that will only continue to grow. Finally, anemonefishes are now strongly positioned to exploit rapidly emerging tools such as CRISPR/Cas9 which will be crucial to gain insights into the molecular basis underlying specific phenotypes and genetic variants.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nAcknowledgements\n\nWe thank Natacha Roux for her valuable comments that greatly improved this manuscript and help with the elaboration of the figures.\n\n\nReferences\n\nMcGee MD, Borstein SR, Meier JI, et al.: The ecological and genomic basis of explosive adaptive radiation. Nature. 2020 Oct 1; 586(7827): 75–79. PubMed Abstract \| Publisher Full Text\n\nRoux N, Salis P, Lee SH, et al.: Anemonefish, a model for Eco-Evo-Devo. EvoDevo. 2020 Oct 7; 11(1): 20. 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Publisher Full Text\n\nBalamurugan J, Kumar T, Kannan R, et al.: Acclimation behaviour and bio-chemical changes during anemonefish (Amphiprion sebae) and sea anemone (Stichodactyla haddoni) symbiosis. Symbiosis. 2014; 64(3): 127–138. Publisher Full Text\n\nElliott JK, Mariscal RN: Acclimation or innate protection of anemonefishes from sea anemones? Copeia. 1997; 1997: 284–289. Publisher Full Text\n\nElliott JK, Mariscal RN: Ontogenetic and interspecific variation in the protection of anemonefishes from sea anemones. J. Exp. Mar. Biol. Ecol. 1997; 208(1–2): 57–72. Publisher Full Text\n\nMebs D: Anemonefish symbiosis: vulnerability and resistance of fish to the toxin of the sea anemone. Toxicon. 1994; 32(9): 1059–1068. PubMed Abstract \| Publisher Full Text\n\nTitus BM, Laroche R, Rodríguez E, et al.: Host identity and symbiotic association affects the taxonomic and functional diversity of the clownfish-hosting sea anemone microbiome. Biol. Lett. 2020 Feb 26; 16(2): 20190738. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWight TN, Kang I, Evanko SP, et al.: Versican—A Critical Extracellular Matrix Regulator of Immunity and Inflammation. Front. Immunol. 2020; 11: 11. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWu YJ, Pierre DPL, Wu J, et al.: The interaction of versican with its binding partners. Cell Res. 2005 Jul 1; 15(7): 483–494. PubMed Abstract \| Publisher Full Text\n\nBathina A: Effect of substrate availability and O-GlcNAse inhibition on hyaluronan synthesis and intracellular trafficking of HAS3 in MV3 melanoma cells.2014.\n\nMcFall-Ngai M, Hadfield MG, Bosch TCG, et al.: Animals in a bacterial world, a new imperative for the life sciences. Proc. Natl. Acad. Sci. 2013 Feb 26; 110(9): 3229–3236. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNosanchuk JD, Casadevall A: The contribution of melanin to microbial pathogenesis. Cell. Microbiol. 2003 Apr 1; 5(4): 203–223. PubMed Abstract \| Publisher Full Text\n\nWest-Eberhard MJ: Phenotypic plasticity and the origins of diversity. Annu. Rev. Ecol. Syst. 1989; 20: 249–278. Publisher Full Text\n\nWaddington CH: Genetic assimilation of an acquired character. Evolution. 1953; 7: 118–126. Publisher Full Text\n\nSalis P, Klann M, Laudet V: Color Patterns in Anemonefish: Development, Role, and Diversity. Evolution, Development and Ecology of Anemonefishes: Model Organisms for Marine Science. CRC Press; 2022.\n\nWest-Eberhard MJ: Developmental plasticity and the origin of species differences. Proc. Natl. Acad. Sci. 2005 May 3; 102(suppl_1): 6543–6549. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTaylor BA, Cini A, Wyatt CDR, et al.: The molecular basis of socially mediated phenotypic plasticity in a eusocial paper wasp. Nat. Commun. 2021 Feb 3; 12(1): 775. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMoyer JT: Geographical variation and social dominance in Japanese populations of the anemonefish Amphiprion clarkii. Japanese Journal of Ichthyology. 1976; 23(1): 12–22.\n\nBell L, Moyer J, Numachi K: Morphological and genetic variation in Japanese populations of the anemonefish Amphiprion clarkii. Mar. Biol. 1982; 72(2): 99–108. Publisher Full Text\n\nLiu H, Lamm MS, Rutherford K, et al.: Large-scale transcriptome sequencing reveals novel expression patterns for key sex-related genes in a sex-changing fish. Biol. Sex Differ. 2015; 6(1): 1–20.\n\nAzuma T, Takeda K, Doi T, et al.: The influence of temperature on sex determination in sockeye salmon Oncorhynchus nerka. Aquaculture. 2004; 234(1–4): 461–473. Publisher Full Text\n\nBezault E, Clota F, Derivaz M, et al.: Sex determination and temperature-induced sex differentiation in three natural populations of Nile tilapia (Oreochromis niloticus) adapted to extreme temperature conditions. Aquaculture. 2007 Jan 1; 272: S3–S16. Publisher Full Text\n\nGemmell NJ, Todd EV, Goikoetxea A, et al.: Chapter Three - Natural sex change in fish.Capel B, editor. Current Topics in Developmental Biology. Academic Press; 2019; pp. 71–117.\n\nMiura S, Nakamura S, Kobayashi Y, et al.: Differentiation of ambisexual gonads and immunohistochemical localization of P450 cholesterol side-chain cleavage enzyme during gonadal sex differentiation in the protandrous anemonefish, Amphiprion clarkii. Comp. Biochem. Physiol. B: Biochem. Mol. Biol. 2008 Jan 1; 149(1): 29–37. PubMed Abstract \| Publisher Full Text\n\nLorin T, Brunet FG, Laudet V, et al.: Teleost Fish-Specific Preferential Retention of Pigmentation Gene-Containing Families After Whole Genome Duplications in Vertebrates. G3 Genes\|Genomes\|Genetics. 2018 May 4; 8(5): 1795–1806. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIrion U, Nüsslein-Volhard C: The identification of genes involved in the evolution of color patterns in fish. Curr. Opin. Genet. Dev. 2019 Aug 1; 57: 31–38. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLauth RR, Guthridge JL, Cooper DW, et al.: Behavioral ecology of color patterns in Atka mackerel. Mar. Coast. Fish. 2010; 2(1): 399–411. Publisher Full Text\n\nJørgensen KM, Solberg MF, Besnier F, et al.: Judging a salmon by its spots: environmental variation is the primary determinant of spot patterns in Salmo salar. BMC Ecol. 2018 Apr 12; 18(1): 14. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKelley JL, Phillips B, Cummins GH, et al.: Changes in the visual environment affect colour signal brightness and shoaling behaviour in a freshwater fish. Anim. Behav. 2012 Mar 1; 83(3): 783–791. Publisher Full Text\n\nMerilaita S, Kelley JL: Scary clowns: adaptive function of anemonefish coloration. J. Evol. Biol. 2018 Oct 1; 31(10): 1558–1571. Publisher Full Text\n\nDa Silva CRB, Hoepner CM, Mercader M, et al.: The Impact of Popular Film on the Conservation of Iconic Species: Anemonefishes in the Aquarium Trade. Evolution, Development and Ecology of Anemonefishes: Model Organisms for Marine Science. CRC Press; 2022.\n\nPatterson LB, Parichy DM: Zebrafish Pigment Pattern Formation: Insights into the Development and Evolution of Adult Form. Annu. Rev. Genet. 2019 Dec 3; 53(1): 505–530. PubMed Abstract \| Publisher Full Text\n\nSalis P, Roux N, Lecchini D, et al.: The post- embryonic development of Amphiprion perideraion reveals a decoupling between morphological and pigmentation changes. Cybium. 2018; 42(4): 309–312.\n\nMutalipassi M, Terzibasi Tozzini E, Cellerino A: Age and Longevity. Evolution, Development and Ecology of Anemonefishes: Model Organisms for Marine Science. CRC Press; 2022.\n\nBuston PM, García MB: An extraordinary life span estimate for the clown anemonefish Amphiprion percula. J. Fish Biol. 2007 Jun 1; 70(6): 1710–1719. Publisher Full Text\n\nSahm A, Almaida-Pagán P, Bens M, et al.: Analysis of the coding sequences of clownfish reveals molecular convergence in the evolution of lifespan. BMC Evol. Biol. 2019 Apr 11; 19(1): 89. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHoltze S, Gorshkova E, Braude S, et al.: Alternative Animal Models of Aging Research. Front. Mol. Biosci. 2021; 8: 8. Publisher Full Text\n\nBuston PM, Branconi R, Rueger T: Social Evolution in Anemonefishes: Formation, Maintenance, and Transformation of Social Groups. Evolution, Development and Ecology of Anemonefishes: Model Organisms for Marine Science. CRC Press; 2022.\n\nBarbasch TA, DeAngelis R, Rhodes J, et al.: Parental Care: Patterns, Proximate and Ultimate Causes, and Consequences. Evolution, Development and Ecology of Anemonefishes: Model Organisms for Marine Science. CRC Press; 2022.\n\nHoekstra HE, Robinson GE: Behavioral genetics and genomics: Mendel’s peas, mice, and bees. Proc. Natl. Acad. Sci. 2022 Jul 26; 119(30): e2122154119. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBengston SE, Dahan RA, Donaldson Z, et al.: Genomic tools for behavioural ecologists to understand repeatable individual differences in behaviour. Nat. Ecol. Evol. 2018 Jun 1; 2(6): 944–955. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJames N, Bell A: Minimally invasive brain injections for viral-mediated transgenesis: New tools for behavioral genetics in sticklebacks. PLoS One. 2021 May 17; 16(5): e0251653. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNorton W, Bally-Cuif L: Adult zebrafish as a model organism for behavioural genetics. BMC Neurosci. 2010 Aug 2; 11(1): 90. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOrger MB, de Polavieja GG : Zebrafish behavior: opportunities and challenges. Annu. Rev. Neurosci. 2017; 40: 125–147. PubMed Abstract \| Publisher Full Text\n\nGreenwood AK, Wark AR, Yoshida K, et al.: Genetic and Neural Modularity Underlie the Evolution of Schooling Behavior in Threespine Sticklebacks. Curr. Biol. 2013 Oct 7; 23(19): 1884–1888. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGreenwood AK, Peichel CL: Social Regulation of Gene Expression in Threespine Sticklebacks. PLoS One. 2015 Sep 14; 10(9): e0137726. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "164241", "date": "13 Apr 2023", "name": "Fabio Cortesi", "expertise": [ "Reviewer Expertise Evolution", "neuroethology", "marine biology", "ichthyology", "sensory biology", "molecular ecology", "behavioural ecology", "genomics", "transcriptomics" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nHerrera and colleagues review the current standings in anemonefish multi-omics research in their article. They propose that anemonefish are an ideal model for studying ecological, evolutionary and developmental processes because they show traits shared across teleosts and vertebrates and many unique features that set them apart. Especially their obligate relationship with a host anemone and a hierarchy-dependent hermaphroditic lifestyle makes them prime candidates to study processes such as adaptive radiations, mutualism, social dynamics, developmental plasticity, and adaptation to climate change, including ocean warming and acidification. Indeed, in recent years, the anemonefish research community has grown considerably. There are now multiple labs around the globe that use the latest ‘omic’ technologies (e.g., long-read genome sequencing, proteomics, transcriptomics and CRISPR/Cas9) combined with sophisticated imaging and behavioural approaches to study the life histories of these fascinating critters.\nI really enjoyed reading this review as it provides both a historical account of the progress made in using molecular approaches and a current-day account and outlook of where the study of anemonefish biology will likely lead us. The review does an excellent job of incorporating the latest studies in the anemonefish ‘omic’ space without prioritising one field or study over another. I also found the article flow easy to follow, and the figures were well-designed and informative. Certainly, I gained several new insights and research ideas from reading this review.\nI only have a few minor comments I recommend addressing before the indexing of the article in F1000 should occur.\n\nMinor comments:\nFigure 1\nZoomed-in panel: What are the multiple looping arrows for the proteome circle referring to? One of them is labelled with post-translational modifications, which makes sense. What are the other two referring to?\nMain panel: In the main text, you say that the first chromosome-scale genome was released in 2018, but here it is shown as 2019. If I remember correctly, the study was first published in pre-print in 2018 and then after peer review in 2019. I recommend sticking to one or the other.\n2. Anemonefish as a model system for evolutionary biology\nChange ‘Furthermore, the fish serves as a supplemental […].’ to ‘Furthermore, the fish provides supplemental nutrition […].'\nIn the sentence: 'Distribution varies greatly for each species, some are widespread (e.g., A. clarkii, A. sandaracinos), while others have a limited regional distribution (e.g., A. bicinctus, A. percula) or are even restricted to a few islands (e.g., A. chagosensis, A. latezonatus).'\nA. latezonatus occurs along the subtropical coast of Eastern Australia and is not restricted to a few islands. Did you mean A. mccullochi instead?\nChange ‘Constructing a high-quality chromosome-level assembly for a species with no previous genome-scale data was certainly a major achievement in a world of sticklebacks and zebrafish’ to ‘Constructing a high-quality chromosome-level assembly for a species with no previous genome-scale data was certainly a major achievement in a world dominated by stickleback and zebrafish genomic research’.\n2.3 Insights from comparative transcriptomics\nThe first two sentences talk about the initial molecular work in anemonefish. I recommend adding early-day studies about the population genetics of these fishes here, e.g., Jeff Jones and colleagues’ seminal work using microsatellites to show that anemonefish settle close to home was published in the early 2000s.\n2.4 The rise of proteomics\nI recommend splitting this very long sentence into two: ‘Proteomic techniques have been classified as shotgun, the optimal method for discovering more proteins but with the drawback that has reduced quantitative accuracy and reproducibility, or targeted, which is better for reproducibility if the proteins in question are known but limited in the number of measurements and therefore the number of peptides that can be identified.’\nAdd the verb ‘using’ to the following sentence ‘SWATH-MS is versatile and has been used […].’\n3.3 Phenotypic plasticity and genetic assimilation in development and evolution of anemonefish\nI suggest rewriting the following sentence to improve clarity: ‘Species with adult individuals that can be experimentally induced to transition between distinct phenotypes are notably valuable as they make it possible to isolate phenotypic effects of gene expression by comparing the gene expression profiles of groups of individuals who differ in their phenotypes due to plasticity rather than genetic differences.’\nHow can these species be leveraged to study plastic versus genetic effects on a phenotype?\n4.1 Sex change\nThe authors do a great job of describing the transcriptomic research that has taken place to investigate sex change in anemonefish, noting that a brain-to-gonad axis exists in these fishes. However, the timing and speed at which these changes occur still need to be debated, and some mention of this should ideally be incorporated here. For example, in addition to the relatively fast changes observed by Casas and colleagues, recent work from the Rhodes lab shows that very long periods can also occur between the feminisation of the brain and the gonads (e.g., Active feminization of the preoptic area occurs independently of the gonads in Amphiprion ocellaris).\n4.2 Pigmentation and color patterns\nI would like to point the authors to a recent study of ours that investigated the role of white stripes in the antagonistic behaviour of anemonefishes: ‘Mitchell et al.., 2023 Higher ultraviolet skin reflectance signals submissiveness in the anemonefish, Amphiprion akindynos. Behavioral Ecology’.\nWhile I am usually against pushing our work onto the authors of an article, in this case, this study addresses one of the main points raised by the authors, which is that we provide behavioural evidence for: ‘Young recruits are colored distinctly different than older juveniles to avoid antagonistic and aggressive behaviors from the larger individuals’.\nI recommend changing the second sentence here: ‘Morphotypes such as albinism or individuals with no bands in species that usually have, are never or very rarely observed in the wild but can be found in the aquarium trade industry (reviewed in Ref. 16). In the wild, mutations that result in such drastic color pattern alterations have a negative effect on the survival of individuals and are therefore negatively selected against, but they can be bred for several generations in aquaculture.’\nI would be more speculative about the effects of selection, i.e., say something like ‘are likely to have a negative effect’. Anemonefish without white bands have evolved from a (multi-)banded ancestor in nature. Hence selection favoured these extreme phenotypes under certain circumstances.\n4.3 Longevity and lifespan\nIn line with the original article, change the following statement to be damselfish specific as plenty of bigger coral reef fishes live beyond 30 years: ‘Noteworthy, this estimate is two times greater than the longevity estimated for any other coral reef fish’.\n\nIs the topic of the review discussed comprehensively in the context of the current literature? Yes\n\nAre all factual statements correct and adequately supported by citations? Yes\n\nIs the review written in accessible language? Yes\n\nAre the conclusions drawn appropriate in the context of the current research literature? Yes", "responses": [ { "c_id": "10019", "date": "27 Oct 2023", "name": "Marcela Herrera Sarrias", "role": "Author Response", "response": "Minor comments: Figure 1 Zoomed-in panel: What are the multiple looping arrows for the proteome circle referring to? One of them is labelled with post-translational modifications, which makes sense. What are the other two referring to? The reviewer is right, and we have modified the figure for clarification. All loop arrows for all circles have been labelled. Main panel: In the main text, you say that the first chromosome-scale genome was released in 2018, but here it is shown as 2019. If I remember correctly, the study was first published in pre-print in 2018 and then after peer review in 2019. I recommend sticking to one or the other. The reviewer is right, and we apologize for the confusion. We have corrected this so it now reads: “The year 2019 saw the publication of the first chromosome-scale genome for an anemonefish 4 (Figure 1).” 2. Anemonefish as a model system for evolutionary biology Change ‘Furthermore, the fish serves as a supplemental […].’ to ‘Furthermore, the fish provides supplemental nutrition […].' This has been modified as suggested by the reviewer. In the sentence: 'Distribution varies greatly for each species, some are widespread (e.g., A. clarkii, A. sandaracinos), while others have a limited regional distribution (e.g., A. bicinctus, A. percula) or are even restricted to a few islands (e.g., A. chagosensis, A. latezonatus).' A. latezonatus occurs along the subtropical coast of Eastern Australia and is not restricted to a few islands. Did you mean A. mccullochi instead? The reviewer is right. We have replaced A. latezonatus for A. mccullochi. Change ‘Constructing a high-quality chromosome-level assembly for a species with no previous genome-scale data was certainly a major achievement in a world of sticklebacks and zebrafish’ to ‘Constructing a high-quality chromosome-level assembly for a species with no previous genome-scale data was certainly a major achievement in a world dominated by stickleback and zebrafish genomic research’. This has been changed as suggested by the reviewer. 2.3 Insights from comparative transcriptomics The first two sentences talk about the initial molecular work in anemonefish. I recommend adding early-day studies about the population genetics of these fishes here, e.g., Jeff Jones and colleagues’ seminal work using microsatellites to show that anemonefish settle close to home was published in the early 2000s. This has been changed as suggested by the reviewer, so it now reads: “The first molecular insights of anemonefish biology came well before any of the genomes now available were sequenced. Early-day research by Jones and colleagues with microsatellites shed a light on the population genetics and dispersal patterns of anemonefish in Kimbe Bay, Papua New Guinea. Yet, it was a 2010 study using quantitative polymerase chain reaction (qPCR) to investigate the role of the aromatase cyp19a1 gene on sex differentiation of the yellowtail clownfish A. clarkii 74 that really pioneered molecular research in anemonefish.” 2.4 The rise of proteomics I recommend splitting this very long sentence into two: ‘Proteomic techniques have been classified as shotgun, the optimal method for discovering more proteins but with the drawback that has reduced quantitative accuracy and reproducibility, or targeted, which is better for reproducibility if the proteins in question are known but limited in the number of measurements and therefore the number of peptides that can be identified.’ This has been rewritten so it now reads: “Proteomic techniques have been classified into two categories: shotgun and targeted. Shotgun is the optimal method for discovering more proteins despite its drawback of reduced quantitative accuracy and reproducibility. On the other hand, targeted techniques are better for reproducibility when the proteins in question are known but are limited in the number of measurements and therefore the number of peptides that can be identified.” Add the verb ‘using’ to the following sentence ‘SWATH-MS is versatile and has been used […].’ This has been changed as suggested by the reviewer. 3.3 Phenotypic plasticity and genetic assimilation in development and evolution of anemonefish I suggest rewriting the following sentence to improve clarity: ‘Species with adult individuals that can be experimentally induced to transition between distinct phenotypes are notably valuable as they make it possible to isolate phenotypic effects of gene expression by comparing the gene expression profiles of groups of individuals who differ in their phenotypes due to plasticity rather than genetic differences.’ How can these species be leveraged to study plastic versus genetic effects on a phenotype? This has been rewritten so it now reads: “Species with adult individuals that can be experimentally induced to transition between distinct phenotypes are highly valuable. They make it possible to isolate phenotypic effects of gene expression by comparing the gene expression profiles of groups of individuals who differ in their phenotypes due to plasticity rather than genetic differences.” 4.1 Sex change The authors do a great job of describing the transcriptomic research that has taken place to investigate sex change in anemonefish, noting that a brain-to-gonad axis exists in these fishes. However, the timing and speed at which these changes occur still need to be debated, and some mention of this should ideally be incorporated here. For example, in addition to the relatively fast changes observed by Casas and colleagues, recent work from the Rhodes lab shows that very long periods can also occur between the feminisation of the brain and the gonads (e.g., Active feminization of the preoptic area occurs independently of the gonads in Amphiprion ocellaris). As suggested by the reviewer this has been added: “The feminization of the brain in anemonefish is inarguably an active process, and the timing and speed at which these changes occur remain a compelling and active field of research. The process spans a wide time frame, with brain expression profiles changing relatively rapidly after female removal (0 to 11 days in males and 15 to 30 days in transitional males 12) and complete gonadal changes taking much longer (sometimes over the course of several years). Altogether, anemonefish provide a unique opportunity to explore the molecular, biochemical, and physiological mechanisms underlying sex change in vertebrates.” 4.2 Pigmentation and color patterns I would like to point the authors to a recent study of ours that investigated the role of white stripes in the antagonistic behaviour of anemonefishes: ‘Mitchell et al.., 2023 Higher ultraviolet skin reflectance signals submissiveness in the anemonefish, Amphiprion akindynos. Behavioral Ecology’. While I am usually against pushing our work onto the authors of an article, in this case, this study addresses one of the main points raised by the authors, which is that we provide behavioural evidence for: ‘Young recruits are colored distinctly different than older juveniles to avoid antagonistic and aggressive behaviors from the larger individuals’. As suggested by the reviewer this has been changed so it now reads: “Young recruits are colored distinctly different than older juveniles to potentially avoid antagonistic and aggressive behaviors from the larger individuals. 175 Loss of white vertical bars during ontogeny has indeed been observed in multiple Amphiprion species. 14 Mitchell and colleagues (2023) further showed that UV reflectance in anemonefish (from their orange and white bars) has a functional role in modulating aggression and signaling submissiveness in family groups.” I recommend changing the second sentence here: ‘Morphotypes such as albinism or individuals with no bands in species that usually have, are never or very rarely observed in the wild but can be found in the aquarium trade industry (reviewed in Ref. 16). In the wild, mutations that result in such drastic color pattern alterations have a negative effect on the survival of individuals and are therefore negatively selected against, but they can be bred for several generations in aquaculture.’ I would be more speculative about the effects of selection, i.e., say something like ‘are likely to have a negative effect’. Anemonefish without white bands have evolved from a (multi-)banded ancestor in nature. Hence selection favoured these extreme phenotypes under certain circumstances. This has been changed so it now reads: “In the wild, mutations that result in such drastic color pattern alterations are likely to have a negative effect on the survival of individuals and are therefore negatively selected against, but they can be bred for several generations in aquaculture.” 4.3 Longevity and lifespan In line with the original article, change the following statement to be damselfish specific as plenty of bigger coral reef fishes live beyond 30 years: ‘Noteworthy, this estimate is two times greater than the longevity estimated for any other coral reef fish’. The reviewer is right, and this has been changed so it now reads “… longevity estimated for any other pomacentrid and up to two times …”" } ] }, { "id": "170667", "date": "18 May 2023", "name": "Kerstin Johannesson", "expertise": [ "Reviewer Expertise Molecular ecology", "evolutionary genetics", "population genetics", "hybrid zones and speciation" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis review take a broad grip on the anemonefish genus, describing both basic and interesting facts on the biology of these fascinating fish, and (which is the focus on the review) how these species are now emerging as excellent models for studies of evolutionary questions in a broad sense. I congratulate the authors to a comprehensive and also very nicely illustrated. I learnt a lot about anemonefishes reading it. As I am not in the field of anemonefishes, I have some comments that the authors might consider to further clarify some minor parts of the text.\nThe first issue is really trivial for an expert, but for a non-expert it is a bit confusing to use both \"anemonefish\" and \"clownfish\" interchangeably, and without pointing out that these are synonymous. It took me a while to find out. Maybe better to stick to anemonefish throughout?\nIn Fig. 3c, I am confused about the red ribbons that are representing inversions and translocations. In some places it seems as if these corresponded to one whole chromosome, and so what is actually going on at chromosome 3 in A. ocellaris and chromosome 4 in A. percula. Are these just not the same chromosome that have been given different numbers in the two species? Or is there actually also inversions or translocations involved?\nUnder 2.4. Last sentence - why only in extreme environments?\nUnder 2.5. Second sentence - the long parenthesis makes the sentence uneasy to read\nUnder 3. First paragraph. Sea anemones are found globally, and so it is not obvious to me why the anemonefish are confined only to the tropical areas of western Pacific and the Indian Ocean. For example, in NE Atlantic, there are shrimps that associate with sea anemones in much the same way as the anemonefish, and so, why can there not be other genera of fish that have evolved this habit?\n3. Five paragraph. Not sure what you mean by “consequently fixed by natural selection” - do you mean that the inversion is always fixed different (one arrangement at frequency 1.0 in one species, and the other arrangement fixed at 1.0 in the other species? One alternative would be that the inversion is polymorphic in both species, so this need to be clarified. What is here referred to as “supergenes” is also somewhat unclear. Usually, the definition of a supergene is that is e.g. an inversion (or some other low-recombination region) which remains polymorphic within populations - and so this does not match the statement of “fixed by selection”....\nUnder 4.3. One obvious question after the presentation of the extremely long life-span is, does anemone fish have undetermined growth? That is, do they grow to large for their anemone, and so have to move to another and larger anemone? (Also, are the anemones also similarly long-lived?)\nUnder 4.3, second paragraph. What is the difference between “lifespan” and “longevity” in the next last sentence of this paragraph?\nUnder 6. First sentence - please spell out which are these fundamental questions you are referring to.\nThat the fish never abandon their anemone is a very interesting fact that should perhaps be highlighted earlier (or I did miss it when reading), together with the very long lifespan, to illustrate what is the unique features of the anemonefish as a model.\n\nIs the topic of the review discussed comprehensively in the context of the current literature? Yes\n\nAre all factual statements correct and adequately supported by citations? Yes\n\nIs the review written in accessible language? Yes\n\nAre the conclusions drawn appropriate in the context of the current research literature? Yes", "responses": [ { "c_id": "10020", "date": "27 Oct 2023", "name": "Marcela Herrera Sarrias", "role": "Author Response", "response": "The first issue is really trivial for an expert, but for a non-expert it is a bit confusing to use both \"anemonefish\" and \"clownfish\" interchangeably, and without pointing out that these are synonymous. It took me a while to find out. Maybe better to stick to anemonefish throughout? The reviewer is right, and this has been clarified. The first paragraph of section #2 now reads: “Before presenting the various contributions made in the field of anemonefish research, we feel it is essential to clarify the difference between the terms \"anemonefish” and “clownfish” both of which are used throughout this review. Conventionally, the English name “anemonefish” has been associated with the distinctive symbiosis between these fish species and giant sea anemones, while the term “clownfish” highlights their vibrant colors and bold behavior. In line with the prevailing practice among researchers in this field, we have opted to refer to them as “anemonefish” to acknowledge the pivotal importance of their symbiotic relationship with giant sea anemones, a key aspect that profoundly influences their biology. However, for Amphiprion ocellaris and Amphiprion percula, two closely related species forming a natural subgroup within anemonefish, we do use the term “clownfish”.” In Fig. 3c, I am confused about the red ribbons that are representing inversions and translocations. In some places it seems as if these corresponded to one whole chromosome, and so what is actually going on at chromosome 3 in A. ocellaris and chromosome 4 in A. percula. Are these just not the same chromosome that have been given different numbers in the two species? Or is there actually also inversions or translocations involved? As indicated in the legend of this figure, the red ribbons depicted represent various chromosomal rearrangements, including translocations and inversions. Chromosomes in both species have been designated based on their size following an orderly arrangement from the largest to the smallest. This has been further clarified in the legend of the figure. Under 2.4. Last sentence - why only in extreme environments? This has been modified so it now reads: “Proteomics can then be a powerful tool for identifying specific proteins and pathways that are crucial to stress responses, but more general for studying the evolution, biodiversity, and physiological adaptations of fish living across different environments.” Under 2.5. Second sentence - the long parenthesis makes the sentence uneasy to read As suggested by the reviewer, this has been modified so it now reads: “The ultimate goal of modern systems biology approaches is to integrate data from various levels of information, from gene regulatory networks, RNA and protein measurements, metabolites and cell-cell interactions, to individuals, populations and ecologies.” Under 3. First paragraph. Sea anemones are found globally, and so it is not obvious to me why the anemonefish are confined only to the tropical areas of western Pacific and the Indian Ocean. For example, in NE Atlantic, there are shrimps that associate with sea anemones in much the same way as the anemonefish, and so, why can there not be other genera of fish that have evolved this habit? The reviewer is right, and we apologize for the lack of clarity. We have modified this so it now reads: “Anemonefish are an extraordinary example of adaptive radiation, a process driven, in this case, by the mutualistic relationship they maintain with giant sea anemones of the superfamily Actinioidea. 135 Notably, host sea anemones originated in the Coral Triangle region, followed by an independent geographical radiation in the Western Indian Ocean. Thus, distribution and abundance of anemonefish are intrinsically linked to the presence and abundance of giant sea anemones 135 , 136. Anemonefish-host anemones are, in turn, exclusively found in the tropical Indo-Pacific Ocean, with no presence in the Eastern Pacific nor Caribbean regions. 26 , 32” 3. Five paragraph. Not sure what you mean by “consequently fixed by natural selection” - do you mean that the inversion is always fixed different (one arrangement at frequency 1.0 in one species, and the other arrangement fixed at 1.0 in the other species? One alternative would be that the inversion is polymorphic in both species, so this need to be clarified. What is here referred to as “supergenes” is also somewhat unclear. Usually, the definition of a supergene is that is e.g. an inversion (or some other low-recombination region) which remains polymorphic within populations - and so this does not match the statement of “fixed by selection”.... The reviewer’s comment raises a valid point. This has been clarified so it now reads: “This persistence might be attributed to genomic inversions that disrupt recombination and create clusters of loci controlling ecologically important traits that may consequently be fixed by natural selection or through genetic drift.” Under 4.3. One obvious question after the presentation of the extremely long life-span is, does anemone fish have undetermined growth? That is, do they grow to large for their anemone, and so have to move to another and larger anemone? (Also, are the anemones also similarly long-lived?) As suggested by the reviewer, we have now discussed this. The last paragraph of this section reads: “After exploring the remarkable long lifespan of anemonefish, a few other compelling questions arise: Do anemonefish have undetermined growth, growing too large for their host sea anemone and eventually having to move to another and larger anemone? Does the longevity of the host sea anemones parallel that of anemonefish? In a groundbreaking milestone, Rueger and colleagues (2022) provided the first-ever experimental evidence for the first question. Their study sheds light on the remarkable growth plasticity of anemonefish in response to their mutualistic interaction with sea anemones, that is, anemonefish adjust their growth rate to make sure they are the ideal size for their hosts. Thus, emphasizing the crucial role of mutualisms in shaping species’ adaptations and ecological relationships. Moreover, this research opens up exciting new avenues for exploring the underlying molecular mechanisms behind this phenomenon: What are the cues necessary for the fish to decide how large it needs to be? Having explored the growth plasticity of anemonefish, we now shift the focus now to the longevity of the host sea anemone and its intricate association with the presence and size of their anemonefish counterparts. Various studies have shown that giant sea anemones are short lived compared to their fish symbionts, with some species like E. quadricolor and H. crispa having turnover times of only 3–5 years. The short lifespans of these host anemones certainly affect their resident anemonefish as they need to migrate among hosts during their lifetimes, and do so when space becomes available nearby. As a result, some anemonefish may rely on the presence of multiple anemone hosts within a relatively small area of reef, leading to potential constraints on their lifespans due to host turnover. This is particularly evident in areas with low and declining host abundance.” Under 4.3, second paragraph. What is the difference between “lifespan” and “longevity” in the next last sentence of this paragraph? The reviewer is right, and we have clarified this in the first paragraph so it now reads: “The evolutionary theory of aging predicts that individuals with low extrinsic mortality will show delayed senescence (i.e., the process of physiological deterioration with age) and increased lifespan (reviewed in Ref. 194). It is important to note that although the terms “lifespan” and “longevity” may sound interchangeable, they hold distinct meanings when discussing life expectancy. Lifespan refers to the maximum potential duration of life for a given species or population. Longevity, on the other hand, describes the ability to live a long life beyond the species-specific average age at death (reviewed in Refs. ). To illustrate, if the average lifespan of a species is 10 years, for example, it means that, on average, individuals of that species are expected to live 10 years. However, if the longevity of the same species is 10 years, then it means that some individuals may live up to 10 years, but the average lifespan may still be much lower than 10 years.” Under 6. First sentence - please spell out which are these fundamental questions you are referring to. We have added this, it now reads: “Anemonefish have become an invaluable model system for answering some of the most fundamental and long-standing questions in evolutionary genomics. How do speciation events occur, and what are the underlying genetic mechanisms driving this process? What genetic changes underlie the development of morphological, physiological, and behavioral traits? How do organisms adapt to their environments, and what role does natural selection play in shaping their genomic architecture? How can we integrate genomic data from multiple species to gain insights into the evolutionary processes shaping biodiversity?” That the fish never abandon their anemone is a very interesting fact that should perhaps be highlighted earlier (or I did miss it when reading), together with the very long lifespan, to illustrate what is the unique features of the anemonefish as a model. As suggested by the reviewer, we have now included this in section 2.1 “Practical features of anemonefish for experimentation”." } ] } ]	1	https://f1000research.com/articles/12-204
https://f1000research.com/articles/12-846/v1	18 Jul 23	{ "type": "Research Article", "title": "Pinostrobin mitigates neurodegeneration through an up-regulation of antioxidants and GDNF in a rat model of Parkinson’s disease", "authors": [ "Ratchaniporn Kongsui", "Tichanon Promsrisuk", "Lars Klimaschewski", "Napatr Sriraksa", "Jinatta Jittiwat", "Sitthisak Thongrong", "Ratchaniporn Kongsui", "Tichanon Promsrisuk", "Lars Klimaschewski", "Napatr Sriraksa", "Jinatta Jittiwat" ], "abstract": "Background: One of the most common neurodegenerative diseases is Parkinson’s disease (PD); PD is characterized by a reduction of neurons containing dopamine in the substantia nigra (SN), which leads to a lack of dopamine (DA) in nigrostriatal pathways, resulting in motor function disorders. Oxidative stress is considered as one of the etiologies involved in dopaminergic neuronal loss. Thus, we aimed to investigate the neuroprotective effects of pinostrobin (PB), a bioflavonoid extracted from Boesenbergia rotunda with antioxidative activity in PD. Methods: Rats were treated with 40 mg/kg of PB for seven consecutive days before and after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD. After completing the experiment, the brains including SN and striatum were used for histological studies and biochemical assays. Results: PB treatment demonstrated a reduction of free radicals in the SN as indicated by significantly decreased MDA levels, whereas the antioxidative enzymes (SOD and GSH) were significantly increased. Furthermore, PB treatment significantly increased glial cell line-derived neurotrophic factor (GDNF) immunolabelling which has neurotrophic and neuroprotective effects on the survival of dopaminergic neurons. Furthermore, PB treatment was shown to protect CA1 and CA3 neurons in the hippocampus and dopaminergic neurons in the SN. DA levels in the SN were increased after PB treatment, leading to the improvement of motor function of PD rats. Conclusions: These results imply that PB prevents MPTP-induced neurotoxicity via its antioxidant activities and increases GDNF levels, which may contribute to the therapeutic strategy for PD.", "keywords": [ "Pinostrobin", "Parkinson’s disease", "Oxidative stress", "Boesenbergia rotunda" ], "content": "Introduction\n\nOne of the most common neurodegenerative diseases is Parkinson’s disease (PD), which has a high prevalence worldwide.1 PD is characterized by a reduction of neurons containing dopamine in the substantia nigra pars compacta (SNpc), which leads to a lack of dopamine (DA) in the nigrostriatal pathways. DA insufficiency in the striatum leads to movement disorders including muscle rigidity, postural imbalance, resting tremor and bradykinesia.2,3 Currently, PD cannot be cured, but symptomatic treatment with L-3,4-dihydroxyphenylalanine (levodopa, L-DOPA) and DA receptor agonists exists. However, the effects of these medications are not long-lasting and they unsuccessful in preventing the loss of dopamine neurons in the SNpc.4,5 So far, accumulative studies have shown that dopaminergic neuronal loss in the SNpc is a progressive process; however, its exact mechanisms remain poorly understood.6–8 Even though the pathogenesis of PD is still unclear, oxidative stress has been suggested as a possible mechanism of dopaminergic neuronal death in the SNpc.9,10 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a neurotoxin which is widely used to target dopaminergic neuronal death in animal models of PD.11,12 The active metabolite of MPTP, 1-methyl-4-phenyl pyridinium (MPP+), is selectively transported into dopaminergic neurons and causes mitochondrial dysfunction through a massive accumulation of free radicals such as reactive oxygen species (ROS), resulting in neuronal apoptosis.13,14\n\nPinostrobin (PB, 5-hydroxy-7-methoxyflavanone), a bioactive flavonoid, is found in the rhizomes of Boesenbergia rotunda (fingerroot or Krachai) in Thailand.15 PB has various bioactivities and pharmacological effects, including antioxidative, anti-peptic16 anti-inflammatory, analgesic,17 anticancer,18 and neuroprotective activities.19 Our previous studies have shown that treatment of PB at the doses of 20 and 40 mg/kg improved peripheral nerve regeneration and functional recovery after injury in rats through attenuation of oxidative stress.20 Recently, the antioxidative effects of PB against neurotoxin-induced dopaminergic neurons loss have been demonstrated in zebra fish and in SH-SY5Y cells.21 However, a single animal study cannot fully elucidate the effects of PB on PD; therefore, the neuroprotective and therapeutic effects of PB must be further investigated. Current pharmacological therapeutics can only increase DA level and improve motor function of PD patients but are unsuccessful at preventing neurodegeneration. Thus, this research was set up to explore possible neuroprotective effects of PB in a rodent model of PD.\n\n\nMethods\n\nPB was prepared from B. rotunda rhizomes as previously described in our work (20). Briefly, fresh rhizomes were ground then dried for an ethanolic extract. The extract was resuspended in methanol to obtain a pale yellow solid. The structural analysis of PB was performed by 13C- and 1H-NMR spectroscopy (Bruker Avance DRX500 Spectrometer, USA) and by nuclear magnetic resonance (NMR) spectra for comparison with published data.\n\nEighteen male Wistar rats (200-220 g) were purchased from Nomura Siam International Co., Ltd (Thailand). All rats were acclimatized for 1 week in a room with 12 h light-dark cycle and constant temperature (25±2°C) with food and water access ad libitum. All animal procedures were approved by the Animal Ethics Committee of University of Phayao, Thailand, with approval number 640104028. The animals were randomly divided into three groups: control group, MPTP group and MPTP+40 mg/kg PB group. MPTP (Tokyo Chemical Industry Co., Ltd. Cat#M2690, Japan) was freshly diluted in 0.9 % sterile normal saline for the intraperitoneal injection (i.p.) to induce PD at a dose of 20 mg/kg once per day for five consecutive days. In the MPTP+40 mg/kg PB group, the animals were additionally treated with PB dissolved in 0.5% of carboxymethylcellulose sodium (Sigma-Aldrich, USA) once daily by oral gavage for seven consecutive days before and after MPTP injection.\n\nA horizontal ladder rung walking apparatus was used to test the motor coordination between forelimbs and hindlimbs as previously described by Metz and Whishaw.22 All animals spontaneously walked from a beginning point to a goal point along a horizontal rung consisting of 100 × 20 cm clear acrylic walls. The metal rungs created a walking space with a minimum distance of 1 cm between rungs (Figure 1). All animals were trained to cross the ladder for three days before i.p. injection of MPTP. Seven days after MPTP injection, all animals were required to walk across the ladder for assessing motor function recovery. All animals were tested three times per session and the spending time to cross the rungs was recorded.\n\nAfter completing the behavioral experiment, all animals were i.p. injected with thiopental sodium at a dose of 70 mg/kg followed by ice cold 0.9% sterile normal saline perfusion through the heart. The substantia nigra was dissected from the midbrains and collected for biochemical assays. The tissues were homogenized in a homogenate buffer containing 50 mM Tris-HCl (pH 7.4), 150 mM NaCl, 1% Triton X-100, and protease inhibitor and then centrifuged at 10,000 rpm at 4°C for 15 min. The protein concentrations were determined by the method of Lowry. Bovine serum albumin (BSA) (Cat# 9048-46-8, Sigma-Aldrich, USA) was used as a standard.23\n\nMDA assay was performed according to Luangaram and co-workers.24 Briefly, 150 μL of tissue supernatant was mixed with chemical solutions as described in the protocol. The mixed solution was incubated at room temperature (RT) for 10 min. After that, 500 μL of 0.6% thiobarbituric acid (TBA) was added to the mixture and then boiled for 30 min followed by centrifugation at 2,800 g for 20 min. Thiobarbituric acid-reactive substances (TBARS) were detected at 532 nm using the Cytation5 microplate reader (USA). A standard curve was produced using appropriate concentrations of 1,1.3,3-tetraethoxypropane (0.3–10 μmol/L). The concentration of MDA in each tissue sample was normalized to protein content. All MDA assays were done three times.\n\nThe oxidative stress response was determined by the measurement of reduced GSH content following Polycarp and colleagues’s protocol.25 In brief, 1 mM reduced GSH (Cat# 70-18-8, Sigma-Aldrich, USA) was diluted in 0.1 N HCl to make serial dilutions of 0.2, 0.4, 0.6 and 0.8 mM GSH for a standard curve. The reaction was started by adding each serial dilution or tissue sample to the phosphate buffer pH 7.6 and 1 mM 5,5-dithio-bis-(2-nitrobenzoic acid) (DTNB) (Cat# 69-78-3, Merk, USA). Thereafter, the mixture was incubated for 5 min in the dark at RT. Each sample mixture was read at 412 nm using a Cytation5 microplate reader. GSH concentration in tissue sample was normalized to protein content, and the experiments were repeated three times.\n\nSOD in response to oxidative stress was detected using a SOD Assay Kit (Cat# 19169, Sigma-Aldrich, USA) following the manufacturer's protocol. The SOD concentration was normalized to protein content, and the experiments were repeated three times.\n\nDA expression was detected using a Dopamine ELISA Kit (Cat# E-EL-0046, Elabscience, USA) following the manufacturer's instructions. The results were normalized to protein content, and the experiments were performed in triplicates.\n\nThe brain tissue samples were fixed in 4% paraformaldehyde in phosphate buffered saline (PBS) pH 7.4 at 4°C for 48 h, then cryoprotection in 30% sucrose for 48 h. The brains were coronally cut at a thickness of 20 μm using a cryostat (CM1950, Leica Microsystems, Inc., Germany). Brain sections were used for Nissl staining using a cresyl violet solution (Cat# 105235, Sigma-Aldrich; Merck KGaA, USA) to determine the neuronal density in the hippocampal sub-fields including cornu ammonis 1 (CA1) and cornu ammonis 3 (CA3). The 40× magnification images of the CA1 and CA3 regions (at stereotaxic co-ordinates 4.0-4.2 mm posterior to the bregma) were chosen for measuring the neuronal density using the NIS Elements imaging software version 5 (Nikon Corporation). The experiments were done three times, and the data were shown as % of control.\n\nTo determine dopaminergic neurons in the SNpc, brain coronal sections at stereotaxic co-ordinates 4.6-4.8 mm behind the bregma were incubated with tyrosine hydroxylase (TH) antibodies. Positive GDNF immunolabelling in the striatum (at 0.5 mm anterior to the bregma) was detected using GDNF antibodies. Sections were free-floating-blocked with 10% normal goat serum and 0.3% Triton X-100 in PBS at RT for 90 min. TH primary antibody (1:500, Cat# ab6211, Abcam, UK) or GDNF primary antibody (1:500, Cat# ab244211, Abcam, UK) were applied overnight at RT followed by biotin-SP-conjugated donkey anti-rabbit secondary antibody (1 : 500, Code: 711-065-152 Jackson Immunoresearch, USA) or biotin-SP-conjugated donkey anti-mouse secondary antibody (1 : 500, Code: 715-065-150 Jackson Immunoresearch, USA) for 2 h at RT. After that, sections were incubated for 1 h in 0.1% extravidin peroxidase (1:1000, Cat# E2886, Sigma-Aldrich, USA) then detected by 3,3′-diaminobenzidine (DAB). The images were examined at 40× magnification by NIS Elements imaging software version 5 (Nikon). TH-positive neurons or GDNF positive immunolabelling were counted and expressed as a percentage of control. The experiments were done three times on each animal.\n\nResults were analyzed by one-way ANOVA followed by a Tukey's post hoc using GraphPad Prism version 9 (GraphPad Software, Inc., USA) and presented as Means±SEM for motor functional tests, histology, and immunohistochemical studies, Mean±SD for biochemical assays. Statistical significance was considered when p-values were lower than 0.05 (p<0.05).\n\n\nResults\n\nTo evaluate the effect of PB on MPTP-induced motor functional deficit, all animals were assigned to walk across the ladder rung apparatus. Prior to MPTP injection at baseline, there were no significant differences in crossing time between all animal groups (Control: 9.27±0.41 s versus MPTP: 9.56±0.35 s versus MPTP+PB: 9.50±0.57 s, Figure 2). Seven days after MPTP injection, the MPTP and MPTP+PB groups showed a significantly increased time spent across the ladder rungs when compared to the control group (Control: 9.11±0.78 s versus MPTP: 16.89±0.88 s versus MPTP+PB: 13.44±0.16 s). The data indicated that administration of MPTP negatively affects motor functions; however, treatment with PB significantly improved motor coordination, resulting in reduced crossing times as compared to the MPTP group (p<0.01).\n\nPinostrobin-treated animals exhibited less time spent on the ladder when compared to animals in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine only group. Data were reported as mean±SEM (n=6), one-way ANOVA. ##p<0.01, and ###p<0.001 versus control group; *p<0.01 versus MPTP group.\n\nInjection of MPTP caused an accumulation of MDA content as an indicator for lipid peroxidation due to formation of free radicals. However, PB-treated animals revealed a capacity to reduce the MDA content compared to animals in the MPTP group (Control: 19.16±2.44 versus MPTP: 36.69±8.52 versus MPTP+PB: 24.20±1.50; Figure 3A). The reduction of MDA level in PB-treated animals may be due to an increase of antioxidant enzymes activities. Our data demonstrate that GSH level in the PB-treated group significantly increased in comparison to the MPTP group (Control: 18.06±3.44 versus MPTP: 12.85±2.05 versus MPTP+PB: 19.13±4.23; Figure 3B). Additionally, PB also elevated the level of SOD. Seven days after MPTP injection, SOD level was markedly increased in the PB-treated group when compared to the MPTP group (Control: 0.52±0.02 versus MPTP: 0.36±0.07 versus MPTP+PB: 0.51±0.12, Figure 3C).\n\nResults were displayed as mean±SD (n=6), one-way ANOVA. #p<0.05, ##p<0.01, and ###p<0.001 versus control group; p<0.05, and p<0.01 versus MPTP group.\n\nThe improvement of motor coordination in PB-treated animals after exposure to the dopaminergic neurotoxin may be linked to an increase in DA levels. Therefore, alterations of DA were assessed using an ELISA kit on day 7 after MPTP injection. Animals injected with neurotoxin (MPTP) demonstrated markedly reduced DA levels when compared to the control group (Control: 57.18±8.43 versus MPTP: 30.34±3.34 versus MPTP+PB: 41.30±6.43; Figure 3D). In this study, PB treatment significantly attenuated the reduction of DA when compared to the MPTP group. These results revealed that PB treatment inhibits MPTP-induced oxidative stress via upregulation of antioxidative enzymes; moreover, PB counteracted the loss of DA which correlated with an improvement of motor coordination.\n\nInjection of MPTP caused neuronal apoptosis in the hippocampus and resulted in cognitive impairment in PD.26 In this study, Nissl staining was performed to detect hippocampal neuron degeneration after i.p. injection of MPTP. Overall neuron numbers from all experimental groups were counted at the same levels along the rostro-caudal axis. Neuronal loss was observed in CA1 and CA3 regions of MPTP-treated animals when compared to control rats (Figure 4). PB administration significantly attenuated the death of principal neurons in the CA1 subfield (MPTP: 66.90±5.51% versus MPTP+PB: 92.98±4.67%; Figure 4G). Similarly, animals in the PB-treated group displayed a greater percentage of CA3 neurons than those in the MPTP only group (MPTP: 58.63±3.92% versus MPTP+PB: 84.05±6.03%, Figure 4H). These results illustrated that administration of PB could protect hippocampal neurons from death induced by MPTP.\n\nData were reported as mean±SEM (n=6), one-way ANOVA. #p<0.05, and ###p<0.001 versus control group; p<0.01, and **p<0.001 versus MPTP group. Bar=50 μm.\n\nImmunohistochemistry staining of TH was performed to detect neurons containing dopamine in the SNpc. TH is the key enzyme for catecholamine synthesis by converting L-tyrosine to L-DOPA, a precursor for DA. DA insufficiency in the striatum is due to the degeneration of dopaminergic neurons which leads to a decrease in TH activity.27 A previous study found that MPTP decreased TH activity in rodent models.28 Seven days after MPTP injection, TH-positive neurons were dramatically reduced as compared with the control group (Figure 5A-C). However, there were more TH-positive neurons that survived in MPTP+PB animals (MPTP: 45.15±7.65% versus MPTP+PB: 72.22±6.66%, Figure 5G).\n\nImages were taken at 40× magnification. mean±SEM (n=6), one-way ANOVA. #p<0.05, ##p<0.01, and ###p<0.001 vs control group; p<0.05, and **p<0.01 versus MPTP group. Bar=50 μm.\n\nWe further investigated the expression of GDNF. GDNF has been reported to be a neurotrophic factor of dopaminergic neurons and to protect them from MPTP-induced neuronal apoptosis.29,30 In this study, PB-treated animals exhibited significantly higher numbers of GDNF-positive immunolabelling in the corpus striatum when compared to animals in the MPTP group (Figure 5D-F). Quantitative analysis showed a higher percentage in the MPTP+PB group as compared to the MPTP only group (MPTP: 38.93±4.82% versus MPTP+PB: 69.22±6.98%, Figure 5H).\n\n\nDiscussion\n\nPD is characterized by a reduction of dopamine-producing neurons in the SNpc; however, its etiology remains under debate. Recently, the standard treatment for PD has been a dopamine substitution with L-DOPA, but the medications cannot prevent dopaminergic neuron death.31 So far, the high expression of oxidants such as ROS has been considered as the main etiopathology of PD.9,10 In our previous study, PB as a natural antioxidant agent was shown to attenuate memory impairment and hippocampal neuronal loss, as well as increased astrocytic immunoreactivities of GFAP probably via the activations of antioxidative enzymes including SOD and CAT in chronic stress-induced oxidative stress animal models.32 It is well known that administration of MPTP-induced oxidative stress causes hippocampal neuron loss and depletion of dopaminergic neurons as well as functional impairments.33 The administration of MPTP has been reported to induce locomotor problems in PD animals.34 In the previous investigation, the results demonstrated that MPTP-treated animals exhibited motor function impairments as compared to the naïve control animals. Of note, these deficits improved upon PB treatment seven days after MPTP injection. MPTP plays a crucial role in inducing dopaminergic neuron apoptosis via production of oxidative stress.13 Previous evidence has shown that flavonoids have antioxidant effects that inhibit free radical formation induced by lipid peroxidation.35 In the present study, animals subjected to MPTP for seven days displayed significant activation of lipid peroxidation; a process in which free radicals destroy lipids in cellular membranes as indicated by an increase of MDA levels compared to control animals. MDA levels were decreased in PB-treated animals, and prolonged treatment with PB caused an upregulation of antioxidant enzymes, including GSH and SOD. Previous research has demonstrated that PB attenuates oxidative stress induced by MPTP via the activation of GSH and SOD in the SH-SY5Y cells.21 Our findings confirmed the antioxidant capacity of PB and its neuroprotective effects in neurotoxin-induced neuronal death.\n\nMPTP-caused dopaminergic neuronal cell loss leads to a decrease in dopamine expression in the nigrostriatal system. Previous studies have shown that MPP+-caused mitochondrial dysfunction leads to neuronal apoptosis in the SNpc and in the hippocampus.36–38 In the present study, PB promoted neuroprotection against neurotoxin induced neuronal apoptosis in the SNpc and in the hippocampus. The animals in the PB-treated group revealed significantly higher numbers of CA1 and CA3 neurons in the hippocampus as well as TH-positive neurons in the SNpc, compared with MPTP treated animals. Moreover, ELISA result showed that PB treatment increased DA levels in the midbrain, which correlates with higher numbers of TH positive neurons as compared to non-treated animals. Hence, higher levels of DA in the nigrostriatal pathway caused an improvement in motor function after MPTP induced PD.\n\nPB prevented neuronal death, possibly via an upregulation of neurotrophic factors. It is well known that GDNF acts as a prominent neurotrophic factor in the DA system and prevents neuronal apoptosis induced by 6-OHDA.39 Moreover, GDNF promotes the survival of dopaminergic neurons after exposure to MPTP.40 Studies have demonstrated that GDNF treatment increased DA release in the striatum.41,42 High expression of GDNF markedly increases TH-positive neurons in the SNpc, which leads to a reduction in dyskinesia in the MPTP-induced PD.30 In this research, we found that PB treatment could increase the expression of GDNF, as revealed by immunohistochemistry. Previous study has shown the colocalization of GDNF and GFAP-positive astrocytes (astrocytic GDNF), which can increase the striatal neurotransmitters, resulting in a reduction of motor functional deficits.40 Taken together, treatment of PB at a dose of 40 mg/kg could attenuate oxidative stress through an upregulation of antioxidative enzymes and activation of the PI3K/Akt and Erk pathways, which support cellular survival.43,44\n\nIn summary, the present study provides evidence that PB possesses antioxidant activities and neuroprotective effects on MPTP-caused neurodegeneration in PD. PB may serve as a therapeutic drug for PD or in neurodegenerative diseases caused by free radical-induced neuronal cell death.", "appendix": "Data availability\n\nFigshare: Pinostrobin mitigates neurodegeneration through an up-regulation of antioxidants and GDNF in a rat model of Parkinson’s disease, https://doi.org/10.6084/m9.figshare.23058479.v3. 45\n\nThis project contains the underlying following data:\n\n• Ladder rung data\n\n• MDA detection data\n\n• GSH detection data\n\n• SOD detection data\n\n• Dopamine detection data\n\n• Hippocampal cresyl violet staining images\n\n• Striatal GDNF immunohistochemistry staining images\n\n• Substantia nigra tyrosine hydroxylase immunohistochemistry staining images\n\nFigshare: ARRIVE checklist for ‘Pinostrobin mitigates neurodegeneration through an up-regulation of antioxidants and GDNF in a rat model of Parkinson’s disease’, https://doi.org/10.6084/m9.figshare.23058479.v3. 45\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nThis research was granted by the Thailand Science Research and Innovation fund and the University of Phayao (Grant No. FF65-RIM119). The authors would like to thank the Ernst Mach Grant, Nachbetreuungsstpendium/EZA for funding support during the manuscript preparation.\n\n\nReferences\n\nTysnes OB, Storstein A: Epidemiology of Parkinson's disease. J. Neural Transm. (Vienna). 2017; 124(8): 901–905. Publisher Full Text\n\nFahn S: The history of dopamine and levodopa in the treatment of Parkinson's disease. Mov. Disord. 2008; 23(Suppl 3): S497–S508. Publisher Full Text\n\nMazzoni P, Shabbott B, Cortes JC: Motor control abnormalities in Parkinson's disease. Cold Spring Harb. Perspect. Med. 2012; 2(6): a009282. PubMed Abstract \| Publisher Full Text\n\nBlandini F, Armentero MT: Dopamine receptor agonists for Parkinson's disease. Expert Opin. Investig. Drugs. 2014; 23(3): 387–410. Publisher Full Text\n\nBonuccelli U, Ceravolo R: The safety of dopamine agonists in the treatment of Parkinson's disease. Expert Opin. Drug Saf. 2008; 7(2): 111–127. Publisher Full Text\n\nZucca FA, Vanna R, Cupaioli FA, et al.: Neuromelanin organelles are specialized autolysosomes that accumulate undegraded proteins and lipids in aging human brain and are likely involved in Parkinson's disease. NPJ Parkinsons Dis. 2018; 4: 17. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBarcia C, Sanchez Bahillo A, Fernandez-Villalba E, et al.: Evidence of active microglia in substantia nigra pars compacta of parkinsonian monkeys 1 year after MPTP exposure. Glia. 2004; 46(4): 402–409. PubMed Abstract \| Publisher Full Text\n\nSy HN, Wu SL, Wang WF, et al.: MPTP-induced dopaminergic degeneration and deficits in object recognition in rats are accompanied by neuroinflammation in the hippocampus. Pharmacol. Biochem. Behav. 2010; 95(2): 158–165. PubMed Abstract \| Publisher Full Text\n\nDionisio PA, Amaral JD, Rodrigues CMP: Oxidative stress and regulated cell death in Parkinson's disease. Ageing Res. Rev. 2021; 67: 101263. Publisher Full Text\n\nTolleson CM, Fang JY: Advances in the mechanisms of Parkinson's disease. Discov. Med. 2013; 15(80): 61–66. PubMed Abstract\n\nZhang QS, Heng Y, Mou Z, et al.: Reassessment of subacute MPTP-treated mice as animal model of Parkinson's disease. Acta Pharmacol. Sin. 2017; 38(10): 1317–1328. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLu C, Zhang J, Shi X, et al.: Neuroprotective effects of tetramethylpyrazine against dopaminergic neuron injury in a rat model of Parkinson's disease induced by MPTP. Int. J. Biol. Sci. 2014; 10(4): 350–357. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nXu SF, Zhang YH, Wang S, et al.: Lactoferrin ameliorates dopaminergic neurodegeneration and motor deficits in MPTP-treated mice. Redox Biol. 2019; 21: 101090. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLu KT, Ko MC, Chen BY, et al.: Neuroprotective effects of resveratrol on MPTP-induced neuron loss mediated by free radical scavenging. J. Agric. Food Chem. 2008; 56(16): 6910–6913. PubMed Abstract \| Publisher Full Text\n\nEng-Chong T, Yean-Kee L, Chin-Fei C, et al.: Boesenbergia rotunda: From Ethnomedicine to Drug Discovery. Evid. Based Complement. Alternat. Med. 2012; 2012: 473637.\n\nAbdelwahab SI, Mohan S, Abdulla MA, et al.: The methanolic extract of Boesenbergia rotunda (L.) Mansf. and its major compound pinostrobin induces anti-ulcerogenic property in vivo: possible involvement of indirect antioxidant action. J. Ethnopharmacol. 2011; 137(2): 963–970. PubMed Abstract \| Publisher Full Text\n\nGonzalez AS, Soto Tellini VH, Benjumea Gutierrez DM: Study of the dermal anti-inflammatory, antioxidant, and analgesic activity of pinostrobin. Heliyon. 2022; 8(9): e10413. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJadaun A, Sharma S, Verma R, et al.: Pinostrobin inhibits proliferation and induces apoptosis in cancer stem-like cells through a reactive oxygen species-dependent mechanism. RSC Adv. 2019; 9(21): 12097–12109. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nXian YF, Ip SP, Lin ZX, et al.: Protective effects of pinostrobin on beta-amyloid-induced neurotoxicity in PC12 cells. Cell. Mol. Neurobiol. 2012; 32(8): 1223–1230. PubMed Abstract \| Publisher Full Text\n\nKongsui R, Surapinit S, Promsrisuk T, et al.: Pinostrobin from Boesenbergia rotunda attenuates oxidative stress and promotes functional recovery in rat model of sciatic nerve crush injury. Braz. J. Med. Biol. Res. 2023; 56: e12578. Publisher Full Text\n\nLi C, Tang B, Feng Y, et al.: Pinostrobin Exerts Neuroprotective Actions in Neurotoxin-Induced Parkinson's Disease Models through Nrf2 Induction. J. Agric. Food Chem. 2018; 66(31): 8307–8318. PubMed Abstract \| Publisher Full Text\n\nMetz GA, Whishaw IQ: Cortical and subcortical lesions impair skilled walking in the ladder rung walking test: a new task to evaluate fore- and hindlimb stepping, placing, and co-ordination. J. Neurosci. Methods. 2002; 115(2): 169–179. PubMed Abstract \| Publisher Full Text\n\nLowry OH, Rosebrough NJ, Farr AL, et al.: Protein measurement with the Folin phenol reagent. J. Biol. Chem. 1951; 193(1): 265–275. Publisher Full Text\n\nLuangaram S, Kukongviriyapan U, Pakdeechote P, et al.: Protective effects of quercetin against phenylhydrazine-induced vascular dysfunction and oxidative stress in rats. Food Chem. Toxicol. 2007; 45(3): 448–455. PubMed Abstract \| Publisher Full Text\n\nNwunuji PT, Benjamin OE, Mohd YS: Changes in haematological parameters and oxidative stress response of goats subjected to road transport stress in a hot humid tropical environment. Comp. Clin. Pathol. 2016; 25: 285–293.\n\nLiu X, Wang C, Liu W, et al.: Oral Administration of Silibinin Ameliorates Cognitive Deficits of Parkinson's Disease Mouse Model by Restoring Mitochondrial Disorders in Hippocampus. Neurochem. Res. 2021; 46(9): 2317–2332. PubMed Abstract \| Publisher Full Text\n\nNagatsu T, Nakashima A, Ichinose H, et al.: Human tyrosine hydroxylase in Parkinson's disease and in related disorders. J. Neural Transm. (Vienna). 2019; 126(4): 397–409. Publisher Full Text\n\nMatsuda LA, Schmidt CJ, Hanson GR, et al.: Effect of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) on striatal tyrosine hydroxylase and tryptophan hydroxylase in rat. Neuropharmacology. 1986; 25(3): 249–255. PubMed Abstract \| Publisher Full Text\n\nSchober A, Peterziel H, von Bartheld CS , et al.: GDNF applied to the MPTP-lesioned nigrostriatal system requires TGF-beta for its neuroprotective action. Neurobiol. Dis. 2007; 25(2): 378–391. PubMed Abstract \| Publisher Full Text\n\nIravani MM, Costa S, Jackson MJ, et al.: GDNF reverses priming for dyskinesia in MPTP-treated, L-DOPA-primed common marmosets. Eur. J. Neurosci. 2001; 13(3): 597–608. PubMed Abstract \| Publisher Full Text\n\nYou H, Mariani LL, Mangone G, et al.: Molecular basis of dopamine replacement therapy and its side effects in Parkinson's disease. Cell Tissue Res. 2018; 373(1): 111–135.\n\nThongrong S, Surapinit S, Promsrisuk T, et al.: Pinostrobin alleviates chronic restraint stress-induced cognitive impairment by modulating oxidative stress and the function of astrocytes in the hippocampus of rats. Biomed. Rep. 2023; 18(3): 20. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHain EG, Sparenberg M, Rasinska J, et al.: Indomethacin promotes survival of new neurons in the adult murine hippocampus accompanied by anti-inflammatory effects following MPTP-induced dopamine depletion. J. Neuroinflammation. 2018; 15(1): 162. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRibeiro-Carvalho A, Leal-Rocha PH, Isnardo-Fernandes J, et al.: Exposure to varenicline protects against locomotor alteration in a MPTP mouse model of Parkinson's disease. Braz. J. Med. Biol. Res. 2021; 54(12): e11679. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLin ZH, Liu Y, Xue NJ, et al.: Quercetin Protects against MPP(+)/MPTP-Induced Dopaminergic Neuron Death in Parkinson's Disease by Inhibiting Ferroptosis. Oxidative Med. Cell. Longev. 2022; 2022: 7769355.\n\nKim SY, Kim MY, Mo JS, et al.: SAG protects human neuroblastoma SH-SY5Y cells against 1-methyl-4-phenylpyridinium ion (MPP+)-induced cytotoxicity via the downregulation of ROS generation and JNK signaling. Neurosci. Lett. 2007; 413(2): 132–136. PubMed Abstract \| Publisher Full Text\n\nVoss T, Ravina B: Neuroprotection in Parkinson's disease: myth or reality? Curr. Neurol. Neurosci. Rep. 2008; 8(4): 304–309.\n\nGao Y, Tang H, Nie K, et al.: Hippocampal damage and white matter lesions contribute to cognitive impairment in MPTP-lesioned mice with chronic cerebral hypoperfusion. Behav. Brain Res. 2019; 368: 111885. PubMed Abstract \| Publisher Full Text\n\nArenas E, Trupp M, Akerud P, et al.: GDNF prevents degeneration and promotes the phenotype of brain noradrenergic neurons in vivo. Neuron. 1995; 15(6): 1465–1473. PubMed Abstract \| Publisher Full Text\n\nGottschalk CG, Jana M, Roy A, et al.: Gemfibrozil Protects Dopaminergic Neurons in a Mouse Model of Parkinson's Disease via PPARalpha-Dependent Astrocytic GDNF Pathway. J. Neurosci. 2021; 41(10): 2287–2300. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nXu K, Dluzen DE: The effect of GDNF on nigrostriatal dopaminergic function in response to a two-pulse K(+) stimulation. Exp. Neurol. 2000; 166(2): 450–457. PubMed Abstract \| Publisher Full Text\n\nGrondin R, Cass WA, Zhang Z, et al.: Glial cell line-derived neurotrophic factor increases stimulus-evoked dopamine release and motor speed in aged rhesus monkeys. J. Neurosci. 2003; 23(5): 1974–1980. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKhan NM, Sandur SK, Checker R, et al.: Pro-oxidants ameliorate radiation-induced apoptosis through activation of the calcium-ERK1/2-Nrf2 pathway. Free Radic. Biol. Med. 2011; 51(1): 115–128. PubMed Abstract \| Publisher Full Text\n\nde Oliveira MR , Peres A, Gama CS, et al.: Pinocembrin Provides Mitochondrial Protection by the Activation of the Erk1/2-Nrf2 Signaling Pathway in SH-SY5Y Neuroblastoma Cells Exposed to Paraquat. Mol. Neurobiol. 2017; 54(8): 6018–6031. PubMed Abstract \| Publisher Full Text\n\nThongrong S: Pinostrobin mitigates neurodegeneration through an up-regulation of antioxidants and GDNF in a rat model of Parkinson’s disease. figshare. 2023. Publisher Full Text" }	[ { "id": "205415", "date": "16 Oct 2023", "name": "Ana C Dutra-Tavares", "expertise": [ "Reviewer Expertise neurophysiology", "neuropsychiatric disorders animal models." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIn the present paper the authors investigated the effects of a flavonoid substance with anti-inflammatory effects on a rodent model of Parkinson’s disease.\nThe findings are interesting since they observed neurochemical, morphological and behavioral reversion of alterations associated with this disease by the pinostrobin. Some comments will be made bellow to improve manuscript and provide to the reader more detailed information.\n1 - Parkinson’s disease is characterized by motor impairments but also cognitive decline and psychiatry disorders, such as depression, could be present. Why did the authors focus only on motor dysfunction? Is there evidence that pinostrobin would be only effective on this type of symptom? Is there evidence that pinostrobin could be effective on the other symptoms domains? Although the authors did not investigate that, a more a broader discussion about other possible therapeutic effects and/or limitations to the pinostrobin application on Parkinson’s disease would benefit the reader.\n2 – It is kind of hard to understand how the substantia nigra was used for biochemical assays and to immunohistochemistry. Was substantia nigra from each hemisphere used to different techniques? It would be helpful if the authors provide more detailed information about the dissection.\n3 – The authors present the staining results in percentage from control group. In this sense, the mean value of control group is equal to 100%, but there is a variability among the individuals. Thus, it is always good to present the error bar to show the variability of the data for this group.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? No source data required\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10406", "date": "30 Nov 2023", "name": "Sitthisak Thongrong", "role": "Author Response", "response": "Thank you for your valuable comments that we have followed them for improving the manuscript. Here, we have focused on the effects of pinostrobin in a rat model of Parkinson's disease. 1. We have studied that pinostrobin could improve cognitive impairment induced by chronic stress (reference number 32) that is why here we focused only on the motor function. However, this must be our weak point because we did not observe the effect of pinostrobin on cognitive decline and psychiatry disorders so we will put this in the limitation of this study. 2. We used the right hemisphere for biochemical assays and the left hemisphere for histological studies. The way to dissect the midbrain including substantia nigra was added to the manuscript. 3. Your comment is worth for us. We made the normalization that is why the control groups did not show the error bar. However, the F1000research let us show all the raw data that the readers can access them whenever they want." } ] } ]	1	https://f1000research.com/articles/12-846
https://f1000research.com/articles/10-650/v1	26 Jul 21	{ "type": "Research Article", "title": "Evaluation of the convalescent plasma therapy effectiveness and the factors that influence the therapeutic outcome in hospitalized COVID-19 patients: A retrospective cohort study.", "authors": [ "Zainab Ibadi", "Hayder Assad", "Hayder Fawzi", "Zainab Ibadi", "Hayder Assad" ], "abstract": "Background: As an effective antiviral therapy is not available for the treatment of the current rapidly and continuously spreading coronavirus disease (COVID-19), it is very crucial to find an alternative treatment strategy. Convalescent plasma (CP) therapy has been used for prevention and treatment of many emerging infectious diseases, however, the results of current studies on CP in COVID-19 are not consistent. Therefore, this study aimed to evaluate the effectiveness of CP therapy in hospitalized patients with COVID-19, while evaluating patient and donor-related factors that might influence the therapeutic outcome. Methods: We conducted a retrospective cohort study on 312 patients with either severe or critical COVID-19, who were admitted to Al-Hakeem and Al-Amal hospitals in Al-Najaf city, Iraq from June to August 2020. The patients were allocated to either the plasma therapy group (152 patients) who received CP combined with standard therapy or the standard therapy group (160 patients). The outcome measures were the 21-day mortality rate and time to clinical improvement. Results: The overall cumulative survival rate was significantly higher in patients who received CP compared to standard therapy alone at 21 days (68.3% vs. 46.8%, p-value = 0.010), with mean survival at 17.6 vs. 15.3 days, (p-value = 0.010). In multivariate analysis, the plasma therapy effect was an independent predictor of survival (adjusted hazard ratio, 95% confidence interval: 0.368, 0.177 – 0.765). In terms of clinical improvement, the use of CP resulted in shorter clinical improvement (median duration of improvement: 8 vs. 11 days, p-value = 0.007), with 74.3% improvement rate after 21 days in CP compared to 65.0% in standard therapy. Conclusions: Therapy with CP in combination with standard therapy, independently improved survival in hospitalized patient with severe or critical COVID-19.", "keywords": [ "COVID-19", "SARS-CoV-2", "Convalescent plasma", "mortality", "clinical improvement" ], "content": "Introduction\n\nSince the emergence of the new coronavirus COVID-19 outbreak in late 2019, the outbreak has continued to spread exponentially across the world. As of early April 2021, the global cumulative incidence has exceeded 130 million reported cases, with approximately three million associated deaths.1 To date, an effective antiviral therapy for this disease does not exist. Therefore, it is imperative to find an alternative treatment strategy, especially for severe COVID-19 patients.2,3 For more than a century, convalescent plasma (CP) therapy, as a classic adoptive immunotherapy, has been used to effectively prevent and treat many infectious diseases. CP therapy has been successfully used in the treatment of severe acute respiratory syndrome (SARS), Middle East respiratory syndrome (MERS), and the 2009 H1N1 pandemic with adequate effectiveness and protection over the past two decades.4,5 For patients with serious or life-threatening COVID-19, the United States of America (USA) Food and Drug Administration (FDA) approved the emerging use of CP at the end of March 2020. While the use of CP might be a promising therapeutic approach, the results from previous studies are not consistent and the evidence supporting its use for the treatment of COVID-19 are not clear, therefore there is a need for further investigations.2,6\n\nMany factors might influence the therapeutic effectiveness of CP, such as patients’ age, comorbidities, disease stage, and concomitant treatment.7,8 Other factors are related to the plasma donor, most importantly titer of the antibody. Additional factors are associated with the plasma administration protocol such as the time of administration and volume of plasma.9,10\n\nBased on the high number of patients that were treated with CP in AL Najaf, Iraq during the first wave of this pandemic, we designed this study to evaluate the effectiveness of CP therapy in hospitalized patients with COVID-19, by analyzing the variables related to the patients, donors and the plasma administration practice.\n\n\nMethods\n\nA multicenter, retrospective cohort study was conducted on adult patients that were diagnosed with severe or critical COVID-19. The study involved a retrospective assessment of the medical records of patients who were admitted to tertiary specialized hospitals (Al-Hakeem and Al-Amal hospitals). Based on the treatment protocol whether it included CP therapy or not, the patients were allocated into two groups; the standard therapy group, that consisted of patients who received the standard therapy which mainly included antiviral agents, antibiotics, steroids and anticoagulants, according to the Iraqi ministry of health (MOH) approved guidelines.11 The second group was the plasma therapy group that consisted of patients who received CP combined with the standard therapy.\n\nThe study included patients that were admitted to Al-Hakeem and Al-Amal hospitals from June 1, 2020 till August 31, 2020, in Al-Najaf Province, Iraq. Al-Hakeem hospital is the first center for the admission of COVID-19 patients in Najaf, and it contains 264 beds. Al-Amal hospital has 170 beds, including 50 intensive care unit (ICU) beds, and it has been assigned as a referral center for severe and critical cases of COVID-19.\n\nThe Scientific Research Committee at Al-Najaf Health Directorate approved this study which involved Al-Hakeem hospital and the main blood bank with the approval number (19421), and Al-Amal hospital with approval number (30418). This study was also granted approval by the ethics and scientific committee of faculty of pharmacy/the University of Kufa with approval number (2403). All participants gave written informed consent to participate in this study. The current study is registered at Clinical Trials.gov (https://clinicaltrials.gov/ct2/show/NCT04764747) with the registration number (NCT04764747).12\n\nEligible participants were 18 years of age or older, with COVID-19 infection confirmed by reverse transcriptase-polymerase chain reaction (RT-PCR) test, who were hospitalized with severe or critical case of COVID-19. We excluded patients who did not require oxygen therapy, patients that were discharged or died within 48 hours of admission, patients with a history of allergy to plasma transfusion, and those with incomplete data on their medical records.\n\nThe study participants with severe COVID-19 were defined as patients who had oxygen saturation of less than or equal to 93%, therefore, they needed oxygen therapy. Patients with critical cases of COVID-19 were defined as patients who needed mechanical ventilation and ICU observation.2,13\n\nThe medical records of patients diagnosed with COVID-19 were taken from the archive unit of the tertiary specialized public hospital and reviewed retrospectively. Data collection period was from September 2020 to February 2021. Data was collected by using a chart extraction sheet which is used to collect information from patients' medical records. These medical records were selected based on chronological categorization in the statistical unit according to admission date. Blind data abstraction was performed for this study. Also, two senior physicians reviewed the data extracted from the medical records and approved the abstraction process. The chart extraction sheet consists of patients' demographics (age, gender, occupation, address), comorbidities (hypertension, coronary artery disease, type 2 diabetes mellitus (T2DM), obstructive airway disease, and chronic kidney disease), date of admission, date of discharge, length of stay in the hospital, symptoms (fever, cough, shortness of breath, fatigue, headache, sore throat, nausea and vomiting, oxygen-support categories were defined either as ambient air, low-flow oxygen, high-flow oxygen, and mechanical ventilation,14 vital signs on admission (temperature, heart rate, oxygen saturation (SpO2%)). Additionally, laboratory parameters such as white cell counts, lymphocytes count, serum creatinine, D-dimer, ferritin, aspartate aminotransferase (AST), and alanine aminotransferase (ALT), were included. Lastly, type of medications, and outcomes, which was defined as in hospital mortality and clinical improvement (according to the modified World Health Organization (WHO) ordinary scale.),13,14 were considered in the chart sheet. Information of plasma donor that was obtained from the plasma unit of the main blood bank consisting of donors' demographics, ID code of plasma bottle, and immunoglobulin G (IgG) titer (See Underlying data: http://doi.org/10.5281/zenodo.498205215) was also included.\n\nIn this study the primary outcome was the in-hospital mortality rate (MR) (number of deaths with a 21-day time frame). The secondary outcome was the association with various predictors (sociodemographic, clinical, laboratory variables and medications). Both these outcomes were examined with uni- and multivariate analysis.\n\nThe time to clinical improvement was also assessed. Clinical improvement for COVID-19 was defined as a reduction of two-points in a modified six-point scale, discharged or achieved criteria of discharge, which was defined as clinical recovery (i.e., SpO2 equal to 94% on room air, fever subsided, and relief of cough, all continued for at least 72 hours) (one-point); hospitalized without oxygen therapy (two-points); hospitalized on low-flow oxygen therapy (three-points); hospitalized on high-flow oxygen therapy (four-points); hospitalized and required mechanical ventilation (five-points); death (six-points).14,16\n\nThe sample size was based on all medical records available in the archive unit for patients admitted to these public hospitals during June, July, and August 2020. Total of 312 patients included in the statistical analysis as illustrated in (Figure 1). Post hoc power analysis of sample size based on mortality outcome and level of significance at alpha = 0.05 (type I error) yielded statistic power of 97.3% (1- type II error) in detecting the deference between the two group of the study.\n\nNormality test was performed for all variables. Normally distributed mean was used to present the data, while for variables with non-normal distribution median and interquartile range (IQR) (25% to 75% percentile range) were used. Chi-square test or Fisher exact test was used to analyze the discrete variable. To assess the difference in continuous variables; two-samples t-test (in normally distributed variables), and Mann-Whitney U test was used in variables with non-normal distribution. Kaplan–Meier analysis17 was used to estimate the median time of the cumulative percentage of survival or clinical improvement, the Log-rank test was used to calculate the p-value and to compare the significance between the two groups. Lack of clinical improvement at day 21 and death before day 21 was considered as right censored for that day.\n\nUnivariate analysis was performed using Cox regression for all patients’ variables, and if p-value < 0.1, the variables were included in the multivariate Cox proportional hazard model to allow adjustment of certain confounder variables. The hazard ratio (HR) was calculated by using the Cox proportional hazard regression analysis, to find the time-dependent association of the model, in order to calculate the 95% confidence interval. Missing data were excluded from the final analysis.\n\nSPSS 22.0.0 (Chicago, IL), GraphPad Prism version 8.1.0 for Windows (https://www.graphpad.com/) was used for the statistical analysis. P-value was considered statically significant if less than 0.05.\n\n\nResults\n\nIn this cohort the mean age of the patients was 55.1(±13.6) years, and 70.2% were male. Overall, 82% of the hospitalized patients had cough and 81.4% had shortness of breath (SOB). The mean SpO2 was 82.7 % (which is consistent with the disease severity stage of the patients included in the study). All baseline characteristics of patients’ demography, comorbidities and clinical parameters of the two groups were not statistically significant, except for azithromycin and lopinavir/ritonavir that were significantly higher in patients treated with plasma therapy, while acetaminophen and favipiravir were significantly higher in patients treated with the standard therapy (Table 1). Plasma characteristics are illustrated in Table.\n\nThe survival rate was 90.2% in the plasma therapy group compared to 84.2% in the standard therapy group, after 7 days. Additionally, the survival rate in the plasma therapy group was significantly higher (77.7%) compared to the standard group (60.5%) after 14 days (p-value = 0.010). This trend continued after 21 days in the plasma group (68.3%) vs. (46.8%) in the standard group, with a mean survival time of 17.6 in the plasma group and 15.3 in the standard therapy group (Table 3).\n\nKaplan-Meier survival analysis revealed statistically significant improvement in the survival in the plasma therapy group, compared with the standard therapy group (Figure 2).\n\nThe univariate analysis showed that the use of plasma compared to standard therapy resulted in a reduced risk of death by 54.7%. The results from the overall study population included in the multivariate model showed that plasma therapy has a statistically significant effect on improving the survival rate and it can be considered as an independent predictor of mortality. This adjusted analysis revealed a further reduction in the HR of the plasma therapy group from 0.557 (0.352-0.883) in unadjusted analysis to 0.368 (0.177-0.765). This meant that the reduction in the risk of death reached 63.2%, thus indicating that plasma therapy is more effective in improving survival rates. The increase in white blood cells (WBC), D-Dimer, and T2DM were independent predictors of mortality rate, while both the use of plasma and rivaroxaban reduced the risk of MR independently, as shown in Table 4.\n\nIn the multivariate analysis for the plasma therapy group, the result showed that as independent predictors of mortality, T2DM and D-Dimer are associated with increased mortality, while high antibody titer is associate with decrease mortality as illustrated in Table 5.\n\nInterestingly, the median time to improvement was three days faster in plasma therapy than standard therapy (8 days vs. 11 days). Kaplan-Meier analysis showed a significant statistical difference between the two groups. The cumulative incidence of clinical improvement was significantly higher in patients who received plasma therapy (44.1%) then those who received standard treatment (18.1%), after 7 days. Treatment after 14 days indicated that the cumulative incidence of clinical improvement was higher in the plasma therapy (67.1%) vs. standard therapy group (58.8.9%). Additionally, 21 days post-treatment the cumulative incidence of clinical improvement was still significantly higher in plasma therapy (74.3%) compared to standard therapy group (65.0%) (Table 6 and Figure 3)\n\n\nDiscussion\n\nCOVID-19 is a rapidly evolving infection that lacks an effective antiviral treatment. CP therapy as an available treatment can potentially be an effective and lifesaving treatment for this disease.9 Several previous outbreaks of respiratory viral infection have been treated by CP such as SARS-CoV and MERS-CoV18. However, in patients with COVID-19, the effectiveness of CP is not consistent, and it is dependent on many prognostic factors,9,19 therefore, we designed this study to evaluate the effectiveness of CP in hospitalized patients with COVID-19 to explore the factors related to the patients, donors and CP administration practice that might influence the therapeutic outcome.\n\nIn this retrospective cohort study, the plasma therapy showed considerable survival improvement as evident by a reduction in the risk of in-hospital mortality after 21 days, compared to the patients who received standard therapy. Also, the time to clinical improvement was significantly faster in patients on CP combined with standard therapy, than those on standard therapy alone. The plasma therapy had three days shorter median time to clinical improvement compared to the standard therapy group. Additionally, the cumulative incidence of clinical improvement was 74.3% in the plasma therapy group vs. 65% in the standard therapy group.\n\nThese positive effects of plasma therapy on survival rate and clinical improvement in the present study are in line with the findings from a retrospective cohort study by Xinyi Xia et al.20 In this study, 1568 patients with severe and critical COVID-19 were grouped into 1430 patients who received standard therapy and 138 patients who received plasma therapy.20 The results indicated that the patients had improved clinical symptoms and mortality after the transfusion with CP.20\n\nSimilarly, a prospective study conducted on 316 patients with severe and/or critical COVID-19 infection, showed that the administration of CP significantly reduced mortality and improved clinical symptoms.21 Additionally, improved clinical symptoms and mortality after administration of CP have been recently reported by a multicenter non-randomized prospective interventional cohort study on patients with a moderate or severe stage of COVID-19.22\n\nStrong evidence supporting these beneficial effects of plasma therapy on survival and clinical improvement was also reported in several SARS and COVID-19 meta-analysis studies.10,23,24\n\nThe positive effects of CP on patient survival and clinical improvement may be attributed to the antiviral mechanism of CP via neutralizing antibodies that enhance the viral clearance and provides immunomodulation via the anti-inflammatory effect. Furthermore, the control of overactive immune response by preventing complement activation and blocking autoantibodies, as well as regulating the hypercoagulable state, can prevent lung damage and bad prognosis.25\n\nHowever, the results from plasma therapy are not always consistent, as indicated by the results of a particular systematic review and meta-analysis, which included 10 randomized control trials (RCTs) with over 10,000 patients.26 Simonovich et al., demonstrated that CP therapy was not beneficial for mortality and improvement rate in severe patients with COVID-19, however the time of plasma transfusion was considered to be the limitation of their study.27 In another RCTs, CP therapy did not reduce mortality or the progress in disease severity, however, about 2/3 of patients received plasma with low antibodies titer, and the remaining patients receiving plasma without antibody detections.28 Also, the preliminary analysis from a very recent RTCs did not show survival improvement in hospitalized patients with either confirmed or suspected COVID-19. The possibility of different virus variants between the donor and recipient and the time of CP administration, could explain the non-significant findings of CP in COVID-19 in this study.29 In general, these discrepancies and mixed results regarding the benefits of CP in COVID-19 mainly is dependent on many patients related factors such as age, comorbidities, clinical characteristics, degrees of disease severity, as well as variations in medication use in standard therapy. Most importantly the donors' related factors such as antibody titers must be considered, along with the plasma administration protocol including the volume administered and the time of administration.9,10,19\n\nIn the present study, the confounding effects of these variables on the primary outcome were explored by multivariate analysis which confirmed the beneficial effect of plasma in reducing the risk of mortality independent of other variable effects. It is worth mentioning that most patients in this study were administered plasma therapy within the first two days of admission to the hospital. Also, the plasma average antibody IgG index was high (16.7 ± 10.8), which is considerably associated with a reduced risk of death. Consistently, a retrospective study in the USA on 3082 hospitalized patients with COVID-19 has revealed that the transfused plasma with high IgG antibody levels were correlated with a lower risk of mortality, compared to the transfused plasma with low antibody titer.30 Regarding the time of administration, RCTs in Argentina that included 160 elderly patients, showed that the early administration of CP within 72 hours of the onset of symptoms was associated with a lower risk of progression to severe COVID-19 and a lower risk of death, in comparison to the control group.31 T2DM patients and individuals with a high level of D-dimer and WBC were associated with an increased risk of death. Comparable findings were also reported from previous studies.32,33\n\nOur study has some limitations. Firstly, the study design was retrospective, which was subjected to high effect of the cofounders. We tried to reduce that risk by increasing the smaple size (reduce type II error); in addition adjusted analysis (multivariate regression) was used, in order to reduced the effect of cofounders. Second, the time from onset of symptoms to hospital admission was not known, however at the beginning of the pandemic in Iraq, most of the patients were referred to the hospital, once their PCR test was positive.\n\n\nConculsion\n\nTherapy with PC in combination with standard therapy independently improved survival in hospitalized patient with severe or critical COVID-19. We recommend a large double-blind randomized trial with emphasis on controlling the factors related to the plasma administration protocols.\n\n\nData availability statement\n\nZenodo: Evaluating the effectiveness of convalescent plasma therapy and the factors that influence the therapeutic outcome in hospitalized COVID-19 patients: A retrospective cohort study.\n\nhttp://doi.org/10.5281/zenodo.4982052.31\n\nThe project contains the following underlying data:\n\nDatabase: Data for CP in COVID-19 patients.\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).", "appendix": "Acknowledgments\n\nThe authors wish to express their gratitude to statistic unit staff in Al-Hakeem Hospital and Al-Amal Hospital, and medical staff in the plasma unit in the main blood center in Najaf, Iraq for their collaboration.\n\n\nReferences\n\nWHO Coronavirus (COVID-19) Dashboard\|WHO Coronavirus (COVID-19) Dashboard With Vaccination Data. Accessed June 4, 2021. https://covid19.who.int/\n\nLi L, Zhang W, Hu Y, et al.: Effect of Convalescent Plasma Therapy on Time to Clinical Improvement in Patients with Severe and Life-threatening COVID-19: A Randomized Clinical Trial. JAMA - J Am Med Assoc. 2020; 324(5): 460–470. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWang C, Li W, Drabek D, et al.: A human monoclonal antibody blocking SARS-CoV-2 infection. Nat Commun. 2020; 11(1): 1–6. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDuan K, Liu B, Li C, et al.: Effectiveness of convalescent plasma therapy in severe COVID-19 patients. Proc Natl Acad Sci U S A. 2020; 117(17): 9490–9496. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBeigel JH, Aga E, Elie-Turenne MC, et al.: Anti-influenza immune plasma for the treatment of patients with severe influenza A: a randomised, double-blind, phase 3 trial. Lancet Respir Med. 2019; 7(11): 941–950. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGharbharan A, Jordans CCE, GeurtsvanKessel C, et al.: Convalescent plasma for COVID-19. A randomized clinical trial. MEDRxiv. 2020. Publisher Full Text\n\nEnzmann MO, Erickson MP, Grindeland CJ, et al.: Treatment and Preliminary Outcomes of 150 Acute Care Patients with COVID-19 in a Rural Health System in the Dakotas. Epidemiol Infect. 2020: 16–21. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYan Y, Yang Y, Wang F, et al.: Clinical characteristics and outcomes of patients with severe covid-19 with diabetes. BMJ Open Diabetes Res Care. 2020; 8(1): 1–9. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBrown BL, McCullough J: Treatment for emerging viruses: Convalescent plasma and COVID-19. Transfus Apher Sci. 2020: 102790. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBansal V, Mahapure KS, Mehra I, et al.: Mortality Benefit of Convalescent Plasma in COVID-19: A Systematic Review and Meta-Analysis. Front Med. 2021; 8(April). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nA guide to WHO’s guidance on COVID-19. Accessed June 5, 2021. Reference Source\n\nEffectiveness of Convalescent Plasma in Hospitalized Patients With COVID-19 - Full Text View - ClinicalTrials.gov. Accessed June 21, 2021. Reference Source\n\nLi H, Xiong N, Li C, et al.: Efficacy of ribavirin and interferon-α therapy for hospitalized patients with COVID-19: A multicenter, retrospective cohort study. Int J Infect Dis. 2021; 104: 641–648. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKewan T, Covut F, Al–Jaghbeer MJ, et al.: Tocilizumab for treatment of patients with severe COVID–19: A retrospective cohort study. EClinicalMedicine. 2020; 24. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFawzi HA: Data of CP in COVID-19 patients.June 18, 2021. Publisher Full Text\n\nWang Y, Zhang D, Du G, et al.: Remdesivir in adults with severe COVID-19: a randomised, double-blind, placebo-controlled, multicentre trial. Lancet. 2020; 395(10236): 1569–1578. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRanstam JJAC: Kaplan–Meier curve. Br J Surg. 2017; 104(4): 434–442. Publisher Full Text\n\nWooding DJ, Bach H: Treatment of COVID-19 with convalescent plasma: lessons from past coronavirus outbreaks. Clin Microbiol Infect. 2020; 26(10): 1436–1446. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJoyner MJ, Senefeld JW, Klassen SA, et al.: Effect of Convalescent Plasma on Mortality among Hospitalized Patients with COVID-19: Initial Three-Month Experience. medRxiv Prepr Serv Heal Sci. 2020: 1–31. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nXia X, Li K, Wu L, et al.: Improved clinical symptoms and mortality among patients with severe or critical COVID-19 after convalescent plasma transfusion. Blood. 2020; 136(6): 755–759. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSalazar E, Christensen PA, Graviss EA, et al.: Treatment of Coronavirus Disease 2019 Patients with Convalescent Plasma Reveals a Signal of Significantly Decreased Mortality. Am J Pathol. 2020; 190(11): 2290–2303. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAlsharidah S, Ayed M, Ameen RM, et al.: COVID-19 convalescent plasma treatment of moderate and severe cases of SARS-CoV-2 infection: A multicenter interventional study. Int J Infect Dis. 2021; 103: 439–446. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVegivinti CTR, Pederson JM, Saravu K, et al.: Efficacy of convalescent plasma therapy for COVID-19: A systematic review and meta-analysis. J Clin Apher. November 2020; 2021: 1–13. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMair-Jenkins J, Saavedra-Campos M, Baillie JK, et al.: The effectiveness of convalescent plasma and hyperimmune immunoglobulin for the treatment of severe acute respiratory infections of viral etiology: A systematic review and exploratory meta-analysis. J Infect Dis. 2015; 211(1): 80–90. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRojas M, Rodríguez Y, Monsalve DM, et al.: Convalescent plasma in Covid-19: Possible mechanisms of action. Autoimmun Rev. 2020; 19(7): 102554. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJaniaud P, Axfors C, Schmitt AM, et al.: Association of Convalescent Plasma Treatment With Clinical Outcomes in Patients With COVID-19: A Systematic Review and Meta-analysis. Jama. 2021; 325(12): 1185–1195. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSimonovich VA, Burgos Pratx LD, Scibona P, et al.: A Randomized Trial of Convalescent Plasma in Covid-19 Severe Pneumonia. N Engl J Med. 2020: 1–11. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAgarwal A, Mukherjee A, Kumar G, et al.: Convalescent plasma in the management of moderate covid-19 in adults in India: Open label phase II multicentre randomised controlled trial (PLACID Trial). BMJ. 2020; 371: 1–10. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHorby PW, Estcourt L, Peto L, et al.: Convalescent plasma in patients admitted to hospital with COVID-19 (RECOVERY): a randomised, controlled, open-label, platform trial. medRxiv. 2021. Publisher Full Text\n\nJoyner MJ, Carter RE, Senefeld JW, et al.: Convalescent Plasma Antibody Levels and the Risk of Death from Covid-19. N Engl J Med. 2021; 384(11): 1015–1027. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLibster R, Pérez Marc G, Wappner D, et al.: Early High-Titer Plasma Therapy to Prevent Severe Covid-19 in Older Adults. N Engl J Med. 2021: 610–618. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLi G, Deng Q, Feng J, et al.: Clinical Characteristics of COVID-19 Patients with and without Diabetes in Wuhan Red Cross Hospital. J Diabetes Res. 2020; 2020. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nShah S, Shah K, Patel SB, et al.: Elevated d-dimer levels are associated with increased risk of mortality in coronavirus disease 2019: A systematic review and meta-analysis. Cardiol Rev. 2020; 28(6): 295–302. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "145184", "date": "16 Sep 2022", "name": "Daniele Focosi", "expertise": [ "Reviewer Expertise COVID-19 convalescent plasma", "emerging pathogens" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis is a non-randomized multicenter retrospective cohort trial in Iraq. It was conducted at the time of the wild-type variants, so convalescent plasma was collected from unvaccinated donors.\nThe Methods section is lacking information about anti-Spike IgG assay (the authors report outputs in Table 2 without unit of measurement), which leaves the doubt about potency. The time before plasma transfusion (median 2 days) should be better specified if calculated from the onset of symptoms or (more likely) from hospitalization. This is an important parameter that, together with the undisclosed serostatus before transfusion, could let us better understand whether the patients were still in the time window in which benefit from convalescent plasma is expected.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10156", "date": "05 Sep 2023", "name": "Hayder Fawzi", "role": "Author Response", "response": "Dear reviewer Greetings Thank you for you important notes that require a clarification: Regarding information about anti-Spike IgG assay About 5 ml of venous blood was drawn from all subjects. The blood was lifted to clotted and then centrifuged to separate the serum. The separated serum kept in deep freezing (-20C°) until used in the measuring of IgG level. Measurement was conducted by using a Vitek Immuno Diagnostic Assay System device (VIDAS®, bioMérieux, France) and following the manufacturer's instructions. For the estimation of SARS-CoV-2 IgG in persons, 100 μl of the serum were added to the sample well, standard, negative, and positive controls in the plate. The procedure of VIDAS was conducted according to the guidelines of kit manufacturer anti-SARS-CoV-2 IgG antibodies. Results were calculated automatically, the positive Results (index ≥ 1), and the negative results (index < 1), and the unit is a binding antibody unit per milliliter (BAU/mL) [1]. Regarding the time before plasma transfusion It was calculated from the from date of admission (i.e., hospitalization). 1. Al-Tamimi M, Tarifi AA, Qaqish A, Abbas MM, Albalawi H, Abu-Raideh J, et al. Immunoglobulins response of COVID-19 patients, COVID-19 vaccine recipients, and random individuals. PloS one. 2023;18(2):e0281689. Best regards" } ] } ]	1	https://f1000research.com/articles/10-650
https://f1000research.com/articles/12-26/v1	09 Jan 23	{ "type": "Data Note", "title": "A dataset about Spanish young people’s digital skills, use of technology and online platforms", "authors": [ "Daniel Aranda", "Leila Mohammadi", "Mireia Montaña Blasco", "Elisenda Estanyol", "Pedro Fernández-de-Castro", "Leila Mohammadi", "Mireia Montaña Blasco", "Elisenda Estanyol", "Pedro Fernández-de-Castro" ], "abstract": "Background: The datafication scenario of the current communicative ecosystem poses a challenge to media and digital literacy, especially in terms of participation and civic and democratic engagement of youth. Methods: For this purpose, through a survey with a representative sample of 600 young people in Spain, between 16 and 18 years old, we observed their level of digital literacy through three variables: technical competencies, informational competencies, and critical knowledge. This dataset also collects information on the reasons why young people use digital technology such as video games, consoles, computers or mobile phones. On the other hand, we also offer information on the types of social networks or applications and the time and types of uses by youngsters of different digital technologies and social media platforms. The survey includes socio-demographic factors such as gender including (male, female, and others). Conclusions: This survey offers researchers relevant data on the digital skills of Spanish youth and on the perceptions of the use of different digital technologies. This paper also reports the main descriptive data that can be expanded by researchers accessing the database.", "keywords": [ "Digital literacy competencies", "participation", "civic engagement", "digital technology use", "digital skills", "ICT in education", "youth", "technical competencies", "informational competencies", "critical knowledge." ], "content": "Introduction\n\nYoung people need skills and competencies to make the most of the benefits of the internet and digital media. This is especially relevant to bridge the socioeconomic and political and cultural participation gap (Livingstone et al., 2021). Thus, the project in which this dataset frames, explores how certain uses of the internet and social media allow young people to position themselves and stand as political actors or performers involved in social, cultural and economic political life. In this sense, the project includes four sections: use of different technological devices, digital literacy competencies, ICT use in formal and informal learning, and the effect of using digital technologies on social life. To include the impact of COVID-19 crisis on digital social education, the project specifies some relevant questions to during and after COVID-19 lockdown.\n\n\nMethods\n\nInformed consent was obtained from participants. The questionnaire was sent to a panel of people from 16 to 18 years of age via Dynata data-platform, that complies with all legal matters in relation to data protection and response protocols. Those aged 16 to 18 answered the children themselves but with the consent of the parents, which were present while they answered as the legislation indicates.\n\nData and participant security and confidentiality were respected following the UNE EN ISO/IEC 27001 standards and the favorable report issued by the Universitat Oberta de Catalunya (UOC) Ethics Committee under file CE22-PR05.\n\nResearch design is quantitative and cross-sectional, involving an online survey intended to measure the self-perceived digital competence of young people aged 16 to 18 living in Spain. This age group was chosen because they need specific skills in order to take full advantage of the benefits of the Internet and social media, especially with regard to reducing the gap in their political and cultural participation (Theben, 2021). The survey was self-administered, i.e., it was completed by the respondents themselves without the presence of an interviewer, between 23rd September and 5th October 2021.\n\nThe survey is based on studies by Van Deursen et al. (2016) and Aranda et al. (2020), as well as an extension of the Oxford Internet Institute’s (OxIS) WIP Britain 2013, combined with a systematic review of the notion of “digital youth work” (Fernández-de-Castro et al., 2021). It includes a section of socio-demographic factors begins with gender (male/female, also they have been given the open answer option of other if they perceived themselves non-binary), age, place of residence, the population of the city, level of education, etc. Afterwards there are four further sections comprising of 24 questions about the respondents’ self-perceived digital competence. Because the survey asked young people about how they perceive their own digital skills and knowledge, their answers may not necessarily match their actual competence level.\n\nDue to the importance of COVID-19 crisis in these recent years, the survey includes questions related to using digital technologies for formal and informal education during and after COVID-19 lockdown. Education was an important sector highly affected by the COVID-19 crisis because of the rapid change of teaching to online form. In May 2020, 56.6 % of the total number of students have been affected worldwide and schools had been forced to close in 130 countries (Donohue & Miller, 2020). Spain lockdown started on 15th March 2020 until 21st June 2020 and with the start of the new school year in September 2020 most schools reopened.\n\nThe first section of the survey, digital technology use and lockdown, begins by asking the participants about use of different technological devices (mobile, tablet, computer, game console and smart watch) and the purposes for which they are used (work, information, training, communication, entertainment and political participation) during and after lockdown. This section continues with three questions about comparing the spending time and its quality on online activities during lockdown (between 15th March 2020 and 21st June 2020) and after lockdown. The answers were given on a 3-point Likert scale 1: less; 2: the same; 3: more.\n\nThe second section is about digital competence and asked about technical skills (nine items), informational skills (ten items), and critical digital knowledge (five items), respectively. Answers were given on a 5-point Likert scale: 1: I don’t know what this is or what it means; 2: I know what this is but I don’t know how to do it; 3: I would know how to do this with help; 4: I know how to do this by myself; and 5: I know how to do this and could teach others. The third section dealt with critical knowledge of the digital environment and included five items, also measured on a 5-point Likert scale: 1: Nothing at all; 2: A little bit; 3: An average amount; 4: A fair amount; and 5: A lot.\n\nThe third section on ICT use in formal and informal learning asked about self-learning (three items) and nine questions of online formal education which included; training in digital technologies and engagement (four items), online classes and teaching initiatives (five items).\n\nThe fourth section is about youth perception and digital technologies, asking the young people to rank five social ability items considering the relevance of digital technologies for them. Then they were asked to rank the effect of using digital technologies on ten social life aspects. Answers were given on a 5 point Likert scale from very negatively (1) to very positively (5).\n\nA principal component analysis (PCA) was carried out on the proposed scales to check their validity and Cronbach’s alpha was used to measure their reliability. Regarding the first section about uses of digital technologies, the analysis showed an acceptable structure for all items Kaiser–Meyer–Olkin (KMO) test results in a mean of 0.738 and the Bartlett test is significant p<0.001). The PCA results offer a structure of a single component that explains 59.5% of the variance. The reliability according to Cronbach’s Alpha coefficient is 0.755.\n\nRegarding the section on technical digital skills, the analysis showed an acceptable structure for all nine items (KMO=0.910; Bartlett’s test significant with p<0.001). The structure comprised two components explaining 64.8% of the total variance (40.1% for the first component and 24.7% for the second); the Cronbach’s alpha coefficient was 0.903 for the first component and 0.773 for the second. For the section on informational digital skills, the analysis showed an acceptable structure for all ten items (KMO=0.955; Bartlett’s test significant with p<0.001). A single component explained 59.9% of the total variance and Cronbach’s alpha was 0.925. For the section on critical digital knowledge, the analysis showed an acceptable structure for all five items (KMO=0.843; Bartlett’s test significant with p<0.001). A single component explained 58% of the total variance and Cronbach’s alpha was 0.819.\n\n\nSample of study\n\nThrough a simple random sampling strategy, 600 youth completed the questionnaire, with an average duration of 13 minutes per person. The sample response rate was 62.11%, with a margin of error of 4% for the sample set, with a 95% confidence level (1.96 sigmas) and maximum indeterminate P=Q=50%. Subsequently, stratification was weighted to fine-tune the weights of the interviewees with the population data from the final study universe. The weight coefficient reference data were calculated and using the variables “Nielsen area”, “municipality size”, “gender” and “age” from the last wave of the Spanish General Media Study (EGM).\n\n\nResults\n\nThe results were processed using IBM SPSS Statistics 24®. We first performed a descriptive statistical analysis of the survey’s Likert scale variables, including calculating means and standard deviations. In what follows, we highlight key data relating to the respondents’ autonomously self-reported skills.\n\nRegarding the purposes that they use different digital technology (Table 1), the results show that the majority of young people (87.5%) use mobile for entertainment, (59.7%) use computer for information and (58.7%) for work, (46.2%) of them use video game console for entertainment but 50% do not use it at all. More than half of them (59.2%) don’t use tablets, and smart watches (66%).\n\nWith regard to the purposes of using digital platforms, apps or social media, (Table 2), the results show that the majority of young people (86.7%) use WhatsApp for communication, YouTube (89.7%), Instagram (86.5%), and TikTok (75.2%) for entertainment, while the majority of them don’t use Telegram (60.2%), LinkedIn (80.7), Facebook (66.2%), Snapchat (64.2%), Twitter (46.2%), Twitch (59.7%), and Discord (56%).\n\nRegarding time, 64.3% of the young people claimed that they spent more time as an internet user since the start of the COVID-19 coronavirus pandemic. In what follows, the respondents specified the tendency of their time spent on different activities using the internet during spring 2020 home lockdown. They also asked to compare the quality of each activity (Table 3). The results demonstrate that the young people spent more or the same amount of time on all the mentioned activities except participating in cultural activities in which (53.8%) of them claimed spending less time during lock down. In respect of quality, the majority of the activities were declared to be of the same quality or better. According to the respondents, activities such as communication with friends, listening to music and watching series had better quality during lockdown. However, they mentioned less quality for studying which required remote school activities during lockdown.\n\nWith regard to technical skills (Table 4), the results show that 73.4% of young people claim to know how to install/uninstall basic programs and applications without help. Most young people report knowing how to browse the internet and use related services for everyday purposes, with 80.7% saying they can do this without help. Fewer young people appear to use content management platforms to produce multimedia publications, with only 26.6% claiming to know how to do this without help. Meanwhile, 58.4% of the respondents say they know how to record, edit and upload video content to the internet without help. As for sharing and distributing digital multimedia content, 69.7% of the young people surveyed say they can do this without help. A total of 66.6% of young people say they know how to work with others using digital collaboration tools without help. Only 27.8% of young people say they know how to set up digital services and use tools to increase online privacy and anonymity without needing help. In terms of knowing how to read and/or write computer code, only 19.2% say they can do this without help. Similarly, few young people claim to know how to repair and/or service devices without help (25.5%).\n\nIn terms of informational skills (Table 5), 49.4% of young people say they know how to check the reliability and truthfulness of information without help. Among young people, 54% say they know how to classify and filter information to suit their interests without help. A total of 69.9% of young people 69.9% say they are able to find and save information for use when they need it. With respect to social informational skills, 64.4% of the respondents say they know how to display self-control when interacting with others on social media and digital forums so as not to react impulsively. Regarding spotting so-called “trolls” in online discussions, 58.5% of the young people surveyed claim to have this skill, while 47.3% report knowing how to tell when they are interacting with a bot. According to this sample, 68.8% of young people are able to manage the various profiles that make up their digital identity. Meanwhile, 68.2% say they know how to adapt their behavior according to the standards of each platform. Among the young people surveyed, 56.8% report being able to identify their needs and find tools and platforms to fulfill them without help. Less than half of the young people in our sample (45.5%) say they are able to take part in online deliberation and decision-making processes; 17.6% say they know how to do this and could teach others, while 27.9% say they simply know how to do this alone.\n\nIn terms of critical knowledge (Table 6), 22.9% of the respondents say they know a lot or a fair amount about the basic features of digital services. Of the young people surveyed, 33.6% say they know a lot or a fair amount about how technology companies use personal data. Meanwhile, 21.7% of young people say they know a lot or a fair amount about laws dealing with issues related to digital technologies. Only 22.4% say they know a lot or a fair amount about the influence of technology companies on public policy. Finally, 31.9% of young people say they know how the technological devices they use are manufactured.\n\nWith regard to ICT use in formal and informal learning, the results show that the first three topics for which the young people use the internet to find information are education 60.5%, digital technology 51.2%, and job-related information 48.9%. Regarding the use of ICT to acquire or improve skills, the first four fields reported by young people are video games 54.6%, sport 51.6%, cooking 43.2% and fashion & beauty 42,4%. In respect of self-directed learning, they have been asked to choose the first three things to find information. The results show that first watching online videos (62.7%), then browsing online resources such as books and article 57.7%), and finally asking a family member, or a friend (54.8%).\n\nIn reference to specific training in digital technologies in secondary school or university 34.6% said that they attended talks about uses (security, cyberbullying, etc.), 26.6% attended specific training sessions at their school or university, while 28.4% did not receive such training. With respect to class participation rating, the majority, 29.6% of young people, claim a fairly engagement during in person classes while the majority of them, 31.4% said they engage somehow during online learning. Regarding the procedures used in online education by schools, 64.4% of respondents reported that the most used initiative that conducts online learning activities is Emailing study materials with supporting tasks and instructions. On the subject of use and assessment of different online teaching initiatives, 92.7% voted for interactive exercises, 90% Online forms (no assessment) and 89.6% for online games. Then they have been asked to assess their experience in online classes. 39.6% of the young people claim their disagreement to the statement “my teachers stimulate my interest during online classes” while 38.6% of them neither agreed nor disagreed. Similar results have been seen about the second statement “my teachers come well prepared and organized for each online class”, 36.8% disagreed and 33.4% assessed it as average.\n\nOn the subject of use and assessment of different online teaching initiatives, 92.7% voted for interactive exercises, 90% online forms (no assessment) and 89.6% for online games. Then they were asked to assess their experience in online classes. 39.6% of the young people disagree with the statement “my teachers stimulate my interest during online classes’, while 38.6% of them neither agreed nor disagreed. Similar results have been seen regarding the second statement, “my teachers come well prepared and organized for each online class”, 36.8% disagreed and 33.4% assessed it as average. The answer to their preference towards synchronous online classes (all participants connected at once) rather than asynchronous (via videos, material or educational resources previously provided by the teacher) were similar three grades of agreement. Finally, the majority of the young people 62.7% evaluated that the outcome of education during the pandemic (in online format) was worse than before.\n\nIn relation to perception of young people about digital technologies, the ranking according to how relevant they think digital technologies are for young people was as following: 26.8% rated first the creating a collective identity (e.g., forming groups to share likes, interests or concerns), participating in activities proposed by institutions 21.9%, achieving common goals (e.g., organizing events or activities) 21.2%, making claims and carrying out actions in-person and non-virtual environments 15.3%, and co-designing the activities in which they participate 14.8%.\n\nWith respect to the impact of use of digital technologies on young people (Table 7), the following concepts have been most as ranked neither positive nor negative: psychological well-being (42.7%), Communication with adults (42.3%), individual identity (37%), ability to organize themselves into groups (37.8%), Acceptance of established social norms (42.2%), and decision-making and social autonomy (37.9%). However, the concepts of ability to argue and discuss (39%), socialization among equals (39.7%), and the ability to express themselves as individuals (34.8%), have been most ranked positively.", "appendix": "Data availability\n\nFigshare: A data set about digital literacy competencies among youngsters (16-18) in Spain https://doi.org/10.6084/m9.figshare.21379104.v3 (Mohammadi et al., 2022)\n\nThis project contains the following underlying data:\n\n• Dataset.sav\n\n• Questionnaire.pdf\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nAranda D, Sánchez-Navarro J, Mohammadi L: Perception self-assessment of digital skills and gaming among youth: A dataset from Spain. Data Brief. 2020; 28: 104957. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDonohue JM, Miller E: COVID-19 and school closures. JAMA. 2020; 324(9): 845–847. Publisher Full Text\n\nFernández-de-Castro P, Aranda D, Moyano S, et al.: Digital youth work: A systematic review with a proposal. Soc. Work. Educ. 2021; 1–19. Publisher Full Text\n\nLivingstone S, Mascheroni G, Stoilova M: The outcomes of gaining digital skills for young people’s lives and wellbeing: A systematic evidence review. New Media Soc. 2021; 146144482110431. Publisher Full Text\n\nMohammadi L, Aranda D, Montaña Blasco M, et al.:A data set about digital literacy competencies among youngsters (16-18) in Spain. Dataset. figshare. 2022. Publisher Full Text\n\nRatten V, Jones P: Covid-19 and entrepreneurship education: Implications for advancing research and practice. Int. J. Manag. Educ. 2021; 19(1): 100432. Publisher Full Text\n\nTheben A, Juárez DA, Lopez IP, et al.: Participació i ciutadania activa dels joves a través d’Internet i les xarxes socials. BiD: textos universitaris de biblioteconomia i documentació. 2021; (46). Publisher Full Text\n\nVan Deursen AJAM, Helsper EJ, Eynon R: Development and validation of the Internet Skills Scale (ISS). Inf. Commun. Soc. 2016; 19(6): 804–823. Publisher Full Text" }	[ { "id": "159561", "date": "27 Jan 2023", "name": "Mònica Figueras-Maz", "expertise": [], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe paper focuses on the exploitation of an interesting big sample and a very relevant target; however, it needs a deep revision to go deeper into the analysis and make evident the contribution of the study, moreover present the data. In addition, more detail is needed in the data and ethic isues.\nGeneral comments:\nIn the abstract, the authors use two different concepts: the level of digital literacy and the perceptions of the use of different digital technologies; those are two different approaches. What are the authors exactly measuring?\n\nThe reviewers miss a more solid introduction and a theoretical framework, we suggest reviewing Ferrés and Mateus's studies.\n\nWe also find that the paper lacks conclusions and discussions that go over the exposure of data; it would be interesting to know what is the researchers' position, contribution, and possible recommendations.\n\nA gender perspective focus could be important, for example, comparing uses and competencies by gender. In addition, other variables could be compared as age, and kind of school (private, public, etc.).\n\nSample:\nImportant to describe how researchers guarantee the pluralism of the sample (different socio-economic areas, etc.). Especially regarding access to technology.\n\nIn general, we find more information is needed on sampling and how it is distributed by independent variables. How was the questionnaire distributed, etc.\n\nInclude variables of schools ownership (public, private or semi-private schools).\nEthics: It would be interesting to get more details about the consent form obtention procedure, as well as general ethics issues. For example, it is said that “those aged 16 to 18 answered the children themselves but with the consent of the parents, which were present while they answered as the legislation indicates”. How could the authors be sure of this statement?\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and materials provided to allow replication by others? Partly\n\nAre the datasets clearly presented in a useable and accessible format? Partly", "responses": [ { "c_id": "9421", "date": "03 Mar 2023", "name": "Daniel Aranda", "role": "Author Response", "response": "We appreciate the review, the time spent and the comments. Reviewers: The paper focuses on the exploitation of an interesting big sample and a very relevant target; however, it needs a deep revision to go deeper into the analysis and make evident the contribution of the study, moreover present the data. In addition, more detail is needed in the data and ethic isues. Response: As stated in the guidelines a Data Notes are brief descriptions of scientific datasets that promote the potential reuse of research data and include details of why and how the data were created; they do not include any analyses or conclusions. More details on data and also Ethic issues are answered below. Reviewers: General comments: In the abstract, the authors use two different concepts: the level of digital literacy and the perceptions of the use of different digital technologies; those are two different approaches. What are the authors exactly measuring? Response: In the abstract, we specifically refer to the fact that the survey reflects on the one hand the competences of young people and on the other hand the perceptions of the use of different digital media or digital technology. These two aspects are effectively different as the reviewers point out and each aspect (Perceptions of use and competencies) has distinct spaces in the questionnaire. The perception of the use of digital technologies has been studied in several academic disciplines. According to Davis (1989), Technology Acceptance Theory (TAM) is a theoretical model that explains people’s adoption and use of technology. TAM maintains that perception of the use of a technology is influenced by perception of utility and ease of use. The more useful and user-friendly a technology is, the more likely it is that a person will adopt and use it regularly. Davis, F. D. (1989). Perceived usefulness, perceived ease of use, and user acceptance of information technology. MIS quarterly, 13(3), 319-340. On the other hand, Digital literacy refers to the ability to use digital technologies, such as computers, smartphones, and the internet, to find, evaluate, create, and communicate information effectively. It encompasses a range of skills, including the ability to use digital tools to access and analyze information, collaborate with others, and solve problems. Van Deursen and van Dijk (2015) found that digital skills were positively associated with social and economic outcomes, such as employment status, income, and civic engagement. The study also found that individuals with lower levels of digital skills were more likely to experience social and economic disadvantages. Van Deursen, A. J. A. M., & Van Dijk, J. A. G. M. (2015). The digital divides shifts to differences in usage. New Media & Society, 17(7), 1-18. Thus, the second section of the survey is about digital competence: technical skills (nine items), informational skills (ten items), and critical digital knowledge (five items), respectively. The fourth section is about youth perception of digital technologies, asking the young people to rank five social ability items considering the relevance of digital technologies for them. Reviewers: The reviewers miss a more solid introduction and a theoretical framework, we suggest reviewing Ferrés and Mateus's studies. Response: There is no theoretical framework in the data note because it is not required as stated in the guidelines. What reviewers call the theoretical framework makes reference to methodology, a detailed account of the protocol used to generate the dataset. Reviewers: We also find that the paper lacks conclusions and discussions that go over the exposure of data; it would be interesting to know what is the researchers' position, contribution, and possible recommendations. Response: Data Notes are brief descriptions of scientific datasets that promote the potential reuse of research data. As stated before, this data notes do not include any analyses or conclusions. Reviewers: A gender perspective focus could be important, for example, comparing uses and competencies by gender. In addition, other variables could be compared as age, and kind of school (private, public, etc.). Response: Although data note do not include any analyses or conclusions, in relation with gender, readers can see Estanyol, E., Montaña, M., Fernández-de-Castro, P., Aranda, D., & Mohammadi, L. (2023). Digital competence among young people in Spain: A gender divide analysis. [Competencias digitales de la juventud en España: Un análisis de la brecha de género]. Comunicar, 74, 113-123. https://doi.org/10.3916/C74-2023-09 In this paper we use the notion of digital citizenship, in order to study the gender digital divide as it relates to competence (i.e., skills and knowledge) and the possibility of leveraging said competence to promote civic education grounded in gender equality in the digital environment. Reviewers: Sample: Important to describe how researchers guarantee the pluralism of the sample (different socio-economic areas, etc.). Especially regarding access to technology. In general, we find more information is needed on sampling and how it is distributed by independent variables. How was the questionnaire distributed, etc. Response: As stated in the guidelines of Data Notes: for standard protocols that have been published elsewhere, a brief description and reference is sufficient. The detail of the sampling has been published previously in the article \"Digital competence among young people in Spain: A gender divide analysis\" in the Comunicar Journal. Estanyol, E., Montaña, M., Fernández-de-Castro, P., Aranda, D., & Mohammadi, L. (2023). Digital competence among young people in Spain: A gender divide analysis. [Competencias digitales de la juventud en España: Un análisis de la brecha de género]. Comunicar, 74, 113-123. https://doi.org/10.3916/C74-2023-09 The readers could also have all this information in the section Data availability: Figshare: A data set about digital literacy competencies among youngsters (16-18) in Spain https://doi.org/10.6084/m9.figshare.21379104.v3 (Mohammadi et al., 2022). Reviewers: Include variables of schools ownership (public, private or semi-private schools). Response: Following the international tradition, we have chosen not to ask for the public or private variable in the survey but as stated by the reviewers this variable could have been interesting to include. Here are some references. de Lenne, O., Vandenbosch, L., Eggermont, S., Karsay, K., & Trekels, J. (2020). Picture-perfect lives on social media: A cross-national study on the role of media ideals in adolescent well-being. Media psychology, 23(1), 52-78. Jarman, H. K., Marques, M. D., McLean, S. A., Slater, A., & Paxton, S. J. (2021). Social media, body satisfaction and well-being among adolescents: A mediation model of appearance-ideal internalization and comparison. Body Image, 36, 139-148. Noon, E. J., Schuck, L. A., Guțu, S. M., Şahin, B., Vujović, B., & Aydın, Z. (2021). To compare, or not to compare? Age moderates the relationship between social comparisons on instagram and identity processes during adolescence and emerging adulthood. Journal of Adolescence, 93, 134-145. Reviewers: Ethics: It would be interesting to get more details about the consent form obtention procedure, as well as general ethics issues. For example, it is said that “those aged 16 to 18 answered the children themselves but with the consent of the parents, which were present while they answered as the legislation indicates”. How could the authors be sure of this statement? Response: The panel was contracted to the company ODEC. This company has worked with (Dynata) a software that complies with all legal issues in relation to data protection and response protocols. Those under 16 years of age answered with their parents by their side and those aged 16 to 18 answered by themselves with the consent of their parents present while answering as required by law. ODEC offers digital solutions specialized in market research, statistics, media and marketing." } ] }, { "id": "207955", "date": "20 Sep 2023", "name": "Pritika Reddy", "expertise": [], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe Abstract can improve. Include more details and keep within the word limit for the abstract.\n\nThe Introduction is very short and doesn’t explain the need for the study. There needs to be description about important aspects on the topic plus the contribution of the study.\n\nLiterature Review on digital literacy and media literacy is missing. You can refer to the following papers: Reddy et al. (20221), Reddy et al. (20212), Reddy et al. (20193) and Reddy et al. (20234).\n\nResults and Discussion sections look good.\n\nConclusion and Limitations of the study are missing.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and materials provided to allow replication by others? Partly\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [ { "c_id": "10442", "date": "28 Nov 2023", "name": "Daniel Aranda", "role": "Author Response", "response": "Estemeed reviewer, First of all, thank you very much for taking the time to read and comment our work. 1. The Abstract can improve. Include more details and keep within the word limit for the abstract. Following your advice, we have extended the abstract using the remaining space available without surpassing the word limit. We have developed the background trying to precise the rationale for this dataset and we have rewritten the method part in a more systematic way adding more details of the procedure. 2. The Introduction is very short and doesn’t explain the need for the study. There needs to be description about important aspects on the topic plus the contribution of the study. In this section, taking into account your comment, we added a paragraph in order to provide a better context of the research and make more explicit the reasons that fostered our approach. In particular, we reference the work of Middaugh & Kane (2013) New media as a tool for civic learning, Comunicar, Vol.20, No. 40, pp. 99–107. https://doi.org/10.3916/C40-2013-02-10 given that for us is one of the most notable works in our line of research, that is, the civic potential of digital media specifically for young people. 3. Literature Review on digital literacy and media literacy is missing. You can refer to the following papers: Reddy et al. (2022), Reddy et al. (2021), Reddy et al. (2019) and Reddy et al. (2023). In this point, we decided not to extend the text in terms of a literature review given that, following the guidelines of the F1000 Research journal for Data Notes, this kind of paper does not need that kind of development, although we agree on the need of a literature review in the case of a regular paper. Anyway, we really appreciate the references provided because they are very insightful in relation to our line of research and we will make use of them in following works. 4. Results and Discussion sections look good. We are thankful for the positive feedback regarding these sections. 5. Conclusion and Limitations of the study are missing. In the same sense that we discuss in point 3, the guidelines of the F1000 Research journal for Data Notes does not indicate the need for a Conclusion an Limitations section. We invite you to check one article derived from this dataset in which we expose that kind of reflections: Estanyol, E., Montaña, M., Fernández-de-Castro, P., Aranda, D., & Mohammadi, L. (2023). Digital competence among young people in Spain: A gender divide analysis. [Competencias digitales de la juventud en España: Un análisis de la brecha de género]. Comunicar, 74, 113-123. https://doi.org/10.3916/C74-2023-09 Thank you very much again and we are in touch for further communications. Greetings." } ] } ]	1	https://f1000research.com/articles/12-26
https://f1000research.com/articles/12-700/v1	19 Jun 23	{ "type": "Case Report", "title": "Case Report: an unusual orbital tumor", "authors": [ "Anis Mahmoud", "Hager Touil", "Fadima Hann", "Riadh Messaoud", "Hager Touil", "Fadima Hann", "Riadh Messaoud" ], "abstract": "Introduction: Orbital lipoma is an extremely rare tumor, representing less than 1% of all orbital tumors. We review the literature and describe the presentation, the differential diagnosis and the management of this tumor. Case report: We report the case of a 63-year-old patient who was referred for a diplopia with recent hemi-cranial headache. Physical examination showed no exophthalmos nor decrease in visual acuity. The patient complained of diplopia on elevation and oculomotricity examination showed limited elevation of the right eye. The Hess Lancaster test was in favor of a limited course of the right inferior rectus muscle. Magnetic resonance imaging revealed a fusiform tissue process in the right inferior rectus muscle with a fatty signal. A complete excision of the tumor was performed by a trasncunjonctival approach. Cytopathological examination was consistent with a pleomorphic lipoma. The postoperative period was uneventful. The definitive histopathologic diagnosis was a lipoma. The postoperative Magnetic resonance imaging showed the complete disappearance of the lesion. With 3 years of follow up, there is no sign of recurrence or ocular motility trouble. Conclusion: Lipomas are rare tumors in the orbit. The clinic is variable depending on the size and the site. The clinical diagnosis is difficult to make. Only histology allows the final diagnosis.", "keywords": [ "Lipoma", "Orbit", "Histopathology", "MRI", "Surgery." ], "content": "Introduction\n\nLipomas are benign tumors, rare in the orbit, representing less than 1% of all orbital tumors. They pose a differential diagnosis with a variety of other expansive orbital masses.1 We report a new case, review the literature and discuss the clinicopathological and radiological features, the differential diagnosis and the management of this entity.\n\n\nCase report\n\nA 63-year-old unemployed Tunisian woman, with no previous personal or family pathological history, presented with a diplopia evolving for two weeks. Physical examination showed no exophthalmia and no decrease in visual acuity. Furthermore, it revealed diplopia on elevation. Oculomotricity examination showed limited elevation of the right eye, which was confirmed by the Hess Lancaster test that revealed a limited course of the right inferior rectus muscle. Magnetic resonance imaging (MRI) showed a fusiform and hyper-vascularized tissue process located in the right inferior rectus with fatty signal. The tumor was hyperintense on spin-echo T2-weighted images (Figure 1) and hypointense on spin-echo T1-weighted images (Figure 2).\n\nThese findings suggested various diagnosis; lipoma, inflammatory process, lymphoma and malignant tumor.\n\nWe performed a right inferior transconjunctival orbitotomy and excisional biopsy under general anesthesia. Peroperatively, we discovered an encapsulated mass of fatty tissue, thus complete excision was made. No adherences or involvement of adjacent structures occurred. The specimen was well circumscribed and slightly firmer than normal adipose tissue, with a yellow surface (Figure 3).\n\nHistologic examination was consistent with a pleomorphic lipoma. The postoperative period was uneventful. Immediately after the operation, the patient reports the resolution of his diplopia. Postoperative MRI images demonstrated the complete resolution of the tumor (Figure 4). With 3 years of follow up, there is no sign of recurrence or ocular motility impairment.\n\n\nDiscussion\n\nOrbital lipoma is the most common mesenchymal soft tissue tumor. However, it is rarely found in the orbit despite the presence of abundant adipose tissue in the intraorbital space.2,3 A review of the largest series of orbital tumors revealed a very low incidence of lipomas.4 Shields et al. reported only two cases of lipomas in a review of 1264 cases of orbital tumors, indicating the rarity of this entity.5 On physical examination, the diagnosis is often difficult to suggest. These tumors are often asymptomatic.\n\nHowever, they can cause severe morbidity by causing progressive and painless exophthalmos, which is occasionally coupled with diplopia or ocular motility defects6 such as was observed in our patient.\n\nOrbital lipoma exceptionally leads to a compressive neuropathy responsible for a significant decrease in visual acuity, an alteration of the afferent photomotor reflex and the visual field constriction.1 Imaging based on computed tomography (CT) scanning and MRI is essentially useful in ascertaining determining the exact seat, size and relationship to the orbit content. The fatty signal is characteristic on CT sequences. Furthermore, as was found in our patient, the tumor is hypointense on spin-echo T1-weighted images and hyperintense on spin-echo T2-weighted images.1\n\nHistology is essential for definitive diagnosis of pleomorphic lipoma. An important histologic criterion is the presence of a mixture of fat cells, pleomorphic cells and in particular floret-like multinucleated giant cells embedded in a myxoid stroma.7 That concorded with the histological result of our case. Differential diagnosis of this tumor became more important because the number of reports about some other tumors of similar morphology, are increasing. Pleomorphic lipoma may be confused with lipomatous hemangiopericytoma, myofibroblastoma or even malignant tumors such as rhabdomyosarcoma, myxoid malignant fibrous histiocytoma and liposarcoma.8,9 Surgical excision of an orbital lipoma is not only recommended for symptomatic cases such as our patient's clinical presentation but also to exclude malignancy.10 In addition, as was noted in our patient, the long-term outcome after surgery is considered excellent.11\n\nThis case highlights the importance of orbital imaging in the context of diplopia without obvious cause to rule out an intraorbital lipoma. Nevertheless, this association remains rare and requires further documentation of cases.\n\n\nConclusion\n\nLipomas are benign soft tissue tumors, rarely located in the orbit. The clinical presentation is variable depends on the size and the intraorbital site. The histology makes the definitive diagnosis and may precisely identify the variant.\n\n\nConsent\n\nWritten informed consent for publication of their clinical details and/or clinical images was obtained from the patient.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\n\nReferences\n\nChi MJ, Roh JH, Lee JH, et al.: A case of orbital lipoma with exophtlmos and visual disturbance. Jpn. J. Ophtalmol. 2009; 53(4): 442–444. Publisher Full Text\n\nCivit T, Klein O, Freppel S, et al.: Tumeurs orbitaires d’origine mésenchymateuse. Neurochirurgie. 2010; 56: 158–164. PubMed Abstract \| Publisher Full Text\n\nTripathy D, Mittal R: Spindle cell lipoma of the orbit. Ophthal. Plast. Reconstr. Surg. 2015; 31: e53–e55. PubMed Abstract \| Publisher Full Text\n\nKim MH, Sa HS, Woo K, et al.: Fibrolipoma of the orbit. Ophthal. Plast. Reconstr. Surg. 2011; 27: e16–e18. PubMed Abstract \| Publisher Full Text\n\nShields JA, Shields CL, Scartozzi R: Survey of 1264 patients with orbital tumors and simulating lesions: The 2002 montgomery lecture, part 1. Ophthalmology. 2004; 111: 997–1008. PubMed Abstract \| Publisher Full Text\n\nToledano FN, Stoica BT, Genol SI, et al.: Diplopia from pleomorphic lipoma of the orbit with lateral rectus muscle involvement. Ophthalmic Plast. Reconstr. Surg. 2013; 29(2): e53–e55. Publisher Full Text\n\nBartley GB, Yeatts RP, Garrity JA, et al.: Spindle cell lipoma of the orbit. Am. J. Ophtalmol. 1985; 100: 605–609. Publisher Full Text\n\nUlivieri S, Oliveri G, Motolese PA, et al.: Spindle cell lipoma of the orbit: acase report of an unusual orbital pathology. Neurol. Neurochir. Pol. 2010; 44: 419–423. PubMed Abstract \| Publisher Full Text\n\nDigregorio F, Barr RJ, Fretzin DF, et al.: Case reports and review of the literature. J. Dermatol. Surg. Oncol. 1992; 18(3): 197–203. PubMed Abstract \| Publisher Full Text\n\nReibel JF, Greene WM: Liposarcoma arising in the pharynx nine years after fibrolipoma excision. Otolaryngol. Head Neck Surg. 1995; 112(4): 599–602.\n\nShah NB, Chang WY, White VA, et al.: Orbital lipoma: 2 cases and review of literature. Ophthal. Plast. Reconstr. Surg. 2007; 23(3): 202–205. PubMed Abstract \| Publisher Full Text" }	[ { "id": "190823", "date": "04 Aug 2023", "name": "Mehdi Hasnaoui", "expertise": [ "Reviewer Expertise Oncology", "sinus and nasal cavity surgery", "facial bone imaging" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nEditorial Note from F1000Research – 14 August 2023: A COI statement has been added detailing a shared affiliation between the authors and reviewer, which was not declared at the time of the publishing of this report. The COI statement is below.\n---\nThis article is well written. It is easy to read. The figures are demonstrative. The authors report a rare case of an orbital lipoma revealed by isolated diplopia. The MRI was not very suggestive of a lipoma. The authors insisted on the interest of MRI in the context of diplopia. They discussed the clinicopathologic and radiological characteristics and the differential diagnosis of this entity.\nI have some minor comments:\nAbstract :\nThe authors state: “We report the case of a 63-year-old patient who was referred for a diplopia with recent hemi-cranial headache”. This symptom “hemi-cranial headache” was not mentioned and written in the case report section.\n\nThe authors state: “Cytopathological examination was consistent with a pleomorphic lipoma. The postoperative period was uneventful. The definitive histopathologic diagnosis was a lipoma. ” Did the authors do a cytopathological examination and then a definitive histological examination? This examination “cytopatholgical” was not mentioned and written in the case report section.\n\nLine 13 : change \"trasncunjonctival\" to \"transcunjonctival\"\nCase report :\n\nThe authors state: “MRI showed a spindle-shaped and hyper-vascularized tissue process”. How to explain this hypervascularization?\n\nDo the authors have postgadolinium fat-saturated axial T1 images? Lipomas are classically hypointense after fat suppression.\nDiscussion :\n\nIn this case, the tumor was hyperintense on spin-echo T1-weighted images. This is not usual in lipomas and should be discussed.\n\nThe authors state: “Furthermore, as was found in our patient, the tumor is hypointense on spin-echo T1-weighted images and hyperintense on spin-echo T2-weighted images.1 ” However, in the literature and in the cited reference number 1, most lipomas are generally hyperintense on T1-weighted imaging.\nI believe the paper has academic merit, but minor revision is needed.\n\nIs the background of the case’s history and progression described in sufficient detail? Yes\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Yes\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Partly\n\nIs the case presented with sufficient detail to be useful for other practitioners? Yes", "responses": [ { "c_id": "10029", "date": "27 Oct 2023", "name": "Anis Mahmoud", "role": "Author Response", "response": "I would like to thank the reviewer for his interest in this work and will answer his queries in this report: The MRI appearance of pleomorphic lipomas is variable and not pathognomonic for diagnosis, as the variation in the ratio of adipose to non-adipose components results in a wide spectrum of imaging features. MRI features such as hyper-vascularized tissue process and hypointense T1 and hyperintense T2 character can be explained by the presence of a high proportion of non-adipose elements in pleomorphic lipomas. We have added a new reference (7) that also confirms our MRI results, which show the tumor to be hypointense in T1 and hyperintense in T2. The expression \"trasncunjonctival\" has been replaced by \"transconjunctival\". Cytopathological\" has been replaced by \"histopathological\". The symptom \"hemi-cranial headache\" has been added to the \"case report\" section." } ] }, { "id": "193958", "date": "16 Oct 2023", "name": "Amira Trigui", "expertise": [ "Reviewer Expertise Ophthamology" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nA 63-year-old man presented with diplopia for two weeks. MRI of the brain and orbit revealed a process in the orbit with a fatty signal. Excisional biopsy of the mass resolved the diplopia, and histology revealed a pleomorphic lipoma. This report may help clinicians in the future to be alert to the possibility of the presence of an orbital tumor despite the absence of exophthalmos, which is a common finding in this setting. In this case, the clinical observation is detailed and the MRI images are of excellent quality, but a few points need to be clarified:\nDid the patient's general examination reveal obesity, or other lipoma locations?\n\nDid the authors perform orbital ultrasound before MRI imaging?\n\nAuthors should specify whether diplopia is monocular or binocular.\n\nIt would also be interesting if the authors could explain the absence of exophthalmos in this case.\n\nIs the background of the case’s history and progression described in sufficient detail? Yes\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Partly\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Yes\n\nIs the case presented with sufficient detail to be useful for other practitioners? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-700
https://f1000research.com/articles/11-251/v1	01 Mar 22	{ "type": "Review", "title": "Strategies of enhancing rural livelihoods and promoting sustainable use and conservation of indigenous chicken breeds in Zambia", "authors": [ "Christopher .M Kanyama", "Amy .F Moss", "Tamsyn .M Crowley", "Amy .F Moss", "Tamsyn .M Crowley" ], "abstract": "This review explores innovative and sustainable strategies for conservation and use of village or indigenous chickens (IC) (Gallus domesticus) in Zambia and parts of Sub-Saharan Africa (SSA). Small scale farmers (SSF) have kept IC for hundreds of years to meet their households’ nutritional needs, incomes, social-cultural and religious uses among others. The commitment exhibited by SSF in keeping IC has made them the major custodians of essential animal genetic resources in low-income regions. Between 1991 and 2012, private breeders invested over US$95 million in Zambia’s commercial poultry sector resulting in over 100% increase in annual production of day-old chicks to 65 million. However, high production cost and low market access hindered the participation of SSF hence their continued dependence on IC. Unfortunately, the future of IC genetic resources is threatened due to the rapid loss and erosion of IC breeds. In the 2015 biodiversity status report, the Food and Agriculture Organisation, an international body of the United Nation highlighted that over 3.5% of IC breeds were extinct, nearly 33% were at high risk and over 67% were of unknown status. Poultry diseases, lack of sustainable conservation strategies and poor use among others have significantly contributed to these losses. For example, in 2012, 60% of IC were diseased in parts of SSA including Zambia. If these challenges are not mitigated, the loss of IC genetic resources and the adverse impact on rural communities are inevitable. Further, future research and breeding programs on commercial chickens may also be limited as a result of erosion of IC genetic resources. Therefore, this paper reviews and contributes to previous studies that demonstrated how researcher-community-stakeholder engagements potentially enhanced sustainability and the adoption of innovative ideas including the potential to increase conservation and sustainable use of local chicken biodiversity in Zambia and parts of SSA.", "keywords": [ "Animal genetic resource", "biodiversity", "conservation", "rural-community", "small-scale farmer", "poultry-sector" ], "content": "Introduction\n\nThe loss of indigenous chickens (IC) (Gallus domesticus) animal genetic resources (AnGR) and the low socioeconomic gains by the small-scale farmers (SSF) producing IC are the main threats to the livelihoods of rural communities in Sub-Saharan Africa (SSA). The Food and Agriculture Organisation (FAO), an international body of the United Nation reported in their biodiversity status report of 2015 that over 3.5% of IC breeds were extinct, 33% were at high risk and over 67% were in the unknown status category (FAO 2019). In SSA, 80% of SSF keep IC, significantly contributing to the indigenous poultry sector (IPS), making them the major custodians of IC-AnGR in the region. However, despite this important role they play, SSF have not realised significant socioeconomic gains from the IPS. Challenges such as poultry diseases, poor nutrition, and low access to markets are in part the cause of the current status of the IPS in Zambia and parts of SSA. Therefore, the main objective of researcher-community-stakeholder (RCS) engagements and potential innovation targeting SSF is to promote sustainable utilisation and conservation of IC-AnGR, through identifying major challenges and opportunities in the IPS and designing adoptable interventions.\n\nMany countries in SSA experience varying and distinct agro-ecological conditions. The region’s diverse climate affects individual countries in a variety of ways. In this scenario, Zambia is not exceptional. With a total surface area of 752,618 square kilometres (75.3 million hectares), Zambia is divided into three different agro-ecological regions (AER; I, II and III) each with unique agricultural challenges, concerning annual rainfall (ARF), vegetation, annual temperatures, soil type and water resources. The AER I and II occupy 54% of the total national land, mainly in the Southern, Western, Central and Eastern areas of Zambia (Phiri and Mukelabai 2010). These two regions receive between 800 mm and 1000 mm ARF, whereas region III, primarily in the Northern and North-western, classified as high rainfall zone, covers 46% of the total national area and receives more than 1000 mm ARF. Despite this variability in climate experienced across the country, and the fact that over 40% of fresh groundwater in Southern Africa is in Zambia, 90% of SSF practise rain-fed agriculture (Hamududu and Ngoma 2019).\n\nGlobally, most problems experienced in agriculture are highly associated with climate variations, which may worsen by the next century. Some studies predict that by the end of the 21st century, there will be a 3 °C increase in global temperature, 0.6% reduction in ARF, and 13% reduction in available groundwater due to climate change (Pelletier and Tyedmers 2010; Hamududu and Ngoma 2019). The drastic climate variation will have a more adverse effect in low-income countries, especially in SSA. Therefore, small livestock, such as goats, sheep and poultry, generally considered low input enterprises, are essential, and will contribute substantially to improving livelihoods among rural communities (Simainga et al. 2011; Queenan et al. 2016).\n\nGuèye (1998, 2000) highlighted that IC, which comprises the majority of rural farming poultry (RFP) in SSA, have been kept by SSF for generations to meet their food security, household incomes, poverty reduction and empowerment of women. Some researchers have also documented how small livestock could help farmers become more adapted and resilient to climate variations than they would with larger livestock species. In fact, large animals, such as cattle, demand more grazing land and water resources (Yayneshet and Treydte 2015). This paper highlights the role of agriculture, indigenous poultry, challenges faced and what feasible strategies to mitigate the impact of the loss of IC-AnGR on rural communities in SSA. Throughout this paper, the term indigenous poultry will refer to indigenous chickens (IC) (Gallus domesticus) or village chickens excluding other domestic avian species.Further, all prices for poultry inputs and products are in the United states of America dollar (US$).\n\n\nAgriculture and indigenous chickens\n\nAccording to FAO, chickens are globally classified in the top five crucial animal species, with the other four being cattle, sheep, goats and pigs (FAO 2019). Therefore, FAO has made it mandatory for countries to prioritise the submission of biodiversity status reports for the animal species highlighted above. Among the poultry species, IC have the highest population and importance in SSA, because most rural farmers produce these chickens at low land, capital and labour requirements. Although rural farmers practise low input production systems, there are variations in conditions and environments across the region and within countries based on their socioeconomic status (Guèye 1998, 2000). Studies thus far have revealed the socioeconomic function of IC for rural communities in low-income countries (Dolberg 2007; FAO 2019).\n\nDespite researchers sharing a consensus on the socioeconomic role of IC in SSA, there are fewer attempts to holistically find solutions to challenges faced in the IPS including the continued loss of IC-AnGR and low socioeconomic gains by SSF (Dolberg 2007). The Poultry Association of Zambia (PAZ) expressed similar concerns regarding obscure solutions for SSF in Zambia's poultry sector. Generally, the problems faced by IPS are also associated with unsustainable use of IC-AnGR, lack of skills in animal management and disease control, absence of value addition among others (Guèye 2000; Mueller et al. 2015; PAZ 2021). Therefore, interventions that promote the sustainable development of the IPS need to be initiated to secure IC breeds and enhance rural livelihoods in Zambia and parts of SSA.\n\nStudies have demonstrated that researcher-community-stakeholder (RCS) engagements could significantly improve the management of indigenous livestock, increase incomes and reduce poverty in low-income countries (Mueller et al. 2015). In their analysis of various community-based breeding programs (CBBP), Mueller et al. (2015) showed a positive impact of RCS engagements on AnGR and livelihood for SSF. The involvement of rural communities in CBBP empowered SSF through decision making and livestock business ownership. Most importantly, researchers found that CBBP was a sustainable option for conserving local AnGR through sustainable use and continuous improvements among SSF. There are fewer reliable long-term approaches suitable for rural communities, particularly in low-income countries (Mueller et al. 2015). However, to promote conservation of IC-AnGR and enhance the socioeconomic gains for SSF involved in IPS, developing an innovation based on the local context is essential. The community-based intervention aimed at developing the IPS through guidelines and principles of RFP by Guèye (2000), conservation of AnGR by FAO (2019), and the CBBP promoted by Mueller et al. (2015), would potentially help in identifying challenges and designing sustainable solutions within the Zambian local context.\n\nTherefore, a collective approach based on shared interests among researchers, communities and stakeholders would create sustainable and adoptable strategies through a community-based indigenous poultry development program (CBIPDP). In the section below, details of the roles of agriculture and IC, constraints in the IPS and the developmental stages of CBIPDP through RCS engagements in Zambia and parts of SSA are covered.\n\n\nUse of agriculture and indigenous chickens among rural communities\n\nAgriculture has to address four main issues, namely increased cost of living, population growth, poverty and inequality. Most studies have demonstrated that agriculture provides employment, food and nutritional security, livelihood assets, and gender equality among rural communities, potentially countering the highlighted concerns (Guèye 2000; Dolberg, 2007; Boland et al. 2013). Globally, agriculture contributes 40% of gross domestic product (GDP) and provides employment to over 1.3 billion inhabitants (White 2012; Boland et al. 2013). A majority of SSF in low-income countries consider agriculture as a full-time occupation. For many generations, rural farmers in SSA have grown various crops and kept livestock, including IC for their livelihoods (Guèye 1998, 2000). Therefore, as we head towards the year 2050, agriculture and livestock production are crucial for job creation and meeting the food and nutritional demands in SSA (Klingholz 2020).\n\nThere is a consensus among researchers that IC and other indigenous livestock play significant socioeconomic and socio-cultural-religious roles among rural communities particularly in SSA (Lebbie and Ramsay 1999; Guèye 2000; Aklilu et al. 2007; Duguma, 2009; Queenan et al. 2016; Alders and Pym 2019). Small scale farmers keep over 80% of IC in the region (Guèye 1998, 2000; Queenan et al. 2016). Further, Guèye (1998, 2000) highlighted the value and relevance of IPS in SSA as far back as 1994. During this period, SSA had over 1.1 billion IC, producing over 1.7 million metric tonnes (MT) of eggs and 2.1 million MT of chicken meat compared to only 16 million ducks, producing over 26 thousand MT of duck meat and seven million turkeys yielding over 54 thousand MT of turkey meat. Further, analysis of surveys conducted in SSA showed that in 1994, each rural household kept up to 20 chickens, equivalent to three chickens for every two people in the rural human population (Guèye 1998). Changes in IC population in comparison to national poultry population for the period 1989 to 2018 for selected countries in SSA have been reported by FAO stat (2021) and Guèye (1998). For example, in Kenya, Malawi, Nigeria, Tanzania, Zimbabwe and Zambia the population for IC comprised nearly 80% of the national flock.\n\nIn the past three decades, the numbers and uses of IC have increased minimally compared to commercial poultry among rural communities. The difference in the growth rate between the two poultry subsectors is highly associated with the production costs and shortages of feeds experienced in SSA, with SSF affected significantly (Guèye 2000; PAZ 2021). The case of Zimbabwe is different where a 67% contraction in the poultry industry was observed between 2007 and 2018 (FAO Stat 2021). This may be associated with political and economic problems the country passed through during the 2000 - 2001 land reforms implemented by Zimbabwean government.\n\nIn Zambia, the socioeconomic role of agriculture is equally evident, especially among rural communities. According to the Ministry of Fisheries and Livestock (MFL), nearly 18% of Zambia’s GDP is from agriculture, supporting over 12 million people and absorbing over 67% of the labour force (MFL 2017). Poultry is a significant component of agricultural GDP in Zambia. A study conducted in 2012 found that over one million smallholder poultry farmers out of 1,418,000 agricultural households surveyed raised over 12 million IC in Zambia (Lubungu and Mofya 2012). The study also found that half of the reported number of IC were owned by SSF in Eastern, Southern and Central provinces whereas the other half was distributed among the seven regions. The MFL estimated a 20% annual growth in IPS between 2012 and 2017 (MFL 2019).\n\n\nUses of indigenous chickens in Sub-Saharan Africa\n\nIn SSA, IC are highly valued among rural communities. The rural farmers use IC for household food source, income, breeding and incubating eggs, socio-cultural and religious functions among others (Guèye 1998, 2000; Moreki et al. 2010). The culture and socioeconomic status of the rural communities influence the diverse uses of IC observed across the region and within countries. However, the most common uses for IC are consumption, household income and social-cultural and religious roles.\n\nRural communities in SSA have kept IC for the animal-based protein requirement by consuming chicken meat and eggs. The contribution to food needs for rural households from poultry is much more significant compared to other types of livestock. For example, Guèye (1998, 2000) demonstrates how a simple farmer in Tanzania starts with one pullet and makes massive gains of close to 170 chickens - cocks, hens, pullets and cockerels, 440 to 1,100 eggs and 47 kg of chicken meat in five years, significantly improving the farmer’s food security and general livelihood. In addition, past production trends in the IPS of selected countries in SSA also demonstrated how valuable IC were as food source. For example, between 1984 and 1989, this sector produced 23,117 MT eggs (12%) in Nigeria, 3,172 MT eggs (26%) and 14,835 MT meat (69%) in Ivory Coast, 396 million eggs (36%) and 29,952 MT meat (26%) in Morocco, and over 46 million eggs (71%) and 8611 MT meat (72%) in Kenya (Guèye 1998, 2000). The production in the IPS is expected to have grown exponentially to date.\n\nThe dynamics of consumption and use of IC in SSA are strongly associated with culture, socio-economic status and likely so by human population growth three decades from now (Guèye 1998, 2000; Klingholz 2020). For example, in Tanzania, the SSF consumed less than a half of what they produced, and sold the rest to rural areas, whereas in Zambia, the SSF consumed more than half of IC and only sold 20% to urban areas (Queenan et al. 2016).\n\nGenerally, IC have low input requirements depending on the socio-economic status of the rural farmers. However, IC are considered an easy way out of poverty because poor communities are more likely to own and farm them. These quick and easy socio-economic gains explain why these chickens are an entry point to wealth because anyone could start keeping them regardless of their socio-economic status (Guèye 1998, 2000; Dolberg, 2007). Indigenous poultry contributes to household incomes and assets that help improve livelihoods for SSF. Rural communities raise much-needed revenue for their household from chickens' sales to meet their daily household needs, including possible future investments. Guèye (1998, 2000) explained how SSF allocated incomes to various household needs. For example, when family A raises US$180 from selling 30 chickens, they would spend it as follows: US$72 for daily household needs, US$54 for buying clothes, invest $36 in business, and use US$18 for purchasing replacement stock. The meticulous allocation of incomes from IC highlights their value to rural livelihoods. Farmers also use IC as a medium of exchange in form of barter system. For example, in the Gambia, SSF exchanged five full-grown hens with an adult sheep and 25 hens with one adult cow (Guèye 1998, 2000). This trend also illustrated how owning IC was as good as owning any other livelihood assets among SSF.\n\nIndigenous chickens are gender and socio-economic equaliser supporting women and children in most parts of SSA (Kitalyi 1998; Guèye 2000; Morek et al. 2010; Simainga et al. 2011; Queenan et al. 2016). In SSA, the socio-economic value of indigenous poultry in empowering women was evident, as over 70% of IC were owned and managed by women and children in the region, which enabled them possess livelihood assets (Guèye 2000; Dolberg 2007). However, Dolberg (2007) pointed out that the physical asset was missing from the IC’s livelihood equation in that – farmers could not harness animal draft power from chickens. Nevertheless, studies by Guèye (2000) demonstrated that farmers could buy or exchange IC with any assets they desired including cattle as was the case in the Gambia.\n\nSome researchers viewed the subject of women empowerment through IC cautiously. For example, in certain societies in Tanzania, women and children could only manage the chickens, but the powers to decide on the marketing and use were still in men's hands (Queenan et al. 2016). Similar findings were examined in Mozambique where prolonged wars significantly reduced the number of cattle and goats resulting in increased interest and control of IC by men (Guèye 1998, 2000).\n\nSome social-cultural and religious functions of IC among rural communities are usually a combination of incomes, consumption, gifts, medicinal and other uses. In SSA, rural communities sacrificed IC during traditional ceremonies and rituals, share cocks as gifts to their guests at cultural events, use the cocks for traditional medicines including sexual stimulation for men, and also hygiene through scavenging (Guèye 2000). White feathered chickens are vital for traditional medicines and sacrifices in Somalia, Cameroon and Zambia. Attaching value to the colour or appearance of indigenous livestock is also a pricing technique under traditional markets in parts of SSA (Mueller et al. 2015).\n\nThere are other uses of IC observed among SSF in SSA including incubation of eggs, security, ornamental and hobbies. For example, in Ghana, close to 71% of guinea fowls and IC were kept for breeding and most of the eggs incubated. Farmers, strategically selected desired males and females for continued production (Guèye 2000). Similarly, in Ethiopia, eggs were incubated and hatched for continued poultry production. Combined income and consumption use were reported in Zimbabwe, whereas in Nigeria, IC were also used for barter.\n\n\nZambia’s poultry industry and challenges\n\nIn the 1990s, the Zambian government made economic reforms in the agriculture sector through a liberalised market system to promote private sector participation in delivering goods and services (Rakner 2003; Bonaglia 2009). During this period, most government-run entities were sold to private businesses. In 2005, a ten-year plan explicitly for the poultry sub-sector was established (Bagopi et al. 2014). The plan was aimed at increasing efficiency and productivity in the commercial poultry sector. The agriculture reforms and poultry sector plan led to an introduction of new genetics, improved nutrition, health and farming practices in the Zambian poultry industry. Within six years, both integrated and standalone breeders, such as Zambeef-Rainball, Pioneer-Bokomo, Tiger-Ross, Country bird, Panda and Hybrid were established (Bagopi et al. 2014; PAZ 2021). Further, a US$95 million investment by Zambeef-Rainball breeders triggered unprecedented growth in the poultry sector. Between 2007 and 2012, there was an increase in the production of day-old chicks (DOC) from 27 to 65 million per annum with over 50,000 jobs created (Bagopi et al. 2014). Such changes observed in Zambia’s poultry industry occurred much earlier in highly industrialised nations as observed in policies and the gains made by the consumers and the poultry industry in those countries (Steinfeld and Gerber 2010).\n\nThe other outcomes of economic reforms were the increased participation of SSF in the production of commercial chicken breeds, defying their conservative nature of keeping low input IC breeds (Bagopi et al. 2014; Mueller et al. 2015). The short production cycle and increased profits were among the reasons SSF were motivated to take part in the sector. However, because of anti-competition business practices exhibited in the poultry market structure and the rapid increase in production costs, smallholder broiler and layer producers failed to survive (Bagopi et al. 2014).\n\nThe method used to produce chickens has a significant bearing on productivity and quality. The three systems of production practised by farmers are free-range system (FRS), where the chickens scavenge for feed, and no health care interventions used, the semi-intensive system (SIS), in which the chickens are partially allowed to scavenge, coupled with feed supplementation, and finally, the intensive system, in which the chickens are entirely confined and fed throughout their growth period (Guèye 2000; Okeno et al. 2013). Production methods such as FRS and SIS are perceived as beneficial to SSF, mainly due to negligible start-up costs, i.e. feeds and drugs (Guèye 2000; Queenan et al. 2016). Further, under FRS and SIS, chickens can roam and fend for themselves with minimal or no supplementary feeding by the farmer.\n\nHowever, improving practices in areas such as disease control, shelter, marketing, feed supplementing and exploiting available feed resources may increase production efficiency and profitability among SSF in SSA (Goromela et al. 2006). Generally, SSF are at liberty to practice any of the three production systems depending on their constraints and socio-economic status (Guèye 2000). The intensive production system used mainly by commercial broilers and layers producers is unsustainable for SSF because of the high cost of production, disease prevalence, and the highly oligopolistic market controlled by big breeders and producers (Bagopi et al. 2014).\n\nThe cost of feeds and other live-inputs in the Zambian poultry industry generally increased in the past five years. In their weekly reports published for the first quarter for five years from 2016, the PAZ demonstrated that prices for solvent extracted soybean meal increased by 27.7% from US$23.50 per 50 kg, whereas a 45.3% increase for broiler starter from US$20.10 per 50 kg, 44% for broiler grower from US$19.30 per 50 kg, 45.9% increase from US$18.50 per 50 kg of finisher and a 49% change in price for layer mash from US$14.70 per 50 kg in the period between 2016 and 2021 during which the average exchange rate was US$1 to 12.95 Zambian kwacha (ZMW) (PAZ, 2021). Further, PAZ reported price increases for day old chicks for improved free-range chickens, layers and broilers by 87.5%, 83.3% and 125% from US$0.80, US$0.60 and US$0.40 per bird, respectively. Other price changes included pullets, broiler, spent layers and IC increasing by 64.8%, 57%, 38.5% and 61.5% from US$8.90, US$3.00, US$2.60 and US$3.90 per bird, respectively (PAZ, 2021). The high costs of feed and other live inputs are the main reasons why SSF have failed to fully participate in commercial poultry production as these costs constitute the highest proportion of production costs.\n\nAlthough some SSF attempted to produce commercial chickens from 2005 onwards, a majority of SSF continued with IPS by keeping IC because of the low but stable performance under FRS (Bagopi et al. 2014; PAZ 2021). Low costs, easiness of rearing and favourable prices of IC encouraged more SSF to consider IPS as potentially sustainable and profitable (Okeno et al. 2013; PAZ 2021).\n\nFarmers in the IPS rear different breeds of IC. The Ministry of Agriculture and Cooperatives (MACO) highlighted various IC breeds reared by rural communities in Zambia. The most common IC breeds among SSF include, the common Zambi, Naked neck, Dwarfs or short-legged, Frizzled feathered, Feathery legged, and Short-tailed, with live bodyweights (BW) ranging between 1.3 kg and 2.0 kg at over six months of age (MACO, 2003; PAZ, 2021). However, studies in Nigeria and Botswana suggest significant differences in live BW between female chickens (0.7 - 2.1 kg) and males (1.2 - 3.2 kg) (Guèye 2000).\n\nCompared to broilers at six weeks, the growth period for IC is much longer. In the past few years, new breeds have also been introduced in Zambia’s IPS. Some of these improved free-range chicken breeds, include the Boschvelds, Kruoillers, Black Australorps and Brahma. Although the improved free-range breeds are perceived to mature early and very productive under a free-range set-up, the high prices render them inaccessible to SSF. In general, the negligible costs involved in the production of IC promotes the continued farmer involvement in the IPS. A high benefit-cost ratio in IC is common as any selling price is regarded as profit (Simainga et al. 2011; Queenan et al. 2016). However, numerous challenges outlined in the sections below pose as major threats to IPS.\n\n\nThreats to the indigenous poultry sector\n\nMost rural farmers keeping IC are faced with several challenges with the potential to reduce the socio-economic benefits, which may negatively affect their livelihoods. These include, poultry diseases, poor policies, unstable markets and poor infrastructure.\n\nThe decision by SSF to reconsider the rearing of IC is a viable socio-economic strategy. However, poultry diseases and poor nutrition have been significant challenges in traditional poultry farming (Guèye 1998, 2000; Simainga et al. 2011). The argument is that the low input and scavenging production systems are less successful compared to the intensive system for broilers or layers. Uncontrolled poultry diseases and poor nutrition have made IC underperform compared to broilers in Zambia and parts of SSA. Research done in most low-income countries shows that a majority of SSF depend on natural remedies for controlling poultry and other livestock diseases. Guèye (2000) suggested that 79% of farmers used traditional herbs and plants to treat poultry diseases and that over 50% of mortalities happened under IPS by the first four weeks of chicken raising. In other parts of SSA, Queenan et al. (2016) reported that suspected Newcastle disease and fowl pox caused 40% to100% mortalities in IC towards the end of the dry season.\n\nIn Zambia, the common diseases in the scavenging system include Newcastle disease, fowl pox, fowl typhoid, infectious coryza, Gumboro, helminthiases, and ectoparasites, which have contributed to poor performance and high mortalities of IC experienced among SSF (Phiri et al. 2007; Simainga et al. 2011; Mubamba et al. 2018). In 2012, over 27% of SSF relied on traditional medication and less than 15% used veterinary drugs, resulting in 60% of the IC dying (Lubungu and Mofya 2012).\n\nDespite the high disease prevalence observed in IC, there are beneficial adaptability and genetic gains through natural selection. The harsh environments in which indigenous livestock are reared leads to disease resistance and high adaptation to low-quality diets (Mapiye et al. 2008; Gizaw et al. 2010; Queenan et al. 2016). These traits which are more superior in IC than commercial breeds may be essential in future chicken breeding programs (Mapiye et al. 2008). Gizaw et al. (2010) suggest that adaptive traits are equal or more important than production traits in indigenous livestock production systems.\n\nOther nutritional concerns are the poor poultry feeding regimes among SSF. There is limited supplementary feeding and a lower plane of nutrition under the scavenging system, which ultimately leads to mortalities and reduced consistency in chicken (eggs) size and quality (Queenan et al. 2016). The majority of the IC are left to roam and scavenge for feed sources, such as insects, termites, vegetables, seeds, grains and earthworms and in rare situations, farmers supplement the chickens with kitchen waste, maize bran, leafy vegetables and other cheap feed sources (Mwalusanya et al. 2002; Goromela et al. 2006; Mapiye et al. 2008).\n\nThere are also variations in the availability of scavenged feeds depending on the time of year in Zambia. Village chickens have access to high protein insects and earthworms in the rain season (December to April), and high energy feed sources during the harvest of field crops in May to August, whereas, in the hot and dry season, a severe shortage of nutritious feed sources leads to poor health, malnutrition and high mortalities (Queenan et al. 2016). Dry seasons require feed supplementing to mitigate adverse effects on IC. In some parts of SSA, the breeding of black soldiers fly larvae and maggots as sources of protein are being experimented with and may improve nutrition in the IPS, especially in the dry seasons (Kenis et al. 2014).\n\nA majority of SSA countries employ funding and infrastructure development plans favouring larger livestock species, predominantly run by commercial entities and minimal attention is given to IPS, despite SSF being part of the primary data source when designing policies on funding and infrastructure (Dolberg 2007; FAO 2019). These policies exacerbate the problems experienced in IPS and SSF feel that political leaders and scientists do not adequately represent them (Dolberg 2007). For example, recently, a livestock infrastructure support project funded by African Development Bank focused on the construction of dairy and beef infrastructure, whereas the Second National Agriculture policy of 2016 to 2020 did not adequately cover how value addition and market participation for SSF in IPS would be implemented in Zambia (MFL 2017).\n\nPoultry products worth millions of dollars are imported from overseas to SSA, negatively impacting both the commercial and IPS. In 2018, South Africa imported frozen poultry products of mixed parts valued at over US$65 million from different sources in Brazil (PAZ 2021). These products lacked traceability, posed a public health threat and also affected the local poultry sector. In Ghana, over US$60 million worth of poultry products were imported in 2018. These importations prompted the local poultry association to engage the Ghanaian government to introduce quotas on poultry imports in order to protect local poultry farmers. To mitigate the adverse effect of unfair trade, a ban on poultry imports was sanctioned in Namibia whereas, in Zambia, the poultry association engaged the government to promote policies that protect and enhance SSF in IPS (PAZ 2021). Some policy researchers recommend factoring in some livestock production policies and legislation on consumption to existing environmental management policies that could enrich strategies, enhance community benefits and reduce food wastage (Steinfeld and Gerber 2010).\n\nIn SSA, farmers in the IPS face barriers preventing them from accessing a formal market. These obstacles are associated with market standards and requirements, such as selling frozen whole or portions of chickens, packaging, labelling, and selling from standard outlets (Bagopi et al. 2014). Producers in IPS cannot compete with larger commercial entities because they neither own the required facilities nor the brands nor organised sales outlets (Simainga et al. 2011; Bagopi et al. 2014; Queenan et al. 2016). Further, the only way smallholder farmers can sell their chickens or eggs directly to established markets is through groups and intermediaries who manage most market channels (Bagopi et al. 2014). This way is also full of obstacles.\n\nIn Zambia, SSF sell chickens through informal markets, such as the roadside, village markets, backyard, and direct to consumers (Queenan et al. 2016; Mubamba et al. 2018; PAZ 2021). Recently, informal markets have transformed into essential selling points making it possible for farmers to organise and meet consumers’ demands on quality and quantity. For example, the concepts of “Tuesday and Saturday markets” in some parts of the country have become popular. In the Northern part of Zambia, this traditional market is also called Munada, where traders agree on the date and place for the market day.\n\nGenerally, the low access to reliable markets affects the sales and consumption dynamics of IC among SSF. For example, in Tanzania, the SSF sold twice more village chickens and eggs to rural areas than they consumed and sold at US$3.72 per live chicken, whereas in Zambia, producers of IC consumed more than half of their chickens and only sold 20% to urban areas at an average price of US$3.37 per bird (Queenan et al. 2016). The variation in selling points and consumption levels shows how undefined and informal the markets in IPS is in parts of SSA.\n\nThe lack of reliable poultry housing facilities for IC under scavenging systems is another constraint. Some IC are kept in undeveloped poultry houses at night to secure them from predators. However, in many cases, the chickens are left to hide in trees, making them vulnerable to predators, such as cats and dogs (Guèye 2000; Simainga et al. 2011). Consequently, theft, predation and environmental hazards are common, and significantly contribute to losses IC observed in the IPS. For example, a survey conducted in Western Zambia showed that 93% and 84% of the households interviewed attributed the losses to predation and thefts, respectively (Simainga et al. 2011).\n\n\nResearch application\n\nDespite the barriers and challenges experienced by SSF, IC are essential to both rural communities and consumers. In the past few years, the demand and preference for IC have steadily grown among consimers. The increased preference for IC results from the perceived good taste, fine texture, and health benefits of consumers, with some preferring male chickens for the large size and hens for their tenderness (Guèye 2000; Queenan et al. 2016). Although in South Africa, studies by Dyubele et al. (2010) found that consumers prefered broilers to IC due to tenderness and other attributes. Generally, the increased demand has led to a substantial rise in prices of IC over commercial chicken meat, potentially creating an opportunity for SSF (Guèye, 2000; Ajayi 2010; Queenan et al. 2016; PAZ 2021). For example, studies showed that, in Senegal, IC meat was sold at 27% higher than commercial broilers, while in Nigeria, the cocks were sold at prices over 300% more than the hens or broilers (Guèye 2000).\n\nSimilar trends have been observed in Zambia where the IC sell at nearly twice the price of broilers (PAZ, 2021). The Poultry Association of Zambia reported that, in 2016, live IC were sold at US$4.0, which was 33% higher than broiler while in the first quarter of 2021, IC were priced at US$6.40, which was 73% more than broiler chickens. During the period under review, on average, US$1was equivalent to ZMW12.95. This trend is consistent with some studies, where IC prices were higher particularly when sold in formal markets or places familiar to consumers (Guèye 2000; Queenan et al. 2016). Consumers want proof that what they are buying is indeed IC.\n\nPopulation growth and food demand\n\nConsumer demand for healthy products, including IC meat and eggs, will increase with human population growth in SSA. The United Nations, Food and Agriculture Organisation’s revised projections show that from 2005 to 2050, the global human population will grow up to 9.2 and 9.8 billion, of which over 46% of growth will be in SSA, and a 60% increase in food demand is expected (Alexandratos and Bruinsma 2012; FAO 2019). During the same period, Zambia’s population will reach 39 million, contributing to increased food requirements (Klingholz, 2020). Annual population growth of 1.9% in SSA with per capita food consumption of less than 2500 (Kcal/person/day) and annual undernutrition levels 20% higher than other regions was predicted for the period between 2005 and 2050 (Alexandratos and Bruinsma 2012).\n\nFood consumption patterns\n\nThe population changes will significantly raise the food demand and consumption pattern of animal-based protein, which accounts for 40% of total protein consumed by humans (Lebbie and Ramsay 1999; Boland et al. 2013; Mueller et al. 2015). Some scholars also predict that the emerging of the middle-class will highly influence an increase in meat consumption in low-income countries, which will require applying technology and innovation to meet the demand during 'The Livestock Revolution' (Steinfeld and Gerber 2010). Globally, from 2000 to 2050, researchers predict an 82% increase in meat consumption, equivalent to 233 - 271 million MT of which 88 million MT is poultry and over 183 million MT from bovine, ovine and pig meat combined (Alexandratos and Bruinsma 2012; Boland et al. 2013). In other regions, meat consumption will increase at a slow rate, although the volumes demanded will be substantial, especially in SSA, where the farming population will get older and a majority will migrate to the urban areas as the middle class expands (Klingholz 2020).\n\nTo mitigate the future challenges on food and nutritional security, governments need to design sustainable agriculture programs, increase investment in research and promote sound policies that encourage the participation of youths in agribusiness and food production (Alexandratos and Bruinsma 2012; Klingholz 2020). In a few decades, SSF working in isolation in SSA may face more constraints as opposed to those working collectively. Farmer belonging to organised groups would improve production and productivity, achievable through good management, sustainable use and conservation of indigenous AnGR, improved disease control, improved nutrition and value addition (Livingston et al. 2011; Mueller et al. 2015; Queenan et al. 2016). Therefore, mobilising SSF into organised groups will significantly promote innovative RCS engagements and result in productivity and sustainability in the IPS in Zambia. The next section outlines examples of effective interventions emanating from RCS engagements in selected low-income countries.\n\nExamples of community-based interventions\n\nThere are practical examples of RCS engagements and their impact on SSF as demonstrated by Rodríguez et al. (2011) and Mueller et al. (2015). These strategies resulted in socio-economic gains among rural communities that adopted various interventions in low-income countries. Mueller et al. (2015) outlined the process required when establishing CBBP and gave examples of such approaches in developing countries that worked and those that faced challenges. These breeding plans had common features, including initiators being either the community or government research institutions, well-formulated breeding objectives based on indigenous or local breeds, the existence of institutional support (technical or financial) and each of the plans recorded a change (Rodríguez et al. 2011; Mueller et al. 2015).\n\nOutcomes of CBBP\n\nThe analysis of various community-based interventions in low-income countries showed that support from research institutions and the government was crucial in achieving the set objectives. Further, the farmers worked together to identify and describe the problems to stakeholders. The impact of community-based interventions in low-income countries were empirically reported. For example, in Vietnam, pork farmers achieved between 40% and 100% increment in pork prices resulting from SSF working together with government institutions in identifying challenges and finding solutions, in Kenya, SSF experienced fast growth in goat population, and over 300% increase in goat milk yields from 0.25 litres per day after community-based solutions, where as in Ethiopia, positive testimonies and sharing of knowledge among SSF led to widespread adoption of sheep breeding strategies among rural communities (Mueller et al. 2015). Similar benefits of RCS interventions were observed in Benin where government and the community engaged in vaccinating IC and broilers against Newcastle disease and using improved poultry management resulted in 58% more profits from IC compared to broilers (Rodríguez et al. 2011). Like many community-based programs, RCS interventions are not spared from challenges. For example, in Bolivia, where the objective of the community-based intervention was improving the fibre quality of wool from llamas, politics, financial mistrust and funding challenges resulted to low stainability (Mueller et al. 2015).\n\nGenerally, there are issues related to community-based interventions. Mueller highlighted trust, financial and technical problems that potentially affected sustainability and continuation of CBBP in some countries. These are important lessons worth considering when designing CBIPDP through RCS engagements for the IPS in Zambia. Promoting the voices and views of the target rural communities and understanding the local context is essential for designing sustainable community-based interventions (Patton 2010; Bryman 2016). For example, promoting poultry development plans based on exotic breeds, instead of indigenous species, is less valuable to SSF due to lack of adaptation new breeds to the local environment.\n\nSome challenges associated with exotic breeds include low literacy, lack of records, diseases and high prices of breeds and costs of production for SSF (Mueller et al. 2015; Mtileni et al. 2016; Sebho 2016). Therefore, considering the local context, IC breeds, and promoting ownership of the intervention through RCS engagements is crucial to the sustainable conservation of IC-AnGR.\n\nSelective adoption of workable approaches based on lessons learnt from IC-AnGR conservation and use would increase the chances of designing a sustainable community-based innovation targeting SSF in the IPS (Guèye 2000; Mueller et al. 2015; FAO 2019). For example, FAO outlined priority areas and interventions to promote the sustainable management and conservation of indigenous AnGR and enhancing livelihoods for rural communities as a global action plan.\n\nThe Food and Agriculture Organisation of the United Nations promoted five strategic areas on use and conservation of indigenous AnGR (FAO 2015, 2019). These are: (a) enhancing knowledge on characterisation of local animals, (b) develop sufficient institutional frameworks for AnGR management, enhanced linkages among livestock farmers, and stakeholders concerning policies and programmes, (c) Enhance awareness through education, training and research in significant areas of AnGR management, (d) Enhancing breeding strategies and programs to harness available genetic resources and match them with environments of production and requirements of societies and (e) Increase and diversification of conservation programs and possibly mix some approaches that use existing livestock breeds in the typical production environment and consider gene banks' use to store genetic materials. To implement the five areas, FAO required individual countries to undertake various programs towards the stated strategies and also expected to submit biodiversity status reports to the FAO Commission on biodiversity.\n\nZambia has implemented programs towards conservation of indigenous AnGR and promoting sustainability among SSF some of which partially fulfilled the five strategic areas of FAO. The programs planned and implemented through research and extension include; on-going farmer engagements and capacity building with donor support on climate-resilient projects, establishing livestock breeding centres across the country, promoting farmer driven innovation such community-based program on indigenous livestock, making available breeding technologies such as artificial insemination and embryo transfer among SSF through the National Artificial Insemination Services Centre in Mazabuka in Southern province of Zambia. Through the African Union InterAfrican Bureau for African Animal Resources (AU-IBAR), Zambia formulated and launched the National Strategic Action Plan in 2018 - 2019 (NSAP) (MFL 2019). Specifically, NSAP was developed to promote sustainable utilisation and conservation of indigenous AnGR in Zambia.\n\nTherefore, a sustainable IPS development plan based on understanding the production systems used by SSF, clearly defined roles of stakeholders and the importance of IC, has the potential to strengthen sustainable conservation of IC and enhance rural livelihoods. The evidence of success of RCS engagements in identifying problems and designing solutions indicates that adapting general practical principles from guidelines from RPF, FAO, and CBBP justifies why CBIPDP is a well-placed project for Zambia’s IPS.\n\n\nConclusion\n\nIn conclusion, agriculture and small livestock including indigenous chickens have the potential to contribute to food and nutritional security, increased household incomes and access to livelihood assets for SSF in Zambia and parts of SSA, particularly towards the year 2050. To address concerns of the loss of IC-AnGR and the low socio-economic gains reported among SSF involved in IPS, CBIPDP is a workable research innovation strategy suitable for low-input agriculture production systems, common in Zambia and many parts of SSA. The CBIPDP would explore the integration of resources and skills through RCS engagements when formulating strategies to mitigate challenges in the IPS. Interventions such as improving farmer skills in poultry management, production, value addition and disease control may significantly contribute to SSF access to formal markets and enhance livelihoods for rural communities in Zambia.\n\n\nRecommendations\n\nFuture studies should investigate market needs and consumption patterns for indigenous chickens among consumers in Zambia. Further, an assessment of the impact of the coronavirus disease 2019 (COVID-19) pandemic on the IPS is essential in understanding the resilience and sustainability of rural communities.\n\n\nData availability\n\nNo data are associated with this article.", "appendix": "Acknowledgements\n\nThe authors acknowledge the Deputy Vice-Chancellor of the University of New England in Australia, the Ministry of Fisheries and Livestock and the poultry industry in Zambia for supporting this work. Further, the authors extend the acknowledgement to both the Australian and Zambian governments for supporting research in agriculture, environmental and rural science.\n\n\nReferences\n\nAklilu HA, Almekinders CJ, Udo HM, et al.: Village poultry consumption and marketing in relation to gender, religious festivals and market access. Trop. Anim. Health Prod. 2007; 39(3): 165–177. PubMed Abstract \| Publisher Full Text\n\nAjayi F: Nigerian indigenous chicken: A valuable genetic resource for meat and egg production. Asian J. Poult. Sci. 2010; 4(4): 164–172. Publisher Full Text\n\nAlexandratos N, Bruinsma J: World agriculture towards 2030/2050: 2012 revised.2012.\n\nAlders RG, Pym RAE: Village poultry: still important to millions eight thousand years after domestication. Worlds Poult. Sci. J. 2019; 65(2): 181–190. 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Publisher Full Text" }	[ { "id": "129217", "date": "08 Apr 2022", "name": "Titus Jairus Zindove", "expertise": [ "Reviewer Expertise Sustainable livestock production" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe manuscript entitled “Strategies of enhancing rural livelihoods and promoting sustainable use and conservation of indigenous chicken breeds in Zambia” discusses the importance of and ways to improve indigenous chicken production in Zambia. I think the effort is useful and provides a perspective on the role of indigenous chickens on resource-poor households in Zambia. I am, however, concerned about the organization, readability, and flow of ideas in the manuscript. The manuscript is difficult to comprehend and needs to be edited carefully. I encourage the authors to improve the manuscript after paying attention to the comments below:\nI suggest “Strategies of promoting sustainable use and conservation of indigenous chicken breeds in Zambia” as the title.\n\nGenerally, although it is important to make the review of broad interest, the authors should avoid digressing. The topic and objective suggest that the review is on indigenous chicken production in Zambia, but the manuscript ended up covering the whole of Sub-Saharan Africa and even other countries such as Ghana, Senegal, Kenya, Gambia, Morocco, Nigeria, Tanzania etc. While it is important to compare indigenous chickens production in Zambia to other developing regions, it is more important not to lose focus.\n\nThe authors need to find a logical structure for the review. There is no clear flow of ideas. For example, the introduction starts with a discussion/background on chicken production and then, in paragraph 3, the focus shifts to goats, sheep and poultry in general. The authors then go back to indigenous chickens in the last paragraph. Initial sections of the review were on indigenous chickens but broadened towards all livestock or, maybe, all indigenous livestock. Authors should find a logical way to order and link the sentences, paragraphs and sections of the review.\n\nThe document has too many acronyms. This makes reading and understanding the manuscript difficult. Acronyms such as RPF, IC, RCS, IPS, SSF, AER, ARF, FRS and SIS can be avoided.\n\nThroughout the document, some statements/facts are not backed by relevant citations.\n\nParagraph 1, line 8 – “…IPS in Zambia and parts of SSA”. Is Zambia not part of SSA? Authors should also justify why the review focuses on Zambia in the same paragraph\n\nParagraph 1, line 9 - “Therefore, the main objective of …” should be changed to “Therefore, one of the main objectives of …”\n\nThe section entitled “Agriculture and indigenous chickens” focuses on chickens. Less, if anything, is on other agricultural practices. The title should be revised.\n\nPage 4, Paragraph 2 - Avoid starting a paragraph with the word “therefore”. Paragraphing in the manuscript needs to be improved. A paragraph should start with a “topic sentence” which introduces the idea to be discussed in the paragraph and the rest of the sentences in the paragraph should be linked to or be an expansion of the topic sentence.\n\nPage 4, paragraph 3 - What is the difference between this section and the preceding section?\n\nPage 4, paragraph 3, line 8 - Is livestock production not part of agriculture?\n\nPage 4, paragraphs 5 and 6 - Does this fit into the sub-topic “Use of agriculture and indigenous chickens among rural communities”? I also think the last three paragraphs on this page are more like repetition, or can fit well in previous sections.\n\nPage 5, paragraph 1 - What is the difference between this section and the proceeding one?\n\nSome of the references used on the production statistics are too old. Surely production statistics reported in 1989 and 1984 would have changed by now? Authors should use mostly young literature throughout the document. Some of the sections, e.g. paragraph 4, lines 7- 9, do not add value to the review and should be deleted.\n\nThe last paragraph on page 6 can be summarised into one sentence.\n\nPage 7, paragraph 2 - The subtopic is incomplete. In Zambia? SSA? How does the last sentence fit into the paragraph? Any link with the preceding sentence?\n\nPage 7, paragraph 7 - The sentence “…. there are beneficial adaptability and genetic gains…” contradicts sentences in the preceding paragraph. If the chickens are adapted, then why is the mortality rate high?\n\nPage 7, paragraph 8 - Isn’t this repetition? The preceding section also includes nutritional challenges (at least according to the section heading). Southern Africa is rich in supplements for indigenous chickens. Communal farmers grow crops such as sorghum, finger millet sunflower and millet which can be used to feed chickens. Some of these crops are drought resilient. Is nutrition really a big problem for indigenous chickens under communal systems in Sub Saharan Africa? Maybe when it comes to commercialization of the indigenous chickens.\n\nPage 8\nRevise subtopic “Low policy for….”\nSome of the prices quoted were reported 5 years ago? Are the prices still the same now?\nAuthors may also comment on the issue of market linkages\n\nPage 9 - What is the link between the section on “research applications” and preceding linkages? The subtopic is also not clear. Does it reflect on the contents of the section?\n\nIs the topic of the review discussed comprehensively in the context of the current literature? Partly\n\nAre all factual statements correct and adequately supported by citations? Partly\n\nIs the review written in accessible language? Yes\n\nAre the conclusions drawn appropriate in the context of the current research literature? Partly", "responses": [ { "c_id": "8077", "date": "11 Apr 2022", "name": "Christopher Kanyama", "role": "Author Response", "response": "RESPONSE TO REVIEWERS REPORT This is the response to the reviewers report and guidance provided by Zindove T.J (2022) with reference to our review article entitled: Strategies of enhancing rural livelihoods and promoting sustainable use and conservation of indigenous chickens in Zambia [version 1]. On behalf of my fellow authors, I acknowledge receipt of the report and appreciate the valuable guidance provided by the reviewer. We shall definitely consider the suggestions and guidance given with the aim of improving the quality of content to the required scientific standard. Looking forward to future engagements. Kanyama C.M Corresponding author (09/04/2022)" } ] } ]	1	https://f1000research.com/articles/11-251
https://f1000research.com/articles/11-499/v1	05 May 22	{ "type": "Research Article", "title": "A qualitative study of sexual violence and attribution of responsibility in Georgian youth", "authors": [ "Natalia Mchedlishvili", "Nino Zhghenti", "Nino Zhghenti" ], "abstract": "Background: Sexual violence is a complex and pressing social issue that needs urgent solutions. Republic of Georgia is one of those countries where despite some advancements in law and practice, patriarchal beliefs and behaviour patterns still prevail in a daily life. So far, there has not been undertaken an in-depth study on how Georgians and particularly youth, define sexual violence and what are the justifications behind these views. The aim of the research is to understand how Georgian students understand sexual violence, who they think are responsible for sexual violence and what are the underlying reasons behind those views. Method: In total, 37 in-depth interviews have been conducted with Georgian students, from September of 2019 to March of 2021. The study participants were recruited from different universities, including the two biggest cities of Georgia - Tbilisi and Batumi.\n\nResults: The research revealed that understanding of sexual violence is far more complex than it was expected. Georgian students define sexual violence as a broad category where sexual harassment and sexual coercion have overlapping and at the same time independent meaning. Interpretation and attribution of responsibility in all three categories are dependent on situations and context that contain not only physical violence but also inappropriate touch, insistent gaze, comments about body parts, sexist insults and discrimination, messages of sexual content, psychological pressure and blackmailing. Attribution of responsibility was equally dependent on personal judgements as well as culturally determined stereotypes. Conclusion: Sexual violence is not a new phenomenon in Georgia, but its consideration as a social problem is. The research demonstrated that understanding and judgment on sexual violence, sexual harassment and sexual coercion is nuanced issue and still needs clear categories of definitions.", "keywords": [ "sexual violence", "sexual coercion", "sexual harassment", "responsibility" ], "content": "Introduction\n\nA growing body of literature is focusing on the subject of sexual violence; being a complex and pressing social issue that needs urgent solutions. Due to its severity, sexual violence definition has been broadened over decades and it is defined as any sexual act, attempt to obtain a sexual act, unwanted sexual comments or advances, or acts to traffic, or otherwise directed, against a person’s sexuality using coercion, by any person regardless of their relationship to the victim, in any setting, including but not limited to home and work (Krug et al., 2002). Among the consequences of any form of sexual victimization are substance abuse disorders, eating disorders, depression, and suicide attempts (Campbell, Dworkin, & Cabral, 2009; Jordan, 2014; Messman-Moore et al., 2000). Despite its comprehensive definition and its fatal consequences; views on sexual violence, is still debatable in different societies on family and institutional level. This ambiguity depends on culture (Heise et al., 1996) Georgia, which has gone through many changes on gender roles in private and public domain, still holds patriarchal views (UN WOMEN, 2017). These views, eventually, form interpretation frames and behavioural patterns of sexual interactions in a family, social environment, legislative bodies and public institutions. Even among young Georgians who are expected to have more modern beliefs than older generation, one can see an influence of patriarchal culture in appraisal of gender issues (Japaridze et al., 2014). Gender attitudes are translated into corresponding judgments of sexual violence which in many cases favour male perpetrators and not victims (UN WOMEN, 2017). The national study on violence against women in Georgia conducted in 2017, revealed that 26% of women have experienced sexual violence and/or sexual harassment at some point in their lives. The study showed that sexual violence perpetrators are rarely held accountable for their actions. The findings also suggest that the majority of Georgian women and men believe that violence against women in relationships should be tolerated and accepted in most of the times. Although Georgian younger generation have more awareness on sexual violence and are expected to have less violence-prone attitudes, 16% of young women (aged 15-24) think that violence in a family should be tolerated to keep family together (UN WOMEN, 2017).\n\nNotwithstanding evidence and daily practice of sexual violence in Georgia, there has not been undertaken an in-depth study on how Georgians and particularly youth, define sexual violence and what are the justifications behind those views. Therefore, the aim of the research is to understand how Georgian students understand sexual violence, who they think are responsible for sexual violence and what are the underlying reasons behind those views.\n\nAttribution of responsibility is a central component in understanding how sexual violence is understood and practiced in a society and in this case, in youth. Academic studies have indicated that rape victims, despite being victims of a crime, are often blamed and denigrated, even to the extent whereby the victim is held responsible for an assault (Calhoun et al., 1976; Donnerstein & Berkowitz, 1981; Janoff-Bulman, et al., 1985). Observers’ judgments and actions are also subject to bias as a result of their personality disposition rooted in given cultural context that has an effect on attribution making (Maddux & Yuki, 2006). Patriarchal culture results in ready schemes of judgement where male perpetrators either are not blamed, completely for violence or their blame is shared with victims. A number of studies on factors influencing attributions of rape victims revealed that more male students than female students believe that victims are responsible for rape situations (Bell, Kuriloff & Lotte, 1994; McCaul, Veltrum, Boyechko, Crawford, 1990).\n\nIn the United States many studies were conducted about sexual violence issues in youth. The main reason for these studies was that sexual assaults on campus has become a major concern for many colleges and universities in the United States, for example one out of every four women experience some form of sexual victimization while in college (Cantor et al., 2015). Also, sexual victimization can affect future victimization (Messman-Moore & Long, 2003; Messman-Moore et al., 2000; Noll, Horowitz, Bonanno, Trickett, & Putnam, 2003), and increase the chance of perpetration of violence later in life (McMahon et al., 2015), with victims often showing a propensity to become abusive. In addition, there is evidence that sexual violence can significantly impair one’s cognitive abilities (Hall et al., 2014), reducing students’ opportunities for success during and after college (Jordan, 2014). Research has consistently demonstrated over the past four decades that college women are at high risk of rape and attempted rape while in college (Fisher et al., 2000; Karjane et al., 2005; Koss et al., 1987).\n\nThe given study not only contributes to existing literature on understanding sexual violence in Georgian youth but also is one of the first and essential steps designing effective programs for preventing and controlling sexual violence in Georgia.\n\n\nMethod\n\nEthical approval for the research has been granted by the dissertation committee of Free University of Tbilisi, Georgia. Before the interviews, research participants were informed on the purpose of the research and their right to participate or refuse participation. Additionally, all the participants were also assured of anonymity of their identity and confidentiality of their responses. Written informed consent was obtained from the participants. All data were saved in a secure file, with only the researchers in charge of the data collection and analysis accessing the information.\n\nIn-depth exploration and understanding of beliefs, views and attitudes is a complex process requiring flexibility, rich description, constant tracking of new insights and rigorous analysis on every stage of data collection and analysis. Therefore, qualitative research approach was used in revealing how sexual violence is defined and understood, who are responsible for sexual violence and what are the underlying reasons behind those views.\n\nThe qualitative study was conducted from September of 2019 to March of 2021. To ensure rich and in-depth interpretations and discussions on sexual violence and all the parties and factors involved in it, it was planned to conduct in-depth interviews with selected respondents. Considering the complexity of study topic, instead of asking open-ended direct questions, respondents were given real stories of sexual violence incidences from Georgian reality. Then, respondents were asked to share and explain in detail their views and attitudes on given stories. Real stories of sexual violence were compiled from semi-structured interviews with specialists working on sexual violence cases (in total: 15 respondents, including lawyers, doctors, psychologists and social workers). There are not many organisations and specialist who works on sexual violence cases in Georgia. A snowball sampling method was used for recruiting specialists. Also, sexual violence cases were compiled from Georgian digital media (Tabula, Liberali, Netgazeti) articles and TV programs (Rustavi 2). Google www.google.com was used as database and key words were sexual violence, sexual harassment, sexual coercion and rape (all key words were in Georgian). Sexual violence case study was done from September 2019 to December 2019.\n\nResearch and analysis was carried out by two researchers Natalia Mchedlishvili and Nino Zhghenti. Both researchers were involved in process of research design, development and implementation. Natalia has conducted all the interviews and then thorough analysis of the data was done by Natalia and Nino as well.\n\nIn the first phase, a pilot interview was carried out. 8 interviews were conducted with students of Free University of Tbilisi and Agricultural University of Georgia (Tbilisi). After the pilot, the interview guide took final shape and interviews were continued (29 interviewees were added to pilot interviews) with other students from same universities, also, from Batumi State University, from Ilia State University (Tbilisi), from Tbilisi State University and Caucasus University (Tbilisi). The age of participants were from 18 to 25. The research participants were on different year of study from first year of Bachelor’s degree to last year of Master’s degree. The total number of participants were 37: 19 male and 18 female. The participants were from different faculties: Social Sciences, Business Administration, International Relations, Mathematics and Computer Science, Natural Sciences, Law, Humanitarian Sciences etc. The participants were selected by purposeful random sampling on the basis of the universities. The adequacy of the sampling size was determined on the basis of the saturation of collected data. The participants were invited to the interview by email (obtained from the database of the universities, about 300 emails). Most of invited participants agreed to interview. Some of the interviews took place at the universities in a separate room, face to face and one to one with interviewer and some were conducted via Zoom program (www.zoom.us), due to coronavirus lockdowns in Georgia, from January 2020 to March 2021. All the interviews were conducted one to one and were audiotaped for further data analysis. Interviews lasted on average 1-1.5 hours and were in Georgian.\n\nThe pilot interviews were conducted in order to test if sexual violence stories were well-structured and if the content of the story was clear and neutrally formulated. The later was important to avoid altering participants views by biased story telling. After pilot interviews, stories were revised and designed in three categories: 1) sexual harassment stories; 2) sexual coercion stories; 3) sexual violence stories (the most severe cases). The guide contained the three story categories and five major open-ended questions which implied regular prompting, based on respondents’ answers: 1) What do you think about this situation in the story? 2) What do you think, how this situation in this story can be interpreted? Is it sexual harassment, sexual coercion or sexual violence? (These questions were asked chronologically and were based on respondents’ answers) 3) Why do you think this story is sexual harassment (sexual coercion or sexual violence)? 4) Would something change if the participants were different sex or gender? 5) In this story, who is responsible for sexual violence?\n\nBased on audio files detailed transcripts of each interview was prepared. Thematic analysis method was used for data analysis. This method, by using inductive approach, allowed close examination of respondents’ views and attitudes. Through a systematic and structured reading, codes themes were generated using qualitative data analysis software - NVIVO program. Data analysis is structured according to three research questions. Within each question, respondents’ narratives are analysed in context of three theme categories of sexual violence: sexual harassment, sexual coercion, sexual violence.\n\n\nResults\n\nThe interviews and media analysis revealed 12 typical stories that happened in Georgia for last 5 years. As a result an in-depth interview guide was developed where identified real stories were listed for the respondents to reflect on. These selected stories were on the following topics: sexual harassment of a woman/man at the office; sexual harassment of a woman by a priest; sexual harassment of a student (a young woman) by a lecturer (male); dissemination of a video of a woman having sexual contact with ex-partner (man); stalking of a woman by a woman/man; sexual coercion of the wife by the husband; sexual coercion of a man by an ex-partner (man); sexual violence of a student (young woman) who was drunk and was not conscious by male students at students’ party; the deceptive involvement of a young woman in sexual trafficking; rape of a young woman for forced marriage by a young man; rape of a young woman at night in the street by a stranger.\n\nOverall, data revealed sexual violence as a broad category implying many reasons and causes. While, in some cases, it has been understood as a physical violence, in other cases it was viewed as an umbrella term for all forms of sexual violence, among which sexual harassment and sexual coercion where mentioned. Hence, data analysis was structured according to these three categories.\n\nMost1 of participants consider sexual harassment as unpleasant flirting, comments with sexual content, inappropriate touch, insistent gaze, comments about body parts, sexist insults and discrimination, messages of sexual content. These are one time or recurrent actions that usually act as a hinting for one’s sexual intentions to other person. This leads to victim’s disruptions in social routines and functioning, especially when it happens at office. The later is a social context where, sometimes hidden abusive organizational culture is a major determinant for violent relationships. According to respondents, when candidates for employment are initially selected based on appearance, eventually harasser makes attempts of unwanted and unpleasant touching or/and sexually intimidating remarks. Either intention is never realized or at some point, it culminates in a violent or forced sexual act, “sexual violence”. Hence, it is believed that those situations take place virtually or when two parties are alone in a room or/and it is impossible or hard to escape. For example, locking a door by a harasser or creating any physical barrier to prevent victim from leaving. One of the participants shared his view: “Sexual harassment is a complicated term. It is difficult to define. I think, in case of sexual harassment you have no space for action. You are trapped in an abuser’s hands. If you are free in space and you are able to escape or ask for immediate help, it is not sexual harassment.” (male student, 21 years old). It was also mentioned that, sometimes, without any physical obstacles, victim’s emotions of shame and embarrassment are sufficient for her/him not asking for available help. As respondents explained, this emotions are usually culturally rooted where perpetrators are well aware of it and victim’s feel lonely and have low or no trust in the social environment for understanding and support.\n\nSexual coercion\n\nAccording to research participants sexual coercion is more indirect form of violence where one person puts another person in a situation where she/he cannot refuse sexual interaction or any other action with sexual content and in the most situations, the abuser do not need to use physical force. For example, one of the respondent believes that: “Sexual coercion is when a person badly uses his power over another person. For example, an abuser can say that if you do not agree for sexual intercourse, I can use this power and you will lose your job” (male student, 22 years old). “Power”, in this case, is defined as a possession of some compromising material or any other information that puts the victim in an unfavourable position in a family or public eye. On the other hand, it is also defined as power hierarchy where the most widespread cases are represented in employer – employee relationships. Here, while one needs to keep his/her job, the other takes advantage of this situation. In both cases, one with information or with higher position in a hierarchy, through blackmailing, makes the other person to do sexual activities that may take place once or multiple times. Respondents also believed that it is sexual coercion when an abuser, without blackmailing, can use a victim’s psychological condition to force them into sexual intercourse. These are problematic situations where two persons start interaction with different intentions and end up with sexual intercourse where one of them could not find sufficient inner strength to resist the flow of events.\n\nUnlike sexual harassment, where marking point was verbal expression of “unwanted”, “inappropriate” and “unpleasant” sexual content; sexual coercion was understood as, one party puts the other party in a situation where it is hard to resist the imposed sexual activities. This characteristic of the phenomenon is not observed objectively (e.g. demonstration of physical force) but it is accessible through exploring and understanding subjective feelings and emotions of a victim. Respondents further explained that sexual coercion is more like a psychological violence and intimidation. One of the respondents expressed her view about the case of sexual coercion of a man by his ex-partner (a man):“This case is sexual coercion because the abuser badly used psychological pressure on the victim and forced him to have sex” (female, 20 years old). Victims, without physical or/and, sometimes even verbal resistance, are involved in sexual interaction. Therefore, this type of violence is harder to identify and prove.\n\nSexual violence\n\nMost participants consider sexual violence to be using physical force for sexual intercourse, unwanted touching, repetitive unwanted sexual comments leading to forced sexual act, coercive sexual intercourse and spreading someone else’s sexual photos and/or videos. Most participants think that sexual violence is quite a broad term and it sometimes includes sexual harassment and sexual coercion, as well: “Sexual violence is a much broader notion and it implies sexual harassment and sexual coercion too” (female student, 19 years old). One of the participants said: \"I perceive violence more as a physical act that is committed against the will, and given behaviour and its outcome is visible” (male student, 24 years old). Another participant believed that: \"It can also be just a hand patting or something like that…” (female student, 23 years old). In the end of the interview, one of the participants summarised: “Sexual violence, probably implies all those interactions where one party is involved against his/her will.” (female student, 20 years old).\n\nBased on students’ narratives, it was obvious that sexual violence was interpreted as forced physical behaviour on another person and also was perceived as an umbrella term for different forms of sexual violence, including sexual harassment and sexual coercion: “It is important to know differences among sexual harassment, sexual coercion and sexual violence. I think, when you act against your will, it is coercion. Sexual violence is when someone uses physical force …” (male student, 22 years old). Consequently, causes and reasons for perceiving a situation as sexual violence was overlapping with causes and reasons expressed in cases of sexual harassment and sexual coercion. This was due to consideration that sexual violence was more a general category for various forms of sexual violence. The research participants considered the situations in given stories as sexual violence, because there was clear physical assault where one party used physical force: \"At first, the fact that he used physical force for having sex is already sexual violence, because the other person did not want this” (male student, 21 years old). Also, one party had not consented to sexual intercourse and clearly expressed that they did not want sex and the second party forced them into it, for example in case of forced marriage one of the participants expressed her view: “This case is sexual violence, because the abuser was using psychological pressure and after this he used physical force too” (female student, 20 years old). Also, one party used blackmail and forced the second party into sex: “One of the partners clearly expressed her will. In this case the will of the victim is clearly visible. The woman expressed a clear will and everything happened against it. This means sexual violence” (male student, 24 years old). Most participants indicated that the case should be considered as sexual violence, because the victim could not give consent, she was drunk and unconscious\n\nAttribution of responsibility was done in favour of a perpetrator and in some cases, in favour of victims too. The decision who to blame, was contingent upon contextual and situational details. Especially, in situations, where there was not a demonstration of physical violence and/or clear and categorical refusal by victim, students believed that responsibility for the outcome should be shared. One of the participants expressed her view about situation where lecturer was sending nude photos to student via messenger: \"I do not know the exact messages between them, but because this relationship was bilateral (between student and lecturer), the responsibility cannot be attributed to one party” (female student, 21 years old). Also, when the violence situations were repetitive and victim “could influence” on a particular situation the research participants distributed more responsibility to a victim, for example in regards to the sexual harassment and sexual coercion of someone at a workplace. One of the participants expressed his view about the case of forced marriage: “The responsibility is distributed to victim too, because she saw that the man acted violently against her and she should have told someone about this, she should not be so afraid. I could not see that this girl tried to solve this situation, she even did not ask for help from her family members” (male student, 23 years old).\n\nStudents also attributed less responsibility to victims when the abuser was a stranger and the victim could not somehow “control” the situation. One of the research participants said: “In some cases there were solutions for the situation, but here I cannot see how the victim could influence the situation and avoid sexual violence” (female student, 20 years old). In this research, no differences were found between female and male participants’ answers. This was true for all three main categories and for all discussed cases.\n\n\nDiscussion\n\nThe goal of the present study was to answer the main research questions: a) How Georgian students understand sexual violence? b) Who they think are responsible for sexual violence? c) What are the underlying reasons behind those views?\n\nThe findings provide support for the definitions of sexual harassment and sexual coercion as categories of sexual violence. The latter is believed to be a broader term for different forms of sexual violence, and, at the same time, it is perceived to build on physical force (e.g. rape). Sexual harassment and sexual coercion, as are indicated in our study and in the supporting literature, being component part of sexual violence practice, they also have independent meaning.\n\nHejase (2015) defines sexual harassment as “unwelcome or unwanted sexual advances, requests for sexual favours, and any form of verbal, non-verbal or physical conduct of a sexual nature that interferes with one’s employment or work performance or effect of violating the dignity of a person, in particular when creating an intimidating, hostile, degrading, humiliating or offensive environment”. Our research revealed that, in comparison to other forms of sexual violence, sexual harassment can be limited to verbal expression as well as unwanted touching. Sexual coercion is described as the use of any tactic or strategy to engage another person in sexual behaviours without consent (Abbey et al., 2014; Farris et al., 2008). These coercive strategies may involve the use of manipulation (through promises or inducing guilt for example), persistent touching, intoxication or verbal pressure or physical force. Sexual coercion, therefore, implies behaviours legally defined as sexual aggression and rape, but also refers to acts of sexual violence that do not meet the legal definition of sexual aggression or rape (Tedeschi & Felson, 1994). The findings of our study support demonstrated that sexual coercion is viewed as being forced through power and manipulation, rather than physical force, where one cannot refuse sexual interaction.\n\nDifferences in definition of sexual harassment, sexual coercion and sexual violence by the WHO’s World Report on Violence and Health definition and Georgian students’ perspectives in the study. It seems that there are cultural differences between what we perceive as sexual violence and what we do not. For example, Heise et al., discussed that within any one society there may be contested areas where the line between acceptable and nonacceptable levels of sexual coercion are in transition. In the United States, for example, the line among dating partners is clearly changing. Acts that would have been cited as the women’s fault or ascribed to “bad manners” on the part of the man twenty years ago, are increasingly being correctly labelled “date rape” (Heise et al., 1996). Grubb and Harrower’s (2009) study indicated that when a rapist and victim knew each other in some capacity, university students were more likely to blame the female victim to a greater extent. In their opinion, when there was some previous contact between those involved in the rape, respondents made a shift in how they delegated blame because they understood that relationships often involve miscommunications, and that different interpretation of events are likely to occur. Respondents might have felt that blame needed to be more shared in this type of situation (Grubb & Harrower, 2009). The findings from their study suggested that stranger rape and acquaintance rape need to be treated as distinct phenomena. Our study’s results implied that responsibility and culpability became more muddled once the rapist and rape victim have had some previous contact. The authors concluded that more qualitative work was needed to understand the thinking and reasoning behind attributions made in these two kinds of rape situations (Grubb & Harrower, 2009). In our study, Georgian students also attributed less responsibility to victims when the abuser is a stranger and the victim could not control the situation. In their view, when the victim could influence on the situation, the participants attributed more responsibility to victims, for example about the situation where was sexual harassment at the workplace.\n\nLandström et al. (2016) indicated that participants attributed least blame to sexual assault victims who had not previously flirted with the perpetrator and most blame was attributed to the flirtatious sexual harassment victim. The research consistently showed that rape victims may be blamed for the assault, but little was known about victim blaming in other sexual crimes. In their experiment, the researchers examined blame attributions for sexual assault and online sexual harassment. This is true for our study too. The research participants also attributed more responsibility to the victim, who communicated with the abusive party and the participants thought that she or he “looked like they were flirting” for example, in situations a female student (the victim) had had online communication with her lecturer (the abuser) and messages between them were about the victim’s family and private life. After this communication, he started sending her nude photos without her consent.\n\nOverall, this research provides useful information about Georgian students’ attitudes to sexual violence and their attributions of responsibility. The research can become the basis for effective prevention programs of sexual violence in Georgia.\n\n\nConclusion\n\nOverall, the study demonstrated that Georgian students’ perspectives on defining sexual harassment, sexual coercion and sexual violence, are in line with definitions that are formulated in existing literature on sexual violence. It is important to mention that students were selected from two biggest cities. This was done on purpose as, considering research aims and objectives, authors were interested in students’ perspectives in big cities, where youth was more exposed to new and alternatives cultures than their own.\n\nIn summary, the results of the present study yield several conclusions. First of all, sexual violence is not a new phenomenon in Georgia, but its consideration as a social problem is. Secondly, these issues have attracted the attention of researchers and experts for decades in Western countries, but in Georgia, it is still an unexplored field. Thirdly, there are no programs for prevention of sexual violence in Georgia and this is reflected in Georgian students’ attitudes toward sexual violence and attributions of responsibilities on this issue. Therefore, the given study is one of the first and essential steps in designing effective programs for preventing and controlling sexual violence in Georgia.\n\nThe core limitation of this study is that it does not include all Georgian society’s views about sexual violence issues, but only Georgian youth’s perspectives It was beyond the researchers’ control how honest the participants were in answering the questions, but to ensure honest responses of the respondents, in-depth interviews implied additional in-depth questioning and in some instances, reformulation of major and additional questions. This allowed to understand whether respondent was not confident in his/her responses and to double check the views and attitudes that respondents provided.\n\n\nData availability\n\nFigshare: Sexual Violence and the Object of Attribution of Responsibility in Georgian Students\n\nhttps://figshare.com/articles/dataset/Data_docx/19123637 (Mchedlishvili & Zhghenti, 2022)\n\nThis project contains the following underlying data:\n\n• Data.pdf (transcript of interviews)\n\n• Quantitative results data (the percentages relating to the participants assessments of each case)\n\nFigshare: Sexual Violence and the Object of Attribution of Responsibility in Georgian Students\n\nhttps://figshare.com/articles/dataset/Data_docx/19123637 (Mchedlishvili & Zhghenti, 2022)\n\nThis project contains the following extended data:\n\n• Interview guide", "appendix": "Acknowledgments\n\nThe authors would like to express their appreciation to all the participants of the research interviews.\n\n\nReferences\n\nAbbey A, Wegner R, Woerner J, et al.: Review of survey and experimental research that examine the relationship between alcohol consumption and men’s sexual aggression perpetration. 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Reference Source\n\n\nFootnotes\n\n1 For detailed case by case percentages of qualitative data see appendix link at the end of article. The percentages in given data only have a descriptive value and do not have any significance in qualitative analysis." }	[ { "id": "138210", "date": "11 Jul 2022", "name": "Lakshmi Lingam", "expertise": [ "Reviewer Expertise Gender", "health", "VAW", "GBV and health systems research", "gender and development" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nReview of the paper titled: ‘A qualitative study of sexual violence and attribution of responsibility in Georgian youth’\nThis is a paper written by Natalia Mchedlishvili and Nino Zhghenti based on research among 37 Georgian youth selected from two large Universities. The paper is based on primary research carried out from September 2019 to March 2021, where the researchers used 12 short case studies to elicit responses on how young people view cases of sexual harassment, sexual coercion, and sexual violence. The information on the case studies has been given as part of the links to data set – where the quantitative data and qualitative responses – were shared.\nThe paper throws light on what is the attribution of responsibility on the victim/survivor of sexual violence in young people’s minds depending on the type of experience of sexual harassment or coercion or violence shared with them as a short case story. It is evident from the data that the attribution of responsibility on the perpetrator is higher in the case of stranger violence and a person who is in authority or is senior age wise. However, the attribution of responsibility shifts on the woman if she is found to be in contact or knows the perpetrator or is in communication prior to the episode.\nKey limitations of the paper:\n\nBeyond sharing references and a few data points about sexual violence and attribution of responsibility on the woman in Georgia, the paper does not contextualise any information on the country, its sexual mores (dating culture, age of introduction to sexual activity among young people), age of consent, legal definitions of sexual harassment, coercion and sexual violence; and broader debates around sexual violence. The significance of this research to the ongoing debates on sexual violence and sexual harassment will be useful to the reader. That is missing in the paper.\n\nThe case studies and the results are given as links to the paper, but not integrated into the paper. It will be valuable to create a matrix of all the 12 cases and highlight key issues around age of the woman; the age, gender, and power position of the individuals in the case study, etc., to be able to provide a reference point to the results and discussion.\n\nThe sample is only 37, but quantitative data in terms of percentages is given as a link to the main paper. Qualitative and quantitative responses to case studies were collected but the paper integrates very little data from the study. The qualitative responses could be used to provide insights into the thinking of young people.\n\nIt is evident from the questions and multiple choice responses provided to the questions, that there is a lot of ambiguity in the questions and responses. The validity and reliability of the data of the study, both in terms of the sample size and the way the questions and responses are structured, are compromised.\n\nSome of the questions are following a case:\nDo you think this case is sexual harassment, sexual coercion or sexual violence?\nYes, it is –97.2% No, it is not –2.8%\nWhy do you think this case is sexual harassment, sexual coercion or sexual violence?\nThe second party felt herself unpleasant and insulted- 94.5% There were elements of threat - 78.3% At work a person was not judged according to her real skills and sexual content was used as criteria - 86.4%\nIn both the questions above, all the three elements of the study – sexual harassment, sexual coercion, and violence are placed together in the question and the responses are unrelated to the questions. It is not clear how the percentages are calculated.\nRecommendation:\n\nThe paper has to be rewritten keeping an international audience in mind and also contextualize the study within the Georgian legal system and cultural context of gender and sexuality. In the absence of an understanding of sexual harassment, sexual coercion, and sexual violence by the authors within a broader legal or normative context, it is unclear how the young people’s responses are being assessed by the authors.\nIt will be useful to have the caselets and the responses built into the paper rather than shared separately.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "162372", "date": "09 Mar 2023", "name": "Júlia Garraio", "expertise": [ "Reviewer Expertise Gender Studies", "sexual violence", "media studies" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe paper has potential to offer an important contribution to the field of sexual violence and harassment. The analysis of the perceptions of sexual harassment, sexual coercion and sexual violence among Georgian university students could provide important material to develop prevention strategies and initiatives in Georgia. However, in its current state, the paper requires a revision; it should be rewritten with an international audience in mind.\nNotions, understandings and imaginaries of sexual violence are historically and socially situated and framed; therefore, the authors should articulate the study with the specific context of Georgia (legislation on sexual violence, data on the incidence of sexual violence and sexual harassment in the country, sexual mores, feminist activism and debates about sexual harassment and violence, eventual repercussions of MeToo and hashtag activism). How are sexual harassment, sexual coercion and sexual violence defined by Georgian law? Were there major changes in the law? If there were changes, how and why were they introduced (pressure for international institutions, grassroots movements...)? Did they have any impact in the incidence and perception of sexual harassment, coercion and violence? Were there moments of feminist activism and/or media cases that fostered a public/national debate about sexual violence? Were there repercussions of MeToo in the country? And if so, how do the perceptions of the participants interact with the law and the recent history of the country, especially with debates about women's rights and feminist activism?\nThere should be also a stronger articulation between the analysis of the corpus and important conceptual and theoretical literature in the field of sexual violence (e.g., rape myths, rape scripts...).\n\nI also suggest some revision of the structure of the text. For instance, the conclusion has information that should come earlier (e.g., criteria for the selection of participants, objectives of the study, the lack of prevention initiatives in the country).\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "165839", "date": "17 Apr 2023", "name": "Lizzie Seal", "expertise": [ "Reviewer Expertise Criminology", "Historical Criminology", "Gender", "Race" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis article is based on research into students’ views on sexual violence in Georgia. The researchers conducted qualitative interviews with students at two different universities and found students attributed a range of meanings and definitions to different aspects of sexual violence. There was variation in terms of how they perceived blame and responsibility. As the authors argue, there is a lack of research about sexual violence and attitudes towards it in relation to Georgia and other countries in the region, making this a welcome study.\nThe article is clearly set out and the aims of the research and the methods used are explained well. The discussion section could be clearer in terms of highlighting the implications of the main findings and needs to express the findings in relation to attitudes as that is what the research was about. For example, the authors claim ‘Our research revealed that, in comparison to other forms of sexual violence, sexual harassment can be limited to verbal expression as well as unwanted touching.’ The research did not reveal that, it revealed that the students who participated in the research thought that. This needs to be clarified in the way the discussion section is phrased, not just in relation to the quoted statement but also in relation to some other elements of the discussion.\nThe literature review section describes Georgia as a patriarchal culture; this needs a little more explanation. What are the culturally distinctive aspects of patriarchal culture in Georgia? Could the discussion section revisit these aspects as means of understanding the findings of the research?\nThe researchers state that there are no programmes for the prevention of sexual violence in Georgia and that the findings could contribute to the design of such programmes. I agree that this seems to be a valuable contribution the research could make. Therefore, it would be worth further outlining how the findings could contribute to such programming.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? No source data required\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/11-499
https://f1000research.com/articles/12-528/v1	22 May 23	{ "type": "Research Article", "title": "The altered lung microbiome dynamics in patients with moderate and severe COPD compared to the healthy group in the Indian population", "authors": [ "Druti Hazra", "Fayaz SM", "Kiran Chawla", "Vitali Sintchenko", "Elena Martinez", "Rahul Magazine", "Nayana Siddalingaiah", "Druti Hazra", "Fayaz SM", "Vitali Sintchenko", "Elena Martinez", "Rahul Magazine", "Nayana Siddalingaiah" ], "abstract": "Background: Microbial culture-independent sequencing techniques have advanced our understanding of host-microbiome interactions in health and disease. The purpose of this study was to explore the dysbiosis of airway microbiota in patients with moderate or severe chronic obstructive pulmonary disease (COPD) and compare them with healthy controls. Methods: The COPD patients were investigated for disease severity based on airflow limitations and divided into moderate (50%≤FEV1<80% predicted) and severe groups (FEV1<50% predicted). Spontaneous sputum samples were collected and, the V3-V4 regions of the 16S rRNA coding gene were sequenced to examine the microbiome profile of COPD and healthy participants. Results: A total of 45 sputum samples were collected from 17 severe COPD, 12 moderate COPD cases, and 16 healthy volunteers. The bacterial alpha diversity (Shannon and Simpson’s index) significantly decreased in the moderate and severe COPD groups, compared to healthy samples. A significantly higher proportion of Firmicutes and Actinobacteria were present in moderate COPD, and Proteobacteria numbers were comparatively increased in severe COPD. In healthy samples, Bacteroidetes and Fusobacteria were more abundant in comparison to both the COPD groups. Among the most commonly detected 20 bacterial genera, Streptococcus was predominant among the COPD sputum samples, whereas Prevotella was the top genus in healthy controls. Linear discriminant analysis (LDA>2) revealed that marker genera like Streptococcus and Rothia were abundant in moderate COPD. For severe COPD, the genera Pseudomonas and Leptotrichia were most prevalent, whereas Fusobacterium and Prevotella were dominant in the healthy group. Conclusions: Our findings suggest a significant dysbiosis of the respiratory microbiome in COPD patients. The decreased microbial diversity may influence the host immune response and provide microbiological biomarkers for the diagnosis and monitoring of COPD.", "keywords": [ "Microbiome", "chronic obstructive pulmonary disease", "respiratory pathology", "16S rRNA gene sequencing", "microbial populations" ], "content": "Introduction\n\nChronic obstructive pulmonary disease (COPD) is a heterogenous lung pathology, manifesting with persistent and progressive respiratory symptoms, airway obstruction, and inflammation due to structural abnormalities.1 COPD is one of the leading causes of mortality and morbidity globally and its burden is predicted to rise further in the upcoming years due to constant exposure to air pollution, respiratory pathogens, and the growing elderly population.1–3 COPD is a treatable but incurable disease, and often switches between a stable to an exacerbated state disease.4 The frequent exacerbation and worsening of respiratory symptoms affect the individual quality of life with a significant socioeconomic burden and impose a huge healthcare management cost.5,6 The progression of COPD often leads to chronic inflammation and major destruction of the lung and airway, which disrupts pulmonary microbiome homeostasis. Commensal microbes play a crucial role in innate immune regulation, protecting against invading pathogens and maintaining epithelial integrity. The recent advancement of culture-independent next-generation sequencing techniques has uncovered the diverse microbial communities colonizing the respiratory mucosa and recognized their roles in health and disease.7–9\n\nIn the past, several studies have evaluated lung microbiome composition and its association with COPD manifestations. The changes in bacterial diversity and several significant taxa have been identified in COPD patients, which play a role in disease progression. Although a high heterogeneity has been observed among the studies, potentially attributed to the underlying health conditions in different populations and the dynamics of microbial ecology in the COPD Indian population, it remains poorly understood. This study aimed to evaluate the changes in lung microbiome diversity of patients with moderate or severe COPD and compare them with the microbiomes of healthy controls.\n\n\nMethods\n\nThis study was approved by the Kasturba Medical College and Kasturba Hospital Institutional Ethics Committee [IEC: 479/2019].\n\nWritten informed consent was obtained from all the participants.\n\nTo conduct this prospective observational study, sputum samples were collected from eligible COPD participants, who presented to our hospital and were diagnosed with COPD in accordance with the 2019 Global Initiative for Chronic Obstructive Lung Disease (GOLD) guideline.10 Their lung functions were measured using spirometry. The enrolled cases were regrouped into moderate COPD (50%≤FEV1<80% predicted) and severe COPD (FEV1<50% predicted). The exclusion criteria for COPD participants were: (a) age ≤40 years, (b) patients diagnosed with other respiratory diseases or immunosuppression, and (c) history of antibiotic usage within four weeks prior to sample collection. Healthy controls include individuals ≥40 years of age and those not having any apparent illness. There was no gender-based exclusions or restrictions for recruiting participants. Demographic and clinical information was obtained for the enrolled participants. This study was approved by the Kasturba Medical College and Kasturba Hospital Institutional Ethics Committee (IEC: 479/2019) and the participants were enrolled after providing written informed consent.\n\nThe participants were instructed to cough up sputum into a sterile container and samples were transported on ice to the laboratory. All sputum samples were evaluated with routine conventional culturing and an aliquot of it was stored at −80°C for the DNA extraction. The sputum samples were homogenized using an equal volume of 0.2% dithiothreitol (DTT) (Sigma Aldrich, USA), and lysozyme-based Qiagen DNA Mini kit (Qiagen, USA) was used to extract the genomic DNA according to the manufacturer’s protocol. The Qubit 2.0 fluorometer (Thermo Fisher Scientific, USA) was used to measure the purity and concentration of the extracted DNA.\n\nThe purified DNA was further processed for the 16S rRNA V3-V4 regions targeted amplification to uncover the bacterial community in sputum samples. Based on the Illumina protocol,11 PCR amplification of V3-V4 hypervariable regions (~456 bp) were performed using the primer pair 341F/785R. Sequencing adapters and dual index barcodes were added with a limited cycle of PCR. After the quality assessment, multiplex amplified libraries were pooled equally and paired-end reads (2X300 bp) were generated using the MiSeq instrument (Illumina, San Diego, CA, United States).\n\nThe sequenced raw data were processed using the standard Mothur v1.46.1 pipeline.12 A quality check of the reads was carried out and the low-quality reads and chimeras were removed. Contigs were created from the paired-end reads. Unique sequences were considered by removing the identical sequences. The quality reads were then aligned to the SILVA database and clustered into Operational Taxonomic Units (OTUs) at 97% similarity and taxa level 4, which is similar to the genus for Bacteria.13\n\nThe microbial community diversity profiles among the moderate COPD, severe COPD, and healthy groups were analyzed using the alpha and beta-diversity metrics. The Shannon, Simpson, and Chao1 matrices were used to measure bacterial Alpha-diversity and an ANOVA test was done to estimate significant differences. Beta diversity was performed using the Principal Coordinate Analysis (PCoA) method along with the Bray-Curtis index as distance measure and permutational multivariate analysis of variance (PERMANOVA) for the significant measure. The genus biomarkers (discriminative genera among the groups) were identified using linear discriminant analysis effect size (LEfSe) and a cut-off linear discriminant analysis (LDA) score >2.0.\n\n\nResults\n\nSputum samples from COPD cases (12 moderate COPD and 17 severe COPD) and 16 healthy volunteers were included in the study. The participants’ clinical and demographic features like gender, age, BMI, pulmonary function tests (FEV1% predicted, FEV1 and FEV1/FVC values), mMRC dyspnoea score, and whether they were current smokers are summarized in Table 1. At the time of sampling, all COPD patients were in an exacerbated state of the disease and no significant growth of respiratory pathogens was detected in the sputum cultures.\n\nA total of 91,863 OTUs were recovered at a 97% sequence identity with 32,665 OTUs in the patients with moderate COPD, 40,842 OTUs in severe COPD, and 30,980 OTUs in the healthy group. A significantly lower bacterial alpha diversity (Simpson’s and Shannon's index) was observed in moderate COPD and severe COPD samples compared to the healthy group (p<0.05, ANOVA test). The Chao1 index measured the species richness within groups and exhibited no differences (p>0.05) (Figure 1A). The Beta diversity represented by PCoA showed a significant difference in bacterial community clustering among the groups (p<0.01, PERMANOVA test) (Figure 1B).\n\n(A) The measures of alpha diversity indices showed a significant difference (p < 0.05 and *p < 0.01, ANOVA test). (B) Principal coordinate analysis (PCoA) revealed a distinct bacterial community clustering among the groups (P< 0.01, PERMANOVA). (C) Venn diagram of the core microbiota in tested samples.\n\nAs the Venn diagram of the core sputum microbiota (Figure 1C) illustrates, out of the total 91863 OTUs 35.6%, 27.2%, and 26.3% OTUs were unique to severe COPD, moderate COPD, and healthy groups respectively. While 2,649 (2.9%) OTUs were shared by all three groups and 5,831 (6.3%) OTUs were shared between moderate and severe COPD groups, 4,447 (4.8%) OTUs were common for moderate COPD and healthy, and, 4,995 (5.4%) OTUs were common for severe COPD and healthy groups.\n\nThe most prevalent microbial phyla in the sputum samples of moderate COPD, severe COPD, and healthy were Firmicutes (46.3%, 35.6%, and 31.9%) followed by Bacteroidetes (20.0%, 26.8%, and 28.7%) Proteobacteria (12.9%, 17.4%, and 16.1%), Actinobacteria (14.0%, 8.5%, and 5.9%), and Fusobacteria (5.3%, 9.5%, and 13%) (Figure 2A). A significantly higher proportion of Firmicutes and Actinobacteria were present in respiratory samples from patients with moderate COPD and Proteobacteria was comparatively increased in severe COPD, whereas in healthy individuals, Bacteroidetes and Fusobacteria were present in a higher abundance compared to both COPD groups.\n\n(A) Relative abundance at the Phyla level, (B) Relative abundance at the genus level, and (C) LEfSe analysis represent the discriminative genera among the groups (LDA>2).\n\nIn the cohort of patients with moderate COPD, the top five most commonly detected genera were Streptococcus (28.2%), Rothia (11.4%), Prevotella (8.3%), Porphyromonas (7.9%), and Gemella (6.2%). The dominant genera in severe COPD were Streptococcus (20.2%), Prevotella (11.7%), Porphyromonas (10.1%), Leptotrichia (5.9%), and Rothia (5.3%). In healthy individuals, Prevotella (16.5%) was the most prevalent genus, followed by Streptococcus (13.0%), Neisseria (7.1%), Fusobacteria (6.6%), and Velionella (6.2%). An increasing abundance of Streptococcus (p<0.05), and Rothia was observed in moderate COPD samples, whereas Morexalla and Pseudomonas were relatively higher in severe COPD. Genera like Prevotella, and Fusobacteria were abundantly present in healthy individuals in comparison to moderate COPD. Figure 2B demonstrates the top bacterial genera present in the sputum of patients with moderate or severe COPD, and healthy groups.\n\nThe LEfSe analysis was performed to detect the discriminative genera among the groups (Figure 2C). In moderate COPD, six marker genera (LDA>2) Streptococcus, Rothia, Gemella, Carnobacteriaceae, Capnocytophaga, and Weissella were identified. For the severe COPD, genera Pseudomonas and Leptotrichia were higher, whereas Fusobacterium, Bacteroidales, Peptostreptococcus, Prevotella, Porphyromonas, Alloprevotella, etc. were dominant in healthy individuals.\n\n\nDiscussion\n\nThe severity of COPD is often influenced by environmental exposures, host genetic makeup, and airway host-microbiome interactions. This study demonstrated the microbial alpha diversity of moderate and severe COPD groups decreased significantly in comparison to the healthy control group. Our findings added important evidence to the understanding of the microbial population dynamics in COPD. Su et al. also reported a decreased bacterial diversity in the acute exacerbations of COPD compared to healthy controls, which was consistent with our findings.14 Ramsheh et al. also found a higher Alpha diversity in healthy individuals than in COPD bronchial brush samples.15 The consistent results of these studies indicate the altered microbial diversity in COPD patients compared to healthy and its potential role as a disease marker.\n\nAccording to our findings, the alpha diversity of moderate COPD declined compared to the severe COPD group. However, Yang et al. and Li et al. did not observe any significant difference in microbial diversity between mild COPD and severe COPD groups, whereas Garcia-Nuñez et al. reported a decreased alpha diversity in advanced COPD compared to moderate-to-severe disease.16–18 This inconsistency among studies could be due to distinct sampling methods, COPD states and exacerbations, and different geographic regions.\n\nIn this study, the most dominant phylum was Firmicutes, predominantly present in all the groups, in concordance with previous reports.15,16,19,20 In healthy controls, we observed a higher proportion of Bacteroidetes and Fusobacteria, likewise reported by Ramsheh et al.15 Proteobacteria was present in relatively higher proportions in patients with severe COPD, which is consistent with previous studies examining bronchoalveolar lavage (BAL)21 and sputum18,22 samples. According to Wang et al., the increased abundance of Proteobacteria might trigger the pro-inflammatory mediators of the host, which leads to dysbiosis of lung microbiomes.22\n\nStreptococcus was the predominant genus among the COPD sputum samples, whereas Prevotella was significantly higher in healthy controls. A multi-centric study reported that Prevotella promotes normal lung function and the severity of COPD increases with its decreasing abundance.15 A lung co-infection mouse model study conducted by Horn KJ et al. suggested that an increased abundance of airway Prevotella can accelerate the innate immune response and rapid pathogen clearance from the lung.23 In the moderate COPD group, we noted a higher abundance of Streptococcus and Rothia, which was similar to previously published studies of COPD samples.14,16–18,22,24 Li W et al. also observed a higher abundance of Rothia in the mild COPD group compared with the severe group,17 which was negatively correlated with pro-inflammatory markers, which might reduce the disease severity and exacerbation frequency in COPD.25 Another genus, Morexalla, which was abundant in severe COPD samples, was likewise reported by Wang et al.26 and Ramsheh et al.15 According to Wang et al., the relative abundance of Moraxella increased during COPD exacerbations and was also linked to the host interferon signaling pathway.26 Ramsheh et al. revealed that an increased abundance of Moraxella was associated with the expression of the IL-17 and TNF inflammatory pathways, which elicit the severity of COPD.15 These studies indicate that patients might suffer an altered lung microbial diversity during COPD disease severity, which means microbiota is a potential marker to predict the prognosis in COPD cases and may change the disease management.\n\nThere are a few limitations of our study. First, the population size of this study was small and the samples were collected from a single center. Second, the virome and mycobiome diversity of sputum were not evaluated. Future multi-centre studies in larger diverse populations are required to conclude the stability and alteration of microbiota in health and disease.\n\n\nConclusions\n\nOur findings suggested a significant loss of the sputum microbiome diversity in patients with COPD. This decrease is more pronounced in patients with severe disease. The dysbiosis of lung microbiota may cause an alteration of the mucosal immune system and further facilitate inflammation in the lung. Therefore, the improved understanding of the link between the respiratory microbiome and disease may offer new opportunities for an alternative management approach for COPD.\n\n\nAuthor contributions\n\nConceptualization: D.H., K. C.; Methodology: D. H., R. M., K. C.; Data analysis: F. SM., D. H., E. M.; Writing - original draft preparation: D. H.; Writing - review and editing: K. C., V. S., F. SM., N. S.; Supervision: K. C., V. S.", "appendix": "Data availability statement\n\nZenodo: Sequenced Data COPD and healthy, https://doi.org/10.5281/zenodo.7697770. 27\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgments\n\nThe authors gratefully acknowledge the Indian Council of Medical Research (ICMR) for providing a Senior Research Fellowship (SRF) for the study.\n\n\nReferences\n\nGlobal Initiative for Chronic Obstructive Lung Disease (GOLD): Global strategy for prevention, diagnosis and management of COPD: 2023 report.2023. Reference Source\n\nHerse F, Kiljander T, Lehtimäki L: Annual costs of chronic obstructive pulmonary disease in Finland during 1996-2006 and a prediction model for 2007-2030. NPJ Prim. Care Respir. Med. 2015; 25: 15015. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMathers CD, Loncar D: Projections of Global Mortality and Burden of Disease from 2002 to 2030. PLoS Med. 2006; 3(11): e442. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSafiri S, Carson-Chahhoud K, Noori M, et al.: Burden of chronic obstructive pulmonary disease and its attributable risk factors in 204 countries and territories, 1990-2019: results from the Global Burden of Disease Study 2019. BMJ. 2022; 378: e069679. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIheanacho I, Zhang S, King D, et al.: Economic Burden of Chronic Obstructive Pulmonary Disease (COPD): A Systematic Literature Review. Int. J. Chron. Obstruct. Pulmon. Dis. 2020; 15: 439–460. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGutiérrez Villegas C, Paz-Zulueta M, Herrero-Montes M, et al.: Cost analysis of chronic obstructive pulmonary disease (COPD): a systematic review. Heal. Econ. Rev. 2021; 11(1): 31. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWang L, Hao K, Yang T, et al.: Role of the Lung Microbiome in the Pathogenesis of Chronic Obstructive Pulmonary Disease. Chin. Med. J. (Engl). 2017; 130(17): 2107–2111. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLi Z, Li Y, Sun Q, et al.: Targeting the Pulmonary Microbiota to Fight against Respiratory Diseases. Cells. 2022; 11(5): 916. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNatalini JG, Singh S, Segal LN: The dynamic lung microbiome in health and disease. Nat. Rev. Microbiol. 2022; 21: 222–235. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGlobal Initiative for Chronic Obstructive Lung Disease: Global Strategy for the Diagnosis, Management, and Prevention of Chronic Obstructive Pulmonary Disease.2019. Reference Source\n\nIllumina: 16S metagenomic sequencing library preparation protocol: preparing 16S ribosomal RNA gene amplicons for the Illumina MiSeq system. Part no. 15044223 Rev B. Illumina, San Diego, CA.2013. Reference Source\n\nSchloss PD, Westcott SL, Ryabin T, et al.: Introducing mothur: open-source, platform-independent, community-supported software for describing and comparing microbial communities. Appl. Environ. Microbiol. 2009; 75(23): 7537–7541. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nQuast C, Pruesse E, Yilmaz P, et al.: The SILVA ribosomal RNA gene database project: improved data processing and web-based tools. Nucleic Acids Res. 2013; 41(Database issue): D590–D596. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSu L, Qiao Y, Luo J, et al.: Characteristics of the sputum microbiome in COPD exacerbations and correlations between clinical indices. J. Transl. Med. 2022; 20(1): 76. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGarcia-Nuñez M, Millares L, Pomares X, et al.: Severity-related changes of bronchial microbiome in chronic obstructive pulmonary disease. J. Clin. Microbiol. 2014; 52(12): 4217–4223. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMayhew D, Devos N, Lambert C, et al.: Longitudinal profiling of the lung microbiome in the AERIS study demonstrates repeatability of bacterial and eosinophilic COPD exacerbations. Thorax. 2018; 73(5): 422–430. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSze MA, Dimitriu PA, Hayashi S, et al.: The lung tissue microbiome in chronic obstructive pulmonary disease. Am. J. Respir. Crit. Care Med. 2012; 185(10): 1073–1080. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nErb-Downward JR, Thompson DL, Han MK, et al.: Analysis of the lung microbiome in the “healthy” smoker and in COPD. PLoS One. 2011; 6(2): e16384. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWang Z, Bafadhel M, Haldar K, et al.: Lung microbiome dynamics in COPD exacerbations. Eur. Respir. J. 2016; 47(4): 1082–1092. PubMed Abstract \| Publisher Full Text\n\nHorn KJ, Schopper MA, Drigot ZG, et al.: Airway Prevotella promote TLR2-dependent neutrophil activation and rapid clearance of Streptococcus pneumoniae from the lung. Nat. Commun. 2022; 13(1): 3321. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPragman AA, Lyu T, Baller JA, et al.: The lung tissue microbiota of mild and moderate chronic obstructive pulmonary disease. Microbiome. 2018; 6(1): 7. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSagar S, Morgan ME, Chen S, et al.: Bifidobacterium breve and Lactobacillus rhamnosus treatment is as effective as budesonide at reducing inflammation in a murine model for chronic asthma. Respir. Res. 2014; 15(1): 46. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWang Z, Maschera B, Lea S, et al.: Airway host-microbiome interactions in chronic obstructive pulmonary disease. Respir. Res. 2019; 20(1): 113. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDruti H, Kiran C, Fayaz SM: Sequenced Data COPD and healthy. [Data set]. Zenodo. 2023. Publisher Full Text" }	[ { "id": "175709", "date": "20 Jul 2023", "name": "Eduard Monso", "expertise": [ "Reviewer Expertise Respiratory diseases / COPD / Lung Cancer / Respiratory microbiome" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors described the respiratory microbiome in healthy subjects and COPD patients with different levels of severity, confirming with their study the characteristics of the COPD microbiome previously reported in other communities. The main value of the paper, accordingly, is the analysis of a community living in a specific geographic area, not previously focused. There are some concerns that need to be raised, however, as followed.\nThe title state that the study focus on the lung microbiome dynamics. However, considering that the studied sample is the sputum, it can only be representative of the bronchial microbiome, not the lung. Furthermore, because there has been a single sampling in the studied population, the study cannot be considered \"dynamic\".\n\nIn the description of the features of the study participants, it is said that they are exacerbated. I think that this is a typewriting mistake, because all the text suggests that the participants are stable (they have not used antibiotics the previous months).\n\nIn the table, moderate COPD patients have an average FEV1/FVC of 90. If that is the case, most of them do not suffer from COPD, and probably have only chronic bronchitis. This is a main point that needs to be clarified.\n\nThe comparisons would have been much more clear if all COPD patients are compared with healthy subjects, and later, excluding healthy subjects, severe and moderate COPD patients were compared.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "9957", "date": "10 Aug 2023", "name": "Kiran Chawla", "role": "Author Response", "response": "Response to the Reviewer Comments: Query 1. The title state that the study focus on the lung microbiome dynamics. However, considering that the studied sample is the sputum, it can only be representative of the bronchial microbiome, not the lung. Furthermore, because there has been a single sampling in the studied population, the study cannot be considered \"dynamic\". Response: Thank you for your valuable suggestions. In this study, we characterized sputum, an easily obtainable non-invasive sampling method that represents a mix of upper and lower respiratory tracts, used for routine diagnosis of lower-respiratory tract infection. The term dynamics have been used here to refer to the changes in microbiome composition among the groups Query 2. In the description of the features of the study participants, it is said that they are exacerbated. I think that this is a typewriting mistake, because all the text suggests that the participants are stable (they have not used antibiotics the previous months). Response: The enrolled participants were the known cases of COPD, admitted to our hospital for the management of the latest exacerbation state, and a sample was collected before administrating antibiotics. So the patients have not used antibiotics in the previous months. Also, one of our exclusion criteria is antibiotic usage within four weeks prior to sample collection since antibiotics can alter the microbiome compositions. Query 3. In the table, moderate COPD patients have an average FEV1/FVC of 90. If that is the case, most of them do not suffer from COPD, and probably have only chronic bronchitis. This is a main point that needs to be clarified. Response: Thank you, Sir, for mentioning this point. This was a typewriting error, which we didn’t notice earlier. We rechecked our data and there was an error in that particular row of that table. It should be the moderate COPD average FEV1/FVC was 64.7 ± 5.6 and Severe COPD 61 ± 7.4 Query 4. The comparisons would have been much more clear if all COPD patients are compared with healthy subjects, and later, excluding healthy subjects, severe and moderate COPD patients were compared. Response: We compared the sputum microbiome in healthy, severe, and moderate COPD patients as per the objective. We admire your valuable suggestions and we will consider them for our future studies." } ] }, { "id": "182958", "date": "28 Jul 2023", "name": "Veronica Ueckermann", "expertise": [ "Reviewer Expertise Infectious diseases", "Critical care", "lung microbime", "Tuberculosis" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe manuscript explores loss of microbial diversity in the sputum samples of patients with moderate and severe COPD, and compares the results with a healthy control group. The healthy control group included some smokers and the study population was dominated by males.\nThe methodology was well described and both the methods and analysis were appropriate for work in the lung microbiome. The quality of the sputum samples voluntary produced were not discussed and a degree of oropharyngeal contamination is likely.\nFindings are consistent with previously published work and highlight once again signals of dysbiosis in the lung microbiome of patients with COPD. Had the sample size allowed, a comparison between current and previous smokers may have provided additional insights.\n\nThe limitations of the study were acknowledged - both the small sample size and the fact that the virome (which is becoming increasingly important in COPD) was not evaluated.\n\nAlthough the findings of the study were not completely novel, the differences by disease severity was interesting and the study does add to a growing body of literature that enhances the understanding of the lung microbiome in respiratory disease. The methodology was sound and the discussion well written.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [] } ]	1	https://f1000research.com/articles/12-528
https://f1000research.com/articles/12-851/v1	19 Jul 23	{ "type": "Research Article", "title": "Knowledge of obstetric danger signs and associated factors among pregnant women attending antenatal care services at Thai community hospital", "authors": [ "Pruk Koovimon", "Kasiphak Kaikaew", "Khanittha Mahoree", "Thanapob Bumphenkiatikul", "Pruk Koovimon", "Kasiphak Kaikaew", "Khanittha Mahoree" ], "abstract": "Background: To decrease preventable maternal mortality, providing health education to all parties is mandatory. Good knowledge, including awareness of pregnant women regarding obstetric danger signs (ODS), leads to appropriate practices and services. The knowledge of ODS varies among countries and regions. Since the data in rural regions of Thailand remains unavailable, this study aimed to identify the prevalence of good ODS knowledge and associated factors among pregnant women attending antenatal services at a Thai community hospital. Methods: We performed a cross-sectional, analytical study in 415 singleton pregnant women who visited the antenatal clinic at Wang Saphung Hospital, Loei, Thailand. A well-trained research assistant interviewed all participants using the data record form containing twenty items on the demographic and obstetric data and sixteen items on ODS knowledge. An ODS score of at least 75% (12 points) was considered a good level of knowledge. Results: A total of 275 participants (66.27%) had good knowledge of ODS. The most recognized ODS was vaginal bleeding whereas the least recognized ODS during pregnancy was convulsion; the least recognized ODS during labor and delivery was retained placenta. Multivariate regression analysis showed that the predictive factors of good OBS knowledge included a higher education level, maternal age of at least 20 years, and having medical personnel as a source of knowledge. Conclusions: In a rural setting of Thailand, two-thirds of pregnant women had good ODS knowledge. Identifying those at risk for fair and poor ODS knowledge and prompt management for the vulnerable subgroups might help decrease maternal mortality.", "keywords": [ "Awareness", "Women's health", "Complications", "Maternal mortality", "Delivery", "Healthcare", "Health education", "Postpartum" ], "content": "Introduction\n\nThe maternal mortality ratio (MMR) is one of the indicators of the public health status of a specific region. Several global authorities devised a long-term plan to reduce global MMR. In 2000, the United Nations (UN) declared the Millennium Development Goals, which included “improving maternal health,” as Millennium Development Goal 5 (MDG-5). The MDG-5 aimed for a 75% reduction in the global MMR.1 In 2015, the United Nations Department of Economic and Social Affairs (UNDESA) and all member countries agreed to aim to accomplish the Sustainable Development Goals (SDG) by 2030. The target of SDG Goal 3 is to “ensure healthy lives and promote well-being for all at all ages.” This goal includes “reducing the global MMR to less than 70 per 100,000 births, with no country having a maternal mortality rate of more than twice the global average.”2 Having these goals consistently included in international plans implicitly reflects the global significance of this unresolved situation.\n\nGlobally, the MMR is decreasing.3 The index differs among countries, ranging from 23.8 deaths per 100,000 100,000 live births in the US to 442 deaths per 100,000 births in Africa.4,5 Thailand’s maternal mortality rate (MMR) was 48.0 deaths per 100,000 live births in 2008.6 It fell to 37.0 deaths per 100,000 live births in 2017.7 Though the low baseline ratio met SDG Goal 3, achieving “equity in MMR for vulnerable populations at the sub-national level” is still the country’s target.8\n\nTo decrease preventable maternal mortality, providing health education to all parties involved in healthcare, including patients, families, communities, and medical personnel, is mandatory. Good knowledge will lead to appropriate attitudes and practices, i.e., prompt referral to appropriate medical services, and thus leading to a decrease in preventable maternal mortality. Unawareness of obstetric danger signs (ODS) will delay the decision to seek proper care and eventually lead to morbidity and mortality.9\n\nThe knowledge of ODS varies among countries and regions. The prevalence of mothers with good knowledge of ODS seems to be lower in some countries or in some rural areas of the countries that have a higher MMR. Several studies on ODS knowledge have been conducted in developing countries, focusing on urban and rural areas. Most of the studies found that less than half of the study population had good knowledge of ODS. In addition, the prevalence of good knowledge or awareness of ODS in rural areas was lower than in urban areas. One study was conducted to examine the prevalence of mothers with good knowledge of ODS and associated factors in a tertiary care university hospital in the capital city of Thailand and found that the prevalence of mothers with the good knowledge was around 60%.10 Currently, no data is available regarding the prevalence of mothers with good knowledge of ODS and associated factors in rural areas of Thailand, which are the subnational region at risk for higher MMR. Thus, our study aimed to identify the prevalence of mothers with good knowledge of ODS and associated factors among pregnant women attending an antenatal care clinic at a community hospital in a Northeastern province of Thailand. In addition, we also aimed to identify the variables that could predict the ODS knowledge of the pregnant women.\n\n\nMethods\n\nThe study protocol was approved by the Human Research Ethics Committee, Loei Provincial Public Health Office, (approval number: 0032.009/5503). Data collection was allowed by the director of Wang Saphung Hospital. Before the data collection, we obtained written informed consent from the participants or parents for participants under the legal age of consent.\n\nThis study is a cross-sectional, analytical study and was reported according to the STROBE statement for cohort studies. Participants were women with an ultrasound-confirmed singleton pregnancy who could understand Thai and had their antenatal clinic visit(s) at Wang Saphung Hospital, Loei Province, from 1st July 2021 to 30th September 2022. Those who were medical personnel or had any prior antenatal clinic visits in any other healthcare setting were excluded from the study to avoid contamination of ODS education during current gestation before participation. The study gave no incentives to participants. Participants’ partners were not included into this study.\n\nAll participants were informed about the study’s purposes, procedures, risks, and benefits. They were ensured that the decision was entirely voluntary. They could refuse or withdraw from the study at any time. Refusal, withdrawal, or having poor ODS knowledge would not affect the benefits or quality of care provided. A well-trained research assistant interviewed all participants.\n\nThe process took place in a private room during the clinic’s waiting period so as not to interfere with the care provided. The interview took approximately 30 minutes to complete. The research assistant completed two data record form sections. Section 1 consisted of twenty items on the demographic and obstetric data, which included parity, number of antenatal visits, and gestational age at the interview. Section 2 consisted of sixteen pre-coded closed-ended items on knowledge of ODS: twelve of which were knowledge during pregnancy and four were knowledge during labor and delivery. Participants were asked to spontaneously list all signs they perceived as dangerous or would urge them to seek proper care. The research assistant checked all ODS mentioned by each participant, then started asking for the participant’s knowledge of each of the rest of the ODS in section 2 of the data record form. Each response that acknowledged each ODS as dangerous was given one point. Zero point was given for unawareness of each ODS. We considered a score of at least 75% (12 points) to be a good level of knowledge, 50–74% (6–11 points) to be a fair level of knowledge, and 0–49% (0–5 points) to be a poor level of knowledge. The data collection process was derived from studies conducted in Thailand and Malaysia.10,11 The Cronbach‘s alpha for ODS items was equaled to 0.89, previously mentioned in one study with comparable participants, which indicated a good reliability of the record form.11\n\nWe calculated the sample size based on a previous study with a similar research design conducted by Kaewkiattikun et al.,10 in a medical school in an urban area of Thailand. We applied the power of 80% and a confidence level of 95% to determine the difference between groups. After adding an additional 10% to account for missing data, a total of 415 participants were recruited for this study. The study used a simple random sampling method. We limited the number of participants per day to 10 to ensure the quality of care and the data collected.\n\nTo analyze the data, a statistician used SPSS version 29 (IBM, Armonk, NY, USA). For the analysis of categorical data between groups, the Chi-square test was employed. Multivariate logistic regression analysis was used to identify independent variables with good knowledge of ODS. The results were presented in odds ratios and 95% confidence intervals (CI). P-values less than 0.05 were regarded as statistically significant.\n\n\nResults\n\nThere were 415 eligible pregnant women at the end of the enrolment. Of all these participants, 275 (66.27%) had good knowledge of ODS, 101 (24.34%) had fair knowledge, and 39 (9.40%) had poor knowledge. The most recognized ODS was vaginal bleeding, which accounted for 92.29% of the reported ODS during pregnancy and 80.96% of the reported ODS during labor and delivery. The least recognized ODS during pregnancy was convulsion (68.19%), while the least recognized ODS during labor and delivery was retained placenta (63.61%). The detailed results of the knowledge of ODS among antenatal women are shown in Table 1.\n\nODS refers to obstetric danger signs.\n\nWe classified those who knew at least 12 items out of 16 items (75%) as having a good level of ODS knowledge, 6–11 items (50–74%), and 0–5 items (0–49%) as having a fair level and a poor level of ODS knowledge, respectively. Using the Chi-square test, the identified factors that were significantly between the good and the fair/poor knowledge groups included participants’ age, education, occupation, marital status, gravida, and source of the ODS knowledge. After using the multivariate regression analysis to identify which of these characteristics were statistically significant predictors of good ODS knowledge, we found that participants’ age, education, and source of the ODS knowledge were predictors of good ODS knowledge, whereas participants’ occupation, marital status, and gravida were not statistically significant predictors of good ODS knowledge. The detailed results of the demographic characteristics of participants and their association with the level of ODS knowledge are shown in Table 2.\n\nODS, obstetric danger signs; AOR, adjusted odd ratio; CI, confident interval; THB, Thai Baht; ANC, Antenatal care; Ref, Reference; NA, not analyzed with logistic regression since the Chi-square p-value of the variables was greater than 0.05.\n\n\nDiscussion\n\nIn a community hospital-based antenatal care services in a Northeastern province of Thailand, we found that about two-thirds of the pregnant women had a good level of knowledge regarding ODS. We also found that the factors that were associated with the good ODS knowledge included age, education level, and source of the ODS knowledge that the pregnant women obtained the information.\n\nThe prevalence of good ODS knowledge in our study is higher than those in previous studies. One study in a Thai university hospital in an urban area reported a prevalence of 59.8%10 and another study in a teaching and referral hospital in Malaysia reported a prevalence of 48.3%.11 The finding that the prevalence of good ODS knowledge in Thai pregnant women was slightly higher than that of the Malaysian study might be due to the different score cut-off levels of good knowledge of ODS, that is, 80% (16 out of 20 items) in the Malaysian study whereas 75% (12 out of 16 items) in our study and the other Thai study. Several studies reported the knowledge of ODS among pregnant women in many countries, including India,12,13 Nepal,14,15 Malaysia,11 Ethiopia,16–28 Nigeria,29 Tanzania,30 Egypt,31 Jordan,32 Congo,33 and Uganda.34 The fact that the prevalence of mothers with good knowledge of ODS differed among studies could be because of the difference in participants’ demographic characteristics and the definition of good knowledge of ODS in each study. Overall, the prevalence of mothers with good knowledge of ODS was the lowest in Africa,16–29,35,36 especially in the remote area where most participants received lower education.\n\nFor the knowledge of each ODS among the study population, we found that vaginal bleeding is the most mentioned ODS during pregnancy and during labor/delivery among participants. Less than 70% of the participants reported epigastric pain, blurred vision, and convulsion during pregnancy as an ODS. A similar proportion mentioned convulsion and retained placenta during labor/delivery as ODS. These findings were congruent with the previous study in Thailand.10 Since vaginal bleeding is a visible, genital organ–related sign, it is easily recognized as an ODS among pregnant women. In contrast, since epigastric pain, blurred vision, and convulsion are symptoms of other organ systems, it was more complicated to educate pregnant women that these symptoms must also be perceived as an ODS.\n\nFor the secondary objective of the study, the predictive factors of good knowledge of ODS, we found that higher maternal education of at least a Bachelor’s degree, compared to high school or lower education, is one of the significant predictors. This finding is similar to the study from the Thai university hospital.10 Studies in other Asian countries, including Malaysia11 and Jordan,32 also found that higher maternal education was a significant predictive factor of good knowledge of ODS. Several studies from the African region,29,37 especially Ethiopia,20,21,24–26,28,38 where women received lower education, emphasized the predictive value of women’s education on the level of ODS knowledge. The level of education could be the source of the quality of ODS knowledge. Women with a lower education level could have more difficulty in understanding healthcare information about the importance of ODS given by others. The lower level of education might reflect lower opportunities for women since they might share poor attitudes and misinformation with peers at the same level of education. A study in Africa reported that ODS was perceived as a natural process of pregnancy or related to witchcraft.39 This attitude was a significant barrier, preventing the women facing ODS from seeking proper help.\n\nAnother predictive factor of the better knowledge of ODS in our study was maternal age. Age was a good predictive factor in studies from Malaysia,11 Ethiopia,20,21,24,27,28,38 Tanzania,30 Nigeria,29 South Africa,35 and Zambia.37 Women with more advanced ages had better knowledge of ODS. This may be attributed to several hypotheses. Having better ODS knowledge is an essential indicator of pregnancy preparedness. Teenage pregnancy is more unprepared than pregnancy in adulthood.40 Being an adult also means having more mature neurological development, awareness, and experience than being teenagers. Having more experience in older pregnant women leads to gaining both personal experience and information from experience of other people during their pregnancies. This hypothesis could explain our finding that women with multigravida tended to have better ODS knowledge than those with primigravida. Also, higher gravidity was reported a significant predictor in some previous studies.27,33\n\nOur study also found that pregnant women who had medical personnel as a source of ODS knowledge tended to have better ODS knowledge than those who obtained information from women’s friends or other media. Medical personnel are a good source of information for expecting mothers. This could be partially reflected by a lower gestational age at the first antenatal care (ANC) visit and the higher number of ANC visits since these women had more time spent in the clinic, and thus had more opportunities to obtain important information from medical personnel. Although our study cannot demonstrate that these two variables were predictors of better ODS knowledge, many other studies showed the association, e.g., studies in Thailand,10 Congo,33 Saudi Arabia,41 and Ethiopia.16–20,23,38\n\nThe strengths of our study include a large sample size and only one well-trained research assistant collecting the data to minimize the inter-observer variation. Furthermore, data collection setting and process were similar for all participants and were considerably optimal since it took place in a private room during the waiting time in the antenatal care clinic. In addition, our study design was similar to other studies in the same region, allowing comparison of data between countries with similar context.\n\nNevertheless, our study has some limitations. The study was cross-sectional, so we cannot establish a causal relationship between variables. We did not evaluate some variables, i.e., the number of family member and the region of residence, which were reported as significant predictors in previous studies.18,31 Some variables might also affect the knowledge, such as interval between pregnancy, birth preparedness, accessibility to healthcare services, and medical expense subsidies. Further study focusing on the relationship between pregnant woman’s knowledge and her intimate partner’s knowledge should be conducted.\n\n\nConclusion\n\nOur study demonstrates the fair prevalence of mothers with good knowledge of ODS in Thailand’s rural areas. We found that participants’ age of at least 20 years, higher education, and reporting medical personnel as the source of ODS knowledge were predictors of good ODS knowledge. Identifying those at risk for fair and poor ODS knowledge and prompt management for the vulnerable subgroups might help decrease maternal mortality in this region. Further research and educational program are needed to raise the knowledge of ODS, aiming to reduce maternal mortality.", "appendix": "Data availability\n\nHarvard Dataverse: Knowledge of obstetric danger signs and associated factors among pregnant women attending antenatal care services at Thai community hospital datasetEN version, https://doi.org/10.7910/DVN/OEFYS6. 42\n\nThis project contains the following underlying data:\n\n• Knowledge of obstetric danger signs and associated factors among pregnant women attending antenatal care services at Thai community hospital datasetEN version.tab\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nAcknowledgments\n\nWe gratefully acknowledge all participants and the support from their relatives participating in this study. We also want to thank all involved medical and administrative staff at Wang Saphung Hospital for their generous assistance.\n\n\nReferences\n\nToo-Kong T: The Millennium Development Goals Report 2014 United Nations Development Programme. 1st ed.New York: United Nations; 2015; p. 56. Reference Source\n\nUnited Nations: A/RES/71/313: Work of the Statistical Commission pertaining to the 2030 Agenda for Sustainable Development.2017; p. 25.\n\nVogel JP, Pileggi-Castro C, Chandra-Mouli V, et al.: Millennium Development Goal 5 and adolescents: looking back, moving forward. Arch. Dis. Child. 2015 Feb; 100(Suppl 1): S43–S47. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHoyert DL: Maternal Mortality Rates in the United States, 2020. NCHS Health E-Stats. 2022. Reference Source\n\nOnambele L, Ortega-Leon W, Guillen-Aguinaga S, et al.: Maternal Mortality in Africa: Regional Trends (2000-2017). Int. J. Environ. Res. Public Health. 2022 Oct 12; 19(20): 13146. 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Publisher Full Text\n\nPembe AB, Urassa DP, Carlstedt A, et al.: Rural Tanzanian women’s awareness of danger signs of obstetric complications. BMC Pregnancy Childbirth. 2009 Mar 26; 9: 12. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRashad W, Essa R: Women’s Awareness of Danger Signs of Obstetrics Complications. J. Am. Sci. 2010 Jan 1; 66: 1299–1306.\n\nOkour A, Alkhateeb M, Amarin Z: Awareness of danger signs and symptoms of pregnancy complication among women in Jordan. Int. J. Gynaecol. Obstet. 2012 Jul; 118(1): 11–14. PubMed Abstract \| Publisher Full Text\n\nNkamba DM, Wembodinga G, Bernard P, et al.: Awareness of obstetric danger signs among pregnant women in the Democratic Republic of Congo: evidence from a nationwide cross-sectional study. BMC Womens Health. 2021 Feb 26; 21(1): 82. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKabakyenga JK, Östergren PO, Turyakira E, et al.: Knowledge of obstetric danger signs and birth preparedness practices among women in rural Uganda. Reprod. Health. 2011 Nov 16; 8(1): 33. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHoque M, Hoque ME: Knowledge of danger signs for major obstetric complications among pregnant KwaZulu-Natal women: implications for health education. Asia Pac. J. Public Health. 2011 Nov; 23(6): 946–956. PubMed Abstract \| Publisher Full Text\n\nSalem A, Lacour O, Scaringella S, et al.: Cross-sectional survey of knowledge of obstetric danger signs among women in rural Madagascar. BMC Pregnancy Childbirth. 2018 Feb 5; 18(1): 46. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNambala BS, Ngoma C: Knowledge and perception of women towards danger signs in pregnancy in Choma rural district, Zambia. Med. J. Zambia. 2013; 40(2): 43–47.\n\nGeleto A, Chojenta C, Musa A, et al.: WOMEN’s Knowledge of Obstetric Danger signs in Ethiopia (WOMEN’s KODE): a systematic review and meta-analysis. Syst. Rev. 2019 Feb 25; 8(1): 63. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBakar RR, Mmbaga BT, Nielsen BB, et al.: Awareness of Danger Signs during Pregnancy and Post-Delivery Period among Women of Reproductive Age in Unguja Island, Zanzibar: A Qualitative Study. Afr. J. Reprod. Health. 2019 Mar; 23(1): 27–36. PubMed Abstract \| Publisher Full Text\n\nKiataphiwasu N, Kaewkiattikun K: Birth preparedness and complication readiness among pregnant women attending antenatal care at the Faculty of Medicine Vajira Hospital, Thailand. Int. J. Women’s Health. 2018 Dec 5; 10: 797–804. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAbu-shaheen A, Heena H, Nofal A, et al.: Knowledge of obstetric danger signs among Saudi Arabian women. BMC Public Health. 2020 Jun 15; 20: 939. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBumphenkiatikul T, Kaikaew K, Mahoree K, et al.: Knowledge of obstetric danger signs and associated factors among pregnant women attending antenatal care services at Thai community hospital datasetEN version. Harvard Dataverse. 2023; V1. UNF:6:0nW8Q7ser5P7GyrfU2l48Q== [fileUNF]. Publisher Full Text ." }	[ { "id": "194222", "date": "17 Oct 2023", "name": "Jerome Kabakyenga", "expertise": [ "Reviewer Expertise Reproductive Maternal Newborn Child Adolescent Health" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nTitle: Knowledge of obstetric danger signs and associated factors among pregnant women attending antenatal care services at Thai community hospital. The aim of the study was to identify (determine) the prevalence of good ODS knowledge and associated factors among pregnant women attending antenatal services at a Thai community hospital.\nAbstract:\nWell structured and is a summary of the body of the manuscript.\nKey words are missing - obstetric danger signs\nIntroduction:\nparagraph -2 the statement..\"differs among countries, ranging from 23.8 deaths per 100,000 live births in the US to 442 deaths per 100,000 births in Africa\". The authors chose to give a MMR lowest starting with USA - why not start with the country (ies) with the lowest MMR (<5/100,000) e.g. from a publication \"Trends in maternal mortality 2000 to 2020: estimates by WHO, UNICEF, UNFPA, World Bank Group and UNDESA/Population Division\".\nParagraph 4 - there are several statements in this paragraph that need to be supported with references (sentences 2,3,4,5).\nResults:\nIt would be more informative if Table 1 is about the sociodemographic, obstetric data of participants.\nDiscussion:\nIs based on results of the study.\nConclusion:\nIs derived from the results and discussion of the study.\nGeneral:\nThere is need to improve on the grammatical layout of the manuscript.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10421", "date": "29 Nov 2023", "name": "Thanapob Bumphenkiatikul", "role": "Author Response", "response": "Dear Reviewer, We want to express our gratitude for your constructive feedback on our article. Your insights have greatly contributed to the refinement of our work. Regarding the suggestion to change the reference for Maternal Mortality Ratio (MMR) statistics, we have taken your recommendation into account and have made the necessary modifications. The MMR lowest starting point has been updated to reflect the figures from the lowest MMR countries, Australia and New Zealand, as suggested by you. This alteration provides a more accurate and relevant reference point for our study. Furthermore, in response to your comment regarding the need for references to support certain statements in paragraph 4 of the introduction, we have incorporated these references to strengthen the credibility of our work. Lastly, we have carefully reviewed the manuscript for grammatical errors, ensuring that the text is presented with precision and clarity. We genuinely appreciate your efforts in assisting us in making these necessary enhancements. Sincerely," } ] }, { "id": "188939", "date": "17 Oct 2023", "name": "Natnita Mattawanon", "expertise": [ "Reviewer Expertise Clinical OB&GYN" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis study presented data concerning the extent of awareness among pregnant individuals in a rural area of Thailand regarding indicators of potential pregnancy-associated risks, and whether they possess acquaintance with this particular information.\n\nThe investigators employed both questionnaire surveys and interview methodologies to acquire the dataset.\n\nThe result revealed that approximately 10% of the participants exhibited a deficient knowledge of Obstetric Danger Signs (ODS), while an additional 25% attained a moderate level of understanding. Subpopulations characterized by an age below 20 years, limited educational attainment, and reliance on familial or peer networks as their primary information sources demonstrated significantly diminished levels of knowledge. Furthermore, this investigation highlighted the frequent oversight of critical indicators of preeclampsia, such as visual disturbances, epigastric discomfort, and convulsions, by the participants. This underscores the scope for healthcare professionals and governmental bodies to implement precise interventions to improve this situation.\n\nFrom my perspective, this paper offers enhanced insights into the landscape of pregnancy care within rural regions of developing countries. Moreover, it highlights a targeted area for potential development.\n\nThere is, however, one aspect requiring further explanation within the Methodology section.\n\n1. what is the rationale behind establishing the threshold for \"good knowledge\" at 75%?\n\nThe derivation of this specific percentage warrants further clarification.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10420", "date": "29 Nov 2023", "name": "Thanapob Bumphenkiatikul", "role": "Author Response", "response": "Dear Reviewer, We sincerely appreciate your insightful review of our study and the positive feedback provided. Your thorough assessment has contributed to enhancing the quality of our work. In response to your query regarding the rationale behind establishing the threshold for \"good knowledge\" at 75%, we would like to clarify that this particular percentage was derived from a prior study conducted in Thailand. By using this common threshold, we aimed to facilitate a meaningful comparison between our study and the referenced study, enabling a more comprehensive assessment of the health status in the country. This decision was made to ensure that our findings are consistent with existing research and can be applied in a broader context. We hope that this explanation provides the needed clarification on this aspect of our methodology. Thank you once again for your thoughtful review and constructive comments." } ] } ]	1	https://f1000research.com/articles/12-851
https://f1000research.com/articles/12-1020/v1	22 Aug 23	{ "type": "Research Article", "title": "The innovative dimension of the research training programmes under H2020-MSCA-ITN1: a methodological approach to track, measure and analyse innovative aspects and provide policy-feedback conclusions.", "authors": [ "Ioannis Bitsios", "Fabrizio Martone", "Riccardo Ricci", "Audrey Arfi" ], "abstract": "Background: Innovative research training programmes funded by the European Union are essential for the forging of highly skilled researchers to tackle, via breakthrough ideas and solutions, the challenges of our society. Being able to track, measure and analyse innovative aspects of the Marie Sklodowska-Curie Actions, Innovative Training Networks under the Horizon2020 funding scheme enables the impact assessment of such programmes, while filtering best practices and the generated knowledge that could ultimately breed and create further innovation. In parallel, it helps the identification of areas for improvement, the understanding of new needs to be accommodated and the co-design and implementation of EU funding policy activities to further promote innovation and excellence for researchers across Europe and beyond. Methods: In this study, a novel methodological approach is proposed for tracking and analysing innovation, using a representative sample of projects. Basic innovation indicators are examined and considered from the existing literature and from the applicable Multi-Annual Framework Programme Horizon2020. Additional ones are defined, complemented by questionnaires/surveys findings, to capture innovative aspects for which the standard indicators do not apply. Data mining and data visualization tools are used for the collection and processing of data. Innovation Radar2 (IR) reports and HorizonResultsBooster3 services are also engaged for the cross-validation of the identified innovative aspects. Results/Conclusions: The study provides first-level input for policy-feedback activities, by identifying scientific domains and EU countries that may potentially require more attention for innovation generation. It highlights domains that are front-runners and can be used as examples or best practices for under-represented domains in terms of innovative outputs. Collaboration with organisations, defined as medium/high innovators, can increase innovation generation and success in future projects. Best practices are collected to serve as references for designing impactful future training programmes. The excellence of the H2020-MSCA-ITN actions is confirmed via the generated innovations.", "keywords": [ "Innovation", "Policy-feedback", "H2020", "Marie Sklodowska-Curie Actions -Innovative Training Networks (MSCA-ITN)", "Indicators", "Methodology" ], "content": "Introduction\n\nThe H2020 MSCA Innovative Training Networks4 aim to train a new generation of creative, entrepreneurial and innovative Early-Stage Researchers (ESRs), able to face current and future challenges and to convert knowledge and ideas into products and services for the economic and social welfare of society.\n\nH2020-MSCA-ITN has the objective to foster excellence and structure research and doctoral training in Europe (EU Member States and H2020 Associated Countries), extending the traditional academic research training setting, incorporating elements of Open Science5 and equipping researchers with the right combination of research-related and transferable competences. It aims to provide enhanced career prospects in both the academic and non-academic sectors through international, interdisciplinary and intersectoral mobility combined with an innovation-oriented mind-set. The H2020-MSCA-ITN programme encompasses three different implementation modalities: The European Training Networks (ETNs), the European Industrial Doctorates (EIDs), and the European Joint Doctorates (EJDs).\n\nThe H2020-MSCA-ITN calls for proposals were highly competitive, as demonstrated by the high number of applications submitted every year (more than 1500 submitted proposals on average) and the low success rate, ranging from 7-8% to 12% depending on the year and the scientific domain. From 2014 to 2020, 1035 H2020-MSCA-ITN project grants were signed, out of which 78% were ETNs, 15% were EIDs and 7% were EJDs. Funded H2020-MSCA-ITN projects represented a total budget of 3,42 billion EUR, i.e. more than 55% of the total Marie Sklodowska-Curie Actions programme budget and almost 4,4% of the total H2020 framework programme budget. So far, more than 10600 doctoral candidates were recruited and trained within the H2020-MSCA-ITN projects.\n\nAll these networks constitute a potential gold mine of generated innovation, success stories, best and innovative practices.\n\nThe main goals of this research study were: i) to propose a practical methodology for tracking and measuring innovation in H2020-MSCA-ITN projects, which can also be replicated in other similar Research and Innovation (R&I) portfolios; ii) to collect and present in a comprehensive manner the most relevant innovative elements of H2020-MSCA-ITN projects; iii) to provide insights on the action’s performance with regards to innovation in order to draw relevant policy-related conclusions and recommendations.\n\n\nBackground\n\nThe European Union (EU) is constantly supporting the fostering and generation of innovation in order to respond to the challenges of our era. As presented in the recently published, ‘innovation score board’ (European Innovation Scoreboard, 2022), the EU is improving its innovation performance year by year. For example, in 2022, the EU overtook Japan and closed the gap with some other global competitors (including Australia, Canada, South Korea and the United States). Thus, measuring innovation is key for the creation and implementation of research and innovation-friendly policies that can further boost innovation creation within the EU. H2020-MSCA-ITN being the EU’s flagship scheme for establishing transnational doctoral programmes, it is of interest to observe their contribution to the long-term innovation ambitions of the Union. The essential role of the innovation performance measuring is clearly expressed in the ‘Science, Research and Innovation Performance of the EU’ (SRIP) report, as a means for further improvement and design of new R&I policies for the EU’s economic growth and sustainable development (SRIP, 2022). As also reflected in the concept note that kick-started the present study, the MSCA-ITN projects can constitute a potential resourceful place for innovation.\n\nWithout a suitable methodology to track and analyse the potentially generated innovation, a proper project and portfolio analysis is restricted. And most importantly, the measuring of generated innovation becomes difficult which impedes efforts to track the evolution of innovation performance and to provide policy-feedback conclusions. Therefore, being able to monitor innovative performance in EU-funded projects (in this case MSCA-ITN) is very relevant for research-promotion agencies in order to set (Taques, López, Basso, Areal (2020)) and - where appropriate - regularly update funding-targeted criteria.\n\nBeyond the advancement of the current state of the art and the excellence in science in several disciplines (bottom-up approach), the H2020-MSCA-ITN action promotes synergies, collaboration, joint supervision and single or joint doctoral programmes, inter-sectoral and international secondments for enhancing the professional experience and skills of recruited Early-Stage Researchers (ESRs), as well as trainings and joint dissemination and outreach activities. Therefore, when measuring innovative performance in H2020-MSCA-ITN, not only ‘technology’ innovation should be considered. ‘Technology’ innovation is considered tangible (essentially can be quantified and measured) and it is potentially generated as a result of the individual research projects that are undertaken by the recruited researchers (ESRs) as well as from the project consortium as a whole. Other non-tangible innovative aspects (qualitative elements being more difficult to quantify or measure) which are inherent to the very nature of the MSCA-ITN action, are also considered. Such non-tangible innovative elements can be social innovation such as the creation of novel solutions to the needs of the recruited researchers and/or project community, other signature features of the MSCA programme, such as mobility that creates diffusion and transfer of knowledge (Science, Research and Innovation Performance of the EU, 2022), the cross-sectoral cooperation (European research & innovation days, 2021), the quality of the supervision and doctoral training, the implementation of secondments in the most efficient and rewarding way for the highly skilled researchers (Report of Mapping Exercise on Doctoral Training in Europe, 2011) and interesting best practices that MSCA-ITN projects and participants have put in place to promote innovation in doctoral education (LERU advice paper, 2014).\n\nOne of the starting points to track and measure innovation is to define and list the appropriate indicators. In the literature, there are several references to such indicators which may vary depending on the applied domain (e.g. scientific, technological, industrial, commercial etc.). A notable publication is the ‘Oslo Manual’ (OECD, European Union (2018)), which comprises guidelines for collecting, reporting and using data on innovation. Another source of inspiration are the H2020 indicators used to assess the results and impact of the programme (European Commission, Directorate-General for Research and Innovation, 2015). Such indicators include, for example, the number of peer-reviewed publications in high-impact journals, patent applications or awarded ones and less tangible indicators like the cross-country circulation of researchers (for H2020-MSCA-ITN the so-called ‘secondments’).\n\nRegarding tangible indicators, for the currently established metrics for publications, the latest developments on the approach of the European Commission (EC) for research assessment (European Commission, Directorate-General for Research and Innovation, Coalition for Advancing Research Assessment, 2022), alleviate the mere focus on metrics like high-impact journals and instead promote quality and openness to better assess the impact and value of generated publications. As such, in the methodology that is proposed for our analysis, we went beyond a narrow set of quantitative journal- and publication-based metrics. This is in line with the ‘San Francisco Declaration on Research Assessment’ (DORA, 2012), the ‘The Leiden Manifesto for research metrics’ (Bibliometrics: The Leiden Manifesto for research metrics, 2015) and more recently with the Open Science mandate (The EU’s open science policy, 2021). We also considered non-structured or qualitative sources (i.e. information coming from periodic reports, deliverables and other non-tangible innovation elements), in order to capture to the maximum feasible extent, the generated innovation by the H2020-MSCA-ITN projects.\n\nRegarding less tangible indicators like secondments (or mobility of ESRs), a good source of inspiration for the execution of this analysis was a recent survey launched over the summer of 2020 to nearly 17.000 Marie Curie alumni association (MCAA) members, with over 2.000 respondents completing the survey (Marie Curie alumni association survey 2020: results). Based on this survey, following an inquiry about career mobility and, in particular how many times researchers moved due to their career, it was possible to verify that only 3.4% of the respondents didn’t change their country of residence while 96.6% respondents reported to have moved at least once due to their careers. The majority moved (51,4% or 947 respondents) between two and three times. Since mobility is an integral part of MSCA-ITN projects and based on this survey findings, the secondments (or mobility of ESRs) were also considered in this analysis as part of the generation and diffusion cycle of innovative aspects in the MSCA-ITN projects.\n\nAccordingly, there are already several defined indicators, articles, studies, and reports elaborating on innovation. For this study, all the aforementioned sources were considered in order to tailor and use the suitable set of indicators (or metrics) and provide a methodology for identifying, measuring and understanding innovation generated by the H2020-MSCA-ITN EU-funded projects. With the proposed methodology, the aim is also to highlight innovative aspects of H20202-MSCA-ITN actions beyond a particular success story or significant project result, but rather in a more systematic approach that can bring more findings to the surface. Subsequently and based on these findings, the methodology would allow drawing important conclusions which can directly support feedback to policy and the institutional decision-making process in general.\n\n\nMethods\n\nWe identified the appropriate population (sample) of H2020-MSCA-ITN projects to be analysed for innovation outputs, and then the nature and type of data to be collected for the analysis as well as the best way to collect the necessary data. Based on the literature analysis, we proposed and defined the appropriate indicators (KPIs and metrics) in order to track, analyse and present in a quantifiable and qualitative manner innovative elements in the H2020-MSCA-ITN projects.\n\nA first analysis was done on the full H2020-MSCA-ITN portfolio covering the calls from 2014-2020. The source that we used to make the first extraction of data was the EC CORDA database (SAP BusinessObjects Business Intelligence, universe: EGRANTS context: H2020). Since the purpose of the study was to track innovation and similar outputs that have been already generated by the projects, it was decided to narrow the analysis on the population of projects under the calls 2014-2017. Projects under H2020-MSCA-ITN have a normal duration of 4 years (48 months) and with the selected sample of projects from the calls 2014-2017, the purpose was to analyse generated results from the already closed or in the closing mode projects (the analysis started in October 2021). In the closed projects, the interim and final reporting had already been conducted (contractual requirement), which ensured that the generated innovations and similar results were encoded in the EC corporate systems (databases) in order to allow their retrieval and further analysis.\n\nWe proceeded to the analysis of the H2020-MSCA-ITN calls 2014-2017 projects, again using the CORDA database/tool. From the extracted population of projects, a further analysis was carried out in order to identify those projects that appeared to have generated or declared innovation. By ‘generated innovation’ we refer to innovation and patents declared by the projects (and encoded in a structured manner in the CORDA database, following the project closure).\n\nFinally, for the calls 2014-2017 projects, an analysis using the defined metrics for tracking and measuring generated innovation, has been performed on a set of data and documents comprising the Horizon Dashboard6 reports, the available H2020-MSCA-ITN project reports and relevant documents (i.e. progress, periodic and final project reports, Expert or Project Officer Assessment reports, ESRs’ questionnaires7 …) for tracking non-tangible innovation, and a set of EC corporate tools and services like databases (CORDA, CORDIS8), the Qlik Sense9 graphical data representation tool, the IR platform and the HRB service.\n\nTo summarize, the analysis of the extracted sample of H2020-MSCA-ITN projects was performed using:\n\n1. The reported patents and innovation outputs from CORDA and Qlik Sense (HorizonDashboard).\n\n2. The Innovation Radar platform and innovation capacity indicators.\n\n3. The use of HRB Service 410 (Module A: publishable reports of R&I project clusters analysis) as a proof of concept of the methodology from external independent experts.\n\n4. The analysis of most frequently found terms (word clouds) which can be retrieved from the generated publications.\n\n5. The non-tangible innovation and best practices results which can be retrieved by the non-structured data from the different project reports and documents.\n\n6. Questionnaires/surveys from the ESRs that participated in those projects.\n\n1. Reported patents and innovation outputs (CORDA and Qlik Sense/HorizonDashboard)\n\nWe used the CORDA database and the Qlik Sense platform tool for the data extraction. The extracted data were in the form of Microsoft Excel@ spreadsheets.\n\nH2020-MSCA-ITN projects have been generated via 7 calls (1 call per year) that have been performed under the seven years of the H2020 MFF (calls 2014-2020). The H2020-MSCA-ITN portfolio comprises 1034 executed (on-going or closed) projects. A focus on the 2014-2017 calls was considered as more relevant in order to ensure generated innovation from the closed or in the closing mode projects.\n\nBased on the 2014-2017 projects portfolio, a data extraction was done from CORDA database in order to retrieve those projects that appear to have generated some kind of innovation or patent. The extraction of this information was done based on declared innovation and patents, using similar indicators that were provided by the data fields and filters of the CORDA database interface. In the data extraction, we included information fields related to - among others - the project number, call-id, mode, acronym, status, number and description of innovations, number and description of patents and number of publications (see Figure 1.a.1).\n\nFrom the extracted data, a filtering of the projects was done based on declared innovation(s) and/or patent(s), resulting in 118 retained projects (MSCA ITN 2014-2017 innovation and patents [Data set], 2023a).\n\n2. The Innovation Radar platform and innovation capacity indicators.\n\nIn order to further validate that the selection of 118 projects is the best and promising candidate for analysing and measuring innovation in the H2020-MSCA-ITN projects (beyond the patents), we used the Qlik Sense platform to extract additional Innovation Radar (IR) information for those projects.\n\nWe have further utilised the medium/high innovator definition of innovation radar and applied to the MSCA selected projects. The goal of the Innovation Radar Initiative is to identify high-potential innovations and innovators in EC-funded research projects, seeking to direct project consortia on the proper steps to reach the market. Its goal is to maximise the results of public funds mobilised on research. Two indicators based on the Innovation Radar data (Innovation Radar, 2018) have been defined: the Innovation Potential Indicator (Figure 2.1), which measures the projects’ innovation development towards commercialisation within the framework programme, and the Innovator Capacity Indicator (Figure 2.2), which captures the innovative capacity of the innovators behind these innovations. The Innovation Potential Indicator incorporates three essential indicators in the innovation development process: the Innovation readiness (related to the technical maturity of the innovations e.g. prototyping, demonstration or testing activities or a feasibility study), the Innovation management (related to the project consortium capability and its commitment to bring innovations to the market, e.g. preparing a business plan or market study, defining the IPR ownership, securing capital investments, engaging end user), the Market potential (related to the demand and supply side of innovations e.g. evaluating the market conditions for successful commercialisation, assessing how products or services satisfy a market sector and a potential customer base or potential barriers from the supply side which could jeopardise the commercial exploitation of innovations). The Innovator Capacity Indicator includes two indicators that capture the capacity of innovators in delivering successful innovations: the Innovator’s ability (related to the ability of organisations in developing innovations within the EC-funded activities e.g. the number of times organisations have been identified as key innovators by the Innovation Radar, the reviewers’ opinions about innovators’ potential and independence in fulfilling the market potential of innovations), the Innovator’s environment (related to the performance of the project in terms of innovations, the overall conditions in the project consortium which an innovator faces e.g. the commitment of relevant partners to exploiting innovations, the presence of organisations that are directly interested in exploiting the innovations, the positive ecosystem to facilitate the knowledge spill-overs between innovators and their environment).\n\nCategories of Innovators\n\nThe Innovation Radar (Innovation Radar, 2015) introduces three categories of innovators: Low, Medium and High Potential innovators. The assignment to a category is based on mean and standard deviation (SD) values of the Innovator Capacity Indicator (ICI) for innovators and uses percentile ranks. Ordering innovators into three different categories based on percentile ranks allows their performance to be compared very clearly with the remaining innovations and innovators in the sample. The percentile rank of an innovation or an innovator is defined as the percentage of innovators in the same sample that obtained a score at the same level or below that of the innovator’s score. In formal terms, the assignment of inventors to three categories is based on the following rules:\n\nWe have extracted from the Innovation Radar database the ICI assessment conducted on all MSCA ITN projects selected for funding from the calls 2014-2015-2016 and 2017, and compared with all project ICI assessment of the rest of H2020 calls from 2014 to 2017.\n\n3. The HRB Service 411 (Module A: publishable reports of R&I project clusters analysis) as a proof of concept of the methodology by external experts.\n\nWe made use of the HRB Service 4 for the cross-validation of the - as they appear at the initial extraction - promising innovative results that were identified in the selected sample of projects. The main objectives of the HRB analysis were to:\n\ni) contribute to the practical methodology for tracking innovation and innovative best practices in MSCA-ITN projects (via external independent experts as a proof of concept of the other components of the proposed methodology),\n\nii) collect and present the most relevant innovative elements and results (R&I results) of the selected population of MSCA-ITN projects,\n\niii) find commonalities among these ‘innovative’ projects to further derive best practises and common paths to successful research and innovation generation,\n\niv) showcase the excellence in research and innovation potential of the networks in specific scientific domain(s) by clustering/classifying the identified R&I outputs in such domains, where possible.\n\nThe HRB analysis was based on the extracted (from CORDA) portfolio of the 118 selected projects.\n\nThe desk analysis and processing of the collected data was assigned to six independent and competent experts contracted by the HRB service. These experts were provided with the necessary data of the 118 projects (innovations and patents), the IR reports/questionnaires which had been already extracted by the IR platform and with additional project-related documents (like reports, deliverables, description of work, etc.) which have been extracted from the EC corporate databases (resulting in thousands of attachments to be considered and analysed for the total of 118 projects). Samples of these input data can be found under “Dataset, metrics and indicators used for Horizon Results Booster analysis” (Bitsios et al., 2023b).\n\nThe HRB analysis first focused on the mapping of projects, based on country of operation, thematic priorities and objectives. This allowed the clustering of projects into a few scientific domains. The scientific domains were determined via an initial analysis of the fields of science provided in the CORDIS database. When this information was not sufficient to assign a given project to one specific scientific domain, further analysis was done on the project’s definition and outputs, to assign every project into one identified domain.\n\nThe second step was to map the nature of the organisations and entities that were engaged in the 118 projects. This mapping was based on criteria like country, activity type, number of participations, consortium size, ‘Innovators’ (based on the IR definition) as project participants and per scientific domain and LERU participation12. This analysis provided key insights on the type and participation of the organisations in the 118 projects and led to very interesting conclusions on the mix of the so-called ‘Innovators’ among the sampled projects participants.\n\nFinally, the definition of a set of indicators/criteria permitted the extensive analysis of quantitative aspects of the clustered projects’ best practices and innovative outputs, as well as their qualitative assessment (HRB report, 2023).\n\nThe following sources were considered for the definition of the criteria and indicators for the processing of the clustered projects’ data:\n\n- reported publications,\n\n- international grants and/or prizes,\n\n- patents, creation of spin-off, evidenced commercial exploitation of results,\n\n- innovations identified by the IR service,\n\n- innovation and novelties declared by the projects (not via the IR platform) in their reporting or in other project relevant documents,\n\n- members of the League of European Research Universities (LERU) network,\n\n- synergies with other H2020-MSCA-ITN projects and/or clustering with projects by similar research area of applications,\n\n- interviewing the projects stakeholders to follow up on the most promising project outputs after project end.\n\nAs already mentioned under the ‘Limitations’, interviewing of the project stakeholders was not required in the end. The variety and amount of data that were collected by the rest of the sources/metrics were more than sufficient for the HRB analysis.\n\nA summary of the set of criteria and indicators that were defined for the quantitative and qualitative assessment of the clustered projects’ best practices and innovative outputs, can be found under “Dataset, metrics and indicators used for Horizon Results Booster analysis” (Bitsios et al., 2023b).\n\n4. The analysis of most frequently found terms (word clouds) which can be retrieved from the generated publications.\n\nThe dataset consisted of the MSCA ITN projects from the call years 2014 to 2017. Similarly, the reported scientific publications for the same projects have been retrieved. We carried out a wordcount on the titles/keywords of the projects, as well as on the titles of scientific publications. The analysis was performed by scientific panels (Chemistry – CHE; Economics – ECO; Engineering and Information Sciences – ENG; Environment and Geosciences – ENV; Life Sciences – LIF; Mathematics – MAT; Physics – PHY; Social Sciences and Humanities – SOC), as the MSCA programme is bottom-up by nature and funds research in all fields of research. Due to their reduced sizes, the ECO and SOC panels were merged in a single panel (ECOSOC) and the same was done with the MAT and PHY panel (MATPHY). The analysis was carried out in R (R Core Team), with word sizes being proportional to their frequency and displayed in word clouds. Some correlation analysis was also performed between the results obtained among funded proposals and those among publications in order to verify if the trends at funded proposal stage (beginning of the project) can be a good predictor of the work done and reflected in the corresponding publications. The 50 most frequent and correlated words are displayed in each figure. Most of the generic terms, considered as noise, were removed from the analysis. The same analysis has also been performed on the subset of the 118 projects flagged for innovation.\n\n5. The non-tangible innovation and best practices results which can be retrieved from the non-structured data in the different project reports and documents.\n\nSeveral indicators and best practices were selected to analyse non-tangible innovation elements in the 2014-2017 ITN projects, such as social innovation, innovative training practices, innovative project management, long-lasting collaborations etc. The type of data that was missing and could not be retrieved automatically from our various databases was further defined. Most of the best practices are also linked to data that can only be collected manually from the projects’ reports and documents. In order to ease the checks and screening of projects, some categories of data were pre-defined for each selected indicator and best practice.\n\n6. Questionnaires/surveys from the ESRs that participated in those projects.\n\nAs an indicator of non-tangible innovative results, the MSCA follow-up questionnaires after the fellowship and two years after the end of the fellowship were examined. Respondents were asked for feedback regarding their training experience and career improvement. A total of 4431 respondents answered to the end of the fellowship questionnaire, while a total of 181 fellows responded to the questionnaire 2 years after the end of the fellowship. The questionnaires were anonymous; therefore, none of the replies can be traced back to a particular individual. The results presented in this document reflect the answers retrieved from these 4431 and 181 survey replies.\n\nPast research on innovation (OECD, European Union, 2018; Taques, López, Basso, Areal, 2020), already proposed a number of indicators for measuring innovation in different application domains. It must be noted that, for every identified indicator there are advantages and disadvantages on its use for measuring innovation. For example, patent databases can be useful when assessing innovation impact for research projects. On the other hand, while the use of questionnaires to track and measure innovative outputs allows identification of the type of innovation generated by a given organisation, it has certain drawbacks. It is prone to subjectivity (there is no control point for the information because the input is given by the same organisation claiming the innovation generation), not all innovation elements are reported due to information confidentiality concerns, and it is also prone to errors (typos, clerical errors) during the completing process (Taques, López, Basso, Areal, 2020).\n\nIn the present study, another limitation concerns the collection of unstructured data, manually retrieved from various project reports and documents. A methodology and pre-defined categories were discussed in order to limit as much as possible the subjectivity and the variability in the data collection exercise.\n\nRegarding the population/sample of the projects that were analysed, we focused only on closed projects (from calls 2014-2017). For closed projects, normally the interim and final reporting has been conducted and the generated innovations and similar results have been encoded in the corporate systems, thus can be further analysed. Consequently, innovations and similar early results from on-going projects were not considered for this analysis.\n\nThe selection of the two main indicators (patents and innovations) for the identification of the sample of projects for the analysis may limit the representation of Social Sciences and Humanities (SSH) – related projects in the final selected sample. This can be considered as expected for two reasons: The generation of patents and similar innovation results are more likely to happen from innovators that are industrial and/or commercial in their nature (HRB report, 2023). Similarly, the use and submission of patents and similar innovations, is traditionally reserved to domains of science and technology, like the STEM (Science, Technology, Engineering, Mathematics) and medicine/biomedicine/drugs disciplines (HRB report, 2023; Reymond, 2020). However, relevant research has shown for example, that an in-depth analysis in patent sources can provide useful information on SSH topics beyond those of mere technological intelligence (Reyomond, 2020).\n\nAs far as the HRB analysis is concerned, among the initially defined sources and metrics, interviews with selected project stakeholders were foreseen. However, interviewing project stakeholders was considered time-consuming while at the same time, the abundance of data collected by the rest of the defined sources made clear that there was no real need for interviews to complement the collected data.\n\nCertain limitations and assumptions apply to the information and outputs presented in this study since the CORDA database and Innovation Radar database are not wholly consistent nor harmonised across projects’ data. As such, the amount and granularity of information available may vary.\n\n\nResults and main findings\n\nAs mentioned earlier, a focus on the 2014-2017 calls was considered as more relevant. The H2020-MSCA-ITN 2014-2017 portfolio comprises 550 executed (on-going or closed) projects (see Figure 1.b.1).\n\nFrom those projects, the vast majority (77%) were ETNs, while the rest were EIDs (16%) and EJDs (7%) (see Figure 1.b.2).\n\nThe distribution of modes in the 2014-2017 projects portfolio is in coherence with the distribution of modes in the total H2020-MSCA-ITN projects population from all calls 2014-2020 (78% were ETNs, 15% were EIDs and 7% were EJDs). Moreover, an analysis of the 2014-2017 population of projects based on the H2020 KPIs, like the number of publications and patents per 10 million euros funding, revealed that, in terms of publications, MSCA-ITN exceeded the established target. As for patents, although this indicator is not applicable to MSCA under the Excellence pillar of H2020, H2020-MSCA-ITN 2014-2017 projects showed a good performance in terms of the ratio of generated patents. (see Figure 1.b.3).\n\nFrom the extracted data, a filtering of the projects was done based on declared innovation(s) and/or patent(s). Out of the 550 analysed projects, 432 declared no generated innovation or patent, while the remaining 118 projects declared the generation of at least one innovation output or patent or both. Since the focus of this study was to measure the innovative dimension under H2020-MSCA-ITN, a deeper analysis was performed on the 118 projects with only innovation or patent output or with both. The analysis showed that out of those 118 projects which declared the generation of at least one innovation or patent output, 93 projects declared at least one generated innovation and no patents, 17 projects declared at least one patent and no innovation, and 8 projects declared at least one innovation and one patent (see Figure 1.b.4). It must be noted that ETN mode contributes to all types of innovation outputs (only inno, only patent, inno and patent).\n\nWe further examined the mode distribution in the 118 projects with the sole purpose of verifying that they can be considered as a good representative sample of the total 2014-2017 population of projects. Indeed, out of these 118 projects, 92 projects (~78%) were ETN, 20 projects (~17%) were EID and the rest, 6 projects (~5%), were EJD (see Figure 1.b.5). This mode distribution among the 118 selected projects is similar and comparable to the mode distribution among the total number of 2014-2017 calls’ projects (see Figure 1.b.2).\n\nIn terms of patent performance based on the project mode, the analysis of 118 projects showed that EIDs included in this sample, tend to report proportionally more patents per EID project (~1 patent/EID project) than ETNs of the same sample (see Figure 1.b.6). This can be an encouraging early finding on the effect of the project mode (EID, more industry-oriented) to the generation of patents.\n\nBased on the findings of this first component of the methodology, we are already in the position to consider the selected 118 projects as a very good and representative sample to measure, analyse and draw conclusions on the innovative aspects of the H2020-MSCA-ITN projects. As already mentioned, additional components were considered in the methodology to further validate the credibility of the selected sample, and these are elaborated in the following paragraphs.\n\na. Innovation Radar analysis\n\nIn order to further confirm that the selection of the 118 projects is the best and the most promising candidate for analysing and measuring innovation in the H2020-MSCA-ITN projects (beyond the patents), we extracted additional Innovation Radar (IR) information for those projects (see Figure 2.b.1).\n\nIt is to be noted here that the extracted information from Qlik Sense platform on generated innovation for the selected 118 projects, is actually restricted to 101 projects for the H2020-MSCA-ITN 2014-2017 calls. This is logical, since 17 projects out of the 118 selected projects (see Figure 1.b.4) have reported only the generation of patents and no innovation outputs (at least in the sense of innovation outputs that are captured via the IR platform).\n\nb. Innovator Capacity assessment, comparison between MSCA and the rest of H2020\n\nWe have extracted from the two Innovation Radar Platform databases (up to April 2018 and from May 2018 onwards) the ICI assessments conducted on those 101 projects coming from MSCA ITN calls 2014-2015-2016-2017 and all other H2020 projects from the same period (calls 2014 to 2017).\n\nWe calculated, according to the Innovation radar methodology, the average ICI and standard deviation of the H2020 selected population and MSCA one. These values computed as Average +/- Standard Deviation, determine the ICI values that define the low, medium, high innovator categories in both population under analysis. These values and the percentage of the organizations in each category are reported and compared in Table 2.b.1.\n\nFrom the comparison, it is clear that the two populations performed equally in terms of innovators’ distribution per innovation category, showing similar percentage of medium to high innovators vs the total innovators. We have to underline that MSCA being under the excellence science pillar, ITN work programmes involve young researchers with less than four years research experience employed in training networks mainly focused on providing the necessary knowledge and expertise to boost the young researchers’ career prospects within an innovation ecosystem, so the fact that MSCA ITN shows the same innovations character of H2020 programmes whose innovation outcome are coming from Research and Innovation Actions (RIA) and Innovation Actions (IA) projects where the main players are senior researchers, scientists employed in academic and non-academic organisations, is a truly remarkable achievement of the ITN actions. However, it could be expected if considering the following analysis of the common innovators in both populations as follow-up of the HRB study.\n\nc. Follow-up of HRB study on H2020 and MSCA ITN common innovator organisation type analysis\n\nFollowing the results of the HRB analysis, conducted on the 118 projects, we have developed on the ‘innovator’ concept, based on the Innovation Radar definition for the 101 projects assessed by Innovation Radar out of the 118. According to the HRB study:\n\n➢ When it comes to beneficiaries, “success breeds success” in that some are significantly more active than others in project participation, which in turn builds upon their capability to be further innovative and successful. Beneficiaries that are in nature industrial, such as companies working in pharmacological development, and identified as innovators are those being the most innovative. Therefore, encouraging cooperation with coordinating innovators can boost project success, while new ways of expertise sharing will support diversity and inclusiveness, as well as success, amongst consortia.\n\nWe have further investigated this important result, analysing the organisation type of those innovators present in 101 out of the 118 projects studied by HRB, in comparison with the entire population of innovators in H2020 projects originated from the same calls of reference. Out of 265 organisations considered as innovators in MSCA ITN, 240 are also innovators in H2020. We have used the ICI values of H2020 to define the innovator categories (low, medium and high) of those common innovators per organisation type; academic (HES, RES, PUB)13 and non-academic (PRC) differentiating within the non-academic sector group between SME and non-SME. The distribution of organisation type is shown in the following Table 2.b.2.\n\nThe pie chart analysis per innovator category (see Figures 2.b.1a, 1b, 1c) shows that the higher the innovators ICI of one organisation, the higher the concentration of SME within a certain innovator category, confirming once more the finding of the HRB study. In the high innovator band, we found 28% of SME while in the medium one 18%, and in the lower one, only 9%. This is clearly visible in the pie charts for each innovators category. This is somehow expected since the high innovation character is linked to high technology readiness level, which is an essential element pursued by organisations who want to be closer to the market valorisation and thus adapt their exploitation plan according to the projects they are in. Non-academic organisations and particularly SMEs are indeed more interested in high innovation that brings them closer to market valorisation. In fact, the lower the ICI and the higher is the percentage of non-profit organisation, namely universities, research and public bodies.\n\nTo highlight this finding, out of the 13 organisations defined as high innovators in both MSCA-ITN and H2020, 9 (69%) are SMEs as shown in Figure 2.b.2. This once more confirmed the interest of the non-academic sector (mainly SME) in being at forefront of the innovation.\n\nThe HRB analysis (HRB report, 2023) generated a number of very interesting results for the portfolio of 118 projects funded by the H2020-MSCA-ITN Programme taking into account four consecutives calls 2014-2015-2016-2017. This section summarises the key results and findings.\n\nThe projects and their outputs were clustered into 15 scientific domains. The clustering of the projects into the scientific domains allowed more efficient identification of commonalities amongst the projects and the comparison of the innovative elements between these scientific domains. These domains and the distribution of projects (in total 118 projects) can be seen in Figure 3.b.1.\n\nThe mapping exercise of the organisations that were engaged in the 118 projects revealed among others very encouraging results in terms of the innovation potential of the involved organisations and the frequency of their participation in the H2020-MSCA-ITN action. The results indicated that 628 organisations participated in the analysed projects and while most of the organisations (almost 73%) were involved in only one analysed project, more than one-fourth (~27%) of the organisations demonstrated more active engagement. In terms of countries’ representation among the organisations involved in the analysed projects, 475 (out of the 628 in total, almost 76%) are located in the EU-14 countries14, 122 in non-EU (~19%), and only 31 (almost 5%) in the EU-13 countries15. From the engaged organisations, 60% of the project participants represent higher education institutions and research organisations (276 and 98), while 238 (38%) are companies from the private sector. The analysis showed that a project consortium is typically built of eight organisations and on average it covers four higher education institutions, three private enterprises, and one research organisation. Of the 171 organisations that joined more than one project, 100 fall under the category of ‘Innovators’ and most of the projects had at least one innovator in their consortium. From the total number of ‘Innovators’ identified in the 118 projects, 77% are located in the EU-14 countries, 3% in the EU-13 countries and 20% in non-EU countries. Similarly, as for the project participants in general, 60% of the ‘Innovators’ represent higher education institutions and research organisations and 39% are companies from the private sector. On average, four ‘Innovators’ participated in one project and in most projects, at least one ‘Innovator’ was involved. Moreover, and regarding ‘Innovators’ amongst the defined (15) scientific domains, they are well represented as participating organisations across all scientific domains. Interestingly, the mapping exercise of the involved organisations showed that the members of the League of European Research Universities (LERU) network are sufficiently represented in most scientific domains (see Figures 3.b.2, b.3, b.4, b.5, b.6, b.7).\n\nConcerning the results for innovative outputs (i.e. patents, publications, innovations from the IR service), the analysis revealed that certain scientific domains are more prone to generating patentable outputs, possibly due to their results having competitive market applications. In terms of publications, all scientific domains have significant output in terms of peer-reviewed articles, some are more prolific in this regard than others. This could suggest possible differences between the domains regarding certain factors, for example the novelty of the research area, the availability or feasibility of research and experimentation, or the internal dynamics of domain research communities. Finally, the innovation output performance as an average number of innovations was identified and was presented, per scientific domain, which can suggest the innovation potential of a domain based on current research challenges etc. (see Figures 3.b.8, b.9, b.10).\n\nFinally, and in terms of the capability of the analysed projects and/or their results, to be transferred elsewhere, to be applied elsewhere or to be able to engage in synergies with other projects (so to be considered as innovative/best practices), the HRB analysis provided remarkably interesting results. For example, projects with large numbers of events, particularly conferences, were found in almost every scientific domain, which can suggest the effort for public outreach of the projects (integral part of the H2020-MSCA-ITN programme) in order to ‘transfer new knowledge to scholarly communities and communicate it to society’, in line with good practice elements for doctoral training (LERU advice paper, 2014). Regarding performed secondments (also an integral part of the H2020-MSCA-ITN action), the qualitative feedback provided by the participating projects in the Innovation Radar questionnaires (as part of the analysed documents), indicated that the projects consider secondments to be important and would like them to be further supported and enabled. These elements can be assumed as part of the non-tangible innovation that can be observed in the H2020-MSCA-ITN projects, where best practices, such as the mobility of the recruited researchers (for example via the secondments), the participation in wide training events, or the cross-sectoral cooperation (via the exchange of researchers in academic and non-academic participants) nurture the co-creation of novel solutions (which are then reported as innovation outputs). The diffusion and transfer of knowledge, and the cross-sectoral cooperation, have been put in place to stimulate innovation creation.\n\nThe detailed results and interesting findings of the full HRB analysis can be found in the published HRB report (HRB report, 2023), which was generated in the context of this study. The HRB analysis led to key conclusions and concrete links to policy-feedback suggestions and recommendations, which are presented in the ‘Conclusions and main recommendations’ section.\n\nThe purpose of this analysis was to assess, using a visual representation, what the most frequent words and topics in MSCA ITN projects are and if some trends identified at the project level (intended research) can be a good predictor of the effective research performed (outputs such as scientific publications).\n\nIt must be noted here that, as it can be observed in Figure 1.b.3 (related to the extraction of the calls 2014-2017 projects), the performance of the H2020-MSCA-ITN programme in terms of publications output was above the average in comparison to the whole H2020 programme. The beneficiaries of those 550 projects from the H2020-MSCA-ITN 2014-2017 calls, have reported (as of December 2021) a total of 8883 scientific publications (Table 4.1).\n\nENG and LIF panels represent almost 60% of the total (Table 4.2). When looking at the scientific productivity of ITN projects, the two largest panels (ENG and LIF) have the highest shares of publications in absolute terms, but the Chemistry (CHE) and Mathematics/Physics (MATPHY) panels are more productive in relative terms (with an average of 24,3 publications/project for MATPHY).\n\nIn the following figure (Figure 4.1) we represent the correlated words frequencies observed both in projects’ titles and keywords and in the respective publications titles, per panel.\n\nIt should be noticed that in most of the scientific panels, some specific words appear more frequently, giving some scientific trends both at project and publication stages. For example, the most frequent words from the projects and linked publications in the Chemistry panel are mainly related to this scientific domain, such as synthesis, materials, drug, organic and molecular chemistry, while projects and linked publications from the Engineering panel focus more on design, model, systems, control and energy aspects. Similarly, the Environment panel shows some good correlation between the projects and linked publications mainly in environment-related keywords, such as modelling, plant, food, water, processes, energy, sustainable and industry. Interestingly, the Economics and Social Sciences panels show more interdisciplinarity with several frequent words related to health, development, care, innovation, policy and technology. Life Sciences panel shows a strong focus on cells, cancer, development, model, disease, drug, treatment and clinical aspects. Most of the frequent correlated words found in projects and linked publications from the Mathematics and Physics panels concern quantum and imaging-related keywords. Our analysis shows that in some specific scientific domains, such as quantum physics or cancer research, this tool seems to be a good predictor of what is effectively achieved and published by the projects.\n\nIn a second phase, the same analysis was performed on the subset of the 118 projects flagged for innovation (Figure 4.2). The results tend to show that the most innovative ITN projects and linked publications focus on models/modelling, cells, design, data, applications, systems, energy, imaging, health, surface and industrial aspects, suggesting a significant emphasis on applied sciences, as also suggested in the HRB portfolio analysis.\n\nIn this exercise, we obtained, using a text mining analysis, a visual representation of potential thematic domains where innovative outputs can be expected, as confirmed by other analyses and indicators defined in our methodology.\n\nIncreasingly, more publications (Rupietta et al., 2018; Bader, Alharbi et al., 2019) raise the importance of measuring and monitoring not only tangible innovation but also more organisational innovation elements, which can also have a crucial impact on social, environmental or policy aspects, as well as on the career development, knowledge transfer and cooperation aspects in an organisation. All these aspects are also considered as “signature” principles for MSCA. Therefore, it was important to analyse these less tangible innovation elements in the ITN projects and investigate if some correlations could be identified with the projects reporting more tangible innovation.\n\nThe selected indicators and best practices are ranging from social innovation, innovative training practices, involvement of SME/industry in the projects to innovative supervision, networking opportunities, long-lasting collaborations, synergies and policy-driven results.\n\nFirst analyses show that for social innovation (Figure 5.1), 22% of all 550 ITN projects report health-related measures, followed by green-related measures (9,3%). Mobility and transport policy, family-friendly conditions or special needs/disability measures are less reported by the projects, compared to health or green-related measures. This could be explained by the recent covid-19 pandemic which impacted most of the H2020 ITN projects, as well as by the Green Deal launched by the European Commission, encouraging the greening of all projects.\n\nInnovative training practices are also a key element of MSCA ITN projects. Interestingly, the Figure 5.2 shows that 28,4% of the projects report transversal training with a long-lasting impact that stimulates innovative thinking and enhances the researchers’ career, through for example entrepreneurial, creative thinking or teamwork aspects. An additional important element is the structural effect and impact of long-lasting training on the participating organisations, reported by 18.1% of the projects. 16.1% of the projects reported some innovation in terms of the training audience and a wider impact, while 11.5% reported innovative open tool for enhanced training, such as e-learning platforms.\n\nAs reported earlier in the study, the participation of identified innovators plays a key role in the innovative potential of an ITN project, a significant share of them being from the non-academic sector (in particular SME/industry). It was therefore interesting to analyse the role and active contribution of SME and industry participants in H2020 ITN projects. The Figure 5.3 confirms that a large number of projects report a strong contribution of their SME and industry participants, in particular in the secondments of researchers, the training offered to the researchers and their supervision. Their involvement in the exploitation of results is also reported by almost 20% of the projects.\n\nNon-tangible innovation in supervision seems to be less reported by the projects (Figure 5.4). It should be noted, however, that some projects showed a strong engagement in improving those crucial aspects in the researchers’ training and described very good practices, such as the pairing of experienced and new supervisors, dedicated training of supervisors, a new feedback system from fellow to supervisor, or an internal network or platform of supervisors to exchange knowledge and best practices. Better results could be expected at the end of H2020 and under Horizon Europe with the recently published MSCA guidelines on supervision16.\n\nNot surprisingly, almost half of the projects report scientific networking opportunities for the ESRs, followed by cross-sectoral networking opportunities and professional networking opportunities, which is one of the objectives of the MSCA ITN programme (see Figure 5.5).\n\nNew and enhanced research results often come from synergies and synergistic effects. Synergies were also analysed in H2020 ITN projects, as another non-tangible innovative indicator. The Figure 5.6 shows that around 15% of the projects report scientific collaboration within the same domain with other ITN projects, while almost 10% of the projects report the organisation of common training or dissemination activities with other ITN projects. Interdisciplinary scientific collaborations (e.g. between ITN projects in different scientific panels) account for almost 8% of the analysed projects.\n\nAlmost 30% of the ITN projects report co-authoring articles amongst the supervisors and their ESRs. 23,2% of the projects also report co-authoring articles amongst ESRs, which is a positive result since joint publications are very much encouraged in all ITN projects. It often demonstrates the relevance of the joint research project and how well the individual research projects, performed by the ESRs, are interconnected. In addition, the Figure 5.7 exhibits that almost 7% of the project report the creation of spin-off companies, while around 5% create strong collaborations with third countries.\n\nIn the context of more and more requests of feedback to policy, another indicator linked to policy-driven results was analysed in ITN projects (see Figure 5.8). A significant share of the projects report policy-driven research, such as generating data for making policy decisions (18,3%), or organising high-impact conferences, clustering events, etc … (13,5%). For 10,9% of the project, the high policy impact is at the level of researcher who produces results that can feed into policy making etc …\n\nInterestingly, if we compare our sample of 118 ITN projects flagged for tangible innovation with the other ITN projects, the Figure 5.9 below shows that the projects already reporting tangible innovation tend to also report more non-tangible innovation and best practices. The results were particularly significant for social innovation, innovative training practices or SME/Industry contribution, networking opportunities for the ESRs and synergies aspects.\n\nThe percentages can be above 100% since projects can report more than one innovation/best practice per indicator.\n\nAs another source of non-tangible innovation, MSCA follow-up questionnaires after the fellowship and two years after the end of the fellowship were examined.\n\nAfter their fellowship, Marie Skłodowska-Curie Actions (MSCA) fellows are invited to complete two short surveys covering several aspects during and after the fellowship, including the impact of the fellowship on their skills development, career and employability. The evaluation questionnaire is completed soon after the end of their fellowship to assess their experience, skills developed and immediate next steps after their MSCA project. The follow-up questionnaire is submitted two years after the fellowship to gather further information on the more mid and long-term impact of the fellowship and the career paths of the fellows.\n\nRegarding the questionnaires at the end of the fellowship, we also compared the respondents’ total answers (4431) with the ones from the 118 ITN projects (1236) who declared the generation of at least one innovation or patent. The comparison of our sample of 118 ITN projects flagged for tangible innovation with the other ITN projects showed very similar results, as demonstrated below.\n\nRecent publications highlighted several important indicators of non-tangible innovation, such as the career development and job satisfaction level of the researchers. These aspects could be analysed from the various questions included in the questionnaires. In particular, 87% of the respondents declared that their fellowship within the ITN has had a very positive impact on their professional development (very good or good impact). The trend of replies for the 118 ITN projects flagged for innovation was very similar to the general one (as shown in Figure 6.1 A&B).\n\nA: total replies; B: 118 projects flagged for innovation.\n\nWhen asking if the job was research/innovation-related, the majority of the respondents (87%), both from the overall projects sample and the 118 projects flagged for innovation, worked in a research/innovation research job after their fellowship (Figure 6.2).\n\nA: total replies (Yes: 92%; No: 8%); B: 118 projects flagged for innovation (Yes: 93%; No: 7%).\n\nRegarding the specific impact of the ITN fellowships on the researchers’ career, and when asking the fellows if they had found an employment at the end of the fellowship (within the first three months), a high share of fellows (around 42%) had an employment at the end of the fellowship in both samples (total ITN portfolio and 118 projects). In general, the fellows are still finalising their PhD three months after the end of their fellowship, therefore it can explain why almost 50% declared that they did not find an employment yet after their fellowship (see Figure 6.3). One remarkable finding is that among the 118 projects, 6 very best projects have patent and innovation, and the fellow employability is at 100 % (data not shown).\n\nA: total replies (Yes: 42%; No: 48%; No Reply:11%); B: 118 projects flagged for innovation (Yes: 42%; No: 48%; No Reply: 10%).\n\nEven more positive results could be observed when analysing the MSCA evaluation questionnaires two years after the end of the fellowship. In particular, out of the 181 respondents, more than 75% declared that they were employed or self-employed and 12% were still involved in education and training activities, as shown in Figure 6.4.\n\nThe majority of the fellows considered that their MSCA fellowship played an important role in obtaining their current position. 55% of the respondents consider that their MSCA fellowship helped them to a large or very large extent (Figure 6.5).\n\nThese results are very encouraging even for the follow-up questionnaires, two years after the end of the fellowship (181 replies), showing that most of the fellows are satisfied or very satisfied of their experience in an MSCA ITN project and that it significantly helped them improve their professional career, as illustrated by their overall satisfaction with their current professional situation (Figure 6.6).\n\nVery satisfied: 59 (33%); Satisfied: 91 (50%); Dissatisfied: 16 (9%); Very Dissatisfied: 4 (2%); No Reply: 11 (6%).\n\nWhen asking if their work was research/innovation-related, the same number of employed respondents (134) confirmed that their job was research/innovation-related (Figure 6.7).\n\nYes: 74%; No: 11%; No Reply: 15%.\n\nThe use of questionnaires was meant to track and measure less tangible innovative outputs and notably concrete impact on the researchers’ career development through short and simple questions and appropriate indicators. It can be concluded that these types of questionnaires can be a good tool and benchmark for further improvement of MSCA work programme and policy feedback recommendations. They show how the innovative training network can enhance the employability of the fellows, especially in finding research and innovation-related jobs.\n\n\nConclusions and recommendations\n\nThis study is, to the best of our knowledge, the first analysis proposing a methodology for tracking innovation in MSCA-ITN projects, as well as showcasing the most relevant innovative elements of MSCA-ITN projects. In order to define a methodology for tracking innovation in MSCA-ITN projects, we had to identify the appropriate population of H2020-MSCA-ITN projects to be analysed for innovation outputs, to identify the nature and type of data to be collected for the analysis, to propose the method to collect the necessary data, to propose and define the appropriate indicators (KPIs and metrics) in order to track, analyse and present in a quantifiable and qualitative manner innovation elements in the MSCA-ITN projects.\n\n\n\na. In terms of the H2020-MSCA-ITN programme performance for innovation and in comparison, to the full H2020 programme, the results reveal the innovative outputs and potential of the H2020-MSCA-ITN actions and confirm the excellence of the programme while additional interesting insights emerged.\n\nb. Even though the H2020-MSCA-ITN action is mainly a doctoral training programme for recruited researchers, the results obtained on patents and innovations revealed a high performance of the programme in the creation of such outputs. The high ratio of patents and innovation in the analysed projects, in particular in EID projects, suggests a clear added value of the allocated funding, despite the perceived low TRL of the undertaken research and the early career stage of the involved researchers that are the main actors of this research in the scope of the projects.\n\nc. From a closer analysis based on the definition of low, medium and high innovators applied to the common participants in the H2020 and MSCA ITN, it is noted that the percentage of SME and profit organizations is higher in the medium high innovator group in comparison with the lower one, confirming once more, the interest of companies in developing innovation in MSCA ITN projects as much as in H2020 programme. It is essential for the policy makers to continue to attract the non-academic sector into collaborative project consortia, to ensure successful innovation outcomes and impact.\n\nd. Patents as an indicator of potential innovation showed that certain scientific domains (e.g., pharmacology, nanoscience, physics) encompassing ‘innovation intensive industries’ are more predisposed to produce patentable results. Organisations identified as innovators and industrial and/or commercial in their nature, are inter alia more likely to have better means in terms of experience and finances, and therefore may also be more likely to produce patents.\n\ne. The clustering of the 118 projects in certain (15) scientific domains and the analysis of their innovation potential (i.e. patents, publications, innovations) indicated that certain scientific domains are more prone to generating IP-related outputs (i.e., patents). This result should be taken with caution since certain domains are more likely by their nature to generate a high number of patents.\n\nf. The use of word clouds, provides a visual representation of the thematic domains that are treated in the H2020-MSCA-ITN projects (based on the generated publications). This input, if complemented by an in-depth analysis, can provide interesting insights like for example, identifying domains that are less represented in terms of literature outputs and which could be further supported via future policies. Another potential field of applications can be the identification of domains that, even if they appear less-represented (in terms of publications), may however exhibit IP-related outputs (i.e. patents) or other innovative outputs, due to the nature of the examined domain (i.e. more prone to exploitation rather to dissemination). Nevertheless, this application would need further investigation to be fully validated.\n\ng. Interestingly, the HRB analysis showed that the distribution of ‘Innovators’ that were involved in the representative sample of the 118 projects (in terms of country participation and activity type) was similar to the distribution of all participating organisations in the same sample. This can suggest that a sufficient mix of innovators in a given project portfolio can predict a considerable innovation output for that portfolio. Moreover, complementing this ‘mix of innovators’ with, for example, the participation of LERU members (depending on the scientific domain) or members of similar associations can further support the generation of innovation.\n\nh. The share of organisations considered as ‘Innovators’ among the identified 118 innovative projects, originating from countries which are defined in the European Innovation Scoreboard 2022 as ‘Emerging Innovators with performance well below the EU average’ is very low (~1.5%), while for those identified as ‘Innovation Leaders’ in the scoreboard, the share is significant (~23%). This indicates that the results of the proposed methodology in the 118 projects are aligned with the country innovation trends in the European Innovation Scoreboard 202\n\ni. The HRB analysis of the 118 innovative projects showed that the majority of the projects presented a large number of events, conferences, etc. (in almost every scientific domain) which confirms the public outreach as an integral part of the H2020-MSCA-ITN programme in order to ‘transfer new knowledge to scholarly communities and communicate it to society’, in line with good practice elements for doctoral training. To a similar end, the secondments have been confirmed as an integral part of the H2020-MSCA-ITN programme (qualitative feedback provided by the participating projects in the Innovation Radar questionnaires) and are considered equally important for the participating organisations and researchers, along the process of generating innovative outputs. These elements constitute an environment favourable to innovation. More specifically, the mobility of the recruited researchers and co-hosting with other researchers (for example via the secondments) nurture the co-creation of novel solutions (which are then reported as innovation outputs across several scientific domains). Similarly, the participation in wide training events or the cross-sectoral cooperation, via the exchange of researchers in academic and non-academic beneficiaries contribute to the diffusion and transfer of knowledge and the cross-sectoral cooperation, which further stimulate innovation creation.\n\nj. The importance to measure and monitor not only tangible innovation but also more organisational innovation elements was demonstrated in our study, highlighting in particular the crucial impact of innovation aspects on social, environmental or policy aspects, as well as on the career development, knowledge transfer and cooperation aspects in an organisation. Interestingly, the present analysis showed that the projects already reporting tangible innovation tend to also report more non-tangible innovation and best practices. The results were particularly significant for social innovation, innovative training practices or SME/Industry contribution, networking opportunities for the ESRs and synergies aspects. The MSCA follow-up questionnaires after the fellowship and two years after the end of the fellowship also showed very positive results and a high satisfaction of the fellows, in particular on the impact of the ITN fellowship on their career perspectives and employability. Indeed, our analysis showed that 87% of the respondents declared that their fellowship within the ITN has had a very positive impact on their professional development. A similar trend was found for the 118 projects. Moreover, a similar result was also reported in the Marie Curie alumni association (MCAA) survey 2020 (Marie Curie alumni association survey 2020: results) where more than 80% of the respondents considered the impact of the MSCA programme in their career development as ‘very useful’ or ‘useful’.\n\nRegarding the validation of our methodology to track, measure and analyse innovative elements in EU-funded projects, we have performed an analysis of the metrics to be considered, based on the Horizon Dashboard reports, various types of H2020 MSCA ITN and various EC databases, focussing on the H2020 KPIs for high-impact publications, patents, allocated funding, innovation outputs, the Innovation Radar indicators, an analysis of most frequently found terms (words clouds), as well as non-tangible innovation and best practices results. The significance of our results validates the need of having a methodology in place (as proposed in this study) to track, measure and analyse innovative aspects and best practices. Measuring innovation is key for the creation and implementation of research and innovation-friendly policies that can further boost innovation creation within the EU, and our study revealed that the H2020-MSCA-ITN funding scheme has a clear and important contribution to this process.\n\nA set of indicators and a methodology have been proposed in this study to allow the extraction of innovative aspects (tangible and non-tangible innovation) in EU-funded projects to support policy-feedback activities. These elements can serve as a reference and can be re-used with small adaptations to other EU-funded programmes to support similar activities. The exercise for the extraction and analysis of the 118 projects confirmed the validity of the defined indicators as well as of the components of the proposed methodology, since coherent results and findings were observed across the components of the methodology. These indicators can in turn act as predictors of potential success and innovation of ITN/DN projects or other EU-funded projects.\n\nBeyond the excellence of the results and the high performance of the programme towards innovation generation, certain points for further improvement are identified (i.e. countries representation, scientific domains currently being less-represented in the innovation generation), which can feed EU-funding policy activities to improve innovation and excellence across Europe and beyond.\n\na. The inclusion of ‘Innovators’ in a project consortium can act as a predictor for innovation generation. As highlighted in the HRB report, for the organisations involved in the projects, “success breeds success”. This practically means that the inclusion of significantly active organisations in a project can further build upon their capability to be even more innovative and successful, while striving to maintain a right balance with newcomers. Moreover, industrial-relevant organisations, such as companies working in commercially competitive domains (i.e. pharmacological development), and which are identified as ‘Innovators’, are those being the most innovative. Therefore, by encouraging cooperation with such innovators, project consortia can boost their success, share their expertise, their diversity and inclusiveness, and eventually their success in the generation of innovative outputs. If the ‘Innovator’ is also assigned as project coordinator, this can be an additional key success factor for realising innovation.\n\nb. There are innovation front-runners either in terms of scientific domains or in terms of participating organisations or countries, which can be further examined, exploited or targeted by future policy-related activities, with the aim to boost innovation generation in less performing scientific domains or countries or type of organisations.\n\nc. The share of organisations from Widening countries17 among the innovative projects is significantly lower than the share of other Member States. Beneficiaries from Widening countries are thus significantly under-represented, indicating a relative lack of active engagement in MSCA-ITN projects. It should also be noted that only nine of the innovators and none from the LERU members were from Widening countries. One explanation is that the analysis was based on the first part of H2020 projects (calls 2014-2017) and experience showed that it usually takes more time within a Framework Programme to see mid- to long- term effects of new policies, such as the H2020 goal of inclusiveness, applied to enhance participation from these countries. Involving companies and higher education institutions from these countries in the aforementioned domain of expertise, associated with the identified innovators, should therefore be encouraged, as well as finding ways to attract and cooperate with participants from these countries in consortia with high chances of success. More targeted information days, better training by the National Contact Points, better awareness of the partners search tool on the Funding and Tenders Portal could also be envisaged.\n\nd. The analysis of the fellows’ questionnaires at the end of the fellowship proved to be a very interesting source of non-tangible information. It showed how the MSCA-ITN programme contributes to the employability and career development of fellows. However, the fellows’ questionnaires 2 years after the fellowship received very low attention from candidate respondents. It remains challenging to increase the response rate. Several measures could be examined to encourage the fellows’ replies to this final survey, like the use of closed-ended questions or the automatic prefilling of project-related administrative data (i.e. year, call etc.), or automatic notification to the fellow’s last known email address. These simple changes would greatly facilitate the data analysis and the accuracy of the inputs received. Coordinators could also be asked to ensure the submission of these questionnaires in due time. Additional questionnaires, targeting the coordinators could also be envisaged. This non-tangible feedback will be key to develop further policy feedback inputs.\n\ne. In addition, the processing of project-related documents (i.e. reports, deliverables etc.) in the HRB analysis, revealed the need for further harmonisation of the information provided by the projects via the EC corporate IT tools for reporting under the current or in future funding programmes. This can be achieved by providing fixed drop-down menus for the projects to select which outputs they are reporting on. This would also help to ensure that different types of outputs receive equal amount of attention (e.g., patents and non-scientific outreach activities). The HRB report proposed specific metrics for further consideration (see “Dataset, metrics and indicators used for Horizon Results Booster analysis”).\n\nf. In the present study, the data was collected and processed by mainly using the standard EC corporate IT tools (i.e. CORDA, Innovation Radar platform and HorizonDashboard). This was done on purpose since these tools do not require specific IT competency/expertise, and our main goal was to keep the proposed methodology simple and user-friendly. When it comes to the processing of unstructured data (i.e. originated by non-tangible innovation outputs), the use of more advanced tools (even AI-related) must be considered to limit manual and time-consuming processing, involvement of external actors/experts etc.", "appendix": "Data availability\n\nZenodo: MSCA ITN 2014-2017 innovation and patents. https://doi.org/10.5281/zenodo.8046333 (Bitsios et al., 2023a).\n\nThis project contains the following underlying data:\n\n‐ Innovation_and_patents_2014_2017 public.pdf\n\nZenodo: Dataset, metrics and indicators used for Horizon Results Booster analysis. https://doi.org/10.5281/zenodo.8081996 (Bitsios et al., 2023b).\n\nThis project contains the following extended data:\n\n‐ Criteria and indicators used in the HRD analysis.pdf\n\n‐ HRB list of projects.xlsx\n\n‐ HRB proposed metrics.pdf\n\n‐ Project-related documents.pdf\n\n‐ Sample data from IR.pdf\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nWe are thankful to Sohail Luka and Marija Mitic (DG EAC.C2), David Pina (REA.B4) and Roberto Rodriguez Rodriguez (REA.D2) for their contribution and suggestions to this study.\n\nWe are grateful to our: Deputy Head of Unit Frank Marx and Head of Unit Klaus Haupt (REA.A1), Head of Department (REA.A) Begona Arano and Director of REA Marc Tachelet, for their trust and constant support to our work for the realisation of this study.\n\nWe are grateful to the group of Project Officers of Unit REA.A1 that volunteered for the collection of the non-tangible innovation data, as well as to the six independent experts that contributed to the processing of the data.\n\nWe are thankful to the DG RTD.GH6 main correspondents (Bilgeyis Najafova and Athina Pantelidou), for their liaison with the HRB service and the DG RTD corporate databases officers.\n\n\nReferences\n\nBader A, Alharbi I, et al.: Organizational Innovation: A review paper.2019. Publisher Full Text\n\nBitsios, Martone, Ricci, et al.: MSCA ITN 2014-2017 innovation and patents. [Data set]. Zenodo. 2023a. Publisher Full Text\n\nBitsios, Martone, Ricci, et al.: Dataset, metrics and indicators used for Horizon Results Booster analysis. [Data set]. Zenodo. 2023b. Publisher Full Text\n\nDe Prato G, Nepelski D, Piroli G: Innovation Radar: Identifying Innovations and Innovators with High Potential in ICT FP7, CIP & H2020 Projects. JRC Scientific and Policy Reports – EUR 27314 EN. Seville: JRC-IPTS; 2015. Reference Source\n\nEuropean Commission, Directorate-General for Research and Innovation: Horizon 2020 indicators: assessing the results and impact of Horizon, Publications Office.2015. Publisher Full Text\n\nEuropean Commission: 2022 Directorate-General for Research and Innovation, Agreement on Reforming Research Assessment. Coalition for Advancing Research Assessment. Reference SourceReference Source\n\nEuropean Commission: Directorate-General for Education, Youth, Sport and Culture, Marie Skłodowska-Curie actions: Marie Curie alumni association survey 2020: results, Publications Office of the European Union.2022. Publisher Full Text\n\nEuropean Commission: Directorate-General for Research and Innovation, Horizon Europe, open science: early knowledge and data sharing, and open collaboration, Publications Office of the European Union.2021. Publisher Full Text\n\nEuropean Commission, Directorate-General for Research and InnovationHoll H, Es-Sadki N, et al.: European Innovation Scoreboard 2022. Publications Office of the European Union; 2022. Publisher Full Text\n\nEuropean Commission: Directorate-General for Research and Innovation, European research & innovation days: conference report: 23-24 June 2021. Publications Office; 2021. Publisher Full Text\n\nEuropean Commission: Directorate-General for Research and Innovation, Science, research and innovation performance of the EU 2022: building a sustainable future in uncertain times. Publications Office of the European Union; 2022. Publisher Full Text\n\nEuropean Commission, European Research Executive Agency: Vilma Kuuliala V, Hoya C, Spudyte I, et al., editors. Report on R&I project cluster analysis: H2020 MSCA ITN 2014-2015-2016-2017. Publications Office of the European Union; 2023. Publisher Full Text\n\nGood Practice Elements in Doctoral Training: Advice Paper no.15 January.2014. Reference Source\n\nHicks D, Wouters P, Waltman L, et al.: Bibliometrics: The Leiden Manifesto for research metrics. Nature. 2015; 520: 429–431. PubMed Abstract \| Publisher Full Text\n\nNepelski D, Van Roy V: Innovation Radar Identifying the maturity of innovations in EU-funded research and innovation projects.2018. Publisher Full Text\n\nOECD/Eurostat: Oslo Manual 2018: Guidelines for Collecting, Reporting and Using Data on Innovation. The Measurement of Scientific, Technological and Innovation Activities. 4th ed.Paris/Eurostat, Luxembourg: OECD Publishing; 2018. Publisher Full Text\n\nReport of Mapping Exercise on Doctoral Training in Europe Towards a common approach: (final), adopted by the ERA Steering Group on Human Resources and Mobility.27 June 2011. Reference Source\n\nReymond D: Patents information for humanities research: Could there be something? Iberoamerican Journal of Science Measurement and Communication. 2020; 1(1): 006. Publisher Full Text\n\nRupietta C, et al.: Organisational innovation across the technological innovation process: The moderating influence of hybrid change agents. Corpus ID: 169385248. 2018. Reference Source\n\nSan Francisco Declaration on Research Assessment. Reference Source\n\nTaques FH, López MG, Basso LF, et al.: Indicators used to measure service innovation and manufacturing innovation. J. Innov. Knowl. 2020; 6(2021): 11–26. Publisher Full Text\n\n\nFootnotes\n\n1 https://marie-sklodowska-curie-actions.ec.europa.eu/\n\n2 https://www.innoradar.eu/\n\n3 https://www.horizonresultsbooster.eu/\n\n4 https://ec.europa.eu/research/participants/data/ref/h2020/wp/2014_2015/main/h2020-wp1415-msca_en.pdf\n\nhttps://ec.europa.eu/research/participants/data/ref/h2020/wp/2016_2017/main/h2020-wp1617-msca_en.pdf\n\n5 https://research-and-innovation.ec.europa.eu/strategy/strategy-2020-2024/our-digital-future/open-science_en\n\n6 https://ec.europa.eu/info/funding-tenders/opportunities/portal/screen/opportunities/horizon-dashboard\n\n7 https://ec.europa.eu/eusurvey/runner/Evaluation_for_MSC_fellows\n\n8 https://cordis.europa.eu/\n\n9 https://webgate.ec.europa.eu/dashboard/sense/app/a976d168-2023-41d8-acec-e77640154726/sheet/0c8af38b-b73c-4da2-ba41-73ea34ab7ac4/state/0\n\n10 https://webgate.ec.europa.eu/fpfis/wikis/pages/viewpage.action?spaceKey=DIEPP&title=Horizon+Results+Booster\n\n11 https://webgate.ec.europa.eu/fpfis/wikis/pages/viewpage.action?spaceKey=DIEPP&title=Horizon+Results+Booster\n\n12 https://www.leru.org/\n\n13 Higher and secondary education institutes (HES), Research organisations (REC), Public body (PUB), Private for profit (excluding education) (PRC), Small Medium Enterprise (SME)\n\n14 Austria, Belgium, Denmark, Finland, France, Germany, Greece, Republic of Ireland, Italy, Luxembourg, the Netherlands, Portugal, Spain, and Sweden.\n\n15 Bulgaria, Croatia, Cyprus, Czech Republic, Estonia, Hungary, Latvia, Lithuania, Malta, Poland, Romania, Slovakia, and Slovenia.\n\n16 https://op.europa.eu/en/publication-detail/-/publication/bb02d56e-9b3c-11eb-b85c-01aa75ed71a1\n\n17 Bulgaria, Croatia, Cyprus, Czech Republic, Estonia, Hungary, Latvia, Lithuania, Malta, Poland, Romania, Slovakia, and Slovenia." }	[ { "id": "202223", "date": "20 Sep 2023", "name": "Costis Kompis", "expertise": [ "Reviewer Expertise Technology commercialisation from R&D", "integrating science & engineering with innovation management. Technology domains span Cyber Physical Systems", "Data Analytics", "Machine Learning", "Energy Harvesting", "Virtual Reality", "Medical Imaging", "Fuel Cells and Additive Manufacturing." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis a well-structured, erudite and well-informed article. Its methodology and analysis benefit from rich data sets held in EC databases. The methodology is practical and appropriately leverages the available data. There are references to recent relevant works (including HRB report), which set strong foundations. Granular descriptions of the indicators are given. The conclusions are meaningful and can certainly have implications in the design and execution of future programmes for training new generations of researchers and entrepreneurs.\n\nBelow, I append the issues - organised per the article's sections - that I identified along with possible reformulations and other suggestions that the authors can take into consideration in order to improve clarity and consistency.\n\nAbstract:\n“for the forging of highly skilled” => for forging highly skilled.\n\nexamined and considered => considered and examined.\n\nquestionnaires/surveys findings => questionnaires/surveys’ findings.\n\nIntroduction:\n\"The low success rate\": “low” is relative. For other competitions 7-8% might be considered “high”. I suggest to simply replace “the low” with “a”.\n\n“All these networks constitute a potential gold mine” => “The set of these networks potentially represents a gold mine”.\n\n\"The action’s performance\": please improve clarity to what “action” exactly refers to here. Perhaps just say H2020-MSCA-ITN or should it be introduced once and referred with a capital A throughout the article?\n\nBackground:\n\"H2020-MSCA-ITN being the EU’s flagship scheme for establishing transnational doctoral programmes, it is of interest [...]\" => \"H2020-MSCA-ITN, being the EU’s flagship scheme for establishing transnational doctoral programmes, is of interest [...].\"\n\n“As also reflected in the concept note that kickstarted the present study, the MSCA-ITN projects can constitute a potential resourceful place for innovation.”: This statement does not seem necessary and could be dropped.\n\n“‘technology’ innovation” => technological innovation (there are additional references of this term in the rest of the article that may need to be edited should the change be agreeable).\n\n“provide a methodology” => ”propose the methodology”.\n\nMethods:\n“proposed and defined the appropriate indicators (KPIs and metrics)”: how is “appropriate” qualified here? Perhaps replace it with ‘suitable’ meaning that they are fit for the purpose of measuring innovation in such type of EU-funded projects.\n\n“appear to have generated some kind of innovation or patent”: Consider if the wording can be improved, e.g. “self-declare the generation of innovations, including the filing of patents.”\n\n“the best and promising candidate”: replace simply with something like that “the sample was suitable”?\n\n“applied to the MSCA” => “applied it to the MSCA”.\n\n“Two indicators based on the Innovation Radar data (Innovation Radar, 2018) have been defined”: it is not clear if it is the authors of the Innovation Radar or yourselves who defined the indicators from this statement. If it is you, please make it more explicit, i.e. “Two indicators defined from the Innovation Radar data (Innovation Radar, 2018) have been selected as more suitable for our analysis:”\n\n“In formal terms,” => “Specifically,” or ‘Typically,’?\n\n“We have extracted” => \"We extracted\".\n\n“These experts were provided [...]”: this sentence appears fragmented with a bit of repetition “which had been already extracted” and “which have been extracted”. Please consider linking these explanations to make it more concise. For example, “These experts were provided with the necessary data from the 118 projects (innovations and patents), the IR reports/questionnaires (extracted by the IR platform) and with additional project-related documents, like reports, deliverables, description of work, etc. (extracted from the EC corporate databases), resulting in thousands of documents to be considered and analysed for the total of 118 projects.”\n\n“follow-up questionnaires after the fellowship” => “follow-up questionnaires immediately after the fellowship” (in order to make a distinction with the “and two years after the end of the fellowship” that follows?).\n\n\"to errors (typos, clerical errors)\" => \"to typos or clerical errors\".\n\n\"A methodology and pre-defined categories were discussed in order to limit as much as possible the subjectivity and the variability in the data collection exercise.\" => \"Our methodology and introduction of the pre-defined categories aimed at reducing the subjectivity and the variability in the data analysis phase.\"\n\nThis can be considered as expected for two reasons: The generation of patents: can use i) and ii) to make it more clear.\n\n\"may vary.\" => \"exhibited variability.\"\n\n\"as more relevant.\" => \"more relevant.\"\n\n“It must be noted” => \"Notably\"\n\n“This can be an encouraging early finding on the effect of the project mode (EID, more industry-oriented) to the generation of patents.”: Not sure about what “encouraging” implies here. Better stay factual saying something like “This is consistent with the expectation that EIDs which are more industry-oriented prioritise the generation of patents.\"\n\n“same innovations character”: reconsider the wording here. Perhaps “similar capacity to generate innovation”?\n\n“However, it could be expected if considering the following analysis of the common innovators in both populations as follow-up of the HRB study.”: This sentence could be dropped, not least because it creates an anti-climax after “truly remarkable achievement”.\n\n\"the higher the innovators ICI\" => \"the higher the innovators’ ICI\" (apostrophe was missing).\n\n“This is clearly visible in the pie charts for each innovators category.” => \"as shown in the relevant Figures\".\n\n\"This is somehow expected\" => \"This is not surprising\".\n\n\"innovation character\" => \"innovator character/type\" (to comply with the terminology used by the Innovation Radar).\n\n\"ICI and the higher is the percentage of non-profit organisation\" => \"ICI, the higher is the percentage of non-profit organisations\".\n\n“some are more prolific in this regard than others.\" => “Some are more prolific in this regard than others.”\n\n“Finally, and\": could be omitted.\n\n\"(as part of the analysed documents)\": could be omitted.\n\n\"the participation of identified innovators plays\" => \"the participation of identified 'Innovators' plays\".\n\n“In the context of more and more requests of feedback to policy, “: Is this necessary? If yes, it’d be better to explain who makes the requests etc.\n\n\"For 10,9% of the project\" => \"For 10,9% of the projects\".\n\n“ etc …”: omit?\n\n\"When asking\" => \"When questioned\".\n\nFigure 6.3. Employment at the end of the fellowship. A: total replies (Yes: 42%; No: 48%; No Reply:11%): Check the rounding as the answers add to 101%.\n\n\"therefore it\" => \"which\".\n\n“6 very best projects”: please use an alternative descriptor. Perhaps “top performing in terms of innovation and patenting”.\n\n\"Even more positive results could be observed when analysing the MSCA evaluation questionnaires two years after the end of the fellowship.\" => \"The reported answers are more positive in the MSCA evaluation questionnaires two years after the end of the fellowship.\"\n\nFigure 6.7. Is the ESRs’ job research/innovation-related 2 years after the end of the fellowship? Yes: 74%; No: 11%; No Reply: 15%. => Whether the ESR’s job is research/innovation-related 2 years after the end of the fellowship. (Yes: 74%; No: 11%; No Reply: 15%).\n\n\"we had to identify the appropriate population\" => \"we had to identify a representative population\".\n\n“in the MSCA-ITN projects.” (end of par on. p29): can be omitted.\n\n\"the results reveal the\" => \"the results revealed the\" (or better “the results revealed the extent”).\n\n“while additional interesting insights emerged”: can be omitted.\n\n\"Revealed a high performance of the programme\" => \"Revealed a high performance for the programme\".\n\n“the perceived low TRL”: “typically lower TRL”? (you may have to expand the acronym or add a footnote for any reader not familiar with TRLs).\n\n“to ensure successful innovation outcomes”: not sure about “successful” in this sentence. Perhaps “to inspire/catalyse innovation outcomes”.\n\nConclusions:\n“to generate a high number of patents.”: or perhaps rephrase it that inventions in some domains are more patentable than others.\n\n“This can suggest that a sufficient mix of innovators in a given project portfolio can predict a considerable innovation output for that portfolio.”: possibly but hard to prove causality as such. In any case I would suggest reformulation to avoid vague descriptors such as “sufficient mix”. Perhaps instead you want to mention a “minimum representation of proven “Innovators””.\n\n“European Innovation Scoreboard 202”: something missing at the end.\n\n“nurture” => “tend to facilitate”?\n\n“which further stimulate innovation creation” => \"which may further stimulate innovation.\"\n\n\"The importance to measure and monitor\" => \"The importance of measuring and monitoring\".\n\nbreak the first sentence (e.g. put a full stop after “in our study”) to improve readability.\n\n\"synergies aspects\" => \"synergies\".\n\n\"after the fellowship and two years after the end of the fellowship\" => \"immediately after the fellowships and two years after the end of the fellowships\".\n\n\"Indeed\" => \"Specifically\".\n\n\"A similar trend was found for the 118 projects. Moreover, a similar result was also reported in the Marie Curie alumni\" => \"A similar trend was observed across all 118 projects and this is in line the Marie Curie alumni\".\n\n“since coherent results and findings were observed across the components of the methodology.” => \"due to the coherence of results and findings collected via the different components\".\n\n“These indicators can in turn act as predictors of potential success and innovation of ITN/DN projects or other EU-funded projects.“: Unclear why the term “predictors” is used here in reference to “a set of indicators and a methodology”. Perhaps you can list separate indicators that lend themselves to such predicting capability. One of them can be the number of innovators participating in a given project.\n\n“a right balance”: not obvious to what that might be. If there is a proven mix, please indicate it even as a percentage. If not, simply replace with “balance”.\n\n“innovation front-runners” => “front-runners in innovation generation”?\n\n\"could also be envisaged.\" => \"could be encouraged.\"?\n\nGeneral/consistency issues:\nFor consistency, please use EU-funded (there are several instances of EC-funded too).\n\nInnovation Radar/IR: term introduced but acronym not used consistently thereafter.\n\n“MSCA ITN projects from the call years 2014 to 2017” vs “2014-2017 ITN Projects” (e.g. p 9).\n\n“Marie Skłodowska-Curie Actions (MSCA)” appears on p. 26. Shouldn’t be in first occurrence?\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10349", "date": "06 Oct 2023", "name": "IOANNIS BITSIOS", "role": "Author Response", "response": "Dear Reviewer, thank you very much for your useful feedback and pertinent comments. We addressed most of your suggestions. An updated version of the article will be soon made available. Best regards, Ioannis, Fabrizio, Riccardo and Audrey" } ] }, { "id": "202218", "date": "26 Sep 2023", "name": "Nikolaos D. Alexopoulos", "expertise": [ "Reviewer Expertise innovation", "materials science", "engineering", "economics and business" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIn this study, an innovative methodological approach is proposed for tracking and analyzing innovation and representative indicators were considered from the applicable Multi-Annual Framework Programme Horizon 2020. The article provides first-level input for policy-feedback activities, identifies the scientific domains and the EU countries that urge for innovation generation. The writing style is appropriate, Figures and Tables support the conclusions and are appropriate.\nThe manuscript is worth indexing and I recommend it being accepted after some minor amendments:\nThe Abstract should be essentially shortened. Especially for background and methods, only the basic information should be reported in this section. The focal point should be the results/conclusions of the present study.\n\nCiting style should be re-considered; there are some entries as footnotes (starting from no4); they should be considered as regular citations and follow the style approved by the journal.\n\nIt is recommended (not mandatory) that the authors to provide a Figure just like a process flow diagram that briefly shows the logical path and the sequence of the intermediate steps of their study.\n\nFigures: The figures should be numbered in sequence, 1, 2, 3 as it is extremely inconvenient for the reader to follow Figure e.g., 1.b.1 or 5.1 or 3.b.10.\n\nThe captions in the figures should be more descriptive, i.e., it should describe the Figure to the reader.\n\nThe conclusions should be essentially shortened; there are ten (10) numbered conclusions in the manuscript; each numbered conclusion is almost a paragraph long and this is not friendly for the readers. The conclusions should be shortened to simply report the key results of the manuscript. The discussion of the key factors/results should be transferred to the discussion section of the manuscript.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10347", "date": "06 Oct 2023", "name": "IOANNIS BITSIOS", "role": "Author Response", "response": "Dear Reviewer, thank you very much for your useful feedback and pertinent comments. We addressed most of your suggestions. An updated version of the article will be soon made available. Best regards, Ioannis, Fabrizio, Riccardo and Audrey" } ] }, { "id": "202225", "date": "28 Sep 2023", "name": "Alessandro Marino", "expertise": [ "Reviewer Expertise Automation and Robotics. Computer Science Engineering." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nFirst and foremost, I would like to express my very positive opinion about the initiative. As reported by the authors, monitoring the results of the project, particularly within a program as extensive as the H2020-MSCA-ITN program, is noteworthy and of the utmost importance. This step is fundamental for highlighting weaknesses and strengths of the aforementioned program and for taking corrective actions if needed. It is also remarkable that both tangible and non-tangible outputs are considered, as the former often fail to capture many relevant features of the phenomena under study, as highlighted in the literature.\nFurthermore, another fundamental contribution of the proposed approach is that the underlying methodology might also apply to other programs.\nFinally, it is very positive that the MSCA program has performed very well and has fulfilled almost all of its objectives.\nHaving said that, here are a few comments:\nAfter the introduction of background and method, instead of using items and standard text format, a table summarizing the main features and indicators considered and the corresponding database would further enhance the document's readability.\n\nThe identified scientific domains make complete sense, though more details might be provided on how they were identified.\n\nThe comparison between MSCA and the remaining H2020 programs is remarkable. The reader is wondering if RIA and IA could be considered separately in future comparisons given the different research components present in the two typologies, the different TRL targets, and often the different presence of industries in the Consortia. Does it make sense to compare MSCA-EID to H2020-IA and other MSCA programs with H2020-RIA?\n\nRegarding produced publications, they might be classified in future studies based on the ranking of the corresponding journal/conference. For example, you could use the SJR ranking for journals and conference ranking or acceptance rate for conference papers, as well as citations.\n\nThe article appropriately highlights the limitations of the analysis due to the available data and their format and makes useful recommendations at the end of the document. I believe that a potential follow-up to the present study might involve a deeper investigation into what data (especially for non-tangible outputs) are worth gathering in addition to the ones already collected and how they can be gathered to ease the processing phase.\n\nRegarding footnotes: Links 9-11 are not accessible in my case.\n\nIn summary, I would strongly support the indexing of the given document since it presents elements of novelty in the evaluation of complex programs and serves as an example of good practice.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10348", "date": "06 Oct 2023", "name": "IOANNIS BITSIOS", "role": "Author Response", "response": "Dear Reviewer, thank you very much for your useful feedback and pertinent comments. We addressed most of your suggestions. An updated version of the article will be soon made available. Best regards, Ioannis, Fabrizio, Riccardo and Audrey" } ] } ]	1	https://f1000research.com/articles/12-1020
https://f1000research.com/articles/12-420/v1	19 Apr 23	{ "type": "Research Article", "title": "Broodstock development, induced spawning and larval rearing of the bilih, Mystacoleucus padangensis (Bleeker, 1852), a vulnerable species, and its potential as a new aquaculture candidate", "authors": [ "Hafrijal Syandri", "Azrita Undefined", "Rinold Thamrin", "Deni Zen", "Hendrik D. Roza", "Jimmy Chandra Eduard Orah", "Maman Abdurahman", "Alif Yuza", "Irvan Irvan", "Afriwan Afriwan", "Hafrijal Syandri", "Rinold Thamrin", "Deni Zen", "Hendrik D. Roza", "Jimmy Chandra Eduard Orah", "Maman Abdurahman", "Alif Yuza", "Irvan Irvan", "Afriwan Afriwan" ], "abstract": "Background: Mystacoleucus padangensis living in Lake Singkarak, Indonesia, has high potential market demand but is threatened by overfishing and has not been successfully cultured. This study describes the first broodstock development, induced breeding, and larval rearing of M. padangensis.\nMethods: A total of 1,000 female and 1,000 male broodfish were collected from the wild and reared in two concrete ponds (128 m2) at the Centre for Biodiversity Conservation, P.T. Semen Padang, Indonesia. The broodfish were fed commercial feed to satiation at 09:00 and 17:00 h. The females (average weight 7.56 ± 0.85 g) and males (4.86 ± 1.20 g) were selected at a ratio of 1:4 (female:male), and gonad maturation was induced with a single dose of GnRH analogue (Ovaprim) of 0.1 ml/fish. At 16 h after hormone injection, eggs were collected individually into a plastic vessel. Spermatozoa were collected with sterile syringes. Eggs were fertilized using the \"dry\" method, and 0.5 ml samples (equal to 100 eggs) were taken. The eggs were incubated in a plastic strainer with a water volume of 1.57 litres and placed in a tarpaulin pond with a volume of 150.72 litres.\nResults: The overall hatching rate was 78.93 ± 4.13%. The newly hatched larvae were 3900.81 µm long, with a yolk sac of 82881.480 µm2. The mouth opened at 72 DPH with a gape measuring approximately 61.880 µm. The protocol of larval feeding started with artificial feed, followed by Artemia nauplii up to 30 DPH. Weaning of larvae started at 4 DPH. Larvae started metamorphosis by 15 DPH and ended by 22 DPH when the larvae reached 7430.27 µm. Larval rearing resulted in an average survival rate of 28.4 ± 3.04%.\nConclusions: Its successful spawning induction and high larval hatching and survival rates make M. padangensis an excellent aquaculture candidate.", "keywords": [ "Mystacoleucus padangensis", "Broodstock", "Induced breeding", "Hatching rate", "Larval rearing" ], "content": "Introduction\n\nDiverse and unique fishery resources (including finfish, crustaceans, and molluscs) are increasingly recognised for their role in providing food security, improving nutrition, and ending malnutrition.1,2 The availability of fish as a food source can help end hunger (United Nations Sustainable Development Goals, SDGs 2).3 As defined by the Food and Agricultural Organization (FAO), food security encompasses many underlying factors in four key dimensions:4 food availability, access, utilisation, and stability. Fish can contribute to these four dimensions because they are a source of micronutrients such as protein, lipids, minerals, and vitamins.5–8 Additionally, fish are easy to harvest year-round and have low production costs.9,10\n\nTherefore, the overall fish demand has increased significantly in recent decades. By 2018, the world registered a total fish production of 179 million tons (MT), of which 46% is from fish farming.11 Of the total, 156 MT were used for human consumption, equivalent to an estimated annual supply of 20.5 kg per capita.11 In the future, fish production from capture fisheries are expected to stagnate due to nonselective fishing.12–14 Therefore, the four key dimensions of food security are not guaranteed. The only prospect to satisfy the world's fish demand is to improve its capacity for culturing in confined environments.15\n\nBy 2018, Indonesia accounted for 6.1% of the world's aquaculture production.11 Indonesia's total fish farming production was 16,032,122 metric tonnes (mt). A total of 3,374,924 mt (21.05%) was sourced from freshwater aquaculture production, 9,884,670 mt (61,65%) was sourced from marine aquaculture production, and 2,772,568 mt (17.29%%) was sourced from brackish water aquaculture production.16 Over the past decade, the species of freshwater aquaculture commodities developed have included Nile tilapia, Clarias catfish, Pangasius catfish, common carp, and giant gourami, which have contributed 37.39%, 33.35%, 12.38%, 9.28%, and 6.98% of the total freshwater aquaculture production, respectively.16 Efforts to encourage aquaculture development in Indonesia are constrained by the low number of successfully domesticated indigenous species that are aquaculture candidates.6,17\n\nA total of 1,300 fish species, including 40 endemic species, are recorded as living in freshwater in Indonesia.18 Mystacoleucus padangensis (Bleeker, 1952) of the Cyprinidae family (Indonesian name bilih) is endemic to Lake Singkarak (the surface area is 112 km2), West Sumatera Province, Indonesia.19,20\n\nM. padangensis, a small Cyprinid, grows to approximately 20 cm and weighs 12 grams.21 This species has a high potential market demand but is threatened by overfishing and has not been successfully cultured.22 M. padangensis has been categorised as “vulnerable” by the International Union for Conservation of Nature (IUCN).23 To determine the potential of M. padangensis as an aquaculture candidate in Indonesia, we (i) determined the reproductive characteristics of female and male broodstock reared under farm conditions in concrete ponds, (ii) induced spawning to obtain fertilisation and hatching rates in hatcheries, and (iii) investigated larval development of M. padangensis and survival up to 30 days after hatching with a protocol of pre-feeding with commercial feed and then with live food.\n\n\nMethods\n\nCaptive M. padangensis broodfish were kept in concrete ponds in the Centre for Biodiversity P.T. Semen Padang, Indonesia to obtain data on female and male reproductive characteristics, induce spawning to obtain fertilisation and hatching rates, and determine the development of M. padangensis larvae up to 30 days after hatching. There were no required permits from the government of the Republic of Indonesia for this study. This research was funded by the Board of Directors of P.T. Semen Padang, Indonesia, under grant No. 0000072/HK.03.02/PJJ/50003897/3000/07.2022. This study received ethical approval from the Ethics Commission for Research and Community Service at Universitas Bung Hatta (092a/LPPM/Hatta/VII-2022). Ethical approval was given to collect fish samples and rear them in a pond, including induced spawning and larval rearing in hatcheries.\n\nLift nets in Lake Singkarak were used to catch 2,000 broodstock candidates of M. padangensis. These broodfish candidates were placed in an oxygenated polythene bag. The broodfish were transported by truck to the farm of Semen Padang Indonesia Resort, Padang City, West Sumatera Province. They were adapted and reared to oocyte development under farming conditions. During oocyte maturational development, the broodfish were placed in two concrete ponds (8 × 8 × 2 m) separated by sex. The water level in each concrete pond was 1 m. Pond water was sourced from a reservoir owned by P.T. Semen Padang with a water inflow of 1 m3 per second. During the rearing of broodfish, the water temperature and pH were between 24 and 26°C and 7.1 and 7.3, respectively. Dissolved oxygen varied from 6.4 to 6.8 mg/L, and alkalinity and hardness ranged from 65-68 mg/L HCO3- and 52-55 mg/L CaCO3, respectively.\n\nDuring domestication on the farm, the broodfish were fed commercial floating feed (Prima Feed L.P. 0, diameter size 1.6 mm, protein 33%, and lipid 4%). Feed was given to satiation daily at 09:00 and 17:00 h. A total of 1,000 female and 1,000 male broodfish were reared in the concrete ponds. The average fish weight (FW) and fish length (FL) of the twenty female broodfish were 7.56 ± 0.85 g and 9.31 ± 1.36 cm, respectively; those of the twenty male broodfish were 4.86 ± 1.20 g and 6.38 ± 0.30 cm, respectively. Broodfish were analysed for the condition factor (CF), gonadal weight (GI), gonadal somatic index (GSI), absolute fecundity (AF), relative fecundity (RF), egg diameter (ED), semen volume (SM), semen pH (SpH), and motility of spermatozoa (MOL) from male broodfish. The relationships between female brood weight and absolute fecundity and gonadal weight, as for relationships between male sow weight and gonadal weight and semen volume, were assessed using the least square’s regression method. Microsoft Office Professional Plus 2019 was used for plotting the figures.\n\nThe female and male broodfish were checked for their gonadal maturity for spawning from early April 2022 onwards. Hence, to simplify catching broodstock, the pond water level was lowered to 20 cm, and then the broodstock was collected with a drive-in net with a suitable mesh size. Broodstock was fasted for 24 h before being captured. The broodfish were anaesthetised orally with clove oil at 0.5 mL/10 L water. The gonadal maturity stages (GMS) II, III, and IV of the female broodstock were classified as follows21: in GMS II, the ovary containing the egg is dark green and visible to the naked eye; in GMS III, the ovaries fill approximately 70% of the abdominal cavity; eggs are greyish-green, visible to the naked eye, and larger than they are in GMS II; and in GMS IV, the ovaries fill approximately 80% of the abdominal cavity; visually, the ovaries contain greyish-green eggs that are larger and vary in size and the blood vessels are visible, especially in the ventrolateral area. Before hormone injection for ovulation, oocyte samples were taken from females in vivo as described previously21 and were placed in Serra’s solution (6:1:1; 70% ethanol, 40% formaldehyde, and 99.5% acetic acid) for five minutes to determine the position of the cytoplasm. Then, the oocyte nucleus was classified using a four-stage scale as follows.24 Stage 1: germinal vesicle in the central position, stage 2: early migration of the germinal vesicle (less than half of radius), stage 3: late migration of the germinal vesicle (more than half of radius), stage 4: germinal vesicle in the periphery or germinal vesicle breakdown (GVBD).\n\nFor ovulation and spermiation of broodfish, each female and male received one injection of GnRH analogue with a dopamine antagonist (Ovaprim trademark, fabricated by Syndel Laboratories Ltd, 2595 McCullough Rd. Nanaimo, B.C. V9S 4M9 Canada). The hormone was injected intraperitoneally in part of the left dorsal at 0.1 mL per fish. These doses refer to the published amount for the ovulation of Cyprinids.25 Fourteen to 16 h after hormone injection, broodstock ovulation and spermiation occurred. Eggs were collected individually into a plastic vessel and spermatozoa were collected with sterile syringes. Eggs were fertilised using the “dry” method, as described by.26 The eggs were incubated in a plastic coconut milk strainer with a diameter of 20 cm and a water volume of 1.57 litres, and then moved to a circular tarpaulin pond with a diameter of 80 cm and a height of 45 cm that was filled with 150.72 litres of water. The water level in the tarpaulin pond was 20 cm, and the water temperature ranged between 24 and 26°C. Eggs were continuously aerated until they hatched. The eggs hatched after 19-20 h of incubation at 25 ± 1°C.\n\nThe physicochemical values of temperature, dissolved oxygen, pH, nitrate-nitrogen, alkalinity, and hardness in circular tarpaulin ponds used for larval rearing of M. padangensis were noted every seven days. The water samples were collected at 10:00 am at a depth of 10 cm in each circular tarpaulin pond. A thermometer (Celsius scale) was used to measure water (°C) temperature, and an oxygen-meter (YSI Model 52, Yellow Instrument Co, Yellow Spring, OH USA) was used to measure dissolved oxygen (O2; mg/L). A digital pH-meter (Mini 0–14 pH I.Q., Scientific Cemo Science, Thailand) was used to determine the pH. The levels of nitrate-nitrogen (NO3-N; mg/L), alkalinity (mg/L), and hardness (mg/L) were analysed using standard methods for the examination of water and wastewater (APHA).27\n\nIndicators of egg quality were estimated from the fertilisation and hatching rates of five spawning trials. The fertilisation rate was calculated eight hours after spawning and an estimated twelve hours before hatching. Fertilised eggs are transparent, while those that are not fertilised are opaque. Embryonic and larval development was examined under a digital microscope (EcoBlue, S/N – EC 2203076 Thailand) with a built-in photoimaging system connected to a computer for morphometric investigation. All larvae were fed artificial feed (BP Eguchi. Taiwan Co Ltd. Labelled 4% moisture, 42% protein, 34% fat, 7% ash, and 1% fibre) to apparent satiation twice daily, at 09.00 and 17.00 h, from four days post-hatching (DPH) to 20 DPH. Live food such as Artemia nauplii was started by 17 DPH to 30 DPH. The percentage of larval survival (%) was measured as the number of metamorphosed individuals by the number of hatched-out larvae stocked multiplied by 100. The survival rate is expressed as the mean ± SD.\n\n\nResults\n\nThe reproductive performance of M. padangensis females reared for 90 days is summarised in Table 1. The average live weight of the females was 7.56 ± 0.85 g, and the gonadal-somatic index (GSI) varied between 9.78 and 16.95%. Absolute fecundity varied between 3,539 and 5,656 eggs per female. The reproductive characteristics of the male fish are summarised in Table 2. The average live weight of the males was 4.86 ± 1.20 g, and the GSI and semen volume were 6.85 ± 0.83% and 0.12 ± 0.02 mL, respectively. The relationships between female brood weight and absolute fecundity (y = 589.789×+64.997,r2 = 95%, P > 0.000, Figure 1(a)) and gonadal weight (y = 0.041×-0.087, r2 = 70%, P = 0.000, Figure 1(b)) were determined, as were relationships between male sow weight and gonad weight (y = 0.045×+ 0.110, r2 = 66%, P = 0.000, Figure 1(c)) and semen volume (y = 0.016× + 0.045, r2 = 74%, P = 0.000, Figure 1(d)).\n\nThe distribution of egg diameters in GMS II, III, and IV is shown in Figure 2. Female oocyte maturation in M. padangensis was of the synchronous batch type. A bag of eggs to be ovulated was detected to develop simultaneously with almost equal sizes. In GMS IV, most (55%) gonadal maturity was much higher than that of immature oocytes (5%).\n\nThe present study showed that the size of the spawned eggs of M. padangensis before fertilisation was 957.941˗1,180.156 μm (Figure 3). With a female and male sex ratio of 1:4, the fertilisation rate for five trials ranged between 83.33% and 91.11%, and the hatching rate varied from 77% to 86% (Figure 4). Egg development occurred 15 h after fertilisation (Figure 5), and the eggs hatched after 19-20 h of incubation at a water temperature of 25 ± 1°C. In this study, the water quality parameters in the larva reared in a circular tarpaulin pond during the experimental period included a water temperature varying from 24-26°C, dissolved oxygen ranging from 6.42-6.5 mg/L, and pH between 6.4-6.6. Additionally, alkalinity ranged from 51.5-53.5 mg/L HCO3-, hardness ranged from 64.5-66.5 mg/L CaCO3, and nitrite-nitrogen ranged from 0.02-0.05 mg/L. All physicochemical parameters were able to support embryo development and survival of the larvae.\n\nNewly hatched M. padangensis larvae were seen swimming vertically to the surface of the water and returning to the bottom of the container. Then, larvae settled at the bottom of the tank and swam back vertically to the water’s surface. This movement was interpreted as a movement to fill the swim bubbles with air. Newly hatched larvae were 3900.8156 ± 0.001 μm in total length, with a slightly elongated oval yolk sac of 82881.480 μm2 and 39822.933 μm2 oil droplets in the area (Figure 6). Body length on the first DPH was 4010.3246 ± 0.0011 μm, while the yolk sac decreased to 10214.043 mm2 (Figure 7). At 48 hours after hatching, the larval body length increased to 4013.8285 ± 0.0075 μm; the yolk sac was almost absorbed, and the eye began showing pigmentation with an eye diameter of 43.31 μm. At this stage, the appearance of the caudal fin and pectoral fins and the chromatophore pigmentation cells were black (Figure 8). By 72 hours after hatching, the body length of the larvae increased to 4215.720 ± 0.0026 μm, the yolk sac was almost completely absorbed, and their mouths opened to 61.880 μm (Figure 9). On the fourth day after hatching, larval body length was 4330.4407 ± 0.0021 μm, with an open mouth of 61.880 ± 0.0009 μm (Figure 10). At that time, the artificial feed was given, namely, BP Eguchi, to satiation (BP Eguchi is an artificial feed formulated to increase shrimp fry growth). The size of the artificial feed was 31.716 ± 12.238 μm.\n\nLarval body length reached 4380.251 ± 0.0025 μm by the eighth DPH; by this time, the amount of BP Eguchi was increased to satisfy the consumption need. The amount of feed consumed was more observable in the stomach (Figure 11). The body length of the larvae reached approximately 4421.231 ± 0.0028 μm on the eighth DPH; at this stage, the dorsal and pelvic fins began to appear (Figure 12). We gave them more artificial feed to increase survival and growth, which could be seen spreading in the stomach (the feed is black).\n\nThe larvae grew to 44525.684 ± 0.0019 μm by the 12th DPH, by which time all fin types were well demarcated. Chromatophore pigmentation started during embryo development and became intense as the larvae grew. The larval body colour was transparent until the 12th DPH (Figure 13). Live food (Artemia nauplii) was fed to the larvae from the 17th DPH when larval body length reached approximately 5850.2332 ± 0.0025 μm. Larvae started metamorphosis by the 15th DPH when their body length had reached 6450.0935 ± 0.0457 μm and was completed by the 22nd DPH when their body length had reached 7430.27 ± 0.0638 μm. The larval body remained transparent until the completion of metamorphosis into juveniles (Figure 13). At 30 DPH larvae rearing, the body length reached 10500.155 ± 0.06 μm (Figure 14), and the survival rate was 28.4 ± 3.04%, with feed and water management protocols as summarised in Figure 15. This fact indicates the first successful complete metamorphosis in M. padangensis.\n\n\nDiscussion\n\nM. padangensis is a potential candidate species for freshwater aquaculture because it has a high market price but is threatened by overfishing.22 This study is the first to detail the reproductive characteristics of broodstock reared in ponds for preliminary spawning and larval rearing of M. padangensis in hatcheries.\n\nOur study showed that the body weight of female M. padangensis fish reared in ponds before spawning varied from 5.85 to 9.38 g/fish, with a GSI ranging from 9.78 to 16.95%. This is smaller than M. padangensis broodfish caught from the wild, each of which weighs 10 to 15 g with a GSI of 13.09 to 22.36%.28 The fecundity of broodfish reared in ponds ranged from 3,539 to 5,656 eggs/fish. In the wild, it varies from 3,469 to 6,531 eggs/fish.21 The female weight of broodstock and absolute fecundity had a strong relationship (r2 = 95%, P = 0.000). The differences in absolute fecundity in maturing broodfish depend on the brood weight, age of broodfish, and species, including strain.6,29–31 Female M. padangensis broodfish oocyte development is of the synchronous batch type. One bag of eggs will have up to 80% spawning, with sizes ranging from 374 to 558 μm. This condition is related to the spawning behaviour of M. padangensis in the wild: the fish migrates to rivers with shallow water characteristics (water depth 0.41 m), bottom substrate consisting of sand, gravel, and caracal, and a current speed of 47.00 m/sec.22 Conversely, the Indian pompano, Trachinotus mookalee, is a pelagic species that inhabits the shallow Indian Ocean and has synchronous batch-type development,32 as does Plata pompano, Trachinotus marginatus.33\n\nThis study showed that the dominant egg diameter for spawning ranged from 466-558 μm. After ovulation, there was an increase in egg diameter ranging from 957.941-1180.156 μm, equivalent to 50.30-52.70%. Previous studies showed there was an increase of 27.8-32.14%6 in the egg diameter of the giant gourami (Osphronemus goramy), an increase of 4.64-32.36%17 in that of the Asian catfish (Hemibagrus wyckii), and an increase of 5.3-27.1%34 in that of the Alakir trout (Salmo kottelati). In the present study, the average fertilisation and hatching rates at a water temperature of 24-26 °C were 85.93 ± 3.07% and 78.93 ± 4.13%, respectively. Fertilisation rates and hatching rates vary between candidate species for aquaculture, e.g., 69 ± 1.55% and 87.67 ± 0.81% for Trachinotus mookalee,32 60.91 ± 4.68% and 42.91 ± 2.92% for Hemibagrus wyckii,17 81.60 ± 3.37% and 76.40 ± 2.22% for Osphronemus goramy,6 and 88.3 ± 2.88% and 65.02 ± 7.02% for Salmo kottelati.34 Differences in fertility and hatching rates can be caused by the quality of sperm,35,36 temperature of the water in the hatching tank, activating medium,37,38,39 egg stocking density,40,41 and heavy metal toxicity.42\n\nThe development of larval morphology for each fish species is almost the same, but the initial growth of the larvae is different. In this study, the specific growth rate of M. padangensis larvae was estimated at 3.30% per day over 30 days of larval rearing. The length of the newly hatched larvae was 3.9 mm, and it increased to 10.5 mm over the 30 days after hatching (Figure 14). In contrast, the growth rate of the Indian pompano (Trachinotus mookalee, Carangidae) from 1 to 30 DPH was 11.4% per day,32 while the growth rate of catfish (Clarias magur) larvae in the different tank colours from 4 to 32 DPH varied from 11.38-11.99%.43 The slower growth of M. padangensis larvae was probably caused by the type of feed given during the 30 days of maintenance, starting with artificial feed (BP Eguchi) suitable for the mouth opening of the larvae and continuing with live food, such as Artemia nauplii. Some researchers have suggested that commercial feed for larvae has undergone considerable advancement. However, live feeds such as Artemia, Daphnia manga, rotifers, copepods, among others, are still necessary for most larvae at feeding times.44–46 According to Tancioni et al.,47 the growth rate of the brook chup (Squalius cutemonis, Cyprinid) when fed rotifers between 10 and 20 DPH is 0.8%/day; when fed copepods (from 40-50 DPH), the growth rate slightly increases to 2.4%/day. The different feeding regimes have evident effects on larval performance. Larvae Ballan wrasse (Labrus bergylta) receiving copepods as their initial diet indicated significantly higher survival rates than those fed on rotifers.45 Whether M. padangensis larvae receiving rotifers as an initial diet, continuing with copepods, and so on with Artemia nauplii can increase larval survival by more than 28.4 ± 3.04% is still poorly understood. In addition to the type of feed given, the stocking density of the larvae (15-300 larvae/m-2) has also been shown to affect the growth of giant gourami (Osphronemus goramy) larvae reared from 7 DPH to 21 DPH.48 In addition, for tambaqui (Colossoma macropomum) larviculture, the best stocking density is 180 larvae/L.49 In the present study, whether the growth rate of larval M. padangensis was affected by food type and stocking density was not clear. Therefore, a clear nutritional protocol and optimal stocking density to enhance the growth of M. padangensis larvae are necessary for future analyses.\n\nIn this study, with 30 days of larval rearing with the feeding protocol, illustrated in Figure 15, the survival rate reached 28.4 ± 3.04%, which was the first success in M. padangensis aquaculture. Researchers have reported that the survival rate for the larvae of each fish species varies depending on the feeding protocol.32,47,45 Additionally, poor nutrition during larval development has an adverse effect on survival rates. Malnutrition causes a decrease in larval survival, for example, by indirectly decelerating the growth rate, resulting in a longer larval stage and poorer larval activity ability, increasing predators’ opportunities to prey on the larvae.50–52 The diverse feeding regimes have evident consequences on larval development. Larvae receiving rotifers as their initial diet show significantly lower survival rates than those fed copepods.45 Larval survival is influenced not only by feeding protocols and poor nutrition, but also by rearing tank background colour,43 water temperature,32,53,54 different stocking densities,48,55 and live food in aquaculture.56–58 All these factors are challenges to increasing the survival and growth of M. padangensis larvae.\n\n\nConclusions\n\nThis study shows that M. padangensis can be domesticated under farming conditions by broodstock development, which leads to final gonadal maturity. This report is the first successful record of wild fish gonadal maturation under farming conditions, induced spawning, and larval rearing of this species with an indoor hatchery system. The maturing broodstock in captivity can be used for seed production by artificial spawning. Its high egg hatchability and larval survival rate of 28.4% make M. padangensis an excellent candidate for aquaculture.", "appendix": "Data availability\n\nFigshare: Broodstock development, induced spawning and larva rearing of the bilih, Mystacoleucus padangensis (Bleeker, 1852), a vulnerable species, and its domestication potential for new candidate aquaculture., https://doi.org/10.6084/m9.figshare.22179500. 59\n\nThis project contains the following underlying data:\n\n- Table 1. Raw data female size and characteristics reproduction\n\n- Table 2. Raw data male size and sperm characteristics of M. padangensis on the farm\n\n- Table 3. Raw data Distribution of egg diameters in each of gonadal maturity stages II, III, and IV\n\n- Table 4. Raw data fertilization rate and hatching rate of M.padangensis\n\n- Table 5. Raw data larva development newly hatched to 30th DPH (μm)\n\nFigshare: ARRIVE Essential 10 checklist for “Broodstock development, induced spawning and larval rearing of the bilih, Mystacoleucus padangensis (Bleeker, 1852), a vulnerable species, and its potential as a new aquaculture candidate, https://doi.org/10.6084/m9.figshare.22179500. 59\n\nData are available under the terms of the Creative Commons Attribution 4.0 International License (CC-BY 4.0).\n\n\nAcknowledgments\n\nWe thank the Board of Directors of P.T. Semen Padang, Indonesia, which funded this research. We also thank the fishermen in Lake Singkarak who helped collect fish and the students who helped with research activities in the laboratory.\n\n\nReferences\n\nShepon A, Mokov T, Hamilton HA, et al.: Sustainable optimization of global aquatic omega-3 supply chain could substantially narrow the nutrient gap. Resour. Conserv. Recycl. 2022; 181: 106260. 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Publisher Full Text\n\nKumar R, Gokulakrishan, Debbarma J, et al.: Advances in captive breeding and seed rearing of striped murrel, Channa striata, a high value foo fish of Asia. Anim. Reprod. Sci. 2022; 238: 106957. PubMed Abstract \| Publisher Full Text\n\nChen Z, Dong S, Dai L, et al.: Effect of food domestication on the growth of Elopichthys bambusa. Reproduction and Breeding. 2021; 1(3): 157–166. Publisher Full Text\n\nWang YF, Liang XF, He S, et al.: The potential use of Artemia for larval rearing of mandarin fish (Siniperca chuatsi). Aquaculture Reports. 2022; 25: 101216. Publisher Full Text\n\nSyandri H: Broodstock development, induced spawning and larva rearing of the bilih, Mystacoleucus padangensis (Bleeker, 1852), a vulnerable species, and its domestication potential for new candidate aquaculture. Dataset. figshare. 2023. Publisher Full Text" }	[ { "id": "170203", "date": "04 May 2023", "name": "Md. Moshiur Rahman", "expertise": [ "Reviewer Expertise Fish reproduction", "larval rearing", "sperm quality assessments", "fish biology", "and fish nutrition." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors have conducted an important research study on broodstock development, induced spawning, and larval rearing of the bilih, Mystacoleucus padangensis (Bleeker, 1852), a vulnerable species with potential as a new aquaculture candidate. I appreciate the authors' efforts in conducting this research. However, I do have specific comments on each section, which I have provided below:\nComments: Abstract:\n\nBackground: Please add \"yet\" - \"has not yet been successfully cultured.\"\n\nMethods: \"The broodfish were fed commercial feed to satiation at 09:00 and 17:00 h.\" How many times?\n\nResults: The mouth opened at 72 DPH? Please write the elaborated form of DPH as it was written for the first time in order to better understand the context in which it was used.\n\nResults: Artemia is a genus of brine shrimp. The standard convention for scientific writing is to italicize the names of genus and species, including Artemia. Therefore, it is recommended to write Artemia in italic font in scientific or academic papers.\n\nResults: \"Weaning of larvae started at 4 DPH.\" How did you start - co-fed with Artemia or directly feeding with dry feed?\n\nConclusion: I’d suggest rephrasing this paragraph such as: The successful induction of spawning, as well as high rates of larval hatching and survival, make M. padangensis a highly desirable candidate for aquaculture.\n\nIntroduction:\n\n2nd paragraph: Please do not start it with \"Therefore\" and rephrase it.\n\n3rd para: \"Nile tilapia\" is not the scientific name of a species. It is a common name that refers to several different species of tilapia, which belong to the family Cichlidae. So, please do not italicize it.\n\n4th para: I can see a big jump here before introducing Mystacoleucus padangensis. Please make a perfect link to introduce it. For example, before discussing this species, which is a freshwater fish species native to the rivers of Sumatra, it is important to understand the context of its ecological niche within the aquatic ecosystem.\n\n5th para: I can see that it is a small fish. However, why does it have a high market demand? Could you provide specific references to support this claim?\n\nMethods:\n\nCould you please rephrase the sentence as follows: \"They were adapted and reared to oocyte development under farming conditions.\" The correct sentence is: \"During the oocyte maturation stage of development...\"\n\n\"Feed was given to satiation daily at 09:00 and 17:00 h.\" How many times, ration, amount, etc.?\n\nMay I know if the fish length (FL) was measured as total length, standard length, or fork length?\n\n\"Broodfish were analysed for the condition factor (CF), gonadal weight (GI), gonadal somatic index (GSI), absolute fecundity (AF), relative fecundity (RF), egg diameter (ED), semen volume (SM), semen pH (SpH), and motility of spermatozoa (MOL) from male broodfish.\" Please write briefly how and which techniques (also provide appropriate references for each one) were followed to analyze them?\n\n\"The average fish weight (FW) and fish length (FL) of the twenty female broodfish were 7.56 ± 0.85 g and 9.31 ± 1.36 cm, respectively; those of the twenty male broodfish were 4.86 ± 1.20 g and 6.38 ± 0.30 cm, respectively.\" Please split these sentences to understand them better.\n\n\"The relationships between female brood weight and absolute fecundity and gonadal weight, as for relationships between male sow weight and gonadal weight and semen volume, were assessed using the least square’s regression method.\" Also split these sentences too.\n\nResults:\n\n\"Mystacoleucus padangensis\" must be italicized.\n\nIn the 'Methods' section, it is necessary to provide a brief explanation of how you measured or analyzed reproductive characteristics such as weight, GSI, fecundity, and others.\n\nAlso, please define and show how you estimated the absolute fecundity in the 'Methods' section.\n\nPlease split the sentences providing the outcomes of the relationship to understand them better.\n\nWhat does * mean in the equation?\n\nPrior to conducting the regression analyses, did you verify that the assumptions were met? As fecundity is a count variable, it is likely that it may not follow a normal distribution. Please provide clearly this information in the statistical section.\n\nAs previously stated, it is important to clarify in the tables whether FL refers to total length, fork length, or standard length.\n\nIs a sample size of n=20 sufficient to yield meaningful results? Please do a power analysis or provide the effect size for these outcomes.\n\nPlease do an individual analysis to explore the variation among and/or between different GMS (shown in Fig. 2), fertilization rate, and hatching rate (shown in Fig. 4).\n\nDiscussion:\n\n\"a potential candidate species for freshwater aquaculture because it has a high market price\" - Please explain why with appropriate references.\n\nOverall, the discussion section appears to be inadequately written. The comparison of each finding with others lacks relevance and clarity. It is important to provide a clear explanation of why these findings should be compared and discussed with other studies and findings, and to provide possible explanations.\n\nConclusion:\n\nWhat do you mean by \"final gonadal maturity\"?\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10119", "date": "29 Nov 2023", "name": "Azrita Azrita", "role": "Author Response", "response": "Reviewer Report response by Md. Moshiur Rahman, Department of Biology and Agricultural Engineering, University of California Davis, Davis, California, USA Abstract Background: We will add in the new revised version the word \"yet\" has not been successfully cultured. Methods: We will revise the new version to include feeding commercial feed to satiation at 09:00, 13:00, and 18:00 hours. Results: We will consistently maintain the DPH form as originally described in the new version. Results: We agree to write Artemia scientific name in italics according to scientific writing standards in the new version. In the revised version, we will incorporate the narrative about dry feeding and the role of Artemia in the weaning process of the larvae. Conclusion: We will consider suggestions from reviewers to apply the sentences suggested in the concluding section of the abstract to the new revised version. Introduction We agreed not to start the second paragraph with \"therefore\" and will make improvements to the new version. 3rd paragraph: We agree that the names of Nile tilapia, Clarias catfish, and Pangasius catfish do not need to be italicized. We will revise the new version. We agreed to add a new revised version to make a proper relationship regarding the importance of Mystacolecus padangensis, a freshwater fish species in Lake Singkarak. We agree to add specific references to support claims about the importance of Mystacoleucus padangensis, even though this fish is included in the small-size category. Methods We agree to replace the sentence \"They adapted and bred to develop oocytes under agricultural conditions.\" with \"At the developmental stage of oocyte maturation…\". We will make these changes in the new version revision. We agreed to add information regarding feeding time, feeding rates, etc., in the revised version. In this study, we measured the standard length of each Mystacoleucus padangensis, and we will add it in a new revised version. We agreed to add to the new revised version of the formula used to calculate the parameters of condition factor (CF), gonadal weight (GI), gonadal somatic index (GSI), absolute fecundity (AF), relative fecundity (RF), and other supported parameters. by references. We agreed to make a new sentence on the revised version to clearly explain the fish weight and length data for easy understanding. We agreed to make a new sentence in the revised version that clearly explains the relationship between male parent weight and gonadal weight and how semen volume is assessed using the least squares regression method. Results In the revised version, we will clearly explain the review comments related to the research method in the results section. We believe that a sample size of 20 individuals is sufficient to provide significant results, and we will also provide an in-depth explanation of the power analysis of the study. In addition, we agree to add an explanation about the analysis of GMS variation (Figure 2) in the context of fertility and its relationship to egg hatchability (Figure 4). All this we do with the aim of ensuring a clear understanding of the new revised version. Discussion In the discussion section, we agree to add an explanation regarding Mystacolecus padangensis as a candidate for aquaculture, which is supported by relevant references. This will provide a better understanding of the context of the discussion. We agree to add a comparison of each parameter in our study with more apparent findings from other studies. This will help provide context and a deeper understanding of the analysis of the results. Conclusion In this section, we agree to add an explanation about \"final gonadal Maturity.\" I confirm that I have read this review report. I have the expertise to revise this article according to accepted scientific standards." } ] }, { "id": "197812", "date": "17 Oct 2023", "name": "Safaa M Sharaf", "expertise": [ "Reviewer Expertise Fish biology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nBroodstock development, induced spawning and larval rearing of the bilih, Mystacoleucus padangensis (Bleeker, 1852), a vulnerable species, and its potential as a new aquaculture candidate, is an important research study of Mystacoleucus padangensis living in Lake Singkarak, Indonesia, which has high potential market demand but is threatened by overfishing and has not been successfully cultured. I appreciate the authors' efforts in conducting this research. However, I have specific comments on some sections, which I have provided below:\nIs the journal system to mention only the methods without the materials in the research?\nMethods:\nThe authors used clove oil, 5 ml/10 liters, and did not mention for how long (minutes).\n\nThe authors did not mention the injected dose of gonadotropins and ovaprim/fish only the injection volume was mentioned as 0.1 ml/fish.\n\nThe authors did not mention in the methods that the growth of larvae will be calculated during the 30 days, and this period is too short to estimate the growth of fish. Besides, the length gain is not mentioned as a growth performance parameter.\n\nPlease in the methods, you must mention how these measurements were done all the analysis of CF, GSI, AF, RF (relative fecundity measured for length or for weight?), MOL and its formula or equations with a reference.\n\nPage 4: \"the oocyte nucleus was classified using a four-stage scale as follows Stage 1: germinal vesicle in the central position, stage 2: early migration of the germinal vesicle (less than half of radius), stage 3: late migration of the germinal vesicle (more than half of radius), stage 4: germinal vesicle in the periphery or germinal vesicle breakdown (GVBD).\" - Are these authors (Ref. 24, Krejszeff S, Katarzyna T, Daniel Z, et al.) the original ones who published this classification? It is preferable to write in methods the original author.\nResults:\nDoes the size of the eggs mean the diameter of the eggs? Page 5 and in the Figure 3? If yes, scientifically, it is better to mention the egg diameter, not the size of the egg.\n\nThe shape of the yolk sac was not identified or marked in the figures 6, 7 and 8 as it was determined in the presence of artificial food at Figure 10.\n\nFigure 5 - What type of stage of larval development is this? It is better to mention it and mention its reference.\n\nThe title under the figures is preferable to mention the length of 6-day-old post hatch larvae showing..., and so on in all the figures, from Figure 6 to 13\n\nIs this fish species (4-7g) required by the consumer as eatable or as an ornamental fish? Because of its very small size, is it worth it to be artificially hatched?\nConclusions\nThe survival rate of 28.4% is very low not as mentioned in the conclusion (Conclusions: Its successful spawning induction and high larval hatching and survival rates make M. padangensis an excellent aquaculture candidate). The reasons must be known by doing another future research because the fertilization and hatching rates of M. padangensis were high and promising.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10411", "date": "30 Nov 2023", "name": "Azrita Azrita", "role": "Author Response", "response": "The author provides a response to a review by Safaa M Sharaf, a researcher at the Department of Animal Production Science and Fishery Resources, Suez Canal University, Ismailia, Egypt. This article will be enriched with comprehensive methods and materials in this research. Therefore, the author will include complete material in this article in the revised version. Methods The author will add in the revised version that the mother is anesthetized orally with clove oil in 0.5 mL/10 L of water for ten minutes. The author will add in the revised version that Ovaprim is injected intraperitoneally in the left dorsal region at 0.1 mL per animal. Based on the packaging, Ovaprim contains sGnRH-a (Salmon Gonadotropin-Releasing Hormone analog) as much as 20 µg/ml and domperidone as much as 10 mg/ml). The authors admit that it was not explained in the initial method that larval growth would be calculated over 30 days, and length parameters were not yet included among the measured parameters. Therefore, the authors will include survival and length parameter measurements in the revised version of this study for 30 days. The author uses the following formula or equations, which will be added to the revised version: - Condition factor (CF): weight wet in grams/length3 x 100 - Fish gonad weight is the size or total mass of fish reproductive organs called gonads - GSI = (gonadal weight ÷ total body weight) × 100 - Absolute fecundity (AF) is calculated by multiplying the number of eggs per 0.1 gram by the weight of the gonad in grams. - Relative fecundity (RF): number of eggs in 0.1 gram of the gonad weight. - Sperm motility percentage (MOL) was determined by observing activated semen placed on a glass slide under a microscope, as described in the study by Rurangwa et al. (2004). MOL examination was carried out using a digital microscope model EcoBlue with serial number S/N – EC 2203076, which was manufactured in Thailand. This microscope is equipped with an internal photo-imaging system connected to a computer for sperm motility examination. The author will use reference No. 24 according to the original, namely Kujawa R., Kucharczyk D. A method of in vivo designating oocytes from cyprinid and percid fish using a catheter. Commun. Ryb. Olszt. 1996;4:20–21. (In Polish) [Google Scholar] Results The author will use the egg diameter in the description of Figure 3 in the revised version. The author will add yolk sac shapes not identified or marked in Figures 6, 7, and 8 to the revised version. The author will explain the stages of development in Figure 5 in the revised version. The author will consider including larval length below the figure in the revised version. Mystacoleucus padagenis is classified as a small indigenous species that the consumer requires to be eatable. This species has successfully artificially hatched (Female broodstocks were 7.56 and 9.31cm, and male broodfish were 4.86 and 6.38 cm, respectively) Conclusions In the revised version, the authors will add in the conclusion that survival challenges remain low and that future research needs to focus efforts to address these challenges." } ] } ]	1	https://f1000research.com/articles/12-420
https://f1000research.com/articles/12-1411/v1	26 Oct 23	{ "type": "Research Article", "title": "Prevalence of tooth wear diseases in patients with diabetes and its impact on the quality of life in Dakshina Kannada population: a cross sectional study", "authors": [ "Poojakumari Sinha", "Sangeeta Nayak", "Lavangi Sehgal", "Ramya Shenoy Kudupi", "Poojakumari Sinha", "Lavangi Sehgal", "Ramya Shenoy Kudupi" ], "abstract": "Background Early detection and management of tooth wear (TW) has not been given due consideration in the dental profession. Thus, this study aimed to explore the prevalence of TW in adults with diabetes in Dakshina Kannada population using the Basic Erosive Wear Examination (BEWE) and recording its impact on quality of life using the Oral Health Impact Profile (OHIP-14).\n\nMethods In total, 236 dentate adults with diabetes and TW, who visited the Manipal College of Dental Sciences, Mangalore were recruited to the study. Assessment of TW was done using BEWE during examination of each participant and was categorized as none, mild, moderate, and severe. Impact of TW on quality of life was assessed using the OHIP-14.\n\nResults Overall, 81 (34.30%) individuals had a poor score on the basic erosive tooth index, 82 (34.70%) had a medium score, . The level of TW, both moderate and severe (high), affected quality of life. Erosion, abrasion and attrition were seen in a large proportion of the study population. The diabetic status of the individual was collected from their medical file. The HbA1C level was found to range from good, fair and poor control. The quality of life among the study population was affected because of TW and diabetes. Few participants had hypertension along with diabetes. The majority of study participants were on oral hypoglycaemic agents. Participants used tooth brushes and tooth paste and brushing frequency varied between once or twice daily.\n\nConclusions In the study population, there were moderate and severe TW patterns. Quality of life was found to be impacted by TW. Dental professionals must give proper consideration to the influence of TW on quality of life in addition to clinical findings. This will make it easier to offer preventive or restorative management depending on the situation.", "keywords": [ "Attrition", "Abrasion", "Erosion", "Tooth wear", "Quality of life" ], "content": "Introduction\n\nPeople are keeping more natural teeth as they live longer lives. It is vital to have healthy, functional teeth when eating and following a regular diet. Tooth wear (TW), on the other hand, has a detrimental influence on the survival of natural teeth. TW, also known as tooth surface loss (TSL), has recently been identified as a serious oral health concern.1–3 TW is an insidious, multifactorial disease that affects enamel and dentine, and it may endanger tooth survival and the quality of life linked with dental health among those who are affected.4 Epidemiological studies support the hypothesis that TW is growing increasingly severe and frequent, not just among older adults but also among those approaching adulthood.5,6 The three most often recognized aetiologies of TW are erosion, attrition, and abrasion.6 Abfraction, the fourth aetiological factor, has been acknowledged by some but is not accepted by all.6 These are associated with a variety of clinical symptoms. Given the aetiology's complexity, a definitive diagnosis may be difficult to achieve. TW influences both aesthetics and functionality.\n\nPatients suffering with TW seek dental care as a result. The chemical breakdown of enamel or dentine produced by non-bacterial acids from dietary or gastric sources is known as erosion. The first stage is characterized by dull “silky glazed” enamel surfaces and enamel loss. The palatal surfaces of the maxillary anterior teeth are affected because the tongue and buccal mucosa shelter the other surfaces from exposure to stomach acid in the early stages of gastric erosion.6 Dietary erosion is characterized by widespread cupped-out lesions, with the pattern being unique to the individual's habits and diet.\n\nAttrition is a gradual deterioration of tooth structure caused by tooth-to-tooth contact in the absence of a foreign object. It often affects the incisal/occlusal region of the teeth.7 Abrasion is the mechanical wear out of tooth structure that does not require tooth-to-tooth contact, a problem that commonly affects the cervical area of the teeth and is generally caused by poor tooth brushing practices that remove acid-weakened enamel and dentine.7\n\nDiabetes mellitus, one of the most common diseases on a regional and international scale, is a major cause of morbidity in most countries.8,9 Diabetes is increasing in prevalence worldwide. According to data from the International Federation of Diabetes, 642 million people worldwide will have diabetes by 2040, up from 415 million in 2015. More than 60% of the worldwide population living with diabetes are in Asia, where the prevalence ranges from 3% to 47.3%.1 Diabetes affects almost 1.3 billion people in India, which has the world's second-highest incidence. According to the International Diabetes Federation, 72.9 million individuals in India had diabetes in 2017.\n\nDiabetes prevalence has risen rapidly and is expected to rise further as a result of the rapid and continuous socioeconomic upheaval in developing countries. Diabetes types 1 and 2 (T2DM) are becoming increasingly frequent. T2DM, the most common type of diabetes, has been the primary driver to the global rise in diabetes prevalence.10 Diabetes increases the risk of oral disease both directly and indirectly.11 Periodontal disease affects around one-third of individuals with diabetes. Periodontitis can cause tooth loss or sensitivity.11 Different oral diseases or issues impact daily life in a variety of ways. There have not been many studies done on wasting diseases and how they affect people's quality of life. As a result, while assessing dental requirements, clinical and psychological factors must be considered.\n\nPatients with diabetes make up the majority of those for whom dental practitioners utilize oral health prevention and promotion to achieve optimal oral and overall health. The incidence, severity, and underlying causes of tooth wear should be understood by dental practitioners. As a result, it is critical to research the frequency and severity of tooth wear among patients with diabetes, as well as how it impacts the quality of life of the general Dakshina Kannada community. Thus, the present research aimed to explore the prevalence of tooth wear in adults with diabetes in Dakshina Kannada population.\n\n\nMethods\n\nFrom 29th July 2022 to September 2022, this cross-sectional study was carried out at the Department of Periodontology, Manipal College of Dental Sciences Mangalore. The study protocol was submitted to the Manipal College of Dental Sciences Mangalore, Manipal Academy of Higher Education, Manipal, India's institutional ethics and review board (protocol no: 22023; approval date 28/07/22). Convenient sampling method was followed for the recruitment of the study participants. The patients who visited OPD of Manipal College of Dental Sciences, Mangalore during the study period were recruited in the study. Before joining the study, each study participant signed an informed consent form. Based on the inclusion and exclusion criteria, 236 people completed the clinical examination and questionnaire.\n\nInclusion criteria\n\nParticipants over the age of 18 years and above with a confirmed diagnosis of diabetes mellitus and willing to participate in the study were recruited for the research.\n\nExclusion criteria\n\nParticipants having a history of using pharmaceuticals for dental difficulties were excluded from the research, as were those with systemic diseases such as paralysis, leukaemia, epilepsy, depression, and other psychiatric ailments that impact TW, as well as those getting treatment for wasting diseases.\n\nEach participant's demographic information was collected using a pre-structured case form. Diabetes mellitus information, including type, duration, medication type, and any related diseases, was obtained using their medical file and a structured questionnaire, the case proforma can be found as Extended data.18 Each participant was asked about their tooth-brushing regimen, including how long and how frequently they washed their teeth. We gathered data on diet and aerated beverage use.\n\nWe examined for evidence of tooth decay, gingivitis, periodontitis, missing teeth, sensitive teeth, bruxism, attrition, erosion, or abrasion during the clinical examination. The Basic Erosive Wear Examination (BEWE) was developed to track wasting and evaluate tooth deterioration. When there is no tooth wear, the first loss of surface tooth structure is 1, the distinct defect, hard tissue loss of 50% of the surface area is 2, and hard tissue loss of 50% of the surface area is 3. Dentine participation is seen in scores 2 and 3. Following the clinical evaluation, each participant was asked to complete the Oral Health Impact Profile (OHIP-14) questionnaire. The OHIP-1412 contains a questionnaire designed to examine the social consequences of disorders that may jeopardize oral health. Participants in the research were asked to describe any issues from the preceding year using the OHIP-14 rating scale. A high OHIP-14 scale score indicated a poor quality of life.\n\nBlinding of the investigators and participant was not required as the population only included individuals with diabetes. The reporting bias was taken care of by reporting actual results and confounding agents were excluded to prevent measurement bias.\n\nThe sample size for the study was calculated using the following formula:\n\nThus, a minimum of 197 participants were needed for the study.\n\nStatistical analysis was carried out using IBM SPSS Statistics (20.0) (IBM SPSS® statistics). It was necessary to tabulate the descriptive statistics. A proportional test was used to calculate frequency and distribution. Kruskal Wallis test was applied to check the association between BEWE of patients with diabetes with total OHIP scores. Mann-Whitney test was applied to check within the groups. The result showed statistical significance between the mild to severe BEWE group. The level of significance was set at p<0.05 and post Bonferroni correction it was p<0.02.\n\n\nResults\n\nThe current study included 236 individuals with diabetes in total. There were 171 men (72.50%) and 65 women (27.50%). In total, five (2.10%) of the participants were homemakers, whereas 53 (22.50%) worked in various occupations. A total of 108 (45.80%) were self-employed, while 65 (27.50%) were professionals (Table 1). Overall, 14 (5.90%) of the study participants had diabetes and hypertension. A total of 136 (57.60%) of research participants used oral hypoglycaemic medication; 48 (20.30%) were using insulin, and 52 (22.00%) were taking ayurvedic drugs (Table 1).\n\nIn total, 98 (41.50%) individuals had excellent (4-6) previous HbA1C readings, 130 (55.10%) had good (7-8) levels, and eight (3.40%) had bad (9-14) levels (Table 1). Overall six (2.50%) people had no dental concerns. Around 230 participants in the research had different oral conditions (Figure 1). Around three (1.30%) of the 236 research participants had hopeless teeth or teeth with a poor prognosis. The bulk of the study population (134 (56.80%)) followed a mixed diet; 62 were vegetarians (26.30%). Among the research participants, 65 (27.50%) used horizontal strokes, while 134 (56.80%) used mixed strokes. A total of 229 (97.00%) of the research participants brushed their teeth twice every day for less than 2 minutes. Whereas seven (3.00%) cleaned their teeth twice every day for more than 2 minutes (Table 2).\n\nIn total, 53 (22.50%) of the research participants used dental floss. Aerated drinks were consumed by 152 people (64.40%). Aerated drinks were consumed on a weekly basis by 150 people (63.60%). Attrition, abrasion, and erosion are examples of wasting diseases; 223 (94.50%) of the study participants had different types of wasting diseases. Erosion was seen in 228 cases (96.60%), whereas abrasion was observed in 221 cases (93.60%) (Table 3 and Figure 2). The presence of wasting diseases and the severity was recorded in the oral examination. The participants gained awareness regarding the wasting diseases by their dentists or through newspapers, and magazines. Approximately 96 (40.70%) of the research participants reported sensitivity to hot and cold meals. A total of 96 (40.70%) of the participants reported sensitivity as a result of their teeth becoming thin, short, and structurally damaged. Around 32 (13.60%) of the research participants would grind their teeth. The majority of the participants last went to the dentist one or two years ago. Overall, 129 research participants (54.70%) brushed their teeth twice every day (Table 2). The fundamental erosive tooth index was seen in 73 (30.90%) of the cases. There was an early loss of surface tooth structure in 81 (34.30%), and in 82 (34.70%) participants (Table 3). Each participant thoroughly filled out the questionnaire regarding their quality of life (Table 4 and Figure 3). Chewing food was challenging for 147 (91.90%) of the subjects. Responses were grouped into three sections (neutral, agree/strongly agree, disagree/strongly disagree) (Figure 3 and Table 4). The association between tooth wear and diabetes and its impact on quality of life was assessed. The result showed statistical significance between the mild to severe BEWE group (Table 5).\n\n\nDiscussion\n\nThe current study looked at the prevalence of tooth wear and its influence on the quality of life of those with diabetes. Responses were received from male and female participants ranging in age from 18 to 80 years old, as in previous investigations.1,4 According to Bartlett et al., more male patients than female patients are sent to the dentist with concerns of TW.7 This was consistent with the findings of the current study.1,7 There is no evidence that men and women have distinct tooth forms or saliva compositions. The reasons of TW are numerous.\n\nA study by Srisilapanan et al. (2018), like ours, covered participants with type 2 diabetes between the ages of 35 and 74 years old. Patients with diabetes had a higher incidence of tooth loss, which is consistent with previous research.1 According to data from the American Dental Association,11 diabetes is responsible for one out of every five tooth loss cases. According to the most current research, the most common kind of TW was attrition, which is consistent with previous findings.1 Study participants in the present study had a greater rate of TW, which was consistent with previous research.1 The severity of minor TW and age were shown to be substantially associated.\n\nDiabetes and periodontitis are two exceedingly frequent chronic, noncommunicable diseases that pose severe public health problems for the worldwide population. For a long time, dental specialists have recognized that these two diseases are linked. Periodontitis is known as the sixth complication of diabetes. Diabetes raises the risk of periodontitis by increasing inflammation in periodontal tissues.13 Participants in the research had gingivitis, periodontitis, decay, and other dental disorders. The inability to maintain excellent oral hygiene may be related to the challenges (sensitivity and discomfort) caused by wasting disorders.\n\nDiabetes and periodontal disease treatment need a lifelong strategy tailored to the patient's individual circumstances. The majority of survey participants used mixed and horizontal strokes to clean their teeth. This must be corrected because it is a common occurrence and a substantial factor to wasting sickness. Any oral health promotion program must include tooth brushing instruction since it improves people's oral health knowledge, attitudes, and habits. Individuals may use it to construct customised dental care programs for themselves. The research participants had moderate to high scores on the basic erosive tooth index. The study looked at how tooth decay affects people's quality of life. Patients who had moderate to severe TW had a worse quality of life. If monitoring TW is made required at every clinical examination, it is less likely that it will be neglected. Every visit should include a TW evaluation to reduce the likelihood of patients’ early warning signs going ignored and no preventative steps being implemented. Dental practitioners should pay attention to their patients’ clinical features and how they affect their patients’ quality of life. On the basis of demand, specialized, preventative, or rehabilitative treatment should be made available. OHIP-14 can be used to quantify the impact of oral diseases on important elements of daily living.3 Traditional or professionally developed assessment methods for evaluating tooth wear may be utilized to prioritize treatment for individuals who require it the most urgently.14,15\n\nFurther study is needed to discover how each of these clinical characteristics influences quality of life. By doing so, it may be feasible to identify the elements that have the greatest impact on people's lives. Future research should look at the link between dental hypersensitivity, aesthetics, tooth deterioration, and quality of life.\n\nSeveral strategies may be used to assess the severity of various oral disorders in the population. They are often used in the community to minimize the occurrence of diseases and other ailments. The current study used an instrument such as the OHIP-14 to assess the prevalence of TW and its impact on well-being in diabetes. This is supported by other material that is currently available.16,17\n\n\nConclusions\n\nIn conclusion, the findings of this study provide evidence for the consequences of dental wear disorders in patients with diabetes and their implications on the standard of life of adults in the Dakshina Kannada community. Setting a goal and combining planning and self-monitoring in specific instructions will help you mark the behaviour change more successfully. These people may benefit from regular dental checkups and rigorous adherence to oral hygiene regulations. Future studies should concentrate on this topic.", "appendix": "Data availability\n\nFigshare: Tooth wear in diabetic population, https://doi.org/10.6084/m9.figshare.23635173. 18\n\nThis project contains the following underlying data:\n\n- Datasheet.xlsx\n\n- data-TW.xlsx\n\nFigshare: Tooth wear in diabetic population, https://doi.org/10.6084/m9.figshare.23635173. 18\n\nThis project contains the following extended data:\n\n- case proforma.docx\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nReferences\n\nSrisilapanan P, Jindarat M, Roseman J: The Prevalence and Severity of Tooth Wear in Type 2 Diabetic Patients. Int. J. Dent. 2018 Dec 11; 2018: 3608155–3608158. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOmar R, Johansson A, Johansson AK, et al.: Tooth wear. Int. J. Dent. 2012; 2012: 1. Article ID 731085. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nO’Toole S, Pennington M, Varma S, et al.: 'The treatment need and associated cost of erosive tooth wear rehabilitation - a service evaluation within an NHS dental hospital. BDJ. 2018; 224(12): 957–961. PubMed Abstract \| Publisher Full Text\n\nAl-Omiri MK, Lamey PJ, Clifford T: Impact of tooth wear on daily living. Int. J. Prosthodont. 2006; 2012(6): 601–605. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBomfim DI: Quality of Life of Patients with Different Levels of Tooth Wear. London, UK: Department Of Prosthodontics, Eastman Dental Institute At The University Of London; 2010[googlescholar]. M.Sc. thesis.\n\nKelleher MG, Bomfim DI, Austin RS: Biologically based restorative management of tooth wear. Int. J. Dent. 2012; 2012: 742509. Epub 2012 Jan 18. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBartlett D, Phillips K, Smith B: A difference in perspective the North American and European interpretations of tooth wear. Int. J. Prosthodont. 1999; 12(5): 401–408. PubMed Abstract \| Publisher Full Text\n\nNanditha A, Ma RC, Ramachandran A, et al.: Diabetes in Asia and the Pacific: Implications for the Global Epidemic. Diabetes Care. 2016 Mar; 39(3): 472–485. PubMed Abstract \| Publisher Full Text\n\nZimmet PZ, Magliano DJ, Herman WH, et al.: Diabetes: a 21st century challenge. Lancet Diabetes Endocrinol. 2014; 2: 56–64. Publisher Full Text\n\nPatiño MN, Loyola R, Medina S, et al.: Caries, periodontal disease and tooth loss in patients with diabetes mellitus types 1 and 2. Acta Odontol. Latinoam. 2007; 21(2): 127–133.\n\nAhmadinia AR, Rahebi D, Mohammadi M, et al.: Association between type 2 diabetes (T2D) and tooth loss: a systematic review and meta-analysis. BMC Endocr. Disord. 2022 Apr 13; 22(1): 100. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSlade GD: Derivation and validation of a short-form oral health impact profile. Community Dent. Oral Epidemiol. 1997 Aug; 25(4): 284–290. PubMed Abstract \| Publisher Full Text\n\nPreshaw PM, Alba AL, Herrera D, et al.: Periodontitis and diabetes: a two-way relationship. Diabetologia. 2012 Jan; 55(1): 21–31.PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPatel J, Baker SR: Is tooth wear associated with oral health related quality of life in adults in the UK?. Community Dent. Health. 2020 Aug 31; 37(3): 174–179. PubMed Abstract \| Publisher Full Text\n\nGanss C, Schlechtriemen M, Klimek J: Dental erosions in subjects living on a raw food diet. Caries Res. 1999; 33: 74–80. Publisher Full Text\n\nRao A, Shenoy R, Rao A: Periodontal health on the quality of life among diabetics. Int. J. Adv. Res. 2014; 2(6): 608–613.\n\nAcharya S, Bhat PV: Oral-health-related quality of life during pregnancy. J. Public Health Dent. 2009 Spring; 69(2): 74–77. Publisher Full Text\n\nSangeeta N, Sinha P, Sehgal L: Tooth wear in diabetic population. [Dataset]. figshare. 2023. Publisher Full Text" }	[ { "id": "218390", "date": "12 Apr 2024", "name": "Menaka Karalwad", "expertise": [ "Reviewer Expertise Periodontal Medicine", "Clinical Periodontics", "Lasers in Periodontics", "Periodontal Microbiology" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nA well written paper addressing the most common clinical conditions related to dental health. The study would be justified if authors can comment regarding selection of diabetic patients only as their sample and explain the influence of diabetes on tooth wear when compared to systemic healthy individuals otherwise. The study design can be described as descriptive cross-sectional single center study. In methodology the details on who performed the clinical examination-single/multiple examiners to avoid bias can be stated. Habit history of bruxism needs to be mentioned as its important in tooth wear. The age group 18-80 years, it would be interesting to know & explore the association between age groups for the main outcomes assessed.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "306570", "date": "12 Aug 2024", "name": "Sabin Siddique", "expertise": [ "Reviewer Expertise dental public health" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIt is a relevant study and its impact on quality of life of diabetic patients , as it is one of the frequently reported lifestyle disease in the country and wasting diseases based research is rarely reported among diabetic patients so it is important for dentists to understand the risk of diabetes on oral health.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1411
https://f1000research.com/articles/12-1410/v1	26 Oct 23	{ "type": "Case Report", "title": "Case Report: Effectiveness of physiotherapy rehabilitation in a patient with compound grade IIIC proximal third tibia fracture along with vascular injury", "authors": [ "Kamya Somaiya", "Subrat Samal", "Manali Boob", "Subrat Samal", "Manali Boob" ], "abstract": "The proximal metaphyseal region of the bone is thought to be the location of fractures of the proximal tibia. The surgical management of proximal tibial shaft fractures and high-energy bicondylar tibial plateau fractures is still challenging . Supra-cutaneous plating, also known as a locking compression plate, has been promoted as a crucial technique in the management of challenging reconstructive scenarios since it has shown to be adjustable, concealed, and well-accepted by patients. Physical therapy rehabilitation is critically necessary for a full recovery from a range of orthopaedic issues. Physical therapy rehabilitation is a vital step in the healing process for patients with lower limb fractures. In our study , we rehabilitated a patient with Compound grade IIIC proximal third tibia fracture right side managed with Closed reduction external fixator application and vascular repair along with proximal fibula fracture. Through this study, we developed a physiotherapy rehabilitation protocol that was very helpful for these patients, produced positive results, and served to enhance the patient's quality of life.", "keywords": [ "Proximal tibia fracture", "Compound Grade III C proximal tibia fracture", "Supracutaneous plating", "Physiotherapy rehabilitation", "Quality of Life." ], "content": "Introduction\n\nFractures of the proximal tibia are traditionally suspected to be those that extend into the bone’s proximal metaphyseal area. Despite the fact that fractures can result from relatively low energy mechanisms like sports injuries or falls from a standing position, high-energy trauma is more frequently linked to fractures. A widely used technique for evaluating fractures is the OTA/AO categorization of peri-articular fractures. A type A fracture is one that is entirely extra-articular. Type B injuries are intra-articular fractures that have a piece of the metaphysis that is still intact (partial articular injuries). Type C intra-articular fractures are those that have complete metaphyseal-diaphyseal separation. Every subgroup is further classified. The risk of concurrent vascular injury is highest in distal femur and proximal tibia displacement fractures, and dislocation of the knee is particularly associated with insufficient collateral circulation to sustain the distal limb. A high index of suspicion, aggressive angiography implementation, and tight collaboration between all trauma team members are necessary for the best care of such injuries.1\n\nHigh-energy bicondylar tibial plateau fractures and proximal tibial shaft fractures are still difficult to treat surgically. Bicondylar tibial plateau fractures frequently result in difficulties with the wound, infections, varus collapse, knee stiffness, and articular mal reductions.2 Among the operative modalities are intramedullary nailing, external fixation with or without restricted internal fixation, open reduction and internal fixation, and external fixation alone. Since supra-cutaneous plating, also known as a locking compression plate, has proven to be adaptable, concealed, and well embraced by patients, it has been advocated as an essential tool in the treatment of difficult reconstructive situations.3\n\nIn patients with lower limb fractures, physical therapy rehabilitation is a highly important part of the healing process. In a study, Iliopoulas et al. demonstrated the value of early weight-bearing and range-of-motion exercises in tibial plateau fracture patients. In order to get better clinical results, continued rehabilitation should be taken into account.4 This review thus clarifies the significance of rehabilitation for people experiencing tibial plateau fractures. We are reporting the case of a 42-year-old man who had a compound grade IIIC proximal third tibia fracture on the right side that was addressed with closed reduction external fixation and vascular repair. Along with that the patient also had fibula fracture. This patient received a suitable physiotherapeutic regimen, and results of its efficacy were observed.\n\n\nCase presentation\n\nWe are reporting a case of a 42-year-old male who is a farmer by occupation. Prior to his road traffic collision, the patient was in good health one month prior to hospital admission. The patient was returning from Yavatmal to Ghatanji. Around 6:30 p.m. (7/04/23) he crashed into a four-wheeler while returning to Tivsa and fell off the bike, suffering a right leg injury. He was unable to put weight on his right limb instantaneously following the injury. After that, he was transferred to Yavatmal government hospital where first wound care was provided. He was referred to Acharya Vinoba Bhave Rural Hospital (AVBRH) for follow-up care after the therapy. Numerous diagnostics were done in this case, including X-rays that showed a compound grade IIIC proximal third tibia fracture on the right side with a vascular deficit. Extracutaneous plating of the right proximal tibia was conducted on 8/04/23 for the same reason. On 10/04/23, subsequent angiogram revealed acute thrombotic blockage of the distal ATA and DPA. On the same day for the precise same reason, right limb intra-arterial thrombolysis was conducted. Since that time, the patient has reported of pain and a difficulty to move their right leg. On 13/04/23, a demand for physiotherapy was made, and since then, sessions have been held. The sequence of events is depicted in Figure 1.\n\nAfter the patient provided his informed consent, the physical examination was conducted. The patient was alert, cooperative, and clearly aware of time, place, and people. A comprehensive musculoskeletal assessment was performed. The history of pain is shown in Figure 2. Patient has a history of diabetes and hypertension for six months. On Observation, there was edema present over the distal leg of the right side (Figure 3). The following findings were discovered during an examination of the right lower limb. On inspection, dressing present over medial aspect of the leg and plate with screws present over the lateral aspect of the leg. On palpation, wound present over the medial aspect of the leg of 7 cm (Figure 4), grade 1 edema present over the right distal leg, grade 1 tenderness present over the completer course of tibia over the medial aspect of right leg and over the complete lateral aspect of leg along the plating. The range of motion and manual muscle testing of the left side were completely normal. The range of motion and manual muscle testing of the right side are shown in Tables 2 and 3, respectively.\n\nDiagnostic examination was done using angiogram, colour doppler and X-ray. Right lower limb angiogram revealed acute thrombotic occlusion of the distal ATA and DPA. Colour Doppler revealed that distal DPA shows no flow. Post-operative X-ray of right knee joint (Figure 5) revealed compound Grade 3 C fracture of tibia and fibular fracture with mild displacement. The X-rays shows extracutaneous plating with three screws at the proximal and three screws at distal aspect of the tibia.\n\nThe therapeutic regimen’s main objective is to boost the patient’s quality of life. We recorded the outcome measures pre- treatment and post-treatment in order to evaluate the effectiveness of our treatment regimen. All exercises are given in 1 set of 10 repetitions each and further progressed.5 Table 1, Table 2 and Table 3 explains the therapeutic regimen for Week 1 to Week 12. Figure 6 depicts patient performing non-weight bearing walker-assisted ambulation.\n\n\n\n• Counselling patients and their families on the value of physical therapy rehabilitation and the necessary maintenance of rehabilitation expectations\n\n\n\n• Prior to the therapy session starting.\n\n\n\n• Deep breathing exercises\n\n• Incentive Spirometry\n\n\n\n• Deep Breathing exercises while recieving the physiotherapist’s visual and auditory feedback. 5 cycles of 5 to 6 breaths each.9\n\n• Using incentive spirometer, inspiratory muscle training is done once in every two hours.10\n\n\n\n• Cryotherapy\n\n• Ankle pumps with leg elevation\n\n\n\n• It is applied for 10 minutes.7\n\n• 1 set of 10 repetitions.\n\n\n\n• Assisted Active Range of motion exercises\n\n\n\n• 1 set of 10 repetitions. Progression to Active ROM exercises further.\n\n\n\n• Muscle-setting exercises of the quadriceps, hamstrings and adductors.\n\n• Assisted progression to active SLRs in supine.\n\n\n\n• Each exercise is performed for 10 repetitions in 1 set\n\n\n\n• The following transitions are taught:\n\n➢ Supine to side lying transition\n\n➢ Side lying to sitting transition\n\n➢ Sit to stand transition\n\n\n\n• The transitions are taught sequentially and progressed as per the patient.\n\n\n\n• Upper limb resistance exercises using 0.5 kg weight cuff.\n\n\n\n• 1 set of 10 repetitions is given. Progression done by increasing weight.\n\n\n\n• Non-weight bearing walker assisted ambulation is taught (Figure 6).\n\n\n\n• As per the tolerance of the patient.\n\n\n\n• Patellar mobilization.\n\n\n\n• 1 set of 10 to 12 glides are administered.\n\n\n\n• Lower limb resistance exercises using 0.5 kg weight cuff.\n\n\n\n• 1 set of 10 repetitions is given. Progression done by increasing weight.\n\n\n\n• Partial weight bearing ambulation by putting 25% weight on the right lower limb.\n\n\n\n• As per the tolerance of the patient.\n\n\n\n• Dynamic Quadriceps\n\n• Dynamic Hamstrings\n\n\n\n• 1 set of 10 repetitions were performed. Progression was done by increasing the repetitions and sets.\n\n\n\n• Core strengthening exercises\n\n➢ Abdominal crunches and Curl up’s\n\n➢ Unilateral Pelvic Bridging\n\n➢ Unilateral Pelvic Bridging using physio ball11\n\n\n\n• 1 set of 10 repetitions were performed. Progression was done by increasing the repetitions and sets.\n\n\n\n• Proprioceptive exercises:\n\n➢ Sitting on Swiss Ball doing weight shifts\n\n➢ Sitting on swiss ball lifting one foot.\n\n➢ Standing and doing weight shifts.12\n\n\n\n• 1 set of 10 repetitions were performed. Progression was done by increasing the repetitions and sets.\n\n\n\n• Partial weight bearing ambulation by putting 50% weight on the right lower limb.\n\n\n\n• As per the tolerance of the patient.\n\n\n\n• Muscle Energy Technique and\n\n• Maitland Mobilization for knee range of motion.\n\n\n\n• 1 set of 10 repetitions were performed.\n\n• 1 set of 10 to 12 glides are administered.\n\nWe utilized various outcome measures to analyse the results of our treatment regimen. Figure 7, Table 4 and Table 5 depicts the pre-rehabilitation scores and post-rehabilitation scores.\n\n\nDiscussion\n\nFor a full recovery from a variety of orthopaedic problems, physical therapy rehabilitation is absolutely essential. In addition to pre-operative conditions, post-operative rehabilitation is also a crucial element for patients’ full recovery and the enhancement of their quality of life. The primary goals of post-operative physical therapy are to enable patients to resume their ADLs without difficulty and to enhance their quality of life. Numerous studies have been conducted to demonstrate the value of physiotherapy treatment plans for a range of orthopaedic disorders. In one of their studies, Lalwani et al. demonstrated the significance of the post-fracture rehabilitation programme with significant improvements in the patient’s quality of life, well-being, and physical functioning.6 Similar studies have been done on lower limb fractures, including proximal tibial fractures, and they reveal that these patients benefit greatly from their physiotherapy routine. Nowadays a novel method called closed reduction external fixation using supracutaneous plating for the proximal tibial fractures is carried out. A specific type of low incidence tibial plateau fracture is the bi-condylar fracture. The traditional surgical strategy and rapid, systematic physical therapy rehabilitation improved the functional goals over time, which is a crucial factor in helping such post-operative patients recover.7 The patient was able to regain his functional independence at home and at work because the rehabilitation protocol showed significant reduction of discomfort and pain, greater range of motion, and increased muscle strength and endurance.8 Therefore, post-operative physiotherapy rehabilitation is crucial for these individuals. In our study, we rehabilitated a patient with compound grade IIIC proximal third tibia fracture right side managed with closed reduction external fixator application and vascular repair along with proximal fibula fracture. Our treatment plan produced positive outcomes and enhanced the patient’s quality of life. We developed a physiotherapy program through this study that will be very helpful for these individuals and could be applied to similar patients in the future.\n\n\nConclusion\n\nHigh energy tibial plateau fractures are treated surgically utilising a variety of techniques, one of which is closed reduction and external fixation with supracutaneous plating. Recovery for such patients after surgery depends greatly on postoperative therapy. Through this study, we developed a physiotherapy routine that was very helpful for these patients, produced positive results, and served to enhance the patient’s quality of life.\n\n\nConsent\n\nWritten informed consent for publication of their clinical details and/or clinical images was obtained from the patient/parent/guardian/relative of the patient.", "appendix": "Data availability\n\nThere are no data related to this topic.\n\nZenodo. CARE checklist. DOI: https://doi.org/10.5281/zenodo.8270235\n\nData are available under the terms of the Creative Commons Attributions 4.0 International License (CC-BY 4.0).\n\n\nAcknowledgements\n\nWe thank the patient who co-operated with us during his treatment and consented to publish his case report for future references and our teachers to motivate us to do so.\n\n\nReferences\n\nCone JB: Vascular Injury Associated With Fracture-Dislocations of the Lower Extremity. Clin. Orthop. Relat. Res. 1976-2007. 1989 Jun [cited 2023 Jul 2]; 243: 30–35. Publisher Full Text Reference Source\n\nCole PA, Zlowodzki M, Kregor PJ: Treatment of Proximal Tibia Fractures Using the Less Invasive Stabilization System: Surgical Experience and Early Clinical Results in 77 Fractures. J. Orthop. Trauma. 2004 Sep [cited 2023 Jul 2]; 18(8): 528–535. Publisher Full Text Reference Source\n\nAhmed S, Deshpande S, Chhatbar K, et al.: Extracutaneous Locking Plate As An External Fixation Device In Juxta Articular Compound Fractures- Review. J. Pharm. Negat. Results. 2022 Nov 27 [cited 2023 Jul 2];2995–2997. Reference Source\n\nIliopoulos E, Galanis N: Physiotherapy after tibial plateau fracture fixation: A systematic review of the literature. SAGE Open Med. 2020 Jan 1 [cited 2023 Jul 2]; 8: 2050312120965316. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMedicine AC of S: ACSM’s Guidelines for Exercise Testing and Prescription. Lippincott Williams & Wilkins; 2013; 481.\n\nLalwani SS, Jain DS, Phansopkar PA, et al.: Physiotherapy Rehabilitation to Recuperate a Patient From an Intertrochanteric Fracture: A Case Report. Cureus. 2022 Aug 3 [cited 2023 Jul 2]; 14: e27660. PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source\n\nBawiskar D, Dhote S, Phansopkar P: Early physical rehabilitation post-surgery in a complex type 5 Schatzker Tibial plateau fracture improves functional outcomes: A case report. Med. Sci. 2020; 24(104): 2675–2682.\n\nNawkhare AV, Jawade S, Wadhokar OC: Effect of Novel based Postoperative physiotherapy rehabilitation on functional outcome in Malunited supracondylar fracture of femur and proximal tibia fracture following implant removal.11(2320).\n\nOrfanos P, Ellis E, Johnston C: Effects of deep breathing exercises and ambulation on pattern of ventilation in post-operative patients. Aust. J. Physiother. 1999; 45(3): 173–182. PubMed Abstract \| Publisher Full Text\n\nEltorai AEM, Szabo AL, Antoci V, et al.: Clinical Effectiveness of Incentive Spirometry for the Prevention of Postoperative Pulmonary Complications. Respir. Care. 2018 Mar [cited 2023 Jul 3]; 63(3): 347–352. PubMed Abstract \| Publisher Full Text\n\nAkuthota V, Nadler SF: Core strengthening11No commercial party having a direct financial interest in the results of the research supporting this article has or will confer a benefit upon the author(s) or upon any organization with which the authors is/are associated. Arch. Phys. Med. Rehabil. 2004 Mar [cited 2023 Jul 23]; 85: 86–92. Publisher Full Text Reference Source\n\nClausen B, Holsgaard-Larsen A, Roos EM: An 8-Week Neuromuscular Exercise Program for Patients With Mild to Moderate Knee Osteoarthritis: A Case Series Drawn From a Registered Clinical Trial. J. Athl. Train. 2017 Jun 1 [cited 2023 Jul 24]; 52(6): 592–605. PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source" }	[ { "id": "243895", "date": "22 Feb 2024", "name": "Uzair Yaqoob", "expertise": [ "Reviewer Expertise Neurological and orthopedic surgery and trauma" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThank you for giving me the opportunity of reviewing your manuscript. Overall, I think it is well formulated, written with not much of the technical issues. However, I have some comments to make and I think these will make your study better. The authors claim that they developed a routine that can be helpful for the patients getting operated after high energy tibial plateau fractures. I believe this routine should be applied to more patients and the results should be presented in an observational study to further make this routine as a standard management measure.\n\nABSTRACT: Adequate and to the point as it should be 1. \"..with Compound grade IIIC proximal third tibia fracture right side managed..\" REPHRASE\nINTRODCUTION: 1. First paragraph talks about the fractures and the second one talks about the surgical treatment. I don't think a dedicated paragraph for the surgical treatment is necessary with this topic, but if we do keep it, there should be some more talk about the rehabilitation process and physiotherapy.. Add more references in first paragraph 2. \"..was addressed with closed reduction external fixation and vascular repair..\", rephrase to \"..was addressed with closed reduction, external fixation, and vascular repair..\"\nCASE PRESENTATION: 1. it was a good idea to share the timeline of events as a chart 2. Good figures 3. \"..revealed compound Grade 3 C fracture of tibia and fibular fracture with mild displacement. ..\". REPHRASE Very well structured .\n\nIs the background of the case’s history and progression described in sufficient detail? Yes\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Yes\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Yes\n\nIs the case presented with sufficient detail to be useful for other practitioners? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1410
https://f1000research.com/articles/12-1409/v1	26 Oct 23	{ "type": "Study Protocol", "title": "A cross-sectional study on the prognostic value of the RAPID score in pleural infection in patients attending a tertiary care hospital in Central India", "authors": [ "Srinivasulareddy Annareddy", "Babaji Ghewade", "Babaji Ghewade" ], "abstract": "Pleural infections pose a significant clinical challenge, with diverse outcomes that are often difficult to predict. The renal (urea), age, fluid purulence, infection source, dietary (albumin) (RAPID) score, a clinical tool designed to assess the risk of adverse outcomes in pleural infections, holds the potential as a prognostic indicator. This study aims to evaluate the prognostic value of the RAPID score in patients with pleural infections attending a tertiary care hospital in Central India. This hospital-based prospective cross-sectional observational study will span from July 2022 to June 2024, enrolling 50 adult patients aged 18 years and older admitted to the Department of Respiratory Medicine, a tertiary care hospital in central India. Data will be collected using a structured proforma, encompassing demographic data, clinical history, and comorbidities. Diagnostic investigations, including pleural fluid analysis, laboratory tests, sputum analysis, and radiological assessments, will be performed upon enrolment. RAPID scores will be calculated at admission, stratifying patients into low, medium, and high-risk categories. Treatment will follow established pleural infection protocols, with patients receiving empirical antibiotics and tailored treatment based on culture sensitivity results. Patients will be monitored for three months post-admission, and outcomes such as hospital stay duration, tube thoracostomy need, intercostal drainage tube duration, medical management success, surgical referral rates, surgical interventions, and 30-day and 90-day mortality rates will be assessed. This study aims to contribute valuable insights into the prognostic value of the RAPID score in pleural infections and factors influencing patient outcomes. The findings may facilitate more informed clinical decision-making and improve the management of pleural infections, ultimately enhancing patient care and outcomes.", "keywords": [ "Pleural infection", "RAPID score", "prognosis", "diagnostic investigations", "treatment protocol", "patient outcomes", "Central India." ], "content": "Introduction\n\nPleural infections, commonly called pleuritis or pleurisy, are inflammatory conditions affecting the pleura, the thin double-layered membrane surrounding the lungs and lining the chest cavity. These infections present a challenging clinical scenario characterised by a spectrum of etiologies, varied clinical presentations, and outcomes that range from mild and self-limiting to severe and life-threatening. Managing pleural infections demands a nuanced understanding of the disease process, prompt diagnosis, and appropriate therapeutic interventions.1\n\nCentral to the effective management of pleural infections is the accurate assessment of disease severity and the prediction of clinical outcomes. Prognostic tools that can reliably stratify patients based on their risk of complications, such as prolonged hospitalisation, the need for invasive procedures like tube thoracostomy, or adverse events like mortality, are invaluable in guiding clinical decision-making and optimising patient care.2,3\n\nThe renal (urea), age, fluid purulence, infection source, dietary (albumin) (RAPID) score (a risk score for adverse outcomes in pleural infection) has emerged as one tool designed to assist clinicians in assessing the risk of adverse outcomes in patients with pleural infections. It encompasses a set of clinical parameters that, when combined, generate a numerical score reflecting the patient's risk profile. While the RAPID score has shown promise in predicting outcomes in pleural infections, its utility in the specific context of Central India remains an open question. The region's unique patient demographics, disease patterns, and healthcare infrastructure may introduce variables that impact the score's accuracy and applicability.4,5\n\nThis study endeavours to fill this critical knowledge gap by conducting a comprehensive investigation into the prognostic value of the RAPID score in patients with pleural infections attending a tertiary care hospital in Central India. By assessing the performance of the RAPID score in this specific population, we aim to provide valuable insights into its reliability as a prognostic tool and its potential role in optimising patient management strategies. The study also seeks to identify other factors that may influence patient outcomes in pleural infections, contributing to a more holistic understanding of this complex clinical entity.\n\nThrough rigorous data collection, analysis, and correlation with clinical outcomes, this research aspires to provide clinicians with a refined prognostic tool to enhance their ability to predict and manage pleural infections effectively. Ultimately, this pursuit of knowledge can improve patient care, reduce complications, and enhance healthcare delivery in the realm of pleural infections in Central India.\n\nTo assess the prognostic value of the RAPID score in patients with pleural infections attending a tertiary care hospital in Central India.\n\n\n\n1. To determine the prognostic accuracy of the RAPID score in predicting outcomes of patients with pleural infections.\n\n2. To identify and analyse factors associated with poor outcomes, such as prolonged hospitalisation, the need for tube thoracostomy, surgical intervention, and mortality in patients with pleural infections.\n\n\nProtocol\n\nThis study will adopt a hospital-based prospective design, specifically a cross-sectional observational study.\n\nThe study will be conducted from July 2022 to June 2024. All eligible patients presenting with pleural infection at the Department of Respiratory Medicine, tertiary care hospital of central India, Wardha, will be considered for inclusion during this period.\n\nThe study population will comprise patients who present with pleural infection and meet the inclusion and exclusion criteria while seeking admission to the Department of Respiratory Medicine, Tertiary Care Hospital of Central India, Wardha.\n\nA total of 50 patients will be included in the study. This sample size has been determined based on feasibility and available resources.\n\nInclusion criteria\n\n1. Adult patients aged 18 years or older.\n\n2. Patients diagnosed with pleural infection.\n\nExclusion criteria\n\n1. Patients who declined to participate in the study.\n\n\n\n\nA structured proforma6 will gather comprehensive patient data. This will encompass demographic information, detailed clinical history, and documentation of any existing comorbidities. Furthermore, a thorough physical examination will be conducted during the data collection.\n\nUpon enrolment in the study, all patients will undergo an extensive battery of diagnostic investigations, including:\n\n1. Diagnostic thoracentesis under ultrasound guidance\n\n• Pleural fluid will be aspirated using ultrasound guidance.\n\n• The collected pleural fluid will undergo analysis, which includes assessment of total cell count, differential cell count, protein levels, glucose levels, lactate dehydrogenase (LDH) levels, adenosine deaminase (ADA) levels, acid-fast bacillus (AFB) staining, cartridge-based nucleic acid amplification test (CBNAAT) or true nucleic acid amplification test (truNAAT), gram staining, culture sensitivity testing, and cytological examination.\n\n2. Laboratory investigations\n\n• A complete blood picture (CBC) will be conducted.\n\n• Renal function tests (RFT), liver function tests (LFT), random blood sugar (RBS), serum albumin levels, and HIV screening will also be performed.\n\n3. Sputum analysis\n\n• Sputum samples will be obtained for CBNAAT/truNAAT testing, gram staining, and culture sensitivity analysis.\n\n4. Radiological assessment\n\n• Chest X-rays will be performed as a standard procedure. If clinically indicated, computed tomography (CT) scans of the chest will also be conducted.\n\nRAPID scores will be calculated for each patient upon admission. Based on their calculated scores, these scores will be used to classify patients into risk groups, including low, medium, and high risk.7\n\nPatients will receive treatment per the standard protocol for managing pleural infections. This includes:\n\n• Empirical antibiotics: Patients will receive an initial course of empirical antibiotics, with subsequent adjustments based on the results of culture sensitivity testing. Initial antibiotic regimens may include penicillins with β-lactamase inhibitors or cephalosporins in conjunction with metronidazole or clindamycin. Tuberculosis patients will be treated with anti-tubercular therapy (ATT).\n\n• Tube thoracostomy: Patients who require tube thoracostomy will undergo this procedure as needed, as determined by clinical evaluation.\n\nPatients will be closely monitored over three months following admission. The study's primary outcomes will be assessed, including:\n\n• Length of hospital stay.\n\n• Incidence of tube thoracostomy.\n\n• Duration of intercostal drainage tube placement.\n\n• Success or failure of medical management.\n\n• Rates of surgical referrals and interventions.\n\n• Mortality rates at 30 days (1 month) and 90 days (3 months).\n\n\n\n1. Selection bias:\n\n• Bias: Patients who consent to participate in the study may differ systematically from those who do not, leading to a biased sample.\n\n• Overcoming bias: To minimise selection bias, ensure a representative sample by striving for consecutive enrolment of eligible patients. Additionally, conduct sensitivity analyses to assess the impact of potential selection bias on the results.\n\n2. Measurement bias:\n\n• Bias: Errors or inaccuracies in measuring variables, such as laboratory tests, can introduce bias.\n\n• Overcoming bias: Implement rigorous quality control procedures for data collection and measurement. Standardise procedures and use well-validated instruments and methods. Regularly calibrate equipment and train personnel involved in data collection.\n\n3. Recall bias:\n\n• Bias: Patients or healthcare providers may have difficulty recalling past events or may selectively remember information, leading to recall bias.\n\n• Overcoming bias: Minimize recall bias by collecting data prospectively whenever possible. Use structured questionnaires and electronic medical records to reduce reliance on memory. Ensure that patients are interviewed or examined by personnel blinded to the study objectives.\n\n4. Information bias:\n\n• Bias: Incomplete or missing data can introduce bias if certain patients are more likely to have missing data.\n\n• Overcoming bias: Implement comprehensive data collection procedures and strive for complete data capture. Use appropriate statistical methods to handle missing data, such as multiple imputation or sensitivity analyses.\n\nIn this study protocol investigating the prognostic value of the RAPID score in pleural infections, various statistical methods will be employed to analyse the collected data and extract meaningful insights. Descriptive statistics will be used to summarise patient demographics and clinical variables, including mean, median, standard deviation, and frequency distributions. Bivariate analyses will explore associations between the RAPID score and different clinical outcomes and factors, employing statistical tests appropriate for the data type. Multivariate analyses, encompassing regression models, will assess the independent prognostic significance of the RAPID score while accounting for potential confounding variables. Survival analysis techniques like Kaplan-Meier curves and Cox proportional hazards regression will be applied for time-to-event outcomes. Receiver Operating Characteristic (ROC) analysis will gauge the RAPID score's discriminative ability. Subgroup and sensitivity analyses will be conducted to investigate variations in prognostic value and the robustness of the results.\n\nThe Institutional Ethics Committee of Datta Meghe Institute of Higher Education and Research (DU) has granted its approval to the study protocol (Reference number: DMIHER (DU)/IEC/2022/18). Before commencing the study, we will obtain written informed consent from all participants, providing them with a comprehensive explanation of the study's objectives.\n\nAfter the completion of the study, we will publish it in an indexed journal or conference.\n\nThe study has not yet started. After the publication of the protocol, we will start recruitment for the study.\n\n\nDiscussion\n\nPleural infections, although relatively common, remain a diagnostic and therapeutic challenge due to their diverse etiologies, varied clinical presentations, and unpredictable outcomes.8 The ability to accurately prognosticate and stratify patients based on their risk of adverse events is crucial for clinicians managing these cases effectively. In this study, we sought to evaluate the prognostic value of the RAPID score in predicting outcomes in patients with pleural infections attending a tertiary care hospital in Central India.5\n\nThe RAPID score, designed as a risk assessment tool for pleural infections, comprises easily accessible clinical parameters that aim to predict adverse outcomes such as prolonged hospitalisation, the need for invasive procedures, and mortality. The assessment of this score at admission allowed us to classify patients into low-, medium, and high-risk groups.9\n\nThe findings of this study will offer insights into the utility of the RAPID score in this specific demographic and clinical context. The ability of the score to reliably identify patients at higher risk of adverse outcomes can significantly impact clinical decision-making. For instance, it can aid in prioritising resources for high-risk patients, intensifying monitoring, and guiding the choice and timing of therapeutic interventions.\n\nBeyond the RAPID score, this study aims to identify additional factors influencing patient outcomes in pleural infections. These factors may include comorbidities, microbial aetiology, response to empirical antibiotics, and specific radiological or laboratory findings. By analysing these variables with the RAPID score, we aim to provide a more comprehensive understanding of the complex interplay of factors in pleural infection management.10\n\nThe results of this study can have important clinical implications. If the RAPID score is a reliable prognostic tool in Central India, it can be integrated into routine clinical practice, aiding in risk stratification and resource allocation. This, in turn, can contribute to improved patient outcomes and resource utilisation in managing pleural infections.11\n\nSeveral limitations should be acknowledged in this study. The relatively modest sample size of 50 patients may affect the generalizability of the findings. Additionally, the study's single-center design may limit the external validity of the results. Moreover, the complex nature of pleural infections, which can involve various pathogens and clinical presentations, may introduce heterogeneity that necessitates careful consideration during data analysis.", "appendix": "Data availability\n\nNo underlying data are associated with this article.\n\nZenodo: A cross-sectional study on the prognostic value of rapid score in pleural infection in patients attending tertiary care hospital of central India. https://doi.org/10.5281/zenodo.8340291. 6\n\nThis project contains the following extended data:\n\n• Tool.docx (Patient Information and Data Collection Proforma)\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nKarkhanis VS, Joshi JM: Pleural effusion: diagnosis, treatment, and management. Open Access Emerg Med. 2012; 4: 31–52. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHassan M, Patel S, Sadaka AS, et al.: Recent Insights into the Management of Pleural Infection. Int J Gen Med. 2021; 14: 3415–3429. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRosenstengel A: Pleural infection-current diagnosis and management. J. Thorac. Dis. 2012; 4: 186–193. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWhite HD, Henry C, Stock EM, et al.: Predicting Long-Term Outcomes in Pleural Infections. RAPID Score for Risk Stratification. Ann. Am. Thorac. Soc. 2015; 12: 1310–1316. Publisher Full Text\n\nRahman NM, Kahan BC, Miller RF, et al.: A clinical score (RAPID) to identify those at risk for poor outcome at presentation in patients with pleural infection. Chest. 2014; 145: 848–855. PubMed Abstract \| Publisher Full Text\n\nAnnareddy S: A Cross-Sectional Study on the Prognostic Value of RAPID Score in Pleural Infection (Version v1). [Dataset]. Zenodo. 2023. Publisher Full Text\n\nRapsang AG, Shyam DC: Scoring systems in the intensive care unit: A compendium. Indian J. Crit. Care Med. 2014; 18: 220–228. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFantin A, Castaldo N, Vailati P, et al.: Pleural effusion aetiology, presentation, treatment and outcome in haematological malignancies, IgG4-related disease, chronic myeloproliferative diseases, and haemoglobinopathias: a review. Acta Biomed. 2021; 92: e2021268. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCorcoran JP, Psallidas I, Gerry S, et al.: Prospective validation of the RAPID clinical risk prediction score in adult patients with pleural infection: the PILOT study. Eur. Respir. J. 2020; 56: 2000130. PubMed Abstract \| Publisher Full Text\n\nGonçalves-Pereira J, Conceição C, Póvoa P: Community-acquired pneumonia: identification and evaluation of nonresponders. Ther. Adv. Infect. Dis. 2013; 1: 5–17. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGirwar SM, Jabroer R, Fiocco M, et al.: A systematic review of risk stratification tools internationally used in primary care settings. Health Sci. Rep. 2021; 4: e329. Publisher Full Text" }	[ { "id": "303411", "date": "24 Sep 2024", "name": "Manas Pratim Roy", "expertise": [ "Reviewer Expertise Public Health" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe scoring system may be detailed, including cut-offs for different categories.\nSince it is yet to be validated for India, there is a need for comparison with the existing standard/ gold standard.\nThe part on bias is theoretical and may be omitted.\n“The study will be conducted from July 2022 to June 2024. All eligible patients presenting with pleural infection at the Department of Respiratory Medicine, tertiary care hospital of central India, Wardha, will be considered for inclusion during this period.” and “A total of 50 patients will be included in the study” – these two sentences are not similar.\n\nIs the rationale for, and objectives of, the study clearly described? Yes\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Not applicable", "responses": [] } ]	1	https://f1000research.com/articles/12-1409
https://f1000research.com/articles/12-1405/v1	25 Oct 23	{ "type": "Case Report", "title": "Case Report: Paroxysmal autonomic instability with dystonia syndrome: a rare complication of tuberculous meningitis", "authors": [ "Lowrence Precious Dichoso", "Gerald Pagaling", "Roland Dominic G. Jamora", "Veeda Michelle M. Anlacan", "Lowrence Precious Dichoso", "Gerald Pagaling", "Roland Dominic G. Jamora" ], "abstract": "Paroxysmal autonomic instability with dystonia syndrome (PAIDS) is a rare and life-threatening complication of neurologic diseases. We report the case of a 20-year-old male with acute severe brain damage from tuberculous meningitis, who eventually developed paroxysmal episodes of spontaneous and inducible tachycardia, tachypnea, hypertension, and decerebrate posturing. We diagnosed the patient as suffering from paroxysmal autonomic instability with dystonia syndrome. The unavailability of morphine and the prohibitive cost of prolonged fentanyl use led to a trial of gabapentin, clonazepam, and propranolol for the patient, resulting in symptom resolution. Brain injury causes dysfunction of autonomic centers leading to paroxysmal autonomic instability with dystonia syndrome. Management includes minimizing stimulation and pharmacotherapy with preventive and abortive medications. Alternatives like gabapentin, propranolol and clonazepam were effective in treating the paroxysmal episodes, proving that they may have a role in resource limited settings. PAIDS requires urgent recognition and treatment to prevent further complications and death.", "keywords": [ "Paroxysmal autonomic instability with dystonia case report", "Tuberculous meningitis", "Dystonia", "PAIDS" ], "content": "Introduction\n\nParoxysmal autonomic instability with dystonia syndrome (PAIDS) is a relatively uncommon complication of various central nervous system diseases presenting as a constellation of episodic autonomic hyperactivity accompanied by dystonia, occurring at least one cycle per day for more than three days.1–3 The autonomic dysfunction is caused by disruption of diencephalic connections causing disinhibition or increased activation of the lower sympathetic activating center.1,4,5 The dystonic posturing is from a midbrain lesion causing disconnection of the normal inhibitory signals to the pontine and vestibular nuclei.4 We report a patient with PAIDS secondary to tuberculous meningitis, successfully managed with gabapentin, clonazepam, and propranolol.\n\n\nCase report\n\nA 20-year-old Filipino male student without comorbidities was admitted to our institution for a three-week history of behavioral changes accompanied by incoherent speech, waddling gait, febrile episodes, anorexia, and generalized weakness. He had two episodes of generalized tonic seizures lysed by diazepam at the emergency room. On examination, he had no eye-opening to pain, non-purposeful movements of all extremities, bilateral extensor toe sign and a supple neck. Chest X-ray revealed reticular opacities on both upper lung fields. A cranial computed tomography (CT) scan with contrast was suggestive of tuberculous meningitis with cerebritis (see Figure 1A-C). The lumbar tap revealed a clear yellow cerebrospinal fluid (CSF) with elevated opening pressure, pleocytosis of 8×106/L (neutrophils 20%, lymphocytes 60%), elevated protein of 687 mg/dL, hypoglycorrhachia of 1% of serum, and a negative Cryptococcal Antigen Latex Agglutination System, Bactigen Latex Agglutination, and fungal culture. He was diagnosed with disseminated tuberculosis with brain and lung involvement and was started on isoniazid/rifampicin/pyrazinamide/ethambutol tablets plus dexamethasone. A therapeutic lumbar tap was done every 2-3 days in the absence of consent for a ventriculoperitoneal shunt (VPS) placement.\n\nCranial computed tomography scan with contrast (A-C): Fairly defined focal areas of hypodensities seen in the deep white matter of the bilateral basofrontal parasagittal lobes, head of the left caudate nucleus, and anterior limb of the left internal capsule (A,B: white arrows), with undue leptomeningeal enhancement and mild communicating hydrocephalus (A,C: blue arrows).\n\nCranial MRI with contrast (D-I): Hyperintense foci with some areas of restricted diffusion and magnetic susceptibility artifacts with surrounding vasogenic edema in the cortical-subcortical junction of the bilateral basal ganglia, thalamus, midbrain and pons, bilateral temporal, bilateral parietal, left occipital lobes and bilateral frontal lobes (D-I: red arrows); with leptomeningeal and pachymeningeal nodular thickening and enhancement.\n\nOn the tenth hospital day, the patient developed spontaneous and inducible paroxysmal episodes of tachycardia (>130 beats/minute), tachypnea (>40 breaths/minute), hypertension (150/90 mmHg) with associated extensor posturing of both upper extremities lasting for 1-3 minutes and occurring 3-5 times a day. These were noted during patient manipulations like suctioning and bed turning. The Paroxysmal Sympathetic Hyperactivity Assessment Measure (PSH-AM) was used. This is composed of an 11-point Diagnosis Likelihood Tool (DLT) and a Clinical Feature Scale (CFS) to guide the diagnosis, which has a corresponding likelihood and severity score, respectively.1 With this criterion, our case was very likely to be PAID syndrome with a total score of more than 17 (DLT = 8 points + CFS = 13 points, with a final PSH-AM score of 21).4 Cranial magnetic resonance imaging with contrast revealed a further extension of the tuberculous abscess and non-resolution of hydrocephalus (see Figure 1D-I). A 21-channel electroencephalogram revealed diffuse slowing of background activity, absence of epileptiform discharges with no electrographic correlate of the extensor posturing of both upper extremities, likely dystonia (see Figure 2A-C). Fentanyl drip was started due to unavailability of morphine. However, this was eventually shifted to gabapentin 600 mg three times a day and propranolol 30 mg four times a day. The uptitration of propranolol was limited by episodes of hypotension hence clonazepam 2 mg twice a day was added. The frequency and duration of the paroxysms was reduced to 1-2 episodes per day. By the fifth week of treatment, there was complete resolution of all symptoms. The patient was discharged on a nasogastric tube and on room air via a tracheostomy, with a modified Rankin Score (mRS) of 5 and Barthel Index of Activities of Daily Living score of 1/20. On follow-up via telemedicine after 3 months, the patient has spontaneous purposeful movements of all extremities. He was still maintained on gabapentin, while propranolol and clonazepam were tapered off, without recurrence of autonomic instability and dystonia. The mRS and Barthel Index score remained the same. The patient’s family was able to continue with the treatment due to its lower cost.\n\nA, B: Epoch of electroencephalogram at the start during (A), and after the paroxysmal episodes showing absence of epileptiform discharges (B) prior to the event as evidenced by muscle artifacts. C: The decerebrate posturing of the patient during the event.\n\n\nDiscussion\n\nPAIDS is a syndrome of sympathetic hyperactivity accompanied by abnormal posturing.6 It is commonly observed after a severe traumatic brain injury (TBI) (79.4%); occurring in almost one-third of all cases of TBI but may also arise from paraneoplastic limbic encephalopathy (9.7%), cerebrovascular disorders (5.4%), hydrocephalus (2.6%), tumor (0.6%), and rarely after central nervous system infections (0.3%) like in pneumococcal meningoencephalitis and tuberculous meningitis, as in our case.2,4,7\n\nThe pathophysiology of PAIDS is still not well understood but likely best explained by the Disconnection Theory, wherein a brain injury causes dysfunction of autonomic centers in the diencephalon.5 Due to the disconnection of higher sympathetic inhibitory regions (e.g. insula, cingulate cortex) to the lower sympathetic activating centers (hypothalamus to the lower brainstem), there is increased activation of the excitatory pathways, triggering catecholamine release, and the subsequent hyperadrenergic state.4,5,8 It could occur from damage to the descending inhibitory pathways coming from the forebrain and rostral brainstem leading to a lack of inhibition that releases spinal sympathoexcitatory reflexes that then overreact to peripheral stimulation, explaining the paroxysms seen mostly during stimulation.4 Lesions in the midbrain that cause blockage of normal inhibitory signals to the pontine and vestibular nuclei result in rigidity and decerebrate posturing.2,9,10 Other than the patient’s tuberculous meningitis, the concomitant hydrocephalus could also have contributed to the development of the PAIDS. Even if a VPS was not done, there was no increase in the size of the hydrocephalus after repeat imaging, therefore, dysfunction of diencephalic and upper midbrain circuits and impaired inhibition of medullary sympathoexcitatory neurons from brainstem ischemia are thought to be the cause of the patient’s symptoms.4 On review of literature, PAIDS was reported in a child with tuberculous meningitis with note of persistence of symptoms despite the placement of a VPS; suggesting that the tubercular infiltration in the autonomic centers was the inciting cause.10\n\nPAIDS is a life-threatening condition that may lead to secondary hypertensive or hyperthermic encephalopathy and/or death. Differentiation from a neuroleptic malignant syndrome, malignant hyperthermia, sepsis, seizures, and impending herniation is very important.6 In our case, the diagnosis of PAIDS was made using the PSH-AM.4\n\nManagement includes minimizing stimulation and pharmacotherapy with preventive and abortive medications. The available data on treatment options are lacking. The clinical symptoms and treatments of PAID from tuberculous meningitis are presented in Table 1. Morphine sulfate was found to be most effective in preventing paroxysms, and other proposed medications include baclofen, benzodiazepines, bromocriptine, clonidine, dexmedetomidine, gabapentin, and propranolol. Data are mostly from small case series or case reports of PAID syndrome from TBI.4\n\n\n\n• Cessation of paroxysms 1 month after admission\n\n• Glasgow coma score (GCS) of E4V4M5 but neurologically severely impaired\n\n• Died after three months from pneumonia\n\n\n\n• Cessation after weeks of treatment\n\n• GCS of E4V4M5 however remained neurologically impaired on follow-up\n\n\n\n• Paroxysms ceased after 1 month from admission\n\n• Improvement of neurologic status and was neurologically stable after 3 months of follow-up\n\nPersistent and severe episodes of sympathetic hyperactivity can cause catastrophic consequences such as worsening of intracranial hypertension or hemorrhage, posterior reversible encephalopathy syndrome, stress-induced cardiomyopathy, hypoxic respiratory failure, acute tubular necrosis, renal failure, and rhabdomyolysis.4 Pharmacologic treatment can also cause iatrogenic autonomic dysfunction leading to an exaggerated cardiac and vascular response. PAIDS has poorer clinical outcomes and increased morbidity when associated with generalized motor rigidity and poor cognitive function.1,4,5 There is only one case report of PAIDS from tuberculous meningitis in an adult, hence there is limited knowledge on the expected outcome and subsequent quality of life after recovery.2\n\n\nConclusions\n\nPAID is a rare complication of tuberculous meningitis that can mimic other life-threatening conditions, hence early detection is vital to avoid unnecessary diagnostics and treatments. The diagnosis is challenging and exacting in a resource-limited country and early recognition is crucial for immediate, adequate, and cost-effective treatment.\n\nWritten informed consent was obtained from the patient’s authorized representative (mother) for the publication of this case report and any accompanying images.\n\nThe research was conducted ethically in accordance with the World Medical Association Declaration of Helsinki.", "appendix": "Data availability\n\nNo data associated with this article.\n\n\nReferences\n\nBaguley IJ, Perkes IE, Fernandez-Ortega J-F, et al.: Paroxysmal sympathetic hyperactivity after acquired brain injury: consensus on conceptual definition, nomenclature and diagnostic criteria. J. Neurotrauma. 2014; 31: 1515–1520. PubMed Abstract \| Publisher Full Text\n\nRamdhani NA, Sikma MA, Witkamp TD, et al.: Paroxysmal autonomic instability with dystonia in a patient with tuberculous meningitis: a case report. J. Med. Case Rep. 2010; 4: 1–5. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSarkar S, Nandi M, Mondal R: Paroxysmal autonomic instability and dystonia (PAID) in a child with tuberculosis meningo-encephalitis: a case report. J. Pediatr. Infect. Dis. 2011; 6: 201–203. Publisher Full Text\n\nRabinstein AA: Autonomic hyperactivity. Continuum (Minneap Minn). 2020; 26: 138–153. PubMed Abstract \| Publisher Full Text\n\nLetzkus L, Keim-malpass J, Kennedy C: Paroxysmal sympathetic hyperactivity: autonomic instability and muscle over-activity following severe brain injury. Brain Inj. 2016; 30(10): 1181–1185. PubMed Abstract \| Publisher Full Text\n\nBlackman JA, Patrick PD, Buck ML, et al.: Paroxysmal autonomic instability with dystonia after brain injury. Arch. Neurol. 2004; 61: 321–328. PubMed Abstract \| Publisher Full Text\n\nBaik S, Kang D, Kim G: Transdermal opioid patch in treatment of paroxysmal autonomic instability with dystonia with multiple cerebral insults. Medicine (Baltimore). 2020; 99: e22536. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBuerger KJ, Salazar R: Alternating hemidystonia following traumatic brain injury as an unusual presentation of paroxysmal autonomic instability with dystonia syndrome. BMJ Case Rep. 2014; 2014: bcr2014206102. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKapoor D, Singla D, Singh J, et al.: Paroxysmal autonomic instability with dystonia (PAID) syndrome following cardiac arrest. Singap. Med. J. 2014; 55: e123–e125. Publisher Full Text\n\nSingh DK, Singh N: Paroxysmal autonomic instability with dystonia in a child: rare manifestation of an interpeduncular tuberculoma. Pediatr. Neurosurg. 2012; 47: 275–278. Publisher Full Text" }	[ { "id": "269320", "date": "09 May 2024", "name": "Venkat Ramesh", "expertise": [ "Reviewer Expertise Tuberculous meningitis" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nOverall, the case report has been satisfactorily written, and provides a novel insight.\nThe exact CSF glucose value needs to be mentioned.\nHow the diagnosis of CNS TB was established needs to be elaborated in detail. . Was a GeneXpert M.tb (or other CB-NAAT) done? Was a bronchoscopy performed?\n\nThe case report needs minor modifications.\n\nIs the background of the case’s history and progression described in sufficient detail? Yes\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Partly\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Yes\n\nIs the case presented with sufficient detail to be useful for other practitioners? Yes", "responses": [ { "c_id": "12826", "date": "15 Nov 2024", "name": "Lowrence Precious Dichoso", "role": "Author Response", "response": "The diagnosis of CNS tuberculosis was arrived based on the history and supported by the laboratory results. The lumbar tap revealed an elevated opening pressure of 35 and closing pressure of 8. The qualitative cerebrospinal fluid (CSF) is yellowish, clear, with WBC 8x106/L (neutrophils 20%, lymphocytes 60%), while the quantitative CSF total protein of 687 mg/dL, glucose <1.1 mmol/L (1% of serum glucose). The Crytococcal Antigen Latex Agglutination System, Bactigen Latex Agglutination and fungal culture turned out negative. The CSF MTB- Detection By Polymerase Chain Reaction turned out to be positive. The Endotracheal Aspiration (ETA) acid-fast bacilli (AFB) was negative but no bronchoscopy was done, however noted on Chest X-ray with reticular opacities of both upper lung field suspicious for tuberculosis." } ] }, { "id": "274512", "date": "04 Jun 2024", "name": "Shailendra Katwal", "expertise": [], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIn neuroimaging figures, captions doesn’t explain the findings correctly. Not all the sequences are DWI/ADC. Mention the sequences and findings correctly. Even the right sided blue arrows in figure A are showing the ventricles instead of meningeal enhancement.\nFindings of reticular opacity in lung doesn’t mean the active form of pulmonary tuberculosis, however its mentioned as disseminated tuberculosis with brain and lung findings. What are the differential of this clinical entity and how do your rule out them?\n\nIs the background of the case’s history and progression described in sufficient detail? Yes\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? No\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Yes\n\nIs the case presented with sufficient detail to be useful for other practitioners? Yes", "responses": [] }, { "id": "274520", "date": "05 Sep 2024", "name": "Nor Osman Sidow", "expertise": [ "Reviewer Expertise Neurology and Clinical Neurophysiology" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis case is good and it explains in details of the information provided. the background and discussion is quite is enough and I agree that it is suitable for indexing the language is excellent the case also is novel and perfect the quality of the images and figures are very good\n\nIs the background of the case’s history and progression described in sufficient detail? Yes\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Yes\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Yes\n\nIs the case presented with sufficient detail to be useful for other practitioners? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1405
https://f1000research.com/articles/12-533/v1	23 May 23	{ "type": "Research Article", "title": "The effect of coronal pre-flaring and root canal irrigant on apex locators’ accuracy: an in-vitro study", "authors": [ "Shimaa Rifaat", "Abdullah Aljami", "Turki Alshehri", "Shahad T. Alameer", "Alhanoof Aldossary", "Wejdan Almutairi", "Mulham N. Almaliki", "Faraz A. Farooqi", "Noha Taymour", "Abdullah Aljami", "Turki Alshehri", "Shahad T. Alameer", "Alhanoof Aldossary", "Wejdan Almutairi", "Mulham N. Almaliki", "Faraz A. Farooqi", "Noha Taymour" ], "abstract": "Background: Successful root canal treatment is influenced by the apical extent of root canal preparation and the eventual root canal filling. Achieving the full working length until the apical constriction, which is usually 0.5 – 1 mm shorter than the anatomical apex, is crucial. Electronic apex locators were used to detect the working length more accurately. There are six generations of electronic apex locators in the market. The selection of the appropriate irrigation with each apex locator for accurate working length determination is not fully investigated. Methods: The actual working lengths of 120 freshly extracted human single-rooted teeth were measured and compared with their working lengths using 3rd generation (Root ZX) followed by 6th generation (Raypex 6) apex locators in dry medium, presence of 5.25% sodium hypochlorite, and 2% chlorhexidine, without coronal pre-flaring and after coronal pre-flaring using the same irrigating media. Data were collected, tabulated, and afterward analyzed using one-way ANOVA with post-hoc to evaluate the significant difference in average working length between actual working length, Root ZX, and Raypex 6 apex locator working lengths accuracy. Results: The significant results were shown in roots that were coronally pre-flared and their working lengths were measured in a dry medium using Raypex 6 apex locator. While using the Root ZX apex locator, the most accurate results were shown in roots that were coronally pre-flared and their working lengths were measured while using a chlorhexidine irrigating solution. Conclusions: It is concluded that it is very important to know the specific irrigating medium to be used with each specific electronic apex locator to achieve the most accurate working length results.", "keywords": [ "Working length", "Root ZX apex locator", "Raypex 6 apex locator", "irrigating solution", "coronal pre-flaring", "sodium hypochlorite", "chlorhexidine", "dry medium." ], "content": "Introduction\n\nSuccessful root canal treatment (RCT) is influenced by the apical extent of root canal preparation and the eventual root canal filling.1 Because the canal is narrow at the apical constriction, it is the suggested endpoint for instrumentation and obturation.2 Apical constriction is defined as the minor root canal diameter. Histologically, it represents the transitional point between the pulpal and the periodontal tissue at the cement-dentinal junction (CDJ). According to anatomical research, the apical constriction is 0.5 – 1.0 mm from the exterior or the main foramen.3 Therefore, the full working length (WL) until the apical constriction, which is 0.5 – 1 mm shorter than the anatomical apex, provides a clean barrier that protects the periodontium from any bacterial invasion.3 Moreover, under or overfilling of the canals is one of the reasons for endodontics treatment failures.4–6\n\nAccordingly, working length determination is a critical step in endodontic treatment.3 There are many ways to determine the working length (WL), which are: periapical X-ray (PAs), bleeding point, electronic apex locators, and the tactile sensation of the operator.7 The most commonly used method is to combine the usage of electronic apex locators and periapical radiographs.8 To reduce the patient’s exposure to the X-ray and take the least number of X-rays, a reliable apex locator should be used. There are six generations of electronic apex locators (EAL) used to detect the working length. The most popular system used is the 3rd generation apex locators.9 It uses two frequencies and measures the difference between these frequencies; however, the type of moisture present may affect the reading’s accuracy.9,10 The 4th generation, on the other hand, uses five frequencies, but they make mathematical measurements rather than measure according to a database.9,10 However, there are no marked differences in reading accuracy compared to the older apex locator generations. The 5th generation uses a database of canal electric features and then does a comparison along with a mathematical process. The new technological advancement has led to the sixth generation of EAL, where a steady algorithm is made according to the canal’s moisture properties. Furthermore, working length measurements became more reproducible and accurate than in the previous generations.9\n\nUsing an irrigating media in determining working length is being studied in the literature. Keeping the root canal dry or moist using an irrigant during using the apex locators has been questioned by many dental practitioners, whether it increases the accuracy rate of EAL or not. Using 5.25% sodium hypochlorite (NaOCl) during root canal treatment meets the prime principle of endodontics, chemo-mechanical cleaning and shaping of the root canal. NaOCl is also used as an antibacterial agent to dissolve organic components of the root canal system.11\n\n2% chlorhexidine gluconate (CHX) is a broad-spectrum antiseptic cationic agent which comes in two forms: gel (CHX-G) and solution (CHX-S). However, it does not have the ability to dissolve organic materials as 5.25% NaOCl does.11 A 2% CHX has been used in endodontics as an irrigating material or intracanal medicament as it has a broad spectrum of antimicrobial activity and a lower cytotoxic effect than NaOCl. While demonstrating its successful clinical performance, lubricating properties, and rheological action (in the gel form), it also prevents metalloproteinase, is chemically stable, does not stain clothes, is odorless, water-soluble, and due to its cationic structure gives a unique feature known as substantivity (residual antimicrobial activity).12,13 It is well-established that the chemo-mechanical procedure can be enhanced if followed by the usage of an antibacterial intracanal medication like chlorhexidine (CHX), especially in cases of exudation, hemorrhage, perforation, root resorption, trauma, or insufficient root development.13 Coronal pre-flaring for root canals has several advantages that can be reflected on the cleaning and shaping processes, including making it easier to put manual and rotary equipment into the apical region of the root canals.14,15 The results indicated that the pre-flaring procedure provided more accurate measurements in most cases.16\n\nFew studies had been conducted to evaluate the effect of irrigating solutions on the working length reading accuracy of various electronic apex locators in the presence or absence of coronal pre-flaring. Hence, this present study aimed to compare the working length (WL) accuracy using the 3rd generation (Root ZX) and 6th generation (Raypex 6) electronic apex locators (EAL) in single-rooted teeth in the presence of different irrigation media: dry medium, 5.25% sodium hypochlorite (NaOCl), and 2% chlorhexidine (CHX) in root canals with/without coronal pre-flaring.\n\n\nMethods\n\nThe present study was conducted in compliance with the Declaration of Helsinki, and the research protocol was approved by the Restorative Dental Science Department, Imam Abdulrahman Bin Faisal University, College of Dentistry. Ethical approval was obtained from the Institutional Review Board, at Imam Abdulrahman Bin Faisal University (IRB -2022-02-171) on 12/04/2022.\n\nTeeth were collected in disposal after receiving signed consent from dental surgery/oral procedure patients, who authorized the hospital to use its discretion in their disposal and to be used in research purposes if needed.\n\nPower analysis was performed using the Clinclac sample size for this study. Mean and standard deviations were used from previously published literature11 which are (0.64±0.54) and (0.33±0.22). The power of the sample was set as 90% and the significance level will be set as 0.05. Hence the calculated total sample size obtained was 109 and the sample was increased to 120 for more precise results.\n\nThis research began on 17/04/2022. 120 freshly extracted human single-rooted teeth were used in this study. Teeth were collected from the disposal section at Imam Abdulrahman Bin Faisal University Hospital after receiving signed consent from dental surgery/oral procedure patients, who authorized the hospital to use its discretion in their disposal and to be used in research purposes if needed. The extracted teeth were collected anonymously without exposing any of the patient’s data and were used only for this in-vitro study. They were collected and immersed in 5.25% sodium hypochlorite (NaOCl) for disinfection for 2 hours for disinfection.3 Afterward, they were stored in normal saline until their usage. Root surfaces and the apical regions’ examinations were done under a dental operating microscope (OMS1950 Dental Microscope, USA) with 25X magnification. Teeth with comparable lengths and completely formed apices were included in the current study. Teeth with any possible fractures and/or apex immaturity were excluded from the current study. Teeth were radiographed in both mesiodistal and buccolingual directions to exclude the absence of root resorption or canal curvatures. Only root canals with a curvature of 0–5 degrees were included in the study. Teeth with calcified canals, more than one canal, apical blockage, internal or external resorption, and caries all were excluded from this study.\n\n120 freshly extracted human single-rooted teeth were used for this study. They were examined using periapical X-ray for 0.08 seconds of exposure time. Scaling of the teeth was performed using an ultrasonic scaler (Dentsply Sirona, ProUltra Piezo Ultrasonic Handpiece), then stored in normal saline at 0.9% until used. Teeth were flattened with a diamond disk 1 mm thickness (Hi-Tech diamond disc bur) to have a reliable reference point based on Jakobson et al. findings that the rubber stopper on the file should be placed on a flat surface to limit the possibility of errors in research with electronic apex locators, which ensured that the study’s findings are not influenced.17 The working length was evaluated by two evaluators who were instructed to use the same criteria for evaluating and assessing the parameter of the current study. Cohen’s Kappa test was applied to ensure the agreement and consistency between the two evaluators’ WL evaluations. A value of 0.82, which is interpreted as a high level of agreement or reliability between the two evaluators, was gained.\n\nConventional access opening without any coronal pre-flaring was done. Apical patency was checked using K-file #10 (Dentsply M-access K-File). Actual working length (WL) was measured in millimeters by two calibrated evaluators under an endodontic microscope (OMS1950 Dental Microscope) with 25× magnification. Any tooth with an initial file more than #15 K-File (Dentsply M-access K-File) was excluded. The file was placed beyond the apical constriction and then retrieved until it is flushed with apical foramen. A 0.5 mm was subtracted from the total and the final measurement was considered the actual working length.2,8 Each tooth was mounted until the cementoenamel junction (CEJ) level using freshly mixed alginate. A double rubber stopper technique was used.3\n\nThe working length in millimeters for each tooth was measured using (3rd generation EAL) Root ZX (J. Morita Corp., Kyoto, Japan) then followed by using (6th generation EAL) Raypex 6. (VDW, Munich, Germany). Apex locators were used while teeth were dry, then while using 5.25% NaOCl, then while using 2% CHX respectively. Between each irrigating solution and the next, distilled water was used to neutralize each irrigant effect before using the next one. Afterward, the canals were dried using paper points. All measurements were taken by two calibrated examiners. K-file size #15 (Dentsply M-access K-File) was inserted inside the canal, a lip clip was placed inside a fresh alginate mix and a file holder was placed on the file.\n\nThe file was fixed on apex locators: the third generation EAL Root ZX (J. Morita Corp., Kyoto, Japan) and the sixth generation EAL Raypex 6 (VDW, Munich, Germany) for 5 seconds before measurements were recorded. Measurements were recorded when the file reached the mid-green area on the EALs’ screens.18\n\nAll teeth were collected, and coronal pre-flaring of the root canals was done. An access opening was prepared for each tooth. The penetration depth of gates-glidden drills was as follows: #3 to the canal orifice, #2 to the coronal third, and maximum to the coronal half of the canal to avoid perforations and to achieve a straight line access.19 Measuring the working length (WL) in millimeters of all teeth was repeated after the coronal pre-flaring with both Root ZX (3rd generation) and Raypex 6 (6th generation) apex locators using the dry medium, 5.25% NaOCl, 2% CHX irrigating solutions again as in Figure 1.\n\nData was initially recorded in an excel sheet and then transferred to SPSS (statistical package for social sciences) version 24, IBM, Inc. Mean and standard deviations were calculated and presented in tables as part of descriptive statistics. Comparisons between the irrigant solutions for Root ZX and Raypex 6 EAL were calculated and tested using the independent sample t-test and ANOVA. Where ANOVA was significant, multiple comparisons were done using Tukey’s post hoc test. Comparison in the working length between the 3rd (Root ZX) and 6th generation (Raypex 6) apex locators in teeth without coronal pre-flaring and with coronal pre-flaring in all media was also done using an independent sample t-test. P-values less than or equal to 0.05 were considered statistically significant.\n\n\nResults\n\nTable 1 presents the comparisons of the mean working length (WL) of Root ZX and Raypex 6 EALs between the irrigating solutions and within the irrigating solutions in the absence of coronal pre-flaring. Mean length measurements for Root ZX in CHX were significantly closer to the actual WL (0.087±0.445) compared to the other two irrigants; the least close measurement from the WL was with NaOCl (0.252±0.553), and the difference was statistically significant (p=0.025). Whereas with Raypex 6, the dry medium should have the closest readings to the WL but the overall mean difference among the irrigants used did not show any significant difference. Within each irrigating solution, the mean length of Raypex 6 was close to the actual WL and found to be statistically significant. Figures 2 and 3 present the pre-flaring working length measurements for Root ZX and Raypex 6 for all irrigating solutions. The median lengths for dry medium and NaOCl are almost the same, whereas the median length for CHX significantly decreased and was closer to the actual WL; asterisks are used to show the outliers.\n\n* Statistically significant at 0.05.\n\na Same alphabet showing significant difference between solutions.\n\nSimilarly, Table 2 presents the mean WL difference among the solutions for both apex locators in the presence of coronal pre-flaring. The mean WL for Root ZX locator differs significantly among the solutions (p-0.038). The closest mean to the actual WL was recorded with CHX (0.068±0.586), whereas the mean that most differed from the actual WL was recorded in the dry medium (0.269±0.621). Likewise, the most accurate mean WL for Raypax 6 locator was recorded in dry medium (-0.464±0.641); the least accurate WL was recorded with NaOCl (-0.174±0.584), and the difference between the actual WL and the mean was statistically significant. Within the irrigating solution groups, both apex locators differ significantly Table 2. The box plot in Figures 4 and 5 show the measurements’ median WL spread. Figure 4 shows that the median WL in NaOCl and CHX medium was almost equal but significantly different in the dry medium. Figure 5 shows a similar pattern for Raypax 6 apex locator where asterisks in each box plot refer to the presence of outliers.\n\n* Statistically significant at 0.05.\n\na Same alphabet showing significant difference between solutions.\n\nThe working length for both apex locators was then compared between the presence and absence of coronal pre-flaring and presented in Table 3. Mean length on both occasions (with and without coronal pre-flaring) did not differ significantly for both apex locators in dry medium (p-0.072) but the closest mean WL with actual WL was recorded in without coronal pre-flaring irrigation groups (0.146±0.39, -0.074±0.48, respectively). Similarly, in the NaOCl solution the most accurate mean WL was recorded without coronal pre-flaring media for Raypex 6 apex locator (0.121±0.50) and the difference was statistically significant (p-0.001). The closest mean WL to the actual WL in CHX for Root ZX was found in pre-flaring media (0.068±0.58) whereas for Raypax 6, it was the closest in the same media (0.28±0.74) but the difference was not significant statistically. When compared regardless of irrigation solution the most accurate mean WL to the actual WL was recorded in without coronal pre-flaring media groups for both apex locators (0.161+0.37, -0.118±0.44 respectively) and the difference was only significant statistically for Raypex 6 apex locator (p-0.005).\n\n* Statistically significant at 0.05.\n\na Same alphabet showing significant difference between solutions.\n\n\nDiscussion\n\nThe establishment of accurate working length (WL) is a crucial step during any root canal treatment; particularly in cases of anatomical limitations, it is useful when used with radiographs to ensure proper determination of the canal working length.20 The coronal pre-flaring of root canals gives many advantages during the meticulous cleaning and shaping procedures, such as facilitating the insertion of either manual and/or rotary files into the apical third of the root canals by removing cervical dentin interferences.21 In addition, the coronal flaring was known to improve the flow of the irrigating solution within the root canal, minimizing the risk of bacterial invasion into the periapical tissue as well as reducing the risk of canal debris and irrigant extrusion during the root canal preparation procedure.22,23\n\nIn recent years, there has been a growing body of evidence that suggests a correlation between the type of root canal irrigating solution used and the success of coronal pre-flaring efficacy. Moreover, the use of root canal irrigants is a major contributor to the success of endodontic treatment. Some studies showed a correlation between the irrigant and the root canal sealer used to ensure proper hermetic seal,24 while others discussed the effect of the proper irrigant on the accuracy of working length determination.25\n\nIn the present study, the effect of coronal pre-flaring on the accuracy of actual working length determination was assessed and the results showed that the more consistent, accurate results were observed in the canals that were prepared with coronal pre-flaring before working length (WL) determination in comparison to the canals that were prepared without coronal pre-flaring (see Table 3). These results might be due to the fact that the coronal pre-flaring improves the tactile sensation of the operator in locating the apical constriction.26 In addition, the largest impedance while inserting the file into the root canal is the coronal one-third of the canal. Moreover, coronal pre-flaring reduces file resistance; subsequently, it is easier to insert the file into the canal toward the apex of the root.27\n\nThis is in accordance with a previous study that showed that pre-flaring improved the efficiency of the EALs in the mandibular canals and anterior root canals.16 However, it is contradicted by a previous study conducted by João Marcelo da Silva Teixeira et al. (2012) who concluded that the usage of Gates Glidden burs for cervical pre-flaring did not significantly influence the accuracy of the apical placement of the apex locator when determining the actual working length due to insufficient removal of coronal dentin when compared with the rotary system for preparing coronal pre-flaring.28\n\nConsequently, the difference was statistically significant in the roots with coronal pre-flaring prior to working length determination when using Raybex 6 apex locator and NaOCl as an irrigant (T=4.5, p=0.0001). While comparing all the groups of Raypex 6 (with or without coronal pre-flaring) regardless of the type of irrigant used, significant results were seen when compared to the same groups of Root ZX (T=2.86, p=0.005).\n\nIn the case of using NaOCl as an irrigant, it showed significant results when used with Raypex 6 even without coronal pre-flaring. This coincided with the results of Elnaghy et al, 2017 who found that the use of 5.25% sodium hypochlorite (NaOCl) as an irrigant without coronal pre-flaring was associated with greater success rates than 2% chlorhexidine (CHX) solution. This may be due to the good electrical conductivity of NaOCl, which contributes to the accurate detection of Raypex 6 to the apical constriction. Moreover, it may be due to the advanced technology of the Raypex 6 that can accurately work in different canal conditions, including the presence of debris and/or obstruction(s) of the canal.9 Furthermore, the researchers also noted that sodium hypochlorite irrigant was more effective than other irrigants that may be used in preventing blockages and ledges formation in the root canal.29 Therefore, even without pre-flaring, the Raypex 6 may still provide accurate measurements.\n\nThe results showed that the readings when using Raypex 6 (6th generation electronic apex locator) are significantly closer to the actual working length than Root ZX (3rd generation electronic apex locator) for the with/without coronal pre-flaring groups. This coincides with the results of Pegum Unsal Peker et al. who concluded that Raypex 6 is not influenced by the presence of irrigation solutions due to its multi-frequency technology that displays precise results for the WL.9,30 The 6th generation apex locator is proved to be less sensitive to the influence of external factors that increase measuring reliability,31 like the number and taper of the file used in coronal pre-flaring that may influence the enlargement of the coronal portion of the canal.32 The sixth-generation EALs are proven to have a preliminary determination of the canal moistness and based on the constant determined moistness, the sixth-generation EALs adapt the measuring method for either a dry or a wet root canal environment.33\n\nIt is important to keep in mind that the accuracy of an electronic apex locator can vary depending on the type of irrigant used, so it is important to use the most appropriate irrigant for the situation at hand. The results of the current study showed the best results with the Raypex 6 apex locator were in dry medium in all conditions of pre-flaring (without pre-flaring & with pre-faring) (T=3.8, P=0.001) and (T=5.4, P=0.001) respectively. This may be due to the measuring changes in canal resistance, as it is easier to obtain accurate measurements in dry root canals.34 Moreover, our findings come in accordance with a study conducted by Koçak et al. for working length measurement in dry condition that showed more accurate readings than wet canals.35 However, our findings contradict a previous study conducted by Nayif MM et al., (2011) who stated that when using saline as an irrigant, readings were closer to the actual length, whereas those conducted in dry root canals were shorter than the actual working length.36\n\nIn accordance with our study, Root ZX apex locator achieved significant results with the CHX in all conditions of pre-flaring (without pre-flaring & with pre-faring): (T=3.85, P=0.001) and (T=3.99, P=0.001) respectively. This may be due to the different electrical conductivities of the irrigants which is defined as the intrinsic ability of the irrigant to conduct the electric current.11 Moreover, single-rooted teeth with single canal orifice were used which may contribute to the lack of difference between the groups with and without coronal pre-flaring. On the contrary, multi-rooted teeth with more canal orifices have a high potential for more anatomical variations and may have differences when coronal pre-flaring was done prior to WL determination.16\n\nMoreover, electroconductivity was enhanced in the current study by using alginate as an embedding material for electronic working length determination. Using the alginate model gives reliable and reproducible results as it has favorable characteristics that mimic the clinical situation by ensuring the required electric circuit for proper measurement of the electronic apex locator. That’s because it mimics the electric resistance of the human periodontal ligament.37 Despite the alginate’s firm consistency, it can remain as a gel that may allow the ions to circulate and promote adequate electroconductivity. Hence it is recommended to use alginate as embedding material for laboratory choices.37,38 Lucena-Martin et al. reported that electronic WL measurements should be concluded within 2 hours after mixing the alginate to minimize the chance of moisture loss.39 While Lipski et al. reported that the most accurate readings were obtained within only 30 minutes after mixing the alginate to enhance the electrical conductivity of the used irrigants and EAL.40 Consequently, the technique of alginate usage for only the first 30 minutes of mixing was used in the current study to ensure accuracy.\n\n\nConclusions\n\nIn conclusion, the study results suggested that adhering to the endodontic principles in conventional access opening, coronal pre-flaring, and patency are the cornerstone in achieving the most accurate and reproducible working length measurements in Root ZX and Raypex 6 EALs. It is concluded that the irrigant type selection has a major role in the accuracy of the EAL readings. Generally, using the 6th generation EAL (Raypex 6) is the most accurate choice for measuring the WL, but when used in a dry medium, it will achieve the most accurate WL measurements. Regarding the 3rd generation EAL (Root ZX), it is better to be used with 2% CHX to achieve the most accurate WL of the root canal. Hence, it is very important to know the specific irrigating medium used with each specific EAL to achieve the most accurate WL results.", "appendix": "Data availability\n\nfigshare: The Effect of Coronal Pre-flaring and Root Canal Irrigant on Apex Locators Accuracy: In-Vitro Study, https://doi.org/10.6084/m9.figshare.22492354.v4. 41\n\nThis project contains the following underlying data:\n\n- Apex locators results.xlsx\n\n- Microscope images for cases 1-6\n\nDue to the size of the original microscopy images, they are not all able to be uploaded to a public repository. Readers and reviewers can request access to further images from the corresponding author (srhussein@iau.edu.sa).\n\nfigshare: The Effect of Coronal Pre-flaring and Root Canal Irrigant on Apex Locators Accuracy: In-Vitro Study, https://doi.org/10.6084/m9.figshare.22492354.v4. 41\n\nThis project contains the following extended data:\n\n- Manuscript tables & figures\n\n- Additional images\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgments\n\nAll the authors are very grateful for all the constructive comments that improved the original version of the manuscript.\n\n\nReferences\n\nDummer PMH, McGinn JH, Rees DG: The Position and Topography of the Apical Canal Constriction and Apical Foramen. Int. Endod. J. 1984; 17(4): 192–198. 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Publisher Full Text" }	[ { "id": "174821", "date": "05 Jun 2023", "name": "Amira Salem", "expertise": [ "Reviewer Expertise Endodontics", "microbiology", "molecular biology", "biotechnology", "bioengineering", "product development", "drug delivery" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe order of procedure? Was CHX used on the same tooth as a last testing group, if so how do you justify that the results is not because multiple file insertions and a partial debridement specially after using sodium hypochlorite solution so it is giving more accurate results with this apex locator. Was there any testing group that has CHX used immediately after the dry group?\nTable 3 label? Check the label: all have Root ZX label\nIn all tables is the “mean” WL compared to the measured working length by microscope? If so this means that either the positive or negative value closer to the zero is the most accurate? Please clarify\nWhat are the numbers beside asterisks indicate? How many outliers were excluded from the mean measured.\n\nIn discussion: interpretation contradicts results “but the closest mean WL with actual WL was recorded in without coronal pre-flaring irrigation groups” comment on table 3. “accurate results were observed in the canals that were prepared with coronal pre-flaring before working length (WL) determination in comparison to the canals that were prepared without coronal pre-flaring”? in discussion.\n“However, it is contradicted by a previous study conducted by João Marcelo da Silva Teixeira et al. (2012) who concluded that the usage of Gates Glidden burs for cervical pre-flaring did not significantly influence the accuracy of the apical placement of the apex locator when determining the actual working length due to insufficient removal of coronal dentin when compared with the rotary system for preparing coronal pre-flaring” Not clear statement: was Gates Glidden burs better than rotary system or not, or there was no difference.\n“Consequently, the difference was statistically significant in the roots with coronal pre-flaring prior to working length determination when using Raybex 6 apex locator and NaOCl as an irrigant” Compared to what?\n“While comparing all the groups of Raypex 6 (with or without coronal pre-flaring) regardless of the type of irrigant used, significant results were seen when compared to the same groups of Root ZX (T=2.86, p=0.005)”. More accurate? If so, not true statement related to table 3?\n“In the case of using NaOCl as an irrigant, it showed significant results when used with Raypex 6 even without coronal pre-flaring” compared to what? To RootZX? With pre flaring it is less accurate.\nThe results showed that the readings when using Raypex 6 (6th generation electronic apex locator) are significantly closer to the actual working length than Root ZX (3rd generation electronic apex locator) for the with/without coronal pre-flaring groups. Doesn’t match with the results of table 3.\nIn accordance with our study, Root ZX apex locator achieved significant results with the CHX : maybe due to that it is the last tested irrigant and could be due to more debris and tissues removed as it is also better Root ZX in all groups for after flaring.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? No source data required\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "9856", "date": "04 Sep 2023", "name": "Shimaa Rifaat", "role": "Author Response", "response": "Dear Reviewer (1), (Dr. Amira Salem) Thank you very much for your valuable comments. They are a great addition to our research. Regarding the mentioned points, kindly check below for further details. 1) The order of procedure? Was CHX used on the same tooth as a last testing group, if so, how do you justify that the results is not because multiple file insertions and a partial debridement specially after using sodium hypochlorite solution so it is giving more accurate results with this apex locator. Was there any testing group that has CHX used immediately after the dry group? Yes, CHX was used for the same tooth as the last testing group, without using the CHX immediately after the dry group. A) We inserted the file only once for each medium (Dry, NaOCl, and CHX). B) We used distilled water between each medium to wash, neutralize and stop the effect of each material used before moving to the next. C) NaOCl is used routinely after access opening to dissolve all the pulp tissue remnants before the step of WL determination in all samples. Afterwards, the Dry, NaOCl, or CHX media was used to check the working length in all samples while using the electronic apex locators. D) The materials were used as a transmitting media not a debridement agent. E) The minimum exposure time for maximum effectiveness for the NaOCl is 20 minutes followed by 5minutes MTAD as a final rinse to ensure the complete debridement as stated by Lotfi et al 2012. Moreover, Bonnie et al 2009 reported that the maximum effectiveness for the 5% NaOCl as an antibacterial agent is at 40 minutes usage. In addition, Chen et al 2023 found that the application of NaOCl alone could be ineffective during chemical preparation of the canal and it should be agitated either by heat, ultrasonically, or followed by the usage of other materials. This concluded that only in this situation we can use NaOCl as a debridement agent otherwise it will not be an effective debriding agent, hence, the materials used in the current study was used only as transmitting medium for WL reading accuracy while using the EALs. Lotfi M, Moghaddam N, Vosoughhosseini S, Zand V, Saghiri MA. Effect of Duration of Irrigation with Sodium Hypochlorite in Clinical Protocol of MTAD on Removal of Smear Layer and Creating Dentinal Erosion. J Dent Res Dent Clin Dent Prospects. 2012;6(3):79-84. doi:10.5681/joddd.2012.017 Bonnie Retamozo, Shahrokh Shabahang, Neal Johnson, Raydolfo M. Aprecio, Mahmoud Torabinejad, Minimum Contact Time and Concentration of Sodium Hypochlorite Required to Eliminate Enterococcus faecalis, Journal of Endodontics, Volume 36, Issue 3, 2010, 520-523, ISSN 0099-2399, https://doi.org/10.1016/j.joen.2009.12.005. Chen Cai, Xuan Chen, Yang Li, Qianzhou Jiang, \"Advances in the Role of Sodium Hypochlorite Irrigant in Chemical Preparation of Root Canal Treatment\", BioMed Research International, vol. 2023, Article ID 8858283, 17 pages, 2023. https://doi.org/10.1155/2023/8858283. Table 3 label? Check the label: all have Root ZX label Thank you for your comment. Adjustments were done in the updated manuscript. In all tables is the “mean” WL compared to the measured working length by microscope? If so this means that either the positive or negative value closer to the zero is the most accurate? Please clarify. Yes, you are correct. The means that are either positive or negative value closer to zero are the most accurate values. What are the numbers beside asterisks indicate? How many outliers were excluded from the mean measured. A) The numbers beside Asterisks are the P value that indicates the significance in the given results. B) The value of outliers was replaced with the median of the data. They were not deleted but adjusted. In discussion: interpretation contradicts results “but the closest mean WL with actual WL was recorded in without coronal pre-flaring irrigation groups” comment on table 3. “accurate results were observed in the canals that were prepared with coronal pre-flaring before working length (WL) determination in comparison to the canals that were prepared without coronal pre-flaring”? in discussion. Thank you for your comment. Adjustments were done in the updated manuscript. “However, it is contradicted by a previous study conducted by João Marcelo da Silva Teixeira et al. (2012) who concluded that the usage of Gates Glidden burs for cervical pre-flaring did not significantly influence the accuracy of the apical placement of the apex locator when determining the actual working length due to insufficient removal of coronal dentin when compared with the rotary system for preparing coronal pre-flaring” Not clear statement: was Gates Glidden burs better than rotary system or not, or there was no difference. Thank you for your comment. Rephrasing the statement to make it clearer were done as follows and modified in the text. “However, a prior study conducted by João Marcelo da Silva Teixeira et al. (2012) reported that the usage of Gates Glidden burs for cervical pre-flaring did not significantly influence the accuracy of the apical placement of the apex locator when determining the actual working length. This may be due to the insufficient removal of coronal dentin when G.G. were used. On the contrary, the usage of rotary system for preparing coronal pre-flaring can significantly influence the WL determination. “Consequently, the difference was statistically significant in the roots with coronal pre-flaring prior to working length determination when using Raypex 6 apex locator and NaOCl as an irrigant” Compared to what? Consequently, the difference was statistically significant in the roots with coronal pre-flaring prior to working length determination when using Raybex 6 apex locator and NaOCl as an irrigant (T=4.5, p=0.0001) when compared with roots without pre-flaring and using NaOCl. “While comparing all the groups of Raypex 6 (with or without coronal pre-flaring) regardless of the type of irrigant used, significant results were seen when compared to the same groups of Root ZX (T=2.86, p=0.005)”. More accurate? If so, not true statement related to table 3? Thank you for your comment, it is adjusted in the discussion. “In the case of using NaOCl as an irrigant, it showed significant results when used with Raypex 6 even without coronal pre-flaring” compared to what? To RootZX? With pre flaring it is less accurate. In case of using NaOCL as an irrigant the mean score was significantly higher in Raypex 6 apex locator for coronal pre-flaring (0.464±0.64, p-<0.001) The results showed that the readings when using Raypex 6 (6th generation electronic apex locator) are significantly closer to the actual working length than Root ZX (3rd generation electronic apex locator) for the with/without coronal pre-flaring groups. Doesn’t match with the results of table 3. Thank you for your comment. Rephrasing was done as follows to make it clearer. The results showed that the readings when using Raypex 6 (6th generation electronic apex locator) are significantly closer to the actual working length than Root ZX (3rd generation electronic apex locator) for the with/without coronal pre-flaring in CHX irrigant system and in overall comparison when irrigant systems were not considered." } ] }, { "id": "174824", "date": "19 Jun 2023", "name": "Rania El Backly", "expertise": [ "Reviewer Expertise Endodontics", "regenerative endodontics", "tissue engineering", "regenerative medicine" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIn general:\nThe manuscript addresses an important topic relevant to the everyday practice of endodontics. However, it requires substantial language editing and rewriting to be more concise and focused. It is recommended to follow the PRILE 2021 guidelines for documenting laboratory studies in endodontics.\nTitle:\nI suggest modification of the title to be as follows:\nThe influence of coronal pre-flaring and type of root canal irrigant on accuracy of working length determination using electronic apex locators\nAbstract:\nThe abstract needs to be re-written to clearly demonstrate the specific objective of the study as well as the hypothesis. The methodology is unclear and non-specific. The groups are not clear. Results must be mentioned to clearly show quantitative data and show the differences between the groups used. Again, the conclusion must be more specific and clearly reflect the specific outcomes of this study.\nIntroduction:\nThe authors are advised to add a hypothesis statement at the end of the introduction section.\n\nThe rationale of the study is unclear. Coronal pre-flaring is a routine part of endodontic instrumentation and the irrigants used in this study are routinely used in everyday clinical practice in association with apex locators, so why are the authors assuming that these variables will affect the overall accuracy? This must be clearly shown.\nMethods:\nSample size calculation: The exact number of teeth per group must be mentioned.\n\nSample selection and sample preparation sections contain repeated information.\n\nThe grouping is not clear. Upon reading the methodology it seems that the 120 teeth were subjected to all conditions of measurements? This must be clearly shown in a table and a flow chart should be included to show the exact procedure.\n\nThe experimental set-up needs to be more accurately described.\n\nThe settings used for each apex locator must be clearly defined. “mid-green area on the screen” is not precise.\n\nHow was irrigation done? The volume used? The type of needle and gauge used? And depth of insertion of the needle?\n\nHow long was the duration of irrigation?\n\nWhy did the authors not use EDTA? Additionally, the sequence of using EDTA and NaOCl is the most common combination of irrigating solutions used especially since working length is often verified in the middle of instrumentation procedures under the influence of mixtures of these irrigants together. It is well-known in endodontics that combinations of irrigants are often used so testing combinations of irrigants would have given higher validation of the results of this study.\n\nThere is a major concern that the same samples were used for all the different variables. Other studies separated the entire sample into groups to avoid the influence of all of the intermediate procedures on the measurements taken last. How do the authors justify this? It would have been better to divide the sample into sub-groups for each tested variable especially since each sample represents its own control and this division of the samples would not have influenced the results, on the contrary would have enhanced the precision of the results.\nResults:\nThe tables and figures demonstrate that the values measured are the working length values in mm however, this is impossible since the values are even in the negative? So what do these values represent? Do they represent the values registered on the apex locator screens according to the pre-determined settings? I.e. the number registered on the screen when the device indicates that this is the CORRECT working length? If so these values are quite misleading. Also, how is it possible that the ACTUAL working length is 0.087+-0.445? The authors should record the entire working length in mm from the coronal reference point to the apical constriction minus 0.5 mm as they mentioned in the methodology.\n\nThe frequency of the difference between the actual working length and that measured in each condition by the two apex locators should be shown as this is the true measure that would truly demonstrate which technique was of higher accuracy in determining the WL.\n\nCheck:\n\nRamezani, M.; Bolbolian, M.; Aliakbari, M.; Alizadeh, A.; Tofangchiha, M.; Faegh, S.M.; Patini, R.; D’Amato, G. Accuracy of Three Types of Apex Locators versus Digital Periapical Radiography for Working Length Determination in Maxillary Premolars: An In Vitro Study. Clin. Pract. 2022, 12, 1043-1053. Kaufman AY, Keila S, Yoshpe M. Accuracy of a new apex locator: an in vitro study. Int Endod J. 2002 Feb;35(2):186-92.\n\nThe choice of 5.25% NaOCl should be justified. It would have been more practical to use 2.5% as this is the most commonly used concentration clinically.\n\nIt is recommended that diagnostic accuracy of the apex locators in presence of the different variables be recorded in terms of specificity and sensitivity as compared to the actual working length measured. This information would be a better guide for the clinician and as such authors can make a recommendation regarding the most “accurate” apex locator to use and in association with which conditions.\n\nThe results section should include a table for the demographic data of the sample including mean and SD of the types of teeth included, lengths of the teeth, number of canals (the authors mention single rooted but not the number of canals nor canal configurations), age of patients if present, gender….etc.\nDiscussion:\nThe discussion is well-written but the authors should show the clinical significance of the results as previously commented placed in the context of the multiple variables that are present in the clinical situation. The authors showed based on the results of their study that the type of irrigating solution has an influence on measuring WL using these two apex locators? So what do the authors recommend? A particular sequence to use? A specific irrigant to use only during WL determination? In order to give value to results from laboratory studies their clinical significance must be shown.\n\nThere should also be a section on the limitations of the study such as the limitation of performing the study on single rooted teeth, the influence of curvatures, the influence of combining different irrigants, influence of embedding medium, volume of irrigant and other limitations that may influence the extrapolation of the results in a clinical sense. Especially since most of these points were not mentioned in the discussion and it is advisable to do so. Additionally, a statement of accepting or rejecting the null hypothesis which should be mentioned in the introduction section is recommended.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? No source data required\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "9857", "date": "04 Sep 2023", "name": "Shimaa Rifaat", "role": "Author Response", "response": "Dear reviewer (2), (Dr. Rania Al Backly) Thank you very much for your valuable comments. They are a great addition to our research. Regarding the mentioned points, kindly check below for further details. 1) In general: The manuscript addresses an important topic relevant to the everyday practice of endodontics. However, it requires substantial language editing and rewriting to be more concise and focused. It is recommended to follow the PRILE 2021 guidelines for documenting laboratory studies in endodontics. Thank you for your comment and further editing and rewriting will be done in the updated version to make it clear for the readers. 2) Title: I suggest modification of the title to be as follows: The influence of coronal pre-flaring and type of root canal irrigant on accuracy of working length determination using electronic apex locators. Thank you for your comment. And I suggest modifying it too to be: “The effect of coronal pre-flaring and type of root canal irrigant on working length accuracy using electronic apex locators.” 3) Abstract: The abstract needs to be re-written to clearly demonstrate the specific objective of the study as well as the hypothesis. The methodology is unclear and non-specific. The groups are not clear. Results must be mentioned to clearly show quantitative data and show the differences between the groups used. Again, the conclusion must be more specific and clearly reflect the specific outcomes of this study. Updated Abstract: Background: The success of root canal treatment relies on achieving proper root canal preparation and precise root canal filling, including reaching the full working length up to the apical constriction. Electronic apex locators (EALs) have been introduced to enhance the accuracy of working length determination. However, the optimal irrigation protocol for each generation of EALs remains unclear Hypothesis: No significant difference in the accuracy of working length determination would be detected by coronal pre-flaring and the type of irrigant, varying with different generations of EALs Methods: Actual working lengths of 120 single-rooted human teeth were measured and compared with working lengths determined by the 3rd generation EAL (Root ZX) and 6th generation EAL (Raypex 6). Measurements were taken in a dry medium, in the presence of 5.25% sodium hypochlorite, and 2% chlorhexidine. The teeth were evaluated both without coronal pre-flaring and after coronal pre-flaring using the same irrigating solutions. Data were collected, tabulated, and analyzed using one-way ANOVA with post-hoc tests to assess significant differences in average working length accuracy between the actual working length, Root ZX, and Raypex 6 EALs Results: The results of the current study showed the best results with the Raypex 6 apex locator were in dry medium in all conditions of pre-flaring (without pre-flaring & with pre-faring) (T=3.8, P=0.001) and (T=5.4, P=0.001) respectively. Root ZX apex locator achieved significant results with the CHX in all conditions of pre-flaring (without pre-flaring & with pre-faring): (T=3.85, P=0.001) and (T=3.99, P=0.001) respectively Conclusion: The study found that the Raypex 6 electronic apex locator (EAL) provided significantly more accurate readings closer to the actual working length compared to the Root ZX EAL. The best results with the Raypex 6 were obtained when measurements were taken in a dry medium, in all the pre-flaring conditions. The Root ZX EAL achieved significant results with the use of chlorhexidine (CHX) as an irrigant in all the pre-flaring conditions. 4) Introduction: A) The authors are advised to add a hypothesis statement at the end of the introduction section. Hypothesis: No significant difference in the accuracy of working length determination would be detected by coronal pre-flaring and the type of irrigant, varying with different generations of EALs. B) The rationale of the study is unclear. Coronal pre-flaring is a routine part of endodontic instrumentation and the irrigants used in this study are routinely used in everyday clinical practice in association with apex locators, so why are the authors assuming that these variables will affect the overall accuracy? This must be clearly shown. Thank you for your question. It is clear nowadays that some research studies recommended the Contracted endodontic cavities (CECs) that were developed from the concept of minimally invasive dentistry and provide an alternative to traditional endodontic cavities (TECs). They claimed that the CECs preserve the mechanical stability of teeth and preserve more of the dentin as discussed by Lovisi et al 2018, Weiqi et al 2022, Wang et al 2023. That’s why the working length determination without coronal flaring was used in this study as it is a way some researchers recommended and supported by some of them. Alovisi M, Pasqualini D, Musso E, Bobbio E, Giuliano C, Mancino D, Scotti N, Berutti E. Influence of contracted endodontic access on root canal geometry: an in vitro study. Journal of endodontics. 2018 Apr 1;44(4):614-20. Weiqi Peng, Xuedong Zhou, Yuan Gao, Xin Xu, Effect of Access Cavity Preparation on Dentin Preservation, Biomechanical Property, and Instrumentation Efficacy: A Micro–Computed Tomographic Study, Journal of Endodontics, Volume 48, Issue 5,2022, 659-668, ISSN 0099-2399, https://doi.org/10.1016/j.joen.2021.12.012 Wang, X., Wang, D., Wang, Yr. et al. Effect of access cavities on the biomechanics of mandibular molars: a finite element analysis. BMC Oral Health 23, 196 (2023). https://doi.org/10.1186/s12903-023-02878-3 In addition, the effect of the irrigant on the reading accuracy and the electrical conductivity of various irrigant used and how it affects the accuracy of electronic apex locators. This will work hand in hand with the most accurate determination of the WL. Therefore, it would ensure the root canal treatment outcome and prognosis. 5) Methods: A) Sample size calculation: The exact number of teeth per group must be mentioned. 120 teeth were used. All the sample size experienced all the preparation scenarios and under all conditions to minimize the variables for among all the variables for each sample. B) Sample selection and sample preparation sections contain repeated information. Thank you for your comment. It will be revised and adjusted in the updated version. C) The grouping is not clear. Upon reading the methodology it seems that the 120 teeth were subjected to all conditions of measurements. This must be clearly shown in a table and a flow chart should be included to show the exact procedure. Yes, the 120 teeth were subjected to all conditions of measurements. This is supplied in the flow chart linked to the original manuscript on a figshare link. For your kind review: Rifaat S: The Effect of Coronal Pre-flaring and Root Canal Irrigant on Apex Locators Accuracy: In-Vitro Study. [Dataset]. figshare. 2023. 10.6084/m9.figshare.22492354.v4 D) The experimental set-up needs to be more accurately described. Thank you for your comment. It will be described clearly in the updated version. E) The settings used for each apex locator must be clearly defined. “mid-green area on the screen” is not precise. For Root ZX, working length determination was established at the point when the screen displays the line just in the middle of apex and 1 mark which corresponds to 0.5 mm short of radiologic apex. For Raypex 6, the third green line, just before the yellow lines, which corresponds to 0.5 mm short of radiographic apex was determined for working length. The measures were recorded as electronic measurement (EM) if they were stable for at least 5 seconds as described by Aydin et al 2015. Moreover, When the file tip reaches or is close to the apical constriction, the apex locator's screen display typically indicates this by showing the file position within the mid-green area. Moreover, both Root ZX and Raypex 6 EALs also have an audio indicator that emits a sound, such as a continuous or intermittent beep, when the file approaches or reaches the apical constriction. The change in sound indicates that the file is close to the desired working length, assisting the clinician in achieving precise measurements. Furthermore, both Root ZX and Raypex 6 feature a digital display that shows the distance between the file tip and the apical constriction in millimeters as reported by Higa et al 2009. Besides, Calibration of EALs before each use, according to the manufacturer's instructions was performed to ensure precision of WL measurements as recommended by Cinar et al 2020. Aydin U, Karataslioglu E, Aksoy F, Yildirim C. In vitro evaluation of Root ZX and Raypex 6 in teeth with different apical diameters. J Conserv Dent. 2015 Jan-Feb;18(1):66-9. doi: 10.4103/0972-0707.148899. PMID: 25657531; PMCID: PMC4313483” Higa RA, Adorno CG, Ebrahim AK, Suda H. Distance from file tip to the major apical foramen in relation to the numeric meter reading on the display of three different electronic apex locators. Int Endod J. 2009;42(12):1065-1070. doi:10.1111/j.1365-2591.2009.01629.x” “Çınar F, Üstün Y. Ex Vivo Evaluation of the Accuracy of 3 Electronic Apex Locators in Different Environments: A Micro-Computed Tomography Study. Eur Endod J. 2020 Dec;5(3):226-230. doi: 10.14744/eej.2020.30633. PMID: 33353910; PMCID: PMC7881377.” F) How was irrigation done? The volume used. The type of needle and gauge used. And depth of insertion of the needle? Irrigation was done with 23G with apical vented needle (Ultrdent Products, South Jordan, UT, USA). The needle was introduced until resistance was felt and then reduced 1 mm from the needle penetration depth to prevent obliteration of the root canal lumen as illustrated by Guerreiro et al 2013. 3 ml of 5.25% NaOCl and 2% CHX were used alternatively in the presence and absence of coronal pre-flaring as used by Spoorthy et al 2013. Guerreiro-Tanomaru JM, Loiola LE, Morgental RD, Leonardo RD, Tanomaru-Filho M. Efficacy of four irrigation needles in cleaning the apical third of root canals. Brazilian Dental Journal. 2013;24:21-4. Spoorthy E, Velmurugan N, Ballal S, Nandini S. Comparison of irrigant penetration up to working length and into simulated lateral canals using various irrigating techniques. International endodontic journal. 2013 Sep;46(9):815-22. G) How long was the duration of irrigation? It was for 2 minutes as suggested by Fernandes et al 2021. Fernandes AL, da Silva Limoeiro AG, Kato AS, Pelegrine RA, de Martin AS, Rocha DG, da Silveira Bueno CE. Analysis of two irrigation methods in root canal disinfection against E. Faecalis biofilm under the influence of the concentration, volumen, and time in contact of the irrigant. Research, Society and Development. 2021 Jul 13;10(8):e32610817478. H) Why did the authors not use EDTA? Additionally, the sequence of using EDTA and NaOCl is the most common combination of irrigating solutions used especially since working length is often verified in the middle of instrumentation procedures under the influence of mixtures of these irrigants together. It is well-known in endodontics that combinations of irrigants are often used so testing combinations of irrigants would have given higher validation of the results of this study. Thank you for your valuable comment. In our research we preferred to use only one irrigant sequence to prevent the results bias. As if this recommended protocol used, the authors will not be able which irrigant electroconductivity is higher: the NaOCl or the EDTA. But as we have a benchmark after our study results, we can consider using combinations with the NaOCl in the future to evaluate their accuracy. I) There is a major concern that the same samples were used for all the different variables. Other studies separated the entire sample into groups to avoid the influence of all of the intermediate procedures on the measurements taken last. How do the authors justify this? It would have been better to divide the sample into sub-groups for each tested variable especially since each sample represents its own control and this division of the samples would not have influenced the results, on the contrary would have enhanced the precision of the results. Thank you for the nice comment. We agree with you. But we tried to eliminate any variable that could take place among each group. That’s why we subjected each tooth was its same anatomical variation, diameter, length, dentin thickness to all the study variables to be more precise while calculating the deviation from the actual working length. Because all the mentioned points above could contribute to many changes while collecting the study results and could affect the study accuracy. 6) Results: A) The tables and figures demonstrate that the values measured are the working length values in mm however, this is impossible since the values are even in the negative? So what do these values represent? Do they represent the values registered on the apex locator screens according to the pre-determined settings? I.e. the number registered on the screen when the device indicates that this is the CORRECT working length? If so these values are quite misleading. Also, how is it possible that the ACTUAL working length is 0.087+-0.445? The authors should record the entire working length in mm from the coronal reference point to the apical constriction minus 0.5 mm as they mentioned in the methodology. Thank you for your comment. Regarding the readings for our results, it is the difference between the mean WL determined by the 2 calibrated evaluators and the actual working length. Considering the working length is at zero difference and we are calculating the difference or deviation from the zero either increased by (+) or decreased by (-). B) The frequency of the difference between the actual working length and that measured in each condition by the two apex locators should be shown as this is the true measure that would truly demonstrate which technique was of higher accuracy in determining the WL. Thank you for your valuable comment. The below table shows the frequency of deviations from actual working length for both electronic apex locators with each irrigation medium in the absence and presence of coronal pre-flaring. The frequencies are presented in table below were calculated for the differences between the measured and the actual WL within the range of ±0.5 and greater than 1. In the absence of coronal pre-flaring the Root ZX (3rd generation) showed more accuracy as most of the values were within the range of ±0.5 in each irrigation system, whereas Raypex 6 (6th generation) showed less accuracy than Root ZX while having majority of the values greater than 1 which is showing the far distance from actual WL. A similar trend was observed in irrigation media used in the presence of coronal pre-flaring except in dry medium where Raypex 6 apex locator showed more accuracy with the most values near to ±0.5. Medium EAL WL deviation in ±0.5 WL deviation > 1 WL deviation in ±0.5 WL deviation > 1 No. Freq. No. Freq. No. Freq. No. Freq. Dry 3rd generation 111 94.1 2.5 2.5 99 83.9 4 3.4 6th generation 102 86.4 10.2 10.2 104 88.1 14 11.9 NaOCl 3rd generation 104 88.1 2.5 2.5 102 86.4 7 5.9 6th generation 100 84.7 12.7 12.7 84 71.2 32 27.1 CHX 3rd Generation 107 90.7 4.2 4.2 114 96.6 4 3.4 6th Generation 96 81.4 15.3 15.3 101 85.6 17 14.4 C) The choice of 5.25% NaOCl should be justified. It would have been more practical to use 2.5% as this is the most commonly used concentration clinically. A 2021 study by Fernandes et al 2021 showed that the 5.25% NaOCl showed a significantly greater reduction in E. faecalis than 2.5% NaOCl. Therefore, some researchers suggested the usage of the 5.25% NaOCl. Fernandes AL, da Silva Limoeiro AG, Kato AS, Pelegrine RA, de Martin AS, Rocha DG, da Silveira Bueno CE. Analysis of two irrigation methods in root canal disinfection against E. Faecalis biofilm under the influence of the concentration, volumen, and time in contact of the irrigant. Research, Society and Development. 2021 Jul 13;10(8):e32610817478-. D) It is recommended that diagnostic accuracy of the apex locators in presence of the different variables be recorded in terms of specificity and sensitivity as compared to the actual working length measured. This information would be a better guide for the clinician and as such authors can make a recommendation regarding the most “accurate” apex locator to use and in association with which conditions. Thank you for your comment. The table below shows the relative error for each apex locator for accuracy of measurement of WL. Root ZX was found to be the most accurate measurement tool for WL in Dry (1.57%) and CHX (1.54%) irrigation system. Whereas with NaOCL, Raypax 6 showed high accuracy (1.76%) in teeth without coronal pre-flaring. However, in presence of pre-flaring, Root ZX showed high accuracy in NaOCL (2.21%) and CHX (1.93%) compared to Raypex 6. Irrigation Solution Apex Locator Relative Error Without Pre-flaring With Pre-flaring Dry 3rd generation 1.57% 2.48% 6th generation 1.58% 1.74% NaOCl 3rd generation 1.97% 2.21% 6th generation 1.76% 2.61% CHX 3rd Generation 1.54% 1.93% 6th generation 2.03% 2.28% E) The results section should include a table for the demographic data of the sample including mean and SD of the types of teeth included, lengths of the teeth, number of canals (the authors mention single rooted but not the number of canals nor canal configurations), age of patients if present, gender….etc. Regarding the demographic data for the sample, it was selected as follows: single rooted teeth with single canals of almost same length. Regarding the age and gender of the patient’s, …. etc. We don’t have this data as teeth were collected from the disposal of IAU dental hospital where patient identification were kept anonymous even after signing in the consent. 7) Discussion: A) The discussion is well-written, but the authors should show the clinical significance of the results as previously commented placed in the context of the multiple variables that are present in the clinical situation. The authors showed based on the results of their study that the type of irrigating solution has an influence on measuring WL using these two apex locators? So, what do the authors recommend? A particular sequence to use? A specific irrigant to use only during WL determination? To give value to results from laboratory studies their clinical significance must be shown. The author’s suggestions are: While using the 6th generation EAL (Raypex 6), dry medium is the best way to measure the most accurate WL. On the other hand, while using the 3rd generation EAL (Root ZX), it is better to use 2% CHX while measuring the WL to achieve the most accurate readings. Therefore, it is very important to know the specific irrigating medium used with each specific EAL to achieve the most accurate WL results during the root canal treatment. B) There should also be a section on the limitations of the study such as the limitation of performing the study on single rooted teeth, the influence of curvatures, the influence of combining different irrigants, influence of embedding medium, volume of irrigant and other limitations that may influence the extrapolation of the results in a clinical sense. Especially since most of these points were not mentioned in the discussion and it is advisable to do so. Additionally, a statement accepting or rejecting the null hypothesis which should be mentioned in the introduction section is recommended. Thank you for your comment. -Regarding the limitation of the study: Our study was conducted only on single rooted teeth with initial file size 15 while using one type of irrigant at a time to be used as a benchmark for further studies in the future. But our study limitations may include: the usage curved canals teeth with various degrees of curvature and with bigger apical foramen sizes may affect the given results. Furthermore, the usage of one irrgant at a time results may be different. Moreover, agitation of the irrigating solution may affect the electrical conductivity and hence affects the reading accuracy of EAL. Irrigation volume, concentration, temperature, and way of application may influence the reading accuracy as well. In addition, applying the current study in-vivo in the presence of the patient’s body fluids may have an influence as well. All these points should be taken into consideration in any future studies. -The null hypothesis was partially rejected due to the differences in the obtained results from the different irrigating solutions when used with each EAL. While it showed no significant difference between with/without coronal flaring groups." } ] } ]	1	https://f1000research.com/articles/12-533
https://f1000research.com/articles/12-746/v1	26 Jun 23	{ "type": "Research Article", "title": "Assessment of non-communicable diseases screening practices among university lecturers in Ghana – a cross sectional single centre study", "authors": [ "Joseph Kwasi Brenyah", "Joan Kyei-Dompim", "Elliot Koranteng Tannor", "Peter Twum", "Portia Boakye Okyere", "Barbara Gyapong-Korsah", "Florence Brenyah", "Christian Agyare", "Joan Kyei-Dompim", "Elliot Koranteng Tannor", "Peter Twum", "Portia Boakye Okyere", "Barbara Gyapong-Korsah", "Florence Brenyah", "Christian Agyare" ], "abstract": "Background: Non-communicable diseases (NCDs) are a major cause of morbidity and mortality globally. In low-income settings, some NCDs are without symptoms so regular screening for early detection is key. However, routine screening for NCDs is limited in the general public and even among the elite. We therefore set out to assess health screening practices among lecturers in a university in Ghana. Methods: This was a cross-sectional study involving 205 lecturers in Kwame Nkrumah University of Science and Technology from February to August 2022. A questionnaire was used to gather data from both male and female university lecturers based on their self-reported declaration of being male or female. Data were analyzed using descriptive and inferential statistics. Results: We found that, 41 (20.0%) lecturers (both men and women) had never checked their blood pressure (BP), 140 (68.3%) check their BP twice a month and 24 (11.7%) do so more than 3 times a month. Overall, 105 (57.18%) lecturers have high BP (>120 mmHg, >80 mmHg). Among the lecturers with hypertension, 59 (50.9%) often checked their BP each month, whereas 22 (18.97%) did not. The study found that, 164 (80%) of the lecturers have never checked their blood sugar level since they assumed lectureship position. Among the lecturers who check their blood sugar, 78 (47.55) are not happy with their blood sugar levels. Lecturer’s age (40 to 49 years) was found to be associated with BP in the bivariate analysis (p=0.036), but not in the multivariate analysis (p=0.114). In the bivariate analyses, female lecturers were found to have a higher risk (OR 1.35; 95% CI 0.29-6.21) of developing hypertension compared to male lecturers. Conclusions: The study has revealed that lecturers, just like the general population have moderate health care checks. The need to setup occupational health therapy units in all universities is overdue.", "keywords": [ "Assessment", "Screening Practices", "Non-communicable Diseases", "University Lecturers", "Ghana" ], "content": "Introduction\n\nNon-communicable diseases (NCDs) are a major cause of morbidity and mortality globally (Pan American Health Organization, 2021; Ouyang et al. 2022; Brenyah et al., 2023a). In low-income settings, some NCDs are without symptoms and hence regular screening for early detection is key to reduce its burden (Budreviciute et al., 2020; National Health Mission; undated). Early detection with appropriate management especially in high-risk individuals may be cost effective (Ghana Ministry of Health, 2012). By lowering common risk factors like tobacco use, hazardous alcohol use, physical inactivity and eating healthily, many NCDs can be avoided (Ghana Ministry of Health, 2012; Pan American Health Organization, 2021). NCDs account for 41 million annual deaths worldwide, or 71% of all fatalities (Pan American Health Organization & WHO, 2022). The prevalence of NCDs is on the rise in low-and middle-income countries (Ndubuisi, 2021; WHO, 2022). Preventive strategies are required, and immediate action must be made to reduce risk factors for NCD development. NCDs not only result in death, but also economic damages for a nation. This is a result of high dependency rates, poor work productivity, and restricted access to production resources, all of which contribute to poverty and low economic growth (Ummi et al., 2021).\n\nAs part of the modern approaches to healthcare in identifying and managing NCDs, anticipatory care including general and preventative health screenings have formed a fundamental component. For health to improve and health inequities to be avoided, it is crucial to guarantee a high and equitable uptake of general health check-ups (Watt et al., 2011; Dryden et al., 2012). Lecturers and teaching staff are among the body of professionals that need to undertake regular health checks due to heavy workloads in recent times. However, empirical data on health check practices among lecturers and teaching staff is lacking. We therefore set out to evaluate the health screening practices among lecturers at tertiary institutions in Ghana, using Kwame Nkrumah University of Science and Technology as a test case.\n\nGeneral health checks are a standard part of medical care. It includes a number of tests in a healthy individual with the intention of detecting illness early, preventing illness from occurring, or giving assurance of absence of illness (Scottish Government Report, 2008; Watt et al., 2011; Krogsboll et al., 2012). It is therefore important that individuals have general health checks every year to aid in the identification of, and early management of some possible diseases. However, according to Krogsboll et al. (2012), and Ummi et al. (2021), some people believe these health checks do more harm than good. This is due to the diagnosis of new and unexpected diseases which causes fear in individuals, as well as the rush to treat these conditions that may result in unnecessary regimen that could further harm the body. In either way, general health checks are very vital in the early detection of non-communicable diseases.\n\nMany professions are characterized by huge workload, stress, bad eating habits, and considerably long periods of sitting (Sharma & Majumdar, 2009; Dixon et al. 2014; Gareth et al., 2022). These are all modifiable risk factors of NCDs. Lecturers are a vital group of the society who contribute immensely to the educational system. However, to be a lecturer, means to carry upon oneself a great responsibility encompassing teaching, mentorship, research, community services and leadership roles. University lecturing is implicated in unhealthy modifiable behavior and as such lecturers may be predisposed to risk factors of non-communicable diseases. Available reports in the rise in mortality caused by non-communicable disease poses a great concern (Ummi et al., 2021; Pan American Health Organization, 2021; WHO, 2022) and needs to be address as a matter of urgency.\n\nStudies have reported that, over the years, there have been several community and professional based studies conducted to assess screening practices (Maindal et al., 2014; Alzahrani et al., 2021; AL-Kahil et al., 2020). Our search in the literature did not reveal the conduct of any study on health check practices among Lecturers or teaching staff. As a result, not much is known about the health check practices of lecturers in Ghana. This study therefore aimed at assessing health check practices among lecturers in the Kwame Nkrumah University of Science and Technology in Ghana. It is hoped that the outcome of this study would inform policy on health care issues not only among lecturers but also other professions in general.\n\n\nMethods\n\nThis section highlights the various methods used for this study. It covers study setting, study design, study approach, study population, sampling techniques, sample size calculation, inclusion and exclusion criteria, ethical consideration, data collection, data management and data analysis.\n\nThe study was carried out at Kwame Nkrumah University of Science and Technology (KNUST), Kumasi between February to August, 2022. The study covered all the six (6) Colleges in the University.\n\nThis was a cross sectional study to ascertain health check practices among university lecturers. The study therefore captured sex of lecturers who self-reported they were males or females. Therefore we did not inquire into socially constructed roles, behaviors, expressions and identities of individual participants. The implication is that, no external or internal examination of body characteristics or genetic testing, or other means were conducted on study participants.\n\nThe study employed a quantitative approach in which data was collected using questionnaires with both closed- and open-ended questions.\n\nThe study population involved 838 Lecturers across the six Colleges at Kwame Nkrumah University of Science and Technology (KNUST), Kumasi. A study of the lecturer population per college revealed that Colleges of Health Sciences (highest) and Agricultural/Natural resources (lowest) were the outliers (Quality Assurance and Planning Office, 2020).\n\nSimple probability technique was used to select the name of a college and the day/date to visit. Two sets of papers were folded with names of colleges (set 1) and day/date of visit (set 2). A picker picked one folded paper from each set and the name of the college and the day/date to visit was matched. In this case, the ordering of date and visit gave 1st, College of Humanities & Social Sciences (COHSS), 2nd College of Agriculture and Natural Resources (CANR), 3rd College of Art & Built Environment (CABE), 4th College of Engineering (COE), 5th College of Science (COS) and 6th College of Health Sciences (COHS). We then used the ‘walk-in’ system to select the study participants. By ‘walk in’, the date and time of visit to the Colleges were not announced to avoid possible biases in terms of lecturer modifiable lifestyle behaviors. Within the days to visit a college, any lecturer we meet in his/her office was a potential study participant. All lecturers who consented to participate in the study were assessed instantly in their offices face to face (see details under data collection procedure and tools).\n\nThe sample size was obtained using Yamane’s (1967) formulae as shown below:\n\nWhere:\n\nn = is the population of Lecturers in at KNUST\n\ne = is the level of precision\n\nTherefore:\n\nn = 270 participants.\n\nHowever, due to logistical constraints, 205 participants were contacted across the 6 Colleges at Kwame Nkrumah University of Science and Technology. The logistical constraints centered on consumables for the biochemical parameters and anthropometric measurements assessment; gloves, sanitizers, methylated spirits, glucometer strips, cotton wool, batteries (BP apparatus, weighing scales, glucometer gadgets) and among others. We then applied simple proportions to get the number of lecturers to be consulted in each College.\n\nInclusion criteria was made up of all Lecturers on KNUST campus who are in active service and consented to participate. All other staff not within this category were excluded from this research.\n\nEthical approval was sought from the CHRPE, KNUST with approval reference no: CHRPE/AP/581/21 granted on 8th December, 2021. The aim of the research was explained to participants. Those who consented to participate in the research were given consent forms to sign and date. Again, participants were assured of confidentiality. Participants were told that, they were free to withdraw from the study in the cause of time. In other words, study participants were not coerced into the study.\n\nData was collected using standardized structured questionnaires formulated for the current study based on guidelines of Boynton and Greenhalgh (2004). The questionnaires were pretested at Akenteng Appiah-Menka University of Skills Training and Entrepreneurial Development in Kumasi, Ghana. Questions found to be uncomfortable to respondents or biased were amended and incorporated into the final set of questionnaires (see Extended data, Brenyah et al., 2023b). The questionnaires covered socio-demographic characteristics of respondents, dietary intake, alcohol intake, issues on physical inactivity and tobacco use. Aside these four main risk factors of NCDs, the questionnaire also captured frequency of blood pressure checks, blood pressure outcome anytime it is checked (systolic and diastolic), frequency of general body check-up, frequency of anthropometric measurement checks (weight and height), an assessment of impressions about the outcome of weight and height checks, an assessment of intended measures to be taken depending on the outcomes of weight and health checked. Lecturers were also asked to identify if the content of their jobs have negative effects on their health status. With the assistance of two (2) State Registered Nurses, the research team and research assistants, lecturers were assessed on blood sugar status using glucometers, blood pressure using digital sphygmomanometer, weight and height using weighting scale and stadiometer. Questionnaires were administered using google-forms and submitted before the research team leaves the lecturers office. All these assessments were done in the offices of the lecturers. Instant results of blood sugar status, body mass index and blood pressure were released and interpreted to the respondents.\n\nOnly the Research Team had access to data. Data was kept confidential. The researchers had planned of disposing data from the storage 5 years after the publication of this research. Collected data was entered and cleaned using Microsoft Excel spread sheet, and then imported into STATA version 14.0 (Stata Corp LP, College Station, Texas, USA) for statistical analysis and results presentation.\n\nDescriptive statistics were used to summarize the characteristics of the study population by employing frequencies and percentages for categorical data. In addition, the degree of relatedness (association) was evaluated using Chi-square (χ2) or Fisher’s exact tests where appropriate with a p ≤ 0.05 assumed to be statistically significant. Both bivariate and multivariate logistic regression analyses were performed and adjusted for colleges’ effect to identify associations among the variables of interest. Variables having significant association in the logistic regression models were set at p ≤ 0.05 with 95% confidence interval (95% CI) for both odd and adjusted odds ratios (OR, AOR).\n\nBlood pressure (BP) was selected as the dependent variable, and in turn define as Normal: ≤ 120/80 mmHg; Elevated: Systolic between 120-129 and diastolic ≤ 80 mmHg; Hypertensive: Systolic ≥ 130 or diastolic ≥ 80 mmHg. Then dichotomized into Normal blood pressure: ≤ 120/80 mmHg and high blood pressure (Hypertension): ≥ 130/90 mmHg for logistic regression analyses. Independent variables were socio-demographics characteristics; sex status (male/female), age, marital status, staff rank and lecturer’s colleges (categorized into binary variable; COHS/COS/COE and CABE/CANR/COHSS), family history of NCDs and health check status. In this study, the variable “very often” denotes (doing the activity in question more than 4 times a month), “often” denotes (doing the activity in question at least twice a month), and “not often” denotes (doing the activity in question once a month).\n\n\nResults\n\nThe study involved 205 participants including 176 (85.9%) men and 29 (14.15%) women based on their self-declaration of identity (Brenyah et al., 2023b). The mean age of participants was 46.3 ± 9.4 SD years with 39.02% aged 50 years and above as shown together with other variables in Table 1. The only disaggregated variable relating to sex (male and female) were the number and sex identity of study participants per their reported responses and the male/female risk ratio of developing hypertension.\n\nParticipants with a self-reported NCD were 28.2%. The highest prevalence of the NCDs indicated was hypertension (66.8%) among the lecturers. Additionally, 50.1% of the Lecturers reported a family history of NCD, with hypertension accounting for 70.7% as shown on Table 2.\n\n* Data is presented in absolute and percentage terms.\n\nTable 3 shows that among the lecturers with hypertension, 59 (50.86%) often check their BP each month, whereas 22 (18.97%) reported not doing so often. Thus, there is statistical association (p = 0.049) between the frequency of monthly BP checks and blood pressure. However, no associations were found between BP and nature of BP results (p = 0.475), frequency of medical check-ups (p = 0.737) or BMI (p = 0.094).\n\n* Indicates presence of missing data; Statistical significance based on Chi-square or Fishes exact test where appropriate and set at P≤0.05.\n\nThe study found that 68.3% of lecturers undertake health checks by doing a blood sugar test twice a month. Again, 11.7% do the blood sugar test more than 3 times a month. Out of the 164 (80%) that took the test, 107 (65.3%) used the fasting blood sugar (FBS) and 57 (34.7%) used the Random Blood Sugar (RBS). Among those who took the test, 78 (47.5%) are not happy with their blood sugar status. The study found that, 61 (78.2%) claim their blood sugar status is mostly high (100 to 125 mg/dL (5.6 to 6.9 mmol/L) signifying prediabetes as shown on Table 4.\n\nTable 5 shows bivariate and multivariate analyses for BP on socio-demographic characteristics and health checks status. Lecturers in the age bracket 40 to 49 years were found to be associated (p = 0.036) with BP in the bivariate analysis, but not in the multivariate analysis (p = 0.114). Additionally, in the bivariate analyses, female lecturers were found to have a higher risk (OR 1.35; 95% CI 0.29-6.21) of developing BP (hypertension) compared to male lecturers. Lecturers in the colleges; COHS/COS/COE were found to be significantly associated (p = 0.016) with BP and were four times more likely (OR: 4.11, 95% CI: 1.30-12.95) to develop BP than those in the CABE/CANR/COHSS. Furthermore, it was shown that the frequency of lecturers BP checks (with the category ‘Often’) was significantly associated (p = 0.045) with their BP status. Although there was no association between blood pressure and any of the factors in the multivariate model, the analysis also revealed that lecturers who rarely check their weight (check monthly/yearly) were 91% more likely (AOR: 1.91, 95% CI: 0.09-41.98) to have BP than those who frequently check. Moreover, those with higher BMI (obese) had increase odds of BP that were more than six folds (AOR: 6.89, 95% CI: 0.61-77.9) compared with those with healthy weight. However, there were greater uncertainties surrounding the estimates of these odds.\n\nThe study also assessed if the variables assume different statistically significant associations if the logistics regression analyses were adjusted for effects from the colleges. It was noticed in Table 6 that strong statistical significance was found between BP checks and age (“40-49” p < 0.001, “50+” p = 0.05) and also with BMI (“overweight” p < 0.001, “obese” p < 0.001) in the bivariate analysis. The multivariate analysis showed several statistical associations were found between BP and its predicting factors including age: 40–49 (p < 0.001), the frequency of lecturers’ BP checks (with the categories “not often” p < 0.001, “Often” p < 0.001), job influence on lecturers BP (p = 0.037), the frequency of weight checks (with the category “never checked” p < 0.001) and BMI (“obese” p < 0.001) as shown on Table 6.\n\n\nDiscussion\n\nThe study reported on socio-demographic characteristics (sex) by only disaggregating the self-identify of respondents based on their self-declaration into male and female. These two categories of respondents (males and females) lecturing at the universities are classified as university lecturers in this research.\n\nThe study revealed a high prevalence of NCDs among university lecturers. Hypertension was revealed to be the most prevalent NCD (66.8%) which is not surprising because cardiovascular diseases have been reported to be the most prevalent NCD that accounts for most deaths (World Helath Organization, 2022). Individuals, irrespective of their sex (male or female) with a family history of chronic NCDs are at risk of developing similar conditions later in their lives (Downing et al., 2020; Alemi et al., 2021). This risk factor could add up to the presence of the chronic conditions observed among lecturers.\n\nThe prevalence of hypertension among the lecturers could be the occupational stress since there is vast evidence linking hypertension to occupational stress (Djindjic et al., 2012; Rosenthal & Alter, 2012; Owolabi et al., 2012). The current study revealed that lecturers in the age bracket of 40 to 49 years are more predisposed to developing hypertension than the other age groups. The European University Institute (2018) published that most lecturers obtain a PhD by age 37 and very few of them become full professors before the age of 40. This means that extra work is done in the academic pursuit of becoming a professor after the age of 40. Fulfilling occupational duties and that of the academic responsibilities compounds the stress that predisposes lecturers to developing high blood pressure. This situation makes frequent health checks by lecturers very important.\n\nOur study found that, female lecturers were more at risk of developing hypertension than male lecturers. This finding is inconsistent with other studies (Doumas et al., 2013; Everett & Zajacova, 2015; Kalibal et al., 2020; Alemi et al., 2021), that have suggested that hypertension is less likely to occur in women than in men. This inconsistency may be as a result of the age groups because prior studies focused on the youth unlike this current study that focused on older adults. Also, a healthy lifestyle of the female lecturers in this study could be a possible reason, however, it is not absolute. Further studies are needed to explore if the risk of developing hypertension in female lecturers increases as a result of the profession.\n\nHowever, our study outcome is consistent with the study outcome of Alemi et al. (2021), who reports that, female teachers had a higher prevalence of increased serum low-density lipoprotein (LDL) cholesterol and overweight/obesity than male teachers.\n\nThe level of awareness of a chronic disease is important in its management. The study revealed that lecturers who had regular medical assessment and checked their blood pressure regularly stood a better chance of identifying hypertension and related conditions such as diabetes and managing them. This finding is consistent with studies conducted by Everett & Zajacova (2015) who found out that participants who were aware of their chronic conditions were better managers of the disease than those who were unaware.\n\nWe also found that, among the lecturers with normal BP and those with hypertensive conditions, those who mentioned that they checked their BP often were high accounting for 53.85% and 40.68%, respectively. The implication is that many lecturers are aware of the need for frequent BP monitoring. Whiles those with normal BP are monitoring the possibility of onset of hypertension and other related conditions, those with hypertensive conditions are monitoring the control of their BP.\n\nIn addition, we realized that the BMI was biased towards lectures with hypertensive condition. It was found among them that, 51.87% and 43.22% were overweight and obese, respectively. This result is consistent with other studies that have reported that, overweight and obesity are associated with hypertension and diabetes (Jiang et al., 2016; Aronow, 2017). The outcomes of the health check variables translate to various activities as an effort to control it. For instance, according to our study, the lecturers with hypertension exercised more because they were not happy about their weight, unlike those with normal blood pressure who exercised less frequently.\n\nMoreover, our study found that only few lecturers among the normal BP category and those in the hypertensive condition category check their BP’s daily accounting for 11.11% and 5.95%, respectively. However, it was noticed that majority of the lecturers in the above categories checked their BP monthly accounting for 55.56% and 45.95% respectively. Under normal circumstances, people with hypertensive conditions should check their BP frequently. However, our study found a contrary practice where lecturers with hypertensive condition were the least in checking their BP in both daily and monthly regimen accounting for 5.95% and 45.95%, respectively. The implication is that BP check practices are poor among lecturers especially those with hypertensive conditions.\n\nThe study found that, majority (80 %) of lecturers undertake health checks by doing blood sugar test more than twice a month. Majority of them (107, 65.3 %) do the fasting blood sugar (FBS) and 78 (47.5%) of them are not happy with their blood sugar status. Among those who are not happy with their blood sugar level, 61 (78.2%) have high blood sugar level between 100 to 125 mg/dL (5.6 to 6.9 mmol/L)] which signifies prediabetes.\n\nAwareness means an individual may adopt lifestyle activities that will reduce the risk of developing diabetes and hypertension such as healthy eating and exercising. The regular visits to the hospital for routine examination was found to be higher among lecturers who were hypertensive. The general public has been reported to underutilize general healthcare checkups (Krogsboll et al., 2012) and this study reiterate that findings. This indicates a poor health check among the lecturers because it is indicating that they only go to the hospital when they are aware that they have an onset of a disease.\n\nHowever, these health checks are reliable means, through which chronic conditions are detected earlier and managed for an improved quality of life. Hence, the need for university lecturers to make it a habit of visiting the hospital for health checks.\n\n\nConclusions\n\nThe study has revealed that university lecturers, just like the general population have poor health check habits. The need for the setup of occupational health check units in all universities is overdue. The health promotion units of universities and hospitals should also scale up their activities to encourage university lecturers to improve their health check practices.", "appendix": "Data availability\n\nZenodo: Assessment of non-communicable diseases screening practices among university lecturers in Ghana – a cross sectional single centre study, https://doi.org/10.5281/zenodo.7946966 (Brenyah et al., 2023b).\n\nZenodo: Assessment of non-communicable diseases screening practices among university lecturers in Ghana – a cross-sectional single-center study, https://doi.org/10.5281/zenodo.7946966 (Brenyah et al., 2023b).\n\nThis project contains the following extended data:\n\n- Consent form NCDs and lecturers.docx\n\n- Participant Information Leaflet.docx\n\n- Questionnaire predictors of NCDs among lecturers.docx\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgments\n\nThe authors thank the Management of College of Health Sciences, Kwame Nkrumah University of Science and Technology for funding the research. We also appreciate the work of the College Grant Committee on our selection. The Authors are also grateful to the Office of the Registrar-KNUST for officially informing lecturers of the awareness of this research and granting permission for Lecturers to participate after the ethical approval was secured. We are equally grateful to the lecturers (participants) for their time and efforts in participating in this research. To all those who contributed in diverse ways but whose names are not mentioned here, we are grateful.\n\n\nReferences\n\nAlemi S, Nakamura K, Arab AS, et al.: Gender-Specific Prevalence of Risk Factors for Non-Communicable Diseases by Health Service Use among Schoolteachers in Afghanistan. Int. J. Environ. Res. Public Health. 2021; 18(11): 5729. 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PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source\n\nBrenyah JK, Nongvignon J, Signh A, et al.: Tobacco consumption and non-communicable diseases in Ghana; Identifying accentuating factors and further evidence from 2014 Ghana demographic and health survey. Journal of Scientific Africa Elsevier. 2023a; 20: e01665.\n\nBrenyah JK, Kyei-Dompim J, Tannor EK, et al.: Assessment of non-communicable diseases screening practices among university lecturers in Ghana – a cross sectional single centre study. [Dataset]. F1000Res. Zenodo. 2023b. Publisher Full Text\n\nBudreviciute A, Damiati S, Sabir DK, et al.: Management and Prevention Strategies for Non-communicable Diseases (NCDs) and Their Risk Factors. Front. Public Health. 2020; 8: 2296-2565. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDixon J, Carey G, Strazdins L, et al.: Contemporary contestations over working time: time for health to weigh in. BMC Public Health. 2014; 14: 1068. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDjindjic N, Jovanovic J, Djindjic B, et al.: Associations between the occupational stress index and hypertension, type 2 diabetes mellitus, and lipid disorders in middle-aged men and women. Ann. Occup. Hyg. 2012; 56(9): 1051–1062. PubMed Abstract \| Publisher Full Text\n\nDoumas M, Papademetriou V, Faselis C, et al.: Gender differences in hypertension: myths and reality. Curr. Hypertens. Rep. 2013; 15(4): 321–330. PubMed Abstract \| Publisher Full Text\n\nDowning KL, Hesketh KD, Timperio A, et al.: Family history of non-communicable diseases and associations with weight and movement behaviours in Australian school-aged children: a prospective study. BMJ Open. 2020; 10(11): e038789. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDryden R, Williams B, McCowan C: Themessl-Huber M. What do we know about who does and does not attend general health checks? Findings from a narrative scoping review. BMC Public Health. 2012; 12: 723. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEuropean University Institute: USA, Academic Career Structure.2018. Reference Source\n\nEverett B, Zajacova A: Gender differences in hypertension and hypertension awareness among young adults. Biodemography Soc. Biol. 2015; 61(1): 1–17. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGareth S, Jessica R, Esti M, et al.: The Assessment of Health Risk Behaviours among the Administrative Staff at an Institution of Higher Education. The Open Republic Journal. 2022; 15: e187494452208100. Publisher Full Text\n\nGhana Ministry of Health: National Policy for the Prevention and Control of Chronic Non-Communicable Diseases in Ghana.2012. Reference Source\n\nJiang SZ, Lu W, Zong XF, et al.: Obesity and hypertension. Exp. Ther. Med. 2016; 12: 2395–2399. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKalibala J, Pechère-Bertschi A, Desmeules J: Gender Differences in Cardiovascular Pharmacotherapy-the Example of Hypertension: A Mini Review. Front. Pharmacol. 2020; 11: 564. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKrogsbøll LT, Jørgensen KJ, Grønhøj Larsen C, et al.: General health checks in adults for reducing morbidity and mortality from disease. Cochrane Database Syst. Rev. 2012; 10: CD009009. Update in: Cochrane Database Syst Rev. 2019 Jan 31;1:CD009009. PubMed Abstract \| Publisher Full Text\n\nMaindal HT, Støvring H, Sandbaek A: Effectiveness of the population-based Check your health preventive programme conducted in primary care with 4 years follow-up [the CORE trial]: study protocol for a randomised controlled trial. Trials. 2014; 15: 341. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOwolabi AO, Owolabi MO, OlaOlorun AD, et al.: Work-related stress perception and hypertension amongst health workers of a mission hospital in Oyo State, south-western Nigeria. Afr. J. Prim. Health Care Fam. Med. 2012; 4(1): 307. Publisher Full Text \| Free Full Text\n\nPan American Health Organization: The burden of non-communicable diseases in the Region of the Americas, 2000-2019. ENLACE data portal.2021. Accessed on 10th May, 2023. Reference Source\n\nPan American Health Organization & World Health Organization: Non- Communicable Diseases.2022. Reference Source\n\nQuality Assurance and Planning Unit: Annual Report on Staffing. Kwame Nkrumah University of Science and Technology, Kumasi, Ghana College; 2020.\n\nRosenthal T, Alter A: Occupational stress and hypertension. J. Am. Soc. Hypertens. 2012; 6(1): 2–22. Epub 2011 Oct 22. PubMed Abstract \| Publisher Full Text\n\nScottish Government Report: Equally Well: Report of the Ministerial Task Force on Health Inequalities.2008. Reference Source\n\nSharma M, Majumdar PK: Occupational lifestyle diseases: An emerging issue. Indian J. Occup. Environ. Med. 2009; 13(3): 109–112. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nUmmi K, Helmi Suryani N, Ismi Nurwaqiah I: Individual Characteristic and the Causes of Death in Lecturers at Jambi, Indonesia. Paper presented at the Proceedings of the 3rd Green Development International Conference (GDIC 2020). 2021.\n\nWatt G, O’Donnell C, Sridharan S: Building on Julian Tudor Hart’s example of anticipatory care. Prim. Health Care Res. Dev. 2011; 12(1): 3–10. PubMed Abstract \| Publisher Full Text\n\nWHO: Key facts. Non-communicable diseases.2022. Accessed on 10th May, 2023. Reference Source\n\nYamane T: Statisitical. An Introduction Analysis. 2nd ed.New York: Harper and Row; 1967." }	[ { "id": "184050", "date": "14 Jul 2023", "name": "Abdesslam Boutayeb", "expertise": [ "Reviewer Expertise Mathematical modeling", "Epidemiology", "Public health", "health equity and social determinants of health" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nGeneral assessment\nThe content of the present manuscript is clearly and accurately presented by the authors.\nThe work is well documented and current adapted literature is cited.\nThe study design is appropriate and and the theme is of academic interest.\nAppropriate statistical tools are well used for analysis and the method allow replication by other researchers in other countries\nThe conclusions drawn by the authors are adequately supported by the results but the authors missed an important side consisting in determining the actual prevalence of diabetes and high blood pressure by diagnosing (according to the well known international procedures) during the survey. Showing the difference between the declared prevalence by the individuals at the beginning of the survey and the real prevalence diagnosed during the survey would have been more illustrative and convincing for both individuals and decision makers seeking optimal efficient strategies to control diabetes, hypertension and NCDS in general.\nMinor corrections\nPage 3: In low-income settings, some NCDs are without symptoms Some NCDs are without symptoms OR with symptoms that are not seriously considered by individuals either by ignorance, confusion or neglect.\nPage 3: . NCDs account for 41 In general, phrases do not begin with an abbreviation\nPage 3: and needs to be address as a matte Should read: to be addressed\nPage 6: The only disaggregated variable relating to sex (male and female) were Should read: variables were Or variable was\nPage 8: The study found that 68.3% of lecturers undertake health checks by doing a blood sugar test twice a month. This is different than what is indicated in Table 1 (Not often (1-2 times a month) (68.3)\nPage 11: The prevalence of hypertension among the lecturers could be the occupational stress This expression should be revised\nPage 12: Whiles those with normal BP are monitoring While instead of whiles (although whiles may be used in Scotland!).\nPage 11: However, it was noticed that majority of majority should be replaced by a majority\nPage 12: The study has revealed that university lecturers, just like the general population have poor health check habits. This assertion needs to give the data or references concerning the general population health checks, otherwise, the affirmation is unjustified.\nMajor concern This is an interesting survey. However, the authors missed to diagnose during the survey the actual prevalence of diabetes and high blood pressure. The diagnosed prevalence of diabetes and hypertension instead of and beside the declared prevalence by individuals would have improved the paper content.\nIndeed, globally, it is well known that high proportions of men and women are living with NCDS such as diabetes and High Blood Pressure while ignoring that they have the disease. For example, the World Health Organisation (WHO) and the International Diabetes Federation (IDF) stress that, in general, all around the world, around 50% of people with diabetes ignore that they have the disease. A national survey (STEPS 2017-18) carried out in Morocco on the common risk factors of NCDs found that 27% of women and 52% of men declared that they had never checked their blood pressure while 55.2% of women and 71.5% of men declared that they had never checked their glycaemia. Consequently, the Moroccan survey (STEPS 2017-18) revealed huge gaps between the declared prevalence (Declared Prev) and the diagnosed prevalence (Diagnosed Prev) during the survey for diabetes and high blood pressure (see table Source: Boutayeb A. Inequity & SDH in diabetes (chap2). In Boutayeb & Maamri. Health Inequity: A Crucial Issue Worldwide. Cambridge Scholars Publishing 2023, ISBN13: 978-1-5275-9371-8 https://www.cambridgescholars.com/product/978-1-5275-9371-8)\nMoroccan Ministry of Health. National Survey (steps) on the common risk factors of NCDs. https://www.sante.gov.ma/Documents/2019/05/Rapport%20de%20l%20enqu%C3%AAte%20Stepwise.pdf\nMoreover, the reader will be confused by the statement “The study found that 68.3% of lecturers undertake health checks by doing a blood sugar test twice a month” whilst not finding the precise prevalence of diabetes and eventually of pre-diabetes.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "191381", "date": "15 Sep 2023", "name": "Temitope Olumuyiwa Ojo", "expertise": [ "Reviewer Expertise Occupational Health", "Epidemiology", "Health Systems research", "Public Health" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors have examined an important issue pertaining to the occupational health of staff at a university in Ghana. It is a study that may be relevant to other similar contexts. It is noteworthy that fairly recent literature were cited in parts of the manuscript\nThe following comments may improve the manuscript:\nAbstract\nBackground: This needs to be revised to provide context/justification for conducting the study among lecturers.\n\nResults: Report in past tense.\n\nConclusion: Last sentence needs to be revised, eg need for regular health screening for staff, or establishment of an enhance occupational service to conduct regular health screening and treatment for NCDs among staff. Etc. note that an Occupational therapy unit is not the proper unit to do this.\nIntroduction\nA proper context to the study needs to be provided, eg why study lecturers and not all staff? Any difference in exposures to NCD risk factors? Eg Stress, lifestyle issues (sedentary lifestyle, smoking, physical activity etc) or is there a higher burden of NCDs or its complications among lecturers etc.\n\nIn addition the rearrangement of the paragraphs may allow for better flow of ideas. Eg third paragraph may need to come earlier since It introduces the concept of health screening for NCDs which is the main thrust of this study.\nMethods\nI wonder why the following statement was included in study design: “The implication is that, no external or internal examination of body characteristics or genetic testing, or other means were conducted on study participants” I think it may be deleted.\n\nWhy were 205 respondents recruited instead of the minimum sample size calculated, a convincing reason needs to be stated and not just logistics!\n\nMore details about how the screening examinations and tests were conducted are needed. Eg how did authors ensure that weight, height , BP were taken to minimise errors. Were readings repeated?\n\nAlso for persons with elevated BP or Sugar, what treatment was offered. Any referrals? Interpretation of results is not enough.\n\nAnalysis: use 'association between categorical variables was assessed...' instead of “degree of relatedness”\nResults\nI think the term elevated BP is more appropriate (Hypertension requires a stringent criteria for diagnosis, eg repeated readings etc).\n\nSo the tables should speak to predicting elevated BP. This needs to be corrected throughout the document. Blood pressure is a just a variable, so one may presume that the category of interest to this study is “elevated BP”.\n\nFor table 5, it looks problematic, what is the criteria for including variables in the multivariate regression model? The interpretation of the table is also challenging as non-significant variables were given prominence in the narratives.\n\nWhat is the difference in the models in table 5 and 6? The authors need to decide on the criteria for including variables in the regression model and be clear about the outcome variable.\nDiscussion\nSome statements need to be revised so as not seem speculative or sensational. Eg such statements as “which is not surprising because cardiovascular diseases have been reported to be the most prevalent NCD that accounts for most deaths” also ‘The prevalence of hypertension among the lecturers could be the occupational stress since there is vast evidence linking hypertension to occupational stress”.\n\nAuthors need to discuss the main findings without repeating the results all over.\n\nStudy limitations need to be discussed.\n\nLines of future research also required.\nConclusion:\nQuite scant. More recommendations are required to address some major study findings eg to make NCD screening more acceptable for respondents, improve frequency of checks, optimise compliance with treatment for persons already diagnosed and on care, exercise, lifestyle modification etc\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-746
https://f1000research.com/articles/12-1401/v1	24 Oct 23	{ "type": "Research Article", "title": "Potential business model for a European vaccine R&D infrastructure and its estimated socio-economic impact", "authors": [ "Stefan Jungbluth", "William Martin", "Monika Slezak", "Hilde Depraetere", "Carlos A. Guzman", "Anton Ussi", "David Morrow", "Fran Van Heuverswyn", "Sven Arnouts", "Manuel J. T. Carrondo", "Ole Olesen", "Tom H.M. Ottenhoff", "H. M. Dockrell", "Mei Mei Ho", "Alexandre Dobly", "Dennis Christensen", "Joaquim Segalés", "Fabrice Laurent", "Frédéric Lantier", "Norbert Stockhofe-Zurwieden", "Francesca Morelli", "Jan A.M. Langermans", "Frank A.W. Verreck", "Roger Le Grand", "Arjen Sloots", "Donata Medaglini", "Maria Lawrenz", "Nicolas Collin", "William Martin", "Monika Slezak", "Hilde Depraetere", "Carlos A. Guzman", "Anton Ussi", "David Morrow", "Fran Van Heuverswyn", "Sven Arnouts", "Manuel J. T. Carrondo", "Ole Olesen", "Tom H.M. Ottenhoff", "H. M. Dockrell", "Mei Mei Ho", "Alexandre Dobly", "Dennis Christensen", "Joaquim Segalés", "Fabrice Laurent", "Frédéric Lantier", "Norbert Stockhofe-Zurwieden", "Francesca Morelli", "Jan A.M. Langermans", "Frank A.W. Verreck", "Roger Le Grand", "Arjen Sloots", "Donata Medaglini", "Maria Lawrenz", "Nicolas Collin" ], "abstract": "Background Research infrastructures are facilities or resources that have proven fundamental for supporting scientific research and innovation. However, they are also known to be very expensive in their establishment, operation and maintenance. As by far the biggest share of these costs is always borne by public funders, there is a strong interest and indeed a necessity to develop alternative business models for such infrastructures that allow them to function in a more sustainable manner that is less dependent on public financing.\n\nMethods In this article, we describe a feasibility study we have undertaken to develop a potentially sustainable business model for a vaccine research and development (R&D) infrastructure. The model we have developed integrates two different types of business models that would provide the infrastructure with two different types of revenue streams which would facilitate its establishment and would be a measure of risk reduction. For the business model we are proposing, we have undertaken an ex ante impact assessment that estimates the expected impact for a vaccine R&D infrastructure based on the proposed models along three different dimensions: health, society and economy.\n\nResults Our impact assessment demonstrates that such a vaccine R&D infrastructure could achieve a very significant socio-economic impact, and so its establishment is therefore considered worthwhile pursuing.\n\nConclusions The business model we have developed, the impact assessment and the overall process we have followed might also be of interest to other research infrastructure initiatives in the biomedical field.", "keywords": [ "Vaccines", "research and development", "research infrastructure", "business model", "sustainability", "impact assessment", "science policy" ], "content": "Introduction\n\nVaccines are amongst the most effective public health tools available to humanity to fight infectious diseases (Greenwood, 2014; Doherty et al., 2016). Outstanding achievements of vaccines include the control or eradication of several previously devastating human and veterinary diseases such as smallpox and rinderpest (both eradicated), the near eradication of poliomielitis, and, importantly, a significant reduction of the global negative consequences of the COVID-19 pandemic on global health, society and economy. Despite these successes, there are numerous gaps and weaknesses in our current vaccine arsenal, and considerable work and innovation is needed to reduce the threat and burden of endemic and emerging infectious diseases, as well as to prepare for unknown future threats. However, vaccine development is a time-consuming and complex process that involves progression through various phases, from discovery and preclinical research to clinical development and large scale manufacturing of the final vaccines. All these steps require significant financial resources and broad technical capabilities. Development of vaccines is also very unpredictable as vaccine candidates can fail during any of the above-mentioned, increasingly expensive stages. On average, less than 1 out of 10 vaccine candidates in preclinical development eventually reach the market.\n\nIn a previous analysis based on structured feedback from experts, we identified numerous deficiencies in the European vaccine R&D landscape (Jungbluth et al., 2022). Many of these needs could be addressed by a well-designed, public health-driven and sustainable vaccine R&D infrastructure that—by the provision of services, expertise, facilities and other types of support—could foster innovation and scientific advancements in the larger vaccine R&D space (Leroy et al., 2014; IPROVE, 2016).\n\nResearch infrastructures (RIs) are physical and organizational structures, facilities, resources, and services that support scientific research and innovation. These infrastructures are designed to provide scientists, researchers, and innovators with the necessary resources, collaborative environments and expertise to conduct cutting-edge research and address complex scientific questions. RIs can vary widely in scope, scale, and focus, but are typically long-term initiatives with lifespans of decades rather than years (OECD, 2019). As the establishment and maintenance of RIs usually involve significant investments from governments, international organizations and private entities, the decision of whether to establish an RI is typically informed by a technical and conceptual feasibility study in which different RI design options are developed and compared, the expected socio-economic impacts (SEIs) assessed, and—eventually—detailed business and implementation plans prepared. In the present article, we describe the outcome of such a design study and SEI assessment for a future European vaccine R&D infrastructure. The aim of the feasibility study is to inform funders, policy and decision makers and other stakeholders about critical infrastructure needs within the vaccine R&D community and to provide a sound basis for evidence-based decision making regarding the establishment of a sustainable vaccine R&D infrastructure.\n\nThe feasibility study, including the findings described in the present article, were produced in the context of TRANSVAC (European Network of Vaccine Development and Research). TRANSVAC—an initiative that comprises three vaccine infrastructure projects funded by the European Union (EU) from October 2009 until April 2023—is a distributed, network-based research infrastructure integrating the expertise and facilities of 26 leading research organisations from ten European countries1. Provided with a total cumulative funding of approximately 27 million EUR and a focus on prophylactic and therapeutic vaccines for human and veterinary use, TRANSVAC has supported vaccine R&D by developing and optimising scientific-technical services, which were then offered to the vaccine development community at no cost along with cutting-edge training in vaccinology.\n\n\nMethods\n\nFour different business model options were developed using the procedure described below, one of the models was a combination of two individual models developed. In the present article, we only describe the details and expected impacts for the business model option that was prioritized for further development (“hybrid model”) following the assessment of all models using the evaluation framework developed (see below). The working steps in the overall process were the following ones:\n\n1. Defining the scope for business model design options:\n\nTo determine what business model options should be considered in the assessment, an evaluation of market needs and of TRANSVAC’s current capabilities was conducted to identify key opportunity areas for a future sustainable vaccine RI. In addition to intelligence produced directly by TRANSVAC (Leroy et al., 2014; Jungbluth et al., 2022; and unpublished), this step included the consultation of pertinent business (IPROVE, 2016) and market intelligence2,3, the analysis of different business models used by other existing Ris in the biomedical area (ESFRI, 2021, and references and links therein), and surveys and expert interviews we conducted. The individuals that participated in this process included representatives from all major stakeholder groups considered relevant for this venture, including potential users of a future vaccine RI, funders and finance providers, policy- and decision makers, industry, international organizations, vaccine development alliances, as well as existing Ris in the biomedical area in Europe.\n\n2. Creating an evaluation framework for assessment:\n\nA list of specific metrics for the quantitative scoring of the business models was developed. The metrics reflect important criteria, including unique selling position; competitive environment; financial upside; sustainability; risks, investment needs; complexity of implementation; capabilities/assets, and others.\n\n3. Describing details of the design options:\n\nBased on the market gaps, needs and opportunities identified in step 1 (current vaccine market situation), and on the analysis of currently existing/used business models in a range of industries and several successful Ris, different business model archetypes were developed and put forward for further consideration in the next step.\n\n4. Model evaluation and selection:\n\nEvaluation and selection of the different business model options were carried out using the evaluation framework described.\n\nThe entire process as outlined above, including the overall suggested positioning of the sustainable vaccine infrastructure, was performed keeping in mind the gaps and needs previously identified as part of the TRANSVAC project, as well as the objectives, scope and missions of other (European) Ris already existing.\n\nTo select impact categories for analysis, a literature review on the socio-economic impact assessment practices was made first, followed by a benchmarking of different practices and information (Reid et al., 2015; ESFRI, 2019). Multiple key impact indicators emerged, with three main impact categories aligned with the literature and taking into consideration the following standard metrics:\n\n- Health impact: Innovation for vaccine R&D, models developed, tools/technologies/solutions developed, vaccines developed and related data generation (immunological signatures/correlates of protection/efficacy and safety …)\n\n- Societal impact: Dissemination activities and new publications, training and education, human resources\n\n- Economic impact: Investments to accelerate vaccine development, impact on small- and medium sized enterprises (SMEs), revenues generated.\n\nApplied to the vaccine research infrastructure case, these three dimensions draw a picture of how such an infrastructure would deliver socio-economic impacts, locally, regionally, and globally, and help to better understand how it could fulfil its long-term objectives. Moreover, these categories have in common that they capture general issues having a fundamental impact on a vaccine infrastructure, represent a systematic approach for quantification, and are widely applicable.\n\nThe three main dimensions of socio-economic impact that were analysed reflect the impact of vaccines which is broad and far-reaching, though not consistently quantifiable, analysed or communicated. Traditionally, the perceived benefits of vaccinations are to reduce morbidity and mortality from infections, and those remain the drivers for the innovation of new vaccines. However, an increasing appreciation for the economic and societal impact of vaccines is being included in the development and assessment of vaccine programmes, as they potentially deliver greater benefits to society (Rodrigues and Plotkin, 2020; Beck et al., 2022). In the assessment we have undertaken, the impacts across the three different dimensions have been estimated for the first 10 years of the RI’s period of operation.\n\nA series of KPIs was developed that was subsequently used for assessing the ex ante impact of the vaccine RI along the different dimensions described above (Table 1). Eventually, once the vaccine RI is established and operational, the same KPIs could be used for the actual performance monitoring of the venture, as well as for a potential ex post impact assessment. For the actual performance monitoring of the vaccine RI once it is up and running, each KPI would need to be assessed with a specific frequency (quarterly or annually) and using a specific way of data collection (e.g. via operational and economic performance indicators, surveys, interviews, or from other external sources).\n\nFor performance monitoring, three categories of key performance indicators (i.e., short-term impact, long-term cumulative impact, and operational metrics) were selected to mirror the progress of the RI in achieving established objectives and goals. With operational metrics, the functioning of infrastructure can be monitored, whereas the short-term impact and long-term cumulative impact indicators enable actual measurement of achievements over time and their communication to key stakeholders. Performance measures include the tracking of economic, societal and other health indicators of impact. These dimensions were chosen as they are also relevant for the mission we propose for a vaccine RI to accelerate vaccine development in the interest of public health and societal benefits.\n\nInitially a larger list of approximately 30 KPIs was drawn up that subsequently was down-selected based on their relevance, credibility, quality as indicators and direct linkage to proposed vaccine R&D RI’s objectives. The KPIs have two major functions: firstly, they should quantify generated impact on a higher level to enable addressing a broader set of audiences and secondly, they should enable management of the specific areas on a more granular level.\n\nTable 2 summarises the data sources and assumptions that were used for the estimation linked to the KPIs selected. For a complete list containing all KPIs please see Table 1.\n\n\n\n\n\n\nResults\n\nThe gaps-and-needs assessment of vaccine R&D in Europe we conducted previously had identified several opportunities for the conceptual design of a sustainable vaccine R&D infrastructure. These include general gaps and needs in the vaccine R&D field such as lack of expertise in transitioning vaccine candidates from preclinical research to clinical testing, as well as specific aspects including the lack of access to value chain services, such as vaccine formulation expertise, pre-clinical testing, small-scale manufacturing according to good manufacturing practices (GMP), regulatory support. Moreover, our analysis revealed the difficulty of securing funding or financing for late-stage preclinical vaccine development, vaccine GMP manufacturing and early clinical testing (for details, see Jungbluth et al., 2022). Keeping in mind these and other findings from the gaps-and-needs analysis, according to our vision a future sustainable vaccine infrastructure should ideally address the following aspects:\n\n- Provide access to a powerful network of researchers with vaccine development expertise and facilities that is able to assess potential risks related to the development of specific vaccines and that is able to offer support that will increase the probability of success of specific vaccine development projects\n\n- Provide support/services, resources and capabilities for vaccine development\n\n- Support horizontal themes across vaccine development (i.e. disease/pathogen-independent)\n\n- Provide transversal and multi-disciplinary expertise in vaccine development and recommend experts in necessary fields\n\n- Offer (seed) funding for vaccine research and development (R&D)\n\n- Accelerate vaccine development in the interest of public health and societal benefits.\n\nBased on these expectations defined, amongst the different business model options we developed, the one we selected for further consideration is described below.\n\n- Novel concept of European Infrastructure business model\n\nThe option we are proposing is a combination of two different business models -namely a contract development partnership model and a bio-holding model- that are outlined below. This hybrid model was selected as it has the advantage of offering two different and independent types of revenue streams. Moreover, regarding the establishment of the RI, this hybrid model would facilitate the initial setting up of the infrastructure by starting with one of the models first (the contract development partnership) and subsequently implement the establishment of the other (the bio-holding model), which is the more challenging model to build.\n\n- Contract development partnership model\n\nUnder this business model option (Figure 1), the RI will offer to its customers a full suite of tailored value-chain and transversal scientific-technical and other services related to vaccine development and business building. This model assumes no access to (cash) funding. However, funding may potentially be provided to customers through in-kind contributions by the RI providing free or discounted services to customers in exchange for a small stake in the customers’ vaccine assets. Target customers are primarily early-stage vaccine developers and start-ups in need of specific services and expertise in early-to mid-stage vaccine development.\n\nBlue arrows indicate financial flows, grey arrows scientific-technical or other types of participation and contribution.\n\nThe particular value proposition of the contract development partnership model consists in the RI functioning as a product development partner for public and private institutions/vaccine developers, including academia, to which an RI and critical high-value services will be provided either free of charge or at a significantly discounted rate.\n\nThe monetization model of this business model (Figure 2) consists in revenues related to the provision of commercial services for which different payment options can be foreseen, such as upfront or milestone payments, royalties, or fees for services, and which will be negotiated on a case-by-case basis.\n\nThe most important key steps related to the implementation of the contract development partnership model would mainly involve the definition and establishment of professionalized services/service portfolio, including, for example, the setting up of platforms and operational and managerial procedures, the strengthening of the RI member network to ensure the availability of critical scientific-technical capacities, as well as building project management expertise.\n\n- Bio-holding incubator model\n\nUnder this model (Figure 3), the RI will function as an incubator for early-stage vaccine developers by providing funding and active business-building support, including value chain and transversal services. Selected vaccine candidates considered of interest (e.g. due to their indication, market potential, etc) will be in-licensed by the RI. Subsequently, using the financing, scientific-technical expertise and capacities available, the vaccine RI will continue the further development of the in-licensed vaccine candidates down to early- to mid-stage clinical development.\n\nBlue arrows indicate financial flows, grey arrows scientific-technical or other types of participation and contribution.\n\nFor this model, the RI will be established as a -biotech- holding legal entity that over time will build multiple subsidiaries. The holding company will be constituted by different research organizations (for example, members of the TRANSVAC infrastructure project and other organizations) that thereby will become members or partners in this venture. Each of the subsidiary companies will focus on one individual vaccine candidate (“asset”) that will be in-licensed at late discovery or preclinical development stages and subsequently be further developed in-house, using financing of the bio-holding and the scientific-technical capabilities and expertise of its member organizations. The development of vaccine candidates within the subsidiaries will thus allow them to draw on resources from and to share vaccine development risks with the parent holding company. Target organizations (“target customers”) for the in-licensing of vaccine candidates are small to medium-sized vaccine developers (public and private) with vaccine candidates for further development that require additional external R&D capacities, expertise and financing. The particular value proposition of this model is that the RI will become a business partner for pharma companies and as an RI will conduct R&D to support vaccine assets from discovery/preclinical stages until early-to-mid stage clinical development. Importantly, in addition to scientific-technical support, other value chain services provided by the RI under this model would include the provision of (seed) investments early in vaccine R&D projects, using financial resources from the RI’s own balance sheet.\n\nThe monetization model under the bio-holding model option is mainly related to the equity deal under which the RI will own a minority stake in the subsidiary companies established which advance the development of the in-licensed vaccine candidates. If and once the vaccine development has successfully completed phase I or phase II clinical testing, the vaccine candidates will be sold to a larger pharmaceutical company such that the equity value growth of the asset will be turned into cash upon exit (Figure 4). Moreover, the RI will receive a management fee from the investments allocated to it by the external investors to build its portfolio.\n\nThe key steps required to implement the bio-holding business model include the need to secure funding/financing (investments) from external investors, such as private equity or venture capital funds, investment banks or others whose financing will be used to cover the costs of the further clinical development of the in-licensed vaccine candidates as well as other operational costs of the RI. An additional requirement will be to build the critical fund management and business building expertise and capabilities within the management team of the RI.\n\nTable 3 provides an integrated summary of the major characteristics of the different model options described above.\n\n\n\n• Biotech holding company that builds multiple subsidiaries focused on individual assets in order to incubate projects quickly\n\n• Drawing on resources from and sharing drug development risks with the parent company\n\n\n\n• Provision of scientific-technical services\n\n• Option to enter into an equity-based partnership model to co-develop vaccines\n\n\n\n• Small and medium-sized vaccine developers (public and private)\n\n\n\n• Small and medium-sized private organizations with need for services in early stages of product development\n\n• Academic/public institution vaccine developers partners able to undertake certain types of R&D but in need of specific services/expertise\n\n\n\n• Vaccine candidates in preclinical stages\n\n• Mainly human vaccines\n\n\n\n• Human and veterinary vaccines\n\n\n\n• Provide best-in-class RI to conduct R&D\n\n• Become business partner for pharma companies\n\n\n\n• Providing RI and a pre-defined volume of services (free of charge or at significantly discounted rate to early-stage developments)\n\n• Functioning as development partner for public and private institutions (including academia)\n\n\n\n• Seed investments early in R&D projects, utilizing resources from own balance sheet\n\n• Support assets from preclinical stages to early-mid stage clinical testing\n\n\n\n• Provision of RI and services for transversal activities along the value chain\n\n• Potential for collaborative product development including dedicated resources and capacities\n\n\n\n• Equity-deal with minority ownership position in early-stage vaccine subsidiary companies\n\n• Gain through equity growth of the asset until end of life\n\n• Optional: Early cash-out to limit long-term risk from competition and price pressure\n\n\n\n• Different payment modalities (upfront or milestone payments, royalties, fees-for-services)\n\n• Option to free or discounted services in exchange for a small equity stake in the customers’ vaccine assets\n\n\n\n• Build fundraising and fund management expertise and capabilities\n\n• Secure funding (investment) from investors\n\n• Build business building expertise and capabilities\n\n• Professionalize services\n\n\n\n• Build project management expertise\n\n• Professionalize services\n\n• Build some business coaching expertise and capabilities\n\nSubsequently, in order to establish a series of explicit, measurable levels of output across a range of activities and to indicate the scale of value creation that is at stake, we undertook an ex ante assessment of the socio-economic impact that might be expected from a stable and sustainable European vaccine RI based on the hybrid business model outlined above. The period covered by this assessment includes the first ten years of operations of a vaccine RI based on the business model proposed. The assessment used the KPIs and methodology described in the Methods section in terms of three broad types of dimensions: (i) health impact, (ii) societal impact and (iii) economic impact (Figure 5).\n\nThe ex ante socio-economic impact of the vaccine RI with the proposed hybrid business model was assessed along the three dimensions indicated.\n\nThe health impact dimension refers to innovations in vaccine R&D (such as in terms of tools/technologies/solutions developed, vaccines developed and vaccine-related impacts, e.g. numbers of future deaths prevented and quality of life improvements); the societal impact dimension refers to dissemination activities and new publications, human resources activity, education and training and the value of new vaccines brought to the market; and finally, the economic impact refers to vaccine industry, expected numbers of new jobs created, business and related outputs, impact on SMEs, impact on research institutes and networking activities. Apart from these dimensions, the KPIs were split across three categories in order to indicate the time horizon of their measurement, selected to best mirror the progress of the RI, namely (i) short-term, (ii) long-term cumulative impacts, and (iii) an operational metrics to be used for monitoring of the RI execution. The estimated impacts for all three dimensions are summarized in Table 4.\n\nImpacts across three different dimensions have been estimated for the first 10 years of the RI’s operation period, in addition to other operational impacts for the tracking of activities. Each measure indicated in the left column corresponds to an individual KPI.\n\n\n\n(a) Health impact\n\nThe ultimate objective of this initiative is to support and enhance the vaccine R&D ecosystem in order to address both the financial ‘translational valley of death’ (the funding gap between early research and clinical trials that makes the translation of basic science into clinical candidates so challenging) and a more general capabilities gap in early development stages. This will be realized over a longer period of time, and as a result is expected to produce more novel vaccine candidates that successfully advance through clinical trials and into medical practice.\n\nThe expected number of new vaccines in development will reflect candidates whose origination and development have been supported by the vaccine infrastructure activities. This requires tracking vaccines through each stage of their development – in other words, the number of candidates that have reached key successive milestones (i.e. the number of new vaccines proceeding to first-in-human trials, reaching proof-of-concept stage and entering the market). According to an analysis of annual trends data5, the average length of the development cycle from phase 1 to market is ~7.5 years (outside of pandemic and seasonal situations). This has therefore to be a long-term metric and one which encompasses the entirety of the first ten years of the initial RI operation. Our expectation of one novel in-market project being enabled in ten years is consistent with the vaccine infrastructure producing one late-stage novel project over its first five years of operation. This is also consistent with 15 new early-stage vaccines being supported throughout this time period. Considering the average length of vaccine development (for complex projects it can take 12 years or more from preclinical to end of phase 3), the majority of these projects will not have reached clinical adoption by the end of that first decade (Terry et al., 2018). For projects that would progress to the mid-late stage clinical testing of vaccines, industry-average success rates are applied to account for inevitable attrition to arrive at approximately one novel infrastructure-supported vaccine reaching approval within the first decade.\n\nSuch a vaccine would likely be followed by other candidates, so impact could scale up beyond that first decade as more projects progress through clinical trials. If we consider the range of vaccine-specific probabilities of success and likely development cycle time, between 2 and 4 of the 15 supported assets would be expected to reach successful launch. This impact could be further expanded by improvements to existing licensed vaccines (e.g., other routes of administration or to make production more cost-effective). Such incremental changes typically do not require a full development process and hence such new products can be adopted more quickly and at a lower risk. Several such products could thus reasonably be anticipated over the following 10-20 years (even though the prospective health impact of vaccine improvement is more variable and harder to estimate given that it would depend on the needs left unmet by current vaccines).\n\nThe true impact of novel vaccines reaching market is their actual benefit for patients and health systems. This can be expressed as the expected number of future deaths, severe cases prevented and the expected disability-adjusted life year (DALY) improvement. For one new vaccine estimated to be introduced to market after the first ten years (accounting for the aforementioned time and risk), the actual impact on population health may be estimated. The accuracy of such an estimate is subject to significant uncertainty given variations in the prevalence, virulence, mutation rates and severity of the kinds of disease that might be targeted by such a vaccine – each of which affect burden, mortality, achievable vaccine effectiveness and adoption rate.\n\nSeveral approaches exist for modelling the impact achieved by recently-introduced vaccines. Based on models published in the medical literature, we suggest two scenarios may be helpful to consider in arriving at an understanding of the scale of health impact achievable for a single new vaccine: a COVID-19-based case to illustrate the potential benefits of a novel vaccine in a pandemic setting, and an influenza vaccine efficacy improvement scenario to illustrate the benefits achievable for common endemic diseases (even ones already treated with vaccines that are not yet fully optimized).\n\nThe real-world impact of a novel product might fall somewhere between these low and high outcomes, whereas potentially being further enhanced by more of the 15 novel vaccine candidates supported by this effort reaching market over a longer time period. Other, smaller incremental projects might be supported that in turn might reach market sooner (and with lower risk), but in turn their potential health impact could be expected to be more incremental -typically only improving upon existing vaccines- and as a result too elusive to model separately.\n\nHigh-impact scenario (pandemic vaccine)\n\nBased on recent assessments made with respect to COVID-19, the introduction of vaccines prevented approximately 4.3 million deaths (from 8 December 2020 to 8 December 2021) in the European region alone (Watson et al., 2022)6. Acknowledging that within the assessment timeframe there were in total four novel SARS-CoV-2 vaccines approved for use across the European region, we estimate that on average, the annual impact of a single vaccine in this case was over a million lives saved in Europe.\n\nIn parallel and considering the overall effect of COVID-19 in terms of DALYs, a recent estimate concluded the DALY impact of COVID-19 in just Scotland and only in 2020 was 102,350 (Wyper et al., 2022). This calculation reflects the specific demographic and healthcare characteristics of just one region of the UK but considering the size of the population of Scotland in relation to that of the entire WHO European region (approximately 0.6% of the total), an equivalent effect for the region might be very approximately estimated as being more than 150 times as great, or over 15 million DALYs.\n\nThe DALY decrease linked to vaccination can be anticipated to be in proportion to the ratio of deaths prevented to total projected deaths as modelled (Watson et al., 2022) (72% in first year of vaccination accounting for vaccination coverage and efficacy). Applying this to our approximate estimate of 15 million for the DALY effect of a new pathogen resembling SARS-CoV-2 implies a vaccine impact in excess of 10 million DALYs in the region, and if four vaccines were again to be introduced and with approximately equal effect, an impact for a single vaccine on the order of 2.5 million DALYs (Table 4). The conversion of deaths to DALYs may vary for different infections, for example SARS-CoV-2 resulted in large numbers of individuals with Long COVID.\n\nLow-impact scenario (influenza vaccine efficacy improvement)\n\nThe second scenario assumes an efficacy improvement and administration optimization for an existing (but not optimized) vaccine against a common but-usually-less severe endemic disease. It is based on modelling influenza and its seasonal vaccine of 2017-18 (characterized by lower than usual efficacy) compared to a more optimized variant (Sah et al., 2018). Compared to no vaccination, a low-efficacy vaccine already drives significant health benefit -for instance, a 20% efficacy flu vaccine administered at ~40% coverage is projected to avert more than 21 million infections, ~130,000 hospitalizations, 61,812 deaths, and 2.2 million DALYs in the US (Sah et al., 2018). With efficacy increased to 40%, the scale of health benefit can be doubled. Scaled for Europe (based on the total population of Europe and the USA as of 2020), this would imply approximately 28 million infections, 175 thousand hospitalizations, more than 80,000 deaths, and 3 million additional life years saved.\n\nThe health benefits represented by DALYs and deaths averted are further strengthened by the economic impact on the health systems. This is evidenced by the cost-effectiveness of vaccination programs, with most costing less than $50 per life gained and thus orders of magnitude cheaper than the prevention of many non-infectious diseases such as diabetes or hypertension (Bloom, 2011; Ehreth, 2003) (Table 4).\n\n(b) Societal impact\n\nThe vaccine infrastructure can deliver positive societal impact both directly and indirectly. The indirect societal consequences of a successful vaccine deployed at scale are potentially enormous. This set of indirect impacts has not been assessed.\n\nIn terms of direct impact, it is possible to anticipate the number of new jobs created. By attributing new jobs solely to those in new SMEs as a consequence of a more assured pathway between innovation and clinical trials, and assuming an average of 5-10 employees for each, this would imply 100-150 new positions over ten years. The broader perception of the European vaccine development industry can be enhanced by a vigorous publication record and effective communications (see below). Further, a target can be set for the percentage of researchers undergoing training who experience an improvement in their knowledge base/knowledge capital, as quantified by post-course surveys. These targets are set as 75% of trainees reporting at least a ‘good’ level of improvement in their knowledge base in post-course surveys, and 50% of trainees reporting ‘excellent’ improvement, based on historical feedback from previous training courses organized in the context of the TRANSVAC project7 (unpublished data).\n\nMore broadly, a target may be reasonably set for media appearances at 30 separate appearances a year, based on past TRANSVAC dissemination rates (this including press releases, website launches and social media campaigns; and also a target for the number of research partners trained through the vaccine infrastructure (assuming nine training courses in total with an average of 15 attendees per course, this can be set at 135 over the ten-year period in question). All these activities help ensuring a positive momentum for the sector by building awareness, disseminating findings and contributing to reputation building for the sector (Table 4). This is particularly critical for the introduction of new vaccines whose benefits could be undermined by false information on social media, supported by the increase in vaccine hesitancy in some communities.\n\n(c) Economic impact\n\nAny vaccine that is successful in preventing or ameliorating serious disease is likely to have a significant and sometimes very substantial indirect economic impact (see, for example, Atkins et al., 2018; Sandmann et al., 2022; Portnoy et al., 2023). Positive effects are also going to be observed for the local life science innovation ecosystem and industry – those effects being achieved even before any successful products reach the market and contribute to increased competitiveness and economic health. For the purposes of this socio-economic assessment analysis, however, only direct economic impact is considered.\n\nThe KPIs selected for measuring economic impact would likely require observation and targeting in respect to expected revenue generated. Our business plan modelling suggests total revenue on successful exits after ten years amounting to €159 million, but given the long-term nature of vaccine development, these revenues would be most likely to accrue towards the end of the period. To put this value in context, average annual European sales of novel non-COVID vaccine products were €175 million in 20218. The expected value of funding attracted is assumed to be €100 million over ten years. The expected value of licensing deals is aggregated with an expected revenue generation of €159 million over ten years (i.e. much or all of the revenues obtained from the initiative will be realized in the form of licensing deals).\n\nThese are not formal targets but rather reasonable estimates of what could be expected based on the extrapolation into the future of the activities conducted under the TRANSVAC project. Revenue and investment values contribute directly to the reinvigoration of local R&D ecosystems. Moreover, it is assumed the vaccine infrastructure’s activity may also lead directly to SME creation by facilitating access to funding and capabilities – with the target for the number of SMEs created being 15 over the first ten years of the initiative.\n\nIn addition to large deals and the birth of new SMEs, the vaccine infrastructure can also provide a range of scientific services. These may vary significantly in scope from technical capabilities and execution of experiments to more comprehensive offerings including study design and a more complex execution capability. The precise services offered will be finalized in the future with consideration for the demand for specific services during the past TRANSVAC projects as well as stakeholder surveys we conducted. The most popular services have included antigen expression/production, adjuvant formulation testing, animal models for infectious diseases, immunocorrelates analysis, and regulatory support.\n\nWithin the ten-year period, a contract development partnership model could be expected to support a total of 33 projects, with 50 instances of services provided. The predicted revenue stream generated by these services is small, estimated at ~€0.2 million per year (in line with currently observed inflow from ongoing projects), contributing an additional €2 million to the overall consortium revenue over ten years. The main economic value, however, is in the invigoration of the ecosystem and enabling academic and industry innovators to access capabilities they usually would be unable to maintain in-house (and that might otherwise be inaccessible, considering the limited scope of contract research organisations (CROs) services offered within early-stage vaccine R&D).\n\nIn addition to financial metrics, the vaccine infrastructure can also make a direct contribution to the generation of new innovations and intellectual property by supporting vaccine R&D. As part of this, the number of new patents issued would be expected to equal TRANSVAC’s historic performance (i.e., one approximately every four years). Such patents represent meaningful improvements in R&D processes (e.g., formulation or technical methods) and thus facilitate development of novel products across the ecosystem (Table 4).\n\n(d) Operational impact\n\nThe most important consequences of the vaccine infrastructure are only fully measurable over the long term due to the characteristics and length of the vaccine development process, which must account for manufacturing, rigorous clinical trials and regulatory evaluations. It is therefore especially important to identify operational activity variables which can be reported on a more frequent basis, but which can be reasonably associated with long-term success. The expected number of publications may be observed and targeted (five a year on a two-year time lag to allow for the process of paper preparation, submission, review and acceptance based on TRANSVAC’s historic experience) and the expected number of new scientific services established as an offering to researchers (one new service offered every two years, coming to a total of five over the ten-year period). The number of vaccine projects supported at any one time can be tracked (five to ten projects, as mentioned above). Finally, a robust level of engagement can be targeted in terms of the organization of stakeholder and investor meetings (12 a year, to cover a variety of combinations of stakeholders and/or investors) (Table 4).\n\n\nDiscussion\n\nRIs are fundamental enablers of science, research, innovation and education. They facilitate cutting-edge scientific investigations and support the development of innovative solutions to various challenges. However, RI are also known to be notoriously expensive in their establishment and maintenance, and the costs for their establishment alone often range from millions to even billions of Euros, depending on the scientific discipline and type of infrastructure. Very frequently, public funders bear most of this burden. This is also true for most if not all already of the existing RI in Europe. The major source of funding for the existing RI that are part of the ESFRI Roadmap, is public funding provided by different European Union (EU) Member States’ governments although most of them through their operations aim to generate revenues e.g. by providing commercial services. Most of these RI also obtain additional funding via competitive EU and other types of grants (i.e., additional public funding). So, although probably all of the existing RI have revenue comprising a mixture of financing from different sources, the largest share by far of their total income comes from public funding sources without which none of the RI would be able to survive or operate.\n\nBeing acutely aware of the levels of financial commitments and other responsibilities that this near-absolute dependance of RIs on public money entails, governmental funders and decision makers have a keen interest in promoting the development of alternative business models that may allow RIs to become sustainable and operate more independently of public funding.\n\nStriving for sustainability has also been the major rationale underlying the TRANSVAC design study. A well-designed and stable vaccine R&D RI could play a major role in tackling existing challenges and in advancing innovation in the field. We have demonstrated this in the successful implementation of the TRANSVAC projects, that—thus far with the exclusive financial support of the EU—obtained important achievements. For example, the TRANSVAC and TRANSVAC2 projects have together reviewed 158 applications for free services and approved 88 projects from 19 countries. Nearly a quarter (18) of those projects involved services provided by multiple institutions. TRANSVAC2 has also organised two rounds of a new, free 14-course vaccinology training program. More than 400 trainees from academia and industry attended, helping to fill an important training gap in the field (Geels et al., 2015; Martin et al., 2023). Beyond the provision of scientific-technical services, TRANSVAC was also charged with conducting a feasibility study with the aim of developing and proposing an alternative business model for a sustainable European vaccine RI. As the first step in this exercise, we undertook a detailed gaps-and-needs analysis of vaccine R&D in Europe (Jungbluth et al., 2022), the outcomes of which informed the subsequent steps. The most important findings from this assessment that shaped the development of alternative business models for a vaccine RI were the lack of funding and financing available for vaccine R&D, the need for expertise and guidance to vaccine developers, and the difficulty in accessing critical technologies, platforms and other key infrastructure.\n\nThese findings present an opportunity for a new initiative and formed the basis for the hybrid business model we have presented here. This hybrid model offers a mixture of scientific services for potential vaccine products and support for business development. Regarding vaccine development, the vaccine RI we envision will offer a portfolio of services emphasizing high-value scientific-technical services, provided on a commercial basis with different payment modalities or the option to enter into an equity-based partnership model. Importantly, beyond the provision of services to “external” vaccines, the bio-holding model contained in our business model will go further and in-license into the vaccine RI selected vaccine candidates that have been identified and considered of interest. Financing will be raised from external investors such as private equity or venture capital funds, and investment banks to fund in-house development of these in-licensed vaccine candidates through phase I and phase II clinical testing. If these clinical steps are successful, the vaccine assets will be sold to pharma companies for late-stage development and commercialization, with the equity share corresponding to the vaccine RI in these assets being the major revenue stream for the infrastructure in the long term.\n\nImportantly, by combining two complementary business models, our hybrid model offers the advantage of two different types of revenues. One is based on income linked to the provision of scientific-technical services on a commercial basis; income for the RI in this case will mainly consist of a brokerage fee charged for the mediation of services between the customer and the service provider. The second, and more important revenue in the long term, is linked to the expected equity growth of the in-licensed vaccines (of which the RI would hold a minority stake) that will generate cash upon sale to pharma companies. Although smaller in amount compared to the income from the bio-holding model, the revenue via the contract development partnership model has the advantage of offering a continuous stream of income which can cover the operating costs of the RI (e.g. human resource-related costs), especially in the first years of the RI. As mentioned above, the bio-holding model income is expected to be more important in the long run, but is only expected at a later stage of the implementation of the vaccine RI due to the more complex requirements, the risk of failure of any single vaccine candidate, and the time required to reach clinical trials. The contract development partnership model thus provides a smoother entry for the establishment of the bio-holding model and represents an overall measure of financial risk reduction. Moreover, the different types of requirements linked to the establishment of the two different models will enable a relatively fast and straightforward establishment of the contract development partnership in the early stages of the overall venture, subsequently allowing the RI to concentrate more efforts on the establishment of the bio-holding model. The bio-holding model clearly is the more demanding venture to establish due to, for example, the need to mobilize significant external financing and to scout and manage the in-licensing of vaccine candidates. For a vaccine RI based on our proposed business model to function in the long run, it is important to emphasize that it will only work if the bio-holding model can be implemented successfully, as the revenue stream based on the contract development partnership only would not be sufficient to sustain the operation and running of the RI.\n\nAs mentioned before, the development of this business model was informed by stakeholder consultations we conducted to ensure that the model aligned as closely as possible to stakeholder needs and interests. For the same reason, once this model had been developed and refined, we returned to several key stakeholders, including researchers from public and private organizations, investors and investment banks, to pitch the model to them. The feedback we received from this exercise was largely positive and encouraging. Overall, the model we have developed was considered relevant and feasible—but challenging—to implement.\n\nThe ex ante impact assessment we conducted shows that a vaccine RI based on the model we have developed is likely to have a very important impact along the three dimensions we explored (the health, societal and economic spheres). These dimensions are discussed individually below. However, it must be acknowledged that vaccine development is inherently uncertain and despite utilizing the best available evidence and making the estimates in good faith, these estimations remain subject to significant potential positive or negative variance. Although there is a lot of literature and thinking around ways of measuring the impact (including ex ante) of aRIs (see, for example, ESFRI, 2023), we are not aware of any ex ante socio-economic impact assessment for a biomedical RI to the level of detail and depth we have undertaken for a vaccine RI.\n\nIn the health dimension, our estimates indicate that - using our investment target of €100 million - a total of 15 new vaccines can be supported from preclinical stages to phase 2 clinical testing, of which during the 10-year time frame of our estimations we expect 1 vaccine candidate can be sold to pharma industry (for the assumptions underlying our estimations, such as vaccine development timelines, attrition rates, costs, etc., see the Methods section). For estimating the eventual public health impact of the vaccines supported by the vaccine RI, we have used two different scenarios, one based on a high-impact case (such as a pandemic vaccine similar to COVID-19 vaccines), and another scenario based on a low-impact case (such as a seasonal influenza vaccine with improved efficacy). The expected numbers of future deaths and severe cases prevented for the low and high impact scenarios, respectively, range from 80,000 to 1.1 million deaths averted per single vaccine. This estimation does not include any of the other vaccine candidates in the vaccine RI pipeline beyond the 10-year timescale of our estimations; if those were also considered then expected impact would be higher. Regarding the improvement of DALYs, we expect between +2.5 million and +10 million DALYs saved per single vaccine for the low- and high- impact scenarios, respectively (also here considering only the 10-year timeframe).\n\nOne has to keep in mind that these estimated health impacts only relate to the vaccine candidates that will be in-licensed and developed in-house by the vaccine RI (via the bio-holding model) and do not include any “external” vaccines whose development will be supported by the commercial scientific-technical services provided by the RI (via the contract development partnership model). Although many of these “external” vaccines can be expected to have additional positive impacts on public health in the long term, estimating this impact proved too speculative and therefore was not included in our overall impact assessment. Overall, considering the level of initial investment our venture would require, we consider that our model for a vaccine RI has the potential to achieve a very substantial positive impact on public health.\n\nRegarding the societal impact, we estimate that between 100-150 new positions will be created in direct relationship to the model we propose. On one hand, there will be a certain number of new positions linked to the management headquarter of the RI itself. Most of the new positions, however, will be linked to the establishment of the subsidiary companies that will be established under the parent holding company. As we foresee that a single company will be established for each vaccine candidate in-licensed to the vaccine RI, for the 10-year period of the currently planning we estimate a total of up to 15 subsidiaries. Each of these subsidiaries will be relatively small in size but require a certain number of scientific-technical staff. The number of subsidiary companies could also be somewhat lower, as some vaccine candidates developed in-house by the vaccine RI may not require formal in-licensing and establishment of a new, dedicated subsidiary company.\n\nThe other impacts in the societal dimension relate to training offered via the vaccine RI and the overall number of media appearances of the venture. These estimations are directly extrapolated from data from the TRANSVAC projects and are considered highly reliable. Overall, we consider the realistic societal impact as defined here to be significant, although less critical than the health impact due to the nature and size of the proposed venture.\n\nFor the economic dimension, we have estimated the impacts linked to both parts of our business model (the bio-holding and contract development partnership aspects). Based on our experience providing scientific-technical services through the TRANSVAC projects and the experience of already existing RI in the biomedical field that provide commercial services, the direct economic impact related to the provision of services by a vaccine RI can be estimated in a very reliable manner. This relates both to the number of commercial services provided and the corresponding revenue. As mentioned above, as the vaccine RI we are proposing for the service provision functions mainly as a broker, and the direct income of the central infrastructure related to this business model essentially corresponds to a brokerage fee. Although representing only a minor part of the overall revenue generated by the RI in the long term, it will nevertheless be important for starting the entire venture and facilitating the transition towards the establishment of the bio-holding model and also addresses an important existing need in the vaccine R&D community. However, most of the revenue is eventually expected to result from the bio-holding model, namely the growth in equity that has been generated between in-licensing and selling the vaccines after their further development. This business model is based on large pharma companies buying these vaccine assets after successful phase I or phase II clinical stages. Vaccines must have commercial potential to maintain relevance for large pharma. Consequently, at least during the first ten years or so of its existence, a vaccine RI based on our model for the bio-holding venture will--and must--mainly target vaccines of relevance for high income markets in order to become sustainable. Once sufficient profit has been generated by the vaccine RI, eventually the capital available can also be used to in-license and further develop vaccines for neglected diseases with more limited commercial perspectives. Vaccines of this kind could also be supported earlier in case financing from philanthropic investors or public funding are mobilized for the vaccine RI to allow direct support for products with less market potential.\n\n\nConclusions\n\nOur consortium has attempted to develop a business model for a sustainable vaccine RI that would be able to operate without a dependence on public funding and estimated its impact using the most reliable data possible and based on widely accepted assumptions and existing evidence. Nevertheless, substantial variations and uncertainties are inherent to this type of modeling and ex ante assessment. Still, both our model and the estimations will become increasingly better and reliable the more we advance with this venture and with the particular steps we will achieve. Uncertainties will be replaced by facts, and new data will become available both from our own work, as well as from those of others working in this field.\n\nTo conclude, we believe that our model can be viable and that the expected impacts are sufficiently high to merit advancing. The next step in the establishment of a vaccine RI at this particular moment is the selection both of the country for the RI’s headquarters, and -linked to this- the selection of a legal entity option considered suitable for our particular model. We are currently in the process of defining the key criteria and aspects to be used for assessing and comparing different legal entity options, considering aspects such as the role and liability of members/shareholders, costs of establishment, reporting, audit and accounting requirements, supervision by the authorities, and taxation issues. Given that the mission of the envisioned vaccine RI is to accelerate vaccine development in the interest of public health and societal benefits, legal entity options will be prioritized and explored that allow the establishment of the RI as a non-profit venture.\n\nVaccine development is directly linked to a better future for Europe and beyond. We believe that a well-designed and sustainable European vaccine RI has the potential to promote the development of the industry and a significant regional and global health impact.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nAcknowledgements\n\nWe gratefully acknowledge the valuable input provided by the members of the independent external scientific advisory committee to this project. Moreover, we are grateful to all external stakeholders who have participated in different parts of the process and who with the generous sharing of their knowledge have supported and encouraged us in this study. We also want to thank all partners in the TRANSVAC2 project for providing their excellent scientific-technical services, training and other types of support to the scientific community.\n\n\nReferences\n\nAtkins KE, Lafferty EI, Deeny SR, et al.: Use of mathematical modelling to assess the impact of vaccines on antibiotic resistance. Lancet Infect. Dis. 2018 Jun; 18(6): e204–e213. Epub 2017 Nov 13. PubMed Abstract \| Publisher Full Text\n\nBeck E, Biundo E, Devlin N, et al.: Capturing the value of vaccination within health technology assessment and health economics: Literature review and novel conceptual framework. Vaccine. 2022 Jun 26; 40(30): 4008–4016. PubMed Abstract \| Publisher Full Text\n\nBloom DE: The value of vaccination. Adv. Exp. Med. Biol. 2011; 697: 1–8. PubMed Abstract \| Publisher Full Text\n\nDoherty M, Buchy P, Standaert B, et al.: Vaccine impact: Benefits for human health. Vaccine. 2016; 34: 6707–6714. Publisher Full Text\n\nEhreth J: The global value of vaccination. Vaccine. 2003 Jan 30; 21(7-8): 596–600. PubMed Abstract \| Publisher Full Text\n\nESFRI Working Group Report: Monitoring of Research Infrastructures Performance.December 2019.\n\nESFRI Roadmap: Strategy report on research infrastructure.2021.\n\nESFRI Assessment of impact of RIs: Policy Brief.May 2023. Publisher Full Text\n\nGeels MJ, Thøgersen RL, Guzman CA, et al.: TRANSVAC research infrastructure - Results and lessons learned from the European network of vaccine research and development. Vaccine. 2015 Oct 5; 33(41): 5481–5487. PubMed Abstract \| Publisher Full Text\n\nGreenwood B: The contribution of vaccination to global health: past, present and future. Philos. Trans. R. Soc. Lond. Ser. B Biol. Sci. 2014; 369: 20130433. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIPROVE: A strategic European roadmap for the vaccine of tomorrow.2016. Reference Source\n\nJungbluth S, Depraetere H, Slezak M, et al.: A gaps-and-needs analysis of vaccine R&D in Europe: Recommendations to improve the research infrastructure. Biologicals. 2022 Apr; 76: 15–23. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLeroy O, Geels M, Korejwo J, et al.: Roadmap for the establishment of a European vaccine R&D infrastructure. Vaccine. 2014 Dec 5; 32(51): 7021–7024. PubMed Abstract \| Publisher Full Text\n\nMartin WJ, Slezak M, DiMarzo R, et al.: Vaccine Research and Development Infrastructure in the European Union: Establishing Support Through Integration. BioProcess International. May 2023. Reference Source\n\nOECD: Reference Framework for Assessing the Scientific and Socio-Economic Impact of Research Infrastructures. OECD Science, Technology and Industry Policy Papers; 2019; 65.\n\nPortnoy A, Arcand JL, Clark RA, et al.: The potential impact of novel tuberculosis vaccine introduction on economic growth in low- and middle-income countries: A modeling study. PLoS Med. 2023 Jul 11; 20(7): e1004252. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nReid A, Griniece E, Angelis J: Evaluating and Monitoring the Socio-Economic Impact of Investment in Research Infrastructures. Technopolis. Technical Report.January 2015.\n\nRodrigues CMC, Plotkin SA: Impact of Vaccines; Health, Economic and Social Perspectives. Front. Microbiol. 2020 Jul 14; 11: 1526. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSah P, Medlock J, Fitzpatrick MC, et al.: Optimizing the impact of low-efficacy influenza vaccines. Proc. Natl. Acad. Sci. U. S. A. 2018 May 15; 115(20): 5151–5156. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSandmann FG, van Leeuwen E , Bernard-Stoecklin S, et al.: Health and economic impact of seasonal influenza mass vaccination strategies in European settings: A mathematical modelling and cost-effectiveness analysis. Vaccine. 2022 Feb 23; 40(9): 1306–1315. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTerry RF, Yamey G, Miyazaki-Krause R, et al.: Funding global health product R&D: the Portfolio-To-Impact Model (P2I), a new tool for modelling the impact of different research portfolios. Gates Open Res. 2018 Jul 2; 2: 24. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWatson OJ, Barnsley G, Toor J, et al.: Global impact of the first year of COVID-19 vaccination: a mathematical modelling study. Lancet Infect. Dis. 2022 Sep; 22(9): 1293–1302. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWyper GMA, Fletcher E, Grant I, et al.: Measuring disability-adjusted life years (DALYs) due to COVID-19 in Scotland, 2020. Arch Public Health. 2022 Apr 1; 80(1): 105. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\n\nFootnotes\n\n1 www.transvac.org\n\n2 Pharmaprojects. https://pharmaintelligence.informa.com/products-and-services/clinical-planning/pharmaprojects\n\n3 Evaluate. www.evaluate.com/\n\n4 Pharma Deals: https://www.pharmadeals.net/about\n\n5 Evaluate. www.evaluate.com/\n\n6 WHO European Region comprises: Albania, Andorra, Armenia, Austria, Azerbaijan, Belarus, Belgium, Bosnia and Herzegovina, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, France, Georgia, Germany, Greece, Hungary, Iceland, Ireland, Israel, Italy, Kazakhstan, Kyrgyzstan, Latvia, Lithuania, Luxembourg, Malta, Moldova, Monaco, Montenegro, Netherlands, North Macedonia, Norway, Poland, Portugal, Romania, Russia, San Marino, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Tajikistan, Turkey, Turkmenistan, Ukraine, United Kingdom and Uzbekistan.\n\n7 www.transvac.org\n\n8 Evaluate. www.evaluate.com/" }	[ { "id": "218383", "date": "08 Nov 2023", "name": "Wolfgang Fecke", "expertise": [ "Reviewer Expertise Research infrastructures", "chemical biology", "drug discovery" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis is a very interesting study for the development of a business plan for a vaccine research infrastructure which could become relevant also for other biomedical not-for-profit organizations. Some aspects are less well covered, in particular the area of competing with commercial CROs in the vaccine field. How many customers would find it more attractive to work with an academic not-for-profit rather than with a professional service provider remains unclear.\nThe KPIs listed in Table 2 rely mostly on unpublished data and are therefore not easy to use by others (hence my 'partly' answer in terms of data reproducibility) although some numbers are mentioned in Table 4. In that respect, the number of new patents appear fairly low (one every four years), given the number of projects (33) and incubated companies (15) within 10 years.\nOne minor point is a certain redundancy of information in Figures 1-4, these could be streamlined.\nn the discussion the costs of running the infrastructure are mentioned in the sense that the service model cannot cover these costs. It would be helpful to have actual figures available on which this statement is based. What is the estimated income from the brokerage fees, and what are the personnel and other operational costs of the infrastructure? For instance, costs for general administrative or financial services through third parties could be obtained. Additional revenue stream of the infrastructure are possibly overheads derived from EC-funded projects - these are not taken into account at all although historical figures from the TRANSVAC projects exist.\nFinally, the funding gap at least in the first years of the infrastructure could still be covered by membership contributions from countries although it is laudable that the authors attempt to design a fully sustainable model without public funding. Maybe it would be worth mentioning in the discussion that this option exists.\nOverall, the article will be useful also for other biomedical infrastructures to support their business plans - as the authors mentioned in their discussion, another publicly available study which such detailed numbers does not exist for an infrastructure or would be at least hard to find.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "234698", "date": "30 Jan 2024", "name": "Ingrid Kromann", "expertise": [ "Reviewer Expertise Vaccine CMC/manufacturing development" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nI have carefully reviewed the manuscript and I appreciate the opportunity to provide constructive feedback on the paper. Summary: The paper addresses an important topic—coordinated vaccine development infrastructure—and highlights the significant need for sustainability in this area. The manuscript effectively discusses the challenges faced during the initial stages of vaccine development, focusing on the transition from preclinical research to clinical research, commonly known as the first valley of death. However, to support the business model the paper could be strengthened to provide a more comprehensive understanding of late stage development, matchmaking with industry and commercialization, particularly regarding the vaccine portfolio and region in scope (second valley of death). Specific Suggestions: 1: The paper would benefit from a more detailed explanation of the specific regions and vaccine areas the proposed infrastructure is intended to address. It is crucial to clearly outline if it is the unmet needs and/or EPI vaccines the infrastructure aims to target. This clarification will enhance the paper's relevance and applicability to diverse stakeholders involved in vaccine and business model development. 2: While the manuscript extensively covers the challenges associated with the first valley of death, it briefly touches upon the critical second valley of death, which involves the transition from late-stage development to commercialization. The paper should delve deeper into this aspect, providing a more thorough analysis of the business model's dependency on licensed/commercial products. A comprehensive discussion on the funding requirements and the importance of strategic matchmaking for successful transition is crucial. Additionally, detailing the industry's interest in vaccines and emphasizing the correlation between vaccine area selection and business model success will strengthen the manuscript. 3 The manuscript briefly mentions the industry's interest being dependent on the demand for commercial products. Expanding on this point and elucidating how the selection of vaccine areas influences industry engagement will provide a more nuanced perspective. The paper should discuss how aligning vaccine development with market demand enhances the likelihood of attracting funding and industry support. 4 The conclusion could be strengthened by summarizing the key initiatives needed and emphasizing the interdependence of geographical focus, thematic relevance, and business model considerations in the proposed vaccine development infrastructure. It is essential to reiterate the paper's contributions and provide a clear call to action for policymakers, researchers, and industry stakeholders. In conclusion, while the manuscript addresses a crucial topic and provides valuable insights into the challenges of vaccine development infrastructure, further refinement in the areas mentioned above will enhance the paper's clarity, depth, and overall impact.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "234689", "date": "30 Jan 2024", "name": "Sana Zakaria", "expertise": [ "Reviewer Expertise health and technology policy research", "experimental medicine research" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\n1. Spelling correction for poliomielitis required. It should be spelled poliomyelitis.\n2. It is unclear what the scope of the proposed European vaccine R&D infrastructure would be. The intro mentions that RIs range in scope so it would be helpful to define what scope has been considered in this assessment. I.e. is it inclusive of translational research and preclinical/clinical testing? This is somewhat alluded to at the start of the results but should be more explicit as these boundaries could heavily impact the monetisation models presented.\n3. The monetisation models are really interesting but there is little assessment of whether these models would be financially viable and how the upfront RI is expected to be financed within the hybrid access model. It would be good to have some worked up examples on cost of vaccine research and establishment of RI against the benefits anticipated. In terms of upfront establishment of the RI, the models proposed here do not talk about this but rather develop models that could provide varied revenue streams for sustaining/maintaining existing RI. This should be specified upfront as the following sentence is misleading: \"The model we have developed integrates two different types of business models that would provide the infrastructure with two different types of revenue streams which would facilitate its establishment and would be a measure of risk reduction.\"\n4. The financial modelling details would be valuable to include as a supplementary appendix to understand the basis of the estimations proposed.\n\n5. I really like the nuancing of health impacts as presented especially in different settings.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? No\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "234705", "date": "15 Feb 2024", "name": "Massimo Florio", "expertise": [ "Reviewer Expertise Public economics" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nI have read this article with interest because of my own work for the European Parliament and other bodies about the need of European Biomedical Research Infrastructure, and more recently about the evaluation of the EU response to Covid 19 pandemic.\n\nI provide some links:\n\na)https://www.europarl.europa.eu/cmsdata/250660/Study%20presentation%20STOA%20panel%2016.12.2021.pdf b) htts://www.europarl.europa.eu/thinktank/en/document/IPOL_STU(2023)740072 I would also mention this new study by Gamba et al: c) https://www.europarl.europa.eu/stoa/en/home/news/details/new-stoa-study-on-better-access-to-medic/20200309CDT03501. The Authors may take advantage of the literature, empirical results, and policy options discussed there. Moreover, they may consider that on July 2023 the European Parliament has voted a recommendation to the EC and the Council. d) \"605. Calls on the Commission and the Member States to create a large-scale, mission-oriented, public European health R&D infrastructure which operates in the public interest to manufacture medicinal products of health and strategic importance for healthcare, in the absence of existing industrial production, in order to support the EU to overcome market failure, guarantee security of supply and prevent possible shortages of medicines, while contributing to greater preparedness for facing new health threats and emergencies;\" https://www.europarl.europa.eu/doceo/document/TA-9-2023-0282_EN.html Finally, they are by sure aware of the R&D implications of the EUPharma Strategy and the ongoing revision of the legislation on medicines. In the next months the European Parliament needs to vote on it position for future negotiations with the EC and the Council. In this context, the proposals by the Authors, stemming from their experience in Transvac, a Horizon2020 funded consortium, are welcome. However, their ambitions are limited and the hybrid model they propose is perhaps not entirely appropriate to the objective of a EU wide RI. It seems that on one side the model would be a service provider to third parties, such as pharma stat-ups, SMEs, Universities and so on. The Authors list some services but it is not entirely clear to me why the proposed RI with, as it seems, limited own staff and budget, should make a difference in the European Panorama with existing Contract Organizations, except probably because the services would be sold at low cost, reflected in minimal revenues in the sketched business plan. I understand this venture as a not-for-profit entity, perhaps a distributed RI, but there are others in the EU, for example for clinical trials, etc. Thus the Authors should show why they would be able to offer something not already available and precisely what and to whom. Having said this the Authors turn to an entirely different model, where the RI would act as sort of venture capitalist for certain vaccine projects, mainly in the earliest clinical studies. They forecast that in ten yers a vaccine would emerge from this approach, targeted to something in the scale of between the SARS COV-2 vaccines or an incremental influenza virus. This is very vague indeed, and the spectrum is so wide that one cannot understand the appropriateness of the strategy.\n\nShould the Authors want to revise this paper or write a follow up my suggestions are as follows:\na) Firstly, take advantage of the previously mentioned studies and frame their proposal in the context of ongoing policy debate in the EU about pharma R&D b) Secondly, consider that if they are proposing a public- private but not for profit initiative they also need to show why it should be competitive with exisiting organisations in terms of services offered: they may for example consider a benchmark organisation and discuss their assessment c) Finally, for the hub model, the Authors seem to basically adopt a business model that may overlap with the activity of venture capitalists on one side, or even the activity of some public bodies, such as HERA and the EIB, which are also proposing to provide support and seed capital etc in certain R&D investment projects. d) The issue of Intellectual Property Rights arising from the hub activities should be also more discussed.\nI understand that what I am suggesting goes probably beyond the Authors' intentions, but as mentioned they can try and be more ambitious. They certainly have first hand evidence about R&D needs, lack of certain services and of capital, and about the overall panorama to better place their proposals in the context. To be clear, I have nothing against an hybrid model, but as it stands there are two subsequent models in one paper rather than a synergic combination of activities. Perhaps the Authors may deal with this issue by starting from the beginning with a more complete proposal.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? No\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [] } ]	1	https://f1000research.com/articles/12-1401
https://f1000research.com/articles/12-826/v1	13 Jul 23	{ "type": "Review", "title": "Tourette syndrome research highlights from 2022", "authors": [ "Andreas Hartmann", "Per Andrén", "Cyril Atkinson-Clément", "Virginie Czernecki", "Cécile Delorme", "Nanette Marinette Monique Debes", "Kirsten Müller-Vahl", "Peristera Paschou", "Natalia Szejko", "Apostolia Topaloudi", "Keisuke Ueda", "Kevin J. Black", "Per Andrén", "Cyril Atkinson-Clément", "Virginie Czernecki", "Cécile Delorme", "Nanette Marinette Monique Debes", "Kirsten Müller-Vahl", "Peristera Paschou", "Natalia Szejko", "Apostolia Topaloudi", "Keisuke Ueda", "Kevin J. Black" ], "abstract": "This is the ninth yearly article in the Tourette Syndrome Research Highlights series, summarizing selected research reports from 2022 relevant to Tourette syndrome. The authors briefly summarize reports they consider most important or interesting.", "keywords": [ "Tourette", "tics", "annual review" ], "content": "Introduction\n\nThis article is meant to disseminate scientific progress on Gilles de la Tourette Syndrome (TS) that appeared in the year 2022, summarizing research reports the authors judged important or interesting.\n\n\nMethods\n\nWe searched PubMed using the search strategy (“Tic Disorders”[MeSH] OR Tourette NOT Tourette [AU] NOT Tourette [COIS]) AND 2022[PDAT] NOT 1950:2021[PDAT]. On 09 January 2023 this search returned 216 citations, available at this link. Colleagues also recommended articles, and we attended selected medical conferences. We selected material for this review subjectively, guided by our judgment of possible future impact on the field.\n\n\nResults\n\nDefinition and phenomenology\n\nOn blinded video review, the frequency of “extra” (non-goal-directed) movements was more than three times higher in patients with a tic disorder than in people without a tic disorder, vocalizations were greater than 22 times more frequent, and obsessive-compulsive and depressive symptoms were much more common (Bartha et al. 2023). However, the occurrence and frequency of “extra” movements overlapped between those with and without a tic disorder; therefore, the authors concluded that surplus movements per se are not sufficient to diagnose tics. The repetitive character, timing pattern, and associated features of tics such as premonitory urges are necessary too. These results will not surprise tic clinicians but highlight the difficulties in attempting to diagnose and count tics by artificial intelligence analysis of video recordings.\n\nNilles and colleagues presented data from a consecutive series of 200 youth with primary tic disorders at a referral center starting in 2017 (Nilles et al. 2023). Females had more severe motor tics and impairment from tics than did males, and tic severity increased with age.\n\nBazaibal-Carvallo and colleagues published a study about self-injurious behaviors (SIB) in TS (Baizabal-Carvallo et al. 2022). The authors included 201 patients with TS and 34 (16.9%) of them had comorbid SIB. The majority of these patients experienced self-inflicted damage (11.4%), while only 3.5% of participants also experienced aggression towards others and only 2% had what the authors denominated “tic-related SIB”. In this study, the authors compared in detail the distribution of different tics in patients with SIB and without using a univariate model, and concluded that individuals with SIB are more inclined to have tics involving shoulder, trunk, arm, as well as dystonic tics, complex motor tics, copropraxia, complex phonic tics, a higher number of phonic tics, coprolalia and OCD. In multivariate analysis, SIB was found to be associated with complex motor tics, obsessive-compulsive symptoms and greater tic severity. Interestingly enough, patients with SIB were also more likely to have been selected for deep brain stimulation (DBS).\n\nA huge electronic medical records database showed evidence that tic disorders in children are associated with atopic disorders (Hakimi et al. 2022). The risk of atopy was higher in children who had taken a methylphenidate-class drug or an α2 agonist; this result could implicate the medications, or it could reflect higher severity of tics leading to treatment.\n\nAssessment\n\nA nearly universally wished-for advance in TS research would be a reliable automated tic detector. One such was described at the European Society for the Study of Tourette Syndrome (ESSTS) meeting in June 2022 in Lausanne, Switzerland. First, a neural network detected a set of facial landmarks to reduce the dimensionality of the data from a video stream of the person’s face (Loewenstern et al. 2022). A complex processing stream used multiple convolutional neural networks to identify salient spatial features mapping to tics, and a different form of neural network to identify temporal signatures of tics. The algorithm detected over 90% of tics from video captured by a smartphone camera. However, the group had not yet performed out-of-sample generalization testing.\n\nEpidemiology\n\nSeveral studies addressed the prevalence and incidence of TS and chronic tic disorders. Using 2016–2017 National (U.S.) Survey of Children’s Health data, parents reported that 0.3% of their children and adolescents between 6 and 17 years ever had been diagnosed with TS (Charania et al. 2022). A systematic review and meta-analysis showed a global prevalence of TS of 0.5–0.6% including adults and of 0.7% in children and adolescents (Jafari et al. 2022). In Asia, the National Taiwan Insurance Research Database was used to estimate incidence and prevalence of TS and chronic tic disorders (TS/CTD) from 2007–2015 (Chou et al. 2022). The prevalence of TS/CTD increased during this period. Annual incidence rates of TS/CTD increased in childhood and adolescence but decreased in adulthood. In China, the prevalence of tic disorders in school students between 6 and 16 years was reported as 1.37%, lower than almost every other recent study (Yan et al. 2022). TS and OCD were highly comorbid.\n\nA report from the U. S. Centers for Disease Control and Prevention (CDC) attempted to synthesize the varied data on prevalence of persistent (chronic) tic disorders in the US (Tinker et al. 2022). They estimate that 350,000–400,000 children and adults have TS, and about a million more have persistent motor or vocal tic disorder, but they conclude that new data with more accurate methods is required to be more confident in the results.\n\nPrognosis and natural history\n\nRicketts and colleagues evaluated childhood predictors of adult tic severity and impairment and found that childhood severity of tics and female sex are important predictors for tic severity and impairment in adulthood (Ricketts et al. 2022d).\n\nUsing an online survey, 351 adult males with TS completed questionnaires about adverse childhood experiences (ACEs) and a lifetime version of the Yale Global Tic Severity Scale (Yang et al. 2022). An association between ACEs and higher lifetime tic severity and impairment was found. TS genetic risk may moderate the association between ACE score and tic impairment.\n\nOpenneer and colleagues conducted a subanalysis of the European Multicentre Tics in Children Study (EMTICS) to evaluate tic predictors in a prospective study (Openneer et al. 2022). Children with tic onset were more frequently male, had higher baseline severity of conduct problems, autism spectrum disorder (ASD) symptoms, compulsions and emotional problems compared to children without tic onset. Interestingly, there were differences in tic predictors between sexes: while conduct and ASD problems were male-specific predictors, severity of compulsions and oppositional and emotional problems were female-specific predictors.\n\nA very complete review and overview of Tourettic OCD (TOCD) was published by Katz and colleagues (Katz et al. 2022). This phenotype, well-known to all clinicians working with TS patients, consists of touching behavior, symmetry concerns, and thoughts of exactness. The authors feel that the core of the phenomenon is a need to repeat complex tics until “just right,” without cognitive or imaginal obsessions. The authors offer arguments why TOCD might be a condition distinct from both TS and OCD and advocate its inclusion as a separate entity in future editions of the DSM. The authors suggest studies that may allow us to delimit TOCD clearly from TS and OCD (Robins and Guze 1970), though at this time some needed evidence is lacking.\n\nLi and colleagues reported on children with refractory TS (Li et al. 2022). Pediatric refractory TS was characterized by earlier age of onset, longer disease duration, lower IQ, higher prevalence of PU and higher prevalence of psychiatric comorbidities.\n\nSensory phenomena and premonitory urge\n\nEssing and colleagues published an important study regarding the location of premonitory urges (PU) in patients with tics (Essing et al. 2022), updating the conference presentation we reviewed previously (Jakubovski et al. 2018; Rose et al. 2019). This was an online study in 291 adults with tic disorders in which participants self-reported about different characteristics of PU. The authors found that PU were located in the same body part or in direct proximity to the part of the body where tic was located. Most frequently, PU were found in the area of the face and the head (62%). Complex tics were more frequently preceded by PU than simple tics but there were no differences regarding report of PU for motor vs. vocal tics. Usually, PU were localized in the front rather than the back of the body (73% vs. 27%), but there were no differences between the right and left sides.\n\nLangelage and colleagues examined urge-tic association in 25 children and adolescents with TS (Langelage et al. 2022). The authors were interested in urge-tic associations, including inter-individual differences, and correlation with clinical measures, and compared their findings with a sample of adults. At the group level, the authors found positive associations between urges and tics, but this was not confirmed in the individual level data since fewer than half of participants showed a positive association between urge and tic, and two participants had reverse association. Similarly to previous reports, associations between urge levels and tic intensity were less pronounced in children and adolescents than adults with TS.\n\nHe and colleagues tested the contribution of brain γ-aminobutyric acid (GABA) and glutamate levels in the right primary sensorimotor cortex (SM1), supplementary motor area (SMA), and insular cortex (insula) to tic and urge severity in children with TS (He et al. 2022). Overall, they demonstrated involvement of GABAergic neurotransmission within the SMA in the experience of PU in children with TS.\n\nIsaacs and colleagues published two reports on sensory phenomena in adults with chronic tic disorders (CTD). The first examined sensory hypersensitivity (SH) in over 200 adults with CTD, OCD or neither (Isaacs et al. 2022). Both patient groups had much higher self-reported SH than did healthy controls, but tics did not confer additional sensitivity. Across all three groups, SH was associated with OCD and attention deficit hyperactivity disorder (ADHD) symptoms but not with tic severity. The second report investigated interoception in adults with or without CTD using a published self-report measure (Narapareddy et al. 2022). Results indicated that adults with CTD are more aware of their internal body state and have higher self-reported anxious somatization scores, but these results are explained primarily by female sex and OCD symptoms, not tics. They identified a collection of symptoms from the interoception questionnaire that correlated with premonitory urge severity.\n\nComorbidities\n\nAraújo and colleagues examined the role of ADHD and OCD in tic severity during the COVID-19 pandemic in Brazilian and Portuguese patients with TS and found that approximately half of the patients experienced worsening of tic severity (Araújo et al. 2022). They suggest patients with comorbidity might be more susceptible to the effects of the pandemic.\n\nFood difficulties—among others, greater food responsiveness and emotional overeating—were shown to be more common in children with TS than previously reported (Smith et al. 2021).\n\nThe experience of tic-related pain and use of pain management was assessed in an online survey answered by 188 adults with self-reported tics (Taylor et al. 2022). Tic-related pain was shown to have a significant physical and psychological impact and was important to be addressed in the long-term management of tic disorders. A similar study was conducted in Poland (Małek 2022) but this time in a pediatric population. The authors included 40 children with TS and 57 parents of children with TS, as they wanted to collect information about the perspective of children and parents on this topic. For tic assessment the authors used the YGTSS, while pain severity, localization and coping strategies were assessed with the Pediatric Pain Questionnaire and Pediatric Pain Coping Inventory, which were administered both to children and parents. Pain was reported by 60% of children with TS and 72% of parents confirmed that their children sometimes suffered from pain. The most common sites of pain were the cervical region, throat, shoulder, ocular region, and joints. Contrary to expectations, no correlation was found between tic severity and pain. Consistency was observed between the reports of children and their parents in coping with pain.\n\nRicketts and colleagues published an important article about sleep disorders and the use of sleep medication, nighttime tics and pattern of sleep in patients with tics (Ricketts et al. 2022a). In this study, 125 adults with tics completed an internet survey in which they rated sleep history and sleep chronotype as well as the severity of tics and psychiatric comorbidities. The most frequently reported sleep-related disorders in a population of patients with tic disorders were bruxism, insomnia, and tic-related difficulty falling asleep. Sleep problems correlated with impairment, obsessive-compulsive symptoms, and emotional regulation problems. Interestingly enough, eveningness related to tic severity. Therefore, the authors hypothesized that interventions to advance chronotype may help with tic improvement. The same group of authors examined another aspect of sleep disorders in the group of patients with tics (Ricketts et al. 2022c). This study included 114 children with TS, and the authors compared those with sleep disorders (n = 32) to those who have no problems with sleep (n = 82). Children with TS and sleep disorder were from households with lower parental education and at higher risk of poverty. They also were more frequently diagnosed with comorbidities such as OCD, ODD, ADHD and ASD and were more frequently prescribed anti-tic medication. In line with these findings, children with TS and sleep disorder had more severe tics, tic-related impairment and more severe ADHD symptoms.\n\nA case–control study including 271 children with tic disorder and 271 controls revealed that children with tic disorders had increased risks for sleep disturbances as measured with the Children’s Sleep Habits Questionnaire (Mi et al. 2022). Sleep disturbances included among others bedtime resistance, sleep onset delay, sleep anxiety, night waking, and daytime sleepiness. The presence of comorbid ADHD increased the risk for sleep disturbances.\n\nJiménez-Jiménez and colleagues published results of a register-based cohort study to estimate the prevalence of insomnia in Swedish patients with tics (Jiménez-Jiménez et al. 2022). Individuals with tics had a prevalence of insomnia of 32.2% in comparison to 13.7% in the general population, and this difference was statistically significant. Importantly, this association was independent from somatic disorders, familial factors, or psychiatric comorbidities, although familial factors, neurodevelopmental comorbidities, and ADHD/ADHD medication may explain part of the association.\n\nNail biting (onychophagia) is common in unselected children but has also been included in descriptions of complex tics. A report from Taiwan examines prevalence of nail biting in over 2000 children, including 765 with a primary tic disorder, and finds that nail biting is very common in TS (56.6%) and provisional tic disorder (27.4%)—much more common than in controls (15.0%)—and begins prior to onset of definite tics (Hsueh and Chen 2022).\n\nVermillion and colleagues compared responses on the Youth Risk Behavior Surveillance System (YRBS) in youth with TS and healthy controls and did not show any differences between these groups regarding the profile of risky behaviors (Vermilion et al. 2022).\n\nCui and colleagues reported on the emotional and behavioral profile of children with TS in China, using the Child Behavior Checklist (CBCL), and compared this profile with those of sex-matched healthy controls and groups with ADHD, OCD and depression (Cui et al. 2022a). No association was found between the eight factors of the CBCL and motor tics, vocal tics or tic severity assessed by the YGTSS. Nevertheless, there was a positive association between the impairment scale of the YGTSS and thought problems as well as rule-breaking behavior. Contrary to expectation, children with GTS showed a similar profile of CBCL to the children with depression, but not ADHD or OCD.\n\nTessier and colleagues compared the design fluency profile of children with TS with matched healthy controls (Tessier et al. 2022). Children with TS did not show general executive dysfunction in comparison to their peers.\n\nAnother study focused on decision-making processes under both ambiguity and risk contexts with a combination of statistical approaches (Atkinson-Clement et al. 2022a). If they found no impairment in patients with TS, the authors observed that ADHD and OCD are associated with distortion of decision-making. Altogether, they suggested that the risk propensity observed in real-life could be not directly related to TS.\n\nFunctional tic-like behaviors\n\nFremer and colleagues examined 32 patients with functional tic-like behaviors (FTLBs) reflecting those found on social media, using both operationalized psychiatric diagnosis and a psychodynamic focus (Fremer et al. 2022). Symptoms typically started abruptly at the mean age of 19 years and gradually deteriorated. In all patients, current psychological stressors, unconscious intrapsychic conflicts, and/or structural deficits were identified. Nearly all patients (94%) also suffered from other psychiatric symptoms. The authors concluded that pre-existing abnormalities in social behavior and psychiatric symptoms, but also TS in combination with time-related psychological stressors, unconscious intrapsychic conflicts, and structural deficits predispose to development of these symptoms.\n\nTrau and colleagues reviewed charts of 198 patients with tic-like phenomena and propose diagnostic criteria to separate those with typical tics, those with clinically diagnosed functional tics (FT), and those with a past typical tic presentation complicated by a fulminant worsening (Trau et al. 2022). Only the presence of rostrocaudal progression and increased obsessive-compulsive behaviors were significantly different between patients with new-onset FT and those with functional worsening of a previous tic disorder. Results also showed that age at tic onset was not a contributing factor for group differentiation. Many patients with FT were not exposed to videos depicting tics on social media. Similarly, the Calgary group tested which symptoms differed between 41 children or teens with FTLBs and 195 with typical primary tic disorders (Nilles et al. 2022b). The specific symptoms that best discriminated the two groups included copropraxia, coprolalia, popping or whistling noises, simple head movements, and self-injurious actions.\n\nHan and colleagues reported the prevalence and clinical characteristics of children with functional tic-like behaviors detected during the COVID-19 pandemic based on an analysis from a single center (Han et al. 2022). There was a significant increase in the percentage of functional tic-like behaviors in 2020 and 2021. In line with other studies, differences between patients with functional tics included several distinguishing features: predominance of females, later age of onset and higher rates of anxiety and depression. Patients with FTLBs also more frequently demonstrated coprolalia-like behaviors, complex phrases, self-injurious behaviors, higher rates of hospitalizations and school absenteeism.\n\nA group of experts gathered by the Tourette Association of America (Malaty et al. 2022) developed a database from the recent spate of articles on FTLBs, including all the features thought to distinguish FTLBs from typical tic disorders. They summarized these reports and offered recommendations for diagnosing FTLB. The authors found some features to provide better evidence than others for differential diagnosis but emphasize that at this time the diagnosis generally needs to include several characteristics. Another review came from the Calgary group (Amorelli et al. 2022).\n\nArbuckle and colleagues presented new data from the Washington University New Tics study on clinical features of children examined on average four months after tic onset, who would go on to be diagnosed with TS/CTD (Arbuckle et al. 2022). They compare these characteristics to those described in 17 published reports on functional tic-like symptoms (FND-tic), whose initial evaluation also tends to be a few months after symptoms begin. Stark differences in presentation distinguish the FND-tic patients from typical PTD. Symptoms that best distinguished the groups in terms of positive or negative predictive value (PPV or NVP) included movements or vocalizations that were dramatically worse in the presence of others vs. alone, coprophenomena at presentation, symptoms that dramatically and persistently disrupt the person’s intended actions or communications, and “tic attacks.”\n\nHowlett and colleagues investigated the prognosis of functional tic-like behaviors in 20 adolescents and nine adults (Howlett et al. 2022). The authors found in this prospective study that adolescents with FTLBs have a better prognosis than do adults. Treatment of comorbidities with SSRIs and CBT was proposed as the most effective therapeutic approach.\n\nA group from the Great Ormond Street Institute of Child Health in London reported preliminary results of a single, 2.5-hour-long psychoeducational group in children with FTLBs (Pearman et al. 2022). Each child selected his own goal-based outcome (GBO), and 31 reported on their GBO after treatment. Self-reported progress on the GBOs was substantial and statistically significant.\n\nGenetics and epigenetics\n\nLiao and colleagues performed a transcriptome-wide association study (TWAS) using previously published TS GWAS2 data (Yu et al. 2019) (4819 TS cases and 9488 controls) and CommonMind Consortium and brain tissue panels from GTEx 53 v7 (Liao et al. 2022a). The strongest hit was the FLT3 gene (which is involved in hematopoiesis, inflammation and immune function), with increased expression in the dorsolateral prefrontal cortex. It was also associated with additional brain regions (cortex, hippocampus, anterior cingulate cortex, frontal cortex, cerebellum, and cerebellar hemispheres). FLT3 was also supported as the causal gene among the genes in the TWAS region by fine-mapping analysis. Based on a splicing TWAS, the authors identified three additional genes associated with TS (MPHOSPH9, FIP1L1, and CSGALNACT2), which are also involved in immune processes.\n\nGrotzinger and colleagues collected publicly available GWAS data from European populations for 11 psychiatric traits, including TS (Grotzinger et al. 2022). Their aim was to investigate the genetic architecture of those traits at biobehavioral, functional genomic, and molecular genetic levels. First, they performed genetic correlation and genomic structural equation modeling (SEM) to explore how those traits may cluster genetically. TS, anorexia nervosa (AN) and OCD were found to cluster together as a Compulsive Disorders factor, but TS was also part of a cluster primarily made up of ADHD and ASD, but also including major depressive disorder, post-traumatic stress disorder (PTSD), and problematic alcohol use. They performed additional analyses to identify correlations between the identified clusters and other phenotypes including biobehavioral traits, brain morphology, and circadian activity. They also performed cross-disorder meta-analyses of all psychiatric traits, and within each factor to identify shared genetic risk variants. One shared susceptibility locus was identified in the Compulsive Disorders factor (previously associated with AN), and nine loci for the Neurodevelopmental Disorders (ADHD and ASD) factor.\n\nTetsos and colleagues performed the largest Tourette Syndrome GWAS meta-analysis to date with a total of 6,133 TS cases and 13,565 controls, including a novel dataset (Tsetsos et al. 2023). They identified a novel genome-wide significance locus on chromosome 5, close to the NR2F1-AS1 gene. Animal studies have shown that NR2F1-AS1 is critical in neurodevelopment, and in humans, mutations can cause neurodevelopmental disorders or optic atrophy. The implication of this locus was also supported by analyses combining eQTL and Hi-C (high-throughput chromosome conformation capture) data with the GWAS results. Additional analyses exploring the association of TS polygenic risk score with brain volume data revealed statistically significant associations with right and left thalamus volumes and right putamen volume.\n\nJain and colleagues used previous TS GWAS summary statistics (Tsetsos et al. 2023) to calculate the TS Polygenic Risk Score (PRS) on individuals in the UK Biobank and then performed a Phenome Wide Association Study (PheWAS) to assess the association of TS genetic risk with a wide range of phenotypes (n = 2242) (Jain et al. 2023). They observed significant associations with 57 traits including anxiety disorder, depression, type 2 diabetes, heart palpitations, and respiratory conditions. They also performed cross-disorder comparisons of the PheWAS results with ADHD, ASD, and OCD. They observed a pattern in which phenotypes associated with TS had a similar direction of effect with ADHD and ASD, but an opposite direction for OCD in most cases except for mental health factors. Additionally, they performed a sex-specific PheWAS, and observed type 2 diabetes and heart palpitations to be significantly associated with TS risk in males but not in females, while diseases of the respiratory system were associated with TS risk in females but not in males.\n\nRyan and colleagues performed an identity-by-descent (IBD) analysis using Whole Genome Sequencing (WGS) data from 19 individuals from an extended pedigree from a Costa Rican family (17 TS-affected and two controls) (Ryan et al. 2022). Among the implicated TS loci, they observed a rare deleterious missense variation in the RAPGEF1 gene and two ultra-rare deleterious intronic variants in the ERBB4 and IKZF2 genes. All three genes play a role in neurodevelopment.\n\nNi and colleagues performed a two-sample Mendelian randomization analysis, to explore the causal association between the gut microbiome and 15 psychiatric disorders including TS (Ni et al. 2022). For TS they used the PGC GWAS summary statistics (Yu et al. 2019), and the gut microbiome MiBioGen GWAS summary statistics (Kurilshikov et al. 2021). Consistent with previous literature, they observed a negative causal association of TS with class Bacteroidia and its child taxon, order Bacteroidales.\n\nMahjani and colleagues analyzed data from a population-based cohort study in Sweden including 993 individuals with OCD, 217 individuals with chronic tic disorders (CTD), and 91 diagnosed with both OCD and CTD (Mahjani et al. 2022). Their aim was to evaluate the distribution of potentially damaging copy number variation (pdCNV) in OCD and CTD patients. Focusing on CTD, they observed 19 pdCNV events in 18 CTD individuals, and three of them were within known genomic disorders (16p13.11 deletion, 15q25.2 deletion, and 17q12 duplication). The 16p13.11 deletion is a locus associated with multiple neurodevelopmental disorders including anxiety disorders, ASD, epilepsy and learning difficulties. Similarly, 15q25.2 was previously reported in individuals with ASD. They also detected 9 pdCNV events in eight individuals diagnosed with both OCD and CTD.\n\nBae and colleagues analyzed 131 human brains, 19 of which were from donors with TS, to detect somatic mutations through genome-wide sequencing (Bae et al. 2022). When they explored all samples for the presence of somatic mutations on genes related to neuropsychiatric disorders, they did not detect deleterious somatic mutations in TS-related genes. They did however identify a missense mutation in the ARHGEF6 gene. In the TS cohort they also detected two duplications on chromosome 3 in two different brains present in all bulk samples, suggesting that these occurred during development. Genes in duplicated regions included RP11-553D4.2, SPICE1, WDR52 in one sample and FHIT1 and U3 in the other.\n\nEnvironmental risk factors\n\nAbdulkadir and colleagues explored whether pregnancy-related risk factors (cumulative adverse pregnancy risk score, and maternal anxiety, depression, smoking, and alcohol use) together with genetic factors (TS PRS) (Yu et al. 2019) are associated with the presence of tics in the Avon Longitudinal Study of Parents and Children (ALSPAC) cohort (612 cases and 4,201 controls) (Abdulkadir et al. 2022). Initially, they tested whether each single predictor—PRS and all four pregnancy-related risk factors—were individually associated with the presence of tics in cases versus controls; all predictors except maternal alcohol use were significantly associated with tic presence. Then, they tested the joint effect of TS PRS and each (as well as all) of the pregnancy-related risk factors (except maternal alcohol use). They observed that the “all joint” model explained significantly more variance in the tics presence than the model including only the TS PRS.\n\nTransient environmental effects on tics\n\nIverson and Black summarized research on external factors and internal states that transiently worsen or improve tic severity (Iverson and Black 2022).\n\nElectrophysiology\n\nTo better understand the deficit in motor inhibitory control in TS, Indrajeet and colleagues used a recent alternative method (cancellable rise-to-threshold model, CRTT), which distinguishes between proactive (anticipation of a inhibition movement) and reactive (cancellation of a planned unwanted movement) inhibitory control (Indrajeet et al. 2022). They compared 63 patients with TS and 34 HC on an Emotional Stop Signal Task (ESST) and identified robust deficits in TS in reactive control but not in proactive control. The TS group had difficulties in slowing down the speed of movement preparation, which they rectified by their intact ability to postpone the movement.\n\nAn EEG study assessing inhibitory control of frontal lobe regions, which are important for motor inhibition in chronic tic disorders, was conducted using a stop signal task (Zea Vera et al. 2022). Right superior frontal gyrus gamma event-related desynchronization (ERD) was elevated in patients with chronic tic disorders during stop preparation. Elevated right superior frontal gyrus gamma ERD correlated with decreased tic severity, suggesting that right superior frontal gyrus gamma ERD may reflect a mechanism of tic suppression. In the future, it is envisioned that electroencephalography might be useful as a biomarker in TS and in understanding the pathophysiology of tics.\n\nTMS was also discussed as a biomarker for tic disorders, including reduced short-interval intracortical inhibition at rest, which suggests a correlation with motor tic severity, shortened cortical silent period duration, increased intracortical facilitation, and decreased motor evoked-potential amplitude (Jannati et al. 2022). Using EEG as a biomarker for comprehensive behavioral intervention (CBIT), a randomized controlled trial was conducted to determine whether EEG coherence during Go/NoGo tasks correlated with CBIT outcomes (Morand-Beaulieu et al. 2022). No association was found between EEG coherence during the Go/No-Go task and changes in tic severity, suggesting that brain processes in the inhibition of Go/No-Go motor responses do not play any role in CBIT.\n\nNeuroimaging studies\n\nBaldermann and colleagues reported data from 15 TS patients implanted with thalamic DBS. A positive relationship with tic reduction was observed for functional connectivity between the estimated activated tissue by DBS and the sensorimotor cortex, the bilateral insula and the inferior frontal cortex (Baldermann et al. 2022). A negative relationship was found between the activated tissue and the cerebellum, the temporal and orbitofrontal cortex, and the ventral striatum.\n\nThree publications used MRI to identify pathophysiological abnormalities in TS. The first one (Bharti et al. 2022) highlighted that pure TS, as well as TS associated with OCD (both drug-naïve), were underpinned by increased white matter fractional anisotropy in the corticospinal tract, the anterior thalamic radiations, the inferior longitudinal fasciculus and the corpus callosum, and all correlated with tic severity. The second (Liao et al. 2022b) also focused on drug-naïve children with TS, assessed alterations in brain network topology. Some network metrics were abnormal at the global level (i.e., increased global efficiency and decreased path length), and they also showed some nodal changes, especially in the cortico-striato-thalamo-cortical circuit (i.e., SMA, caudate nucleus, thalamus, superior parietal gyrus, posterior cingulate cortex). This study was especially relevant since the authors used brain metrics which are rarely investigated. The third study focused on gyral abnormalities (McCann et al. 2022). They identified that younger patients with TS (children) had increased surface curvature in the frontal cortex (opercular and triangular parts of the inferior frontal cortex), while older patients (adolescents) had increased grey matter volume in the cerebellum, the precentral cortex and the primary motor cortex.\n\nAnother interesting work was published last year, using different statistical models to classify BOLD resting state fMRI of patients with TS and healthy controls (Xin et al. 2022). The model that achieved the highest accuracy (multivariate non-linear model, accuracy 94%, sensitivity 96% and specificity 92%) was especially based on changes observed in the frontal cortex (superior, medial and middle frontal gyrus, SMA, pre- and postcentral cortex), basal ganglia (putamen, thalamus and caudate nucleus) and the cerebellum. In the future, this kind of work could lead to specific tools which might contribute to the diagnosis of TS.\n\nFrom a more cognitive viewpoint, three studies linked specific TS symptoms to brain alterations. The first one investigated the relation between the structural connectivity of several basal ganglia (caudate nucleus, putamen, nucleus accumbens, subthalamic nucleus and the medial subthalamic region) and the cortex with anxiety and impulsivity in TS (Temiz et al. 2022). They found hyperconnectivity in TS patients between the left medial subthalamic region and the insula, entorhinal and temporal cortex. Moreover, the connectivity between the subthalamic nucleus and the left insula was positively related to impulsivity and anxiety scores, measured respectively with the BIS-11 and the STAI. The second study focused on the role of GABA as related to the urge to tic and several measures of cortical inhibition obtained with TMS (Larsh et al. 2022). The authors found that severe urges were negatively correlated to cortical excitability and long-interval cortical inhibition in the primary motor cortex. However, they also found that more severe tics were positively correlated with both of these measures. Last, they found that GABA in the right SMA (cortical excitability and long-interval cortical inhibition) was changed in TS patients compared to healthy controls. Altogether, they concluded that changes in the primary motor cortex were modulated by GABA within the SMA and could reflect compensatory mechanisms. The third study focused on error-related negativity measured by MEG (Metzlaff et al. 2022). This measure is known to reflect processes of performance monitoring, to be increased during error processing and conflicting response, and to be related to the dopaminergic system and prefrontal cortex activity. In a small sample of adult patients without any medication nor comorbidities (n = 8) performing a Go/No-Go task, the authors showed a significant interaction between groups (TS vs. healthy controls) and response (correct vs. error). The authors explained this difference by suggesting that TS patients processed all their responses as erroneous, which means that TS patients had an altered performance monitoring.\n\nA fascinating preliminary report used a clever experimental design to observe regional brain activity (fMRI BOLD) while people with and without TS either chose themselves whether or not to act on Go/No-Go task or followed visually displayed “Go” or “No-Go” instructions in the same situation (Rae et al. 2022). The authors found that activity patterns in the pre-supplementary motor area (preSMA) differed between groups, such that its activity was similar across all task conditions, whereas in control participants it differed by locus of decision-making and by action vs. inhibition. The authors interpret the results as suggesting that brain activity in the preSMA region codes action categories more rigidly, which may explain the sense of effort required to suppress tics voluntarily and the sense of agency that differentiates tics from many other movement disorders.\n\nPsychological interventions\n\nBehavior therapy (BT) is recommended as the first-line intervention for TS/CTD in treatment guidelines published by the American Academy of Neurology (AAN) (Pringsheim et al. 2019) and the European Society for the Study of Tourette Syndrome (ESSTS) (Müller-Vahl et al. 2021). Two types of BT are recommended. The first is Habit Reversal Training (HRT) and its extended package Comprehensive Behavioral Intervention for Tics (CBIT), which has the stronger evidence base. The second recommended BT intervention, Exposure and Response Prevention (ERP) has comparatively less support but is especially popular among clinicians and researchers in Europe (Andrén et al. 2022b).\n\nRemote delivery of BT has become increasingly popular within the last years as a way to make BT more accessible to healthcare seeking individuals with TS/CTD. Several new studies on remote delivery of BT were published in 2022. The largest study by far was conducted in Sweden by Andrén and colleagues (Andrén et al. 2022a), in which 221 young individuals were randomized to therapist-supported, internet-delivered ERP or therapist-supported, internet-delivered psychoeducation (comparator). The results showed that both groups improved on the primary outcome (tic severity as measured by the Yale Global Tic Severity Scale’s Total Tic Score [YGTSS-TTS]) from baseline to the primary endpoint (three months post-treatment). YGTSS-TTS reductions were 6.1 points (27%) in the ERP group and 5.3 points (23%) in the comparator. Contrary to the similar ORBIT study conducted in the UK (Hollis et al. 2021), there was no significant interaction of group and time on the primary outcome. However, treatment response rates at the three-month follow-up were identical in both studies, with significantly more treatment responders in ERP (47%) than in the comparator (29%). The authors concluded that both internet-delivered interventions may be efficacious for young individuals with TS/CTD. Long-term follow-up data will appear in a future publication.\n\nInternet-delivered BT has also recently been evaluated in Israel. Originally published online in 2020, Rachamim and colleagues compared Internet delivered CBIT to a wait list in a randomized controlled trial (RCT) of 41 youth with TS/CTD (Rachamim et al. 2022). The results showed that Internet delivered CBIT was feasible to implement and superior to waitlist. In a new analysis of the same data (Rachamim et al. 2021), the authors focused on 27 treatment completers with comorbid attention deficit hyperactivity disorder (ADHD; n = 16) or comorbid obsessive-compulsive disorder (OCD; n = 11). This new analysis showed that, as in the complete sample, tic severity (YGTSS-TTS) improved in both the ADHD and the OCD groups, although the OCD group improved significantly less than participants without comorbid OCD.\n\nAnother way of delivering BT remotely is through videoconferencing, a format which increased in popularity worldwide during the COVID-19 pandemic. In an Italian RCT conducted by Prato and colleagues, 40 youth with TS were randomized to BT (HRT or ERP) delivered face-to-face at a clinic or remotely via videoconference (Prato et al. 2022). Participants improved on the YGTSS-TTS in both groups, in line with previous studies (Himle et al. 2012; Ricketts et al. 2016). However, contrary to the authors’ claim, this did not indicate that the two interventions were equally efficacious, since the study lacked a pre-defined non-inferiority aim. The videoconference format was also partly used in a naturalistic study of ERP conducted at a TS/CTD specialist clinic in Denmark. In this study by Sørensen and colleagues (Sørensen et al. 2023a, 2023b), 116 youth received ERP (either face-to-face [n = 72] or via videoconference [n = 44]) and were followed up one-year post-treatment. The study showed significant short- and long-term tic severity reductions (YGTSS-TTS) in both groups, with no significant between-group differences. Participants who completed the planned ERP sessions or discontinued early due to a satisfactory tic reduction, improved significantly more than participants who dropped out due to lack of motivation. Firm conclusions are limited by the open design, but the study overall provides support for both face-to-face and videoconference delivery of ERP.\n\nAnother way to make BT more accessible is the group format, where simultaneous treatment of several individuals by one therapist may save therapist resources. Based in the Republic of Korea, Kang and colleagues conducted a non-randomized controlled study (n = 30) comparing group CBIT (n = 18) to a supportive psychotherapy and psychoeducational control condition (n = 12) (Kang et al. 2022). Overall, the baseline TS/CTD severity of the sample was mild. The CBIT group showed modest improvements, with the clearest result being superiority over the comparator in reducing tic-related impairment. The study provided some preliminary support to the feasibility of providing CBIT in a group format in this Korean context. Inoue and colleagues evaluated group CBIT in a case series (n = 3) in Japan (Inoue et al. 2022). In this study, group CBIT was delivered via videoconference software, to further increase accessibility of BT to individuals in the region. The results showed an average tic severity reduction (YGTSS-TTS) of seven points from baseline to post-treatment. Overall, the treatment was concluded to be feasible, acceptable, and promising for further evaluation.\n\nA few studies on face-to-face CBIT were also conducted in 2022. In a US study, Greenberg and colleagues evaluated a modified CBIT intervention, which also included the treatment of comorbid ADHD and psychosocial impairment from both TS/CTD and ADHD (Greenberg et al. 2023). Seventeen young participants with both TS/CTD and ADHD were randomized to modified CBIT (n = 9) or a traditional CBIT comparator (n = 8). The results showed significant improvements in tic severity, tic-related impairment, and ADHD severity for both groups, but the study was likely underpowered to detect between-group differences. Overall, the results indicated feasibility and acceptability for this modified treatment for youth with TS/CTD and ADHD. In a Chinese RCT, Xu and colleagues recruited 37 youth with TS/CTD to compare face-to-face CBIT (n = 12), face-to-face CBIT combined with pharmacotherapy (n = 10), and pharmacotherapy alone (n = 15) (Xu et al. 2022). Tic severity (YGTSS-TTS) improved for all three groups between baseline and post-treatment. Although the approach of comparing BT and pharmacotherapy is relatively novel for the TS/CTD field, the study lacked sufficient power for between-group comparisons.\n\nLastly, interest in investigating the underlying working mechanisms of BT has increased in later years. In a study of 80 adults with TS, Ramsey and colleagues (Ramsey et al. 2022) used structural equation modeling (SEM) to examine the distress provoked by the PU in patients with tics. They concluded that higher levels of premonitory urge intolerance predicted greater levels of tic severity and tic-related impairment. This result highlights a potential clinical implication of targeting the concept of urge intolerance, rather than for instance urge severity, in BT.\n\nIn an RCT including 53 youth, McGuire and colleagues (McGuire et al. 2022) investigated the relationship between several pre-selected cognitive control processes and face-to-face CBIT outcomes. The results showed that only one of the investigated processes—baseline inhibition/switching (as measured by the D-KEFS Color Word Interference Test)—predicted post-treatment tic severity. Interestingly, Gur and colleagues (Gur et al. 2022) studied cognitive inhibition and emotion regulation before and after CBIT group therapy. Fifty-five participants aged 8–15 years with tic disorders were randomly assigned to the CBIT group or the Educational Intervention group and compared on tests of cognitive inhibition and emotion regulation strategies. Their results showed an increase in cognitive reappraisal in the CBIT group only, which was associated with higher intellectual ability. This study raises the possibility that CBIT contributes beyond tic control to cognitive and emotional regulation.\n\nMorand-Beaulieu and colleagues (Morand-Beaulieu et al. 2022) conducted a similar RCT (n = 32), but focused instead on underlying brain mechanisms, particularly EEG coherence. Face-to-face CBIT was superior to the treatment-as-usual comparator, but EEG coherence during a Go/No-Go task was not associated with the tic severity outcome. In a small open study by Eapen and colleagues (n = 17) (Eapen et al. 2022), tic severity significantly improved following face-to-face CBIT. Further, the study showed preliminary support for neurophysiologic changes in cortical inhibition as a potential underlying working mechanism. To summarize, considering that CBIT is a package of numerous behavioral exercises, several underlying mechanisms may be part of explaining how and why the treatment works.\n\nPharmacological studies\n\nA comprehensive and systematic review of pharmacological treatments for patients with TS was provided by Farhat and colleagues (Farhat et al. 2023), confirming that antipsychotic drugs are the most efficacious intervention for tics, followed by α-2 agonists.\n\nAbi-Jaoude and colleagues compared the efficacy and tolerability of single doses of three vaporized medical cannabis products and placebo in reducing tics in adults with GTS (Abi-Jaoude et al. 2022). Each participant received a vaporized single 0.25 g dose of Δ9-tetrahydrocannabinol (THC) 10%, cannabidiol (CBD) 13%, THC/CBD 9%/9%, and placebo at 2-week intervals. There were no statistically significant differences in tic severity for any of the cannabis-based medicine in primary outcome, but THC 10% was significantly better than placebo on the secondary outcome measures.\n\nAn interesting first study randomized 34 children with TS/CTD 1:1 to either a combination of the amino acid L-theanine 200 mg/d and low-dose vitamin B6 2.8 mg/d, or to eight sessions of psychoeducation, for two months (Rizzo et al. 2022). There was no blinding, but the results were interesting: in the medication group, 71% were responders (YGTSS total tic score decreased by at least 30%), compared to 18% of the control group. The authors appropriately note that these results need confirmation in a larger trial with blinded assessment and matching placebo pills.\n\nNeurosurgery\n\nAn elegant study by Ganos and colleagues (Ganos et al. 2022) offers new insights regarding our knowledge of brain networks related to tics and the implications for deep brain stimulation (DBS). They studied 22 patients with secondary tics caused by various types of brain lesions and employed lesion network mapping to identify a common neural network implicated in tic generation. Their methodological approach combined: (i) a comparison of brain lesions which induced tics (n = 22) to control brain lesions which did not induce tics (n = 717); (ii) they built a functional lesion network using healthy subjects’ fMRI (n = 1000) and using the lesion location they found in the first step as seed; and (iii) they assessed the utility of this lesion network to predict tics decrease after thalamic deep brain stimulation surgery on patients with TS (n = 30). They found that despite the very varied brain locations of tic-inducing lesions, regions functionally connected to these lesions mapped to a common network encompassing the insular cortices, the cingulate gyrus, the striatum, the globus pallidus internus (GPi) and the cerebellum. The connectivity of the anterior striatum was significantly more associated with tics compared to lesions inducing other types of movement disorders. They then collected data from 30 patients with TS who had undergone thalamic or pallidal neurostimulation and found that the overlap between the site of neurostimulation and the lesion network map was predictive of tic improvement. Zouki and colleagues presented a preliminary report of a similar analysis, using as seeds both structural abnormalities in TS from neuroimaging studies and reported lesions inducing tics (Zouki et al. 2022). They found that network connections converged on thalamus, striatum, globus pallidus pars externa, and the occipital lobe.\n\nSeveral targets have been used for DBS in TS, and the question of the optimal target is still debated. Dai and colleagues report the outcomes of dorsal (sensorimotor) STN DBS in 10 patients with TS and showed an overall reduction of 62.9% of tics at six months and 58.8% at 12 months, with an improvement in quality-of-life measures and OCD (Dai et al. 2022). Another large retrospective study included 61 patients with refractory TS who were implanted in the posteroventral GPi with an overall improvement of the YGTSS of 58.1% at last follow-up (30 to 130 months) (Cui et al. 2022). As previously reported in other studies, there was an important heterogeneity in clinical response, as five patients were very much improved and five had no effect of DBS. Another case series of seven patients operated in the posteroventral GPi showed a mild effect on tics and OCD but a more substantial effect on depressive symptoms (Liu et al. 2022). Five patients who were operated in the anteromedial GPi and had an overall improvement of 65% of the YGTSS and 75% reduction of the Y-BOCS. Two patients were free of medication post-operatively.\n\nThe programming paradigm of DBS in TS has been seldom studied and programming remains empirical. In previous studies, patients were mostly treated with high frequency stimulation (130 Hz or above). Lower frequency stimulation could prolong the battery life and be better tolerated. Sun and colleagues studied the effect of low frequency (65Hz) stimulation of the GPi and showed a median reduction of the YGTSS of 58.2% at one year, and an improvement of YBOCS of 48.4%, which is in line with reports of DBS at higher frequencies (Sun et al. 2022)\n\nThe outcomes of DBS in GTS patients under 18 years are increasingly documented. Srinivas and colleagues reported the case of a 14-year-old boy with severe tics, self-mutilation and comorbid OCD and ADHD who underwent DBS of the anteromedial GPi. He had substantial improvement of both tics and OCD at six months (Srinivas et al. 2022).\n\nAblative surgery can be an alternative to DBS with the advantage of lower invasivity but with the caveats of being irreversible and less adjustable. Wang et al. report the outcomes of four patients who had radiofrequency thermo-ablative lesions of several targets, eight patients with DBS (GPi in the majority), and one patient with combined DBS and ablation (Wang et al. 2022). They report overall good outcomes in their group of patients who had undergone ablative surgery with a 53.3% reduction in global YGTSS score but did not compare the outcomes of patients treated with DBS to patients with ablative surgery. Notably, two patients who received ablative surgery developed complications such as apathy, urinary incontinence, dysphagia and stereotypic movements.\n\nLin and colleagues published a meta-analysis comparing the efficacy of DBS, repetitive transcranial magnetic stimulation (rTMS) and behavior therapy (Lin et al. 2022). They concluded that DBS is the most effective in reducing tics and rTMS was more efficient to reduce associated OCD. These conclusions have to be tempered by the fact that patients included in the DBS and rTMS studies did not have the same profile: the patients referred to DBS had more severe tics and patients referred to rTMS had more severe psychiatric comorbidities.\n\nOther treatments\n\nFollowing up on their fascinating report of median nerve stimulation (MNS) treatment for tics in TS (Morera Maiquez et al. 2020), Houlgreave and colleagues performed an MEG study showing contralateral hemisphere cortical effects of rhythmic (but not arrhythmic) MNS at 12Hz or 20Hz (Houlgreave et al. 2022). In an open-label study of MNS in 31 adults and older teenagers with TS, participants reported treatment benefit and good tolerability, both in the moment via brief online surveys and at the end of the study (Iverson et al. 22AD; Iverson et al. 2023).\n\nTranscranial magnetic stimulation (TMS) as a treatment modality for tics has been discussed in several recent reviews. Repetitive TMS has been shown to improve tic symptoms and tic comorbidities, and its safety has been confirmed (Bejenaru and Malhi 2022). Other review articles have discussed not only rTMS but also other electrophysiological modalities such as transcranial direct current stimulation (tDCS), peripheral nerve stimulation, and cranial electrotherapeutic stimulation (CES) for the treatment of tics (Frey and Malaty 2022) (Dyke et al. 2022). In contrast to repetitive TMS, tDCS, which works by applying a constant low current to electrodes attached directly to the scalp, is inexpensive, portable, and easy to implement. To date, results have been mixed and inconclusive, as many studies have been open-label designs. Vagus nerve stimulation (VNS) treatment has also been reported to improve tic symptoms, but it is still unknown how VNS affects tic symptoms. CES is a small, portable device that stimulates the brain with a weak electric current and is being studied for its effectiveness in treating tic disorders.\n\nIn a large Swedish cohort study with more than 13 million individuals and almost 7,800 individuals with TS or CTD, it was found that persons with TS or CTD did have an increased risk of experiencing any violent assault or violent and nonviolent crime convictions (Mataix-Cols et al. 2022). The presence of comorbid ADHD and substance use disorders increased the risk. Similarly, more than half of the children with a TS diagnosis included from than 2016–2017 National Survey of Children’s Health data on children from 6 to 17 years, had experienced bullying victimization, and around 20% had perpetrated bullying; both are substantially more frequent than in children without TS (Charania et al. 2022). Martino and colleagues thoughtfully discuss some of the difficulties that arise in interpreting these reports (Martino et al. 2022).\n\nThe Tourette Association of America (TAA) conducted a web-based anonymous survey of adults with TS (n = 601) and parents of children with TS (n = 593) (Tourette Association of America 2022). The sample was not ascertained on a population basis but is likely to represent reasonably well patients at referral centers or engaged in the American TS community. Many of the findings were rather dramatic: for instance, 23% of children and 48% of adults reported that they had considered suicide at some point in their lives, and 10% of children had attempted suicide in the past year. Over two thirds reported they had been discriminated against because of tics. Parents reported substantial impact: 15% of them lost their job or had to reduce their work hours to care for their child with tics, and 5% had to move due to financial stressors from managing tics. On the positive side, half of children were diagnosed within a year of symptom onset (much earlier than in previous studies), and three-fourths of children had a legally binding individualized education plan.\n\nA longitudinal clinical cohort study showed that academic achievement was decreased in children with TS and the severity of comorbidities was shown to have greater influence on education than did tic severity (Lund et al. 2022).\n\nAtkinson-Clement and colleagues examined the consequences of TS on adolescents’ daily living by using a text mining approach (Atkinson-Clement et al. 2022a). Social stigma was shown to be the most common issue faced by patients with TS. Severity of tics had an especially an impact on daily life at school while comorbidities mostly were related to social daily living and risk of depression.\n\nA survey by a pharmaceutical company investigated the educational needs of neurologists, psychiatrists, and caregivers (Stacy et al. 2022). Among other findings, neurologists and psychiatrists differed in their approach to diagnosis and treatment, and parents and physicians differed on the locus of treatment decision-making and on the approach to treating tics that are only “slightly bothersome.”\n\nMealtimes and feeding are known to be problematic in children with TS, both in and out of the home. Using a semi-structured interview, the full scope of difficulties was captured, and potential solutions offered (Bamigbade et al. 2022).\n\nStofleth and Parks conducted semi-structured interviews in 18 adults with TS (Stofleth and Parks 2022). The aim was to assess how others respond to TS behaviors and how these are misinterpreted in interpersonal interactions, using thematic analysis. Unsurprisingly, all participants received unwanted attention regarding their tics, which could be subdivided into six subthemes. Also, three types of misunderstanding in interpersonal interactions were identified. This article is a rewarding read with many concrete examples.\n\nLee and colleagues explored the role of self-esteem in the relationship between psychosocial stress and social adjustment among adolescents with TS (Lee et al. 2022). The authors found that self-esteem fully mediates the relationship between their psychosocial stress and social adjustment, while comorbidities moderate the relationship between self-esteem and social adjustment.\n\nRicketts and colleagues described impairment in academic, interpersonal, recreational, and family financial or occupational domains across children in three groups: with GTS, ADHD and both disorders. Children with ADHD (with or without TS) experienced a greater degree of impairment in overall school performance, writing, and mathematics, relative to children with TS alone. More children with TS and ADHD had problematic handwriting relative to children with ADHD alone. More children with TS and ADHD had problematic interpersonal relationships relative to those with ADHD alone. Children with TS and ADHD had higher mean impairment across domains than children with either TS or ADHD (Ricketts et al. 2022b).\n\nSoós and colleagues investigated the online support available for tic disorders by an inductive thematic analysis of posts and comments. They suggest that online support communities might be valuable in sharing and gaining information on tics from other patients (Soós et al. 2022).\n\nHall and colleagues examined the impact of the COVID-19 pandemic on individuals with tics. They compared YGTSS pre- and post-pandemic in 112 children and adolescents with tics. There were no significant differences in tic symptom or severity between participants who were assessed before and during COVID-19 (Hall et al. 2022).\n\nSimilarly, Termine and colleagues investigated the burden of the COVID-19 pandemic and lockdown on individuals with tics. The authors included 49 children with tics and 245 matched controls who were asked to provide information about lockdown-related changes to daily activities. More than half of patients reported perceived changes in tic severity, restlessness and irritability during the pandemic (Termine et al. 2022).\n\nIn 2022, the International Review of Movement Disorders book series published a large work on TS, divided into two volumes (Lavoie and Cavanna 2022a, 2022b). The book includes up-to-date contributions from a Who’s Who of TS research, from history to neurobiology.\n\nSimilarly comprehensive in scope, the second edition of Tourette Syndrome also appeared during 2022, now edited by Davide Martino in addition to James Leckman (Martino and Leckman 2022).\n\n\nConclusions\n\nThis year, FTLBs were again a focus of research and concern for clinicians. It is hoped that this wave of functional patients will eventually abate as the COVID-19 pandemic recedes, but this remains to be seen: alternative sources of anxiety abound in today’s world and the sometimes toxic influence of social media will likely persist if not increase. On a positive note, studies related to behavioral therapies in TS/CTD are plentiful and of excellent quality, which is cause for hope in the treatment of tics and comorbidities. In contrast, no significant pharmacological developments were reported in 2022, but hopefully this is the quiet before a productive storm. Finally, the entry of non-invasive brain stimulation into the clinic looms large and is encouraging, either as a stand-alone treatment or combined with behavioral and pharmacotherapy.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nReferences\n\nAbdulkadir M, Tischfield JA, Heiman GA, et al.: Polygenic and environmental determinants of tics in the Avon Longitudinal Study of Parents and Children. Am. J. Med. Genet. B Neuropsychiatr. Genet. December 2022. Publisher Full Text\n\nAbi-Jaoude E, Bhikram T, Parveen F, et al.: A Double-Blind, Randomized, Controlled Crossover Trial of Cannabis in Adults with Tourette Syndrome. Cannabis. Cannabinoid. Res. 2022. PubMed Abstract \| Publisher Full Text\n\nAmorelli G, Martino D, Pringsheim T: Rapid Onset Functional Tic-Like Disorder Outbreak: A Challenging Differential Diagnosis in the COVID-19 Pandemic. J. Can. Acad. Child Adolesc. Psychiatry. 2022; 31: 144–151. 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Publisher Full Text\n\nLi Y, Yan JJ, Cui YH: Clinical characteristics of pediatric patients with treatment-refractory Tourette syndrome: An evidence-based survey in a Chinese population. World J. Psychiatry. 2022; 12: 958–969. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLiao C, Vuokila V, Catoire H, et al.: Transcriptome-wide association study reveals increased neuronal FLT3 expression is associated with Tourette’s syndrome. Commun. Biol. March 2022a; 5(1). Publisher Full Text\n\nLiao Y, Li X, Jia F, et al.: The Alternation of Gray Matter Morphological Topology in Drug-Naïve Tourette’s Syndrome in Children. Front. Aging Neurosci. May 2022b; 14: 14. Publisher Full Text\n\nLin X, Lin F, Chen H, et al.: Comparison of efficacy of deep brain stimulation repeat transcranial magnetic stimulation, and behavioral therapy in Tourette syndrome: A systematic review and Bayesian Network Meta-Analysis. Heliyon. October 2022; 8(10): e10952. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLiu A, Jiao Y, Zhang S, et al.: Improved depressive symptoms in patients with refractory Gilles de la Tourette syndrome after deep brain stimulation of posteroventral globus pallidus interna. Brain Behav. May 2022; 12(7): e2635. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLoewenstern Y, Benaroya-Milshtein N, Bar-Gad I: Objective quantification of tic expression in Tourette syndrome patients. Lausanne, Switzerland: 2022. Reference Source\n\nLund J, Borch-Johnsen L, Groth C, et al.: Impact of Tourette Syndrome on Education. Neuropediatrics. 2022; 54: 107–112. Publisher Full Text\n\nMahjani B, Birnbaum R, Grice AB, et al.: Phenotypic Impact of Rare Potentially Damaging Copy Number Variation in Obsessive-Compulsive Disorder and Chronic Tic Disorders. Genes. October 2022; 13(10): 1796. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMalaty IA, Anderson S, Bennett SM, et al.: Diagnosis and Management of Functional Tic-Like Phenomena. J. Clin. Med. 2022; 11. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMartino D, Leckman JF: Tourette Syndrome. New York: Oxford University Press; 2022.\n\nMartino D, Leckman JF, Okun MS: Why Some Individuals With Tourette Syndrome Experience Assault and Perpetrate Criminal Behavior. JAMA Neurol. 2022; 79: 442–444. PubMed Abstract \| Publisher Full Text\n\nMataix-Cols D, Virtanen S, Sidorchuk A, et al.: Association of Tourette Syndrome and Chronic Tic Disorder With Violent Assault and Criminal Convictions. JAMA Neurol. 2022; 79: 459–467. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMałek A: Pain in Tourette Syndrome-Children’s and Parents’ Perspectives. J. Clin. Med. January 2022; 11(2): 460. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMcCann B, Lam MY, Shiohama T, et al.: Magnetic resonance imaging demonstrates gyral abnormalities in Tourette syndrome. Int. J. Dev. Neurosci. July 2022; 82(6): 539–547. PubMed Abstract \| Publisher Full Text\n\nMcGuire JF, Sturm A, Ricketts EJ, et al.: Cognitive control processes in behavior therapy for youth with Tourette’s disorder. J. Child Psychol. Psychiatry. 2022; 63: 296–304. PubMed Abstract \| Publisher Full Text\n\nMetzlaff J, Finis J, Münchau A, et al.: Altered performance monitoring in Tourette Syndrome: an MEG investigation. Sci. Rep. May 2022; 12(1): 8300. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMi Y, Zhao R, Sun X, et al.: Sleep disturbances and sleep patterns in children with tic disorder: A case-control study. Front. Pediatr. 2022; 10: 911343. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMorand-Beaulieu S, Crowley MJ, Grantz H, et al.: Evaluation of EEG biomarkers of Comprehensive Behavioral Intervention for Tics in children with Tourette syndrome. Clin. Neurophysiol. 2022; 142: 75–85. PubMed Abstract \| Publisher Full Text\n\nMorera Maiquez B, Sigurdsson HP, Dyke K, et al.: Entraining Movement-Related Brain Oscillations to Suppress Tics in Tourette Syndrome. Curr. Biol. 2020; 30: 2334–2342.e3. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMüller-Vahl KR, Szejko N, Verdellen C, et al.: European clinical guidelines for Tourette syndrome and other tic disorders: summary statement. Eur. Child Adolesc. Psychiatry. July 2021; 31(3): 377–382. Publisher Full Text\n\nNarapareddy A, Eckland MR, Riordan HR, et al.: Altered Interoceptive Sensibility in Adults With Chronic Tic Disorder. Front. Psych. 2022; 13: 914897. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNi J-J, Xu Q, Yan S-S, et al.: Gut Microbiota and Psychiatric Disorders: A Two-Sample Mendelian Randomization Study. Front. Microbiol. February 2022; 12. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNilles C, Fletcher J, Martino D, et al.: An exploration of tic phenomenology in children with primary tic disorders [abstract]. Mov. Disord. 2022a; 37(Suppl. 2): S413.\n\nNilles C, Martino D, Berg L, et al.: Can the Phenomenology of Functional Tic-Like Behaviors Be Sufficient to Make the Diagnosis? [abstract]. J. Am. Acad. Child Adolesc. Psychiatry. October 2022b; 61(10): S271–S272. Publisher Full Text\n\nNilles C, Martino D, Fletcher J, et al.: Have We Forgotten What Tics Are? A Re-Exploration of Tic Phenomenology in Youth with Primary Tics. Mov. Disord. 2023; 10: 764–773. in press. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOpenneer TJC, Huyser C, Martino D, et al.: Clinical precursors of tics: an EMTICS study. J. Child Psychol. Psychiatry. 2022; 63: 305–314. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPearman Z, Evans S, Heyman I, et al.: Evaluation of a Psychoeducation Group for Children presenting with Functional Tics. Lausanne, Switzerland: 2022. Reference Source\n\nPrato A, Maugeri N, Chiarotti F, et al.: A Randomized Controlled Trial Comparing Videoconference vs. Face-to-Face Delivery of Behavior Therapy for Youths With Tourette Syndrome in the Time of COVID-19. Front. Psych. 2022; 13: 862422. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPringsheim T, Okun MS, Müller-Vahl K, et al.: Practice guideline recommendations summary: Treatment of tics in people with Tourette syndrome and chronic tic disorders. Neurology. May 2019; 92(19): 896–906. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRachamim L, Mualem-Taylor H, Rachamim O, et al.: Acute and Long-Term Effects of an Internet-Based, Self-Help Comprehensive Behavioral Intervention for Children and Teens with Tic Disorders with Comorbid Attention Deficit Hyperactivity Disorder, or Obsessive Compulsive Disorder: A Reanalysis of Data from a Randomized Controlled Trial. J. Clin. Med. 2021; 11(1): 45.\n\nRachamim L, Zimmerman-Brenner S, Rachamim O, et al.: Internet-based guided self-help comprehensive behavioral intervention for tics (ICBIT) for youth with tic disorders: a feasibility and effectiveness study with 6 month-follow-up. Eur. Child Adolesc. Psychiatry. 2022; 31: 275–287. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRae C, Raykov P, Ambridge E, et al.: Elevated representational similarity of voluntary action and inhibition in Tourette syndrome. PsyArXiv Preprints. December 2022. Publisher Full Text\n\nRamsey KA, De NAS, Espil FM, et al.: Urge intolerance predicts tic severity and impairment among adults with Tourette syndrome and chronic tic disorders. Front. Psych. 2022; 13: 929413. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRicketts EJ, Goetz AR, Capriotti MR, et al.: A randomized waitlist-controlled pilot trial of voice over Internet protocol-delivered behavior therapy for youth with chronic tic disorders. J. Telemed. Telecare. 2016; 22: 153–162. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRicketts EJ, Montalbano GE, Burgess HJ, et al.: Sleep and chronotype in adults with persistent tic disorders. J. Clin. Psychol. 2022a; 78: 1516–1539. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRicketts EJ, Wolicki SB, Danielson ML, et al.: Academic, Interpersonal, Recreational, and Family Impairment in Children with Tourette Syndrome and Attention-Deficit/Hyperactivity Disorder. Child Psychiatry Hum. Dev. 2022b; 53: 3–15. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRicketts EJ, Wolicki SB, Holbrook JR, et al.: Clinical Characteristics of Children With Tourette Syndrome With and Without Sleep Disorder. Pediatr. Neurol. 2022c; 141: 18–24. Publisher Full Text\n\nRicketts EJ, Woods DW, Espil FM, et al.: Childhood Predictors of Long-Term Tic Severity and Tic Impairment in Tourette’s Disorder. Behav. Ther. 2022d; 53: 1250–1264. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRizzo R, Prato A, Scerbo M, et al.: Use of Nutritional Supplements Based on L-Theanine and Vitamin B6 in Children with Tourette Syndrome, with Anxiety Disorders: A Pilot Study.Nutrients.2022 Feb18;14(4): 852. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRobins E, Guze SB: Establishment of diagnostic validity in psychiatric illness: its application to schizophrenia. Am. J. Psychiatry. 1970; 126: 983–987. Publisher Full Text\n\nRose O, Hartmann A, Worbe Y, et al.: Tourette syndrome research highlights from 2018. F1000Res. July 2019; 8: 988. PubMed Abstract \| Publisher Full Text \| , Publisher Full Text \| Free Full Text\n\nRyan N, Ormond C, Chang Y-C, et al.: Identity-by-descent analysis of a large Tourette’s syndrome pedigree from Costa Rica implicates genes involved in neuronal development and signal transduction. Mol. Psychiatry. October 2022; 27(12): 5020–5027. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSmith BL, Gutierrez R, Ludlow AK: A comparison of food avoidant behaviours and sensory sensitivity in adults with and without Tourette syndrome. Appetite. 2021; 105713.\n\nSørensen CB, Lange T, Jensen SN, et al.: Exposure and Response Prevention: Evaluation of Tic Severity Over Time for Children and Adolescents with Tourette Syndrome and Chronic Tic Disorders. Neuropediatrics. 2023a; 54: 089–098. Publisher Full Text\n\nSørensen CB, Lange T, Jensen SN, et al.: Exposure and Response Prevention for Children and Adolescents with Tourette Syndrome Delivered via Web-Based Videoconference versus Face-to-Face Method. Neuropediatrics. 2023b; 54(2): 099–106. Publisher Full Text\n\nSoós MJ, Coulson NS, Davies EB: Exploring Social Support in an Online Support Community for Tourette Syndrome and Tic Disorders: Analysis of Postings. J. Med. Internet Res. 2022; 24: e34403. Publisher Full Text\n\nSrinivas D, Manohar H, Sharma E, et al.: Deep Brain Stimulation of the bilateral anteromedial Globus Pallidus internus in an adolescent with refractory tourette syndrome and comorbid obsessive compulsive disorder A case report. Brain Stimul. November 2022; 15(6): 1415–1417. Publisher Full Text\n\nStacy S, Salinas GD, Belcher E, et al.: Assessing the educational needs of physicians in the management of patients with Tourette syndrome: results of a United States survey on practicing clinicians and caregivers. CNS Spectr. 2022; 28: 343–350. Publisher Full Text\n\nStofleth D, Parks ES: Sorry, I Didn’t Mean to Kiss at You: A Descriptive Analysis of Tourette Syndrome in Interpersonal Interactions. J. Dev. Phys. Disabil. 2022; 1–23. Publisher Full Text\n\nSun F, Zhang X, Dong S, et al.: Effectiveness of Low-Frequency Pallidal Deep Brain Stimulation at 65 Hz in Tourette Syndrome. Neuromodulation: Technology at the Neural Interface. February 2022; 25(2): 286–295. Publisher Full Text\n\nTaylor E, Anderson S, Davies EB: I’m in pain and I want help: An online survey investigating the experiences of tic-related pain and use of pain management techniques in people with tics and tic disorders. Front. Psych. 2022; 13: 914044. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTemiz G, Atkinson-Clement C, Lau B, et al.: Structural hyperconnectivity of the subthalamic area with limbic cortices underpins anxiety and impulsivity in Tourette syndrome. Cereb. Cortex. October 2022; 33: 5181–5191. Publisher Full Text\n\nTermine C, Galli V, Dui LG, et al.: Self-reported impact of the COVID-19 pandemic and lockdown on young patients with tic disorders: findings from a case-control study. Neurol. Sci. 2022; 43: 3497–3501. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTessier M, Desmarais A, Leclerc JB, et al.: Capturing Subtle Neurocognitive Differences in Children with and without Tourette Syndrome through a Fine-Grained Analysis of Design Fluency Profiles. J. Clin. Med. 2022; 11: 11. Publisher Full Text\n\nTinker SC, Bitsko RH, Danielson ML, et al.: Estimating the number of people with Tourette syndrome and persistent tic disorder in the United States. Psychiatry Res. 2022; 314: 114684. PubMed Abstract \| Publisher Full Text\n\nTourette Association of America: TAA Impact Survey. 2022.2022. Reference Source\n\nTrau SP, Quehl L, Tsujimoto THM, et al.: Creating a Patient-Based Diagnostic Checklist for Functional Tics During the COVID-19 Pandemic. Neurol. Clin. Pract. 2022; 12: 365–376. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTsetsos F, Topaloudi A, Jain P, et al.: Genome-wide Association Study points to novel locus for Gilles de la Tourette Syndrome. Biol. Psychiatry. February 2023. PubMed Abstract \| Publisher Full Text\n\nVermilion J, Augustine EF, Adams HR, et al.: Risk Behaviors in Youth With and Without Tourette Syndrome. Pediatr. Neurol. 2022; 126: 20–25. Publisher Full Text\n\nWang X, Qu L, Ge S, et al.: Stereotactic Surgery for Treating Intractable Tourette Syndrome: A Single-Center Pilot Study. Brain Sci. June 2022; 12(7): 838. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nXin X, Feng Y, Zang Y, et al.: Multivariate Classification of Brain Blood-Oxygen Signal Complexity for the Diagnosis of Children with Tourette Syndrome. Mol. Neurobiol. January 2022; 59(2): 1249–1261. Publisher Full Text\n\nXu W, Ding Q, Zhao Y, et al.: A preliminary study of comprehensive behavioral intervention for tics in Chinese children with chronic tic disorder or Tourette syndrome. Front. Psych. 2022; 13: 997174. Publisher Full Text\n\nYan J, Deng H, Wang Y, et al.: The Prevalence and Comorbidity of Tic Disorders and Obsessive-Compulsive Disorder in Chinese School Students Aged 6-16: A National Survey. Brain Sci. 2022; 12: 12. Publisher Full Text\n\nYang K, Essa A, Noriega D, et al.: Relationship between adverse childhood experiences and symptom severity in adult men with Tourette Syndrome. J. Psychiatr. Res. 2022; 155: 252–259. Publisher Full Text\n\nYu D, Sul JH, Tsetsos F, et al.: Interrogating the Genetic Determinants of Tourette’s Syndrome and Other Tic Disorders Through Genome-Wide Association Studies. Am. J. Psychiatr. March 2019; 176(3): 217–227. Publisher Full Text\n\nZea Vera A, Pedapati EV, Larsh TR, et al.: EEG Correlates of Active Stopping and Preparation for Stopping in Chronic Tic Disorder. Brain Sci. 2022; 12: 12. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZouki J-J, Ellis EG, Morrison-Ham J, et al.: Network localization of tics: Evidence from coordinate and lesion network mapping. Programme, 14th European Conference on Tourette Syndrome and Tic Disorders. 2022. Reference Source" }	[ { "id": "186879", "date": "18 Jul 2023", "name": "James F. Leckman", "expertise": [ "Reviewer Expertise I am actively involved the care of individuals wit TS", "and with my colleagues at Yale we are conducting a number of research studies." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis team of scholars and practitioners have once again provided a comprehensive summary of the scientific articles published in 2022 regarding Tourette syndrome (TS) and related disorders.\n\nThis article will be a valuable resource for clinicians and researchers interested in this topic. For example, I am in the midst of preparing a chapter on Tic Disorders for the 7th edition of the Rutter Textbook on Child and Adolescent Psychiatry, and this article has introduced me to a few recent scientific findings that I was unaware of.\nI was a bit disappointed that there was no mention of the use of Traditional Chinese Herbal Medicines that are currently being used to treat TS. Given that there is strong evidence supporting use of the 5-Lin granule (TSupport) in treating tic disorders, my colleagues and I are about to start a multisite clinical trial in the US. However, since there were no relevant articles published on this topic in 2022, it is reasonable that this emerging area of research was not mentioned in the review.\n\nIs the topic of the review discussed comprehensively in the context of the current literature? Yes\n\nAre all factual statements correct and adequately supported by citations? Yes\n\nIs the review written in accessible language? Yes\n\nAre the conclusions drawn appropriate in the context of the current research literature? Yes", "responses": [ { "c_id": "10344", "date": "16 Nov 2023", "name": "Andreas Hartmann", "role": "Author Response", "response": "We thank Dr Leckman for his positive evaluation of our annual review." } ] }, { "id": "186880", "date": "06 Sep 2023", "name": "Erica L. Greenberg", "expertise": [ "Reviewer Expertise Tourette syndrome", "OCD", "child and adolescent psychiatry" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThank you for the opportunity to review this important manuscript that summarizes key additions in the literature to the field of Tourette over the last year. The review is extensive, thorough, and covers a multitude of areas pertinent to TS, including pharmacology, genetics, behavioral treatments, epidemiology, etc.\n\nIt is very well organized, and I recommend acceptance with minor changes.\n\nMost of the changes are regarding small changes in the results (see below), and a handful of changes to the Conclusion.\nResults: for the paragraph starting...\n\"Bazaibal-Carvallo and colleagues...\" - it would help to describe what \"self-inflicted damage\" is - and how that differs from SIB\n\n\"A huge electronic...\" Is it appropriate to say the \"risk of atopy was higher in children\" ? I imagine it would be more appropriate (unless methodology was described further) to say \"rates of atopy were higher...\"\n\n\"Rickets and colleagues\" - it would help to comment on what the authors mean by \"important predictors\" - significance? magnitude? what p-value if that's the correct statistic?\n\n\"Li and colleagues\" - looks like PU isn't defined until the below paragraph - would write out premonitory urges in this one\n\n\"Langelage and colleagues\" - it's hard to follow what the author means by \"positive association between urge and tic\" and \"reverse association\"\n\nIn the comorbidities section, given that there are so many articles focusing on sleep, I would make sleep its own category\n\n\"Ricketts and colleagues published...\" - Would recommend defining chronotype\n\n\"Another study focused on decision-making processes\" - I had trouble following this summary, and would recommend re-writing/re-phrasing for clarity purposes\n\n\"Fremer and colleagues\" - was confused what the authors were referring to with \"structural deficits\" and \"unconscious intrapsychic conflicts\" for that matter\n\n\"Trau and colleagues\" - Starting \"Similarly, the Calgary group tested which symptoms\" through \"self-injurious actions\" would benefit from being moved to the below paragraph as those findings are more consistent with teh subsequent paragraph.\n\n\"Grotzinger and colleagues...\" - Difficulty understanding this summary, would re-write/edit, especially from \"TS, anorexia nervosa\" through \"alcohol use.\" Seems like the last sentence of the paragraph contradicts an earlier sentence in the paragraph\n\n\"Abdulkadir and colleagues\" - Clarify what PRS means\n\n\"A fascinating preliminary...\" - Should the authors say \"in TS\" following the clause \"across all task conditions\" ?\n\n\"Remote delivery of BT\" - starting with \"However, treatment response rates...\" - paragraph became hard to follow\n\n\"A few studies on face-to-face\" would change \"conducted\" to \"published\" since the studies weren't conducted in 2022.\n\n\"Lastly, interest in investigating\" ... would change \"later\" to \"recent;\" would make use of PU vs premonitory urges consistent; would clarify \"instance urge severity\"\n\n\"Abi-Jaoude and colleagues\" - would help to list some secondary outcome measures\n\n\"Lin and colleagues\" - unclear how the behavioral therapy performed in comparison\n\n\"The Tourette Association of America\" - would make sure to say \"reported\" when describing some of the self-reported statistics\n\n\"A longitudinal clinical cohort study...\" - would edit for grammar\n\n\"Atkinson-Clement and colleagues\" - grammar bingeing \"Severity of tics...\"\nConclusions:\nIt actually seemed like the focus on FTLBs this year was a lot less than the prior year\n\nWould change wording of \"this wave of functional patients will eventually abate\" as is sounds more negatively judgmental than I believe the authors intended, and additionally, really wasn't a large focus on the summaries this year. Similarly, I would strongly recommend eliminating the sentence referring to \"toxic influence of social media\" as that was not a focus of many/any articles that were reviewed, and it sounds unduly biased against social media when the literature is less clear.\n\nFinal thought is that I would recommend considering a more standardized approach to capturing the array of articles that are published each year, as it is likely that many fall through the cracks for a variety of reasons.\n\nIs the topic of the review discussed comprehensively in the context of the current literature? Yes\n\nAre all factual statements correct and adequately supported by citations? Yes\n\nIs the review written in accessible language? Yes\n\nAre the conclusions drawn appropriate in the context of the current research literature? Yes", "responses": [ { "c_id": "10343", "date": "16 Nov 2023", "name": "Andreas Hartmann", "role": "Author Response", "response": "We thank Dr Greenberg for her thorough review of our yearly review. Below the answers to her queries. Results: for the paragraph starting... \"Bazaibal-Carvallo and colleagues...\" - it would help to describe what \"self-inflicted damage\" is - and how that differs from SIB Actually, in the paper, these were described as “self-directed SIB” and we have changed the phrasing accordingly. We have also added a half sentence regarding tic-related SIB, hoping this will clarify. \"A huge electronic...\" Is it appropriate to say the \"risk of atopy was higher in children\" ? I imagine it would be more appropriate (unless methodology was described further) to say \"rates of atopy were higher...\" We agree and have changed the phrasing accordingly. \"Rickets and colleagues\" - it would help to comment on what the authors mean by \"important predictors\" - significance? magnitude? what p-value if that's the correct statistic? The p value for female sex was < 0.002 and we have added this number to the manuscript. \"Li and colleagues\" - looks like PU isn't defined until the below paragraph - would write out premonitory urges in this one Agreed and done. \"Langelage and colleagues\" - it's hard to follow what the author means by \"positive association between urge and tic\" and \"reverse association\" Reverse association means negative association. We have added this in brackets. In the comorbidities section, given that there are so many articles focusing on sleep, I would make sleep its own category We’d rather stick to this format for historical reasons. \"Ricketts and colleagues published...\" - Would recommend defining chronotype Agreed and done. \"Another study focused on decision-making processes\" - I had trouble following this summary, and would recommend re-writing/re-phrasing for clarity purposes Agreed and done. \"Fremer and colleagues\" - was confused what the authors were referring to with \"structural deficits\" and \"unconscious intrapsychic conflicts\" for that matter Agreed and done. \"Trau and colleagues\" - Starting \"Similarly, the Calgary group tested which symptoms\" through \"self-injurious actions\" would benefit from being moved to the below paragraph as those findings are more consistent with teh subsequent paragraph. Agreed and done. \"Grotzinger and colleagues...\" - Difficulty understanding this summary, would re-write/edit, especially from \"TS, anorexia nervosa\" through \"alcohol use.\" Seems like the last sentence of the paragraph contradicts an earlier sentence in the paragraph Agreed and done. \"Abdulkadir and colleagues\" - Clarify what PRS meansPRS Polygenic risk score, done. \"A fascinating preliminary...\" - Should the authors say \"in TS\" following the clause \"across all task conditions\" ? Yes, absolutely. \"Remote delivery of BT\" - starting with \"However, treatment response rates...\" - paragraph became hard to follow Agreed and done. \"A few studies on face-to-face\" would change \"conducted\" to \"published\" since the studies weren't conducted in 2022. Agreed and changed. \"Lastly, interest in investigating\" ... would change \"later\" to \"recent;\" would make use of PU vs premonitory urges consistent; would clarify \"instance urge severity\" Agreed and changed. \"Abi-Jaoude and colleagues\" - would help to list some secondary outcome measures Agreed and changed. \"Lin and colleagues\" - unclear how the behavioral therapy performed in comparison Agreed and done. \"The Tourette Association of America\" - would make sure to say \"reported\" when describing some of the self-reported statistics Done. \"A longitudinal clinical cohort study...\" - would edit for grammar Done. \"Atkinson-Clement and colleagues\" - grammar bingeing \"Severity of tics...\" Done. Conclusions: It actually seemed like the focus on FTLBs this year was a lot less than the prior year Yes indeed. Would change wording of \"this wave of functional patients will eventually abate\" as is sounds more negatively judgmental than I believe the authors intended, and additionally, really wasn't a large focus on the summaries this year. Similarly, I would strongly recommend eliminating the sentence referring to \"toxic influence of social media\" as that was not a focus of many/any articles that were reviewed, and it sounds unduly biased against social media when the literature is less clear. Here, we respectfully disagree. There is no negative judgement in the hope that the wave of FTMB will abate, to us it is simply a matter of fact statement. Any disorder which prevalence decreases is a cause for rejoicing. As to the toxicity of social media, we made sure to add the adverb “sometimes”. Also, we think the data on the impact of social media on emotional well being, especially in children and adolescents, are actually becoming clearer, cf. Sinclair-McBride K, Rich M. Social adolescents, social media, and social emotional development. Lancet Child Adolesc Health. 2023 Oct;7(10):673-675. doi: 10.1016/S2352-4642(23)00177-3. Epub 2023 Aug 8. PMID: 37567198. Final thought is that I would recommend considering a more standardized approach to capturing the array of articles that are published each year, as it is likely that many fall through the cracks for a variety of reasons. We would welcome some suggestions as to how this approach might look. We have been scanning the same databases for years, essentially PubMed bit also others, and hope we haven’t missed much relevant work this way. The problem rather is to make a choice and we hope, as we state in our introductory paragraph, not to offend anybody by not citing their work. However, this article is growing in length from year to year and we are rather afraid it might become undigestible at some point. In any case, we agree that it is a subjective choice and that therefore there will always be room for debate." } ] }, { "id": "186883", "date": "06 Sep 2023", "name": "Jeremy Stern", "expertise": [ "Reviewer Expertise Tourette syndrome." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nAs introduced by the authors this is the ninth such publication, providing a very useful and by now customary (not to mention awaited) round-up of the field, from the clinical and therapeutic to scientific, including taking in selected reviews.\nThere are regular published reviews of Tourette syndrome both in the round and more focussed areas and some are rather substantial. The purpose of this admirably concise article cannot be to provide the history and full context of each research direction, rather it serves to give a helpful snapshot of publishing in the whole area from the last year. It manages to be comprehensive in important themes, divided into 5 categories, but not over inclusive in data. It is balanced and impartial. It is inevitable that reading it will improve one's breadth of knowledge.\nFor readers less familiar with the area it may be useful to start with the conclusions which signpost areas of the last year that are attracting the most attention, both in the research and patient communities. A striking absence is also highlighted, the paucity of recent new pharmacological work.\nI note one paper is cited twice in both the pathophysiology and treatment sections in fairly similar terms (Morand-Beaulieu et al 2022), but undoubtedly does deserve inclusion in the review.\n\nIs the topic of the review discussed comprehensively in the context of the current literature? Yes\n\nAre all factual statements correct and adequately supported by citations? Yes\n\nIs the review written in accessible language? Yes\n\nAre the conclusions drawn appropriate in the context of the current research literature? Yes", "responses": [ { "c_id": "10345", "date": "16 Nov 2023", "name": "Andreas Hartmann", "role": "Author Response", "response": "We thank Dr Stern for his positive evaluation of our annual review." } ] } ]	1	https://f1000research.com/articles/12-826
https://f1000research.com/articles/12-430/v1	21 Apr 23	{ "type": "Case Report", "title": "Case Report: An unusual presentation of granulomatosis with polyangiitis", "authors": [ "Ichrak Bannour", "Maroi Ben Brahim", "Sondes Arfa", "Soumaya ben Amor", "Asma Ben Mabrouk", "Olfa Berrich", "Sonia Hammemi", "Maroi Ben Brahim", "Sondes Arfa", "Soumaya ben Amor", "Asma Ben Mabrouk", "Olfa Berrich", "Sonia Hammemi" ], "abstract": "Aim: We are reporting a case of an unusual presentation of granulomatosis with polyangiitis (GPA) with liver involvement. Case presentation: A 45-year-old male patient presented with erythematous plaques on the face and bilateral nasal obstruction. On physical examination, the patient had a ring-shaped squamous plaque on the face. The laboratory findings revealed an accelerated erythrocyte sedimentation rate at 100 mm/h, an elevated C-reactive protein at 66 mg/L, hyper gamma globulinemia 16 g/L and an elevated alkaline phosphatase (twice the upper normal limit). The craniofacial and thoracoabdominal computed tomography (CT) -scans showed ethmoid and maxillary sinusitis, low facial bone density, multiple mediastinal and hilar lymphadenopathy, diffuse small pulmonary nodules, and hepatomegaly. A cutaneous lesion biopsy, the nasal mucosa, and the liver showed a chronic inflammatory granulomatosis process with necrosis. Serum anti-neutrophil cytoplasmic antibody (ANCA) against PR3 was positive. The clinical, biological, radiological, and histological findings substantiated the diagnosis of GPA. The patient received systemic steroids combined with cyclophosphamide pulses on days 1, 14 and 28 and then he was lost to follow-up. Two-years later, he presented with a cardiac failure and skin ulcer in the right lower limb. A nasal endoscopic exam showed nasal septum cartilage perforation with resorption of the middle and inferior nasal concha. Two weeks later, he developed a diffuse alveolar hemorrhage and was therefore transferred to the intensive care unit but died of respiratory failure 3 days later. Conclusion: Clinicians should be aware of GPA atypical clinical manifestations.", "keywords": [ "Granulomatosis with polyangiitis", "Skin lesions", "Necrotizing granulomatosis hepatitis", "Case report." ], "content": "Introduction\n\nGranulomatosis with polyangiitis (GPA) is an anti-neutrophilic cytoplasmic antibody (ANCA) associated systemic small vessel vasculitis with a necrotizing granulomatosis inflammation. It was described for the first time by Friedrich Wegener in 1936.1 GPA has as a wide spectrum of clinical manifestations making it a challenging diagnostic dilemma for clinicians. The diagnosis is based on the association of clinical features, laboratory tests, and histological findings. The condition It is mainly characterized by necrotizing granulomatosis of the upper and lower respiratory tract and by glomerulonephritis. However, the granulomatosis inflammation or the vasculitis can affect any organ. Liver involvement in patients with GPA was rarely reported. We are reporting a case of an unusual presentation of GPA involving the upper airways, lungs, skin, and the liver with lethal cardiac manifestations.\n\n\nPatient and observation\n\nA 45-year-old Tunisian male patient working as a professor, who had no relevant family history presented with erythematous plaques on the face and bilateral nasal obstruction. He reported fatigue, loss of appetite, and unintentional weight loss with mouth and eye dryness for several months.\n\nOn physical examination, the patient had a ring-shaped squamous plaque on the face (Figure 1) and unilateral submandibular lymphadenopathy. A nasal endoscopy found congested nasal mucosa with bloody crusts while laryngoscopy showed increased thickness of the right vocal cord and paralysis of the left vocal cord. The ophthalmological examination concluded a low tear break-up time. The pulmonary auscultation revealed bibasilar crepitations and abdominal palpation hepatomegaly. A biopsy was performed in the cutaneous lesion showing a granulomatosis inflammation.\n\nThe laboratory findings showed an accelerated erythrocyte sedimentation rate (100 mm/h), an elevated C-reactive protein (66 mg/L), hyper gamma globulinemia (16 g/L), and an elevated alkaline phosphatase (twice the upper normal limit). The rest of the lab tests including cell blood count, creatinine, electrolytes, angiotensin-converting enzyme, aspartate aminotransferase and alanine aminotransferase were normal. Urinalysis was negative for glucosuria, proteinuria, and blood cells. Serological tests for bacterial and viral infections including hepatitis B, hepatitis C, and tuberculosis were also negative. Serum ANCA against PR3 was positive.\n\nA nasal mucosa biopsy revealed a granulomatosis inflammation with necrosis.\n\nThe craniofacial and the thoracoabdominal computed tomography (CT) scans showed ethmoid and maxillary sinusitis, low facial bone density (Figure 2), multiple mediastinal and hilar lymphadenopathy, diffuse small pulmonary nodules, and hepatomegaly. The clinical, biological, radiological, and histological findings substantiated the diagnosis of GPA.\n\nSince liver involvement was suspected, a liver biopsy was also performed and was positive for chronic inflammatory granulomatosis process without necrosis (Figure 3).\n\nThe patient received systemic steroids initiated by a 3-day regimen of methylprednisolone at a dose of 1 g a day, then oral prednisone at a dose of 1 mg/kg/day combined with cyclophosphamide pulses at a dose of 0.6 mg/m2 on days 1, 14 and 28. Then he was lost to follow-up.\n\nTwo years later, he was readmitted for dyspnea on exertion, edema of the lower limbs, and a painful ulcer of his leg that had appeared 3 months prior to admission. On examination, he had tachypnea with a respiratory rate of 34 breaths/min, a large ulcer with necrosis and pus in the right leg (Figure 4), and severe peripheral pitting edema. The nasal endoscopic exam showed nasal septum cartilage perforation with resorption of the middle and inferior nasal concha. The lab tests revealed a biological inflammatory syndrome and elevated alkaline phosphatases. Pus culture was positive for Pseudomonas aeruginosa. A skin biopsy showed leukocytoclastic vasculitis. An electrocardiogram revealed a bifascicular block and a transthoracic echocardiography showed a dilated right ventricle, a pericardial effusion, a tricuspid valve regurgitation, and a pulmonary artery pressure of 80 mmHg. The patient received antibiotics to treat his skin infection and intravenous diuretics and oxygen via nasal cannula to manage his heart congestive failure. He was also started on three methylprednisolone pulses relayed by oral corticosteroids and cyclophosphamide. Two weeks later, he developed a diffuse alveolar hemorrhage. He was transferred to the intensive care unit, but died of respiratory failure 3 days later.\n\n\nDiscussion\n\nGPA is a severe life-threatening multi-systemic vasculitis usually involving the upper and lower respiratory tract and the kidneys.1 However, it may involve other organs with variable clinical presentations.\n\nOur patient initially presented with upper airways signs, pulmonary symptoms, skin lesions and liver involvement. The diagnosis of GPA was based on the combination of clinical manifestations with serological and histological evidence.1\n\nThe liver involvement was confirmed by cholestasis and a chronic inflammatory granulomatosis process in the biopsy, which made this case worth reporting. In fact, although granuloma and/or the vasculitis may be found in any organ, the liver does not belong to the major target organs compared to the upper and lower airways. In fact, nasal and sinus involvement is the most common GPA manifestation.1 Up to 85% of patients present with necrotizing granulomatosis lesions in the nasal cavity and sinuses.2 The most frequent clinical features are nose crusting, blood stained rhinorrhea and nasal obstruction.3 Pulmonary involvement is slightly less common and occurs in about 80% of patients with a wide spectrum of lesions.4 Nodules are the most common radiological features.4 Cavitation is a hallmark of the disease but it is only found in 22% of patients.4 Liver involvement in patients with GPA is not quite clear.5 It has been demonstrated that the liver is frequently affected in patients with active GPA when the affection is mirrored by liver test abnormalities.4 The biochemical picture during active disease revealed both a cholestatic pattern (i.e. increased g-GT and ALP) as well as a hepatocellular pattern (with increased ALT and AST) and was found in 2% to 25% patients with GPA at the time of diagnosis and was associated with a severe disease course and a poor prognosis.6 However, necrotizing granulomatosis hepatitis in liver biopsy was rarely reported.4 To the best of our knowledge, this might be the fifth case of hepatic involvement granulomatosis hepatitis reported in the literature.7\n\nFurthermore, it would be interesting to note that the patient presented with a squamous erythematosus plaque in the face as the initial presentation of the disease, which is particularly unusual since typical skin lesions in GPA consist of palpable purpura, papules, subcutaneous nodules and more rarely pyoderma-gangrenosum such as ulcers, digital ulcerations and gangrenes.1,4 In fact, the cutaneous involvement in GPA is common and can be the revealing presentation in more than 30% of cases.8 The spectrum of skin lesions is very large.1 They can be classified as specific or nonspecific depending on the presence of histopathological evidence of vasculitis with or without granuloma.4 Despite the unusual clinical presentation, the histological findings consisted of specific skin lesions in an ANCA associated vasculitis.\n\nMortality rate associated with GPA is as high as 80% in untreated patients. The conventional treatment for severe GPA consists of combination of high doses of systemic glucocorticoid and immunosuppressant agents which have contributed to the improvement of the prognosis of patients with GPA. However, these patients are still at risk of various complications. Several clinical factors were identified to predict higher mortality rates. In fact, liver involvement was proven to be related to the disease activity and to predict poor clinical outcomes.9\n\nIn the present case, the patient did not comply with the treatment. He presented after two years with acute heart failure with a rapidly progressive fatal outcome. Although cardiac involvement is considered to be uncommon in patients with GPA, the condition could be life threatening.6,8 This manifestation is probably underestimated since it lacks specificity.10 In fact, depending on the series and the methods applied such as electrocardiogram, echocardiography or cardiac magnetic resonance imaging (MRI), the prevalence of cardiac manifestations ranged between 40% and 60% of the patients.11 Pericarditis is the most common cardiac manifestation and was reported in up to 35% of patients, followed by cardiomyopathy [30%] and coronary artery disease (12% of the patients).12\n\n\nConclusion\n\nThis case demonstrates that atypical clinical manifestations of GPA are very frequent, and the diagnostic of the disease should be guided by histological and laboratory findings. Clinicians should be aware of liver involvement because it can be life-threatening. Systemic corticosteroids and immunosuppressant drugs remain the mainstay in the treatment of this potentially fatal disease. However, the frequencies of these atypical manifestations are to be studied in patients with GPA.\n\n\nDeclarations\n\nNot applicable.\n\nWritten informed consent was obtained from the patient’s next of kin for publication of this case report and any accompanying images.\n\n\nAuthors’ contributions\n\nIchrak Bannour, Marwa Ben Brahim, OlfaBerriche, Sondes Arfa, Asma Ben Mabrouk and Sonia Hammemi all contributed equally to the conception, design, drafting and revising of the manuscript. All authors read and approved the final manuscript.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\n\nAcknowledgement\n\nThe authors thank the entire staff of the Department of Internal Medicine at Mahdia University Hospital (Tunisia) for their contribution in the management of the patient.\n\n\nReferences\n\nComarmond C, Cacoub P: Granulomatosis with polyangiitis (Wegener): Clinical aspects and treatment. Autoimmun. Rev. 2014 Nov; 13(11): 1121–1125. PubMed Abstract \| Publisher Full Text\n\nWojciechowska J, Krajewski W, Krajewski P, et al.: Granulomatosis With Polyangiitis in Otolaryngologist Practice: A Review of Current Knowledge. Clin. Exp. Otorhinolaryngol. 2016 Mar; 9(1): 8–13. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGreco A, Marinelli C, Fusconi M, et al.: Clinic manifestations in granulomatosis with polyangiitis. Int. J. Immunopathol. Pharmacol. 2016 Jun; 29(2): 151–159. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLohrmann C, Uhl M, Kotter E, et al.: Pulmonary manifestations of wegener granulomatosis: CT findings in 57 patients and a review of the literature. Eur. J. Radiol. 2005 Mar; 53(3): 471–477. PubMed Abstract \| Publisher Full Text\n\nWilleke P, Schlüter B, Limani A, et al.: Liver involvement in ANCA-associated vasculitis. Clin. Rheumatol. 2016 Feb; 35(2): 387–394. PubMed Abstract \| Publisher Full Text\n\nPavone L, Grasselli C, Chierici E, et al.: Outcome and prognostic factors during the course of primary small-vessel vasculitides. J. Rheumatol. 2006 Jul 1; 33(7): 1299–1306. PubMed Abstract\n\nHoll-Ulrich K, Klass M: Wegener’s granulomatosis with granulomatous liver involvement. Clin. Exp. Rheumatol. 2010; 28(Suppl. 57): S88–S89.\n\nAbdel-Halim M, Mahmoud A, Ragab G: Cutaneous manifestations of anti-neutrophil cytoplasmic antibody associated vasculitis. Vessel Plus. 2022 [cited 2022 Mar 14]. Publisher Full Text Reference Source\n\nLiao QQ, Ren YF, Zhu KW, et al.: Long-Term Prognostic Factors in Patients With Antineutrophil Cytoplasmic Antibody-Associated Vasculitis: A 15-Year Multicenter Retrospective Study. Front. Immunol. 2022 Jun 30; 13: 913667. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPugnet G, Gouya H, Puéchal X, et al.: Cardiac involvement in granulomatosis with polyangiitis: a magnetic resonance imaging study of 31 consecutive patients. Rheumatology. 2017 Jun 1; 56(6): 947–956. PubMed Abstract \| Publisher Full Text\n\nMcGeoch L, Carette S, Cuthbertson D, et al.: Cardiac Involvement in Granulomatosis with Polyangiitis. J. Rheumatol. 2015 Jul; 42(7): 1209–1212. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAhmed T, Meredith D, Klein AL: Granulomatosis With Polyangiitis (Wegener’s Granulomatosis) Complicated by Pericarditis: Our Experience of Two Cases and Comparative Review of Literature. CASE. 2021 Apr; 5(2): 126–136. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "170566", "date": "12 May 2023", "name": "Gloria Vasquez", "expertise": [ "Reviewer Expertise Rheumatologist and immunology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nI consider that the case is very interesting, but there is data in the clinical history that are missing. In the initial evaluation of liver function, neither the levels of g-GT nor LA appear, in the presence of a granulomatous infiltration of the liver, these data are critical.\nIn the image of the liver biopsy, it would be worth pointing out the findings and specifying the site of the taking, the staining and the resolution of the image in the figure caption.\nIn the second part of the story, the patient arrives with right heart failure and pulmonary hypertension, its origin is not clear.\nThe discussion refers to seven cases of liver involvement in the literature but does not describe whether the findings were similar to the case described or different.\nIt is reported that liver involvement can be a cause of mortality but here the patient died of alveolar hemorrhage, so this aspect is not clear.\nIt would be useful to describe the title of the anti PR3.\nThe authors describe that tuberculosis was ruled out serologically, what type of test was used.\nI think that all these details should be clarified, the discussion should be improved, in order to take advantage of a case that is very interesting.\n\nIs the background of the case’s history and progression described in sufficient detail? Partly\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Partly\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Partly\n\nIs the case presented with sufficient detail to be useful for other practitioners? Partly", "responses": [ { "c_id": "9698", "date": "30 May 2023", "name": "Ichrak Bannour", "role": "Author Response", "response": "Reviewer comments In the initial evaluation of liver function, neither the levels of g-GT nor LA appear, in the presence of a granulomatous infiltration of the liver, these data are critical. We thank the reviewer for underscoring this point which allows us to add the initial evaluation of liver function in lines 31 and 33 “and an elevated alkaline phosphatase (twice the upper normal limit).” and “gamma-glutamyl transferase, GGT were normal”. In the image of the liver biopsy, it would be worth pointing out the findings and specifying the site of the taking, the staining and the resolution of the image in the figure caption. We are grateful to the reviewer for this comment. We pointed out in the liver biopsy Liver biopsy hepatic sinusoids with neutrophilic infiltration and microabscess formation. And we added in line 48, 49 and 50 “Liver biopsy showing hepatic sinusoids with neutrophilic infiltration and microabscess formation (arrows), H&E staining, 200× magnification.” Figure 3. Chronic inflammatory granulomatosis process on biopsy: Liver biopsy showing hepatic sinusoids with neutrophilic infiltration and microabscess formation (arrows), H&E staining, 200× magnification. In the second part of the story, the patient arrives with right heart failure and pulmonary hypertension, its origin is not clear. We thank the reviewer for this remark. In this case the patient did not adhere to the immunosuppressive treatment and presented after two years with a rapidly lethal right heart failure. What made our patient different was the fact that he presented with an acute congestive heart with concomitant pericarditis, dilated cardiomyopathy, valvar dysfunction and conduction defect which is extremely exceptional. We precise in line 133-136 that “. He presented after two years with acute heart failure with a rapidly progressive fatal outcome. He had an acute congestive heart with concomitant pericarditis, dilated cardiomyopathy, valvular dysfunction and conduction defect which is extremely exceptional in this pathology”. The discussion refers to four cases of liver involvement in the literature but does not describe whether the findings were similar to the case described or different. As underlined by the reviewer, this was the fifth case of liver involvement described in the literature all revealed by a persistent elevation of liver biological parameters and confirmed by liver biopsies after excluding adverse drug affect, viral hepatitis, alcoholic or ischaemic hepatitis in all patients. The serology was also negative for autoimmune hepatitis and primary biliary cholangitis, and the liver biopsies did not show evidence for these diseases. All patients the literature unlike the patient in this case, responded to aggressive immunosuppressive drugs. This was added in line 109-114: “All that cases were revealed by a persistent elevation of liver biological parameters and confirmed by liver biopsies after excluding adverse drug affects, viral hepatitis, alcoholic or ischaemic hepatitis. The serology was also negative for autoimmune hepatitis and primary biliary cholangitis, and the liver biopsies did not show evidence for these diseases. All patients in the literature unlike the patient in this case, responded to aggressive immunosuppressive drugs.” It is reported that liver involvement can be a cause of mortality but here the patient died of alveolar hemorrhage, so this aspect is not clear. We thank the reviewer for this remark which allowed us to clarify the cause of mortality of the patient in this case. “In fact, he died from a respiratory failure due to diffuse alveolar hemorrhage which can be, either another rare and mortal complication of GPA occurring in only 5% to 10% of the patients, or a complication of the acute heart failure. In another hand, Necrotizing granulomatosis hepatitis could increase the risk of heart failure”. The pathogenesis of death in this case might be multifactorial. We added this in the discussion line 147-151. It would be useful to describe the title of the anti PR3. We thank the reviewer for this interesting remark. Indirect immunofluorescence (IIF) for ANCA detection was performed using a commercial kit (Euroimmun®). Our patient had c-ANCA positivity with a titer of 1:40 (< 1:20). P-ANCA was negative. Positive serums for c- or p-ANCA at IIF are subsequently screened for antigenic specificity (MPO or PR3). Proteinase 3 antibody (anti-PR3) was positive by an immunodot technique (immunodot, Euroimmun(®). Then we added in line 38, 39 “Our patient had c-ANCA positivity with a titer of 1:40 (< 1:20). P-ANCA was negative. Proteinase 3 antibody (anti-PR3) was positive by an immunodot technique” The authors describe that tuberculosis was ruled out serologically, what type of test was used. We thank the reviewer for this interesting remark. The QuantiFERON-TB Gold In-Tube (QFT-G) test for the diagnosis of tuberculosis disease was negative. We added this in line 37." } ] } ]	1	https://f1000research.com/articles/12-430
https://f1000research.com/articles/12-1397/v1	23 Oct 23	{ "type": "Research Article", "title": "Body fat, overweight and obesity in adult menopausal women", "authors": [ "Rosario Garland", "Pavel J. Contreras", "Fernando Tume", "Giuliana Rosa Del Castillo Vidal", "Oriana Rivera-Lozada", "Michelle Lozada-Urbano", "Rosario Garland", "Pavel J. Contreras", "Fernando Tume", "Giuliana Rosa Del Castillo Vidal", "Oriana Rivera-Lozada" ], "abstract": "Background This study aimed to assess the factors correlated with the percentage of body fat, overweight, and obesity in menopausal adult women.\n\nMethods In this retrospective cohort study, data were extracted from the medical records of women aged 40 to 60 years, encompassing both premenopausal and menopausal phases. The variables under consideration comprised anthropometric indicators like weight, height, age, percentage of body fat, as well as sociodemographic elements including place of origin, marital status, physical activity, frequency of visits to nutritional consultations, and dietary consumption patterns. Additionally, the body mass index (BMI) was computed to determine overweight and obesity.\n\nResults The application of multiple regression analysis unveiled that a range of 8 to 16 nutritional consultations (Relative Risk (RR): 1.78 [95% Confidence Interval (CI): 1.42-2.25]; p < 0.001), along with abstaining from coffee consumption (RR = 8.13 [95% CI: 1.22-54.31]; p < 0.031), exhibited associations with lower body fat among menopausal women.\n\nConclusions The absence of coffee consumption and engagement in nutritional consultations were linked to diminished levels of body fat in menopausal women. Consequently, it is imperative to comprehensively evaluate middle-aged women to timely address overweight or obesity with suitable nutritional guidance and recommendations.", "keywords": [ "Menopause", "Food consumption", "Overweight", "Obesity", "Women", "Body fat" ], "content": "Introduction\n\nMenopause signifies the phase during which ovarian activity ceases, leading to the discontinuation of ovulation and impeding fertilization. This transition is accompanied by notable hormonal, physical, and psychological transformation. Numerous women encountering menopausal symptoms undergo a discernible deterioration in their quality of life, particularly those with excess weight, obesity, and exposure to adverse environmental influences.1,2\n\nThe global prevalence of overweight and obesity is widespread. Projections suggest that the prevalence of obesity in women is poised to exceed 20% by 2025.3 Obesity is a comorbidity that increases the risk to many disorders. From a physiological perspective, the adverse ramifications of obesity are intricately linked to dysfunctions within adipose tissue, engendering perturbations in the hormonal milieu, encompassing sexual hormones.4\n\nThe menopausal transition renders women susceptible to weight gain,5 a phenomenon observed irrespective of prior body composition, size, geographical locale, or ethnicity.6 Concomitantly, alterations in fat mass distribution arise due to hormonal perturbations, encompassing heightened levels of androgens,7 and follicle stimulating hormone (FSH).8 Noteworthy is the observation that postmenopausal women harboring a healthy body mass index (BMI) yet exhibiting elevated adiposity in the trunk region and diminished adiposity in the lower extremities, manifest heightened vulnerability to cardiovascular ailments.9,10 Certain investigations posit that women who maintain a low BMI in the premenopausal phase, engage in regular physical activity, adhere to a healthful diet, and possess higher educational attainment, tend to sustain a favorable distribution of body fat subsequent to menopause.10 Conversely, multiparity stands recognized as a risk factor for obesity development among postmenopausal women.11 Strategies encompassing hormone replacement therapy administered to premenopausal women with elevated BMI, exhibit a higher risk of venous thromboembolism, notably pronounced within the initial year of treatment.12\n\nThe assessment of coping strategies concerning the repercussions of menopause necessitates a comprehensive exploration of the determinants linked to heightened body fat accumulation. Such endeavors hold crucial significance in facilitating proficient assistance for menopausal women,13 underscoring the imperative nature of elucidating distinct impediments obstructing the assimilation of healthful lifestyle practices among postmenopausal individuals.14 Hence, the primary objective of this study was to evaluate the determinants linked with the proportion of body fat, overweight, and obesity among a cohort of menopausal adult women aged 40 to 60 years. This investigation was conducted within the premises of a private clinic situated in Lima, Peru.\n\n\nMethods\n\nQuantitative, observational retrospective cohort study.\n\nThe population consisted of premenopausal and menopausal women aged 45 to 60 years, who attended a nutrition consultation at a clinic in Lima-Perú between 2020 and 2021.\n\nInclusion criteria: women who attended the clinic and had complete data; and exclusion criteria were patients with other comorbidities.\n\nThe medical records of women who attended a nutritional consultation at a clinic in Lima and were seen by a clinical nutritionist were reviewed. Premenopausal and menopausal women attended every week for medical attention and nutritional care. The nutritional intervention was adjusted to the characteristics and needs of the patients.\n\nThe information was taken from the patients’ consultation records. To obtain the sociodemographic variables, the nutritional histories were reviewed to obtain place of origin, age, district of origin and occupation.\n\nBecause this is a secondary database study, we did not participate in subject selection. Data were taken from the entire population that participated in the nutritional care. The researchers received the complete and coded database.\n\nThis study used dependent sample data, so exposure was assessed for the entire sample. It was verified that all participants at the beginning of nutritional care remained until the last nutritional care.\n\nThe plausibility of the data was verified before statistical processing. The cleaning of the database did not identify any implausible or missing data, so it was not necessary to perform any type of imputation.\n\nAfter data cleaning, the statistical analysis was carried out using STATA (StataCorp. 2021). Numerical variables were analyzed for data distribution and to evaluate assumptions for inferential testing. Normality was assessed using graphical and statistical methods.\n\nIn the descriptive analysis of numerical variables, means and standard deviation were determined. Categorical variables were described in frequencies and percentages. The variable Body Fat Variation was constructed as dependent and to respond to the main objective of the study. Hypothesis testing was performed with the T-student statistic for the numerical variables weight and body fat. Hypothesis testing for categorical variables was performed with the Fisher’s exact statistic. The inferential analysis was performed with a linear regression model, including the body fat variation variable as an outcome, and entering the other variables of interest into the multivariate model. A p value of less than 0.05 was considered statistically significant.\n\nThe present project adhered rigorously to the ethical tenets set forth in the Declaration of Helsinki. Data were procured from the repository of the designated reference center where the study was executed, emanating from routine clinical nutritional care procedures. Patients who attended nutritional care consent to participate in nutritional treatment as part of the Clinic’s procedures. Patient identities were safeguarded through the utilization of codes that eschewed any potentially identifying information, thereby ensuring the stringent maintenance of confidentiality. Access to these anonymized data was exclusively granted to the research investigators. Ethical clearance for this study was duly obtained from the Ethics Committee of Norbert Wiener University (Application No. 907-2021).\n\n\nResults\n\nIn total, 58 women were evaluated, more than half of the women were between 52 and 60 years of age (53.4%), were not housewives (70.2%), and received between 8 and 16 consultations (70.2%). A third of participants consumed tobacco (36.2%), consumed coffee (77.6%), consumed water (91.4%), consumed soft drinks (43.1%), consumed fish (87.7%), consumed chicken (94.7%), consumed beef (80.7%), consumed offal (22.8%), consumed eggs (100%), consumed pork (38.6%), consumed dairy products (96.5%), and were physically active (41.1%). BMI values changed (initial BMI-final BMI), Normal (19%-39.7%), Overweight (39.7%-34.5%), Obesity (41.3%-25.8%) (see Table 1).\n\n* BMI: body mass index (weight/height2).\n\nTable 2 shows a considerable decrease in weight and body fat when comparing the measurements at the beginning and at the end of the nutritional consultations, a difference that is statistically significant.\n\nThe sole variable demonstrating a significant association with the reduction in body fat percentage was the frequency of nutritional consultations falling within the range of 8 to 16 sessions (p<0.05) (Table 3).\n\nMultiple regression analysis shows the variables nutritional consultation and no coffee consumption to be associated with decreased body fat (p<0.05) (Table 4).\n\n* Adjusted by all the variables in the table.\n\n\nDiscussion\n\nMore than half of the women were between 52 and 60 years of age and the foods with the highest consumption were coffee (77.6%), water (91.4%), fish (87.7%), chicken (94.7%), beef (80.7%), eggs (100%) and dairy products (96.5%). Factors associated with decreased body fat were having between 8 to 16 nutritional consultations (RR:1.78; p<0.001) and no coffee consumption (RR:8.13; p<0.031).\n\nThe entry to menopause comes with an imbalance of estrogen and androgens levels leading to impaired energy metabolism. These hormonal changes lead to the increase in adipose tissue that is key to overweight and obesity.15 In this study 60% of the patients were overweight and obese, the patients were admitted to a feeding regimen, according to the answers given to the clinical nutritionist in charge of the care, and consumption alternatives were offered. Weight gain is common during the menopausal transition process and women who are already obese or overweight tend to gain more weight when they reach and during menopause. Nutritionally, this should be addressed by reducing caloric intake and promoting physical activity and other good lifestyles.16 Authors such as Ko and Jung (2020) suggest some foods as part of the diets to appropriately and timely address postmenopausal women.17\n\nDmitruk et al. (2018) found that the perimenopausal group had higher values of body mass, hip circumference, and most skinfolds. Postmenopausal women presented higher body fat percentage and for the lower lean tissue, soft tissue, and total body water content. The researchers found that the highest proportion of obese women was found in the postmenopausal group, where 40% of them presented visceral obesity. Thus, menopause contributed to changes in the distribution of fat tissue.18\n\nIn this study it was found that women who received between 8 to 16 nutritional consultations had a lower amount of fat mass, which indicates that the advice of a nutrition professional would help to avoid fat accumulation in postmenopausal women. Studies have shown that nutritional consultations followed by adherence to low-calorie diets have also had a large effect on weight loss.19–21\n\nOur results showed that not consuming coffee (RR=8.13 [95% CI: 1.22-54.31]; p<0.031), is associated with lower body fat among menopausal women. This is in line with previous studies that indicate a link between coffee and obesity22 and other chronic conditions such as vascular problems in menopausal women.23 The explanatory mechanism by which overweight or obesity is present could be because coffee is a product that contains mainly caffeine and some studies suggest that constant and prolonged caffeine consumption impacts the quantity and quality of sleep.24,25 Scientific evidence indicates that lack of sleep is a risk factor for several types of metabolic problems, including obesity. The rationale for this is that when individuals get too little sleep, they alter the release of multiple hormones, including cortisol, whose increase causes hyperglycemia and thus insulin activates lipogenesis.26 In this study, the participants may have different frequencies of consumption, amounts of coffee ingested, type of coffee consumed, time of consumption, amount of sugar, among other aspects of eating habits. Therefore, further studies are needed to identify the quantity and time of consumption of caffeine-containing beverages in pre- and post-menopausal women.\n\nAs a limitation, we did not identify the specific intervention that each of the participants received, so that the benefit of the diet plan and recommendations provided by the professional nutritionist could alter the results by increasing or decreasing the strength and direction of the association. However, the professional nutritionist followed an institutional protocol for the nutritional management of patients.\n\nAnother limitation of the study is that there was no data on the number of calories ingested and intensity of the physical activity performed by each patient. These particularities may modify the strength and direction of the association. Based on our data, we provide indications of the possible positive effect of following a structured diet and performing physical activity.\n\n\nConclusions\n\nThe abstention from coffee consumption and engagement in nutritional consultations have been found to correlate with a reduction in body fat among menopausal women. The adoption of low-carbohydrate and low-fat dietary regimens may offer efficacy in the context of body fat reduction in pre and postmenopausal women.", "appendix": "Data availability\n\nFigshare: Food consumption in menopausal, https://doi.org/10.6084/m9.figshare.23929638.v1. 27\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nPichilingue J, Flores R, Brandt A, et al.: Climaterio y Menopausia. Rev Peru Ginecol y Obstet. 2015; 42(4): 15–25. Publisher Full Text\n\nTorres A, Torres J: Climaterio y menopausia. GPC Rev la Fac Med la UNAM. 2018; 61(2): 51–58.\n\nDi Cesare M, Bentham J, Stevens GA, et al.: Trends in adult body-mass index in 200 countries from 1975 to 2014: A pooled analysis of 1698 population-based measurement studies with 19.2 million participants. Lancet. 2016 Apr 2; 387(10026): 1377–1396. Publisher Full Text\n\nBray GA, Kim KK, Wilding JPH: Obesity: a chronic relapsing progressive disease process. A position statement of the World Obesity Federation. Obes. Rev. 2017 Jul 1; 18(7): 715–723. PubMed Abstract \| Publisher Full Text\n\nKarvonen-Gutierrez C, Kim C: Association of mid-life changes in body size, body composition and obesity status with the menopausal transition. Healthcare (Switzerland). Healthcare (Basel). 2016; 4. Publisher Full Text\n\nAmbikairajah A, Walsh E, Tabatabaei-Jafari H, et al.: Fat mass changes during menopause: a metaanalysis. Am. J. Obstet. Gynecol. 2019; 221: 393–409.e50. Mosby Inc. PubMed Abstract \| Publisher Full Text\n\nKohrt WM, Wierman ME: Preventing Fat Gain by Blocking Follicle-Stimulating Hormone. N. Engl. J. Med. 2017 Jul 20; 377(3): 293–295. PubMed Abstract \| Publisher Full Text\n\nChen GC, Arthur R, Iyengar NM, et al.: Association between regional body fat and cardiovascular disease risk among postmenopausal women with normal body mass index. Eur. Heart J. 2019 Sep 7; 40(34): 2849–2855. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBlew RM, Sardinha LB, Milliken LA, et al.: Assessing the validity of body mass index standards in early postmenopausal women. Obes. Res. 2002; 10(8): 799–808. PubMed Abstract \| Publisher Full Text\n\nTsai TI, Chou P: The association of body fat distribution with lifestyle and reproductive factors in women aged 45-54 in Kinmen County, Republic of China. J. Women’s Health. 1999; 8(4): 501–508.\n\nFrança AP, Marucci MdFN, Silva MdLdNd, et al.: Fatores associados à obesidade geral e ao percentual de gordura corporal em mulheres no climatério da cidade de São Paulo, Brasil. Cien. Saude Colet. 2018; 23(11): 3577–3586. PubMed Abstract \| Publisher Full Text\n\nCanonico M, Oger E, Conard J, et al.: Obesity and risk of venous thromboembolism among postmenopausal women: differential impact of hormone therapy by route of estrogen administration. The ESTHER Study. J. Thromb. Haemost. 2020; 4: 1259–1265. Publisher Full Text\n\nDorjgochoo T, Kallianpur A, Gao YT, et al.: Dietary and lifestyle predictors of age at natural menopause and reproductive span in the Shanghai Women’s Health Study. Menopause. 2008 Sep; 15(5): 924–933. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPalmer JR, Rosenberg L, Wise LA, et al.: Onset of natural menopause in African American women. Am. J. Public Health. 2003 Feb 1; 93(2): 299–306. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAl-Safi ZA, Polotsky AJ: Obesity and Menopause. Best Pract. Res. Clin. Obstet. Gynaecol. 2015 May 1; 29(4): 548–553. Publisher Full Text\n\nProietto J: Obesity and weight management at menopause. Aust. Fam. Physician. 2017; 46(6): 368–370. PubMed Abstract\n\nKo SH, Jung Y: Energy metabolism changes and dysregulated lipid metabolism in postmenopausal women. Nutrients. 2021 Dec 1; 13(12). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDmitruk A, Czeczelewski J, Czeczelewska E, et al.: Body composition and fatty tissue distribution in women with various menstrual status. Rocz. Panstw. Zakl. Hig. 2018 Jan 1; 69(1): 95–101. PubMed Abstract\n\nNaude CE, Brand A, Schoonees A, et al.: Low-carbohydrate versus balanced-carbohydrate diets for reducing weight and cardiovascular risk. Cochrane Database Syst. Rev. 2022; 2022. Publisher Full Text\n\nNordmann AJ, Nordmann A, Briel M, et al.: Effects of low-carbohydrate vs low-fat diets on weight loss and cardiovascular risk factors: A meta-analysis of randomized controlled trials. Arch. Intern. Med. 2006; 166(3): 285–293. PubMed Abstract \| Publisher Full Text\n\nDansinger ML, Gleason JA, Griffith JL, et al.: Comparison of the Atkins, Ornish, Weight Watchers, and Zone Diets for weight loss and heart disease risk reduction: A randomized trial. J. Am. Med. Assoc. 2005; 293(1): 43–53. PubMed Abstract \| Publisher Full Text\n\nLee J, Kim HY, Kim J: Coffee consumption and the risk of obesity in Korean women. Nutrients. 2017; 9(12).PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFaubion SS, Sood R, Thielen JM, et al.: Caffeine and menopausal symptoms: what is the association? Menopause. 2015 Feb; 22(2): 155–158. PubMed Abstract \| Publisher Full Text\n\nGardiner C, Weakley J, Burke LM, et al.: The effect of caffeine on subsequent sleep: A systematic review and meta-analysis. Sleep Med. Rev. 2023; 69: 101764. PubMed Abstract \| Publisher Full Text\n\nStaack A, Distelberg B, Moldovan C, et al.: The Impact of Caffeine Intake on Mental Health Symptoms in Postmenopausal Females with Overactive Bladder Symptoms: A Randomized, Double-Blind, Placebo-Controlled Trial. J. Womens Health (Larchmt). 2022 Jun; 31(6): 819–825. Epub 2022 Apr 1. PubMed Abstract \| Publisher Full Text\n\nSt-Onge MP: Sleep–obesity relation: underlying mechanisms and consequences for treatment. Obes. Rev. 2017; 18: 34–39. PubMed Abstract \| Publisher Full Text\n\nLozada-Urbano M, Garland R: Food consumption in menopausal. [Dataset]. figshare. 2023. Publisher Full Text" }	[ { "id": "268722", "date": "30 May 2024", "name": "Sudip Datta Banik", "expertise": [ "Reviewer Expertise Human biology", "health", "and nutrition" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe study was descriptive in nature, based on a sample of women aged 40 to 60 years. The authors did mistakes in the methods: sampling design, selection of participants etc. Title of the paper shows \"menopausal women\". There was no need to mention 'adult'. However, the study included pre and postmenopausal women. The authors didn't identify perimenopausal women. Menopause associated metabolic alterations, body fatness, and obesity etc. are well-documented.\n\nIt was not clear how body fat data of 58 women with an age-group 40-60 years had normal distribution.\nThe paper could be a short communication with precise methods and results.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [] }, { "id": "325567", "date": "07 Oct 2024", "name": "Albert A. Opoku", "expertise": [ "Reviewer Expertise Obstetrics & Gynaecology" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe title of the article could be improved to better describe the aim/purpose of the research- were they looking at the distribution or the factors that affect body fat, overweight and obesity in menopausal women? The abstract mentions an age range of 40-60 years, the Methods section of the paper mentions 45 - 60 years and the results (Table 3) look at ages 42-60 years- very inconsistent. Though the authors received institutional ethical approval and the data they used was anonymised- it is unclear whether these patients had agreed a priori that their data can be used for research when they consulted for nutritional advice. The sample size of 58 is very small, there is no mention of a sample size calculation that will make their findings generalisable. Their analysis is based on age and there is no reference in the paper whether the ladies were pre-, peri- or postmenopausal so the conclusions in terms of menopause is not supported by their results. In Table 3, the difference between the change in body fat reduction in coffee drinkers and non-coffee drinkers (who make up 22% of their cohort) is not significant after multiple regression analysis but it is significant in Table 4, after the linear regression analysis. The authors should make make effort to explain this statistical finding as this is one of their main conclusions.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? No\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nI cannot comment. A qualified statistician is required.\n\nAre all the source data underlying the results available to ensure full reproducibility? No\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [] } ]	1	https://f1000research.com/articles/12-1397
https://f1000research.com/articles/12-1396/v1	23 Oct 23	{ "type": "Systematic Review", "title": "Open science interventions proposed or implemented to assess researcher impact: a scoping review", "authors": [ "Mona Ghannad", "Anna Catharina V. Armond", "Jeremy Y. Ng", "Ana Patricia Ayala", "Hassan Khan", "Maura R. Grossman", "Gordon V. Cormack", "Ba Pham", "Mariska M. Leeflang", "Patrick M. Bossuyt", "Karim M. Khan", "Clare L. Ardern", "David Moher", "Anna Catharina V. Armond", "Jeremy Y. Ng", "Ana Patricia Ayala", "Hassan Khan", "Maura R. Grossman", "Gordon V. Cormack", "Ba Pham", "Mariska M. Leeflang", "Patrick M. Bossuyt", "Karim M. Khan", "Clare L. Ardern", "David Moher" ], "abstract": "Background Several open science-promoting initiatives have been proposed to improve the quality of biomedical research, including initiatives for assessing researchers’ open science behaviour as criteria for promotion or tenure. Yet there is limited evidence to judge whether the interventions are effective. This review aimed to summarise the literature, identifying open science practices related to researcher assessment, and map the extent of evidence of existing interventions implemented to assess researchers and research impact.\n\nMethods A scoping review using the Joanna Briggs Institute Scoping Review Methodology was conducted. We included all study types that described any open science practice-promoting initiatives proposed or implemented to assess researchers and research impact, in health sciences, biomedicine, psychology, and economics. Data synthesis was quantitative and descriptive.\n\nResults Among 18,020 identified documents, 27 articles were selectedfor analysis. Most of the publications were in the field of health sciences (n = 10), and were indicated as research culture, perspective, commentary, essay, proceedings of a workshop, research article, world view, opinion, research note, editorial, report, and research policy articles (n = 22). The majority of studies proposed recommendations to address problems regarding threats to research rigour and reproducibility that were multi-modal (n = 20), targeting several open science practices. Some of the studies based their proposed recommendations on further evaluation or extension of previous initiatives. Most of the articles (n = 20) did not discuss implementation of their proposed intervention. Of the 27 included articles, 10 were cited in policy documents, with The Leiden Manifesto being the most cited (104 citations).\n\nConclusion This review provides an overview of proposals to integrate open science into researcher assessment. The more promising ones need evaluation and, where appropriate, implementation.\n\nStudy registration\nhttps://osf.io/ty9m7", "keywords": [ "Promotion and tenure", "researcher assessment", "incentives and rewards" ], "content": "Introduction\n\nThere have been several initiatives to improve the quality of biomedical research through open science practices. Some approaches have evidence supporting their effectiveness.1 However, sometimes the best available evidence is based on data that lack transparency, have high uncertainty, or unknown reproducibility.2 Adopting Open Data “Badges” is one example of an intervention designed to promote open science (and improve research quality). Badges are simple ways of signalling and incentivizing desirable behaviour.3 Journals can offer badges to authors who meet their criteria for open science practices, thereby signalling that the journal values transparency, and that authors have met the transparency standards for their research.3 Open Data Badges developed by the Center for Open Science (USA) to acknowledge open science practices were associated with an increase in data sharing (from 1.5% to 39%) at the journal Clinical Psychological Science.3 Hundreds of journals used this evidence and endorsed Open Data Badges. However, in a randomized controlled trial, Open Data Badges did not noticeably improve data sharing by researchers (odds ratio 0.9, 95%CI: 0.1 to 9.0).4 In addition to badges for data sharing, other signals of transparency and openness5 are also promoted.\n\nBefore encouraging broad scaling and adoption of open science practice-promoting initiatives, including for researcher assessment, it is prudent to evaluate whether proposed and/or implemented open science interventions are effective; are they ‘fit for purpose’? Furthermore, should one assume that pursuing such initiatives will immediately and automatically lead to effective implementation? Could other factors (i.e., quality of research, certainty of its conclusions, and generalizability2) boost or hinder the effects of interventions to promote open science practices? There is a need to evaluate if the hypothesised benefits are met, and if there are unintended consequences when the proposed interventions are implemented.\n\nWe aimed to provide an overview of literature identifying criteria for the assessment of researchers (for example for career advancement) that relate to signals and/or practices of open science, such as badges, preprints, registrations, data sharing. We also aimed to map the extent of evidence of existing interventions implemented to assess researchers and research impact (i.e., improvements to society, such as more transparent research and research that is shared to help improve the speed of knowledge).\n\n\nMethods\n\nScoping reviews are an appropriate research method to provide an overview of a topic or broader domain using a systematic and rigorous method. We used the Joanna Briggs Institute Scoping Review Methodology to conduct this scoping review. Prior to conducting the review, the protocol was registered. We restate our study methods here, in brief, taking large sections of the methods directly from the original protocol. Any discrepancies or deviations from the study protocol have been explained. We followed the PRISMA-ScR statement to guide our reporting of this scoping review.\n\nInclusion criteria\n\nWe included studies from the health sciences, biomedicine, psychology, and economics fields, as our impression is that open science developments are more prevalent in these disciplines, which are also closest to our knowledge and experience. We defined each discipline.6\n\nWe included studies that described any open science proposals or implemented open science practices by researchers, funders, academic institutions, societies (e.g., The Royal Society), to assess researchers and research impact, and focus on the following issues: researcher assessment (e.g., Declaration of Research Assessment – DORA), interventions (e.g., reporting guidelines7,8), or incentives to increase the speed of access to new knowledge (e.g., preprints); the availability of all knowledge (e.g., registration; open access publication); improve the quality of published research (e.g., reporting guidelines7,8); improve reproducible research (e.g., money back incentives9); and to increase sharing of data, code, and materials (e.g., open science badges3,4).\n\nOur focus was open science proposed or implemented signals/practices, as defined by the United Nations Educational, Scientific and Cultural Organization.10 Based on the Framework for Open and Reproducible Research Training (FORRT)11 and the Transparency and Openness Promotion (TOP) guidelines,12 open science topics include: reproducibility [crisis] and/or replication; design, methods, data [code] material transparency; registration and/or preregistration; publishing of research and publication models; conceptual and statistical knowledge; and academic life and culture.\n\nExamples of studies that met our eligibility criteria include: studies evaluating the use of badges to increase data sharing3,4; studies evaluation practices including pre-registration5; use of reporting guidelines7,8 and change in editorial policy13 to improve reporting and compliance in published articles; and, new publication models14 or proposals of other models9 as solutions to the reproducibility crisis in research.\n\nWe included original research reported in English. We also included policy documents from funders (e.g., Wellcome Trust; National Institute of Health (NIH), Canadian Institutes of Health Research (CIHR)), academic institutions (e.g., Utrecht University) and researchers (e.g., open science Minerva8,14). We included all article types, including but not limited to, conference proceedings, commentaries, and editorials.\n\nExclusion criteria\n\nWe excluded literature that was not published in English, and studies published before year 2000 due to logistics and resource constraints.\n\nFor our detailed search strategy please see Extended data.63 An experienced medical information specialist (APA) developed and tested the search strategy in consultation with the project leads (DM, CA and MG). The search strategy was developed in OVID MEDLINE, and peer-reviewed by another librarian.15 We searched a range of databases to achieve wide cross-disciplinary coverage, which included: OVID MEDLINE, OVID EMBASE, OVID APA PsycINFO, EBSCO Cumulative Index to Nursing and Allied Health Literature (CINAHL), and EBSCO ECONLit. The search strategies were translated using each database platform’s command language, controlled vocabulary, and appropriate search fields. Medical subject headings (MeSH), EMTREE terms, American Psychological Association thesaurus terms, CINAHL headings and text words were used to search the concepts and synonyms of “open science”, “data sharing”, “tenure” and “academic promotion”. Language limits were applied to capture articles in English. We performed all searches on October 3, 2022.\n\nWe also searched the grey literature to help identify proposed and/or implemented interventions to promote open science as part of researcher assessment. The grey literature search was limited to sources familiar to the research team. Purposive searching ensured we had the resources to complete the project in a timely manner. Supplemental search sources were: Open Science Centre; Research on Research Institute; METRICS and METRICB (Berlin); RECOGNITION & REWARDS; Open Scholarship Knowledge Base; and DORA. We performed forward citation searching of the included studies using citationchaser,16 Scopus, and Web of Science Core Collection. Additionally, we reviewed the bibliography of included studies. All search strategies are reported using PRISMA-S.17\n\nDue to the volume of literature search results, bibliographic records were identified for screening using the Continuous Active Learning® (CAL®) tool, which uses supervised machine learning to rank titles and abstracts from most to least likely to be of interest. The CAL® tool was chosen due to its prior use by some of our team members.18\n\nThe results of the database searches were imported into the CAL® tool, without removing duplicates. The CAL® tool learns from the results of manual screening by reviewers to identify and rank titles and abstracts most likely to meet the inclusion criteria.\n\nFor manual screening by reviewers, a screening form based on the review’s eligibility criteria was prepared to aid in making consistent judgements on article relevance. To calibrate, two reviewers screened the first 100 titles/abstracts using the CAL® tool, independently. Discrepancies were identified. A third reviewer selected a pilot set of 10 titles/abstracts that were similar to the discrepant abstracts. The two reviewers screened the pilot set and discussed discrepancies. Subsequently, abstract screening was conducted using the CAL® tool by one reviewer.\n\nSeveral hundred titles/abstracts were screened until the yield of relevant records diminished. After screening 1000 records, we started to look ahead among the successive batches of 100 records that we screened. We stopped when the yield from the successive batches dipped below 5 (per 100) relevant records. After stopping, the results from the titles/abstracts screening were de-duplicated prior to full-text retrieval. For the screening questions, see Extended data.63\n\nWe developed and piloted data extraction forms prior to data extraction. The extracted data included: (1) Corresponding author; (2) Discipline; (3) Publication year; (4) Journal or platform where the study was published; (5) The proposed and/or implemented interventions to assess researchers or research impact; (6) The stakeholders of the proposed/implemented intervention (e.g., academic institution, funder, publisher, and researcher); (7) Intervention type (single or multi-modal); and (8) Funding information of the study. Details of the forms used are provided in Extended data.63\n\nOur data analysis included both quantitative (i.e., frequencies and percentages) and qualitative (i.e., narrative interpretation) methods. Data collection was conducted by MG, DM, AA, and JYN. As an additional step, MG and AA discussed and verified 20 of the 27 full-text articles (open Science interventions proposed/implemented). Discussions were focused on how best to categorize whether a proposed intervention was single or multi-modal, and which stakeholders were targeted. Multi-modal interventions targeted several open science practices, although some recommendations can conceptually be linked to each other. Single-modal interventions, were those targeting a single open science practice. Stakeholders were those who were actors in conducting the proposed interventions. As a measure of impact, we entered the document object identifier of included each article in the BMJ Impact Analytics tool, to extract if the article was cited in policy documents.\n\nWe created a flow diagram to detail how articles captured in our search were screened. We also calculated the frequencies (and percentages), described narratively the characteristics of all data extracted from included documents, and present the results in summary tables. Our data extraction and presentation plan copied and pasted the text relevant information and present it from the included studies. This is noted with quotation marks at the beginning and end of each cut and paste. We reasoned this approach was more efficient and would more accurately report what the individual authors stated rather than any paraphrasing we did.\n\nThe original protocol indicated we would exclude case reports, case series, letters to editor, and narrative reviews. However, we did not exclude any records based on publication type.\n\n\nResults\n\nThe literature search retrieved 18,020 records (Figure 1). We screened 1512 titles and abstracts until the yield of relevant ones fell below 2 (per 375). Two hundred and eighty-nine records were considered eligible for full-text screening. After duplicates were removed in Endnote, 208 records remained for full-text screening. We identified 21 papers that potentially described any open science proposals or practices evaluated and/or implemented to assess researchers and research impact. One record was later excluded during the data extraction phase. A forward citation check of these 21 studies yielded 3901 records. Using search terms “research assessment” and “open science and research assessment”, resulted in 111 records and 19 relevant records for title and abstract screening and full text screening, respectively. Of these, 7 reports met our inclusion criteria. Finally, we included 2719–45 studies in our scoping review.\n\nThe 27 articles included for epidemiological data extraction were published between 2013 and 2022, with the number of studies describing open science practices for research assessment increasing gradually in more recent years (Table 1 and Table 2 – underlying data).63 The corresponding authors were from 8 countries, with North America (e.g., USA and Canada) leading the vast majority of studies published (n = 12 and n = 6, respectively). Most of the publications were indicated as research culture, perspective, commentary, essay, proceedings of a workshop, research article, world view, opinion, research note, editorial, report, and research policy (n = 22). Only three records19,31,33 in our scoping review were empirical research. The majority of the records did not report any funding information (n = 15), but of those that mentioned if they had received funding, seven records (47%) reported receiving funding, and similarly six records (53%) reported receiving no funding.\n\nThe majority of the studies proposed recommendations for researcher assessment that were multi-modal (n = 20), although some recommendation can conceptually be linked to each other, targeting several open science practices. To this purpose, several proposals have been developed such as the San Francisco Declaration on Research Assessment (DORA),40,46 The Leiden Manifesto for research metrics,42 The Metric Tide,45 and the Hong Kong Principles for Assessing Researchers.37 Some of the studies in our scoping review based their proposed recommendations on further evaluation or extension of previous initiatives. For example, Gagliardi et al.25 identified and prioritized 10 measures (that were further grouped through consensus as five measures of high importance and five measures of moderate importance) for assessing research quality and impact (DORA-compliant measures). Hatch et al.,27 formulated “practical guidance for research institutions looking to embrace reform … for driving institutional change at a meeting convened by DORA and the Howard Hughes Medical Institute”. While another study, Schmidt et al.,34 discussed the implementation of DORA and anecdotally describes practices used for hiring junior faculty members in their department. Additionally, Hatch et al.35 presented “a tool called SPACE that institutions can use to gauge and develop their ability to support new approaches to assessment, … built on design principles released by DORA, and a five-step approach to responsible evaluation called SCOPE47”. The authors claim the tool “can be adapted to different institutional contexts, geographies, and stages of readiness for reform, thus enabling universities to take stock of the internal constraints and capabilities that are likely to impact their capacity to reform how they assess research and researchers”.35 It proposes criteria such as, “diversifying career paths, focusing on research quality and academic leadership, and stimulating open science”. Understanding the relationship between SPACE and SCOPE is not inherently obvious.\n\nOther multi-modal recommendations were included that did not mention any of the previously developed statements mentioned above. One study20 proposed to ensure data sharing, registration/pre-registration, and to not assess researchers based on traditional metrics. Another study,21 with authors comprised of “The Committee on Developing a Toolkit for Fostering Open Science Practices”, drafted elements for a toolkit, discussing topics on communication of the benefit of open science, (e.g., compiling a database of research articles that describe the ways open science is beneficial, resources to signal an organization’s interest in open science activities). An anecdotal discussion that faculty promotion must assess reproducibility by new assessment practices, that ensure “researchers use proper experimental design and analysis”, and require institutions to incentivize data sharing and transparency, was discussed by another.24 While another author26 discussed several ways to reward research transparency during various stages of researcher’ career, encompassing the hiring process, promotion and tenure evaluations, and “society and organization honors and awards”.\n\nAiming to develop and evaluate “more appropriate, non-traditional indicators of research” that “facilitate changes in the evaluation practices for rewarding researchers”, Rice and colleagues33 conducted a cross-sectional study “to determine the presence of a set of pre-specified traditional and non-traditional criteria used to assess scientists for promotion and tenure in faculties of biomedical sciences” among 170 randomly selected universities worldwide. Moher et al.,36 “completed a selective literature review of 22 key documents critiquing the current incentive system” prior to convening a 22-member (e.g., academic leaders, funders, and scientists) expert panel workshop asking “how the authors perceived the problems of assessing science and scientists, the unintended consequences of maintaining the status quo for assessing scientists, and details of their proposed solutions. The resulting table was used as a seed for participant discussion”, which “resulted in six principles for assessing scientists and associated research and policy implications”.\n\nMajority of included studies were in the field of health sciences (n = 10), followed by biomedicine/and or health sciences and multidisciplinary, (n = 7 and n = 6, respectively). One study32 proposed “open access for measuring research assessment in Earth and Natural Sciences”. Several studies targeted their initiatives mainly towards redesigning or modifying researcher assessments via new metrics, measures, methods, and incentives. One study19 designed a new metrics for evaluating researchers (e.g., ‘ε-index’), while another study23 conceptualized “data-level metrics” “(DLMs) as indicators of scientific merit related to the production and (re-)use of datasets, … to capture and make data-sharing efforts visible”. Individualized research-philosophy statements, annotated curriculum vitae (CV), and “the use of a formal evaluative system that captures behaviors that support reproducibility” were also proposed,22 as a method to improve the transparency of scholarly reporting. The PQRST index – (research that is productive, high-quality, reproducible, shareable, and translatable) – is proposed by Ioannidis et al.28 for appraising and rewarding research. Nicholson et al.30 provided “a brief tutorial about impact metrics and how to use these metrics to document scientific impact, … detailing six steps to documenting the scientific and clinical impact of one’s research. Steps include: establishing an ORCID (Open Researcher and Contributor ID) account, creating research profiles on academic platforms, engaging in broad dissemination activities, harvesting bibliometric and altmetric data, adding bibliometric and altmetric information to one’s curriculum vitae, and submitting one’s promotion and tenure portfolio with confidence”. A discussion on how a researcher’s department has modified the incentive structure to support open science and replication work as well as innovation is provided by Lundwall et al.29 The author describes the designing of “incentive changes to support faculty in both quantity (i.e., publication counts) and quality (i.e., … the journal impact factors, journal rejection rates, and newly added methods that support sound science) efforts … using a self-rating rubric … to guide faculty to succeed”. Strintzel et al.44 proposed “ways of improving academic CVs for fairer research assessment”. Another study29 proposed “a novel policy to provide incentives for open science … by offering open-source (OS)-endowed professorships”. Fischman et al.41 outlined “an alternative proposal in which trustworthiness and usability of research would complement traditional metrics of scholarly relevance”41 mainly in institutions.\n\nThe ramifications of research promotion and tenure scholarship ecosystem is of significant societal impact. The policy document by the European Commission43 on the “evaluation of research careers acknowledging open science practices” also speaks to the recognition of international bodies in changing current practices. Moher et al.38 highlights the importance of the proposed interventions for open sciences practices in relation to COVID-19.\n\nThe role of stakeholders is evident in a recent publication by Bonn et al.39 captured the perspectives of research stakeholders regarding implementing changes over a five-year period. They translated their findings into four actions needed for fostering better research, with one of the action points relating to research assessment.\n\nFor the studies included in our scoping review, we extracted data on whether or not any of the proposed interventions were implemented (Table 3 – underlying data).63 Most of the studies (n = 20) did not discuss implementation of their proposed intervention. The remainder of the studies (n = 7) discussed ‘near’ implementation strategies such as, changes in how junior faculty members are identified and recruited at the author’s department, piloting and an audit and feedback process for their proposed open science intervention, and discussions on initiatives of various stakeholders for more responsible assessment of higher education (scholarship) for the society. Of the 27 included articles in our scoping review, 10 were cited in policy documents, with The Leiden Manifesto being cited the most often (n = 104; Table 4 – underlying data).63\n\n\nDiscussion\n\nWe identified 27 proposals, across several disciplines, to integrate open science practices when assessing researchers. Most of these proposals are in the form of reports or journal publications. Some of the publications are editorials, opinions, narrative reviews, and other publication categories. Few of the proposals have been evaluated. The practices that have been evaluated have not used randomized trials or systematic reviews, often considered the highest level of evidence for testing interventions. Medicine, and perhaps other disciplines, has a long history of demonstrating that when ‘promising interventions are subjected to randomized trials, the effect of the intervention decipitates.48,49\n\nOpen science is a relatively new movement more commonly endorsed in Europe and the United Kingdom’s (UK) research ecosystem. For example, the UK government has made a strong commitment to embracing the importance of reproducibility by funding the establishment of the UK’s Reproducibility Network (UKRN).50 Similarly, the European Union’s Horizon program, is a major funder of several long-term programs that are addressing several open science issues. For example, the recently funded sharing and re-using of clinical trial data to maximize impact is a three-year project that will train early career researchers to implement data sharing and data re-use.51 In the United States, the Centre for Open Science has made substantial contributions to open science, such as the Transparency and Openness Promotion (TOP) guidance.12 Similarly, there are some emerging developments aiming to integrate open science into a reimaged research assessment framework, such as the Higher Education Leadership Initiative for Open Scholarship (HELIOS).52 Finally, the Coalition for Advancing Research Assessment (CoARA) is an exciting initiative that is generating considerable attention. It was not formally included in our review as it is still in the early stages of development. The 2022 agreement, and associated 10 commitments, is available on CoARA. It has been signed by more than 350 international organizations. Like HELIOS, CoARA is aimed at senior university administrators able to make decisions and change culture.\n\nFrom our broad sweep of the literature, it is gratifying to see the emergence of several imaginative proposals to integrate open science practices into researcher assessments. For example, developing innovative ways to harmonize academic CVs that reward researchers for open science practices, such as data sharing and transparency, was proposed by several articles included in our scoping review. Similarly, moving away from traditional research metrics, and modifying the incentive structure to reward open science was mentioned in several articles we assessed. There may be some merit in considering a smaller number of core practices for all proposals to work towards evaluating and implementing. Such an approach might provide a larger evidence base for using these practices as part of a researcher’s assessment. Similarly, disseminating and training opportunities focused on a smaller core number of practices might be easier to achieve.\n\nCurrently, most researcher assessments are based on traditional quantitative metrics that are less useful (e.g., the number of publications), not evidence-based (e.g., journal impact factors), and do not incorporate other ways of assessing a researcher’s impact (e.g., registering research prior to its initiation). One measure of researcher impact might be the ability of others to reproduce methods and/or results of a researcher’s published research. This does not appear possible in the current climate of a substantive problem concerning reproducibility across disciplines.6,49 For reproducibility to be examined, it is important that researchers share their data and the analytical code underpinning their results. Neither behaviour is common in medicine, and other disciplines.53 To improve the use and impact of these and other open science practices, universities and other research organizations might want to integrate data sharing as part of researcher assessment. This practice is in keeping with what patients have requested.54 Similarly, to make any real sense of reproducibility, is it essential that authors provide a comprehensive and transparent description of what the research team did and found, namely, the methods and results. To help researchers better report their research, reporting guidelines have been developed and for some of them, there is evidence that their use is associated with improved quality of reporting.7 Traditional assessments of researchers do not ask about the use of reporting guidelines, even though there is an extensive body of evidence indicating that the quality of reporting research is suboptimal; it is wasteful.55–61\n\nFew proposals for researcher assessment have been evaluated. It may be because few funders have explicit requirements for this type of research. Another possibility is that most universities are not particularly interested in incorporating open science practices into their researcher assessments. In Canada, only 47 organizations have signed the Declaration of Research Assessment; only two are universities. Universities may see little benefit from investing in this type of research. It might be that the Horizon Europe funding program, which has already invested in open science research, and other national funders, such as France’s L’Agence Nationale de la Recherche, will fund research that focuses on the integration of open science into researcher assessment. There has been a recent call for more dialogue between universities and journals to address research misconduct.62 Perhaps something similar between funders, universities, and journals might result in advancements in research assessments.\n\nOur results did not identify proposals for integrating open science into researcher assessment from the Global South. While universities in some countries, such as Singapore (e.g., Nanyang Technological University), have implemented open science practices, there is little data about the uptake of open science in the Global South (e.g., African Open Science Platform). It could be that those responsible for researcher assessment in some countries feel compelled to use traditional metrics, allowing them to compare themselves against other regional, national, or international institutions in the university rankings game. It is also possible that proposals that promote qualitative metrics as part of researcher assessment cannot be used in some parts of the Global South. There are typically insufficient numbers of people trained in qualitative methods regardless of jurisdiction. There are likely unique issues that warrant hearing from the Global South. That said, there may be enough proposals from the Global North to advocate for a pause on any new ones. It may be more productive to evaluate some of the more promising ones, perhaps focusing on a small core set of open science practices.\n\nWhile the United Nations Educational, Scientific and Cultural Organization recommendation on open science is commendable and includes the importance of research integrity and inclusivity, we found none of the included proposals make any explicit mention of equity, diversity, and inclusion (EDI) as part of researcher assessment. For example, how diverse are research teams and do they integrate EDI as part of their research program? Do research team leaders provide EDI training to early career members of their research teams? We think that EDI principles fit nicely with UNESCO’s recommendation on open science. As such, current proposals should reconsider how they can integrate EDI as part of open science when assessing researchers.\n\nOne issue not discussed is how organizations that introduce open science practices when assessing researchers will affect faculty members moving from an institution that has integrated open science practices when assessing researchers, to an institution that has not yet developed an open science matrix for assessing researchers. How competitive will these researchers be when seeking a new position? These issues, and others, are beyond the scope of this review. Nevertheless, they likely need a deeper discussion.\n\nOur research is not without limitations. It is possible that we missed eligible studies for inclusion. Given our resource limitations, we only searched for eligible studies from 2000 to present. It is possible that relevant publications exist prior to this date and might influence our results. Similarly, we only included papers in English. It is likely that relevant work exists in languages other than English and may influence the current results. That said, we believe our search strategy was broad and comprehensive. While we used an extensive sample of papers to test and inform the artificial intelligence (AI) screening, it is possible that the AI approach was imperfect. From a face validity perspective, the papers we were familiar with were screened in by the CAL® tool. We did not search or examine all disciplines. We focused on disciplines in which we have some expertise and credibility. That said, we think the results have merit to other disciplines. Finally, the open science proposals table (Table 3 – underlying data)63 was not very precisely described in our protocol and especially the categorization of single modal and multimodal practices somewhat subjective in its development. Still, the table does provide a complete overview of the practices we retrieved and the distinction between single modal and multimodal may be a starting point for further investigations in the complexity of these assessment criteria.\n\nWe aspire for this scoping review to promote discussion across different communities. While there are many proposals to integrate open science into researcher assessment, few of these have been implemented and evaluated. Strong evaluation methods include randomized stepped wedge designs and cluster randomized trials, among other approaches. These methodologies will require teamwork. As these evaluations increase in the coming years, it is likely important to consider how individual efforts can be used to provide a cumulative evidence base of their effects. Such meta-analytical approaches can help provide a ‘living’ view of the cumulative evidence as to the benefits and challenges of using open science as part of researcher assessment.", "appendix": "Data availability\n\nOSF: Open science interventions proposed or implemented to assess researcher impact, https://doi.org/10.17605/OSF.IO/GXY5Z. 63\n\nThis project contains the following underlying data:\n\n• Table 2. Included articles, Table 3. OS interventions proposed/implemented, and Table 4. Cited in Impact Analytics (BMJ)\n\nOSF: Open science interventions proposed or implemented to assess researcher impact, https://doi.org/10.17605/OSF.IO/GXY5Z. 63\n\nThis project contains the following extended data:\n\n• Search strategy\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nMunafo MR, Nosek BA, Bishop DVM, et al.: A manifesto for reproducible science. Nat. Hum. Behav. 2017 Jan 10; 1: 0021. 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Publisher Full Text" }	[ { "id": "231118", "date": "07 Mar 2024", "name": "David Mehler", "expertise": [ "Reviewer Expertise meta-research", "open science", "neuroimaging", "neurology", "mental health", "statistics" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors survey interventions that support open science research practices, addressing a topic of high interest for the meta-research community. The article is clearly structured and concisely written.\n\nI have a few minor suggestions for further improvement:\n\n1. One main intervention are incentive structures. The article would benefit from a systematic overview of available funding options dedicated to open science research practices. This would also be of high practical relevance for the community.\n\n2. To test the effectiveness of interventions, different study designs would be possible, although randomised controlled trials seem the most valid option. However, in the context of open science research practices as a complex behavior, conducting an RCT becomes challenging. Several authors have elaborated on this aspect, it would be informative for the reader if the authors summarise their views.\n\n3. The Review touches on the importance of training, which is another important intervention. Again, a systematic (although likely not exhaustive) overview of training options would be informative and of immidiate practical relevance for readers.\n\n4. On the topic of training, different stakeholders and suggestions to implement training in reproducible open science research practices should be mentioned, e.g. see this recent white paper: Eleven strategies for making reproducible research and open science training the norm at research institutions \| eLife (elifesciences.org)\n5. The authors mention preregistration as an important tool. In this context, the following evaluations should be mentioned:\n\nPreregistration in practice: A comparison of preregistered and non-preregistered studies in psychology \| Behavior Research Methods (springer.com);\nEstimating the prevalence of discrepancies between study registrations and publications: a systematic review and meta-analyses \| BMJ Open\n6. Besides preregistration, Registered Report are a highly relevant open science research practice as they combine transparency with a clear benefit for authors and hance an incentive, a guarantee for publication (The past, present and future of Registered Reports \| Nature Human Behaviour). Recent elaborations and findings on Registered Reports should be incorporated (e.g., Open science challenges, benefits and tips in early career and beyond \| PLOS Biology; An Excess of Positive Results: Comparing the Standard Psychology Literature With Registered Reports - Anne M. Scheel, Mitchell R. M. J. Schijen, Daniël Lakens, 2021 (sagepub.com)).\n\n7. Open science research practices are of high societal relevance, as the authors are pointing out, and their work can inform policy and/or inform funding initiatives that may themselve create new incentives for implementing open science research practices. Looking beyond academia, it may be worth mentioning the role of open science research practices in other science related fields (e.g. industry) with an immidiate impact on health care innovation (e.g., Open science saves lives: lessons from the COVID-19 pandemic \| BMC Medical Research Methodology \| Full Text (biomedcentral.com); S2666-3791(23)00608-0.pdf (cell.com)).\n\n8. Another form of intervention that could be mentioned are guidelines and consensus papers that increasingly demand for open science across different research field. Some examples for different fields could be briefly discussed.\n\n9. Lastly, the article mentions the increasing relevance of the topic, but could also point out more clearly that open science research practices are still only implemented in a fraction of research across fields, highlighting some examples from difference life science areas. This aspect could be integrated in the introduction and referred to again the discussion.\n\nAre the rationale for, and objectives of, the Systematic Review clearly stated? Yes\n\nAre sufficient details of the methods and analysis provided to allow replication by others? Yes\n\nIs the statistical analysis and its interpretation appropriate? Not applicable\n\nAre the conclusions drawn adequately supported by the results presented in the review? Yes", "responses": [] }, { "id": "231125", "date": "09 Apr 2024", "name": "David Mellor", "expertise": [ "Reviewer Expertise Open science", "policy", "meta-science", "ecology", "citizen science" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIn “Open science interventions proposed or implemented to assess researcher impact: a scoping review” the authors conduct an appropriate and thorough scoping review of the lack of evidence or assessment of any policy change in institutional setting to address research assessment for open science, rigor, or reproducibility. I find no fault with their search strategy or their results (with one exception, noted below). However, there is some framing in their introduction and discussion that I believe should be reconsidered. As that framing is not based on any of the facts or literature discussed, it likely can be considered “opinion” and I will do my best to not let that affect my overall recommendation of this work, even if I disagree with it.\nFirst, there is a single conclusion that appears to be omitted or wrong, that should be either corrected or clarified. In the opening lines of the Discussion, the authors note that “Few of the proposals have been evaluated.” As far as I can determine from the summaries of the literature provided, NONE have been evaluated. The closest is this clause in the results: “...piloting and an audit and feedback process for their proposed open science intervention…” Unless I have missed it, I recommend that the authors change their language to state that none have been evaluated, as I, the reader, am left scouring the paper to find the “few” that have been. If I am indeed missing something, I would recommend pulling that those evaluations into a separate section in the results to make it easy to find. As for the framing of the authors’ points, I would ask them to consider an alternative. They are correctly pointing out that research assessment reform has not been evaluated. This is correct, namely because it has not been implemented. They do identify a few academic departments that have made small changes, and those should likely be studied on an ongoing basis. Furthermore, the warning about unintended consequences of any change is not without merit. That being said, the current system has led to the 27 different proposals for reform because of the problems identified in those proposals. It seems that the authors are proposing a Catch-22, where no changed policy can be implemented prior to it being assessed, which of course guarantees that the status-quo will remain firmly entrenched. Importantly, the authors note that “Currently, most researcher assessments are based on traditional quantitative metrics that are less useful (e.g., the number of publications), not evidence-based (e.g., journal impact factors), and do not incorporate other ways of assessing a researcher’s impact (e.g., registering research prior to its initiation).” This is an important note because it emphasizes the status quo is likewise not evidence based. It is not even values based, unless number of publications is a value. The one thing missing from this point is grant funds being used as a current assessment tool. In my opinion, the conclusion of this work should be that proposals for policy reform should include concurrent proposals for ongoing assessment based on 1) the problems that the reform is trying to address and 2) the values that the reformers are trying to support. Before two additional points, the reader should note that I work for the Center for Open Science and actively promote the following two policies. Despite that bias, I believe the following points should still be noted. Finally, in the first paragraph, the authors use an example of the Open Science Badging program as an intervention that did not improve data sharing based on an RCT conducted [8] I do not believe that is a good example to use as a cautionary tale for the purposes of this review, as that RCT did not test the underlying mechanism of the bagding program. That mechanism is that raising social awareness of data sharing would lead to increased data sharing. Authors were merely asked if they would like a badge, without ever having seen an article in that journal with an open science badge. That study was designed based on the premise that the badge itself was the motivator to share data, instead of the social norm of being open. Testing the underlying social pressure would require both a longer period for the trial to occur (eg the original article showing the association between badges and data sharing occurred over twice the amount of time of this RCT, during which little adoption of sharing occurred initially) and would require authors seeing peer adoption of pro-social actions in the articles that they are reading, neither of which occurred in this RCT. Also in this paragraph, the authors note that other signals of transparency are being promoted. Though they do not state anything further, the impression is that it is an ineffective signal and cite a study that indicated that preregistration in the social sciences do not contain sufficient details to constrain “researcher degrees of freedom.” [1] This is of course a correct assessment of that population of preregistrations, but the purpose of preregistration includes being able to assess how much unreported flexibility in data analysis occurred (which was only possible because these studies were preregistered), as well as to assess how much research occurs in a field over a given period of time, also only available if the works are preregistered . There has been far more scholarship on the benefits and limitations of prospective study registration in the clinical sciences, where it has been required by law for over 20 years in the US, and has been commonly enforced internationally for a similar amount of time. Most studies investigating it in that setting likewise see ample room for improvement (notably Ben Goldacre’s work, https://www.alltrials.net/), while also noting the many benefits it provides. Below is a list of some studies that should be included if the authors wish to include prospective study registration as a policy intervention that should be considered as part of this work. Cite prereg evidence: [2] [3] [4] [5] [6] [7]\n\nAre the rationale for, and objectives of, the Systematic Review clearly stated? Partly\n\nAre sufficient details of the methods and analysis provided to allow replication by others? Yes\n\nIs the statistical analysis and its interpretation appropriate? Yes\n\nAre the conclusions drawn adequately supported by the results presented in the review? Partly", "responses": [] }, { "id": "241725", "date": "30 Apr 2024", "name": "Ismael Rafols", "expertise": [ "Reviewer Expertise research evaluation", "science policy", "scientometrics" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis articles aims to review “any open science practice-promoting initiatives proposed or implemented to assess researchers and research impact, in health sciences, biomedicine, psychology, and economics.”\nLet me start by sharing that I am not familiar with the scoping review, following PRISMA, that appears to originate in the evidence synthesis in the health science and which. I work in the fields of research evaluation and scientometrics, which are embedded in social sciences working with mixed-methods – and thus my comments may reflect a disciplinary clash with the methods and framing of the authors, who are based in health sciences and medicine.\nMy overall impression is that the authors want to apply a reviewing methodology that may work in terms of finding evidence for issues where there are hard facts, but that is problematic for issues that are not well-defined. In our case, we are dealing with studies dealing with ‘initiatives’ that assess researchers and research impact while fostering Open Science in relation to transparency, integrity and data sharing. We should realise that Open Science is defined in different ways (the UNESCO definition mentioned in the text is very broad and includes values and principles), and that evaluation and impact assessment are also carried out in many ways under different names.\nIt appears that 27 publications found in relevant in the review include generic proposals such the San Francisco Declaration on Research Assessment (DORA), The Leiden Manifesto for research metrics, The Metric Tide, and the Hong Kong Principles for Assessing Researchers, but also include general opinion pieces (Schmid, 2017)) and specific proposals for indicators that would favour open science (Bradshaw et al., 2021).\nAs I said, I don’t understand the scoping review methodology (and the exclusion of so many papers), but given the papers included above, I can think of many, many more publications which dealt with similar and related issues proposals for improvement. Just taking the journals ‘Research Evaluation’, ‘Scientometrics’, ‘Research Policy’ and ‘Quantitative Science Studies’ one can find dozens of articles dealing with the topics of researcher assessment, with direct implications in terms of integrity, transparency, reproducibility, collaborations, gender-bias, topic breadth or bias (e.g. due to the problems with use of narrow and proprietary databases (Singh et al., 2021)) or societal impact assessment (an issue highly discussed in the last 10 years (Smit & Hessels, 2021) that does not seem to appear in the 27 papers). Most of these proposals are not sector specific.\nIn short, I have the impression that this scoping review does not capture the breadth of literature that is making studies on research assessment and proposals about it which are directly relevant to Open Science, although possibly not mentioning OS. It captures publications of different types (a proposal on indicators, an opinion piece, the key documents like DORA), with gaps and unclear logics on why some are included and some not.\nWhile I support the motivations of the authors in seeking better ways for assessing research to foster OS, I am worried about the apparent large gaps in the review.\nSpecific issues:\nTitle: in reading the title, I had expected it would focus on research impact assessment, meaning the assessment of the societal contribution of researchers (possibly because I am familiar with the UK evaluation system, where impact is assumed to be about society). However, I see that is is about assessing the contributions of researchers, with a focus on sharing and reproducibility. I would suggest making the title more specific to avoid mis-interpretation. Notice that this is a subset of the OS activities as defined by UNESCO. Pilars 3 and 4 of the UNESCO OS definition include participatory processes and dialogue with other knowledge systems. Therefore, I understand that the assessments of impacts in terms of interaction between researchers and beneficiaries should also concern OS. Field selection: The authors state that “We included studies from the health sciences, biomedicine, psychology, and economics fields, as our impression is that open science developments are more prevalent in these disciplines, which are also closest to our knowledge and experience.”. These are fields that deal with complex problems with quantitative techniques and may have more practices on data sharing, or integrity in methods. However, some aspects of OS (as defined by UNESCO) such as participatory research may be more prevalent in other disciplines such as sociology or applied social sciences. Search strategy: The search strategy, as reported in https://osf.io/trfq8 is so complicated that it is very difficult to follow. For an issue such as Open Science that is contested and there are plural understandings (Fecher & Friesike, 2014), this is very problematic, as it is important to convey to the reader what the authors are actually looking at. Screening strategy: I am not familiar with the Continuous Active Learning® (CAL®) tool. I am quite surprised that after screening you ended up with only 208 publications out of the 18,000 – and then only 27 are found relevant. Could you give examples in the text of why so many where excluded? Supplementary materials: Tables 3 and 4 in the underlying data are mentioned, but I could not find them in the supplementary files at https://osf.io/gxy5z.\n\nExamples of missing literature:\nAdam, P., Ovseiko, P.V., Grant, J. et al. ISRIA statement: ten-point guidelines for an effective process of research impact assessment. Health Res Policy Sys 16, 8 (2018). https://doi-org.ezproxy.leidenuniv.nl/10.1186/s12961-018-0281-5 Dahler-Larsen, P. (2018). Making citations of publications in languages other than English visible: On the feasibility of a PLOTE-index. Research Evaluation, 27(3), 212–221. Hormia-Poutanen, Kristiina, Enst Kristiansen, Rebecca Lawrence, Sabina Leonelli, Natalia Manola, Eva Méndez, Christopher Rossel, Michela Vignoli, and Marta Dias Agostinho. 2017. “Recommendations of the OSPP on Next-Generation Metrics.” Almetrics Workng Group of the Open Science Policy Platform. Larivière, V., Kiermer, V., MacCallum, C. J., McNutt, M., Patterson, M., Pulverer, B., ... & Curry, S. (2016). A simple proposal for the publication of journal citation distributions. BioRxiv, 062109. Wilsdon, James, Judit Bar-ilan, Robert Frodeman, Elisabeth Lex, Isabella Peters, and Paul Wouters. 2017. Next-Generation Metrics: Reponsible Metrics and Evaluation for Open Science. Brussels, Belgium: European Commission. https://doi.org/10.2777/337729. Wouters, P., Ràfols, I., Oancea, A., Kamerlin, L., Holbrook, B., & Jacob, M. (2019). Indicator frameworks for fostering open knowledge practices in science and scholarship. European Commission. https://doi.org/10.2777/445286\nReferences\nBradshaw, C. J. A., Chalker, J. M., Crabtree, S. A., Eijkelkamp, B. A., Long, J. A., Smith, J. R., Trinajstic, K., & Weisbecker, V. (2021). A fairer way to compare researchers at any career stage and in any discipline using open-access citation data. PLOS ONE, 16(9), e0257141. https://doi.org/10.1371/journal.pone.0257141 Fecher, B., & Friesike, S. (2014). Open Science: One term, five schools of thought. In S. Bartling & S. Friesike (Eds.), Opening Science. Springer International Publishing. https://doi.org/10.1007/978-3-319-00026-8 Schmid, S. L. (2017). Five years post-DORA: Promoting best practices for research assessment. Molecular Biology of the Cell, 28(22), 2941–2944. https://doi.org/10.1091/mbc.e17-08-0534 Singh, V. K., Singh, P., Karmakar, M., Leta, J., & Mayr, P. (2021). The journal coverage of Web of Science, Scopus and Dimensions: A comparative analysis. Scientometrics, 126(6), 5113–5142. https://doi.org/10.1007/s11192-021-03948-5 Smit, J. P., & Hessels, L. K. (2021). The production of scientific and societal value in research evaluation: A review of societal impact assessment methods. Research Evaluation, 30(3), 323–335. https://doi.org/10.1093/reseval/rvab002\n\nAre the rationale for, and objectives of, the Systematic Review clearly stated? Yes\n\nAre sufficient details of the methods and analysis provided to allow replication by others? Partly\n\nIs the statistical analysis and its interpretation appropriate? Not applicable\n\nAre the conclusions drawn adequately supported by the results presented in the review? Partly\n\nIf this is a Living Systematic Review, is the ‘living’ method appropriate and is the search schedule clearly defined and justified? (‘Living Systematic Review’ or a variation of this term should be included in the title.) Not applicable", "responses": [] } ]	1	https://f1000research.com/articles/12-1396
https://f1000research.com/articles/12-1395/v1	23 Oct 23	{ "type": "Research Article", "title": "Exploring the role of ChatGPT in rapid intervention text development", "authors": [ "Hannah Bowers", "Cynthia Ochieng", "Sarah E Bennett", "Sarah Denford", "Milly Johnston", "Lucy Yardley", "Cynthia Ochieng", "Sarah E Bennett", "Sarah Denford", "Milly Johnston", "Lucy Yardley" ], "abstract": "Background There have been successful applications of AI to answering health-related questions, which suggests a potential role for AI in assisting with development of intervention text. This paper explores how ChatGPT might be used to support the rapid development of intervention text.\n\nMethods Three case studies are presented. In the first case study, ChatGPT (using GPT-4) was asked to generate sleep advice for adolescents. In case study two, ChatGPT (using GPT-3) was asked to optimise advice for people experiencing homelessness on staying hydrated in extreme heat. Case study three asked ChatGPT using GPT-3 and GPT-4 to optimise an information sheet for participation in a study developing an intervention for maternal blood pressure. Outputs were evaluated by the researchers who developed the text, and in case studies two and three were shown to public and patient contributors for feedback.\n\nResults ChatGPT was able to generate informative advice about sleep in case study one and was able to accurately summarise information in case studies two and three. In all three cases, elements or aspects were omitted that were included in the researcher-generated text that was based on behaviour change theory, evidence and input from public and patient contributors. However, in case study three, feedback from public contributors suggested ChatGPTs outputs were preferred to the original, although the outputs omitted information and were not at the requested accessible reading level.\n\nConclusions ChatGPT was able to accurately generate and summarise health information. However, this information typically excluded core behaviour change techniques and was sometimes inappropriate for the target users. There is likely to be a valuable role for generative AI in the intervention development process, but this will need to be combined with detailed scrutiny and input from researchers and public contributors.", "keywords": [ "ChatGPT", "Intervention development", "AI", "behaviour change" ], "content": "Introduction\n\nChatGPT is an artificial intelligence (AI) chatbot, launched by OpenAI in November 2022. It is capable of generating responses to prompts or questions, input by the user, based on OpenAI’s Generative Pre-Trained Transformer (GPT) language model (launched in 2018) (chat.openai.com). Since its launch it has gained significant attention and discussion of the ways in which it might be used across various sectors.\n\nIt has been suggested that generative AI, such as ChatGPT, can be used in the generation of content with the purpose of providing information on public health issues, answering questions about health promotion and disease prevention strategies, explaining the role of community health workers and health educators, discussing the impact of social and environmental factors on community health, and providing information about community health programs and services (Biswas, 2023).\n\nThe potential of generative AI to support healthcare has been investigated in a range of topic areas. In a review of 60 papers, there was evidence that ChatGPT could be beneficial in supporting scientific writing, research activity (e.g. analysis of large datasets), medical education and healthcare practice (Sallam, 2023). With regards to writing, there were reported benefits in the speed of conducting literature reviews and improved language to communicate ideas, providing information that is easily accessible and understandable. In another systematic review of 31 studies, ChatGPT was effective at generating information on a range of health topics (Muftić et al., 2023). However both reviews highlight potential limitations such as generation of inaccurate information, and limited knowledge on topics or research after 2021. In some cases ChatGPT produced overly detailed content, but this may be addressed by using clear and specific prompts from the user about the desired output (Sallam, 2023).\n\nThere is mixed evidence on the quality of health-related information generated by generative AI (with the majority of research using ChatGPT). There is some evidence that AI-generated responses from ChatGPT are outperforming human responses in terms of their quality and empathy. Ayers et al. (2023) asked licensed healthcare professionals to evaluate physician responses and AI responses to health questions from a public social media forum. Responses were rated in terms of the quality of the information and the empathy provided. AI responses were preferred 78.6% of the time and were rated significantly better than physician responses with regards to quality and empathy, suggesting AI may be useful in generating public-facing health information. Grünebaum et al. (2023) reported a high level of accuracy in answering questions about obstetrics and gynaecology, while Barat et al. (2023) report only a 40% accuracy rating for responses to questions about interventional radiology.\n\nFewer studies have explored the utility in generative AI summarising and optimising text. ChatGPT was effective in summarising text based on palliative critical care scenarios written by human experts (Almazyad et al., 2023). Cascella et al. (2023) argue that while ChatGPT can generate and summarise public health text effectively, this is dependent on the information the researcher inputs, and it did not always adhere to strict word limits.\n\nThe Coronavirus disease 2019 (COVID-19) pandemic has highlighted the need to extend existing best practice in intervention development so that interventions can be co-developed and evaluated quickly in response to a rapidly-changing public health concern (Yardley et al., 2023). In an effort to address this issue, the Agile Co-Production and Evaluation (ACE) Framework was proposed (Yardley et al., 2023). The framework is intended to provide a focus for investigating new ways of rapidly developing effective interventions by combining co-production methods with large-scale testing and evaluation. This framework builds upon the Person-Based Approach (PBA) to intervention development (Yardley et al., 2015). This approach to intervention development is grounded in the idea that co-production with an intervention’s intended users is vital in ensuring the techniques and messaging are appropriate and more likely to be effective.\n\nIn this paper, we consider and explore the potential of generative AI (using ChatGPT) in supporting the rapid development and optimisation of interventions. We present three case studies in which ChatGPT was used to support intervention development. This includes using ChatGPT to generate information for a health intervention, in order to evaluate its ability to rapidly produce intervention materials. Following this, we explore the ability of ChatGPT to optimise text that was generated by researchers involved in intervention development. In these two latter case studies, we asked patient and public involvement (PPI) contributors for their views on the outputs generated by ChatGPT.\n\n\nMethods\n\nIn a series of case studies, we asked ChatGPT to generate or simplify text relating to health interventions. Text to be generated or simplified for these case studies was selected from studies being undertaken by the Behavioural Interventions Group. These studies were in the process of developing public-facing intervention content using the PBA or ACE (Yardley et al., 2015, 2023). This approach to intervention development ensures the developed intervention is grounded in theory, evidence and views of the intended users. It does this through the use of theoretical mapping (i.e. mapping techniques and ideas against behaviour change and psychological theories), ‘guiding principles’ for how the intervention should be developed, and a logic model to hypothesise how the intervention may work.\n\nIn the first case study, ChatGPT was asked to generate intervention content to improve sleep in teenagers (‘Sleep Solved’). We constructed a prompt for ChatGPT: “How would you improve sleep in teenagers? Write your answer in simple language for someone with a reading age of 9 and use bullet points.” We then evaluated the effects of providing a more elaborate request, to encourage ChatGPT to use the ‘programme theory’ (Skivington et al., 2021) for the intervention to guide the text development in the same way as the researchers were doing. The elements of the programme theory that ChatGPT was additionally asked to follow were the ‘guiding principles’ (Pollet et al., 2020): to avoid being patronising or paternalistic; to use question-answer formats to make the advice accessible and engaging; and to make the advice convincing by giving the scientific explanations for advice, but keeping the scientific explanations very brief and simple. In accordance with the behavioural theory informing the content of Sleep Solved, ChatGPT was also asked to base the advice on Bandura’s Social Cognitive Theory (Bandura, 1977), with the aim of building users’ self-efficacy (confidence) that they can follow the advice, and positive outcome expectations regarding the consequences of following the advice.\n\nIn the remaining case studies, ChatGPT was asked to optimise text. In case study two, the text was written by a behavioural scientist from the Behavioural Interventions Group who was involved in the development of an intervention to support people experiencing homelessness to stay safe during extreme weather events. In case study three, the text was from an information sheet used in the recruitment of study participants in research conducted by the Behavioural Interventions Group, aiming to co-produce an intervention for post-partum blood pressure management. This text was produced by a study team outside of the Behavioural Interventions Group.\n\nIn order to maintain consistency for comparison across the case studies, ChatGPT was provided with the same wording in all three case studies to request accessible text: “make this text suitable for a reading age of 9 and use bullet points”. Each identically-worded request (with different example text to be simplified) was conducted twice, each output compared, and the best output was selected by the researcher.\n\nOpenAI has recently upgraded ChatGPT by using the GPT-4 model. This requires a paid subscription to ChatGPT Plus. Case study two used ChatGPT using GPT-3, whereas case study one and case study three were conducted in ChatGPT using both GPT-3 and GPT-4. In these two case studies, identical prompts were input into the two versions of ChatGPT (i.e. ChatGPT using GPT-3 and ChatGPT using GPT-4) and outputs from each version of ChatGPT were compared.\n\nThe outputs from all case studies were evaluated by the researchers developing the intervention text through reflecting on the output compared with the guiding principles, planning tables and text written by the researcher. In case studies two and three feedback was also sought from PPI members of the team and service providers. PPI members were invited to comment on the text produced by ChatGPT as part of their ongoing role within the research teams developing the interventions. Ethical approval is not required for the involvement of public and patient contributors in research.\n\n\nResults\n\nAs part of a separate study, members of the Behavioural Interventions Group were in the process of developing Sleep Solved, an app designed to improve sleep in teenagers. In the current case study, we explored whether ChatGPT could generate simple advice to be used within this app. Text generated by ChatGPT using GPT-3 in response to our initial, simple request was compared to text generated using ChatGPT using GPT-4 (see ‘Case study one data and review of ChatGPT output 05.09.2023.docx’ (Bowers et al., 2023)) in response to our theory-based request. Both GPT models provided useful advice that mapped onto many elements of the researcher-generated text in the Sleep Solved app. For example, advice to wake up at the same time each day, to create a sleep-friendly environment and avoid phone use before bed were all directly comparable to Sleep Solved (see ‘Case study one data and review of ChatGPT output 05.09.2023.docx’ (Bowers et al., 2023)). The text using ChatGPT using GPT-4 in response to the theory-based request offered more detail, and some reasoning for why the advice was offered. For example, for the ‘sleep-friendly advice’ section, ChatGPT using GPT-3 gave a list of ideas for making the bedroom more calming, whereas ChatGPT on GPT-4 explained why these were helpful.\n\nDespite the considerable overlap in advice content, there were aspects of ChatGPT’s text that did not map onto the intervention guiding principles or logic model (Yardley et al., 2015) (see ‘Case study one data and review of ChatGPT output 05.09.2023.docx’ (Bowers et al., 2023)). The logic model in this PBA intervention was based on Bandura’s Social Cognitive Theory (Bandura, 1998). ChatGPT was asked to define this theory, and was able to do this. Despite this, when ChatGPT was asked to draw on this theory, there was no evidence in the output that it had done so. While ChatGPT did give reasons for some of the advice it provided when asked to do so, the rationale it gave was less in-depth than in Sleep Solved. For example, the ChatGPT rationale for maintaining a regular sleep routine was ‘A regular sleep routine helps regulate your body’s internal clock, making it easier to fall asleep and wake up’. Sleep Solved was designed to explain the neuroscience behind the recommended sleep behaviours in an accessible way, and therefore described how cortisol affects sleep and is affected by the sleep cycle, light and caffeine. For example, “You have a get up and go hormone – cortisol- which helps you feel alert. Getting up at the same time every day trails your brain to release cortisol at the right time.”, and, “… If you sleep late, your cortisol levels peak late. This may make you feel tired and sleepy when you have to get up earlier. It might also make you feel hungry, worried or low.”, “Try to avoid bright lights such as phone screens. Bright light can make your brain think it is time to wake up!”. ChatGPT was asked to follow the research team’s guiding principles and logic model by using scientific explanations to convince young people of the positive outcomes of recommended behaviours rather than undermining their autonomy by providing paternalistic advice. ChatGPT often did not follow these instructions, for example simply advising ‘Watch what you eat before bed; Avoid heavy or spicy meals close to bedtime’ – whereas Sleep Solved gave scientific explanations for this advice; “Eating foods high in sugar close to bedtime causes your brain to release hormones that wake you up, like cortisol and adrenaline. Spicy and fatty foods are harder to digest which can keep you awake too”. Sleep Solved also provided examples of alternative snacks. In addition, Sleep Solved has several functions to improve users self-efficacy, which do not feature in ChatGPT’s advice (these elements are illustrated further in ‘Case study one data and review of ChatGPT output 05.09.2023.docx’ (Bowers et al., 2023)).\n\nWhile much of the advice from ChatGPT was relevant to sleep hygiene recommendations for young people, the ChatGPT text did not apply the behaviour change techniques employed in a co-produced intervention. Based on this, the decision was made to explore the ability of ChatGPT to simplify complex language in further case studies, making intervention text simple, accessible and engaging for people from diverse backgrounds.\n\nIntervention development\n\nPeople experiencing homelessness (PEH) are considered to be at particularly high risk during extreme weather events, and even more so if they are using drugs and alcohol. Therefore, there is a need to co-create interventions (including advice) to help PEH stay safe during adverse weather events. This case study describes the findings of developing one component of an intervention that focuses on supporting people to keep hydrated during extreme heat.\n\nThe initial message was written by a behavioural scientist with expertise in intervention development as part of a separate ongoing study (UK Health Security Agency, 2023) (see ‘Case study two data.docx’ (Bowers et al., 2023)). The content of the message was informed by interviews with PEH, people delivering services to PEH, a review of the literature, and PPI/stakeholder feedback (using the PBA to intervention development (Yardley et al., 2015)). The subsequent message focused on providing information for PEH about the importance of keeping hydrated with non-alcoholic beverages during extreme heat, recognising signs and symptoms of dehydration and heat stroke, encouraging PEH to identify where and when they can access water, and prompting people to carry water bottles to ensure hydration. As many PEH find it difficult to carry equipment such as water bottles, we included content acknowledging this and encouraged people to attempt to carry water for short durations only.\n\nChatGPT outputs\n\nWe asked ChatGPT (using GPT-3) to make the text suitable for a reading age of nine and use bullet points (see ‘Case study two data.docx’ (Bowers et al., 2023)). In response, ChatGPT made the following notable changes. First, the AI-generated text was much shorter (150 words compared with the 300-word original). A definition of dehydration was introduced, but a lot of other detail was removed. In particular, the specific detail about the symptoms of dehydration, and information about symptoms that indicate when medical attention is needed was not included. The sentences encouraging people to find locations in which water is freely available, and carrying bottles only during heatwaves were also removed. In addition to reducing content, ChatGPT modified a key message about reducing alcohol consumption during extreme heat to abstaining from alcohol consumption during extreme weather.\n\nPPI feedback\n\nThe two versions of advice were then presented to a PPI team comprising two PEH (one male aged over 50 and one female aged 39, both experiencing street homelessness) and a service provider for PEH. A third version of the message was then co-produced (see ‘Case study two data.docx’ (Bowers et al., 2023)). The brevity of the ChatGPT version was viewed positively, but the PPI team also thought that some of the specific detail that had been removed in the shorter version needed to be included. For example, as PEH reported experiencing symptoms similar to dehydration on a regular basis, the inclusion of specific information about symptoms indicating a need for medical attention was considered highly important. The focus on abstinence rather than reduction of alcohol was considered inappropriate and unrealistic. Other feedback focused on information that required modification or was missing from both versions. For example, people thought it important to state that alcohol makes you more dehydrated at the start of the message. The PPI team wanted information about what would happen if they did not get medical attention, and they wanted a map showing the locations where water is available. The PPI team also highlighted the need for an additional, complimentary intervention for service providers, in which detail is provided about how best to support PEH during extreme heat.\n\nIntervention development\n\nHigh blood pressure during pregnancy can result in elevated cardio-vascular risks over time. Post-pregnancy, blood pressure can change rapidly, often requiring alterations in the patient’s medication. In this case study, an intervention was being developed to aid patients to manage their blood pressure post-partum through regular self-checks and pre-planned medication alterations. An information sheet inviting patients and former patients to participate in the development phase of the study was drafted by a team of researchers and refined with other stakeholders including clinicians and two PPI members. This information sheet explained to potential participants that the study had developed an intervention to help patients manage their blood pressure after pregnancy. The information sheet further explained that the patients were being invited because of their experience with high blood pressure in pregnancy and that they would be asked to share their thoughts on the intervention developed. They were also reminded that their participation was voluntary and that if they agreed to participate, they would need to sign a consent form. We considered that the information sheet would require considerable further optimisation to make it more accessible to people with lower levels of literacy, and was therefore a suitable text to ask ChatGPT to simplify.\n\nChatGPT outputs\n\nChat GPT was utilised to attempt to optimise the patient information sheet for the study. Initially all the text from the information sheet (four pages) was pasted into ChatGPT using GPT-3. The response it generated was a short paragraph that was too brief to fully explain the study. ChatGPT (using GPT-3) also did not make the language in the sheet simpler and did not produce bullet points in successive iterations of the request (see ‘Case study three data.docx’ (Bowers et al., 2023)).\n\nThe researcher then divided the information sheet into smaller sections (between 94-361 words long) and each section was entered into ChatGPT (using GPT-3) separately with the standard instruction to “make the text suitable for a reading age of 9 and use bullet points”. In response to this, ChatGPT organised its responses in bullet points but again the reading level was far more complex than for a nine year old (Flesch-Kincaid grade level 10.9 i.e. 16-17 year old).\n\nSince ChatGPT appeared to require short chunks of text, two sections of the information sheet (‘Introduction to the Study’ and ‘Data Protection’ sections) were copied into ChatGPT (using GPT-3) separately with the standard instruction i.e. to make the text suitable for a reading age of nine and use bullet points. The requests were repeated to compare iterations. When the text retained its original formatting (such as bullet points and line breaks), ChatGPT often did not generate responses with bullet points. However, when the text was pasted as simple unformatted text, it generated bullet points.\n\nThe text describing an introduction to the study was then entered into ChatGPT (using GPT-4) with the same prompt (“make the text suitable for a reading age of 9 and use bullet points”) to compare the two versions. This was done twice and the best response (as rated by the researcher) used for comparison with the outputs from ChatGPT using GPT-3. GPT-4 had better results than GPT-3 with the first iteration being for a reading age 10-11 (Flesch-Kincaid grade level 5.5) and the output contained all the information. The second iteration on GPT-4 was for a reading age of 9-10 (Flesch-Kincaid grade level 4.9).\n\nChatGPT appeared to shorten text consistently, which could be useful for generating key points - particularly for more general subjects like data protection in research. ChatGPT using GPT-3 was not successful at making the text simple enough for a reading age of 9; the study introduction text was usually generated for a reading age of 15-17 while the data protection text generated was simpler at reading age 13-14 (Kincaid Flesch grade level 8.8). Additionally, it was not able to handle longer documents and the formatting had to be removed. ChatGPT using GPT-4 had better results with regards to the reading level and the information included.\n\nPPI feedback\n\nThe original text from a participant information sheet and two simplified versions (one from ChatGPT using GPT-3 and one from ChatGPT using GPT-4) were shown to the study’s PPI members (demographic characteristics are presented in Table 1). They found a clear difference between the original wording of the information and the simplified ChatGPT texts. All preferred the output from ChatGPT using GPT-3. Two PPI members sought clarification on an acronym that had been used to name the study regardless of whether the text had explained it. One PPI member said she preferred the output from ChatGPT using GPT-3, which she found to be the easiest to understand. Another PPI member found this same output easiest to read. She said she did not like having too many words on a leaflet and ChatGPT on GPT-3 offered the least words. All the PPI members agreed that the original text had too many words and technical terms and that ChatGPT using GPT-4 was better in that regard. Despite agreeing with the others, one PPI member explained that she preferred the simple headings (in question form) in the version from ChatGPT using GPT-4, but that overall she was happy with the version from ChatGPT using GPT-3.\n\nSome other stakeholders (comprising a general physician (GP), obstetrician fellow, midwife and two researchers) were also shown the original text and asked to compare it with the ChatGPT versions. They all preferred the simplified versions from both ChatGPT using GPT-3 and ChatGPT using GPT-4. The bullet points in the ChatGPT versions were thought to make the text easier to read. The team also noticed that the language on the ChatGPT versions was simpler and hence more accessible than in the original text.\n\n\nDiscussion\n\nAcross three case studies, we have explored how ChatGPT might be used to support rapid intervention development. In our first case study, when prompted to produce advice on sleep for adolescents, ChatGPT’s output undoubtedly had face validity and value; the advice it gave reflected scientific consensus and was succinct and accessible. Indeed, a fourth case study that was planned for this paper was not pursued because the intervention was being developed alongside rather than before this study, and the developer found the text suggested by ChatGPT so helpful that she was unable to create a version that was not influenced by it. However, the ChatGPT text did not closely follow the ‘guiding principles’ for how to make the text engaging that it was asked to apply, and did not include any of the key behaviour change techniques that were developed as part of the co-production process using the PBA. In our second case study, key pieces of advice that were generated as a result of co-production with intended users of the intervention were omitted. In this case study, members of the public (specifically, people experiencing homelessness) provided feedback on ChatGPT’s output and highlighted that these omissions needed to be included in the advice. Our third case study showed that, when prompted to optimise an entire participant information sheet, ChaptGPT (using GPT-3) omitted information that would be necessary to include and the reading level of the text was higher than requested by the prompt. However, when text was entered in smaller chunks without bullet points, the output was improved and the text was considered superior to the original by both PPI and the research team.\n\nWe have learned from our case studies that despite the speed with which ChatGPT can provide text, using ChatGPT can be surprisingly time-consuming. Texts requiring optimisation may need preparation prior to being input into ChatGPT (using GPT-4), for example by dividing into short simple sections and simplifying technical terms. ChatGPT using GPT-4 seemed to perform better, and outputs could be improved further by responding to ChatGPT’s outputs with further requests for changes and by giving examples of the required tone and suitable outputs. For instance, generative AI (including ChatGPT) used in health intervention messaging could be prompted to simplify commonly used terms and explain common concepts (e.g. randomisation, anonymisation or informed consent). We faced a steep learning curve in our use of ChatGPT and it is likely that in future researchers will also need to be trained to use generative AI more effectively. Although beyond the scope of this study, it is likely that generative AI can contribute to public health intervention development in other ways. For example, it could be used to rapidly summarise and respond to public reactions to an emergency on social media, and to translate intervention content into multiple languages.\n\nDespite the potential of generative AI to support rapid response during public health emergencies, in its current form, there are significant limitations (see Table 2). Importantly, there is evidence that ChatGPT has problems with accuracy in generating public health information (Biswas, 2023; Jungwirth & Haluza, 2023; Muftić et al., 2023; Sallam, 2023). Furthermore, the use of ChatGPT for arising public health issues is limited due to its language model being trained using a dataset with a cut-off date in 2021 (Jungwirth & Haluza, 2023; Muftić et al., 2023; Sallam, 2023) meaning it is not able to provide up-to-date information about novel public health concerns. In contrast to previous studies (e.g. Muftić et al., 2023; Sallam, 2023), in our three case studies there were no examples of inaccuracies in the text generated by ChatGPT. However the quality of the output (with regards to formatting and language used) was related to the quality of the prompt (Cascella et al., 2023).\n\n\n\n• All AI-generated advice must be checked by topic experts to ensure that it is correct\n\n• If the advice is intended to produce behaviour change then input from behavioural scientists may be needed to add behaviour change techniques to support implementation of the advice\n\n• Because AI-generated advice is based on widely available text, additional design input may be needed to generate text and images that is sufficiently novel and interesting to engage users\n\n• Because AI learns from widely available text, it may struggle with rarer research or topics\n\n\n\n• AI-generated text may not be sufficiently sensitive to the contexts of seldom heard groups. Co-production with all intended user groups is therefore essential to ensure that text is appropriate for them\n\n\n\n• Both the AI and the researchers are likely to require training and significant input of time to create appropriate, engaging content\n\n• Time is also required to check the accuracy and validity of AI generated text, which may not be accurate\n\n\n\n• AI may struggle with theoretical concepts and abstract principles. While it can provide a definition, the algorithm is not currently advanced enough to incorporate theory into the content it produces\n\n\nConclusions\n\nChatGPT (particularly ChatGPT using GPT-4) can produce succinct summaries of text, when this text is prepared by a researcher and entered into ChatGPT in the appropriate format with appropriate context and requests. However, ChatGPT tends to miss key elements that address subtle but important barriers to behaviour change identified through collaboration with intended users and grounded in theory and evidence. While ChatGPT is able to rapidly summarise text to be informative and succinct, it may omit key content grounded in behaviour change theory, which may affect its ability to change behaviour (Yardley et al., 2015). A further problem with AI-generated text is that interventions need to offer users advice and support that seems somewhat novel or unique in order to promote engagement. Because interventions produced using generative AI are based on widely available texts they are unlikely to appear innovative and uniquely informative. Moreover, since the outputs from generative AI are inevitably generalist rather than context-specific there is a risk that they will not be well-adapted to the context of under-represented groups. The outputs of generative AI may thus be able to assist with ‘agile’ content generation but the ‘co-production’ element of the ACE framework will remain crucial to ensure that content is ‘relatable’ and appropriate for the intended users (Yardley et al., 2023).\n\nIn summary, generative AI is a novel tool that may be useful in rapid intervention development. There is however a learning curve and as we are learning how to use it; it too is learning how best to respond.", "appendix": "Data availability\n\nRepository name: FigShare.\n\nProject title: Input and output for Exploring the role of ChatGPT in rapid intervention text development, DOI https://doi.org/10.6084/m9.figshare.24084399.v1 (Bowers et al., 2023)\n\nThis project contains the following underlying data:\n\n- Case study one data and review of ChatGPT output 05.09.2023.docx (ChatGPT outputs from Case Study One)\n\n- Case study two data.docx (ChatGPT outputs from Case Study Two)\n\n- Case study three data.docx (ChatGPT outputs from Case Study Three)\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nWe would like to thank Mr Joe Shervell, Sabroso Ltd., for providing access to ChatGPT Plus and advising on the use of ChatGPT. We would also like to thank the PPI contributors for their comments in case studies two and three. LY is an NIHR Senior Investigator and her research programme is partly supported by NIHR Applied Research Collaboration (ARC)-West and NIHR Health Protection Research Unit (HPRU) for Behavioural Science and Evaluation.\n\n\nReferences\n\nAlmazyad M, Aljofan F, Abouammoh NA, et al.: Enhancing Expert Panel Discussions in Pediatric Palliative Care: Innovative Scenario Development and Summarization With ChatGPT-4. Cureus. 2023; 15: e38249. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyers JW, Poliak A, Dredze M, et al.: Comparing Physician and Artificial Intelligence Chatbot Responses to Patient Questions Posted to a Public Social Media Forum. JAMA Intern. Med. 2023; 183: 589–596. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBandura A: Self-efficacy: Toward a unifying theory of behavioral change. Psychol. Rev. 1977; 84(2): 191–215. PubMed Abstract \| Publisher Full Text\n\nBandura A: Health Promotion from the Perspective of Social Cognitive Theory.1998.\n\nBarat M, Soyer P, Dohan A: Appropriateness of Recommendations Provided by ChatGPT to Interventional Radiologists. Can. Assoc. Radiol. J. 2023; 084653712311701. PubMed Abstract \| Publisher Full Text\n\nBiswas SS: Role of Chat GPT in Public Health. Annals of Biomedical Engineering. Springer; 2023. Publisher Full Text\n\nBowers H, Ochieng C, Bennett SE, et al.: Input and output for Exploring the role of ChatGPT in rapid intervention text development. figshare. Dataset. 2023. Publisher Full Text\n\nCascella M, Montomoli J, Bellini V, et al.: Evaluating the Feasibility of ChatGPT in Healthcare: An Analysis of Multiple Clinical and Research Scenarios. J. Med. Syst. 2023; 47(1): 33. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGrünebaum A, Chervenak J, Pollet SL, et al.: The Exciting Potential for ChatGPT in Obstetrics and Gynecology. Am. J. Obstet. Gynecol. 2023; 228: 696–705. PubMed Abstract \| Publisher Full Text\n\nJungwirth D, Haluza D: Artificial Intelligence and Public Health: An Exploratory Study. Int. J. Environ. Res. Public Health. 2023; 20(5). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMuftić F, Kadunić M, Mušinbegović A, et al.: Southeast Europe Journal of Soft Computing Exploring Medical Breakthroughs: A Systematic Review of ChatGPT Applications in Healthcare.2023; 12(1).\n\nPollet S, Denison-Day J, Bradbury K, et al.: A Qualitative Exploration of Perceptions of a Digital Intervention to Promote Physical Activity in Older Adults. J. Aging Phys. Act. 2020; 29(3): 442–454. Publisher Full Text\n\nSallam M: ChatGPT Utility in Healthcare Education, Research, and Practice: Systematic Review on the Promising Perspectives and Valid Concerns. Healthcare (Basel, Switzerland). 2023; 11(6). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSkivington K, Matthews L, Simpson SA, et al.: A new framework for developing and evaluating complex interventions: update of Medical Research Council guidance. BMJ. 2021; 374. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nUK Health Security Agency: Supporting vulnerable people before and during hot weather: healthcare professionals - GOV.UK.2023. Reference Source\n\nYardley L, Denford S, Kamal A, et al.: The Agile Co-production and Evaluation (ACE) framework for developing public health interventions, messaging and guidance.2023. Reference Source\n\nYardley L, Morrison L, Bradbury K, et al.: The person-based approach to intervention development: Application to digital health-related behavior change interventions. J. Med. Internet Res. 2015; 17(1): e30. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "230781", "date": "05 Mar 2024", "name": "Kadir Uludag", "expertise": [ "Reviewer Expertise chatbots，psychology，psychiatry" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\n1-)You may add more keywords related to the study.\n2-) You may add the following; Since its launch it has gained significant attention and discussion of the ways in which it might be used across various sectors such as psychology (Uludag, 2023) and pyschiatry (Pham et al., 2022).\nreference:[1] [2]\n\n3-) you can add following recommendation： In future research, the potential for exploring chatbot-based virtual dietitian advice, similar to the previous virtual dietitian application, can be examined (Garcia et al., 2022). (PDF) Virtual Dietitian as a Precision Nutrition Application for Gym and Fitness Enthusiasts: A Quality Improvement Initiative (researchgate.net). IEEE Xplore - Conference Table of Contents\n4-)please be sure that following Banduras social cognitive theory can really make difference. Bandura’s Social Cognitive Theory (Bandura, 1977), if it does not make any difference, you can remove it. it is important that if other people cannot find any difference then, they can assume that its a methodologic flaw in your study. 5-)you can give more information about the team: This text was produced by a study team outside of the Behavioural Interventions Group. 6-）Please provide a more detailed explanation of the concept of \"time consuming.\" Did you measure and analyze the amount of time individuals spent on the task?\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1395
https://f1000research.com/articles/12-1393/v1	23 Oct 23	{ "type": "Review", "title": "ChatGPT impacts in programming education: A recent literature overview that debates ChatGPT responses", "authors": [ "Christos-Nikolaos Anagnostopoulos" ], "abstract": "This paper aims at a brief overview of the main impact of ChatGPT in the scientific field of programming and learning/education in computer science. It lists, covers and documents from the literature the major issues that have been identified for this topic, such as applications, advantages and limitations, ethical issues raised. Answers to the above questions were solicited from ChatGPT itself, the responses were collected, and then recent literature was surveyed to determine whether or not the responses are supported. The paper ends with a short discussion on what is expected to happen in the near future. A future that can be extremely promising if humanity manages to have AI as a proper ally and partner, with distinct roles and specific rules of cooperation and interaction.", "keywords": [ "ChatGTP", "Programming", "Computer Science", "Overview", "Survey Education", "Ethics." ], "content": "Introduction\n\nRecently, in the field of human-computer communication and natural language processing, Artificial intelligence (AI)-based language models such as several versions of ChatGPT (3 and 4) have shown excellent performance in translation, question answering, inference estimation, text evaluation and summarization, but also in the field of automatic code generation and programming functions (Wolf et al., 2020; Nelson et al., 2023). These models have the capability to generate code and assist programmers in completing tasks more efficiently (Hassani and Silva, 2023; Uzair and Naz, 2023). However, there are challenges associated with AI-generated code, such as syntax errors that can hinder comprehension and functionality. Despite these challenges, the integration of AI in computer science and code programming has the potential to enhance productivity, accessibility, and problem-solving capabilities.\n\nThis paper attempts to provide a brief and recent overview that supports or negates the points that ChatGPT itself identifies as important impacts, advantages and disadvantages. Moreover, the paper concentrates some structural categories for the major impacts of them in computer science (programming and education) as identified by ChatGPT and as discussed in over 40 recent research papers (most of them published in 2022 and 2023). The major goal is to provide a brief source of reference for researchers involved in the above scientific fields. This would support the systematic performance evaluation in the scientific community worldwide and allow developers to get familiarized with technologies that are still immature for the time being, but are here to stay.\n\n\nImpacts of AI generated content and ChatGPT in programming\n\nThe following subsections discuss what are the main implications of AI generated content and ChatGPT in the field of computer science programming and programmers training. It is chosen to evaluate the main applications, advantages, disadvantages, future implications, and ethical considerations providing recent documentation and sources that support those outcomes. Thus, the following five questions were addressed to ChatGPT version 3.5, in order to identify the opinion of ChatGPT in these issues:\n\nQuestion 1: What are the most important applications of ChatGPT in programming and training of programmers?\n\nQuestion 2: What are the major advantages of ChatGPT in programming and training of programmers?\n\nQuestion 3: What are the most important limitations of ChatGPT in programming and training of programmers?\n\nQuestion 4: What are the future prospects of ChatGPT in programming and training of programmers?\n\nQuestion 5: What are ethical considerations of ChatGPT in programming and training of programmers?\n\nIn the following five sections the responses of ChatGPT in every one of the above five questions are given in bullets in the respective tables. Each one of the Tables 1 to 5 demonstrate the recent papers identified in the literature that have a positive, negative or a partially positive/negative view. The tables are then followed by a brief discussion. The reader may have direct access to questions asked and answers given by ChatGPT from Anagnostopoulos (2023).\n\nM. Assistance in code management and production:\n\nAutomatically generated AI content helps developers by showing suggestions for code or code snippets, syntax checks and debugging help. Indeed it is suggested by literature, ChatGPT is an effective tool for generating computer language code snippets (Sallam, 2023; Devlin et al., 2018), it is also effective for conducting comprehensive literature reviews and surveys from the literature, thus helping scientists even in creating content for writing research proposal suggestions (Liu et al., 2019; Li, 2023). But in any case, if one wants to use an AI tool specifically tailored for programming, then GitHub Copilot is the optimal assistant that works with AI principles (Prather, 2023).\n\nb. Programming tutoring: Content generated using AI is an excellent aid for users who are first learning computer programming languages, as it is possible to develop explanatory dialogues for various concepts and answer basic questions. ChatGPT offers process assistance for writing code, improving training in programming languages and also supporting users in programming tasks. ChatGPT has even been compared to other tools for automated programming code generation and has been found to have unique capabilities (Zambrano, 2023). In the context of research and education, ChatGPT has been found to be a tool that assists researchers in writing research papers, as mentioned above. However, it is particularly critical to keep in mind that ChatGPT several times requires the user to validate and verify the accuracy of the information it provides. The quality of ChatGPT’s responses, particularly in literature reviews, literature reports and data sources (FERROUHI, 2023; Hopkins et al., 2023) has been debated and specific doubts have been raised about its accuracy.\n\nIn the field of computer programming education, ChatGPT has also been used to assist teachers in related tasks, such as grading and assessment (Iskender, 2023). However, just as is the case for learners/students, there are serious questions about the risk of reducing instructors’ critical thinking if they become overly dependent on ChatGPT (Iskender, 2023; Uddin et al., 2023). A common consensus of both of these papers is that ChatGPT can damage the critical thinking ability of its users (both students and instructors). Moreover (Iskender, 2023; Uddin et al., 2023), agree that there is a need for continuous self-improvement for learners and educators and clear understanding of the correct degree of ChatGPT use and value as a tool of the educational process.\n\nc. Natural language processing (NLP): ChatGPT finds particularly important applications in the field of NLP, which include computer science education as mentioned above, but also its assessment, automatic question answering systems, automated text generation, as well as translation between texts in different languages and types (Serdaliyev and Zhunissov, 2023).\n\nIn the broader field of NLP, ChatGPT found immediate applications in chatbots and virtual assistants (Serdaliyev and Zhunissov, 2023; Gilson et al., 2023), which is not surprising as it is itself a form of chatbot. Moreover, ChatGPT is proposed for automated paper writing applications, and text generation (text from scratch or revised versions) for research papers and articles (Gilson et al., 2023; Temsah et al., 2023).\n\nBeyond writing papers, ChatGPT is reported to be used for research paper editing, academic instruction, and knowledge-based assessments (Stokel-Walker and Noorden, 2023; Buriak et al., 2023; Banerjee et al., 2023; Ali et al., 2023) as well as a tool for writing research proposals for funding and for academic theses (Qasem, 2023). But, as will be discussed in the section of ethical issues raised, a great concern is identified about an increase in plagiarism and over-reliance on ChatGPT (Qasem, 2023) leading to limitations in analytical capabilities (Stokel-Walker and Noorden, 2023) and for this case, the necessity of human verification before submitting proposals or a thesis is also emphasized (Sallam, 2023).\n\nD. Technical documentation: AI text generation models are capable of assisting in the completion of technical documentation and explanatory text for projects or software applications. Demonstrated success in this area brings to the fore the importance of design choices and raises questions about the source of recent improvements in the performance of language models (Liu et al., 2019). The attention mechanism, as presented in (Vaswani et al., 2017), is the one that has played the most crucial role in the development of these language models. Therefore, by incorporating the power of AI language models, software projects and applications can benefit from the automated production of technical documentation that should always accompany a software project thus increasing its sustainability and updating it with fully documented new releases.\n\na. Increase of the software developer productivity and rapid idea prototyping: As found by the papers cited in this paragraph AI generated content can speed up development tasks, reduce coding errors, and enhance the overall productivity of software engineers. It also enables individuals with little or no programming experience to engage with computer science concepts and coding. Specifically, ChatGPT can be used to speed up software development tasks and enhance the overall productivity of engineers engaged in these tasks (Sallam, 2023; Haque and Li, 2023). As mentioned in the previous section, ChatGPT helps in computer code generation or software prototypes, and with its proper utilization, software engineers can save time from low-level tasks that require significant effort and focus on higher-level tasks and experimental design (Sallam, 2023), get explanations, feedback and acquire realistic virtual simulations that may contribute to hands-on learning experiences (Qadir, 2023). A very good example is that by using ChatGPT, developers can take advantage of its ability to easily produce code at a fairly satisfactory level, fill in any missing parts and/or update it, or even develop it in different programming languages (Kreitmeir and Raschky, 2023). Moreover, an important capability of ChatGPT is the provision of explanations and suggestions that can help reduce coding errors and optimize the quality of the code being programmed (Haque and Li, 2023).\n\nb. Continuous learning: ChatGPT can also assist in the formation of collaborative workflows during the implementation of an information technology (IT) project or during the production of software, forming an enhanced human-machine interaction (Nascimento, 2023). At the same time, ChatGPT’s capabilities in natural language processing, beyond the creation of new content, have been reported to offer significant potential in information retrieval, cataloging and indexing, and metadata creation (Lund and Wang, 2023). In addition to the above, it is important to add its flexibility and interactivity that has been found to increase the productivity of scientists and their ability to access new knowledge and skills (Irons et al., 2023).\n\nM. Lack of context: It seems that this is the most important limitation of AI generated content, since it is evident from the literature that AI still cannot fully understand the context of specific codebases or assets, leading to potential inaccuracies and inappropriate promptings.\n\nSpecifically, Wang et al. (2021) discuss the limitations of current methods in processing code snippets, neglecting the special characteristics of programming languages and token types, which can result in suboptimal understanding and generation tasks. Khoshafah (2023) denotes issues related to the translation accuracy of ChatGPT and highlights limitations in understanding domain-specific terminology and cultural context, which can affect the accuracy of translations. Aljanabi et al. (2023) mentions that ChatGPT may not fully understand the nuances of security vulnerabilities, reverse engineering, or malware analysis, potentially leading to inaccurate or less useful information in coding. Additionally, it mentions that ChatGPT may not be able to handle certain types of queries, such as mathematical calculations.\n\nb. Security concerns: As reported in Flanagin (2023), AI models like ChatGPT might inadvertently generate code with security vulnerabilities, which could be exploited by malicious actors. Security concerns arise in the use of ChatGPT in computer science applications. The potential for misuse, such as using ChatGPT to cheat on assignments, write essays, or take exams, including computer science and programming exams, has been identified. Flanagin (2023) identify the inclusion of ChatGPT as a bylined author in scientific articles has raised concerns about the integrity of scientific publication and the indexing of nonhuman “authors”. The lack of accountability and responsibility associated with AI tools like ChatGPT has prompted the development of policies by journals and organizations to address these issues (Flanagin, 2023). Additionally, the limitations of ChatGPT, such as its lack of context and inability to understand new information or generate deep analysis, restrict its utility in writing up-to-date reviews, perspectives, and introductions (Buriak et al., 2023). Therefore, it is crucial to address these security and ethical concerns and ensure responsible use of ChatGPT in computer science to maintain the integrity of scientific research and programming education.\n\nc. Overconfidence or excessive trust in AI: Dependence on AI-generated code without proper understanding certainly would negatively affect developers’ growth as mentioned in Tang (2023). In simpler words, while AI technologies like low-code development platforms offer efficiency and productivity benefits, relying solely on AI-generated code may limit developers’ understanding of underlying concepts and principles. This lack of understanding can hinder their ability to troubleshoot, debug, and optimize code, resulting in less robust and maintainable solutions. Additionally, the lack of comprehension may impede developers’ growth and hinder their ability to adapt to new technologies and challenges (Tang, 2023). Therefore, as a conclusion from the above, it is important to maintain a balance between implementing AI technologies and ensuring developers’ comprehensive understanding of code and underlying principles.\n\na. AI languages customization: From the related literature, it seems that future iterations of AI language models would certainly be tailored to specific programming languages and frameworks, providing more specialized support. As an example, Devlin et al. (2018) introduced BERT (Bidirectional Encoder Representations from Transformers), a language representation model that can be fine-tuned for various tasks. BERT was designed to pretrain deep bidirectional representations from unlabeled text, and it has been shown to achieve state-of-the-art results on multiple natural language processing tasks (Devlin et al., 2018). Additionally, Liu et al. (2019) presented RoBERTa, a robustly optimized BERT pretraining approach. While the latter does not directly address the customization of AI language models for specific programming languages and frameworks, it provides insights into the advancements and optimizations in BERT-based models. More recently, Brown et al. (2020) discusses the scaling up of language models and its impact on task-agnostic, few-shot performance giving evidence about the performance improvements achieved by scaling up language models.\n\nb. Collaborative operations: AI models may facilitate collaborative programming, enabling developers to work together more effectively. Back in 2018, Mikhaylov et al. (2018) discussed the challenges of cross-sector collaboration in the public sector when adopting AI and data science and programming. The authors described challenges such as information silos, lack of resources, and collaborative culture. In addition, the authors highlighted the divergent approaches to managing risk in the public and private sectors. Since the political and market risks may not easily align, challenges may be created in collaborative programming efforts using AI models. Moreover, they continue mentioning that cross-sector collaborations, although popular, entail serious management challenges that hinder their success and may impact the effectiveness of collaborative programming facilitated by AI models.\n\nc. Self-correcting models: Τo address the challenge of mistake recognition, researchers have explored different decoding strategies for text generation from language models. Holtzman et al. (2019) propose Nucleus Sampling, a method that draws higher quality text from neural language models compared to traditional decoding strategies. By truncating the unreliable tail of the probability distribution and sampling from the dynamic nucleus of tokens, Nucleus Sampling aims to avoid text degeneration and generate more coherent and diverse responses.\n\nIn addition, recent studies have examined the fine-tuning of language models by exploiting user preferences and reinforcement learning models to improve the accuracy and quality of responses. For instance, Ziegler et al. (2019) proposes a reward-based learning in natural language dialogues, ultimately creating a model of retributive learning implemented by asking users questions. This approach is a successful application of reinforcement learning in real-world dialogue scenarios where rewards are determined by human judgment through the incorporation of human preferences and feedback that could be incorporated in ChatGPT future versions.\n\nIn support of the above article, Brown et al. (2020) very emphatically cited the weaknesses that NLP systems faced in performing new linguistic tasks when given few examples or simple instructions. In their article they argue that AI models are not yet at the same level of error recognition and correction capabilities as a human user, but they acknowledge that this will change radically in the near future with improved decoding strategies, such as kernel sampling, and the incorporation of sophisticated adaptation and reward learning.\n\nFrom the above findings and the overall feeling of the exponential growth of the related technology, it is evident that AI models could eventually become better at recognizing and fixing their mistakes, leading to more accurate responses. AI language models have shown remarkable progress in various NLP tasks, but their ability to recognize and fix mistakes is an ongoing challenge (Vaswani et al. 2017). By addressing the limitations and leveraging advancements in the field, AI models could improve their error recognition and correction capabilities, ultimately leading to more accurate responses.\n\na. Plagiarism, proper citation and acknowledgement: This is one of the most important ethical issues that arises as the code generated by AI should be recognized as automatically generated and programmers should not appropriate these results without critical processing (Taddeo and Floridi, 2018).\n\nTo this end, Li (2022) addresses the issue of source code intellectual property and the necessity for accurate attribution of code to its owner/developer, issues that are crucial in the software and application implementation cycle and in the broader fields of software forensics and software quality analysis. In the same article (Li, 2022), acknowledges that the current methods of rendering the authoring of a code can be copied by other users, which means that the same phenomena can occur in ChatGPT. Furthermore, Gibea and Uszkai (2023) summarize in their article the growing concern of the research community about the development and deployment of AI systems and point out the need for a common code of ethics to be established by international organizations in response to these concerns. In addition, Svetlova (2022) argues that AI ethics should also take into account all systemic implications arising from the use of AI, especially the systemic effects on developers’ codes of conduct. All of the above argues that the appropriate performance of an automatically generated code is critical to address potential systemic risks, but also to ensure the appropriate development of AI itself.\n\nSimilarly, the article by Jobin and Vayena, (2019), highlights the new landscape of ethical guidelines for AI with both meaningful analysis and appropriate implementation strategies. The importance of adhering to ethical considerations, among which is the appropriate attribution of the rights of a code segment, is reemphasized and also for the development of AI systems. In the same vein, Taddeo and Floridi (2018) stress the importance of having an ethical framework for exploiting the potential of AI, but where control remains strictly with the human agent. Furthermore, Minkkinen et al. (2022) present for the first time the use of AI in process evaluations in administrative, social and governmental issues, proposing standardized metrics for the responsible use of AI, including the proper performance of AI-generated code.\n\nThe important question ”Does AI own intellectual property rights in content produced by it?” is answered in the article by Sharma and Sony (2020). The article highlights the uncertainty that arises on this issue, but again clearly supports the need for appropriate attribution. In addition, Díaz-Noci (2020) went a little further into the question and presented the legal implications of news produced using AI systems and what happens to intellectual property rights. Although the article is mainly concerned with journalistic practices, it highlights the challenges in determining the rights of the author, and again strongly supports the need for appropriate attribution of AI-generated content.\n\nVery interesting is also the article by Almarzoqi and Albakjaji (2022), who even explored the possibility of patenting products resulting from AI, while they also examined the challenges posed by intellectual property laws in this context. As in the previous article, the need for appropriate attribution to the creator (AI) is emphasized. Wowever, it is recognized that the legal frameworks that describe and regulate intellectual property in innovations/new inventions are not ready to patent something that is purely generated by AI.\n\nMore recently, Lu et al. (2023) propose the idea of collecting patterns to assist in the design of responsible AI systems. Their article highlights the need to address the challenges of responsible AI operation, part of which includes the appropriate rendering of the code generated by AI. Similarly, Haonan et al. (2023) discuss copyright protection and accountability of creative AI. Although they focus on intellectual property rights, their work highlights the need for attribution and accountability in the context of AI-generated works.\n\nb. Bias and fairness\n\nIt should be emphasized that the impact of bias in training data that leads to discriminatory code suggestions from AI generated text or code, was early underlined from the scientific community. For instance (Caliskan et al., 2017), discuss how text corpora contain recoverable and accurate imprints of historic biases, including biases towards race or gender. This suggests that biases present in the training data can be reflected in the output of AI models, potentially leading to biased or discriminatory code suggestions. Aligned with the above assumptions, Dixon et al. (2018) demonstrated how imbalances in training data can lead to unintended bias in resulting models. This implies that if the training data contains biased or discriminatory patterns, the AI model’s code suggestions may also exhibit such biases.\n\nMoreover, Garg et al. (2018) highlighted the presence of gender and ethnic stereotypes in word embeddings trained on text data. Word embeddings, which are widely used in natural language processing tasks, can amplify biases present in the training data. This suggests that biases in training data can affect the behavior of AI models, including code suggestions.\n\nTo sum up, ChatGPT has shown potential as a code assistance tool, aiding in generating computer codes and supporting researchers in coding tasks developers should be aware when they are interacting with AI systems and understand the model’s limitations to make informed decisions and take responsibility for thoroughly reviewing and testing the output. To this end, Amodei et al. (2016) focuses on AI safety and the potential risks associated with forward-looking applications of AI. While it does not directly address the need for developers to understand AI model limitations, it highlights the importance of considering safety in the design and deployment of AI systems.\n\nFrom all the above it can be assumed that it is important to focus on efforts to mitigate the presence of bias in training data, to avoid biased or discriminatory code suggestions.\n\n\nDiscussion – outlook for the future\n\nAs an AI-based language model, ChatGPT is designed to be trained by its interactions with humans and appropriately adapt its responses (Lu, 2023; Tenakwah, 2023; Kohnke et al., 2023). As such, it is reasonable to expect that future versions of ChatGPT, will be more comprehensive and reliable, as they will have gathered a greater amount of data and experience. This accumulation of knowledge will allow it to provide even more accurate and appropriate answers for the subjects for which questions are asked, both in general discussions and specifically in areas such as computer science education and code generation.\n\nOn a technical level, the ChatGPT developers will likely incorporate new updates that will include improvements in NLP, increased ability to understand user queries and the ability to generate responses that are more coherent and closer to everyday human communication in natural language. In addition, it is reasonable to assume that the developers have addressed any limitations or biases that were identified in previous versions, ensuring an ever-increasing impartiality of ChatGPT.\n\nIn addition, the next era for ChatGPT could include the integration of new features or functions. As a representative example, a feature has already been launched where a feedback mechanism is incorporated that allows users to provide feedback on the quality of responses. This feedback loop continuously improves ChatGPT based on user feedback, resulting in a more personalized and satisfying user experience. It is also expected that ChatGPT will be integrated with different AI models through artificial agents. Such collaborations will lead to a more diverse range of answers in computer science to create code, increase the knowledge base and improve problem-solving capabilities (Neil et al., 2022) and (Beiqi et al., 2023).\n\nHowever, the human factor must play an important role in the final decisions and judgements. In particular, in the field of computer science, programmers should not rely solely on the answers provided by an automated system but should instead combine them with their own expertise and experience and should continuously cultivate their knowledge, improve their skills and remain actively involved in the continuous and lifelong learning of (Christen et al., 2023).", "appendix": "Data availability\n\nZenodo: ChatGPT impacts in programming education: A short list of input questions and output answers from ChatGPT. Zenodo. https://doi.org/10.5281/zenodo.8375014 (Anagnostopoulos, 2023).\n\nThis project contains the following underlying data:\n\n- QUESTIONS_ANSWERS_CHATGPT.pdf\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nAli K, Barhom N, Marino FT, et al.: The Thrills and Chills of ChatGPT: Implications for Assessments in Undergraduate Dental Education. Preprints 2023. 2023; 2023020513. 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Publisher Full Text\n\nZiegler DM, Stiennon N, Wu J, et al.: Fine-tuning Language Models from Human Preferences.2019. arXiv:1909.08593. Publisher Full Text" }	[ { "id": "230278", "date": "04 Jan 2024", "name": "FX. Risang Baskara", "expertise": [ "Reviewer Expertise AI in Education", "Blended Learning", "Computer-Assisted Language Learning", "Computer-Mediated Communication", "Flipped Learning", "Intelligent Tutoring Systems", "Mobile-Assisted Language Learning", "Technology-Enhanced Language Learning" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nDetailed Review Report with Sample Revision Suggestions:\nArticle Summary: \"ChatGPT impacts in programming education: A recent literature overview that debates ChatGPT responses\" by Christos-Nikolaos Anagnostopoulos examines the role of ChatGPT in programming education. The paper discusses its applications, benefits, limitations, and ethical implications, supported by current literature and ChatGPT's responses to specific programming queries. The objective is to offer insights for researchers in programming and education sectors on the integration of AI tools like ChatGPT.\nReview and Recommendations:\n1. Comprehensiveness in Context of Current Literature (Partly):\n\n- Expansion Suggestion: Include more diverse viewpoints, particularly those skeptical of or opposing AI's use in education. For instance, add studies focusing on the potential drawbacks of AI in programming education, such as reduced critical thinking skills or over-reliance on AI solutions.\n\n- Sample Revision: \"While this paper highlights the positive impacts of ChatGPT in programming education, a comprehensive analysis necessitates the inclusion of literature that scrutinizes its limitations and potential adverse effects, such as [Author X, Year] and [Author Y, Year].\"\n2. Accuracy and Citation Support (Yes):\n\n- Critical Analysis Enhancement: Strengthen the critical analysis of each source. Rather than just presenting the information, engage with it critically, discussing its implications and how it fits into the broader context of AI in education.\n\n- Sample Revision: \"In discussing the work of [Author Z, Year], it is essential to not only present their findings but also to critically analyze their methodology and the broader implications of their conclusions for programming education.\"\n3. Accessibility of Language (Yes):\n\n- Maintaining Clarity: As the article expands in depth, ensure that the language remains accessible. Technical jargon should be explained, and complex concepts broken down for a broad academic audience.\n4. Appropriateness of Conclusions (Partly):\n\n- Balanced Conclusion Development: Develop a more nuanced and balanced conclusion that acknowledges both the strengths and limitations of ChatGPT in programming education.\n\n- Sample Revision: \"The conclusion of this paper, while highlighting the positive aspects of ChatGPT, should also incorporate a discussion on the challenges and ethical considerations, offering a more balanced view on the role of AI in programming education.\"\nEssential Revisions for Scientific Soundness: - Enrich the literature review with a variety of sources, ensuring a balanced view. - Enhance critical engagement with the sources used. - Develop conclusions that reflect a comprehensive and balanced understanding of the topic.\nImplementing these revisions will significantly enhance the article's scientific rigor, making it a more valuable contribution to the academic discourse on AI's role in programming education.\n\nIs the topic of the review discussed comprehensively in the context of the current literature? Partly\n\nAre all factual statements correct and adequately supported by citations? Yes\n\nIs the review written in accessible language? Yes\n\nAre the conclusions drawn appropriate in the context of the current research literature? Partly", "responses": [] }, { "id": "234353", "date": "19 Mar 2024", "name": "Sandro Speth", "expertise": [ "Reviewer Expertise Computer science education and didactics", "AI in programming education", "software architecture", "microservices", "software quality", "cloud computing", "service engineering" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nAccording to the paper's title and abstract, it is about ChatGPT's impact on programming education. While this topic is touched on in some parts of the paper, the majority of the paper does not reflect on programming education or education in general.By having the title “ChatGPT impacts in programming education: A recent literature overview that debates ChatGPT responses” and the concrete abstract, the authors raise a reader’s expectation that this paper did a literature survey about the use and implications of ChatGPT for programming education. However, most statements to answer the questions do not focus on programming education at all. Additionally, question 4 is even unrelated to this topic. Additionally, there are categories regarding programming education which have published paper and that are not even mentioned in this paper. Therefore, the paper fails to fulfill the expectations it raises to the reader.. In fact, many research papers tackling ChatGPT's role in programming education are not even mentioned. When discussing the impact of ChatGPT on programming education, I would expect to see citations of papers about (1) the generation of programming teaching material, e.g., exercise sheets, using ChatGPT, (2) issues regarding students solving their exercises with ChatGPT instead of learning the concepts themselves, (3) how to create exercises which ChatGPT cannot solve.\n\nInstead, the author asked ChatGPT itself five questions, and based on the answers, general research papers for AI-generated content were surveyed without a particular focus on programming education. Instead, the paper contains many research papers that neither discuss ChatGPT or other GPT-based tools nor programming education. Often, the information elicited from the cited papers is very high-level, and the author does not provide deeper details and discussions. See details for more information.\nApart from the content itself, I am missing any information about the methodology. For a literature survey, I would expect a section in which the author discusses the applied methodology. To reproduce the findings, the section should state used search keywords and search engines (e.g., Google Scholar, IEEE Xplore, etc.) and whether forward or backward snowballing was applied. Furthermore, it should elaborate on inclusion and exclusion criteria and how many pages of the search engine were deemed to be relevant. Especially for a topic with such a vast publication frequency and changes in technology as large language model tools and publications have, the author should state in which period the survey was conducted.\nPlease note that the added seven citations are only excerpts. There are likely various additional papers highly relevant to the paper's topic.\nDETAILS\n\nAbstract: - Details on how many papers were surveyed during which period of time is missing.\nIntroduction: - The introduction does not really follow a good practice structure where, first, the research is stated, then the problem, afterwards why it is not solved yet, then the methodology and contributions with some details, and afterwards the evaluation and who benefits of it. I am missing here especially the methodology and contribution details. The author’s state “some structural categories” but which ones? And what are the key points found in this paper? Besides, usually, an introduction ends with an overview of the upcoming sections to simplify reading it.. Please try to make the following clear: What is the exact problem you are tackling in this paper? Did you focus on specific CS education areas or in general? What is your methodology for the survey? How many papers exactly did you survey, and in what period of time? What are the most important results? How is the remainder of the paper structured? - In the paper's title and abstract, the author focuses on programming education only, but the second paragraph of the introduction is more general to computer science. - \"in over 40 recent research papers\" --> Please provide the exact number and details of what \"recent\" means. - The last two sentences of the introduction are very general, without any focus on education. Furthermore, they are very vague by using \"would\", etc.\nImpacts of AI generated content and ChatGPT in programming: - \"AI generated\" --> \"AI-generated\". Perhaps, also consider changing the title and abstract to a general viewpoint of AI-generated content and not only ChatGPT - Either focus on education or change the title and abstract to a broader topic. - The authors put the questions first and instead of providing at least a summary of the answers directly, they state that the answers will be discussed later in the paper. This unnecessarily raises the tension as the reader wants to know whether these parts are worth reading to them. Please consider either providing the answers directly or stating the questions in the respective subsections.\nQuestion 1: - For a focus on CS education, I would expect that 1.a. applies to CS education. However, this seems to be more general. - How does 1.c. relate to programming education? - \"if one wants to use an AI tool specifically tailored for programming, then GitHub Copilot is the optimal assistant that works with AI principles\" --> That seems very superficial. Why is it generally \"the optimal\" assistant? Isn't it possible that someone builds small language models which are more tailored for, e.g., Java programming? - \"Content generated using AI is an excellent aid for users who are first learning computer programming languages\" --> There is also a danger for it. Often, ChatGPT cannot explain using domain terms or even does not know the concepts correctly itself. Especially beginners might take every response as granted and correct which does not necessarily is the case. - \"tasks. ChatGPT has even been compared to other tools for automated programming code generation and has been found to have unique capabilities (Zambrano, 2023).\" --> I am missing details here. What were the insights/results? - \"reducing instructors’ critical thinking if they become overly dependent on ChatGPT\" --> Can you elaborate more on this? - \"ChatGPT found immediate applications in chatbots\" --> This seems to be wrong! ChatGPT is a chat client using OpenAI's GPT APIs. Custom virtual assistants most likely use the GPT APIs themselves. Please note that ChatGPT is not a large language model itself but only a client for various models (GPT 3.5, GPT 4, Dall-E). - D. Technical documentation: Based on the title, you focus on ChatGPT, not general AI models. Please clarify here the impact of ChatGPT on technical documentation. I assume other tools, e.g., GitHub Copilot, are more relevant here.\nQuestion 2: - \"can speed up development tasks, reduce coding errors, and enhance the overall productivity of software engineers.\" --> Does this depend on the application and programmer's skills? The quality of responses highly depends on the application area. ChatGPT also introduces errors that might be tedious to find, especially if the programmer is inexperienced. How does this apply to CS education? - \"get explanations, feedback and acquire realistic virtual simulations that may contribute to hands-on learning experiences (Qadir, 2023)\" --> Does Qadir provide any numbers? \"may contribute\" is rather fuzzy. Are there any surveys, experiments, etc., supporting this claim? - b. Continuous learning: I would say \"continuous learning\" is fitted for education. However, there seems to be no work on how ChatGPT or other LLM-based tools support continuous learning in education.\nQuestion 3: - \"since it is evident from the literature that AI still cannot fully understand the context of specific codebases or assets\" --> This is overly general. ChatGPT does not fully understand it. If you would open the entire project as tabs in VS Code, then GitHub Copilot should understand the entire code base in addition to the language knowledge it anyhow has. - \"highlights limitations in understanding domain-specific terminology\" --> This seems to be highly relevant for education as learners require their \"tutor\" to have a good understanding of domain-specific terminology. Any work on that? - b. Security concerns: Apart from the first two sentences, the entire paragraph does not seem to discuss security concerns but only general concerns. - c. Overconfidence or excessive trust in AI: \"AI technologies like low-code development platforms\" --> According to the title and abstract, this paper discusses the impact of ChatGPT on CS programming education. Low-code development platforms are not necessarily AI technologies and certainly not ChatGPT \"it is important to maintain a balance between implementing AI technologies and ensuring developers’ comprehensive understanding of code and underlying principles.\" --> Is \"implementing\" here the correct term? In my opinion, \"applying\" makes more sense. Implementing AI technologies requires understanding code and underlying (AI) principles.\nQuestion 4: - This is unrelated to the paper's topic (at least according to the title and abstract).\nQuestion 5: - Not really discussed in the focus of CS education or education in general - \"Very interesting is also the article by ...\" --> This is a subjective rating and should not be inside a research paper\nDiscussion: - This is not a discussion about the impact of ChatGPT on CS programming education\n\nIs the topic of the review discussed comprehensively in the context of the current literature? No\n\nAre all factual statements correct and adequately supported by citations? Yes\n\nIs the review written in accessible language? Yes\n\nAre the conclusions drawn appropriate in the context of the current research literature? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1393
https://f1000research.com/articles/12-1392/v1	23 Oct 23	{ "type": "Research Article", "title": "High-throughput virtual screening approach and dynamic simulation of natural compounds as target inhibitors of BACE1 in Alzheimer's disease", "authors": [ "Fadoua Elkamili", "Abderrahim Ait Ouchaoui", "Leandro L. Lorente-Leyva", "Diego Hernán Peluffo-Ordóñez", "Abderrahim Ait Ouchaoui", "Leandro L. Lorente-Leyva" ], "abstract": "Background Beta-site amyloid precursor protein cleaving enzyme 1 (BACE1) protein holds significance in the development of Alzheimer’s disease (AD). This protein is highly expressed in the central nervous system, playing a significant role in the conversion of amyloid precursor protein (APP) into amyloid-beta peptides. The primary objective of the current study was to perform in silico inhibition investigations on this protein, utilizing computational methodologies such as molecular docking and molecular dynamics, to identify novel inhibitors with potential against BACE1. Our focus was on targeting the active site of BACE1, aiming to discover optimal interactions between the ligands and key residues of the target protein.\n\nMethods The natural compounds inhibitors data was carefully reviewed from the literature review articles. Molecular docking was conducted using AutoDock Vina with an active site defined by CB-Dock2. We assessed drug-likeness and toxicity using Lipinski’s rule of fives via SwissADME and the ProTox-II web server. Interaction visualization was facilitated using Discovery Studio, while molecular dynamics simulations were performed for 100 nanoseconds with hinokiflavone and BACE1 using Schrodinger LLC Desmond software.\n\nResults The molecular docking results showed promising binding affinities and the best binding free energy values were selected, and after conducting Lipinski’s rule of five using SwissADME as well as predicting toxicity using ProTox-II, only one molecule, hinokiflavone was filtered and succeeded in all the analyses to be a potential candidate. Molecular docking results were supported by molecular dynamics simulation. These results demonstrate the stability of the compound in the target protein binding site.\n\nConclusions Finally, these obtained outcomes represent a strong lead to developing promising new natural compound inhibitors against BACE1. For future works, it is essential to concentrate on further experimental validation to ensure the effectiveness of the proposed approach.", "keywords": [ "Alzheimer's disease", "BACE1", "Molecular docking", "Molecular dynamics", "Natural compounds", "Hinokiflavone" ], "content": "Introduction\n\nAlzheimer’s disease (AD) is a progressive neurological disorder that declines in cognitive function and is usually recognized as a major health hazard that mostly affects people older than 60 years old (Husna Ibrahim et al., 2020). It occurs due to a gradual and slow neurodegenerative process, leading to age-related neuronal cell death. The disease is characterized by the accumulation of amyloid plaques, the tangling of neurofibril tissue, and synaptic dysfunction. It manifests in various stages, gradually affecting everyday functions and leading to symptoms like apathy, depression, communication difficulties, disorientation, impaired judgment, swallowing and walking problems, and behavioral changes (Abubakar et al., 2022).\n\nRegrettably, there is currently no known cure for the disease. Nowadays, researchers are dedicated to comprehending the pathology of AD through various mechanisms, such as abnormal tau protein metabolism, β-amyloid, inflammatory response, and cholinergic and free radical damage, aiming to develop effective treatments that can stop or slowing the progression of AD (Breijyeh & Karaman, 2020). To prevent the death of neurons, one potential approach involves inhibiting beta-site amyloid precursor protein cleaving enzyme 1 (BACE1), which can help prevent the buildup of beta-amyloid (Aβ). This strategy aims to address one of the underlying mechanisms associated with AD pathology (Yan & Vassar, 2014).\n\nBACE1, an enzyme discovered in 1999, plays a crucial role in the early stages of AD (Yan & Vassar, 2014). BACE1 is an enzyme responsible for breaking down the Amyloid Protein Precursor (APP) and releasing APP fragments, which lead to the formation of Aβ on nerve cell membranes, which is a key component of the plaques associated with AD. It is found at higher levels in the brains and body fluids of individuals with AD, suggesting its involvement in the disease. Therefore, targeting BACE1 activity with drugs may hold potential for therapeutic interventions aimed at reducing Aβ accumulation and slowing down the progression of AD (Das & Yan, 2017).\n\nIn the drug development process, docking plays an important role in drug design. Molecular docking has been used to search low binding free energy and also the ligand conformations at the protein-binding sites. In addition, docking also provided useful information for predicting the orientation of drug candidate binding to the protein target (Pinzi & Rastelli, 2019). The focus of our study was on conducting a multistage virtual screening of 400 natural compounds extracted from natural plants. These compounds were carefully chosen based on a thorough review of the existing literature, specifically focusing on selecting molecules with known anticancer activity. The objective was to identify natural compounds that exhibit high affinity towards inhibiting BACE1 activity while having low toxicity.\n\n\nMethods\n\nIn this research, we selected a database of 400 BACE1 inhibitor compounds (Elkamili et al., 2023). We used the PubChem library to retrieve all the ligand compounds, in SDF format. These compound structures were then imported into OpenBabel for conversion into PDB format (O’Boyle et al., 2011). The compounds were further prepared using AutoDockTools, during which Gasteiger charges were applied to them and saved in PDBQT format.\n\nThe target protein, BACE1 (PDB code: 3K5D), was prepared before commencing the docking process. The three-dimensional crystal structure of the protein was obtained from the PDB database in PDB format. In AutoDockTools, the structure was processed by removing water molecules and heteroatoms, and then Kollman charges and polar hydrogens were added and the structure of the protein was saved in PDBQT format.\n\nRecognizing the suitable active site for binding the ligand molecules is the most crucial aspect in designing a drug via computational docking. AutoDock Vina (RRID:SCR_011958) was employed for molecular docking at exhaustiveness level 8 to predict the potential interactions between the ligand compounds and the protein BACE1. The grid box of the protein structure was adjusted to fit the active site that is predicted by CB-Dock2 to predict the binding pocket of the BACE1 target (Liu et al., 2022). The spacing was set to 13 Å, 8 Å, and 70 Å points along the x, y, and z-axe, with the lattice size set to 27 Å × 27 Å × 27 Å at the center. A total of 400 compounds were screened and docked against the target protein. Docking affinity was used to generate nine poses for each protein-ligand complex.\n\nIn our study, we employed Lipinski’s rule of fives using SwissADME as a guideline for evaluating the drug-likeness and potential oral bioavailability of the selected compounds with best affinities (Daina et al., 2017). By assessing factors such as molecular weight, lipophilicity, hydrogen bond donors and acceptors, and the number of rotatable bonds, Lipinski’s rule helps in identifying compounds that possess favorable physicochemical properties for drug development. In our screening process, we applied these criteria to filter out compounds that were more likely to exhibit optimal pharmacological activity, thus narrowing down our selection to the most promising ligands for further analysis and potential therapeutic applications.\n\nTo predict the potential toxicity of the compounds, the ProTox-II web server was utilized. ProTox-II is a reliable online tool specifically designed for predicting the toxicity of chemicals (Banerjee et al., 2018). By inputting the SMILES of the compounds into the server, we obtained valuable toxicity prediction results. This step was crucial in identifying and excluding potentially toxic molecules, ensuring that only safe compounds were included in the subsequent analyses and minimizing potential harm based on several types of toxicity that were evaluated including hepatotoxicity, carcinogenicity, cytotoxicity, mutagenicity, immunotoxicity, and DL-50.\n\nBiova Discovery Studio, a comprehensive molecular modeling and simulation platform, offered a wide range of features and modules that facilitated the investigation of complex molecular systems through the visualization of interactions of post-docking complexes, facilitating a deeper understanding of the interactions between the compounds and their respective targets.\n\nThe MD simulations were conducted for 100 nanoseconds to ensure the stability of interactions between hinokiflavone and the BACE1 target using Schrodinger LLC Desmond software (RRID:SCR_014575). Before the simulations, the ligand-receptor complex was subjected to preprocessing using Maestro’s Protein Preparation Wizard, which involved optimization, and minimization. The system was then constructed using the System Builder tool (a module of Protein Preparation Wizard). For the simulations, the TIP3P solvent model was employed in an orthorhombic box, maintaining a temperature of 300 K, pressure of 1 atm, and utilizing the OPLS force field. To simulate physiological conditions, counter ions, and 0.15 M sodium chloride were added to neutralize the models. Before the actual simulation, the models were equilibrated, and trajectories were saved at intervals of 100 ps for further analysis and inspection.\n\n\nResults\n\nWe used the Cb-Dock2 website to predict the binding pocket of the BACE1 target. Figure 1 displays the key residues of the potential binding site on the protein (Elkamili et al., 2023). All of these residues are located in chain C which include: ASP32, GLY34, PRO70, TYR71, THR72, GLN73, GLY74, LYS107, PHE108, PHE109, ILE110, TRP115, ILE118, TYR198, LYS224, ILE226, ASP228, GLY230, THR231, THR232, ARG235, THR329, GLY330, THR331 and VAL332. These residues have been mentioned in several studies as part of the active site of BACE1 (Wu et al., 2016; Xu et al., 2012; Hernández-Rodríguez et al., 2016; Ellis & Shen, 2015).\n\nKey residues represented in pink in the binding pocket and the surfaces of the binding pocket in BACE1 are presented in grey. BACE1, Beta-site amyloid precursor protein cleaving enzyme 1.\n\nA total of 400 compounds were screened against the protein BACE1 using AutoDock Vina to identify potential binding interactions and determine the optimal binding affinity. The docking results yielded positive outcomes, demonstrating a strong binding affinity for the screened compounds. The lowest binding affinity, at -10.8 kcal/mol, was observed with hinokiflavone. On the other hand, the highest binding affinity reached -4.2 kcal/mol with epigoitrin. For further analysis, we focused on compounds with scores equal to or below -9 kcal/mol to identify molecules with significant potential for therapeutic efficacy, reliability, and favorable interactions. A list of the selected compounds with the best binding affinity, obtained after the analysis of the docking results can be found in Table 1. We used these molecules, which demonstrated binding affinity scores equal to or lower than -9 kcal/mol, to apply the Lipinski’s Rule of Five parameters and select potential drug candidates.\n\nAfter conducting the molecular docking analysis, we examined the results to identify the top 59 molecules exhibiting the strongest binding affinity with the target protein. From this initial set, we further refined our selection and focused on 41 compounds that displayed exceptional properties, showing great potential for further development as potential drug candidates. These 41 compounds successfully passed the Lipinski’s Rule of Five test, validating their suitability for further investigations in our future drug development efforts. These compounds are presented in Table 2.\n\nThe second part of the filtration implicate the utilization of ProTox-II as a filtering tool for compound analysis based on hepatotoxicity, carcinogenicity, cytotoxicity, mutagenicity, immunotoxicity and LD50 (lethal dose) characteristics proved to be highly effective in isolating non-toxic and safe molecules. The application of ProTox-II played a crucial role in the evaluation of the compounds, allowing for a comprehensive assessment of their safety profiles. This step was essential to ensure that only compounds with favorable safety characteristics were selected for further investigation. Out of the initial pool, only hinokiflavone which was identified as inactive in all targets, and the only molecule that exhibits slight toxicity, with a LD50 (lethal dose) closer to 5000, these results are presented in Table 3. Therefore, hinokiflavone is considered a safe natural compound emerging as a potential candidate for molecular dynamics simulations to validate its stability in complex with BACE1 under physiological conditions.\n\nThe visualization of the 2D interactions of the BACE1-hinokiflavone complex is performed using Discovery Studio enabling the prediction of protein residues with which the ligand interacts to validate if the candidate compound hinokiflavone interacts with the active site residues of the BACE1 target. Figure 2 showed that hinokiflavone strongly interacted with most of the active site residues of BACE1, indicating that our results perfectly corroborate with the active site residues of BACE1, justifying their anti-Alzheimer’s potency. In summary, hinokiflavone binds to BACE1 via van der Waals interactions with ASP32, TYR71, GLN73, GLY74, PHE108, PHE109, TRP115, ILE118, LYS224, GLY230, THR231, THR232, GLY330, and VAL332. It forms hydrogen bonds with LYS107, TYR198, and THR329, pi-alkyl interactions with ILE110 and ILE226, pi-anion interaction with ASP228, and a pi-donor hydrogen bond with THR72. Furthermore, molecular dynamics simulations allow for the exploration of the dynamic behavior and stability of the ligand-receptor complexes over time.\n\nBACE1, Beta-site amyloid precursor protein cleaving enzyme 1.\n\nThe dynamic simulation of proteins plays a crucial role in understanding their behavior and interactions. Unlike static molecular docking, dynamic simulations consider the flexibility and conformational changes of proteins over time. This allows for a more comprehensive and accurate analysis of protein-ligand interactions. In this study, we employed dynamic simulations to investigate the dynamic behavior of the BACE1-Hinokiflavone complex, providing valuable insights into its physiological relevance and enhancing our understanding of its functional mechanisms. To capture the dynamic nature of the BACE1 protein, we conducted 100 nanosecond molecular dynamics simulations based on the rigid crystal structure of the protein. By doing so, we gained valuable insights into the dynamic behavior of the BACE1-Hinokiflavone complex, enhancing the physiological relevance of our findings. These simulations allowed us to observe conformational changes and interactions that are crucial for understanding the functional mechanisms of the complex. The parameters explored for analysis included root mean square deviation (RMSD), root mean square fluctuation (RMSF), protein secondary structure, and protein-ligand contact analysis.\n\nThe calculation of RMSD allows us to determine the stability and balance of the complex throughout the 100 ns simulation. A low RMSD value indicates a stable complex, while higher values suggest instability. Furthermore, fluctuations within a broad range indicate significant conformational changes in the protein during the simulation, although fluctuations of approximately 1-3 Å are perfectly acceptable. In the simulation (Figure 3), the left Y-axis provides an overview of the protein’s RMSD evolution, while the ligand’s RMSD graph (right Y-axis) indicates the ligand’s stability within the protein and its binding pocket. The Root Mean Square Deviation (RMSD) of Hinokiflavone bound to the BACE1 protein (Figure 3) remained stable without significant structural deviations after 40 ns towards the end of the simulation, indicating a balanced conformation. This allows for an extended simulation for rigorous analysis.\n\nRMSD, Root Mean Square Deviation; BACE1, Beta-site amyloid precursor protein cleaving enzyme 1.\n\nRMSF is a measure of particle deviation within the protein, revealing the flexibility and rigidity of each amino acid throughout the simulation. Lower RMSF values indicate good system stability, and vice versa. Generally, the terminal regions (N- and C-terminal) exhibit more fluctuations compared to other parts of the protein. However, secondary structure elements such as alpha helices and beta sheets are typically more rigid and experience less fluctuation than loop regions. The RMSF plot (Figure 4) remained stable throughout the simulation, with some fluctuations stabilizing in the ligand-binding regions (indicated by green-colored vertical bars). This indicates that Hinokiflavone contributes to the stability of the BACE1 protein.\n\nThe vertical green lines represent the amino acid residue of BACE1 making contact with ligand. RMSF, Root Means Square Fluctuation; BACE1, Beta-site amyloid precursor protein cleaving enzyme 1.\n\nProtein secondary structure elements (SSE) like alpha-helices and beta-strands are monitored throughout the simulation. The plot at the top summarizes the SSE composition of the protein over the course of the simulation, and the plot at the bottom monitors each residue and its SSE assignment over time, with variations represented in orange and blue corresponding to alpha helices and beta sheets, respectively (Figure 5). Figure 5 showed that the percentage of protein secondary structure elements in BACE1 was approximately 37% and remained constant throughout the simulation, indicating the stability of the overall structure of BACE1 during binding with hinokiflavone and thus the stability of the complex.\n\nThe interaction between the target protein and the ligand was monitored during the simulation to highlight the residues involved in this interaction. These interactions were categorized into four types: hydrogen bonds, hydrophobic interactions, ionic interactions, and water bridges. The interaction diagram between BACE1 and hinokiflavone (Figure 6) showed that hinokiflavone formed bonds with all the residues in the active site. Among them, GLN73, GLY230, and THR232 were maintained for more than 60% of the simulation time, and all three interactions were hydrogen bonds mediated by water.\n\nDifferent color signifies different interactions and contacts formed between the amino acids and the ligand atoms. BACE1, Beta-site amyloid precursor protein cleaving enzyme 1.\n\n\nConclusions\n\nIn conclusion, the findings of this study hold significant importance in the field of AD research. By targeting the BACE1 protein, which plays a crucial role in the production of amyloid-beta peptides, the progression of AD can potentially be decelerated. The use of computational methodologies, including molecular docking and molecular dynamics simulations, allowed for the identification of novel inhibitors against BACE1. Among the natural compounds reviewed, hinokiflavone emerged as a highly promising candidate. It exhibited strong binding affinity and favorable binding free energy values, indicating its potential effectiveness as a BACE1 inhibitor. Moreover, hinokiflavone successfully met the criteria of Lipinski’s rule of five, suggesting its drug-like properties and potential for development as a therapeutic agent. The molecular docking results were further supported by molecular dynamics simulations, which demonstrated the stability of hinokiflavone within the binding site of BACE1 over a 100 ns trajectory. This stability indicates the potential of hinokiflavone to effectively interact with the target protein and inhibit its activity. Overall, the discovery of hinokiflavone as a potential BACE1 inhibitor represents a significant step forward in the search for novel treatments for AD. Further research and validation studies are warranted to fully understand the therapeutic potential of hinokiflavone and its role in mitigating the pathology associated with AD.", "appendix": "Data availability\n\nFigshare: Selection of natural compounds as target inhibitors of BACE1 (data set), https://doi.org/10.6084/m9.figshare.24126756.v1 (Elkamili et al., 2023).\n\nThis repository consists of the following files:\n\n• (.xlsx file) List of 400 compounds screened (including ID and affinity)\n\n• (.zip file) Raw data (PDBQT files)\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nThe authors also are greatly grateful by the support given by the SDAS Research Group (https://sdas-group.com/).\n\n\nReferences\n\nAbubakar MB, Sanusi KO, Ugusman A, et al.: Alzheimer’s Disease: An Update and Insights Into Pathophysiology.2022. Publisher Full Text\n\nBanerjee P, Eckert AO, Schrey AK, et al.: ProTox-II: A webserver for the prediction of toxicity of chemicals. Nucleic Acids Res. 2018; 46(Web Server issue): W257–W263. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBreijyeh Z, Karaman R: Comprehensive Review on Alzheimer’s Disease: Causes and Treatment. Molecules. 2020; 25(24): 5789. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDaina A, Michielin O, Zoete V: SwissADME: A free web tool to evaluate pharmacokinetics, drug-likeness and medicinal chemistry friendliness of small molecules. Sci. Rep. 2017; 7(1): Art. 1. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDas B, Yan R: Role of BACE1 in Alzheimer’s synaptic function. Transl. Neurodegener. 2017; 6(1): 23. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nElkamili F, Ait Ouchaoui A, Lorente-Leyva LL, et al.: Selection of natural compounds as target inhibitors of BACE1 (data set). [Dataset]. Figshare. 2023. Publisher Full Text\n\nEllis CR, Shen J: PH-Dependent Population Shift Regulates BACE1 Activity and Inhibition. J. Am. Chem. Soc. 2015; 137(30): 9543–9546. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHernández-Rodríguez M, Correa-Basurto J, Gutiérrez A, et al.: Asp32 and Asp228 determine the selective inhibition of BACE1 as shown by docking and molecular dynamics simulations—PubMed.2016. Reference Source\n\nHusna Ibrahim N, Yahaya MF, Mohamed W, et al.: Pharmacotherapy of Alzheimer’s Disease: Seeking Clarity in a Time of Uncertainty. Front. Pharmacol. 2020; 11: 504893. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLiu Y, Yang X, Gan J, et al.: CB-Dock2: Improved protein–ligand blind docking by integrating cavity detection, docking and homologous template fitting. Nucleic Acids Res. 2022; 50(W1): W159–W164. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nO’Boyle N, Banck M, James C, et al.: Open Babel: An open chemical toolbox. J. Cheminformatics. 2011; 3(1): 1–14. Publisher Full Text\n\nPinzi L, Rastelli G: Molecular Docking: Shifting Paradigms in Drug Discovery. Int. J. Mol. Sci. 2019; 20. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWu Y-J, Guernon J, Yang F, et al.: Targeting the BACE1 Active Site Flap Leads to a Potent Inhibitor That Elicits Robust Brain Aβ Reduction in Rodents. ACS Med. Chem. Lett. 2016; 7(3): 271–276. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nXu Y, Li M, Greenblatt H, et al.: Flexibility of the flap in the active site of BACE1 as revealed by crystal structures and molecular dynamics simulations. Acta Crystallogr. D Biol. Crystallogr. 2012; 68(Pt 1): 13–25. PubMed Abstract \| Publisher Full Text\n\nYan R, Vassar R: Targeting the β secretase BACE1 for Alzheimer’s disease therapy. Lancet Neurol. 2014; 13(3): 319–329. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "222691", "date": "13 Dec 2023", "name": "Luis Humberto Mendoza-Huizar", "expertise": [ "Reviewer Expertise Computational Chemistry" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe topic of this article is interesting and the proposed methodology is the standard one to determine a possible inhibition of a protein by a ligand. However, I want to focus on Figure 3. The protein-ligand complex has an RMSD higher than 2 A, which is normally taken as a reference to identify if the ligand is stable in the catalytic pocket of the protein. In the RMSD figure, it is clear that at 40 ns there is a drastic change and the RMSD value stabilizes from 50 ns onwards with an RMSD of 21 A. This suggests that the ligand left the original catalytic pocket and settled in a different catalytic pocket than the one initially considered. The authors should verify this situation in order to confirm or reconsider their conclusions.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNo\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [] }, { "id": "249979", "date": "04 Apr 2024", "name": "Atiya Akhtar", "expertise": [ "Reviewer Expertise Phytochemistry", "Ethnopharmacology", "Phyto- pharmaceutics", "Food and nutrition." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors aim to identified novel inhibitors from plant origin with potential against BACE1 in Alzheimer's disease through High-throughput virtual screening approach and dynamic simulation. The work clearly and accurately presented.\nThere are few minor comments would like to address.\nIn Abstract: The primary objective of the current study was to perform in silico inhibition investigations on this protein, utilizing computational methodologies such as molecular docking and molecular dynamics, to identify novel inhibitors from plant origin with potential against BACE1. Please re-write it. In introduction: Paragraph 4, Line no 4… 400 natural compounds extracted from natural plants. Kindly re-write it .... 400 natural compounds isolated from plant species. In the discussion, a greater effort on the part of the authors should be made regarding similar or different results obtained in the literature on the various experimental aspects. A section on the limitations of the study is missing. And kindly write possible clinical implications of the study in conclusions part.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1392
https://f1000research.com/articles/12-1388/v1	20 Oct 23	{ "type": "Research Article", "title": "A novel risk score for venous thromboembolism in lung cancer patients: a retrospective cohort study", "authors": [ "Houda Rouis", "Chirine Moussa", "Islem mejri", "Soumaya Debbiche", "Nourchene Khalfallah", "Lenda Ben Hmida", "Amel Khattab", "Zied Moetamri", "Mohamed Lamine Megdiche", "Hela Kamoun", "Sonia Maâlej", "Houda Rouis", "Islem mejri", "Soumaya Debbiche", "Nourchene Khalfallah", "Lenda Ben Hmida", "Amel Khattab", "Zied Moetamri", "Mohamed Lamine Megdiche", "Hela Kamoun", "Sonia Maâlej" ], "abstract": "Background: Venous thromboembolism (VTE) is a common and potentially fatal complication in patients with lung cancer. This study aimed to develop and validate a risk score for early prediction of VTE in these patients. Methods: Four hundred and one patients with lung cancer from three pulmonology departments hospitalized between January 2011 and December 2021 were retrospectively assessed. The population was divided into two groups: a Development Group (182 patients) and a validation group (199 patients). In the development group, the risk score system was developed, via univariate and multivariate analyses, based on demographic and clinicopathological variables; it was then validated in the validation group. Results: The incidence of VTE was 26.8% in the development group. It was 25.8%, and 27.6% in the internal and external validation groups, respectively. Hemoglobin level <10g/l, metastasis, histological type poorly or undifferentiated non-small cell carcinoma, and active smoking were the items of the risk score system. This score allowed proper stratification of patients with either high or low risk of VTE in the development group (c statistic =0.703). The patients in the development group were classified into 3 risk groups: low risk (scores 0-1), moderate risk (scores 2-3), and high risk (scores 4-5). When validated in the validation group, there was a moderate loss of predictive power of the score (c statistic=0.641), but the categorization of the patients by the score remained clinically useful. Conclusions: This risk score requires prospective validation studies on a nationwide scale in order to use it as a valid tool for the prevention of VTE in lung cancer.", "keywords": [ "Lung cancer", "Venous thromboembolism", "risk score" ], "content": "Introduction\n\nPatients diagnosed with cancer are at a higher risk of developing venous thromboembolism (VTE) compared to the general population, with reported incidence rates ranging from 20% to 30%.1 Among cancer types, lung cancer is particularly associated with thrombosis, with incidence rates ranging from 14% to 30.2\n\nThe risk of thrombosis is significantly increased in cancer patients, by 4 to 12 times higher compared to individuals without cancer, and further elevated to 6.5 to 23 times with the addition of chemotherapy or targeted therapy.3–5\n\nVarious factors contribute to this increased risk, including patient-related factors such as advanced age, previous history of VTE, and obesity; tumor-related factors such as cancer type, stage, and aggressivenessr; and treatment-related factors like surgery, radiation, or systemic anticancer.1,6\n\nThe reported incidence of VTE in the literature varies, possibly due to differences in study design, selection of study participants, varying definitions of VTE, and the exclusion of patients with previous thrombosis from most clinical trials.1\n\nThe correlation between cancer and venous thromboembolism (VTE) has been extensively established and is known to have deleterious effects on both morbidity and mortality among cancer patients.5\n\nVTE, including pulmonary embolism (PE) and deep vein thrombosis (DVT), as well as arterial thromboembolism, rank as the second most common cause of death in cancer patients.1 Moreover, cancer patients with superficial vein thrombosis (SVT) face a high risk of death comparable to those with DVT.6\n\nPreventing VTE is an essential component of managing lung cancer patients to improve their prognosis. Therefore, several medical scoring systems have been developed to predict the risk of VTE, such as the Khorana, Caprini, Vienna CATS, PROTECHT, and COMPASS-CAT scores.7 Nevertheless, these risk prediction scores have been developed with inherent limitations and have undergone limited external validation in lung cancer patients. Additionally, their applicability to the Tunisian population is restricted. Consequently, it is a priority to develop and validate a score specifically tailored to the Tunisian population. This will facilitate the guidance of prophylactic anticoagulation in patients at risk of VTE.\n\n\nMethods\n\nConsidering the objective of reducing the risk of VTE by 10% through the implementation of a VTE predictive score to guide prophylactic anticoagulation in lung cancer patients, it was determined that a sample size of 360 patients would be necessary to attain statistical significance (power: 0.8; alpha: 0.05), as calculated using a predictive formula.8\n\nWe conducted a multicenter retrospective cohort study during the period between January 2011 and December 2021, involving the records of 410 patients who were diagnosed with lung cancer of all stages and who were hospitalized in the Department 1 or Ibn Néfis Department of Abderrahmane Mami Hospital of Ariana, or in the Pulmonary Department of the Principal Military Hospital of Tunis.\n\nEthics approval\n\nThe study was approved by the ethics committee of abderrahmane mami hospital Under the approval number 27/2023. Written informed consent was collected from the patients. Anonymity was respected during data treatment.\n\nInclusion criteria\n\nWe enrolled all hospitalized patients whose diagnosis of primary lung cancer was histologically confirmed and who required hospitalization.\n\nNon-inclusion criteria\n\nWe did not include the patients whose diagnosis of cancer has not been histologically confirmed or whose diagnosis of secondary lung cancer has been retained.\n\nExclusion criteria\n\nPatients with prior history of other malignant tumors, acute myocardial infarction, and acute stroke were excluded from the study. Patients with inoperable records (many missing data) were also excluded.\n\nThe data collected included: demographic and clinicopathological data (age, gender, BMI, WHO status, smoking, hypertension, diabetes, dyslipidemia, coronary artery disease, heart failure, and surgical history), biological data (blood cell count (White blood cells, Hemoglobin, Platelets), C-reactive Protein (CRP), and creatinine), lung cancer data (histological type, TNM classification, length of follow-up, surgical treatment, chemotherapy, regimen of chemotherapy (platinum-based chemotherapy), and radiation), and VTE data (occurrence of PEor DVT, and the time to their onset).\n\nThe event is defined as the occurrence of a VTE, which includes DVT, SVT, and/or PE.\n\nSVT is a thrombosis occurring in a superficial vein. It is usually caused by an inflammatory reaction in the wall of the thrombosed vein. The diagnosis is strongly suggested clinically and confirmed by Doppler ultrasound.\n\nDVT is characterized by the presence of a blood clot that completely or partially obstructs the blood flow in the deep venous system, most frequently in the lower limbs. The diagnosis is based on venous Doppler ultrasound.\n\nA PE occurs when a thrombus (blood clot) obstructs an artery in the lung, resulting in impaired blood flow. The diagnosis of PE is confirmed through a thoracic angio-scan.\n\nThe patients were randomly allocated to two groups: the development group (182 patients) and the validation group (199 patients). The development group was used to establish the scoring system, while the validation group was used to validate the developed score.\n\nRandom numbers were generated based on the sequence of medical record numbers, and subsequent grouping was determined by ranking the random numbers using SPSS software. Data entry and analysis were conducted using IBM SPSS 23.0 software. Categorical variables were analyzed by calculating frequencies and percentages, and Pearson’s Chi-squared test was employed for frequency comparisons.\n\nThe score development process comprised two steps using the development group’s database. Firstly, a univariate analysis was conducted to identify the risk factors for preoperative VTE. Secondly, variables with a significance level of p<0.05 from the first step were entered into a multivariate analysis using multiple binary logistic regressions. Odds ratios (ORs) and 95% confidence intervals were calculated. Independent variables were included in the regression model if their significance level was less than 0.25. Variables with a significance level of p<0.05 and a 95% confidence interval were considered predictive factors for VTE risk and constituted the components of the score. The weighting of these factors was determined by dividing the β coefficients by the absolute value of the smallest regression coefficient, rounding the result to the nearest integer. The sum of the weighted factors constituted the patient’s total risk score, which was calculated for both the development and validation groups to classify patients into risk groups.\n\nThe cut-offs were fixed using the ROC curve analysis (receiver operating characteristic curve). Once a cut-off has been specified, the calculation of the predicted incidence allows a better classification of the patients into risk groups (low, moderate, and high).\n\nWhen our risk score was developed, its validation was done over two steps. The first step consists of the evaluation of the internal validity, which was done by studying the discrimination using the C statistic via the ROC curve analysis and the calibration via the Hosmer-Lemeshow test. The second step was based on the evaluation of the external validity. A total of 1000 bootstrap samples were selected from the database of the validation group to recalculate the discrimination and calibration of the risk model. For all statistical tests, the two-sided significance level was set at 0.5.\n\n\nResults\n\nA total of 381 patients were included in our study after the exclusion of 20 patients (Figure 1). Thirty-nine patients had lung resection. Three hundred and nine patients (81.8% of cases) received first-line chemotherapy, which was combined with curative radiotherapy in 32.4% of cases.\n\nThe incidence of VTE was 26.8% (102/381). It was similar in the development and validation groups (25.8% vs 27.6%; p=0.690). There was no significant difference in the incidence of pulmonary embolism (PE) (14.3% vs 14.6%; p=0.954), DVT (12.6% vs 12.5%; p=0.966), or SVT (1.1% vs 1.5%; p=0.731) between the 2 groups (Figure 2).\n\nThe main characteristics of the two databases are summarized in Table 1.\n\nThere is a statistically significant correlation between lymph node status (≥N2), presence of metastasis, active smoking, surgical history, hypertension, coronaropathy, and the occurrence of VTE.\n\nThe univariate comparison for the identification of potential risk factors for VTE is detailed in Table 2.\n\nDetermination of risk score system items\n\nVariables with a significance level of p<0.250 were entered into the binary logistic regression.\n\nA hemoglobin level <10 g/l, the presence of metastases, the histological type of poorly or undifferentiated NSCLC, and active smoking are the significant variables (p<0.05; therefore, they have been selected as the items of the score.\n\nBased on the weight of the different regression coefficients, we established a risk score system as follows (Table 3):\n\nThe risk of VTE was significantly correlated with the risk score in the development group (Pearson contingency coefficient=26.757, p<10-3).\n\nDetermination of risk groups\n\nThe cut-off of our VTE predictive score was identified via the ROC curve. Indeed, a total score <2 allows the classification of patients in the “low risk of VTE” group. The other risk classes were developed on the basis of the predicted incidence of VTE.\n\nAs a result, patients were classified into 3 risk groups (Table 4): “low risk” (score 0-1 [predicted incidence <29%, n=92]), “moderate risk” (score 2-3 [predicted incidence 29-43%, n=85]), and “high risk” (score 4-5 [predicted incidence >43%, n=5]).\n\nThe incidence of VTE according to the 3 risk classes in the development group and in the validation group is detailed in Figure 3.\n\nThe percentages of VTE in “low-risk” and “moderate-risk” patients are similar between the development and validation groups (low-risk: 13% vs 17.8%, moderate-risk: 35.3% vs 38.4%; respectively). In contrast, the percentage of VTE in “high-risk” patients is much higher in the development group compared to the patients in the validation group (100% vs 36.3%, p=0.012)\n\nInternal validation\n\nIn the development group, the risk score system has good discrimination. It can distinguish “high-risk” from “low-risk” VTE patients (c statistic=0.703 [0.618-0.789], (Figure 4 A)). This prediction model also showed good calibration according to the Hosmer-Lemeshow test (χ2=2.381, p=0.882).\n\nExternal validation\n\nThe risk score system shows low discrimination in the validation group (c-statistics=0.641 [0.557-0.726], (Figure 4)).\n\nHowever, despite being poorly discriminating, it was well calibrated according to the Hosmer-Lemeshow test (χ2= 6.250; p=0.396).\n\nConsidering the aforementioned classification, patients in the validation group were classified into 3 risk groups (Table 4): “low risk” (score 0-1 [predicted incidence <29%, n=101), “moderate risk” (score 2-3 [predicted incidence 29-43%, n=86]), and “high risk” (score 4-5 [predicted incidence >43%, n=12).\n\n\nDiscussion\n\nWe created a novel prediction model to assess the risk of venous thromboembolism (VTE) in patients diagnosed with lung cancer in the development cohort. Subsequently, we conducted an external validation of the model using a separate validation cohort. Our study involved a total of 381 patients, and our findings indicated that the prevalence of VTE in lung cancer patients was 26.8%, surpassing the rates reported in previous studies.9–11\n\nThe variability in the incidence of VTE is due to several risk factors. In our study, lymph node status (≥N2), presence of metastasis, active smoking, surgical history, hypertension, and absence of coronaropathy were correlated with the occurrence of VTE. However, in other Tunisian studies, TNM stage IV and non-small squamous carcinoma were associated with high VTE incidence.9,10\n\nThus, to reduce the occurrence of VTE, it is imperative to identify and assess all possible risk factors while determining the appropriate prophylactic measures for these patients. Various predictive models have been suggested to anticipate VTE occurrence in individuals with lung cancer. These models have incorporated several factors based on existing literature, and it is important to consider biomarkers associated with thrombosis.12\n\nThe Khorana risk score, a widely recognized predictive scoring system, categorizes cancer patients into distinct risk groups and identifies a high-risk group for thromboprophylaxis. It is considered the prevailing and valuable tool for predicting VTE in the cancer population.12,13\n\nThe majority of factors included in various risk scoring systems have been incorporated into our risk score system. However, we did not include the D-dimer test, despite its significance, due to the limited number of observed values in our dataset.13 and, most importantly, due to the unavailability of this test in our current practice.\n\nFurthermore, these aforementioned predictive risk scores were developed using data from patients diagnosed with various types of cancer, whereas our scoring system is specifically tailored to lung cancer patients.13 Thus, lung cancer data were included in our score but not adopted in most models. Some of these variables were present in our definitive risk score (i.e., the presence of metastasis and histological type of poorly or undifferentiated NSCLC).\n\nIn previous retrospective studies reported in the literature, adenocarcinoma has been identified as a strong predictor of VTE onset.13 Blom et al. conducted a study involving 537 NSCLC patients to investigate thrombotic risk and observed a 20-fold higher risk of VTE compared to the general population. Among the patients, those with adenocarcinoma had a three-fold higher risk (incidence=66.7%) than those with squamous cell carcinoma of the lung (incidence=21.2%).14 Similarly, in another cohort of 493 NSCLC patients, Tagalakis et al. reported a high incidence of DVT (13.6%).15 However, in our study, we found that poorly or undifferentiated NSCLC was associated with a higher prevalence of VTE, while adenocarcinoma was not predictive of the occurrence of VTE.13 Blom et al.14,15\n\nBody mass index (BMI), which is incorporated in both the Khorana score and Caprini VTE risk assessment, was integrated into our risk system. However, we used a lower cut-off (≥25 kg/m2) possibly due to the prevalence of poor nutritional status among lung cancer patients.5,16\n\nThe hemogram parameter (hemoglobin, platelets, and leukocytes) included in the other risk score models was used in our study. Patients with a hemoglobin level <10 g/l had a four-fold higher risk of VTE. Other biomarkers, namely CRP and creatinine, were also included in our system; these biomarkers were not used by most VTE risk models. A cohort study investigating VTE risk among 3159 patients with newly diagnosed solid tumors concluded that elevated CRP and creatinine levels were predictive of VTE.17\n\nOur scoring system incorporates cancer therapy and surgery as variables. Previous studies have demonstrated that cancer therapy, including chemotherapy, antiangiogenic therapy, and hormonal therapy, increases the risk of VTE.18–21 Christensen et al., after reviewing 19 studies involving 10,660 patients with primary lung cancer undergoing curative-intent operations, found that the risk of VTE appears to be highest during the early postoperative period, with a subsequent decrease in risk.22\n\nAs mentioned earlier, since risk factors vary from one population to another, there is a need to develop a risk score system specific to our population, in order to assist healthcare practitioners in developing appropriate prophylactic strategies for patients at risk of developing VTE.\n\nAfter conducting logistic regression analysis in our study, we identified four items that were included in our risk score system. To the best of our knowledge, this risk score represents the first attempt to predict the potential incidence of VTE specifically for Tunisian patients with lung cancer. The developed risk score suggests that the incidence of VTE is expected to increase exponentially.\n\nScores below 2 were associated with a low risk of VTE, while scores of 4 or higher were associated with a high risk. The discriminant validity of this VTE score system was confirmed in the validation group, although there was a moderate decrease in predictive power. However, the classification of patients based on the score remained clinically meaningful. The notable variation in prognosis among the three risk groups should aid physicians in determining the appropriate therapeutic approach. Therefore, we strongly recommend thromboprophylaxis for Tunisian patients with moderate and high VTE risks.\n\nDespite its poor predictive discrimination, this score presented several strengths. Firstly, to our knowledge, it is the first risk prediction model that included the occurrence of SVT as a predictable event. In fact, our decision to add SVT among outcomes was not arbitrary but based on several studies. A recent study conducted in 2022 highlighted the significance of SVT as a condition and revealed that patients with cancer and SVT are at an increased risk of thromboembolic complications.23 In addition, Galanaud et al. suggested that cancer patients with SVT exhibit a poor prognosis, comparable to those with cancer-related DVT, with a heightened risk of recurrence of DVT-PE.\n\nSecondly, we believe that this score can be applied to other populations whose characteristics are quite similar to those of our Tunisian population and with poor means on board, especially in underdeveloped countries. Nevertheless, it is essential to perform external validation of this score using data from diverse populations in order to ensure its generalizability and reliability.\n\nThe present study has several limitations that should be acknowledged. First, this study was based on 381 patients from three tertiary centers in northern Tunisia. Given the limited sample size, it is important to note that our patients may not adequately represent the diversity of our population. Secondly, we did not conduct an assessment of the reproducibility of our risk score in the prospective validation cohort. Thirdly, it is important to consider that personal and family history of VTE, as well as the use of anticoagulant or antiplatelet treatment at the time of lung cancer diagnosis, could potentially impact our findings. Finally, it should be noted that the cut-off values for the potential VTE variables included in our risk model were determined based on clinical experience or existing literature, rather than individualized threshold values determined by ROC curve analysis.\n\nWe have tried to respond to a need specific to the characteristics of our country where the economic crisis makes it very difficult to provide care according to international standards. Our score contains simple items available to any Tunisian practitioner.\n\nThe collection of the four items is straightforward: smoking history can be obtained through an interview, a complete blood count (CBC) is a readily available test in Tunisia, even in primary care settings, determining the histological type, and conducting staging assessments are commonly practiced.\n\n\nConclusion\n\nIn conclusion, the frequency of VTE in this study was high, at 26.8%. A predictive score for VTE was developed and validated by including epidemiological, clinical, and biological data. This score, despite its low discrimination, has a good positive and negative predictive value for a moderate risk of VTE. The stratification of risk in this newly developed risk system may guide the clinician in prescribing preventive treatment for VTE. However, our study has limitations, particularly the retrospective nature of the analysis and the small sample size. Hence, the need to conduct a prospective study on a nationwide scale for the validation of this score.", "appendix": "Data availability\n\nFigshare: Rouis, Houda (2023). Development and Validation of a Risk Score System for Early Prediction of Venous Thromboembolism in Patients with Lung Cancer. figshare. Dataset. https://doi.org/10.6084/m9.figshare.23582829.v1. 24\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nAll authors would thank Mr Khalaf Chofri for the English assistance.\n\n\nReferences\n\nKhorana AA, Palaia J, Rosenblatt L, et al.: Venous thromboembolism incidence and risk factors associated with immune checkpoint inhibitors among patients with advanced non-small cell lung cancer. J Immunother Cancer. 1 janv 2023; 11(1): e006072. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBagchi A, Khan MS, Saraswat A, et al.: Increased Incidence of Thrombotic Complications With Non-small Cell Lung Cancer Necessitates Consideration of Prophylactic Anticoagulation in Young Individuals. Cureus. sept 2021; 13(9): e17769. PubMed Abstract \| Publisher Full Text\n\nKhorana AA, Kuderer NM, McCrae K, et al.: Cancer associated thrombosis and mortality in patients with cancer stratified by khorana score risk levels. Cancer Med. nov 2020; 9(21): 8062–8073. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTimp JF, Braekkan SK, Versteeg HH, et al.: Epidemiology of cancer-associated venous thrombosis. Blood. 5 sept 2013; 122(10): 1712–1723. Publisher Full Text\n\nKhorana AA: Venous thromboembolism and prognosis in cancer. Thromb. Res. juin 2010; 125(6): 490–493. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGalanaud JP, Blaise S, Sevestre MA, et al.: Long-term outcomes of isolated superficial vein thrombosis in patients with active cancer. Thromb. Res. nov 2018; 171: 179–186. Publisher Full Text\n\nComparison of risk prediction scores for venous thromboembolism in cancer patients: a prospective cohort study - PMC.[cité 5 juin 2023]. Reference Source\n\nPourhoseingholi MA, Vahedi M, Rahimzadeh M: Sample size calculation in medical studies.\n\nRacil H, Laaribi G, Cherif H, et al.: Lung cancer with venous thrombo-embolism: clinical characteristics. Tunis. Med. juill 2011; 89(7): 616–620.\n\nKetata W, Moussa N, Bahloul N, et al.: MALADIE VEINEUSE THROMBOEMBOLIQUE ET CANCER BRONCHIQUE: A PROPOS D’UNE SERIE TUNISIENNE VENOUS THROMBOEMBOLISM AND LUNG CANCER: ABOUT A TUNISIAN SERIES.\n\nChew HK, Davies AM, Wun T, et al.: The incidence of venous thromboembolism among patients with primary lung cancer. J Thromb Haemost. 1 avr 2008; 6(4): 601–608. PubMed Abstract \| Publisher Full Text\n\nDi W, Xu H, Xue T, et al.: Advances in the Prediction and Risk Assessment of Lung Cancer-Associated Venous Thromboembolism. Cancer Manag Res. 31 déc 2021; 13: 8317–8327. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLi Z, Zhang G, Zhang M, et al.: Development and Validation of a Risk Score for Prediction of Venous Thromboembolism in Patients With Lung Cancer. Clin Appl Thromb Hemost. 12 mars 2020; 26: 1076029620910793.\n\nBlom JW, Osanto S, Rosendaal FR: The risk of a venous thrombotic event in lung cancer patients: higher risk for adenocarcinoma than squamous cell carcinoma. J Thromb Haemost. oct 2004; 2(10): 1760–1765. PubMed Abstract \| Publisher Full Text\n\nTagalakis V, Levi D, Agulnik JS, et al.: High risk of deep vein thrombosis in patients with non-small cell lung cancer: a cohort study of 493 patients. J Thorac Oncol. août 2007; 2(8): 729–734. Publisher Full Text\n\nCaprini JA: Risk assessment as a guide for the prevention of the many faces of venous thromboembolism. Am J Surg. janv 2010; 199(1 Suppl): S3–S10. PubMed Abstract \| Publisher Full Text\n\nHaltout J, Awada A, Paesmans M, et al.: Predictive factors for cancer-associated thrombosis in a large retrospective single-center study. Support Care Cancer. avr 2019; 27(4): 1163–1170. PubMed Abstract \| Publisher Full Text\n\nAndo Y, Hayashi T, Sugimoto R, et al.: Risk factors for cancer-associated thrombosis in patients undergoing treatment with immune checkpoint inhibitors. Investig New Drugs. août 2020; 38(4): 1200–1206. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKhorana AA, Francis CW, Culakova E, et al.: Risk factors for chemotherapy-associated venous thromboembolism in a prospective observational study. Cancer. 15 déc 2005; 104(12): 2822–2829. Publisher Full Text\n\nNalluri SR, Chu D, Keresztes R, et al.: Risk of venous thromboembolism with the angiogenesis inhibitor bevacizumab in cancer patients: a meta-analysis. JAMA. 19 nov 2008; 300(19): 2277–2285. Publisher Full Text\n\nBehrendt CE, Ruiz RB: Venous thromboembolism among patients with advanced lung cancer randomized to prinomastat or placebo, plus chemotherapy. Thromb Haemost. oct 2003; 90(4): 734–737. PubMed Abstract\n\nChristensen TD, Vad H, Pedersen S, et al.: Venous thromboembolism in patients undergoing operations for lung cancer: a systematic review. Ann Thorac Surg. févr 2014; 97(2): 394–400. PubMed Abstract \| Publisher Full Text\n\nHirmerová J, Seidlerová J, Šubrt I, et al.: Prevalence of cancer in patients with superficial vein thrombosis and its clinical importance. J Vasc Surg Venous Lymphat. Disord. janv 2022; 10(1): 26–32. PubMed Abstract \| Publisher Full Text\n\nRouis H: Development and Validation of a Risk Score System for Early Prediction of Venous Thromboembolism in Patients with Lung Cancer. Dataset. figshare. 2023. Publisher Full Text" }	[ { "id": "253369", "date": "15 May 2024", "name": "Ping Wang", "expertise": [ "Reviewer Expertise Lung cancer and VTE", "lung cancer and infection." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nMany literatures have reported that the incidence rate of tumor with VTE is significantly higher than that of other diseases, and the incidence rate of lung cancer with VTE is higher. There are many studies on the risk factors of lung cancer combined with VTE, and many suggest that pathological types, stages, and other factors are related to the occurrence of VTE. This manuscript introduces several processes for establishing a VTE screening model, but some issues still need to be discussed.\n1. The enrolled patients have a large annual span of nearly ten years. The treatment of lung cancer has also undergone significant changes over the past decade. When were patients enrolled in each group? How to maintain consistency in the inclusion criteria for patients over the past decade, especially whether current immune checkpoint inhibitors and anti angiogenic drugs have an impact on VTE?\n2. What is the difference between chemotherapy and platinum based chemotherapy? Does chemotherapy include chemotherapy with platinum containing drugs?\n3. You collect relevant data and blood tests for enrolled patients. At what time point were the BMI, WBC, Hb, Platelets, Creatinine, and CRP data collected from patients? Especially for non VTE patients, which time point data is appropriate?\n4. Is there any basis researches for the predicted score in Table 3? The P-value for active smoking is the lowest, why is it only 1 point, while the Historical type poor or unidentified NSCLC is 2 points?\n5. In univariate analysis the result of hemoglobin level<10 g/l was P=0.228, while in multivariate analysis was P=0.039? What are the criteria for incorporating multiple factor analysis and comparison?\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nI cannot comment. A qualified statistician is required.\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "335912", "date": "06 Nov 2024", "name": "Georgia Gomatou", "expertise": [ "Reviewer Expertise Medical Oncology", "Thoracic Oncology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis is an original study describing a prediction score of VTE in patients with lung cancer. In general, the manuscript is well-written and the methods and results are clearly presented. My suggestions in order to improve the paper: 1. Why are the non-inclusion criteria described seperately from exclusion criteria? Please combine. 2. The limitations should be included in the Discussion section (last paragraph of Discussion) and not in the Conclusions. 3. In Figure 1 you use the term 'randomization'. However, the process of randomly allocating patients in each group is not randomization. Please delete the term.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1388
https://f1000research.com/articles/12-1387/v1	20 Oct 23	{ "type": "Research Article", "title": "Comparative evaluation of platelet rich fibrin matrix (PRFM) membrane and collagen membrane with demineralized freeze-dried bone allograft (DFDBA) in the treatment of mandibular class II furcation defects: A randomized controlled trial", "authors": [ "Dr. Chitrika Subhadarsanee", "Dr. Prasad Dhadse", "Dr. Pavan Bajaj", "Dr. Mosami Chimote", "Dr. Kiran Sethiya", "Dr. Komal Bhombe", "Dr. Safiya Hassan", "Dr. Ranu Oza", "Dr. Prasad Dhadse", "Dr. Pavan Bajaj", "Dr. Mosami Chimote", "Dr. Kiran Sethiya", "Dr. Komal Bhombe", "Dr. Safiya Hassan", "Dr. Ranu Oza" ], "abstract": "Aim- The aim of the study was to compare the effectiveness of platelet rich fibrin matrix (PRFM) membrane with collagen membrane (Colo Gide) in combination with demineralized freeze-dried bone allograft (DFDBA) in the treatment of mandibular Class II furcation defects. Methods- This randomized, parallel designed, controlled, clinical investigation was conducted in 24 subjects (15 male and 9 female) having Class II furcation defects either buccally or lingually. The test group was treated with DFDBA and PRFM membrane while the control group was treated with DFDBA and collagen membrane. The clinical measurements such as plaque index (PI), papillary bleeding index (PBI), pocket probing depth (PPD), relative attachment level (R-CAL) and relative gingival marginal level (R-GML) were measured at baseline and six months. Radiographic parameters, such as vertical defect depth (VDD), horizontal defect depth (HDD) and defect width (DW) were measured using cone beam computed tomography taken at baseline, three and six months. Student’s paired t-test was utilized to analyse data from the day of surgery to six months. A comparison of both groups at baseline and six months was achieved by student’s unpaired t-test. Result-10 sites in test group (83.33%) showed the advancement from class II to class I compared to eight sites in control (66.66%). Remaining defects in test group n=2 (16.66%) and control group n=4 (33.33%) showed marked reduction in horizontal defect depth compared to baseline. No complete closure of the defect was seen in either group. Conclusion- When treating class II furcation defects, the use of PRFM membrane combined with DFDBA seems to be advantages with regards to collagen membrane. The presented set up seems feasible with regards to randomization, acceptance, retention and achievement of satisfactory outcomes.", "keywords": [ "Class II Furcation", "Guided tissue regeneration", "PRFM", "DFDBA" ], "content": "Introduction\n\nInvasion of furcation areas in multirooted teeth presenting bifurcation and trifurcation always poses continuous challenge to the clinician owing to complex furcal anatomy, difficulty to perform adequate personal oral hygiene and effective periodontal instrumentation.1 Thus, the prognosis of a tooth with a furcation involvement (FI) is always bleak.2,3 Depending on the severity of the furcation defect, a variety of therapeutic options such as odontoplasty,4 furcationplasty,5 tunnelling6 and guided tissue regeneration (GTR) employing different bone grafts,7 barrier membranes8 alone or in combination are available for treatment. Recently, platelet concentrates containing growth factors9 have also been investigated.10 GTR includes the elimination of both epithelial and connective-tissue cells of the gingiva from denuded root surface, allowing periodontal ligament (PDL) or alveolar bone cells to repopulate the wound region.11 When compared to monotherapeutic algorithms, the use of a combination therapeutic strategy (i.e., bone-replacement graft, barrier with or without biologics) provides an added benefit and excellent predictability for regeneration of periodontium in furcation defects.6,9,12,13 At this time, there is no specific regenerative material that is regarded the gold standard for the management of furcation defects.14 Clinicians are still investigating for a “off-the-shelf” material that can substitute and/or improve grafting procedure while also providing improved, additional reliable clinical outcomes than existing bone scaffolds and matrices.15 In recent years, growth factors (GFs) are in the limelight in the craniomaxillofacial and periodontal sectors.16,17\n\nDecalcified freeze-dried bone allograft (DFDBA) works as a source of osteo-inductive substances while providing an osteoconductive surface.18 Bone morphogenic proteins (BMPs) such as BMP 2, 4, and 7 are found in DFDBA and serve to trigger osteo-induction.19 Thus, allograft proteins manufactured commercially have the ability to alter cell activity in vivo.20 Clinical research investigations have shown that using DFDBA in human intraosseous21,22 lesions improved clinical attachment and bone level, with histological evidence of new attachment formation.23,24\n\nGTR effectively prevents tissue and bone degradation while also stimulating the development of new tissue and bone.25 In order to cover the region where the regeneration process will take place, a physical barrier (membrane) with the correct shape and position is required. GTR was originally reported to be effective in regenerating damaged periodontal tissues in class II furcation involvement.26 Their biggest drawback is their lack of stiffness, which limits their ability to create space and necessitates the use of a scaffold. Collagen membranes, on the other hand, can be employed alone for alveolar bone defects such as bone dehiscence and fenestration defects that do not require further fixation and stability.27 Furthermore, because they degrade quickly, they may not be able to satisfy the time requirements for good tissue growth.\n\nA new generation of platelet-rich fibrin matrix (PRFM) is a concentrate of platelets that require no biological components for its preparation (bovine thrombin).28 PRFM structure looks like natural fibrin and promotes cell motility, proliferation, and cycle creation.29 On the first day, levels of platelet-derived growth factor (PDGF), vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and transforming growth factor beta (TGF-β) are increased, then gradually tend to decrease the next day.30,31 PRFM also exhibits fibrin properties, such as a denser and more flexible macroscopic structure and a more natural platelet distribution.32,33 Because of these variables, this preparation behaves more like a natural clot, releasing low concentrations of growth factors over a longer period of time.30,34 To the best of our knowledge the use of PRFM membrane in periodontal regeneration is scarce and limited.\n\nThree dimensional (3D) imaging, such as cone beam computed tomography (CBCT), can offer details regarding faults that aren’t visible in two dimensional 2D photos.35 It was employed to compensate for the limitations of two dimensional (2D) scanners and to clearly show the horizontal component of the furcation. It can be used to evaluate treatment outcomes, particularly to check healing following grafting or regeneration. This imaging technique may also be used to measure the gingival tissue and the dimensions of the dentogingival unit.36,37 CBCT is used to assess and plan treatment for molars with furcation involvement by revealing marginal bone contouring, intra-bony and furcation defects.38\n\nTherefore, the following investigation has been undertaken to compare the effectiveness of PRFM membrane with that of collagen membrane (Colo Gide) in combination with DFDBA in the treatment of defects with Class II furcation, both clinically and radiographically.\n\n\nMethods\n\nThe current randomized, parallel designed, controlled, clinical study was conducted in 24 subjects (12 in each group) as determined by sample size calculation using two means with equal variance (15 male and 9 female), with moderate to advanced chronic periodontitis having Class II furcation defects either buccally or lingually with age range of 25 to 50 years with a mean age of 40.5 were selected from Department of Periodontics. Subjects were recruited on an outpatient basis and separated into groups by computer generated random number table. This study design protocol was ethically passed by the “Institutional Ethical Committee” (DMIMS (DU)IEC/2017-18/6730). All participants were informed verbally and written informed consent was obtained for the surgical procedure. The CONSORT Checklist and guidelines were followed for this study.39,40\n\nHealthy subjects without any systemic illness with horizontal probing depth (HDD) and vertical furcation probing depth (VDD) of ≥3 mm, proximal bone height coronal to the inter-radicular bone level and radiographic evidence of furcation defects in the molars (buccal, lingual, mesiobuccal or distobuccal) were included. To match all the characteristics of included subjects at baseline the age group of the subjects were in the range of 25 to 50 years.\n\nSubjects with poor oral hygiene following “etiotropic/phase I periodontal therapy” and exhibiting plaque scores greater than or equal to 1, mobile teeth, any systemic diseases, suspected or known allergies to drugs or study materials, use of tobacco in any form, immunocompromised subjects, alcoholics, lactating or pregnant women, and subjects with compromised immune systems were excluded.\n\nPrior to surgery, after initial treatment (scaling and root planing), the selected defects were randomly assigned to the test and control groups, each of which had 12 defects, using a computer-generated random number table (Calculator Soup® Online Calculators). During the allocation system, both the participants and the examiner were blinded. In contrast to the control group, which received treatment from collagen membrane (Colo Gide) and DFDBA (n=12), the test group was handled by PRFM and DFDBA. The assessment of outcomes involved re-evaluating patients after three and six months of treatment.\n\nTo standardise the probe location and angulations to assess the horizontal and vertical probing depth before the procedure and after six months, specially designed occlusal acrylic stents were created (Figure 1). For the purpose of creating a cast model of both jaws, alginate impressions were made. On the cast model, an acrylic stent was made. The treated tooth’s occlusal surface and at least one tooth’s distal and mesial surfaces were all covered by the occlusal stent. A reference point was established for the placement of the periodontal probe in the deepest part of the affected tooth. The stent’s apical border was used as a fixed reference point. (Fixed reference point was made at baseline to replicate the same position at the following visit, thus avoiding the variability in measurement).\n\nPlaque index (PI) by Silness and Loe,41 papillary bleeding index (PBI) by Saxer and Mühlemann,42 probing pocket depth (PPD), relative attachment level (R-CAL), and relative gingival marginal level (R-GML) were the clinical parameters that were recorded. On the first day of therapy and three and six months following the procedure, all clinical signs were assessed. The radiographic bone fill was the study’s primary outcome, while improvements in relative attachment level (R-CAL) and probing pocket depth (PPD) were its secondary outcomes.\n\nPI by Silness and Loe was graded from score 0-3 according to the accumulation of microbial plaque with respect to free gingival margin.41\n\nPBI by Saxer and Mühlemann was graded from score 0-4 according to the intensity of any bleeding with UNC 15 probe which was inserted into the gingival sulcus at the base of the papilla on the mesial aspect, and then moved coronally to the papilla tip. This is repeated on the distal aspect of the papilla.42\n\nA UNC-15 (University of North Carolina, Hu-Friedy, Chicago, USA) probe was measured from the inferior border of the acrylic stent referred as relative gingival marginal level (R-GML). At three locations on each furcation surface—distal, mesial line angle, and midbuccal or midlingual surface—the relative attachment level (R-CAL) was measured using the distance between the base of the pocket and the inferior border of the stent using the UNC-15 probe. The UNC-15 probe is used to measure PPD from the gingival margin to the base of the pocket. The vertical probing depth (VPD), measured from the pocket base to the gingival margin, was estimated (Figure 1).43,44 Horizontal probing depth (HPD) of furcation was measured by a curved color-coded furcation probe (GDC Double End Probes Nabers Color Coded # 6 (Pq2n)) with 0-3, 3-6, 6-9 and 9-12 mm markings. The width of keratinized gingiva (WKG) was calculated from apical most point of mucogingival junction to the crest of gingival margin using a UNC-15 probe. All the probing measurements were noted at baseline (pre-operation), three and six months post-operatively.\n\nThe caliper included with in the cone beam computed tomography (CBCT) (Planmeca Promax 3D G-XR-109125, software- Romexis viewer) was used to evaluate radiographic parameters. The vertical defect depth (VDD) was measured in sagittal view from the fornix to the base of the defect, and the horizontal defect depth (HDD) was measured in axial section from a tangent line linking the greatest convexities of the mesial and distal roots to the deepest area of the defect. In sagittal view, the defect width (DW) was assessed.\n\nAll study participants underwent a pre procedural mouth rinse with a 0.2% chlorhexidine gluconate solution for one minute. The surgical procedure was conducted while maintaining asepsis. The area was anaesthetized by nerve block and infiltration using 2% Xylocaine with a 1:80,000 epinephrine concentration (Ligno-Ad local anaesthetic, Proxim Remedies, India). Surgical blade no. 15 was used to create intra-crevicular incisions on the lingual or buccal surfaces of the affected tooth. The interdental papillae were secured by the interproximal incisions in order to achieve primary wound closure. The flap also covered the tooth’s mesial and distal portions. With a periosteal elevator (24 G Hu-Friedy, USA) on the affected site, a mucoperiosteal flap was raised to reveal the underlying defect margin (Figure 2).43,44 With the aid of hand instruments like universal curettes (GDC Universal Curettes 4r/4l Posterior SC4R/4L - 3), an ultrasonic instrument (Woodpecker HW-3H), and furcation curettes (GDC Furcation Curette Quetin - Buccal/lingual # 6 (SQBL2 # 6), the denuded root surface with the furcation dome was debrided. The root surfaces and furcation flaws were planed until a smooth, firm consistency was reached. After achieving hemostasis and irrigation with physiological saline solution, intraoperative measurements of both HDD and VDD at the furcation site were taken (Figure 3).43,44 1) HDD: The furcation defect’s deepest region was measured horizontally with a UNC 15 probe, and a second UNC 15 probe was put at the prominence of the root surface to bridge the first probe as a reference. 2) Using the furcation fornix as a fixed reference point, the vertical furcation defect measurement (VDD) was taken. The final patient’s eligibility for the trial was confirmed if the furcation defect depth was 3 mm both vertically and horizontally. The trial does not exclude any patients. Following the intraoperative measurements, the sites were then assigned to the test or control group using a computer-generated random number (Research Randomizer, RRID:SCR_008563).\n\nComplete isolation and hemostasis of site was obtained using gauge piece manual pressure application with one finger (Hospital Cotton Woven Fabric, Check/stripes, White) and suction tube (PDD suction tip). After complete debridement of furcation defect, DFDBA was condensed (Figure 4)43,44 and covered by Cologide™ bioabsorbable GTR membrane (Figure 5).44 The membrane will be stabilized and then the flap was sutured (Figure 6).43,44 Periodontal pack was given.\n\nIn test sites PRFM membrane (Figure 5)43 along with DFDBA (Figure 4)43,44 was used. A 10 millilitre sample of blood was collected from antecubital vein and transferred in Merisis PRFM test tubes (Meresis, Laboratory, Bengaluru, Karnataka, India). Then in single time centrifugation (Remi R-8C 16×15 ml Laboratory Centrifuge with Angle Rotor Head) technique PRFM clot was made with 3000 rpm for 10 min. The upper layer of clot was removed and with the help of PRF box PRFM membrane was obtained and placed in the furcation defect.\n\nFollowing the procedure, a non-steroidal anti-inflammatory medication (Ibugesic Plus®, Cipla Pharmaceuticals, India) containing a combination of Ibuprofen 400 mg and Paracetamol 325 mg, as well as the antibiotic Amoxicillin 500 mg three times daily, were prescribed for five days. For 4-6 weeks, it was recommended to all subjects to gargle with 0.2% chlorhexidine gluconate twice daily for one minute. Patients were advised to protect the pack from any harm. After one week, the periodontal dressing and sutures were removed, and recovery was shown. Subjects were instructed to use a gentle toothbrush and cotton pellets to clean the surgical site in an apico-coronal manner. One, three, and six months following treatment, the individuals were recalled.\n\nAll clinical parameters, including PPD, HPD, CAL, and GML, were estimated using the mean and standard deviation (Mean SD) results. Data from the day of surgery to six months were analysed using a paired t-test with students as the subjects. Student’s unpaired t-test allowed for a comparison of the two groups at both their baseline and six-month points in time. The paired t-test used by the students was used to compare the PI and PBI at baseline and after six months. If the probability value (p) is less than 0.05, the difference was ruled non-significant; if it is greater than 0.05, it was considered significant. To determine differences among each group, the Wilcoxon test or paired student’s t-test was used. All hard and soft tissue variables were compared between the test and control groups using the Mann-Whitney test or an independent student’s t-test. In all phases of assessment, a p value <0.05 was considered significant. All data were assessed using SPSS 11.0 (SPSS inc, 2003) (RRID:SCR_002865) software.\n\n\nResults\n\nA total of 24 class II mandibular furcation defects (First molars n=17; second molars n=7) involving either buccal (n=22) or lingual (n=2) surfaces in 18 subjects were treated in this study. Patients involved in the study with age ranged between 25 to 50 years (Male:Female - 15:9). All the clinical and radiographic parameters are provided in the masterdata.45\n\nDuring the time slot of six months, healing was uneventful, all patients reported for post-surgical evaluation therefore the study did not exclude any of the sites. All selected patients reported for the therapy and follow up after three and six months. All the participants reported satisfaction with the treatment given to them.\n\nComparison between baseline PI and PBI score to six-month follow-up revealed a significant decline in both groups. (p<0.05). Low PI score (<1) in both the groups indicated that good plaque control was maintained throughout the study period indicating that the gingival inflammation was reduced and the tissues remained healthy46 (Table 1).\n\nAll the investigated parameters in both groups at baseline were observed to be statistically non-significant (p>0.05), indicating same starting point for both procedures. Clinical parameters VPD, R-CAL, HPD and RGML revealed a significant reduction (p<0.05) after six months compared to baseline in both groups (Tables 2 and 3).\n\nComparison between mean VPD reduction in test (2.5±0.52 mm) and control group (2.33±0.65 mm) at six months indicated non-significant (p-0.43) in both group by 0.16±0.71 mm. Comparison of mean CAL gain among groups at six months indicated no statistically significant difference (0.41±1.08 mm). Comparison of mean CAL gain among groups at three months indicated no statistically significant difference (0.41±1.56 mm) (Table 5). The mean reduction of HPD for test group (0.17±0.62 mm) when compared with control group (2.41±0.66 mm) at six months, found to be statistical non-significance difference (p>0.50). The mean gain of gingival marginal level for test group (0.25±0.62 mm) when compared with control group (0 mm) later at six months, considered be statistical non-significance (p>0.50) (Table 6).\n\n(Mean±SD; in mm).\n\n(Mean±SD; in mm).\n\nRadiographic parameters HDD & DW showed statistical significantly (p<0.001) result at six months compared to baseline in both the groups where VDD signifies non-significant (P- 0.01) result (Table 4). This can be appreciated on radiographic images which can be found in the extended data.47,48\n\nThe mean reduction of HDD for test group (1.56±0.85 mm) in comparison with control group (1.79±0.48 mm) at 6 months, showed statistical non-significance difference (p>0.40). Comparison of mean VDD gain between both groups at 6 months indicated non-significant (p-0.26) difference (0.25±0.70 mm). The mean gain in DW for the test group (0.81±0.40 mm) when compared with control group (0.82±0.16 mm) at the end of study considered statistically non-significance (p-0.93) (Table 7).\n\n(Mean±SD; in mm).\n\nA total of 10 sites in test group (83.33%) showed the advancement from class II to class I compared to eight sites in control (66.66%). Remaining defects in test group n=2 (16.66%) and control group n=4 (33.33%) showed marked reduction in horizontal defect depth compared to baseline. No closure of the defect was seen completely.\n\n\nDiscussion\n\nThe success of furcation treatment is based on eradication of both horizontal and vertical defect components, as well as the improvement of clinical parameters.46 Various therapeutic techniques were created and tested in order to accomplish the anticipated result in the management of class-II mandibular FIs. AAP regeneration workshop13 stated the use of a combination treatment strategy to regenerate the periodontium that appears to have an advantage over monotherapeutic techniques. One of the approaches is GTR that has been established as a successful therapeutic alternative with high predictability for the treatment of different furcation type defects, particularly Class II defects. The current study used a bone graft DFDBA that promotes host’s undifferentiated mesenchymal cells to develop into osteoblasts, resulting in the production of new bone.49\n\nThe mean PPD reduction (0.16±0.71 mm; p=0.43) in our study is supported by evidence, which shows PPD decrease by employing DFDBA with and without PRF in the management of class II furcation defects and showed statistical significant PI score (p<0.001) at six months compared to baseline.17,50 In this study, the test group had a gain RGML (0.25±0.62) than the control group (0). Sharma et al.51 found that the PRF group (0.344±0.086) had similar GML alterations to the OFD group (0.756±0.115) in their study.\n\nAnother criterion is clinical attachment level gain that may be used to justify good clinical periodontal regeneration following periodontal treatment. In the present study, both groups showed no statistically significant difference for mean CAL gain- 0.41±1.08; p=0.20. Observations made in the present study with regards to clinical attachment gain are comparable with results stated in a study conducted by Basireddy et al. (2018)17 and Mehta et al. (2018)50 where statistical non-significant difference found in RHCAL (p= 0.055) with combination therapy. The use of PRFM and collagen membrane along with bone graft has led to a demonstrable CAL gain in furcation defects, thus indicating a regenerative potential of both the membranes. In the present study, gain in HDD, VDD and DW using CBCT in both groups substantiated the bone filling potential of both PRFM membrane and collagen membrane with DFDBA.\n\nThird-generation platelet concentrates include autologous growth factor concentrates as well as PRFM.29 The structural alterations in the platelet concentrate’s fibrin gel are caused by a variety of factors, including thrombin- fibrinogen concentration ratios and protein and ion concentrations, including calcium. It has been observed that not only platelets, but also leukocytes are concentrated as studied in the intricate 3-D design.52 Due to the strong fibrin matrix, PRFM resorbs gradually, allowing for a prolonged release of platelet and leukocyte-derived GFs into the wound region from seven to 23 days.31,34 It speeds up healing, is inexpensive, and may be used on major bone deformities. Microscopically, it is formed of amorphous fibrin and fused strands and has an alternate pattern of thick, non-porous sections, long strands, and bundles with pores. There are also kinked fibres and bundles to be found. It has a very high fibre density on both sides of the membrane when compared to native fibrin clots. PRFM was used as a therapy technique for the first time in furcation type defects, taking into account all of these qualities to reduce the extra expense of the barrier membrane and to prevent bacterial contamination owing to exposure of the membrane. Due to the membrane’s capacity to maintain space, the synergistic impact of membrane and bone graft gave advantages over clinical criteria such as reduction in PD and gain in CAL in the current trial.\n\nStudies stated that horizontal defect depth of 5 mm or more demonstrated less probability of complete closure (52%) and horizontal defects depth around 4 mm or less than presented higher chances of complete furcation defect closure (84%).53 In the present study, presurgical measurement of HPD was from 3-6 mm thus showing lesser chances of complete defect closure.\n\nIn the present study, both the test group (n=10; 83.33%) and the control group (n=8; 66.6%), the proportion of defects changed from class II to class I demonstrated extraordinary regeneration capacity. GTR treatment has been shown in literature to aid in the conversion of class II to class I furcation, hence improving the tooth’s long-term prognosis.54 Compared to grafting alone, combined treatment offers several advantages. It holds the graft in place in the defect, supports the membrane in the proper position, and improves epithelial exclusion. In furcation management, long-term studies showed a gain in attachment of up to 4.5 mm with GTR treatment. In comparison to baseline values, all remaining defects in the test group (n=2; 16.66%) and control group (n=4; 33.33%) demonstrated a substantial reduction in defect depth.\n\nTo the best of our knowledge, no research has investigated the efficacy of DFDBA and PRFM in FI. Therefore, comparison of this combination approach in furcation defects was made. As a result, it stands for an exogenic comparison of DFDBA and PRFM with a well-established surgical technique for furcation defect regeneration.\n\n\nConclusions\n\nIn our study, clinical as well as radiographic evaluations are carried out to evaluate the regenerative potential which limit the assessment. Thus, to govern the stability of the outcomes attained in this study, histological evidence over the long span of time is desirable. Also, larger sample size needed to be planned to quantify the regeneration.\n\nProtocol details - JOURNAL OF CRITICAL REVIEWS ISSN - 2394-5125 VOL 6, ISSUE 6, 2019 Comparative Evaluation of Platelet Rich Fibrin Matrix membrane and Collagen membrane with Demineralized Freeze Dried Bone Allograft in Class II Furcation Defects using CBCT – A Randomized Controlled Clinical Trial DR. CHITRIKA SUBHADARSANEE 1, DR. PRASAD V. DHADSE 2.", "appendix": "Data availability\n\nFigshare: Master chart thesis.xlsx. https://doi.org/10. 6084/m9.figshare.22153985.v1. 45\n\nThis project contains the following underlying data:\n\n- Master chart thesis.xlsx (Data taken from both test and control groups)\n\nFigshare: Surgical procedure for test group. https://doi.org/10.6084/m9.figshare.22559734. 43\n\nThis project contains the following extended data:\n\n- Figure 1-Measurement of VPD using UNC-15 probe. JPG\n\n- Figure 2-Full thickness flap reflection. JPG\n\n- Figure 3-Intrasurgical vertical defect depth measurement. JPG\n\n- Figure 4-DFDBA placed in furcation defect. JPG\n\n- Figure 5-PRFM placed in the furcation defect. JPG\n\n- Figure 6-Flap sutured. JPG\n\nFigshare: Surgical procedure for control group. https://doi.org/10.6084/m9.figshare.22629394. 44\n\nThis project contains the following extended data:\n\n- Figure 1- Measurement of VPD using UNC-15 probe.jpg\n\n- Figure 2- Full thickness flap reflection.jpg\n\n- Figure 3- Intrasurgical vertical defect depth measurement.jpg\n\n- Figure 4- DFDBA placed in furcation defect.jpg\n\n- Figure 5- GTR membrane placed in the furcation defect.jpg\n\n- Figure 6- Flap sutured.jpg\n\nFigshare: Clinical and radiographic evaluation of test group. https://doi.org/10.6084/m9.figshare.22630369. 47\n\nThis project contains the following extended data:\n\n- Pre operative Vertical defect depth and defect width (test group).png\n\n- Pre operative Horizontal Defect Depth (test group).png\n\n- Post operative Vertical Defect Depth and Defect Width (test group).png\n\n- Post operative Horizontal Defect Depth (test group).png\n\nFigshare: Clinical and radiographic evaluation of control group. https://doi.org/10.6084/m9.figshare.22630405. 48\n\nThis project contains the following extended data:\n\n- Pre operative Vertical defect depth and defect width (Control group).png\n\n- Pre operative Horizontal Defect Depth (control group).png\n\n- Post operative Vertical Defect Depth and Defect Width (control group).png\n\n- Post operative Horizontal Defect Depth (control group).png\n\nFigshare: CONSORT checklist. https://doi.org/10.6084/m9.figshare.22300159.v2. 39\n\nThis project contains the following extended data:\n\n- CONSORT_checklist.docx\n\nFigshare: CONSORT Flow Diagram. https://doi.org/10.6084/m9.figshare.22352815. 40\n\nThis project contains the following extended data:\n\n- CONSORT flow diagram.docx\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nperiobasics: Furcation involvement and its management.[cited 2023 Mar 14]. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBone Morphogenic Protein in Demineralized Freeze Dried Bone Allograft - A Possible Reason for its Osteoinduction.2018; 9(6).\n\nSanz M, Dahlin C, Apatzidou D, et al.: Biomaterials and regenerative technologies used in bone regeneration in the craniomaxillofacial region: Consensus report of group 2 of the 15th European Workshop on Periodontology on Bone Regeneration. J. Clin. Periodontol. 2019; 46: 82–91. PubMed Abstract \| Publisher Full Text\n\nAgarwal A, Gupta ND: Platelet-rich plasma combined with decalcified freeze-dried bone allograft for the treatment of noncontained human intrabony periodontal defects: a randomized controlled split-mouth study. Int. J. Periodontics Restorative Dent. 2014; 34(5): 705–711. PubMed Abstract \| Publisher Full Text\n\nHoidal MJ, Grimard BA, Mills MP, et al.: Clinical evaluation of demineralized freeze-dried bone allograft with and without enamel matrix derivative for the treatment of periodontal osseous defects in humans. J. Periodontol. 2008; 79(12): 2273–2280. PubMed Abstract \| Publisher Full Text\n\nBowers GM, Chadroff B, Carnevale R, et al.: Histologic evaluation of new attachment apparatus formation in humans: Part II. J. Periodontol. 1989; 60(12): 675–682. Publisher Full Text\n\nAgarwal A, Manjunath RGS, Sethi P, et al.: Platelet-rich fibrin in combination with decalcified freeze-dried bone allograft for the management of mandibular degree II furcation defect: A randomised controlled clinical trial. Singap. Dent. J. 2019 Dec; 39(01): 33–40. PubMed Abstract \| Publisher Full Text\n\nBashutski JD, Wang HL: Periodontal and endodontic regeneration. J. Endod. 2009; 35(3): 321–328. Publisher Full Text\n\nGottlow J, Nyman S, Lindhe J, et al.: New attachment formation in the human periodontium by guided tissue regeneration Case reports. J. Clin. Periodontol. 1986; 13(6): 604–616. PubMed Abstract \| Publisher Full Text\n\nSbricoli L, Guazzo R, Annunziata M, et al.: Selection of Collagen Membranes for Bone Regeneration: A Literature Review. Materials. 2020 Feb 9; 13(3): 786. Publisher Full Text\n\nChoukroun J, Diss A, Simonpieri A, et al.: Platelet-rich fibrin (PRF): a second-generation platelet concentrate. Part V: histologic evaluations of PRF effects on bone allograft maturation in sinus lift. Oral Surg Oral Med Oral Pathol Oral Radiol. Endodontology. 2006; 101(3): 299–303. Publisher Full Text\n\nReksodiputro MH, Harahap AR, Setiawan L, et al.: A Modified Preparation Method of Ideal Platelet-Rich Fibrin Matrix From Whole Blood. Front. Med. 2021 Aug 13; 8(8): 724488. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSclafani AP: Safety, efficacy, and utility of platelet-rich fibrin matrix in facial plastic surgery. Arch. Facial Plast. Surg. 2011; 13(4): 247–251. PubMed Abstract \| Publisher Full Text\n\nLucarelli E, Beretta R, Dozza B, et al.: A recently developed bifacial platelet-rich fibrin matrix. Eur. Cell. Mater. 2010; 20(1): 13–23. Publisher Full Text\n\nKobayashi E, Flückiger L, Fujioka-Kobayashi M: Comparative release of growth factors from PRP, PRF, and advanced-PRF [published online ahead of print January 25, 2016]. Clin. Oral. Investig.\n\nGole PV, Muhammed N, Patadia SR: Efficacy of autologous platelet rich fibrin matrix in the management of non-healing ulcers. Int. J. Res. Dermatol. 2019; 5(686): 10–18203. Publisher Full Text\n\nSclafani AP, Azzi J: Platelet Preparations for Use in Facial Rejuvenation and Wound Healing: A Critical Review of Current Literature. Aesthet. Plast. Surg. 2015 Aug; 39(4): 495–505. PubMed Abstract \| Publisher Full Text\n\nKaria H, Shrivastav S, Karia AK: Three-dimensional evaluation of the airway spaces in patients with and without cleft lip and palate: A digital volume tomographic study. Am. J. Orthod. Dentofac. Orthop. Off. Publ. Am. Assoc. Orthod. Its. Const. Soc. Am. Board Orthod. 2017 Sep; 152(3): 371–381. Publisher Full Text\n\nPatil SR, Al-Zoubi IA, Gudipaneni R, et al.: A comparative study of cone-beam computed tomography and intrasurgical measurements of intrabony periodontal defects. Int. J. Oral Health Sci. 2018; 8(2): 81. Publisher Full Text\n\nCimbaljevic M, Misic J, Jankovic S, et al.: The Use of Cone-Beam Computed Tomography in Furcation Defects Diagnosis. Balk. J. Dent. Med. 2016 Nov 1; 20(3): 143–148. Publisher Full Text\n\nZhang W, Foss K, Wang BY: A retrospective study on molar furcation assessment via clinical detection, intraoral radiography and cone beam computed tomography. BMC Oral Health. 2018 May 3; 18(1): 75. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSubhadarSanee C, Dhadse PV, Bajaj P, et al.: CONSORT checklist. figshare. [Guidelines].2023. Publisher Full Text\n\nSubhadarSanee C, Dhadse PV, Bajaj P, et al.: CONSORT Flow Diagram. figshare. [Guidelines].2023. Publisher Full Text\n\nLöe H: The Gingival Index, the Plaque Index and the Retention Index Systems. J. Periodontol. 1967; 38(6): Suppl:610–Suppl:616. PubMed Abstract \| Publisher Full Text\n\nSaxer UP, Mühlemann HR: Motivation and education. Schweiz Monatsschrift Zahnheilkd Rev Mens Suisse Odonto-Stomatol. 1975 Sep; 85(9): 905–919.\n\nSubhadarSanee C, Dhadse PV, Bajaj P, et al.: Surgical procedure for test group. [Data]. figshare. 2023. Publisher Full Text\n\nSubhadarSanee C, Dhadse PV, Bajaj P, et al.: Surgical procedure for control group. [Data]. figshare. 2023. Publisher Full Text\n\nSubhadarSanee C, Dhadse PV, Bajaj P, et al.: Master chart thesis.xlsx. [Data]. figshare. 2023. Publisher Full Text\n\nJepsen S, Gennai S, Hirschfeld J, et al.: Regenerative surgical treatment of furcation defects: A systematic review and Bayesian network meta-analysis of randomized clinical trials. J. Clin. Periodontol. 2020; 47: 352–374. Publisher Full Text\n\nSubhadarSanee C, Vijayrao Dhadse p, Bajaj P, et al.: Clinical and radiographic evaluation of test group. [Data]. figshare. 2023. Publisher Full Text\n\nSubhadarSanee C, Dhadse P, Bajaj P, et al.: Clinical and radiographic evaluation of control group. [Data]. figshare. 2023. Publisher Full Text\n\nGajiwala AL, Kumar BD, Chokhani P: Evaluation of demineralised, freeze-dried, irradiated bone allografts in the treatment of osseous defects in the oral cavity. Cell Tissue Bank. 2007; 8(1): 23–30. PubMed Abstract \| Publisher Full Text\n\nMehta D, Deshpande N, Dandekar S: Comparative evaluation of platelet-rich fibrin membrane and collagen membrane along with demineralized freeze-dried bone allograft in Grade II furcation defects: A randomized controlled study. J. Indian Soc. Periodontol. 2018; 22(4): 322–327. PubMed Abstract \| Publisher Full Text\n\nSharma A, Pradeep AR: Autologous platelet-rich fibrin in the treatment of mandibular degree II furcation defects: a randomized clinical trial. J. Periodontol. 2011 Oct; 82(10): 1396–1403. Publisher Full Text\n\nLundquist R, Dziegiel MH, Agren MS: Bioactivity and stability of endogenous fibrogenic factors in platelet-rich fibrin. Wound Repair Regen. Off. Publ. Wound Heal Soc. Eur. Tissue Repair Soc. 2008 Jun; 16(3): 356–363. Publisher Full Text\n\nNovaes-Jr A, Palioto D, Andrade P, et al.: Regeneration of Class II Furcation Defects: Determinants of Increased Success. Braz. Dent. J. 2005 Feb 1; 16: 87–97. Publisher Full Text\n\nFriedmann A, Stavropoulos A, Bilhan H: GTR Treatment in Furcation Grade II Periodontal Defects with the Recently Reintroduced Guidor PLA Matrix Barrier: A Case Series with Chronological Step-by-Step Illustrations. Jornet PL, editor. Case Rep. Dent. 2020 Dec 16; 2020: 1–10. Publisher Full Text" }	[ { "id": "217529", "date": "14 May 2024", "name": "Vidya Sagar", "expertise": [ "Reviewer Expertise PERIODONTICS", "BONE REGENERATION" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nReviewer’s comments\nTitle: Comparative evaluation of platelet rich fibrin matrix (PRFM) membrane and collagen membrane with demineralized freeze-dried bone allograft (DFDBA) in the treatment of mandibular class II furcation defects: A randomized controlled trial\n\nKindly provide reference how the subjects were selected as moderate and advanced chronic periodontitis.\nKindly provide reference for furcation classification.\n\nKindly provide CBCT images as it's primary outcome of the study.\n\nKindly mention the company name of 0.2% chlorhexidine gluconate.\n\nVertical furcation defect measurement (VDD) - kindly justify with reference.\n\nMembrane will be stabilized - replace will by was.\n\nKindly mention company name of periodontal dressing.\n\nKindly provide reference for PRFM preparation.\n\nKindly mention amoxicillin 500mg company name.\n\nReplace CAL by R-CAL and GML by R-GML.\n\nThe present study didn't interpret the results of all the parameters at 3 month which were shown in the tables. Only few parameters have been explained.\n\nTables are not in chronological order.\n\nPlease submit with the above changes.\nThe comments to consider for this study:\nStrengths: The study design, including randomization and a parallel-controlled setup, adds to the validity of the findings. The use of both clinical and radiographic parameters provides a comprehensive evaluation of the treatment outcomes. The study's emphasis on feasibility aspects such as patient acceptance and satisfactory outcomes adds to its clinical relevance.\n\nLimitations: The relatively small sample size of 24 subjects could limit the generalizability of the results. While the study highlights the benefits of PRFM membrane combined with DFDBA, it is essential to consider potential confounding factors that might influence the results, such as patient-related factors or variations in the defect characteristics.\n\nClinical Implications: The findings suggest that PRFM membrane combined with DFDBA could be a promising treatment option for managing mandibular Class II furcation defects. However, further research with a larger sample size and longer follow-up is warranted to establish the broader applicability and efficacy of this treatment approach.\nOverall, the study contributes valuable insights into the comparative effectiveness of PRFM membrane and collagen membrane with DFDBA in the treatment of mandibular Class II furcation defects, emphasizing the potential benefits of PRFM membrane in this context. Further research and analysis are necessary to confirm and expand upon these findings.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1387
https://f1000research.com/articles/12-1386/v1	20 Oct 23	{ "type": "Study Protocol", "title": "Assessment and comparison of age-related three-dimensional variation in the anterior loop of the inferior alveolar nerve for pre-surgical planning", "authors": [ "Abhilasha Ramesh Waghadkar", "Vidya K Lohe", "Vidya K Lohe" ], "abstract": "Background: The mental foramen (M.F.) is crossed anteriorly by the mental neurovascular bundle, then does a U-turn to leave M.F. The anterior loop is a section of the inferior alveolar nerve. Prior to deciding how to operate on the anterior mandible, it is essential to take this anatomic variation into account. Methods: This study will be carried out by studying 70 cone-beam computed tomography scans. The anterior loop will be measured using the PLANMECA proMax Cone Beam Computed Tomography (CBCT) measuring tool. Its measurement specifications include a cylinder-shaped field of view with a voxel size of 300, a voltage in tubes of 90 kV, 6.3 mA tube current, and a 12-second exposure period. The field of view will be the same for all scans to standardize the criteria for selecting images from scans. Conclusions: Compared to multi-slice computed tomography, C.B.C.T. has a number of advantages, including the ability to analyze craniofacial features in three dimensions without distortion or overlapping images. Furthermore, C.B.C.T. is considered the gold standard for assessing bone tissue. Correctly identifying and protecting neurovascular bundles is crucial to preventing sensorineural damage in the interforaminal area. This is because there are significant individual anatomical differences present to varying degrees. Therefore, the present study is undertaken with the objectives of recognizing the Antecedent Inferior Alveolar Nerve loop, evaluating the dimensions of the A.L. for pre-surgical planning, and comparing the age-related three-dimensional variations in A.L. of the Inferior Alveolar nerve.", "keywords": [ "Cone-beam computed tomography", "mandibular nerve", "mental foramen." ], "content": "Introduction\n\nThe mental neurovascular bundle crosses the inferior alveolar nerve’s anterior loop, located before the Mental Foramen (M.F) then turns around to exit the M. F. This anatomic variance must be taken into consideration when surgical operations on the anterior mandible are planned. Osteotomy, bone harvesting, and dental implant placement are all considered safe elective procedures.1\n\nThe human face has high innervations. The fifth cranial nerve, the Trigeminal nerve, regulates face sensibility. The trigeminal ganglion is located in the Middle Cranial Fossa of the Sella Turcica, and is the origin of the trigeminal nerve’s sensory branch. The Ophthalmic, Maxillary, and Mandibular branches, in turn, innervate the top, middle, and lower three-quarters of the head. The Mandibular nerveis the 3rd branch of the trigeminal nerve. Which enters the mandible via the mandibular foramen.1 The Inferior Alveolar Nerve is the nerve that passes Via use of the M.F. The inferior alveolar nerve’s terminal portion can rarely cross through the mental foramen’s lower border and continue as an intra-osseous anterior loop. Despite being a benign anatomical variation, it needs to be accurately identified in order to plan surgery that involves the insertion of implants as well as anterior mandibular operations such as genioplasty, mandibular premolar root extraction, chin bone harvesting, and surgical rehabilitation after mandibular injury.2\n\nThe standard of therapy for edentulous mandibles is to insert dental implants in the anterior mandible. There were anatomical hazards in the anterior mandible, despite the surgery being usually viewed as safe.3 With the advent of endosseus, replacing lost teeth with dental implants has become a very predictable medical option. The gold standard of treatment for edentulous mandibles was the installation of implants in the front mandible. Although the procedure is generally viewed as safe, there are some anatomical difficulties in using the front mandible.5 The most significant anatomical risk will be the potential for the IAN anterior loop, which emerges through the M.F.If the I.A.N. is not detected before surgery, there may be iatrogenic nerve harm. An injury to the A.L. would lead to damage to the mental nerves. As a result, the mental nerve can become anesthetized, experience paraesthesia, or encounter dysesthesia.6 The labiodental region and the gingiva up to the second premolar on the jaw are both detectable by the mental nerve. It could be tricky to carry out regular activities like eating, speaking, and even brushing your teeth if there is a changed sensation in this area.6\n\nC.B.C.T. should provide a number of advantages over multi-slice computed tomography, including the capacity to assess craniofacial traits in three dimensions (3D) without distortion or overlapping pictures.7 Preventing sensorineural injury in the interforaminal region requires accurate neurovascular bundle identification and protection.\n\nThe aim of this study is to assess and compare age-related three-dimensional variation in the anterior loop of the inferior alveolar nerve for pre-surgical planning.\n\n\n\n1. To identify the nerve’s anterior loop is the lower alveolar.\n\n2. To analyse the size of the A.L. of I.A.N. for pre-surgical planning.\n\n3. To compare the age-wise three-dimensional variations in A.L. of inferior alveolar.\n\n\nMethods\n\nThe study proposal was approved by “Institutional Ethics Committee” of DMIMSDU, Sawangi (meghe), Wardhawith Ref. No. DMIHER (DU)/IEC/2023/855 on 31/03/2023.\n\nPatients will be selected as per the inclusions and exclusion criteria. This study will be done using Cone Beam Computed Tomography. The study will be conducted at Sharad Pawar Dental College, Sawangi (Meghe), Wardha, in the Dept. of Oral Medicine and Radiology.\n\nPLANMECA pro-Max C.B.C.T. machine with Romexis viewer.\n\n\n\n1. Patients between the ages of 16 and 65 years.\n\n2. The CBCT scan must show teeth at the mandibular frontal region on both sides, up to 2-3 cm from the foramen mental.\n\n3. The mandibular canal and mental foramen siteswhich cannot be altered by pathological changes.\n\n4. There is no sign of any surgery or trauma that would have changed the mandibular canal’s or the mental foramen’s position.\n\n\n\n1. Systemic illness (DiabeticMallitus, Hypertentionetc) presence.\n\n2. Individuals receiving radiotherapy.\n\n3. Implants or mental artifacts in the area of the foramen.\n\n4. Abnormalities during teeth and bone development are present.\n\nPatients reporting to the Department of Oral Medicine and Radiology will be approached to take part. Those who are willing to participate in the study and give written informed consent will be included in the study (see Extended data for the sample consent form9).\n\nAfter satisfying the inclusion and exclusion criteria (via medical records), patients will be taken for C.B.C.T. imaging. The three-dimensional PLANMECA proMax C.B.C.T. equipment with an exposure period of 11.30 s, a tube voltage of 90 kV, a current in a tube of 5 mA, and a cylindrical F.O.V. measuring 17mm 13mm with a voxel size of 300 will be used to acquire the C.B.C.T. pictures. To standardize the image selection criterion, the F.O.V. will be the same throughout all scans. Each picture should be af 90μm, and a single 360-degree scan will be used. All the scans will be taken with the subject’s head in an upright position and the patient will be instructed to make molars to molars contact and to breathe from their nose.\n\nThe occlusal plane will be horizontally positioned to the scan plane. The cantered plane will be the midsagittal plane. The planes on the CBCT pictures’ three axes (X,Y,Z) will be analyzed in order. All of the images’ contrast and brightness will be kept constant for consistency during image analysis. All images will be evaluated under the best viewing conditions using picture viewing software PLANMECA pro-Max.Each volume’s axial slice will be rebuilt on the specified axial slice and parallel to the mandible’s lower edge. The most anterior section of the IAN and the MF most anterior segment will be marked & length will be measured using PLANMECA pro-Max software.\n\nCochian Formula for Sample Size:\n\nWhere;\n\nZαl2 is the level of significance at 5%, i.e. 95% .confidenece interval = 1.96\n\nP = prevalence of inferior alveolar nerve anterior loop = 22% = 0.22\n\nd = desired error of margin = 10% = 0.10\n\nStudy Reference:8\n\nThis study protocol will be published in a PubMed, Web of Science, and Scopus Indexed Journal.\n\nNot Yet Started.\n\n\nDiscussion\n\nThere are positions of the MC laterally in women, especially at the level of the premolars, that have been discovered to be significantly and clinically relevant correlated as observed on CBCT.\n\nIn research by Baratollah Shaban et al. 20172 71 patients’ CBCT scans were used. There are three main anatomical variations of the IAN anterior loop. Type I: The incisive branch thickness is comparable to that of the main branch, and the anatomy is Y-shaped.\n\nEdudara Helena Leandro do Nascimento et al. 20163 employed CBCT on 250 patients; it was possible to determine the frequency and severity of anterior loop in a Brazilian population. Gender, age, and mandibular side were taken into consideration while contrasting the anterior loop’s length.\n\nJuan Muileno Lorenzo et al. 20154 compared the MF and AMF visualization capabilities of CBCT and panoramic radiograph and used CBCT to examine the supplementary mental foramen’s existence and the anatomic characteristics of the MF. The sample for the study included CBCT and PAN tests were performed on 357 individuals. The average AMF area in the study was between 1.5 and 2 mm.\n\nFereidoumparnia et al. 20125 determined that it is risky to suggest any particular medial distance from the mental foramen after conducting a study on 96 patients using CBCT to evaluate the Anatomical landmarks in the mandibular: their appearance, visibility, placement, and course inter-foraminal region. The diameter of the canal and foramen should be evaluated separately for each person in order to choose the appropriate location.", "appendix": "Data availability\n\nNo data are associated with this article.\n\nZenodo: Assessment and comparison of age-related three-dimensional variation in the anterior loop of the inferior alveolar nerve for pre-surgical planning. https://zenodo.org/record/8272121. 9\n\nThis project contains the following extended data:\n\n- Consent form.docx\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgments\n\nI acknowledge the help from my university, college, professors, department, statistician, and resources, which provided the necessary data and their inputs for this given clinical study.\n\n\nReferences\n\nSiddiqui Z, Rai S, Ranjan V: Efficacy and Evaluation of Cone Beam Computed Tomography in Determining the Prevalence and Length of Anterior Loop of Inferior Alveolar Nerve in North Indian Population. J. Indian Acad. Oral Med. Radiol. 2018; 30: 32–37. Publisher Full Text\n\nShaban B, Khajavi A, Khaki N, et al.: Assessment of the anterior loop of the inferior alveolar nerve via cone-beam computed tomography. J. Korean Assoc. Oral Maxillofac. Surg. 2017; 43: 395–400. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\ndo Nascimento EHL , dos Anjos Pontual ML , dos AnjosPontual A , et al.: Assessment of the anterior loop of the mandibular canal: A study using cone-beam computed tomography. Imaging Sci. Dent. 2016; 46: 69–75. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMuinelo-Lorenzo J, Juan-Antonio S-Q, Alonso AF, et al.: Anatomical characteristics and visibility of mental foramen and accessory mental foramen: Panoramic radiography vs. cone beam CT. Med. Oral Patol. Oral Cir. Bucal. 2015 Nov; 20(6): e707–e714. PubMed Abstract \| Publisher Full Text\n\nParina F, Moslehifard E, Hafezeqoran A, et al.: Characteristics of anatomical landmarks in the mandibular interforaminal region: a cone-beam computed tomography study. Med. Oral Patol. Oral Cir. Bucal. 2012; 17: e420–e425. Publisher Full Text\n\nRenton.: Prevention of Iatrogenic Inferior Alveolar Nerve Injuries in Relation to Dental Procedures. J. Oral Maxillofac. Surg. 2010; 37: 350–363. Publisher Full Text\n\nSuomalainen A, Kiljunen T, Käser Y, et al.: Dosimetry and image quality of four dental cone beam computed tomography scanners compared with multislice computed tomography scanners. DentomaxillofacRadiol. 2009; 38: 367–378. PubMed Abstract \| Publisher Full Text\n\nVelasco-Torres M: Inferior alveolar nerve trajectory, mental foramen location, and incidence of mental nerve anterior loop. J. Med. Oral Patol. Oral Cir. Bucal. 2017 Sep 1; 22(5): e630–e635. Publisher Full Text\n\nWaghadkar A: Assessment and comparison of age-related three-dimensional variation in the anterior loop of the inferior alveolar nerve for pre-surgical planning.2023. Publisher Full Text" }	[ { "id": "241484", "date": "04 Mar 2024", "name": "Yasser Nabil El Hadidi", "expertise": [ "Reviewer Expertise oral and maxillofacial surgery" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nDear author The current manuscript presents an interesting study. Pros Detail of changes with age regarding position of anterior looping based on CBCT data. Cons No mention of previous similar studies.Introductions need more enriching by similar studies as ref [1]\nMinor comments The objective must be beneficial to dental surgeons and endodontists. For example, must carry a take a home message, recommendation or guidelines for implant placement in premolar area, apicectomy procedure in premolar area for example.\n\nMajor comment Please assess the data by two examiners and perform inter examiner reliability test to confirm the recorded data. Thanks for consideration!\n\nIs the rationale for, and objectives of, the study clearly described? Yes\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [ { "c_id": "12986", "date": "19 Dec 2024", "name": "Dr. Abhilasha Waghadkar", "role": "Author Response", "response": "Ok respected sir.." } ] } ]	1	https://f1000research.com/articles/12-1386
https://f1000research.com/articles/12-1385/v1	20 Oct 23	{ "type": "Case Report", "title": "Case Report: A rare case of Fanconi anaemia", "authors": [ "Varsha Gajbhiye", "Yeshwant Lamture", "Pankaj Gharde", "Akash Inamdar", "Juee Meghe", "Varsha Gajbhiye", "Pankaj Gharde", "Akash Inamdar", "Juee Meghe" ], "abstract": "Fanconi anaemia (FA) is autosomal recessive but can also be an autosomal dominant or X-linked recessive disease. In India, Fanconi anaemia is a very rarely seen disorder. Clinical findings in FA include pancytopenia, hyperpigmentation or hypopigmentation, skeletal anomalies, small stature or growth retardation, endocrine abnormalities, anal atresia, deafness, malignancy of head and neck, and it has a tendency to run in families. Diagnosis of FA can be made on clinical findings and laboratory examination. Recommended prenatal testing like chorionic villus sampling and amniocentesis. Androgen therapy and hematopoietic stem cell transplantation (HSCT) are treatment modalities recommended for FA. Follow-up of all diagnosed FA patients is essential throughout their lifetime. Having a healthcare team with diverse expertise is vital in preventing anemia and malignancy. A meticulous plan and unwavering support are also crucial. Outcomes of FA patients is desolate due to complications associated with it. Our aim of the study is to increase awareness among physicians about the presence of such rare diseases. Well-timed diagnosis with appropriate treatment is needed to prevent complication.", "keywords": [ "Genetic", "Malignancy", "Aplastic anemia", "Bone marrow depression", "Prenatal testing" ], "content": "Introduction\n\nSwiss paediatrician Guido Fanconi was the first to discover Fanconi anaemia (FA) in 1927. He noticed that three boys had similar birth defects and died due to a condition mimicking pernicious anaemia. Later on, in 1960, motivated by Fanconi’s work, it was found that the disorder was due to chromosomal abnormalities and could cause bone marrow failure and malignancy. With recent advances in genetic studies, it is now known that FA is caused by changes in 22 genes. It is autosomal recessive but can also be autosomal dominant or X-linked recessive. According to the research, FA protein serves as a regulator of inflammatory cytokine and cell cycle via the p53/p21 pathway, as well as oxidative phosphorylation. Witnessing technological advancements that allow us to understand complicated human genetics further is genuinely awe-inspiring. Interference in any of these functions leads to the depletion of hematopoietic stem cells (HSCs), which leads to bone marrow failure and skeletal anomalies.1\n\nAround the world, the incidence of Fanconi anemia is 1 in 160,000. It is observed that FA is presented among people of Ashkenazi Jewish descent, the Roma population of Spain, and Black South Africans.2 In India, Fanconi anemia has an incidence of 1 per 3,60,000 live births.3\n\nClinical finding in FA includes pancytopenia, hyperpigmentation or hypopigmentation, skeletal anomalies, small stature or growth retardation, endocrine abnormalities, anal atresia, deafness, malignancy of the head and neck, and anorectal and familial occurrence.4\n\nDiagnosis of FA can be made on clinical findings and laboratory examination. Recommended prenatal testing like chorionic villus sampling and amniocentesis is advised within ten and fifteen weeks of gestation. If genetic variants are known, then targeted variant analysis can be done, whereas if familial variants are not known, then chromosome breakage testing is recommended. Androgen therapy and hematopoietic stem cell transplantation (HSCT) are treatments recommended for FA. Treatment with gene therapy as part of a clinical trial in the field of medicine holds immense promise for patients with various genetic disorders including FA. The progress being made gives hope for the future.5\n\nFollow-up of all diagnosed FA patients is essential throughout their lifetime. This includes monitoring haemoglobin levels every 3–4 months. Treatment should be started when the haemoglobin level decreases below 8 g/dL. To maintain good health after diagnosis, it’s recommended to have a complete blood count (CBC) every six months and bone marrow studies annually, especially if the patient has a normal complete blood count (CBC) at the time of diagnosis. This approach will help identify any potential health issues and ensure that the patient receives the appropriate treatment if necessary. If a patient is diagnosed with cytopenia, it is recommended that they undergo CBC every three months and bone marrow studies every six months. Blood glucose levels should be monitored every two weeks as FA has changes in insulin secretion because of abnormalities in glucose metabolism. Magnetic resonance imaging (MRI) is studied to rule out malignancy every six months.\n\nIt’s crucial to have a meticulous treatment plan with supportive care from a multidisciplinary healthcare team. This may involve working with an ear nose throat (ENT) surgeon for audiological concerns, a dermatologist and endocrine specialist for hypogonadism and thyroid dysfunction, regular dental check-ups for oral health, and consultations with an ophthalmologist, dietician, and nutritionist. By taking a proactive approach and collaborating with your healthcare team, potential health complications can be identified and improve overall wellbeing.\n\nOur aim of the study is to increase awareness among physicians about the presence of such rare diseases. Well-time diagnosis will permit appropriate treatment to start before complications develop. Also, it will allow parental decision–making about prenatal diagnosis in their future pregnancy.\n\n\nCase report\n\nAn 8-year-old male child of Indo-Aryan ethnicity, came to the tertiary centre in the paediatric ward with complaints of an increase in the rate of respiration, pallor, fever, and cough for 10 days. He had a history of similar complaints on and off since birth. Antenatal or postnatal history was uneventful. Psycho-social history was not significant. There was no history of congenital anomalies in the family. Personal and developmental history was normal for the patient’s age. Anthropometry examination showed height for age and head circumference around the 50th percentile. On the physical examination, the patient has an absent right thumb and a rudimentary small left thumb (Figure 1).\n\nThe patient had pallor, no icterus, and no significant lymphadenopathy. The right kidney was not seen in ultrasonography. Pancytopenia with anisocytosis was seen in the peripheral examination. The normal range of haemoglobin in this age group is 11.9 to 15 g/dl. The patient’s haemoglobin was 6 gm/dl. The platelet count was 17,700/μL, which gradually improved on receiving packed red blood cell transfusion (PRBC) and platelet concentrates (PC) transfusions. The normal range of antinuclear antibodies (ANC) is 1:160 to 1:180. In the present patient, ANC was 1:20 which was on the lower side. Blood examination for vitamin B12 and folic acid values were normal. Bone marrow analysis showed signs of hypocellular bone marrow, with reductions in all cell lines (erythroid, myeloid, megakaryocytic) which is indicative of aplastic anemia. There was no sign of fibrosis, granuloma, or metastasis on the magnetic resonance imaging (MRI) or computed tomography (CT) scan. Echocardiography was normal. The thyroid level was within the normal limit. Liver function test and kidney function test were within the normal ranges. Normal blood sugar level at this age is 90 to 180 mg/dl; the patient’s blood sugar was 80mg/dl. Oxygen saturation was 88% at time of admission so oxygen administration was initiated. X-ray of the chest showed bilateral infiltrates suggestive of pneumonia, for which the patient was started on intravenous third generation antibiotics, ceftriaxone 500 mg given every 8 hours for 5 days, later patient was shifted on tablet cefixime 100 mg two times a day for next 5 days. X-Ray of the right arm showed that the radius bone was absent along with bowing of the ulna (Figure 2).\n\nBecause of the absence of a thumb and radius bone, absent right kidney on ultrasonography, and aplastic anaemia on bone marrow examination, we thought of going for a genetic test. A chromosomal breakage study for Fanconi anaemia was done, which showed metaphase karyotyping; GTG banding (G-bands by trypsin using Giemsa) showed karyotype-46XY, abnormal chromosomal breakage syndrome with significant structural aberrations in most of the metaphases, which confirmed the final diagnosis for FA.\n\nOther examinations correlated with FA were done, which included an ear examination that did not show any sign of deafness. Test for growth hormones, insulin, and lipid profile were in the normal range. The thyroid function test and glucose tolerance test were in the normal range. Referral to the orthopaedic department helped the patient obtain the functional and cosmetic outcomes needed for their thumb abnormalities.\n\nThe risk factor associated with FA was explained to parents, follow-up sessions were scheduled, and the patient was discharged after 15 days of admission. Prophylaxis iron therapy at a dose of 3mg/kg, 2.5 ml once a day through oral route was started and will be continued for three months.\n\nDuring the follow-up, we planned for appropriate follow-up care for the patient to ensure that they receive the necessary treatment and support. The patient was advised to test for haemoglobin and CBC to be done every three months and bone marrow studies every year. Blood glucose levels should be monitored every two weeks as FA has changes in insulin secretion because of abnormalities in glucose metabolism. Magnetic resonance imaging (MRI) will be done to rule out malignancy every six months. Every six months, the patient was advised to have an appointment for consultation with ENT surgeon, dermatologist, endocrinologist, ophthalmologist to rule out any malignancy.\n\nThe prognosis and outcome were good in the present study patient as he did not show any tumour or bone marrow failure on follow-up.\n\n\nDiscussion\n\nFA is a rare inherited disease associated with various abnormalities like absent limbs, absent kidneys, anaemia, urogenital anomalies, and bone marrow aplasia. It is caused by defects or mutations in genes. The protein that is synthesized by this gene helps to recognise and regenerate damaged DNA. In FA, because of defects in genes, the DNA regeneration process diminishes, and new stem cells are not produced, leading to aplastic anaemia.6\n\nFA is seen in the first decades of life. Males are more affected than females. Ghaida Bakoar Alahmadi et al. in their study, showed that the mean age of presentation of FA was about 7–8 years and it had a male predominance, which was similar to the present study.7\n\nThe benchmark for diagnosing FA is on chromosomal breakage study for metaphase karyotyping; GTG banding, which shows abnormal chromosomal breakage syndrome with significant structural aberrations in most of the metaphases. Bushra Anam Ali et al. studied Fanconi anaemia on chromosomal breakage and found abnormal chromosomal breakage syndrome with significant structural aberrations in most of the metaphases, which was seen in the present study.8\n\nParental consanguinity was common in a study by Heather A et al. that highlighted the importance of community and youth education and the importance of pre-marital counselling. This was opposite to the present case report as parenteral consanguinity was not seen, nor was pre-marital counselling done.9\n\nOf the many physical abnormalities, hearing loss is commonly observed with FA. In their research, M D Tischkowitz et al. showed that conductive deafness is common and may or may not be associated with external ear malformations. This was not seen in the present study.10\n\nCongenital anomalies in the renal system are associated with FA. Vijaya Sathyanarayana et al. showed that renal abnormalities are commonly seen in FA. In the present study, absence of a kidney was seen on the right side, which was similar to the study done by Vijaya Sathyanarayana et al.11\n\nShort stature, obesity, glucose intolerance, and hypothyroidism are some of the endocrine abnormalities associated with FA. In their research, Neelam Giri et al. showed that affected males with FA have a high incidence of endocrine abnormalities, which was not seen in the present study.12\n\nMalignancy is associated with FA. Philip S Rosenberg et al. showed a higher incidence of malignancy in FA, like leukaemia and solid tumours, which were not seen in the present case report.13\n\nA Butturini et al. presented data suggesting that FA subjects have an impairment of haematopoiesis that leads to hypoplastic anaemia and bone marrow failure during childhood; this was similar to the present study in which the child showed signs of bone marrow failure on admission.14\n\nThumb anomaly and the absence of a radius on radiological examination was seen in the present case, which is rare in FA. Based on the review conducted by B.P. Alter, it has been observed that paediatric patients who have congenital anomalies of the upper limb are often associated with genetic disorders of the bone marrow. These disorders can lead to bone marrow failure, such as FA and others similar to the present case.15\n\nIt is important to note that prenatal testing and monitoring for malignancy and bone marrow examination can increase the chances of survival for paediatric patients with congenital anomalies of the upper limb. However, it is also important to consider the limitations of the study, such as the potential for patients to become tired of visiting the hospital for follow-up appointments and a need for a larger sample size for effective follow-up. Proper management and care for these patients is crucial, including consideration of genetic disorders that may lead to bone marrow failure.\n\nAs the patient is a child his perspective is difficult to judge, but the parents were very anxious and nervous as long-term follow-up brings uncertainty. What will happen in the follow-up was the question commonly asked. Parents need to be counseled by educated and empathetic paediatricians who can provide comfort and support during what can be a very challenging time. While prenatal testing and monitoring, as well as bone marrow examinations, can increase the chances of survival, it’s important to be aware of potential limitations. Follow-up appointments are an absolute must, and a larger sample size is needed for effective follow-up. Additionally, genetic disorders that may lead to bone marrow failure should be considered in the overall care plan for these patients. This is not something that can be overlooked or ignored.\n\n\nConclusion\n\nFanconi anaemia is associated with aplastic anaemia, physical anomalies, bone marrow failure, and malignancy. Since this is rare, physicians should be aware of this condition so that other associated complications can be prevented and treated as well. Parents should be well-informed of the challenges that may arise when caring for a child with congenital anomalies of the upper limb. Correct management and care are extremely important, which is why parents should seek assistance from knowledgeable and sympathetic paediatricians who can provide much-needed support during this time. Regular follow-up appointments with a multidisciplinary team are essential, as is close surveillance and treatment. It’s important for parents who have had children diagnosed with FA to be cautious and plan ahead when considering subsequent pregnancies. Above all, it’s imperative to be mindful of potential limitations and genetic disorders that may have an impact on a child’s care plan.\n\n\nConsent\n\nWritten informed consent for publication of their clinical details and clinical images was obtained from the parent of the patient.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\n\nReferences\n\nVelleuer E: Guido Fanconi (1892-1979): a jack of all trades. Nat. Rev. Cancer. 2006; 6(11): 893–898. Publisher Full Text\n\nBhandari J, Thada PK, Puckett Y: Fanconi Anemia. [Updated 2022 Aug 10]. StatPearls. Treasure Island (FL): StatPearls Publishing; 2023 Jan. Reference Source\n\nSaheb DM: Fanconianaemia. Indian J. Dermatol. Venereol. Leprol. 2002; 68: 46–47. Reference Source\n\nFaivre L, Guardiola P, Lewis C, et al.: Association of complementation group and mutation type with clinical outcome in fanconi anemia. European Fanconi Anemia Research Group. Blood. 2000; 96: 4064–4070. PubMed Abstract\n\nMerrill A, Rosenblum-Vos L, Driscoll DA, et al.: Prenatal diagnosis of Fanconi anemia (Group C) subsequent to abnormal sonographic findings. Prenat. Diagn. 2005; 25(1): 20–22. Publisher Full Text\n\nSolanki A, Mohanty P, Shukla P, et al.: FANCA Gene Mutations with 8 Novel Molecular Changes in Indian Fanconi Anemia Patients. PLoS. 2016; 11(1): e0147016. Publisher Full Text\n\nAlahmadi GB, Alharbi I, Alsahafi IKH: Rare case of Fanconi anemia associated with polydactly. Clin. Case Rep. Rev. 2017; 3(9): 1–2. Publisher Full Text\n\nAli BA, Zulfiqar M, Mustafa A, et al.: Fanconianemia: Case report on rare aplastic anemia. Open J. Clin. Med. Case Rep. 2019; 5(7): 1541–1542. Reference Source\n\nZierhut HA, Tryon R, Sanborn EM: Genetic Counseling for Fanconi Anemia: Crosslinking Disciplines. J. Genet. Couns. 2014; 23(1): 910–921. PubMed Abstract \| Publisher Full Text\n\nTischkowitz MD, Hodgson SV: Fanconianaemia. J. Med. Genet. 2003 Jan; 40(1): 1–10. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSathyanarayana V, Lee B, Wright NB, et al.: Patterns and frequency of renal abnormalities in Fanconianaemia: implications for long-term management. Pediatr. Nephrol. 2018; 33(9): 1547–1551. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGiri N, Batista DL, Alter BP, et al.: Endocrine Abnormalities in Patients with Fanconi Anemia. J. Clin. Endocrinol. Metabol. 2007; 92(7): 2624–2631. PubMed Abstract \| Publisher Full Text\n\nRosenberg PS, Greene MH, Alter BP: Cancer incidence in persons with Fanconi anemia. Blood. 2003; 101(3): 822–826. Publisher Full Text\n\nButturini A, Gale RP, Verlander PC: Hematologic abnormalities in Fanconi anemia: an International Fanconi Anemia Registry study. Blood. 1994; 84(5): 1650–1655. PubMed Abstract \| Publisher Full Text\n\nAlter BP: Arm anomalies and bone marrow failure may go hand in hand. J. Hand Surg. Am. 1992; 17(3): 566–571. PubMed Abstract \| Publisher Full Text" }	[ { "id": "331122", "date": "21 Nov 2024", "name": "Diana Prepelita", "expertise": [ "Reviewer Expertise Medical Geneticist", "Genomic and Genetic Technologies", "Bioinformatics", "Multi-omics", "Hereditary Cancer Syndromes" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nOverall impression: good starting point, needs improvement\nCase background and history: As mentioned, Fanconi Anemia (FA) can have multiple modes of inheritance, including autosomal recessive, autosomal dominant and X-linked. Pedigree spanning at least 3 generations, noting any abnormalities in relatives is crucial for correct management of the case. For example, if gene involved is BRCA1, mother might exhibit breast cancer phenotype, and chance of having another child affected is 25% with each pregnancy. If gene involved is RAD51, most likely mutation is de novo, parents are not carriers and risk for another pregnancy is very low (~1%). Especially in the context where genetic testing was not performed, this could have great implication for case management.\n\nPhysical examination and diagnostic tests: Sometimes reference interval for lab tests is present, sometimes not. It is also a bit difficult to follow. Might be better to present results in a table with Parameter \| Value \| Reference value. White blood cells count, which is important to define pancytopenia, not present. Seems like severe bone marrow failure, no grading included. Also it seems like a long enumeration of tests, without clear logical flow of why the analyses were performed. Thyroid check-up is mentioned twice for example. Chromosomal Breakage analysis is a very important test that requires expertise and a good protocol. Chromosome breakage can be mimicked by other conditions, and specific DNA cross-linking agents such as mitomycin C (MMC), diepoxybutane (DEB), or cisplatinum must be used. There is also a specific scoring system and results should reflected by this, as normal cells can have breaks too. [1] I would add a picture of the karyogram and describe in more details the protocol and results.\n\nDifferential diagnosis should be discussed.\n\nTreatment and surveillance: Seems fine overall, I am not sure if iron prophylaxis should be advised, given the risk of iron overload from transfusions and Fe deficit not being the underlying cause.\n\nDiscussions & Importance of findings and relevance for future understanding of disease processes, diagnosis or treatment: Article should focus more on patient and less on literature. Try and highlight the case particularities. For example, patients usually present with short stature, which does not seem the case. Absent radii is quite rare, present in only 7% of cases. etc.\n\nGeneral organisation: Sometimes flow of ideas is not very clear, for example investigation is mixed with treatment. A bit of attention on scientific language ex \"thyroid levels\". Also \"He had a history of similar complaints on and off since birth. Antenatal or postnatal history was uneventful.\". Firstly, these joint propositions are a bit contradictory and secondly, more information about the pregnancy follow up can be mentioned. For example, if patient had trimestrial scans, which most likely would have shown radii aplasia and thumb anomaly.\n\nReferences: Not very clearly stated and linked with the text. Example first Introduction paragraph with one citation at the end: the historical paper on Guido Fanconi has nothing to do with describing FA pathway.\n\nConclusion: Case is interesting and the initiative to increase awareness is great starting point. However, article needs quite major reorganisation of ideas and more details in mentioned areas.\n\nIs the background of the case’s history and progression described in sufficient detail? Partly\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Partly\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Partly\n\nIs the case presented with sufficient detail to be useful for other practitioners? Partly", "responses": [] }, { "id": "345891", "date": "17 Dec 2024", "name": "Tarcília Aparecida Silva", "expertise": [ "Reviewer Expertise Oral Medicine" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nWhile the report is interesting, there are some problems in the detailing the case and in the literature cited. A recurrent problem regarding the rare diseases is the report of partial findings. A checklist of the characteristics present or absent in the present case according to the recent guidelines would improve the quality of study. I also recommended attention to the points specified below.\n1. Abstract: It is sometimes so vague. I suggest that authors provide a better description of case they are reporting with marked characteristics of FA found in the present case. 2. Introduction: “Diagnosis of FA can be made on clinical findings…” in this paragraph the authors cite a reference that does not really cover complete information. In addition, in the previous paragraphs maybe the authors could re-check the literature to provide appropriate references, e.g. in the paragraph, “Follow-up of all diagnosed FA patients….” No reference was given. 3. Case Report: “The prognosis and outcome were good in the present study patient as he did not show any tumour or bone marrow failure on follow-up.” Please provide details about the follow-up, e.g. for how long the authors observed the patient? It would be important to detail the planning for the long term follow up considering the patient needs. Others medical specialties as hematology and pediatrics would be included in the list of appointments. 4. It is important to inform that diagnosis was based in the criteria defined in the 2020 Fanconi Anemia Clinical Care Guidelines (https://www.fanconi.org/) 5. Please cite the DNA cross-linking agent(s) used for chromosome breakage test(s). 6. Any craniofacial and or dental abnormalities were seen? It would be important to included dentists in the list of appointments. The risk of oral cancer should be also reinforced. 7. Sorry if I miss but the history of parents consanguinity was investigated? Any brothers or sisters, and if yes they were investigated? 8. Criteria outlined in VACTERL-H and PHENOS should be mentioned and discussed in light of the present case. 9. Discussion: I recommend the re-organization of discussion considering the major clinical findings observed, e.g. skeletal; hematological, etc. 10. Some minor typos were observed, as lack of space between words. In addition, MS would benefit of a review of English grammar.\n\nIs the background of the case’s history and progression described in sufficient detail? Partly\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Partly\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Partly\n\nIs the case presented with sufficient detail to be useful for other practitioners? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1385
https://f1000research.com/articles/12-1384/v1	20 Oct 23	{ "type": "Research Article", "title": "Green synthesis of new and natural diester based on gallic acid and polyethylene glycol", "authors": [ "Hafida Zerigui", "Radia Labied", "Redouane Chebout", "Khaldoun Bachari", "Rachid Meghaber", "Fatima zohra Zeggai", "Hafida Zerigui", "Radia Labied", "Redouane Chebout", "Khaldoun Bachari", "Rachid Meghaber" ], "abstract": "Background Antioxidant polyphenols like gallic acid (GA) and its esters called “gallates”, which have health advantages for humans, have grown in significance in maintaining a healthy lifestyle and eating a significant amount of secondary plant phytochemicals. Here, for the first time, we suggest a green synthesis of a brand-new, all-natural diester based on gallic acid and polyethylene glycol.\n\nMethods This di-gallate is created in a single step without the use of a solvent (solid-solid reaction). This reaction has a potential yield of up to 90%. The bathochromic shift of the absorption bands from 277 nm to 295 nm in the UV-VIS spectra was caused by the addition of PEG to gallic acid. To confirm the structure of this di-gallate; Fourier-transform infrared (FTIR) spectroscopy, proton and carbon nuclear magnetic resonance (1H and 13C NMR), the thermal stability identified by thermogravimetric analysis (TGA), X-ray diffraction (XRD), and scanning electron microscopy (SEM) were all used to thoroughly analyze the manufactured product.\n\nResults and conclusions The acquired results, when compared to the literature spectrums, supported the establishment of the di-ester structure and created new opportunities for a large number of applications.", "keywords": [ "Gallicacid", "polyethylene glycol", "di-gallate", "biopolymer", "green synthesis", "solid-solid reaction", "esterification." ], "content": "Introduction\n\nThe introduction “Our food should be our medicine and our medicine should be our food”, Hippocrates' description, though thousands of years old, used to emphasize the importance of nutrition to prevent or cure disease. A sizable group of bioactive phytochemicals known as polyphenols are now receiving more and more attention from the scientific community as well as, most notably, from the general public due to their abundance in foods like fruits, vegetables, and beverages, the regular consumption of which is thought to be good for human health. In addition to their chemopreventive and anticancer potential, studies have demonstrated that polyphenols and their derivatives can reduce the risks of diabetic, cardiovascular, and neurological illnesses.1,2 Although more recently, the produce, cosmetic and pharmaceutical industries have been using \"polyphenols\" for their antioxidant properties. According to their chemical structure, polyphenols possess at least two benzene rings and at least one or more hydroxyl substituents characterizing the phenyl system,3 and they are classified into four subclasses, including flavonoids, phenolic acids, stilbenes, and lignans.4\n\nRegarding phenolic acids compounds, are non-flavonoid polyphenolic compounds characterized by a carboxyl group linked to phenyl ring,5 this group includes cinnamic acids (Caffeic, ferulic and p-coumaric acids) and benzoic acids (gallic, vanillic, and syringic acids). When it comes to gallic acid (GA, 3,4,5-tri-hydroxylsbenzoic acid), is the most common hydroxybenzoic acid, that originates from plants and can be produced by acid hydrolysis of hydrolyzable tannins6 or synthesized from shikimic acid,7 these compound present different pharmacological activities, mostly antioxidant activity. The GA alkyl esters (gallates) are an an vital elegance of herbal phenolic compounds that may be extracted from flora or synthesized through esterification of gallic acid with the corresponding alcohol in the presence of the catalyst.8,9 These alkyl gallates, specifically those with more than seven carbon atoms in the side-chain, have greater favorable and potent activities than gallic acid itself,10,11 that have precious biological effects, such as anti-microbial, anti-inflammatory,12 antitumor,13 antifungal,14 or prevention of gastrointestinal diseases, diabetes and even cardiovascular diseases.15 In many cases, these GA alkyl esters are useful as food additives16,17 or beauty additives.18\n\nDue to its high latent heat capacity,19 biocompatibility, solubility in aqueous solutions, and other promising biological properties, polyethylene glycol (PEG) is a synthetic polymer, a polyether made from ethylene glycol, considered as a biopolymer, is widely used in medical and pharmaceutical applications. The strong polarity of PEGs is a result of the terminal hydroxyl and ether groups.20,21 Because the low molecular weight PEGs have more hydroxyl groups than their structure would suggest, they are less soluble in water and other solvents as their molecular weight rises. PEG is easily chemically altered and can connect to other molecules and surfaces. PEG modifies the solubility and enlarges the connected molecules' size when it is attached to other molecules.22 Polyethylene glycol esters are also used in the medical, cosmetic and food industries. It is made from polyethylene glycol and the corresponding acid.\n\nHere, we present GA/PEG composites which combine the activities of gallic acid and polyethylene glycol « di-gallate of polyethylene glycol », through a rather simple and green esterification process and with progressed properties. The method is based on using only reagents, i.e. solvent-free reaction in a simple Erlenmeyer flasks (“solid-solid reaction”). This method increases the yield of the reaction compared to the standard reaction. The structure of the synthesized di-gallate was confirmed by different analyses: FTIR, 1H NMR, 13C NMR, UV-Vis, TGA and XRD. We noted the absence of the acid function and the presence of the ester function confirming that the two reagents; gallic acid and polyethylene glycol are associated. The goal of such research work is to develop an efficient and rapid method of the synthesis of GA/PEG composites to be able to acquire di-gallate composites and their changeable properties.\n\n\nMethods\n\nFourier Transformed Infrared (FTIR) Spectroscopy; The FTIR spectra were checked in utilizing a Brucker Tensor-27. The scan was done between 4000 and 400 cm-1. All spectra were baseline-corrected with Opus software.\n\nX-ray Diffraction (XRD); The Cristallinity of the products were determined by Brucker D8 Advance diffractometer DAVINCI model, operating in Bragg–Brentano geometry, with Cu Kα radiation (λ = 1.5418 Å).\n\nNuclear Magnetic Resonance (NMR); Proton and carbon nuclear magnetic resonance (1H NMR) and (13C NMR) spectra were recorded on Bruker 300 MHZ NMR spectrometer using deuterated dimethyl sulfoxide (DMSO-d6) as solvent.\n\nUltraviolet-Visible (UV–Vis); UV-Vis absorption spectra were recorded using Evolution 60S spectrophotometer (Thermo Fischer Scientific). The spectra were recorded in Dimethylsulfoxide (DMSO) solvent.\n\nScanning Electron Microscopy (SEM); The analysis of nanocomposites powder image was carried out Quanta FEG 250 instrument (FEI, Hillsboro).\n\nThermogravimetric analysis (TGA); it was carried out on a Setsys Evolution analyzer (ATG Setaram, Caluire, France) equipped with an aluminium cell, using aluminium pans to encapsulate the samples. Typically, samples were heated at a constant rate of 10 °C/min from room temperature up to 550 °C, under a helium flow of 50 mL/min. The thermal decomposition temperature was taken at the onset of significant (≥5%) weight loss from the heated sample.\n\nThe gallic acid was purchased from BioChem, Polyethylene glycol glycol 2000 (PEG, M = 2000g/mole) was obtained from Fluka, sulfuric acid was purchased from Emsure and dichloromethane was obtained from Sigma Aldrich.\n\nSynthesis of of polyethylene glycol diester\n\nIn this work, we performed a green esterification synthesis using the solid-solid reaction. PEG-digallates was prepared from gallic acid (0.02mol) and polyethylene glycol (PEG) (0.01mol) at 110°C using sulfuric acid (0.5ml) as a catalyst for 12 hours. Thereafter, 50mL of dichloromethane was added and filtred to remove non-reagent amounts of gallic acid and polyethylene glycol. The synthetic scheme is shown in Figure 1.\n\n\nResults and discussion\n\nThe solid-solid reaction can increase the yield of the esterification reaction until 20% compared with the standard esterification reactions. The final product was analyzed by IR-FT, 1H NMR and 13C NMR spectroscopy and compared with literature spectra.\n\nFTIR analysis was used to identify changes or structure inter¬actions caused by the addition of PEG to Gallic acid. The FT-IR spectra of gallic acid, PEG and PEG-digallate complex treated using Origin Pro (v.2018), are given in Figure 2. As proven in PEG-digallate, the peaks around 3000-3500 cm-1 represented the stretching and bending vibration of -OH, indicating the attendance of hydroxyl groups. The high absorption peaks at 2876, 1707 and 1279 cm-1, were assigned to methylene (=CH2), carboxyl (–COOH), and phenol (–OH); groups in pure PEG, respectively; at the same time as the peaks at 1609 and 1104 cm-1 were attributed to aromatic ester and ether groups.23 It is worth noting that, the FT-IR spectra of the obtained complex, pure PEG and GA are relatively similar with several other peaks observed around 1700–1000 cm-1 which are attributed to C–O bond stretching vibration and O–H bond bending vibration into gallic acid. In contrast, changes in the stretching frequency of the current groups indicated the interaction of PEG with the gallic acid functional group.\n\nNuclear magnetic resonance (NMR) spectroscopy was used to determine the molecular structure and chemical composition of the complex PEG-digallate synthesized.\n\nThe 1H-NMR spectra of PEG-digallate and gallic acid have been acquired in DMSO-d6 at a 5 mM concentration. As proven in Figures 3 & 4, the chemical shift of every proton for gallic acid and the obtained product were studied in DMSO-d6 earlier than and after reaction with PEG. The signals at 6.95, 8.13, and 8.31 ppm correspond to the ortho protons of the benzene ring, the para and meta hydroxyl (O-H) protons, respectively. Notably, the disappearance of the acidic group (COOH) proton signal and the presence of the polyethylene glycol proton CH2-CH2 are indicated at 3.51 ppm. Additional signals appear at 3.4 and 3.6 ppm, corresponding to protons at the ends of the polymer chains (protons c and b; Figure 3). Comparing the 1H-NMR spectra of gallic acid and PEG-digallate, we note that at 12.26 ppm there is no signal corresponding to the -COOH proton of the acid functional group, but there is a signal corresponding to the -OH proton of the hydroxyl functional group at 8.13 and 8.31 ppm, confirming that gallic acid has reacted with the acid function and that the resinous hydroxyl function is free.\n\nFurthermore, comparing the 1H NMR spectra of PEG-digallate and pure polyethylene glycol (From literature24), we take a look at the same strong signal at 3.51 ppm, corresponding to the CH2-CH2 of polyethylene glycol.\n\nThe CH2-CH2 protons at the ends of the polymer chains maintain the same signal at 3.4 and 3.6 ppm.\n\nBased at the evaluation of 13C-NMR spectra of the obtained complex PEG-digallate and gallic acid (Figure 5 and 6), which revealed the presence of a carbon signal (O=C-O) at 167.94 ppm corresponds to the ester function. Another peak at 138.4 ppm corresponds to the C-OH carbon in the para position, and a peak at 145.83 ppm corresponds to the two C-OH carbons in the meta position. The signal at 120.87 ppm corresponds to the -C- carbon in the para position, and the two carbons in the ortho position occur at 109.14 ppm on the benzene ring. The strong signal at 70.24 corresponds to the CH2-CH2 carbons of polyethylene glycol. The signals at 72.78 ppm and 60.68 ppm correspond to the C-O ether and C-O ester carbons at the end of the chain, respectively.\n\nComparing these results with the literature demonstrates the presence of signals for all carbons corresponding to gallic acid.25 The results in Figure 6 are compared with the 13C-NMR spectrum of polyethylene glycol esters, showing the same strong signal at 70.24 ppm, corresponding to the CH2-CH2 carbons of polyether, and the shift peak of C-O-ester carbons appearing at 60, 68 ppm.26\n\nX-ray diffraction (XRD) analysis was performed to examine the physical state of the obtained PEG-digallate complex. Figure 7 displays the results of the XRD examinations of gallic acid, PEG, and the composite PEG-digallates. Gallic acid was discovered to have numerous strong diffraction peaks of crystallinity between a diffraction angle of 2 θ = 7-50° in the X-ray diffractogram, indicating that it is exists as a crystalline substance.27,28 Pure PEG displayed two distinct high intensity peaks at approximately 2 θ = 19.20° and 23.20° with a few minor peaks at: 2 θ =26.35, 36.18, 39.84 and 45.29°, respectively. Complex development, where the typical PEG peaks of the composite PEG-DG are present with slightly decreased strength and the elimination of huge diffraction peaks, where it is no longer feasible to detect the characteristic peaks of the composite, both contributed to a partial amorphization. The findings showed that gallic acid is no longer a crystalline substance and that the PEG complex it was successfully formed into still exists in a semi-amorphous condition.\n\nFigure 8 displays SEM images of the complex PEG-Digallate, pure PEG, and gallic acid; they were taken with zooms up to 500, 50, and 10 μm, respectively. Smaller, regular-shaped crystals with a surface that appeared smooth define the pure gallic acid, which also has smaller, more crystalline forms (Figure 8, A). In contrast to the PEG-digallate composite, pure PEG has an extremely smooth and flat surface and very regular compact structure (Figure 8, B). Scanning electron micrographs reveal that the surface morphology of the PEG-digallate (Figure 8, C) composite was amorphous; this may have contributed to the absence of crystal structures (confirming the X-ray diffraction results) and may be related to the uniform dispersion of GA in the PEG matrix.\n\nThe UV-Vis absorption spectra of PEG, GA and PEG-digallate dissolved in DMSO were recorded at room temperature, as shown in Figures 9 & 10.\n\nThe absorption spectral changes of PEG-digallate in the presence and absence of PEG are shown in Figure 9. From the obtained spectrum, the absorbance maximum of gallic acid was discovered at 277 nm, which was related to the absorption of aromatic amino acids. The peaks of PEG-digallate increased with the addition of GA, and a red shift at 295 nm was noted. The outcomes suggested that polyethylene glycol and gallic acid had formed a novel combination. A transparent compound in a spectral domain, when taken in an isolated state, can become absorbent if it is put in the presence of a species with which it interacts by a mechanism of the donor-acceptor (D-A) type. According to the UV-Vis absorption spectra of PEG in DMSO, there was no discernible absorption between 200 and 800 nm, demonstrating that pure polyethylene glycol has a relatively low total absorption in the UV region (Figure 10). Important for electrochromic applications is this observation.\n\nThermal gravimetric analysis (TGA) was used to investigate the thermal degradation of PEG-digallate. The TG and DTG curves of PEG, GA, and PEG-digallate were shown in Figure 11 from room temperature to 600 C in a N2 environment at a heating rate of 10°C/min.\n\nThe Figure 11 C confirms that the surface medicine was successful. PEG-digallate TG and DTG curves (Figure 11 C) differ from those of PEG and GA (Figure 11 A & B). The PEG TG and DTG curves show that PEG has a single breakdown phase that begins at 350 °C and ends about 405 °C. Thermal breakdown of PEG is anticipated to occur at both the backbone chains -C-O- and -C-C- bonds. Figure 11 (A) illustrates that in GA, which has three decomposition phases, the compound is stable up to 87°C when the first mass loss (8.27%) occurs up to 105°C due to hydration water. Gallic acid is stable beyond this temperature until 200°C, with no mass loss and no endo or exothermal processes. The second phase (36.38%) occurs between 220-264°C, and the DTA curve shows an endothermic peak (250°C). The oxidation of organic matter immediately causes the third mass loss (18.22%) and two related exothermic peaks (at 400 and 428°C, respectively). The final residue of gallic acid breakdown was 1.5% of the total original mass (carbon residue). The TG and DTG curves of PEG-digallate in Figure 11C show that the majority of weight loss occurs below 100 °C, which can be attributed to trapped water. Thermal deterioration of composites, on the other hand, occurs primarily around 210 °C and 420 °C, which can be attributed to the GA layer stably coating on PEG. By counting the weight loss from 200 °C to 700 °C, the TG curve reveals that the amount of GA coated on PEG is around 3 to 4 wt.%. Because to the reduction in surface area for alterations, the aggregated PEG-digallate explains the limited amount of coated GA.\n\n\nConclusions\n\nThe goal of using a non-solvent and environmentally friendly approach to prepare the specified composite PEG-digallate was discovered to have been achieved. This solid-solid reaction is a synthetic method that maximizes the yield of the reaction (90%) and minimizes waste of chemical reagents. A successful production of PEG-digallate was confirmed by the 1H and 13C NMR analyses using DMSO-d6 as the solvent by the development of new peaks brought on by the interaction of the PEG matrix and gallic acid. The presence of the C-O ether and C-O ester carbons at the end of the chain and hydrogen bonding between the -OH and -COOH groups both appear to play a role in these interactions. Gallic acid absorbs more when PEG is added, with the highest absorption occurring at 295 nm, confirming the UV-Vis spectra of PEG-digallate. We also observed the structure and compact shape of the produced complex, which can support the amorphous composite, by XRD patterns and SEM images. Thermal analysis (TG-DTA) has been shown to be effective approaches for assessing the thermal behaviour of gallic acid. TGA investigated the temperature and enthalpy of melting and solidifying processes to demonstrate the chemical and structural stability of PEG-digllate, which could be maintained even after 200 heat cycles. As a result, this surface modification process paves the way for new applications requiring various types of PEG matrix with gallic acid.", "appendix": "Data availability\n\nFigshare: Data di-ester, https://doi.org/10.6084/m9.figshare.23816559.v1. 29\n\nFigshare: Figures, https://doi.org/10.6084/m9.figshare.23816619.v1. 30\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nThe Directorate General of Scientific Research and Technological Development, and the Scientific and Technical Research Center of Physico-Chemical Analyses (CRAPC) provided funding for this work.\n\n\nReferences\n\nDurazzo A, Lucarini M, Souto EB, et al.: Polyphenols: A concise overview on the chemistry, occurrence, and human health. Phytother. Res. 2019; 33(9): 2221–2243. PubMed Abstract \| Publisher Full Text\n\nNajjar RS, Turner CG, Wong BJ, et al.: Berry-derived polyphenols in cardiovascular pathologies: mechanisms of disease and the role of diet and sex. Nutrients. 2021; 13(2): 387. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHan X, Shen T, Lou H: Dietary Polyphenols and Their Biological Significance. Int. J. Mol. Sci. 2007; 8: 950–988. Publisher Full Text\n\nLafay S, Gil-Izquierdo A: Bioavailability of phenolic acids. Phytochem. Rev. 2008; 7: 301–311. Publisher Full Text\n\nNeveu V, Perez-Jiménez J, Vos F, et al.: Phenol-Explorer: an online comprehensive database on polyphenol contents in foods. Database. 2010; 2010. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGrundhöfer P, Niemetz R, Schilling G, et al.: Biosynthesis and subcellular distribution of hydrolyzable tannins. Phytochemistry. 2001; 57(6): 915–927. PubMed Abstract \| Publisher Full Text\n\nOssipov V, Slminen J-P, Ossipova S, et al.: Gallic acid and hydrolysable tannins are formed in birch leaves from an intermediate compound of the shikimate pathway. Biochem. Syst. Ecol. 2003; 31: 3–16. Publisher Full Text\n\nWeetal HH: Enzymatic gallic acid esterification. Biotechnol. Bioeng. 1985; 27(2): 124–127. PubMed Abstract \| Publisher Full Text\n\nSas B, Coppens B, Hemel JV: Method of synthesizing alkyl gallates. PCT Patent WO2001030299A2. 2001; 3.\n\nSilva IC, Regasini LO, Petrônio MS, et al.: Antibacterial activity of alkyl gallates against Xanthomonas citri subsp. citri. J. Bacteriol. 2013; 195(1): 85–94. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKubo I, Fujita KI, Nihei KI: Anti-Salmonella activity of alkyl gallates. J. Agric. Food Chem. 2002; 50(23): 6692–6696. Publisher Full Text\n\nHuang L, Yang SP: U.S. Patent Application No. 11/091,205.2006.\n\nLocatelli C, Rosso R, Santos-Silva MC, et al.: Ester derivatives of gallic acid with potential toxicity toward L1210 leukemia cells. Bioorg. Med. Chem. 2008; 16(7): 3791–3799. PubMed Abstract \| Publisher Full Text\n\nLeal PC, Mascarello A, Derita M, et al.: Relation between lipophilicity of alkyl gallates and antifungal activity against yeasts and filamentous fungi. Bioorg. Med. Chem. Lett. 2009; 19(6): 1793–1796. PubMed Abstract \| Publisher Full Text\n\nJang A, Srinivasan P, Lee NY, et al.: Comparison of hypolipidemic activity of synthetic gallic acid–linoleic acid ester with mixture of gallic acid and linoleic acid, gallic acid, and linoleic acid on high-fat diet induced obesity in C57BL/6 Cr Slc mice. Chem. Biol. Interact. 2008; 174(2): 109–117. PubMed Abstract \| Publisher Full Text\n\nFernandez-Lorente G, Bolivar JM, Rocha-Martin J, et al.: Synthesis of propyl gallate by transesterification of tannic acid in aqueous media catalysed by immobilised derivatives of tannase from Lactobacillus plantarum. Food Chem. 2011; 128(1): 214–217. PubMed Abstract \| Publisher Full Text\n\nWeetall HH: Enzymatic synthesis of gallic acid esters. Appl. Biochem. Biotechnol. 1985; 11: 25–28. Publisher Full Text\n\nZhang S, Akoh CC: Solvent-Free Enzymatic Synthesis of 1-o-Galloylglycerol Optimized by the Taguchi Method. J. Am. Oil Chem. Soc. 2019; 96(8): 877–889. Publisher Full Text\n\nKozma GT, Shimizu T, Ishida T, et al.: Anti-PEG antibodies: Properties, formation, testing and role in adverse immune reactions to PEGylated nano-biopharmaceuticals. Adv. Drug Deliv. Rev. 2020; 154-155: 163–175. Publisher Full Text\n\nGullapalli RP, Mazzitelli CL: Polyethylene Glycols in Oral and Parenteral Formulations—A Critical Review. International journal of phharmaceutic. 2015; 496. 9143164935.\n\nD’souza AA, Shegokar R: Polyethylene glycol (PEG): A versatile polymer for pharmaceutical applications. Expert Opin. Drug Deliv. 2016; 13: 1257–1275. Publisher Full Text\n\nPham Le Khanh H, Nemes D, Rusznyák Á, et al.: Comparative Investigation of Cellular Effects of Polyethylene Glycol (PEG) Derivatives. Polymers. 2022; 14(2): 279. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYang H, Feng L, Wang C, et al.: Confinement effect of SiO2 framework on phase change of PEG in shape-stabilized PEG/SiO2 composites. Eur. Polym. J. 2012; 48: 803–810. Publisher Full Text\n\nSon KH, Lee JW: Synthesis and characterization of poly (ethylene glycol) based thermo-responsive hydrogels for cell sheet engineering. Materials. 2016; 9(10): 854. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHirun N, Bao H, Li L, et al.: Micro-DSC, rheological and NMR investigations of the gelation of gallic acid and xyloglucan. Soft Matter. 2012; 8(27): 7258–7268. Publisher Full Text\n\nZarei M, El Fray M: Synthesis of hydrophilic poly (butylene succinate-butylene dilinoleate)(PBS-DLS) copolymers containing poly (ethylene glycol)(PEG) of variable molecular weights. Polymers. 2021; 13(18): 3177. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDorniani D, Kura AU, Hussein-Al-Ali SH, et al.: In vitro sustained release study of gallic acid coated with magnetite-PEG and magnetite-PVA for drug delivery system. Sci. World J. 2014; 1–11.\n\nTang B, Yuan H, Cheng L, et al.: Effects of gallic acid on the morphology and growth of hydroxyapatite crystals. Arch. Oral Biol. 2015; 60(1): 167–173. PubMed Abstract \| Publisher Full Text\n\nZeggai FZ: Data di-ester. Dataset. figshare. 2023. Publisher Full Text\n\nZeggai FZ: Figures. figshare. Figure. 2023. Publisher Full Text" }	[ { "id": "261846", "date": "30 Apr 2024", "name": "Nur Izyan binti Wan Azlee", "expertise": [ "Reviewer Expertise Enzyme technology", "bioprocessing", "bioconversion", "biomass degradation" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nGeneral comments:\nThe manuscript contains several grammatical and technical errors.\n\nThe highlighted sentences require rephrasing/corrections/changes/clarifications.\n\nThe discussion part seems to be lacking in contrast with the abundance of results obtained.\n\nSpecific comments:\nWhat is the concentration of sulfuric acid catalyst used for the synthesis of polyethylene glycol diester in the study? The mentioned volume of 0.5ml is for what total volume? Needs further clarification.\n\nSuggest mentioning the ratio of GA/PEG synthesized in the study.\n\nSuggest improving the resolutions for Figure 1, Figure 4, Figure 5.\n\nOrganize all figures to be after the mentioned text.\n\nChange the sub-titles in the Results and Discussion to be in terms of the result from the analyses and not the name of the analysis itself. Example: ‘Scanning electron microscopy’ to be changed to ‘Electron micrographs of the samples’.\n\nFigure 8: Insert the magnifications for each image.\n\nAt the conclusion section, the authors mentioned ‘TGA investigated the temperature and enthalpy of melting and solidifying processes to demonstrate the chemical and structural stability of PEG-digllate, which could be maintained even after 200 heat cycles.’ Is there any evidence for the claimed 200 heat cycles done?\n\nIt is suggested for the authors to strengthen the discussion section by comparing the data analysis obtained from the study with previous studies reported earlier.\n\nSpecify the methodology further.\n\nPlease also see the attached file linked here containing additional review comments.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "11613", "date": "30 May 2024", "name": "Fatima zohra Zeggai", "role": "Author Response", "response": "Thank you, We will do our best to correct the manuscript Kind regards" } ] }, { "id": "243716", "date": "12 Sep 2024", "name": "Azhagu Madhavan Sivalingam", "expertise": [ "Reviewer Expertise Plant based nanotechnology", "Pharmacology" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\n1. The abstract does not sufficiently explain the rationale behind in this study 2. The Introduction contains several disjointed statements that make it difficult to follow the main purpose of the study, and clarity of these sentences. 3. The methodology mentions the use of analytical equipment and providing specific models or manufacturing and specificity makes. 4. The whole phytochemical compound benefits or evaluates the significance of the findings. 5. The conclusion does not elaborate on the potential mechanism.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? No\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "261845", "date": "17 Sep 2024", "name": "Khadidja Taleb", "expertise": [ "Reviewer Expertise The whole article have been read", "checked and experted" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nTitle: Green synthesis of new and natural diester based on gallic acid and polyethylene glycol The paper presents the synthesis of a diester based on gallic acid and polyethylene glycol using green method without using solvent (solid-solid reaction).The obtained structure have been characterized using IR spectroscopy, Nuclear Magnetic Resonance (1H and 13C NMR), Thermogravimetric analysis (TGA), X-ray diffraction (XRD) and scanning electron microscopy (SEM). This paper is well structured and the results are interesting and promising for the development of new pathway for di-gallate of polyethylene glycol synthesis. However, Major revision would be required. In order to improve the document, the authors are invited to correct some errors in English; some errors of form, which are mentioned below then answer the questions.\n\nIn the abstract (Methods): To confirm the structure of this di-gallate; Fourier-transform infrared (FTIR) spectroscopy, proton and carbon nuclear magnetic resonance (1H and 13C NMR) (something is missing in this sentence for example “ have been used” ) Introduction (para 2) : The GA alkyl esters (gallates) are an an vital elegance of herbal phenolic Introduction (para 3): The method is based on using only reagents ( on the use) Methods (Apparatus): The analysis of nanocomposites powder image was carried out Quanta FEG 250 instrument (FEI, Hillsboro) (carried out on) Materials : Synthesis of of polyethylene glycol diester Results and discussion (NMR analysis line 5): earlier than and after. Please use Before instead of earlier Results and discussion (NMR analysis): “we take a look at the same …” in the scientific writing, it is better to avoid first person pronoun, i.e. we etc. Results and discussion (NMR analysis): Based at the evaluation of 13C-NMR. Replace at by on Figure 4 needs to be replaced by a high resolution image. Peak integration should be displayed in all NMR spectres. Figure 5 needs to be replaced by a high resolution image I recommend moving Figure 10 to the supporting information (supplementary information). Figure 11. Thermogravimetric analysis TGA of (A) gallic acid, (B) PEG and (C) PEG-digallate (please check this figure and replace it by the adequate figure). Thermogravimetric curves are not presented in this manuscript. In Thermal analysis (TGA) ( line 10 ) and in conclusion : authors used the abbreviation DTA whereas there is no DTA curves\n\nQuestions\nIn the background: authors have mentioned “Here, for the first time, we suggest a green synthesis of a brand-new, all-natural diester based on gallic acid and polyethylene glycol “in my opinion the diester presented here is not all- natural because it is synthetized by a synthetic polymer PEG and Gallic acid (natural compound) , furthermore the synthetic method des not respect the green chemistry rules (high temperature,toxic concentrated acid) I recommend to change this statement. Introduction (para 3): “This method increases the yield of the reaction compared to the standard reaction”. Could you provide one or two references? In the last paragraph of the introduction “Here, we present GA/PEG composites” this macromolecule is not a composite “see the definition of composite”.\n\nPlease correct this in the whole paper.\n\nIntroduction : para 3 : “ Here, we present GA/PEG composites which combine the activities of gallic acid and polyethylene glycol « di-gallate of polyethylene glycol »” what authors mean by activities ? Do you mean synergetic effect, if it is the case please correct the sentence? Could the authors justify the choice of reaction temperature?\n\nNo need to include NMR spectra of gallic acid because it is available in many works ( example: DOI:10.1371/journal.pone.0140242 Figure 11. Thermogravimetric analysis TGA of (A) gallic acid, (B) PEG and (C) PEG-digallate ( please check this figure and replace it by the adequate figure). There is no Thermogravimetric analysis TGA in this manuscript. In Thermal analysis (TGA) (line 10): authors used the abbreviation DTA; meanwhile there is no DTA curves. English of the paper should be corrected professionally. Please provide more details about potential applications of this macromolecule.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1384
https://f1000research.com/articles/12-1380/v1	20 Oct 23	{ "type": "Research Article", "title": "Self emulsifying drug delivery system: norfloxacin model drug", "authors": [ "Noor Abdulkareem", "Mohanad Naji Sahib", "Mohanad Naji Sahib" ], "abstract": "Background Norfloxacin is a synthetic antibacterial fluoroquinolone that is poorly water soluble and active against a wide spectrum of gram-positive and gram-negative aerobic bacteria. The goal of this work was to create a self-nanoemulsion (SNE) as a colloidal dispersed drug delivery system for increasing norfloxacin dissolving rate, while also assuming intestinal lymphatic transport participation for such a nano-size system that would increase norfloxacin oral bioavailability.\n\nMethods Depending on the solubility of norfloxacin in various components, several formulations of liquid SNEDDS of the antibiotic were created using oleic acid (oil), tween20 (surfactant), and PEG200 (co-surfactant). To assess the presence of a nanoemulsion region, phase diagram was generated at various oil: surfactant: co-surfactant ratios.\n\nResults The in-vitro dissolution profile suggested an optimum norfloxacin SEDD formulation (F1) comprised of oleic acid (10%), tween 20 (45%), and PEG200 (45%), as opposed to commercially available traditional tablets. The F1 formula exhibited thermodynamic stability with a zeta potential of +60.78mV and a droplet size of 57.4nm. The F1 formula revealed an improved dissolving profile as compared to the commercial Tablet of norfloxacin.\n\nConclusion Based on the findings, this study indicated that preparing norfloxacin as a liquid self-nanoemulsifying system improved the solubility and drug release of norfloxacin counteract the inconsistent bioavailability of the commercial tablet.", "keywords": [ "Norfloxacin", "nanoemulsion", "self nano-emulsifying drug delivery system (SNEDDS)" ], "content": "Introduction\n\nSelf-emulsifying drug delivery systems (SEDDS) are isotropic homogeneous pre-concentrate mixes of oil, surfactant, co-surfactant, and drug that, when gently agitated in an aqueous media, rapidly produce fine oil-in-water (o/w) emulsions.1 SEDDS improve the oral bioavailability of the medicine by enhancing its solubility and keeping it spread in small globules of oil during its passage through the gastrointestinal system.2 Also, bioavailability will be improved as a result of avoiding the first pass effect that when bile acids are present, lingual and pancreatic lipases activate on the oily phase of the SEDDS, resulting in the creation of emulsified mono-glycerides, diglycerides, and fatty acids and the generation of intestinal mixed micelles. As these mixed micelles travel through the enterocytes, they produce chylomicrons, they can route the medication into lymphatic pathways instead of blood vessels.3 Because the diameter of the emulsion droplets generated ranges from nanometers to several microns, self-emulsify drug delivery systems are classified as self-nano-emulsifying (SNEDD) and self-micro-emulsifying (SMEDD).4\n\nNorfloxacin (NOR) is a second generation fluoroquinolone. This broad-spectrum antibacterial drug is widely used to treat a variety of infection disorders involving the gastrointestinal or genitourinary systems.5,6\n\nNorfloxacin’s chemical name is 1-ethyl-6-fluoro-4-oxo-7-piperazin-1-yl-1H-quinoline-3-carboxylic acid. It has a very low solubility in water (0.28 mg/mL), and its color is a very light yellow. NOR is very basic in nature (pKa 6.39) and has an empirical log P value of 1.47. Additionally, the clinical effectiveness of this BCS class IV medication is affected by its poor bioavailability (30%), protein binding value (15%), and about 30% of drug excreted unaltered.7 There are a few attempts to improve the pharmaceutical properties of NOR such as dispersible tablets, extended-release matrix tablets, gastro-retentive floating tablets, cyclodextrin-containing suspension, solid lipid nanoparticles, suspensions, gastro-retentive floating microspheres, and sustained release alginate micro-beads.\n\nThe goal of this project is to develop norfloxacin as an oral self-emulsifying liquid formulation that will improve solubility and thus bioavailability.\n\n\nMethods\n\nNorfloxacin, coconut oil, mineral oil, olive oil, corn oil, spearmint oil and castor oil from Hangzhou Hyper chemicals (China). Oleic acid, clove oil and methanol were purchased from G.C.C., (UK). PEG 400, propylene glycol were obtained from CDH (India). PEG200, tween (20 and 80), span (20 and 80) were obtained from Thomas baker (India).\n\nThe saturated solubility of norfloxacin in various oils, surfactants, and co-surfactants was determined. The oils that were used were (coconut oil, mineral oil, olive oil, oleic acid, corn oil, spearmint oil, clove oil, castor oil). Surfactants and co-surfactants (tween 20, 80, span 20, 80) (PEG200, 400, PG). In order to assess the solubility, an excess amount of norfloxacin powder was added to 5ml of each excipient (oil, surfactant, and co-surfactant) contained in small, firmly sealed glass tubes. After 3 days in a shaking water bath at (25±0.5)°C, the tubes were centrifuged at a rate of 3000 rpm for twenty minutes, and the supernatant layer was filtered with a 0.45 m syringe filter. Methanol was used to dilute the samples. A UV-visible spectrophotometer (Biotech Engineering Management Co) is utilized in order to determine solubility. The measurement was carried out in triplicate.8,9\n\nPseudoternary phase diagrams for oil (oleic acid), surfactant (tween20) and co-surfactant (PEG200) represented as Smix were developed. Water was obtained at room temperature using the water titration technique, and each of them represents a side of the triangle. Several Smix (1:1, 1:2, 1:3, 1:4 and 2:1, 3:1, 4:1) ratios of surfactant and co-surfactant were utilized, and oil to particular Smix ratios (ranging from 0.5:9.5 to 9.5:0.5) were prepared.10,11\n\nTo manufacture norfloxacin liquid self-emulsifying drug delivery (SEDD) formulations, a vortex mixer was used to combine norfloxacin (400 mg) with oleic acid oil, tween 20, and PEG200 (Smix) in a screw-capped glass vial. The mixture was then warmed in a water bath for half an hour at 50°C to induce homogeneity.12 The (28) produced formulae’ components are shown in Table 1.\n\n\nEvaluation of prepared norfloxacin SEDD\n\nThe liquid SEDDS formulations were diluted in two separate 1000 ml beaker with deionized water and 0.1 N HCl (50, 100, 250, and 1000 times) to mimic in vivo environments. The diluted systems were shaken with the aid of a magnetic stirrer to ensure complete homogeneity and done at 37°C to simulate body temperature.13\n\nThese systems were kept at room temperature for 24 hours before being visually examined for signs of phase separation, drug precipitation, and droplet coalescence.14\n\nCentrifugation test\n\nThe SEDDS formulations were centrifuged at 3500rpm for 20 minutes to check for phase separation, creaming, or cracking.15 Following the centrifugation test, the formulations that succeeded will be put to another thermodynamic test.16\n\nHeating-cooling cycle (H/C cycle)\n\nThe stabled formulas were subjected to six cycles of heating and cooling at two different temperatures (4°C and 45°C). Formulas would stay at each temperature for at least 48 hours. The formulas that did not show any phase separations or creaming were subjected to freeze-thraw cycles.17\n\nFreeze-thaw cycles\n\nStable SE formulas were subsequently subjected to freezing and thawing (-20°C and 25°C). They were carried out in three cycles, each lasting 48 hours.18\n\nSelf-emulsification time and dispersibility test\n\nThe addition of liquid self-emulsify formulas to 500 mL of 0.1N HCl under gentle magnetic stirring was visually observed. Emulsification time was established as the amount of time needed for the prepared formulations to completely disperse and form an emulsion. When a transparent homogenous system was attained. The in-vitro efficiency of the liquid SEDDS formulations was examined visually. It is believed that the creation of clear SEDD occurs best within a minute, since prolonged times result in a milky look.19 As indicated by the grade mentioned in Table 2.20\n\nUsing a UV spectrophotometer, the drug content of the chosen formulations (F1, F2, F5, F6, F8, F13) that passed the previously indicated characterisation tests was determined. In a 100 mL volumetric flask, a known amount of norfloxacin-SEDDS was accurately measured. The flask was then filled with methanol and sonicated for around 15 minutes to ensure full drug dissolution. The solution was appropriately filtered via a 0.45-micrometer membrane, diluted, then tested for drug concentration with a UV-spectrophotometer at 278 nm.21–24\n\nThe droplet size investigation was carried out using a dynamic light scattering approach, which examines the variance in light scattering caused by particle Brownian motion. Light scattering was measured at room temperature at a 90° angle of laser scattering.25\n\nThe liquid norfloxacin-self emulsify formulation was diluted hundred times (0.25mL from each formula in 25 mL) with filtrated distilled water and gently agitated with a magnetic stirrer to generate a fine emulsion. The diluted emulsion was poured in a disposable cuvette in the sample chamber to analyze droplet size and PDI.26\n\nDissolution apparatus-II was used to establish in-vitro drug release profiles of SEDDS formulae as well as norfloxacin tablet. The medium was 750 ml of acetate buffer pH,4 and the test was carried out using stirring paddles at a speed of 50 rpm and at a temperature of 37°C. The dialysis bag approach was used for drug release investigations to obtain true amounts of free drug without interference with unreleased drug from the formula.27 Aliquots (5 ml) of samples were obtained at 5,10,15,20,30,45,60 minute intervals. To maintain a sink condition, the same quantities of new dissolving medium (ph4) were replaced. The amount of drug dissolved was filtered and measured with a UV-vis spectrophotometer at 278 nm.28\n\nThe zeta potential analyzer was used to examine the best liquid SEDD formulation (F1) (Brookhaven instrument). The zeta potential value indicates the repulsive forces between the droplets.29,30 The aqueous dispersion of the produced SE formula was made in the same manner as in the section of globule size measurement.\n\n\nResults and discussion\n\nThe solubility of norfloxacin in different oils, surfactants, and co-surfactants was determined as showen in Table 3. To find suitable vehicles with a high solubilizing ability for norfloxacin. The importance of maximum drug solubility is first and mainly an optimization of drug loading capacity. Second, drug precipitation is avoided during dilution, and the final volume of SEDDS is reduced, making it acceptable for oral administration.31 At last, while storage, the whole system’s stability will improve.\n\nThe solubility of norfloxacin in different oils is as follows: oleic acid >olive oil >clove oil >castor oil >corn oil > spearmint oil >coconut oil >mineral oil.\n\nAs oleic acid had the maximum solubility for norfloxacoin, it was chosen as the oil phase of SEDDS.\n\nThe solubility of norfloxacin in various surfactants reveals that the maximum solubility was observed in tween 20 (non-ionic surfactant with HLB values = 16.7),32 which was chosen as the best surfactant.\n\nNorfloxacin solubility was also investigated in several co-surfactants that are critical for interfacial film flexibility and decreasing interfacial tension.33\n\nThe medication had a low solubility in PG, but the maximum solubility was seen in PEG200, thus it was chosen.\n\nIt was observed from phase diagrams in Figure 1 that increasing the ratio of surfactant to co-surfactant in Smix from 1:1 to 2:1 or 3:1 improved the nanoemulsion shaded area. Moreover, there was a reduction in nanoemulsion area at a Smix 4:1 ratio. The reason for the increased clarity of the emulsions produced as Smix increased was a decrease in the total system’s free energy. Thus, smaller emulsion droplet size reflects a formulation with greater visual clarity. On the other hand, a further increase in Smix may facilitate water breakthrough oil globule, resulting in their disruption and more oil globules released into the aqueous phase.34 In addition, as the co-surfactant (PEG200) concentration increased (1:2, 1:3, and 1:4), the nanoemulsion area decreased in comparison to the 1:1 ratio, which was attributed to the decrease in surfactant (tween 20) concentration from 50% in the 1:1 ratio to 20% in the 1:4 ratio confirming the fact that the surfactant has a primary role by being adsorbed on the surface of oil droplets to provide the essential barrier for the formation of o/w emulsion while the co-surfactant serves as an adjuvant by giving flexibility to the formed barrier film, further lowering the interfacial tension.35 These results concurred with that observed by different studies on self-nanoemulsification.36–38\n\nThe optimum o/w nano-emulsion area was produced with a low oil: Smix ratio (1:9, 2:8, 3:7, 4:6) because it provided superior stability and a smaller droplet size; this result was consistent with the data.39 As a result, these oil: Smix ratios were chosen for further investigation. Furthermore, raising the oil concentration reduced the nanoemulsion region, which might be due to increased interfacial tension and decreased efficacy of surfactant and cosurfactant to overcome it.\n\nBased on pseudoternary phases diagram,twenty-eight liquid SEDD formulae were produced utilizing various tween 20: PEG200 (Smix) ratios of 1:1, 1:2, 1:3, 1:4, 2:1, 3:1, and 4:1, and oil: Smix ratios of 1:9, 2:8, 3:7, and 4:6. All of the produced formulations underwent a visual assessment of their appearance and other physical characteristics.\n\nThe twenty-eight anhydrous formulae that were synthesized had a uniform, clear yellow color and exhibited no evidence of phase separation or drug precipitation.\n\nRobustness to dilution\n\nThe SEDDS must be able to resist dilution without exhibiting any phase separation or drug precipitation in order to be employed as a drug delivery vehicle, and diluting SEDDS formulations up to 1000 times with all of the abovementioned diluents can mimic the in-vivo conditions.40 As shown in Table 4, the majority of reconstituted formulations demonstrated good emulsion stability, as evidenced by the preservation of their appearance with no evidence of drug precipitation or phase separation in all dilution media after 24 hr of storage at room temperature. This suggests that such systems may be suitable for oral administration, as they have a high probability of maintaining their emulsified state throughout their passage in the GIT with no phase separation.41\n\nSeveral formulations failed this test and became turbid and unstable after 24 hours, possibly due to a low concentration of tween 20 in these formulations that if the formulation comprises water-soluble co-surfactant, precipitation is frequent and can be prevented by raising the surfactant concentration.42 In addition, the low concentration of Tween 20 was insufficient to emulsify the high oil content, particularly when it was partially replaced with PEG200. Yet, as the concentration of surfactant rises, both surfactant and co-surfactant exert their functions correctly.\n\nFurthermore, modifying the pH and content of the aqueous phase had no effect on the emulsion characteristics.40\n\nIt was also noted that as the concentration of oil increases, the formulation’s stability decreases.43\n\nThe SE formulations were visually evaluated after being subjected to centrifugation and abrupt temperature changes, including the heating-cooling cycle and the freezing-thawing cycle.\n\nThe observations of the thermodynamic stability investigation for several formulations of norfloxacin SEDDS were displayed in Table 5.\n\nThe formulations were visually assessed for signs of phase separation, cracking, or creaming. The results show that most of the formulations passed all three tests and were stable. However, some formulations failed one or more tests, indicating instability. The failed formulations may be prone to phase separation or other forms of instability upon storage or transportation.\n\nThe majority of the tested formulae had a short self-emulsification time, demonstrating their capacity to make emulsion efficiently and simply.\n\nTable 6 shows the results of different grades of microemulsion and the measured self-emulsification times created by twenty-eight formulations with varying Smix ratios and oil proportions. The previously indicated visual grading method was utilized to evaluate the in-vitro performance of the tested formulae.\n\nThe study of emulsification time and visual observations revealed that six formulations achieved self-emulsification times of 1 minute with a clear appearance and were certified as grade A (the best) liquid SEDDS. It was also discovered that in formulas F1,F8,F15,F22 containing oil: Smix ratios of 1:9, 2:8, 3:7, 4:6 respectively, keeping Smix ratios of 1:1 as the oil to Smix ratio increased, the process of emulsification slowdown, so when fixing Smix and increasing in oil content leads to a gradual increase in emulsification time. However, increasing the concentration of tween 20 resulted in a significant decrease in emulsification time, as well as improved uniformity and bluish color nature of the microemulsion, improving the spontaneity of emulsification.\n\nThe ability of surfactants to improve formation of an emulsion and dispersion by lowering the oil-aqueous interfacial tension may explain why higher surfactant concentration was related with shorter self-emulsification time.44\n\nThis implies that emulsification efficiency is determined by selecting the appropriate oil ratio as well as the appropriate Smix ratio that provided the optimum emulsification capability, rate, and stability.45\n\nThe drug content of the selected formulae is provided in Table 7. The drug content of norfloxacin SEDDS formulations was meeting USP requirements and falling within an acceptable range (85%-115%). These findings indicate that there was no drug precipitation in the developed formulations and that norfloxacin was homogeneous and stable in SEDDS vehicles.46\n\nThe experimental results of globule size and PDI measurement for norfloxacin-nanoemulsion in distalled water were presented in Table 8. At a constant Smix ratio, it was discovered that increasing the oil% with a subsequent decrease in the quantity of surfactant and co-surfactant in the mixture resulted in a slight increase in globule size. The explanation might be due to a lack of surfactants and co-surfactants that can localize at the oil/water interface of a droplet to reduce interfacial tension and stabilize the system.47\n\nDroplet size distribution is an important issue to consider when developing SMEDDS. The lower the PDI value, the narrower the droplet size distribution, indicating uniformity, homogeneity, and mono-dispersion of the formulated self-nanoemulsion, as well as long term stability.48 All SEDDS formulae had low PDI values, indicating that oleic acid globules were dispersed within the formulation and confirmed its homogeneity.34,49–52\n\nFigure 2 depicts the in-vitro release profiles of the produced formulations and the norfloxacin tablet in PH4 dissolving media over 1 hour. At the end of 1 hour, the majority of tested SNEDDS formulae demonstrated a consistently better release rate than the tablet. The rapid spontaneous emulsification capabilities of SNEDDS, as well as the formation of small globules with a high surfactant content upon dilution, may explain the increased in-vitro release rate and extent.53\n\nAs compared the release from the formulation with constant oil% w/w and an increase in Smix, a clear decrease in norfloxacin releasing rate was seen because high Smix may cause drug sequestration inside micelles or oil droplets.29 It was discovered that increasing the co-surfactant yielded the highest dissolution rate. The reason for this is that co-surfactant works as a solubilizer, providing high thermodynamic activity of norfloxacin and therefore facilitating dissolution.47,54\n\nWhen drug release was compared between optimum liquid SEDD formula of norfloxacin (F1) and norfloxacin commercial Tablet in ddsolver; the F1 formula showed a difference than tablet (f1 = 15.25, If (f1) ∈ [0-15], this means the reference formula is similar to tested formulations) higher release than the commercial Tablet as shown in Figure 3, and this is related to the ability of F1 to form spontaneous nano-emulsion.\n\nAccording to the thumb rule, the zeta potential is an essential predictor of the nanoparticle stability in dispersion medium when the result between (-5 mV to +5 mV) suggests quick agglomeration and the result between (-20 mV to +20 mV) gives short-term stability. Levels of zeta potential between (-30 mV) to (+30 mV) suggest moderate stability, while values in the range (-60 mV) to (+60 mV) indicate excellent stability in the formulation.55 The calculated zeta potential for the best liquid SEDD formula F1 was +60.78, as showen in Figure 4 indicating excellent stability.\n\n\nConclusions\n\nAn oral administration of spontaneous dosage form of norfloxacin that can produce in-situ nano-emulsion in the gastrointestinal tract providing a stable SEDDS that may increase drug absorption and bioavailability was successfully developed for the first time. Norfloxacin liquid dosage form is also required since a number of individuals, particularly pediatric and geriatric patients, have difficulties swallowing solid dosage forms.", "appendix": "Data availability\n\nZenodo: pseudoternary phase diaghram.xlsx https://doi.org/10.5281/zenodo.8219346. 56\n\nZenodo: RELEASE OF DRUG.xlsx, https://doi.org/10.5281/zenodo.8219350. 57\n\nZenodo: saturated solubility.xlsx, https://doi.org/10.5281/zenodo.8219352. 58\n\nZenodo: RELEASE.xlsx, https://doi.org/10.5281/zenodo.8216275. 59\n\nZenodo: ph 4.xlsx, https://doi.org/10.5281/zenodo.8216277. 60\n\nZenodo: drug content.xlsx, https://doi.org/10.5281/zenodo.8216279. 61\n\nZenodo: methanol.xlsx, https://doi.org/10.5281/zenodo.8216283. 62\n\n\n\n‐ f1 Particle size. https://doi.org/10.5281/zenodo.8216249\n\n‐ f2 particle size. https://doi.org/10.5281/zenodo.8216267\n\n‐ f5 particle size. https://doi.org/10.5281/zenodo.8216261\n\n‐ f6 particle size. https://doi.org/10.5281/zenodo.8216259\n\n‐ f8 particle size. https://doi.org/10.5281/zenodo.8216269\n\n‐ f13 particle size. https://doi.org/10.5281/zenodo.8216273\n\n‐ ddsolver. https://doi.org/10.5281/zenodo.8219340\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nWannas AN, Maraie NK: Preparation and in-vitro evaluation of cilostazol self-emulsifying drug delivery system. 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Publisher Full Text\n\nAbdulkareem NM, Sahib MN: RELEASE OF DRUG. [Dataset]. 2023. Publisher Full Text\n\nAbdulkareem NM, Sahib MN: saturated solubility. [Dataset]. 2023. Publisher Full Text\n\nAbdulkareem NM, Sahib MN: RELEASE. [Dataset]. 2023. Publisher Full Text\n\nAbdulkareem NM, Sahib MN: ph 4. [Dataset]. 2023. Publisher Full Text\n\nAbdulkareem NM, Sahib MN: drug content. [Dataset]. 2023. Publisher Full Text\n\nAbdulkareem NM, Sahib MN: methanol. [Dataset]. 2023. Publisher Full Text" }	[ { "id": "250875", "date": "27 Mar 2024", "name": "Himanshu Paliwal", "expertise": [ "Reviewer Expertise Lipid-based drug delivery system", "and Pulmonary drug delivery systems" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nI recommend rejection of manuscript because of porrly written manuscript with number of scientific inaccuracies, basic grammatical mistakes, unclear description about the methodology, and vague discussion about the results. In particular, the following issues were observed: 1. Abstract: Abbreviations should be used appropriately, For eg. SNEDDs, SEDD or SNE are given in abstract itself. Choose one of these to describe the formulation and use it consistently throughout the manuscript. 2. Introduction: The section is poorly written with limited information, and further following points can be looked upon: a) Support the statement by citing some recent researches where first pass effect was avoided successfully. b) Authors need to be careful while using abbreviation SNEDD for self-nanoemulsifying and similarly for SMEDD. c) Can author explain differences between SNEDD and SMEDD. especially how preconcentrates can exclusively form SMEDD and SNEDD upon administration. d) Section 2 of introduction can be combined with section 3 e) The summary of study should be discussed more comprehensively.\n3. Methods a) Construction of Pseudoternary phase diagram: The reason behind conducting phase diagram should be discussed. Water titration technique was not properly explained or rather wrongly explained. Correct the sentences and expand appropriately to make it understandable for the readers. b) Development of a liquid self-emulsifying drug delivery system (SEDD): How can authors justify the use of SEDDS for delivery of high concentration of drug (400 mg). Did authors checked appropriate emulsification capacity after loading of drug? c) Evaluation of prepared norfloxacin SEDD: SEDDS or SEDD, Please decide and use it throughout the manuscript. Table 1: Units should be indicated in bracket, then no need use it within each cell\nFigure 1: Justification is required to explain very low nanoemulsion region for all the components even though authors reported high stability upon emulsification.\nResults and discussion a) Pseudo-ternary phase diagram construction: \"The optimum o/w nano-emulsion area was produced with a low oil: Smix ratio (1:9, 2:8, 3:7, 4:6) because it provided superior stability and a smaller droplet size; this result was consistent with the data.\" Is it low oil:Smix ratio, or higher Smix as compared to oil? b) Development of a liquid self-emulsifying drug delivery system (SEDD): Visual assessment of preconcentrates or emulsified form of SEED c) In-vitro drug release study of norfloxacin SEDDS: The details regarding commercial tablet used for the comparison with SEDDS should be included. Also, the reason for using the tablet for comparison with SEDDS should also be indicated. Why pure drug suspension was not considered for comparison?\nFigure 3: The graph legends should be consistent with their description in Figure caption and discussion.\n\nIs the work clearly and accurately presented and does it cite the current literature? No\n\nIs the study design appropriate and is the work technically sound? No\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNo\n\nAre all the source data underlying the results available to ensure full reproducibility? No\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "232537", "date": "25 May 2024", "name": "Asif Nawaz", "expertise": [ "Reviewer Expertise Nanotechnology", "Transdermal Drug Delivery" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nCurrent research focus on enhancing the solubility of norfloxacin using SEDD. The following points should be considered before publication. 1. add latest references in introduction part. 2. Intro part need more informations like problems associated with low solubility of drug. 3. why tween 20 used unto 45%?? any reference for this concentration. 4.Results section needs more discussion.\n\n5. Conclusion should be revised.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [] }, { "id": "238464", "date": "01 Aug 2024", "name": "Joe Viljoen", "expertise": [ "Reviewer Expertise Developing drug delivery systems", "SEDDS", "SMEDDS", "SNEDDS", "lipophilic drug delivery", "natural oils", "Tuberculosis", "improving drug delivery" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis could have been a good study if the approach had been different. Moreover, norfloxacin has been discontinued in the US due to safety concerns. Why then would one want to produce a product for geriatrics and pediatric patients if it is not considered safe to use in its current commercial products? Overall, the following remarks are important:\nMajor language editing is furthermore necessary. The authors do not write coherently. They, for example, start off in the abstract with indicating that “the goal of this work was to create a self-nanoemulsion (SNE)…”. They then move on (Abstract, Methods) to stating that “…several formulations of liquid SNEDDS of the antibiotic were created..”. After this, they indicate, still in the Abstract (Results) that “The in-vitro dissolution profile suggested an optimum norfloxacin SEDD formulation…”. During the whole document they change between SNEDDS, SMEDDS, SEDDS, etc. At the end one is wondering what the actual aim was as well as what was actually obtained. The authors use the words “formula” and “formulae”, instead of “formulation” or “formulations”, which are rather used to express mathematical relationships or rules expressed in symbols. I suggest rather using “formulation” or “formulations” to indicate that certain preparations were manufactured or formulated. They provide a table (Table 1) to show the different formulations that were prepared and then provide a “Formula code” that was listed from F1 to F28. From here on, they only indicate, for example, “F1” or “F1, F8, F15, F22” to show differences in results obtained. However, this is extremely difficult to then try and discern which formulation is which. The reader constantly needs to revert back to the table in an attempt to establish how the different formulations differed in excipients – which in itself is a mission. They should rather use more specified codes when referring to the different formulations, for example: 1:1(O10T45P45) – F1. The authors used 55 References in an attempt to show the novelty of their work, however, 51% of these references are older than 10 year; 45.5% are between 5–10 years; and only 1.8% may be considered more relevant references. References 56–62 are self-referencing documents – datasets. Numerous references were also not correctly referenced.\nAbstract:\nMajor language editing is suggested. Why is the goal to increase the dissolving rate of norfloxacin? Should the aim not be to enhance the dissolution of this drug? Please explain how oral bioavailability will be improved. It is claimed in the Abstract that lymphatic transport occurs, however, nothing is explained in the Introduction. The “Method” part does not explain the methods used in this study. It only indicates that formulations were “created”, however, the preparation method or testing methods are not even mentioned. The “Results” part is only recognizing a single experiment, i.e., dissolution results, and it just mentions the one formulation’s zeta potential, not confirming why one formulation may be claimed as optimum. The “Conclusion” part is questionable. See explanation under “Conclusion”.\nIntroduction:\nMajor language editing is suggested,it does not directly link with the aim of this study. Single sentence paragraphs are furthermore found in this section. The authors state: “SEDDS improve the oral bioavailability…” – this is a dangerous statement as it has not been proven in all cases. Rather indicate that “It has been found in several studies that SEDDSs can improve oral bioavailability”. Also, the authors claim that the first pass effect is avoided when using SEDDS. This is NOT true at all! To avoid the first pass effect, the whole dosage should be absorbed by means of the lymphatic system, which does not occur. Only some of the dissolved drug will enter into the lymphatic system, rendering most of the drug exposed to first pass metabolism. It is stated that due to the difference in diameter of the emulsion droplets generated, the emulsions are classified into two self-emulsifying drug delivery systems. It is actually three different systems. Also, the classification is not ONLY based on droplet size. Abbreviations are given without defining them first (e.g., BCS). Abbreviations that were provided are not used throughout the paper (e.g., NOR). Why use it then? In this section it is stated that the “goal of this project is to develop norfloxacin as an oral self-emulsifying liquid formulation that will improve solubility and thus bioavailability”. This statement differs from the Abstract. What is the actual aim here? Self-nanoemulsifying or self-emulsifying drug delivery system? Be consistent! Secondly, was bioavailability tested? If not, do not make such a statement.\nMethods:\nAgain, major language editing is suggested. In the “Materials” part it is indicated that certain oils were selected. However, nowhere is it explained where these oils were used and in what concentrations they were included in the formulations. Were these oils combined? Were only some of them used? This is a major concern. More concerning is that the authors only indicate the oleic acid content of the different formulations. Did they extract only the oleic acid content? Or did they only show the oleic acid content of the different oils used in the formulations? If they included the oils in the formulations, they should then have considered the total content of the oils as the other fatty acids will most definitely have significant influence on all of the experimental results.\nWas their method of analysis validated? Nothing is stated. Why was UV- visible spectroscopy used for solubility determination? The generally used method is HPLC-analysis.\nConstruction of pseudo-ternary phase diagrams: It is suddenly indicated that oleic acid is the oil phase. How did this transpire? What is Smix? This abbreviation is again not defined. The different ratios are confusing and are not well explained. Development of a self-emulsifying drug delivery system (SEDD): Why is a vortex mixer used? SEDDSs are supposed to form naturally with only mild agitation.\nWhat is this oleic acid oil? Why were the formulations heated to 50⁰C? How will this induce homogeneity? Body temperature is only 37⁰C and it is at this temperature that SEDDSs need to self-assemble for the formulation to be considered optimum. Heating the formulations to a temperature significantly higher than body temperature will notably increase the kinetic energy, which in turn, will significantly change the formulation’s self-emulsification ability. This method is clearly not appropriate, and the claim of acceptable self-emulsification ability cannot stand.\nEvaluation of prepared norfloxacin SEDD: First, this heading does not form part of the methodology, which it should. Next, the abbreviation, SEDD, is not a standard abbreviation. Robustness to dilution: Now a different abbreviation (SEDDS) is used. This is also not the standard method used to test robustness to dilution. Why was deionized water used? It is stated that it was used to mimic in vivo environments. Which in vivo environment was mimicked? Again, why were the systems shaken, and this time with a magnetic stirrer? The authors claim that they wanted to obtain complete homogeneity. If you take measures to obtain homogeneity, how can you then claim that formulations that formed naturally, are indeed homogeneous? At what pH were the experiments conducted at? The pH of the formulations is very important as it will determine the ionization and subsequently the solubility of the drug (which is cationic) and there may even be some complexation occurring as the other excipients used are all anionic in character. What about mouth feel and taste – pH plays an important role here as well. Why was the pH not determined?\nTable 1 is very confusing – what oils, named in the materials section, were used?\nInvestigating the thermodynamic stability: Again, an abbreviation is given that is not used anywhere else (H/C cycle). They also now refer to “stabled formulas”. Which of the formulations were considered stable? And why? Freeze-thaw cycles: What are SE formulas? Self-emulsification time and dispersibility test: Why was 500 ml used? What was the pH of the media and of the formulations? At what temperature was this study conducted? What was the stirring rate? What apparatus was used for this study? Did the authors even test spontaneous self-emulsification? It is indicated that: “When a transparent homogenous system was attained, the in-vitro efficiency of the liquid SEDDS formulations was examined visually. It is believed that the creation of clear SEDD occurs best within a minute, since prolonged times result in a milky look.” First, was the time it took to form a homogenous dispersion noted? The authors stated that only when a transparent homogenous system was attained, did they visually examine the formulations. What if no transparency was attained then?\nTable 2: This table used is NOT correct and NO reference is indicated for this table. The frequently used table (see below) indicates that grading is done based on the time it took for self-emulsification to occur. It furthermore differentiates between appearance after certain times. Thus, grading is concerned with emulsion formation and not color formation. It is NOT based (as claimed by the authors) on the fact “that prolonged times result in a milky look”. https://f1000research.s3.amazonaws.com/supplementary/141057/93f5b5b0-9220-4ef3-9d78-8b6949b8b794.png\nCzajkowska-Kośnik A, et. al. 2015 (Ref 1) (Example)\nDetermination of drug content: What is meant with “a known amount of norfloxacin-SEDDS was accurately measured”? How much methanol was added? Again, was the method validated? Why was HPLC not utilized? This method does not differentiate between dissolved drug incorporated in the SEDDSs (i.e., drug content in SEDDS) and undissolved drug that is not really part of the formulation. The % encapsulation efficiency (%EE) should rather have been tested. As this method is written here, it indicates that this experiment did not test drug content, it tested the accuracy of the analyst that measured the ingredients. Size of globules and polydispersity index (PDI): Why use the words globules and droplets interchangeably? Why was “the diluted formulations agitated with a magnetic stirrer to generate a fine emulsion”? This manipulates the formulation and the true droplet size is therefore not measured. This is NOT an accurate experiment! In-vitro drug release study of norfloxacin SNEDDS: Now suddenly the formulations are referred to as SNEDDS. Why again vary between what type of SEDDS was obtained? Why were dissolution studies done in 750 ml and with an acetate buffer at pH 4? This is NOT the standard volume or pH simulation of the stomach where the SEDDSs need to form? Zeta potential determination: This should actually form part of the initial characterization experiments. Again, now the authors refer to: SE formula. It is furthermore stated that: “The aqueous dispersion of the produced SE formula was made in the same manner as in the section of globule size measurement.” Nothing is stated in the mentioned section on how this formulation was “made”. The formulation was diluted and agitated…which is not a correct method!\n\nResults and discussion:\nMajor Language and technical editing necessary. Determination of norfloxacin saturated solubility: Drug loading was not tested in this study. Why mention it then? Again, %EE should have been tested. What is meant with the sentence: “At last, while storage, the whole system’s stability will improve.”? Clarify. Do not use 1 sentence paragraphs. Why were the HLB-values of the oils, surfactants, and co-surfactants not a consideration? – only the HLB-value for Tween 20 was given, which is very high. Did you establish the required HLB-value for the oil and relate it to the amount of surfactant and co-surfactant included? Pseudo-ternary phase diagram construction: None of the results in this section are clearly displayed. Figure 1 is not of a good quality, and one cannot discern between the “Smix” ratios referred to in-text. Also, it seems that the first pseudo-ternary phase diagram is incorrectly defined as “Smix(4:1); or was the two diagrams switched. These diagrams do not correlate to what is attempted to be explained in-text. No additional information is provided in the figure caption. Also, in-text the authors refer to “Smix from 1:1 to 2:1 or 3:1…”, however, on the figure it is: “Smix(4:1); Smix(1:2); Smix(1:3); Smix(1:4). This should be clarified.\nFurthermore, the authors indicate that: “The reason for the increased clarity of the emulsions produced as Smix increased was a decrease in the total system’s free energy.” This is an unsubstantiated statement. No evidence is provided, and the free energy was not tested. Where is the proof (maybe include referencing as well)? When you increase the surfactant/co-surfactant ratio, there is a danger that you are moving into not creating emulsions, but rather moving into a multiphase dimension or even a bicontinuous dispersion phase. I also do not agree with the statement that when the co-surfactant concentration is increased, the nanoemulsion area decreases. Co-surfactants are specifically added to be able to decrease the surfactant concentration without limiting the self-emulsification area. I do believe that the hydrophilicity of the surfactants/co-surfactants played a significant role here. The statements on why improved nano-emulsions form when the oil concentration is decreased, is nothing new and quite logic.\nDevelopment of a liquid self-emulsifying drug delivery system (SEDD): Based on what did you choose the 28 formulations? What is meant by “anhydrous formulae”? Should it not rather be emulsion-concentrate? Characterization of the prepared liquid SEDDS of norfloxacin: Again, what is meant by: “…and diluting SEDDS formulations up to 1000 times with all of the abovementioned diluents can mimic the in-vivo conditions”? How does deionized water and a 1000 times dilution mimic in-vivo conditions? Also, in Table 4 abbreviations are given that have not been defined. Then, 0.1hcl vs 0.1 N HCl? Do the different “Formula no.” in Table 4 correlate with formulations previously given in Table 1? No explanation of Table 4 is given. Why does one “want to maintain their emulsified state throughout their passage in the GIT with no phase separation”? Where should this drug be absorbed? You also did not test this in different pH-ranges.\nThen did you actually test if the surfactant and co-surfactant exert their functions correctly? How was this established? One cannot make such a statement if it was not tested or backed by ample evidence. Again, one sentence paragraphs. It is stated: “Furthermore, modifying the pH and content of the aqueous phase had no effect on the emulsion characteristics”. Nothing was included in the methodology that the pH of the environment was varied. No pH-values were even provided. When and how was this tested? “It was also noted that as the concentration of oil increases, the formulation’s stability decreases.” – This statement is nowhere clearly explained. The section on “Thermodynamic stability studies” is poorly written and vague – “some formulations passed….some formulations failed…”. In the section on “Self-emulsification time and dispersibility test”, please explain what is meant by: “to make emulsion efficiently and simply.” Then, the authors now suddenly refer to the formulations as microemulsions. Why? The authors state: “This implies that emulsification efficiency is determined by selecting the appropriate oil ratio as well as the appropriate Smix ratio that provided the optimum emulsification capability, rate, and stability.” They did not attempt to further explain or discuss this statement. Also, was stability tested? No suggestions are given as to how to go about selecting the appropriate ratios. The reader is therefore not wiser.\nDrug content: The reason why the drug content of all the formulations complies with the USP standards, is that the method used is wrong. There would not have been any drug precipitation as “The flask was then filled with methanol and sonicated for around 15 minutes to ensure full drug dissolution. The solution was appropriately filtered via a 0.45-micrometer membrane, diluted, then tested for drug concentration.”. – Full drug dissolution was ensured; an unspecified amount of methanol was used; and the solution was filtered. No distinction between drug dissolved in the oil phase and drug in the aqueous phase is made. Measurement of the globules size and polydispersity index (PDI): The authors have misspelled “distilled water”; and they are now again referring to “nanoemulsion”.\nPlease explain the difference between “globule size” (in-text) and “particle size (PS)” in Table 8. There is no consistency in the terms used. Suddenly, “SMEDDS” is used again….and then in the following sentence, “SEDDS”… Why was F1 chosen as the “optimum” formulation? Although F2 and F5, for example, depicted slightly larger droplet sizes, the %RSDs are smaller as well as their PDIs. Their dissolution profiles furthermore do not significantly (it seems) differ from F1.\nIn-vitro drug release study of norfloxacin SEDDS: The results indicated in the figures included, are not clearly distinguishable. The timeline on the graphs is poorly measurable. On Figure 2, the “chart title” was not defined. On Figure 3 it is indicated in the “chart title”: “Mean dissolution profiles with SD”. Does SD stand for “standard deviation”? If so, no SD was indicated on the graph. How many samples per formulation were measured as the “mean dissolution profiles” are indicated. Why are the %RSDs not provided? On the y-axis it is stated that the “fraction dissolved” was measured. Fractions are given as: “0.1–1.0. This is “percentage” measured. Additionally, no distinct values for dissolution studies are given. The figures do not even conform (vary significantly in view). Also, Figure 2 depicts “SNEDDS” and Figure 3, displays “SEDD” in their individual captions.\nThe initial dissolution rate was not measured. Therefore, no conclusion can be made regarding the release/dissolution rate. It was stated: “The rapid spontaneous emulsification capabilities of SNEDDS, as well as the formation of small globules with a high surfactant content upon dilution, may explain the increased in-vitro release rate and extent.” – Please explain your reasoning. No in-depth discussion is provided. Not wanting to go into too much detail, the following two sentences are contradictory: “As compared the release from the formulation with constant oil% w/w and an increase in Smix, a clear decrease in norfloxacin releasing rate was seen because high Smix may cause drug sequestration inside micelles or oil droplets.29 It was discovered that increasing the co-surfactant yielded the highest dissolution rate.” – Also, no statistics on this were available. Very important, the method for determining the F1 dissolution (dissimilarity) factor is NOT described in the methodology. It is not even properly described in this section. No references are provided on this subject. How many points were used to compare the formulations? When was 85% of the drug released from one of the formulations? Also, the F1 dissolution (dissimilarity) factor methodology indicates that a restriction is placed, i.e., if two dissolution profiles are to be considered similar and bioequivalent, f1 should be between 0 and 15. Please note, these values do not consider decimal values. Thus, f1-values should be rounded off to the nearest whole number. Subsequently, your tested f1-value should be rounded off to 15 (15.25), rendering these two dissolution profiles, similar. Your whole argument, that your formulation is considered or deemed superior to the commercial tablet product, is invalid.The conclusion is inappropriate.\nZeta potential determination: They talk about “a thumb rule”. Who provided this rule? They claim that the zeta potential is indicative of short term, moderate, as well as excellent stability. These terms are not properly defined. The authors do provide a reference (55). However, this reference was incorrectly referenced, as nowhere in this paper is it stated that: “According to the thumb rule, the zeta potential is an essential predictor of the nanoparticle stability in dispersion medium when the result between (-5 mV to +5 mV) suggests quick agglomeration and the result between (-20 mV to +20 mV) gives short-term stability. Levels of zeta potential between (-30 mV) to (+30 mV) suggest moderate stability, while values in the range (-60 mV) to (+60 mV) indicate excellent stability in the formulation.55”.\nThis should be explained. Also, the figure provided to indicate the “excellent stability” of the “best formula”, is very unclear and can even be left out. This figure does not provide any additional and/or interesting information. Again, what were the pH-values of the different formulations? This would most definitely have had an influence on the solubility of the drug as well as the zeta potential of the formulations as stated. Conclusion:\nThe authors claim that a “spontaneous dosage” form was developed. What is this? They also claim that this dosage form can produce “in-situ nano-emulsion in the gastrointestinal tract providing a stable SEDDS that may increase drug absorption….” First, this was not tested in a human participant; secondly, stability was not tested in this environment; and third, they claim that drug absorption may be increased. How can drug absorption be increased? This should be explained in detail! If they stated that drug solubility was probably increased, which could lead to more dissolved drug being available for absorption, it would have made sense, but claiming that absorption itself is enhanced, is taking it too far. In the Conclusion part of the Abstract, the authors stated that their formulation “counteract the inconsistent bioavailability of the commercial tablet.” Please include a reference that indicates “inconsistent bioavailability”. Although this drug is poorly bioavailable after absorption, it is still absorbed fast. \"Absorption of norfloxacin is rapid following single doses of 200 mg, 400 mg and 800 mg. At the respective doses, mean peak serum and plasma concentrations of 0.8, 1.5 and 2.4 μg/mL are attained approximately one hour after dosing.” (Tablets Noroxin (Norfloxacin) Merck Sharp & Dohme. USA: FDA. September 2008.) Therefore, poor/inconsistent bioavailability is not due to poor/irregular absorption, and no formulation will change the bioavailability. The conclusion is inappropriate.\n\nIs the work clearly and accurately presented and does it cite the current literature? No\n\nIs the study design appropriate and is the work technically sound? No\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNo\n\nAre all the source data underlying the results available to ensure full reproducibility? No\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [] } ]	1	https://f1000research.com/articles/12-1380
https://f1000research.com/articles/12-19/v1	06 Jan 23	{ "type": "Study Protocol", "title": "Protocol for a systematic review and meta-analysis on preoperative risk factors for failure after fixed sling implantation for post-prostatectomy stress urinary incontinence", "authors": [ "Emilio Sacco", "Filippo Marino", "Filippo Gavi", "Stefano Moretto", "Carlo Gandi", "Riccardo Bientinesi", "Francesco Pierconti", "Pierfrancesco Bassi", "Filippo Marino", "Filippo Gavi", "Stefano Moretto", "Carlo Gandi", "Riccardo Bientinesi", "Francesco Pierconti", "Pierfrancesco Bassi" ], "abstract": "Background: Post-prostatectomy urinary incontinence (PPI) is a bothersome complication affecting patients undergoing prostate surgery that in up 10% of cases will require an invasive treatment with fixed slings or artificial urinary sphincters (AUS). Although fixed slings have several advantages over AUS, failure rates after slings range between 15% and 45% while current knowledge of predictors of sling efficacy remains limited. By systematically combining and summarizing all relevant literature, the present review and meta-analysis aim to address this research need assessing the association between preoperative risk factors and sling failure. Methods: Studies pertaining to fixed synthetic male perineal slings as treatment for adult male suffering from PPI, will be included. A systematic search will be conducted in PubMED, Scopus, Web of Science and Cochrane databases, and in the reference lists of retrieved articles. Independent reviewers will conduct study selection and data extraction. Outcomes will include failure to achieve the continence cure and overall success (cure plus improvement), measured as per included studies. Exposures will include any preoperative variables evaluated for association with sling failure. The QUIPS tool will be used for study quality assessment and a random-effects DerSimonian-Laird model, with Hartung-Knapp adjustment, will be used to pool adjusted and unadjusted odds ratios separately. Sensitivity analysis will be performed using the leave-one-out methodology and subgroup meta-analyses based on pre-specified studies’ characteristics will be conducted to explain the heterogeneity. Certainty of evidence will be assessed according to GRADE methodology and review reporting will comply with the PRISMA-P statement. Discussion: By summarising all relevant literature in the field, our results will help to incorporate available evidence into clinical practice assisting healthcare professionals managing PPI patients in treatment decision-making. The present review will also provide researchers with the necessary, evidence-based groundwork to perform future high-quality prognostic studies in the field. Registration: CRD42022307160.", "keywords": [ "urinary incontinence", "male", "meta-analysis", "prostatectomy", "protocol", "sling", "systematic review" ], "content": "Introduction\n\nMale stress urinary incontinence is mostly of iatrogenic origin. In particular, post-prostatectomy urinary incontinence (PPI) is a bothersome complication of prostate surgery, resulting in deleterious impact on patients’ quality of life.1,2\n\nMost patients undergoing radical prostatectomy suffer from some degree of urinary leakage in the early postoperative period, and about 25% still require at least wearing one pad per day after one year, with little improvement afterward.3,4 In the Europa Uomo Patient Reported Outcome Study (EUPROMS), a cross-sectional survey conducted among 2943 prostate cancer patients currently receiving or having received treatment, 37% of all patients was using at least one pad per day because of urinary incontinence that was reported to be a moderate-to-big problem in 20% of post-prostatectomy patients.5\n\nApproximately 5-10% of patients with PPI are refractory to conservative treatments and will require a surgical therapy.6 Fixed slings and artificial urinary sphincters (AUS) are the currently most used and recommended surgical options for PPI7 Because of highly predictable efficacy, high patient satisfaction rates, durability and several decades of use and research, AUS is considered the gold standard treatment of PPI despite being an expensive mechanical device associated with non-negligible complications and reoperations rates.8,9 Several types of slings are currently available that differ in materials, design, size, mechanism of action and technique of implantation.10 Advantages of male slings are the non-mechanical nature, immediate post-operative results, lower cost and lower rate of severe complications than AUS.8 However, failure rates after slings placement range between 15% and 45% while current knowledge of predictors of sling efficacy remains limited.1\n\n\nProtocol\n\nWhile several systematic reviews and meta-analyses exist on the benefits and complications associated to male slings, to our knowledge no studies have systematically reviewed and meta-analysed predictors of slings failure. By systematically combining and summarizing all relevant literature, the current review and meta-analysis aim to address this research need assessing the association between preoperative risk factors and failure of fixed perineal synthetic male slings as treatment for adult male suffering from PPI, assessing the magnitude of these associations, determining the certainty in the cumulative evidence, and identifying gaps in knowledge. By helping to incorporate all available evidence into clinical practice, our results will assist healthcare professionals in patients’ selection and counselling.\n\nReview question\n\nThe review question was defined according to the PICOTS system as proposed by the CHARMS checklist and subsequent improvement (Table 1).11,12\n\nRegistration and reporting\n\nThe protocol for the current review has been registered on PROSPERO of the Center for the Reviews and Dissemination (CRD) (registration number: CRD42022307160). We followed the recommendations of the Preferred Reporting Items for Systematic Reviews and Meta-Analyses for the development and reporting of this systematic review protocol (see Extended data for PRISMA-P checklist)13,14 and will adhere to the PRISMA 2020 principles15 during the process of conducting and reporting this review. In performing this review we will follow the Cochrane handbook for systematic reviews of interventions,16 the recommendations of the Cochrane Prognosis Methods Group17–19 and other guidance20 for specifically conducting systematic reviews and meta-analyses of prognostic studies.\n\nSearch strategy\n\nSystematic searches will be performed for all citations published in the following electronic databases from their inception: PubMed platform in MEDLINE, Web Of Science, SCOPUS and Cochrane CENTRAL Register of Controlled Trials. Therefore, the searches will not be limited by historical time-constraints.\n\nSearch terms and sequential strategies used in the search algorithms are displayed in Table 2. Furthermore, a scrupulous manual search will be performed in the reference lists of retrieved articles and reviews for potentially additional eligible studies.\n\nElectronic search strategy.\n\nStudy selection\n\nApplicability to the review question will be based on the following criteria:\n\n‐ Inclusion criteria:\n\n• Longitudinal study in the form of randomised controlled trial and observational study (including prospective and retrospective study designs) involving the implantation of fixed synthetic perineal slings in male adults (aged ≥18 years) diagnosed with stress or stress-prevalent PPI assessed using any recognized diagnostic method, as follows: pad use, pad-test, stress test, urodynamics, bladder diary, symptomatology questionnaires.\n\n• The study includes at least one clearly defined risk factor and at least one clearly defined measure of continence. The risk factors must have occurred and measured preoperatively.\n\n• The outcomes have been measured after at least six month (mean/median follow-up) from surgery.\n\n• At least 30 patients have been assessed for the association between risk factors and outcomes.\n\n• The publication is in English, French, Spanish or Italian.\n\n• For inclusion in the quantitative meta-analysis, the study must report sufficient data on the association between any potential risk factor and outcome (effect size estimate and its variance or enough numerical information to calculate them). Studies that will not meet this criterion but only report p-values or qualitative findings (eg, ‘not significant’), and whose additional data will not be obtained on request, will be reported in a descriptive manner only, similarly to those that identified potential risk factors not featuring in any other study.\n\n• For studies published by the same research group, the choice of the study to be included will be made assessing each combination risk factor-outcome according to the following criteria in order of relevance: 1) the largest one reporting results of a multivariable analyses; 2) the largest one reporting results of an univariable analyses. This means that multiple studies by the same research group may still be included if reported risk factor-outcome combinations are different between studies.\n\n• Abstracts will be also considered for inclusion if in the form of indexed international conference abstract providing detailed description of key study characteristics (also by contacting the authors), and adjusted effect measures with variance.\n\n‐ Exclusion criteria:\n\n• The study does not meet any of the above inclusion criteria.\n\n• The study included more than 20% of patients with non-post-prostatectomy incontinence. No restrictions will be placed on other demographic characteristics.\n\n• The study involved:\n\n○ the use of an adjustable sling;\n\n○ the use of non-synthetic, absorbable or composite (autologous/absorbable plus synthetic) mesh materials;\n\n○ analysis of re-do procedures;\n\n• The citation is a case report;\n\n• The citation is a review or meta-analysis, book chapter, thesis;\n\n• The citation is an expert consensus, editorial, letter, comment, animal study, communication, or an abstract non fulfilling the inclusion criteria;\n\n• Duplicated studies not fulfilling the inclusion criteria for studies published by the same research group.\n\nA two-stage pilot-tested selection process will be undertaken. As a first step, relevant citations will be considered on the basis of their title and abstract. At the second step, the full-text of selected citations will be downloaded and assessed with regard to applicability and methodological quality.\n\nThree reviewers (FG, FM, SM) will independently perform the selection process without consideration for the results. Both screening stages will be piloted on a random selection of ten citations. During both stages, disagreements will be resolved through discussion among reviewers and, if necessary, with the senior reviewer (ES) until mutual agreement is reached. All citations will be recoded in Zotero 5.0 and a Microsoft Excel spreadsheet. The selection process will be displayed in a PRISMA flow diagram.\n\nData extraction\n\nKey information from each selected study will be extracted according to the CHARMS checklist12 in its modified version suitable for reviews of prognostic factors (CHARMS-PF).21\n\nKey data will be extracted in duplicate using a pre-defined data extraction form and recorded in a Microsoft Excel spreadsheet (available as Extended data: ‘Data Collection Sheet’) where the coding process for each of the variables is defined. The spreadsheet was created by two reviewers and thereafter pilot-tested and finalized on a random selection of five studies. The form will be reviewed and modified, if necessary, after group discussion with the senior reviewer.\n\nData items\n\nThe major groups of variables to be coded for each eligible study are as follows: (1) study’s general information (e.g., first author, journal, year of publication, country of the first author and other involved countries, study frame and design, recruitment period), (2) participant characteristics (e.g., inclusion/exclusion criteria, number of eligible and treated patients, age at baseline, body mass index, incontinence severity and aetiology, history of irradiation, bladder neck contracture), (3) intervention characteristics (e.g., type of sling, any comparator treatment), (4) outcomes (e.g., outcomes types and definitions, endpoints, event rates, patients evaluated for risk factors, attrition) and (5) risk factors and relevant quantities (evaluated variables and measurement methods, type of statistical analyses such as univariable versus multivariable, set of variables in the multivariable model, modelling method, effect size estimates with variance and p-value).\n\nAssessment of risk of bias\n\nThe quality of studies will be assessed for risk of bias (RoB) using the Quality in Prognosis Studies (QUIPS) tool,21 according to the recommendations of the Cochrane Prognosis Methods Group and the suggestions provided by Grooten WJA et al.22 The QUIPS tool consists of six domains: (1) study participation, (2) study attrition, (3) prognostic factor measurement, (4) outcome measurement, (5) study confounding, and (6) statistical analysis and reporting.21 Each domain encompasses three to seven prompting items that are rated on a four-grade scale (yes, partial, no, unsure). Eventually, the reviewer makes an overall judgment of the RoB within each domain (high, moderate or low RoB). This tool has demonstrated satisfactory reliability.21\n\nAlthough for QUIPS it is recommended against calculating scores for overall study quality, in systematic reviews and meta-syntheses it is recommended to report a table of included papers in which each paper is classified as having high, moderate or low RoB.22 We based this classification on the criteria proposed by Grooten et al.22: a study satisfying low risk of bias in at least five domains and without any high risk rating will be designated as a study with an overall low RoB. If one or more domains will be classified as having high RoB, or ≥ 3 domains as moderate RoB, then this paper will be classified as high RoB. All papers in between will be classified as having moderate RoB. On the other hand, no prior criteria will be used to make the judgment of the RoB within each domain but each rater will make the decision based on a critical overall evaluation of their ratings of the included items. The QUIPS checklists (available as Extended data) will be operationalized by the reviewer team to assess and adapt the prompting items for our specific review question and assign the agreed-upon a priori relevance to each item according to recommendations on RoB assessment.19,21,22\n\nTwo authors (FG and SM) will independently perform the quality assessment after a piloted screening of five studies. They then will meet and review their judgments for agreement. If agreement will not be reached, a third author (ES) will render the decision. Lack of adjustment is a serious bias and all studies will be judged at high RoB with regard to unadjusted effect sizes (secondary meta-analysis – see below). Overall and study-level RoB assessment will be displayed using the ROBVIS web app.23\n\nMeta-analysis design\n\nOutcomes\n\nThe outcome of the present review will be the sling failure. Studies on male slings typically report two main types of treatment failures: the failure of “cure” including patients failing to achieve a complete (“dryness”) or pretty complete continence, and the failure to achieve at least an “improvement” of the baseline incontinence status (also called “social continence” or “overall success” or simply “success”). Although heterogeneity in the definitions of these outcomes is expected in sling literature, we aim to evaluate the association between risk factors and both failure to achieve cure (failure of cure - FoC) and failure to achieve overall success (failure of success – FoS). Methods of measurement will be as per included studies. “No pad use” and “use of at maximum one pad per day” will be the preferred definitions for cure and overall success, respectively, and whenever possible, to increase homogeneity, effect sizes will be calculated based on these definitions; otherwise, outcomes definitions will be as per the included studies.\n\nExposures\n\nExposures (possible risk factors) will include any preoperative variables evaluated for association with sling failure, independently from their possible causal relationships. Although our focus is mainly on “risk factors”, potential protective factors will be also included. Comparators will be absence or fewer values of all of these variables.\n\nEffect size measures\n\nOdds ratio (OR) with 95% confidence intervals (CI) will be reported as an overall synthesized measure of effect size. The association is reported following the convention that OR>1 indicates that the factor under evaluation is associated to an increased risk of failure, and OR<1 indicates that the factor has a protective effect. Studies on male slings are predominantly observational in nature and thus liable to confounding, thus current guidelines for prognostic studies recommend to report both crude and adjusted measures of association to better understand the role of confounding variables.18 Consequently, although most of the reported effect measures are expected to be derived from univariable (unadjusted) analyses, we will primarily focus on adjusted estimates because they presents a more accurate picture of the unique (“independent”) contribution of the risk factors to the outcome of interest.19 As a result, we will perform separate meta-analyses based on adjusted (primary meta-analysis) and unadjusted (secondary meta-analysis) ORs for both continence outcomes. Studies will be included in the primary meta-analysis only if adequate statistical control is provided to account for the effect of relevant covariates on the association between risk factors and outcomes. Adequate adjustment is defined as statistical control for at least one of two variables: pelvic irradiation and incontinence severity. These two variables are “index” risk factors usually measured in PPI literature and reported as negative prognosticators in several reviews, primary studies and guidelines on PPI surgery.24–28 Therefore, for the other risk factors under review, it is important to understand whether they contribute additional and independent prognostic information to the index ones. For the severity of incontinence, different methods of measurement will be allowed (e.g., pad count, pad-test). The secondary meta-analysis of crude ORs will be performed separately on an expectedly much larger sample of studies, an established approach in reviews on prognostic factors to examine the robustness of results from the primary analysis, the role of comparator risk factors, and to find other risk factors not yet evaluated by multivariable analysis.18,29–31 Meta-analyses will be performed if ORs from at least two studies could be pooled.32 Each study will contribute only one OR for metagroup and if a study reports effect sizes on several follow-ups or with different type of adjustment, we will choose based on the following criteria in order of importance: 1) sample of at least 30 patients; 2) latest follow-up; 3) the most adjusted one. Descriptive data will be used to characterize our study population. The meta-analysis design is schematically displayed in Figure 1.\n\nMain calculations\n\nThe type of effect size measure is expected to vary between studies, as a result, harmonization will be achieved by converting all effect measures into a common metric, the OR. Effect size conversion will be performed using RevMan calculator and Psychometrica, an online tool33 The reported standard errors will be used to calculate the inverse variance weights. When not directly reported, standard error will be calculated using confidence interval or p-value, according to Altman and Bland.34 For studies reporting hazard ratios (HR), it would be incorrect to take them as an approximation of OR because the outcome (treatment failure) is not a rare event. Therefore, HRs will be converted to ORs using baseline control risk estimates from the individual studies and the methods as outlined by Symons et al.35 and Shor et al.36 If studies do not provide baseline risks, we will use the average risk, prevalence of the risk factor, and the relative effect to estimate the baseline risk, according to Kooter et al.37 Whenever possible, in the absence of a reported effect size, we will calculate the crude OR and its standard error for dichotomous variables based on the reported or reconstructed 2 × 2 contingency table. When zero counts occurs in a cell of a 2 × 2 contingency table, the Haldane-Anscombe correction will be applied.38 This is a method to avoid error in the calculation of odds ratio by adding 0.5 to all the cells of a 2 × 2 contingency table if any of the cell expectations would cause a division by zero. Continuous outcome measures such as mean difference and standardized mean difference, will be converted to OR using the Suissa’s method39,40 or the Hasselblad and Heidges’ method,40 respectively. Considering that continuous risk factors are sometime handled as unit (or range) of exposure and sometime are classified into distinct exposure categories, to maximize the number of studies included in each metagroup, all ORs for continuous risk factors will be converted into estimates of the OR per unit (or range) of exposure by using the Hamling’s method for linear trend calculation.41 The values (doses) assigned to each exposure category will be based on the best information available: in particular, assignments will be made according to a slightly modified parametric method proposed by Shim et al., if medians of the categories were available or computable42; otherwise, a nonparametric method will be applied, using the categories midpoints.43 The method used is as follows: for open smallest category, the median (or the midpoint) is set as the dose assuming that the beginning is zero (e.g., 5 if <10) or the lowest value in the study population, as appropriate; when the categories are closed by specific values, then the median (or the midpoint) is set as the dose; for open largest category, the median (or the midpoint) of the previous category minus the starting value of the previous category is added to the starting value of the last category, and the ensuing value is set as the dose. All calculations will be performed in duplicate and using Excel spreadsheets.\n\nStatistical methods\n\nRandom-effects models\n\nPower to detect heterogeneity is low in meta-analyses of observational studies.44 Furthermore, inherent between-study clinical and methodological heterogeneity is expected a priori due to the characteristics of the male slings literature: different study designs (e.g., retrospective vs prospective studies), different population characteristics (e.g., aetiology of PPI) and different outcome definitions (e.g., more or less stringent definition of success and cure). As a result, in our meta-analysis it is unlikely that studies will be functionally equivalent and effect size is assumed to vary from one study to the next. Therefore, we will take a conservative approach and use random-effects models to derive the summary exposure effects and form CIs, because random-effects models account for any observed heterogeneity regardless of whether the heterogeneity is statistically significant, and allows statistical inferences to be made to a population of studies beyond those included in the meta-analysis.45–47 The random-effects model weights the natural logarithm of each study's effect estimate by the inverse of its variance plus an estimate of the between-study variance (tau-squared) in the presence of between-study heterogeneity.47,48 Inverse-variance weighting is a method of aggregating two or more random variables that are weighted in inverse proportion to their variance in order to minimize the variance of the weighted average. The inverse variance is roughly proportional to sample size, but is a more nuanced measure, and serves to minimize the variance of the combined effect.49\n\nThe DerSimonian and Laird’s (DL) method of moments estimator, implemented in RevMan software, will be use to estimate the between-study variances.50 The pooled estimates will be then represented in forest plots. The p-values will be 2-sided and a statistically significant effect will be claimed at p-value <0.05.\n\nBecause the DL approach is suboptimal and may lead to too many statistically significant results (high type I error) when the number of studies is small and there is moderate or substantial heterogeneity,51 the approach described by Hartung and Knapp (HK method) will be applied in providing the final CIs around the pooled point estimates when less than 20 studies will be available.52,53 It has been showed that this approach consistently results in more adequate error rates (and CIs) and statistical tests, and consequently lower type I error than the DL method, more appropriately accounting for uncertainty in variance estimates, although the pooled effect estimates remain equal.52,54 For DL model, CIs and p-values are based on the normal distribution, whereas for the HK method, they are based on the t-distribution with the degrees of freedom equal to the number of trials minus one, and a weighted version of the DL standard error.52 This simple and robust modification to the common random-effects meta-analysis has been recommended in current guidelines19,55 to improve the summary results and was performed using the Excel spreadsheat provided by IntHout et al.52 Generally, results are more conservative with the HK method, giving wider CIs and larger p-values for the overall treatment effect, particularly in the scenario of less than 10 studies and moderate or substantial heterogeneity (I2 ≥ 30%).56\n\nHowever, the HK approach is not without limitations.56–58 When the heterogeneity is small and study sizes are imbalanced, the HK method may produce too wide, overly conservative CIs.56 Similarly, with very few studies (<5), especially if sample sizes are quite unequal, the HK method is often over-conservative, yielding spuriously wide and uninformative CIs, thereby leading to a loss of power and increasing the chance that relevant risk factors may be missed.51–53,58,59 Furthermore, in case of very few studies and homogeneous study results, CIs using the HK method may be also misleadingly narrower (anti-conservative) than that of the random-effects and fixed-effect method as well.58 As a result, we will use an adaptive, hybrid meta-analytic approach,58 by applying both the random-effects DL and HK methods and selecting the more conservative (widest CIs) analysis to provide the final summary estimates, thus rejecting artificially narrow CIs. Furthermore, the HK method will be discarded in favour of the DL method if yielding inconclusively wide and erratic CIs. DL and fixed-effect models will be also reported together with the HK method as sensitivity analysis (see below).51,58,59\n\nIn a random-effects meta-analysis it is also important to consider the potential effect of a treatment or an exposure within an individual study setting, as this may be different from the average effect.60,61 This can be achieved by calculating the prediction interval (PI) that tells us how much the effect size varies in the same units as the effect size and using absolute values.46 The term ‘prediction interval’ relates to the use of this interval to predict the possible underlying effect in a new study that is similar to the studies in the meta-analysis. A more useful interpretation of the PI is as a description of the range (dispersion) of observed effect sizes. PIs have proved a popular way of expressing the real amount of heterogeneity in a random-effects meta-analysis.61 Actually, PI is considered the most important outcome in a random-effects meta-analysis when heterogeneity is substantial, reflecting the uncertainty we expect in the summary effect if a new study is included.60,61 The PI can be also used to estimate in a future setting the probability that a treatment or an exposure will have a true-positive or true-negative effect, and to perform better power calculations.62\n\nPIs are, however, strongly based on the assumption of a normal distribution for the effects across studies, and can be very imprecise when based on only a few studies and if these studies are small, in which case they can appear spuriously wide or spuriously narrow.56,62 Accordingly, the use of PI is encouraged when the number of studies is reasonable, e.g. more than ten.17 Furthermore, PIs are inaccurate when there is low heterogeneity.56 As a result, in the present meta-analysis we plan to calculate the 95% PIs when at least 10 studies are available and I2 is at least 20%, using the Higgins-Thompson-Spiegelhalter equation and DerSimonian and Laird estimator of tau-squared and variance (with Hartung–Knapp variance estimator if Hartung–Knapp adjustment is applied).46\n\nAssessment of heterogeneity\n\nHeterogeneity between studies will be estimated statistically by the tau-squared, the χ2-based Cochran’s Q test and the I2 inconsistency Higgins’ statistic.63 Statistical significance is set at the 10% level (p-value <0.1) because of low power of the Q test. I2 statistic is the ratio of between-study variance over the sum of the within-study and between-study variances; it indicates the percentage of variance in observed effects that reflects variance in true effects rather than sampling error and is a recommended indicator of heterogeneity. I2 statistic ranges between 0% and 100% and, in agreement with the Cochrane handbook, 0 to 40% will be interpreted as “might not be important“, 30 to 60% as “may represent moderate heterogeneity”, 50 to 90% as “may represent substantial heterogeneity and 75 to 100% as “considerable heterogeneity”.16 Clinical heterogeneity is assumed to be mainly derived from the different types of treatment (e.g., different slings and implantation techniques) and different patients characteristics (e.g., age, incontinence aetiology and severity) among included studies, while methodological heterogeneity is caused by a variety of study designs (e.g., prospective and retrospective), and diverse follow-up lengths and sample sizes. Sub-group analysis and meta-regression will be used to address the cause of heterogeneity (see below).\n\nAnalysis of subgroups or subsets\n\nWhen at least 10 studies will be available on a specific risk factor, subgroup analyses and random-effects meta-regressions will be carried out, also in absence of substantial inconsistency, based on the (pre-specified) following study characteristics: sling type (Advance vs. non-Advance), baseline incontinence severity (pad use, using cut-offs of three pads per day), sample size (cut-off of 100 patients), follow-up length (cut-off of 36 months), study design (retrospective vs. prospective), outcome definition (more vs. less stringent), level of confounder adjustment (cut-off of 4 covariates) and RoB (high vs. low-moderate).19,64 Subgroup analyses performed in the secondary meta-analysis will be useful to assess the robustness of statistically significant differences between subgroups in the primary meta-analysis.\n\nMetabiases\n\nTo evaluate whether summary estimates are prone to small-study effects (e.g., arising from publication bias), funnel plots will be generated and Egger’s regression test will be used to evaluate the presence of asymmetry.65 Duval and Tweedie’s nonparametric trim-fill analysis will be used to determines how many studies would need to be included in the meta-analysis to make the funnel plot symmetrical, and calculate a meta-analytic effect size that adjust for publication bias.66 Briefly, this method initially removes the studies that are in the asymmetric part of the funnel plot, performing a new estimate of the pooled effect size. Then, using the new estimate as the axis of symmetry, studies previously removed are added again to the funnel plot, together with the same number of putative symmetric studies. A final pooled estimate is calculated based on the filled funnel plot.66 All these analyses will be performed only on meta-analyses featuring 10 or more studies.67\n\nSensitivity and influence analyses\n\nSeveral sensitivity analyses will be conducted to test the robustness of the main findings. According to published recommendations, when the HK method is applied, sensitivity analyses should always be performed using the fixed-effect and/or the random-effects models to assess that HK model based estimates of CIs are actually the most conservative51,68 Therefore, results obtained applying the HK model will be reported together with those estimated with the fixed-effect and the random-effects DL methods. Fixed-effect model result will be also reported as sensitivity analysis when only the DL method is applied, unless no heterogeneity (I2=0) was detected (in this case both methods provide the same summary estimate). As stated above, a sensitivity analysis using the fixed-effect model is particularly useful in case of very few studies where both conventional random-effects methods and other modified methods (such as the HK method) become inaccurate.48,51,58,59,62\n\nThe publication bias-adjusted pooled estimates will be also used as a sensitivity analysis to assess whether the crude quantitative syntheses is robust or not (i.e. the result is reversed) to publication bias.16\n\nA leave-one-out analysis69 will be performed to assess the robustness of the summary estimates by removing one study at a time from meta-analyses and recalculating the pooled effect estimates to evaluate whether the meta-analysis results were biased by any individual study. The results will be considered robust if no changes in direction and statistical significance of the effect size are observed on the exclusion of any studies, and the estimates in each case are well within the CIs of the overall estimate.\n\nA sensitivity analysis will be also performed excluding studies in the form of randomized controlled trial or published as abstract only.\n\nFurthermore, for the assessment of potential outliers and influential studies that may influence the robustness of the findings, multiple diagnostic measures will be applied when at least three studies will be available to ensure that the conclusions do not hinge on a few unusual studies.70 Outliers will be identified analysing the studentized deleted residuals (the deleted residual divided by its estimated standard deviation). Influential studies will be identified based on the following parameters: difference in fits value (DFFITS), which essentially indicates by how many standard deviations the predicted average effect for the ith study changes after excluding the ith study from the model fitting; Cook’s distance (Di), a value that summarizes how much all the fitted values change when the ith study is removed; COVRATIOi value (change in the variance–covariance matrix of the parameter estimates when excluding the ith study from the model fitting (a COVRATIOi value below 1, therefore, indicates that removal of the ith study actually yields more precise estimates of the model coefficients, or equivalently, that addition of the ith study actually reduces precision); QE statistics.70 If any outlier/influential studies will be identified, as further sensitivity analysis we will examine the effect of removing them from the meta-analysis.\n\nMissing data\n\nFor missing data, study investigators will be contacted for unreported key data or additional details via electronic mail and researchgate.net if possible. Missing mean values and/or standard deviations are a common feature of meta-analyses of continuous outcome data. Calculation of missing mean values and standard deviation will be based on Wan’s methods71 or Cochrane method,16 as appropriate. Data displayed in graphs will be extracted when not retrieved otherwise using the software WebPlotDigitizer (https://automeris.io/WebPlotDigitizer/). If necessary, imputation of the average value borrowed from one or more studies in the meta-analysis will be used for standard deviation.72\n\nSoftwares\n\nAll statistical analyses will be performed using the statistical softwares Review Manager 5.4, (RevMan) [Computer program]. Version 5.4. The Cochrane Collaboration, 2020), Meta-Essentials73 for metaregression and PI calculation, JASP (The JASP Team (2017). JASP (Version 0.8.1.0) [Computer software]) for publication bias and outliers/influence analyses, and Microsoft Excel for MAC (version 2016) for all other calculations.\n\nAssessment of confidence in cumulative evidence\n\nTo interpret the confidence in cumulative evidence, we will follow the Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) guidance regarding the determination of certainty in estimates of association between risk factors and outcomes.74 This approach has been found to work well in assessing individual risk factors.74 The quality of evidence on each risk factor and from each type of analysis (primary and secondary) will be assessed separately. A summary of finding table will be provided. Grade assessment will be conducted in duplicate with disagreements resolved through discussion and, if necessary, with the senior reviewer (ES) until mutual agreement is reached. The assessment of a body of longitudinal observational studies in the field of prognostic research begins as high certainty in the evidence.74,75 The GRADE approach considers five factors that can decrease the confidence in estimates of effects: (1) RoB, (2) inconsistency, (3) indirectness, (4) imprecision and (5) publication bias; and three factors that can increase our confidence in estimates of effects from observational studies: (1) large estimates of treatment, (2) a dose–response gradient and (3) plausible confounding that would increase confidence in an estimate.76 Certainty is ultimately designated as high, moderate, low, or very low. We will downgrade for RoB if ≥50% of the studies will be at high RoB or if the results are no longer significant or even reversed after removing high RoB studies. An OR higher than 2.5 will be considered as a large estimates for dichotomous or dichotomized risk factors.77,78\n\nThe result of the review will be presented at national and international conferences and will be reported in peer-reviewed medical journals following the PRISMA statement.15\n\nThe research team performed a pilot search in the relevant databases in Mars 2022. This search was used to select the keywords and delineate the final search strategy, start the pilot phase of the studies selection process, customize the QUIPS tool, and create the data extraction form. The first full search and study selection has been completed. Data extraction, risk of bias assessment and data analysis are ongoing. A second full search is scheduled to be conducted in January 2023. After completion of the review, a last search will be performed before the submission of the final report.\n\n\nDiscussion\n\nThis planned review and meta-analysis will systematically explore the evidence available on the association between potential risk factors for failure after treatment of PPI with synthetic fixed perineal slings implantation. By collecting and summarizing information about possible mediating and moderating factors that can clarify why a non-negligible proportion of patients fail to achieve the cure or social continence after sling placement, the results from this review will provide healthcare professionals treating and managing such patients with data useful in everyday practice for decision-making and patient counselling. By helping to stratify patients according to the risk of sling failure, this information may streamline the selection of candidates who are most likely to benefit from sling and assist clinicians in directing high risk patients towards other therapeutic options, or in setting more realistic patients’ expectations regarding slings results. The present review will also provide researchers with the necessary, evidence-based groundwork to perform high-quality prognostic studies and to identify areas for future investigations in the field. A more in-depth knowledge of prognosticators of sling efficacy is expected to add valuable information to the understanding about the mechanism of action of slings. Furthermore, our findings may potentially help to formulate improved practice guidelines and recommendations for the prevention of sling failures and to develop statistical models for predicting individual risk and prognostic endpoints.\n\nTo our knowledge, this is the first systematic review evaluating risk factors for male sling failure and to present them in an evidence-based framework. The strengths of the present review are 1) use of a comprehensive search strategy with broad inclusion criteria (e.g., limited restrictions based on language and study design, no restrictions based on date of publication, possibility of abstracts inclusion), in order to ensure a wide-reaching coverage of relevant research including as much studies as possible, thus limiting the publication bias, 2) the conservative approaches used, aiming to limit type I error (false positive results), 3) use of rigorous methodologies based on latest available guidelines on performing a systematic review and meta-analysis on prognostic studies, 4) separate pooling of adjusted and unadjusted effect measures, 5) assessment of the study quality and of the certainty of evidence following current standards for RoB assessment in prognostic research and the GRADE methodology, 6) use of several methods for influence analyses and extensive sensitivity analyses to assess the robustness of the results to changes in methods and model assumptions.\n\nHowever, we anticipate some methodological limitations in the conduct of this review. Being a review of observational studies, the clinical meaningfulness of summary result may be limited by real study diversity (heterogeneity), especially regarding type of sling, patients’ populations and methods of measurement of both exposures and outcomes. Studies are also expected to differ by design (e.g., retrospective and prospective) and by the number of covariates adjustment, which could potentially alter the magnitude of the associations. We will address this issue by subgroup analysis and meta-regression, when possible. Similarly, missing or imprecise data are frequent in such review, although we expect this issue to be solved by requesting authors to offer relevant data. The measurement methodologies and effect estimates from the individual studies will be assumed to be unbiased, although they may be biased due to flaws in the design or conduct of the studies. Several biases commonly affect observational studies reducing the significance of the review findings; biases of primary studies will be evaluated through the RoB assessment to provide a more insightful interpretation of the results.\n\nIn conclusion, the rationale and methodology of a systematic review and meta-analysis of risk factors for sling failure in patients suffering from PPI were described. Although there are some methodological limitations in conducting such kind of review, we believe they will not be serious enough to affect its value. The results of the review are expected to be a future guide for both clinicians and patients to choose the best treatment for this very bothersome condition.\n\nEthical approval is not required for this systematic review and meta-analysis because it will be based on secondary analysis of data already available in scientific databases and individual patient data will not be obtained or accessed. Even if authors of included studies will be asked to provide relevant missing data, any clinical information connecting with an individual patient will not be revealed.", "appendix": "Data availability\n\nNo data is associated with this article.\n\nZenodo: ‘Protocol for a systematic review and meta-analysis on preoperative risk factors for failure after fixed sling implantation for post-prostatectomy stress urinary incontinence. https://doi.org/10.5281/zenodo.7396318. 79\n\nThis project contains the following extended data:\n\n• QUIPS operationalized.xlsx\n\n• Data Collection Sheet.xlsx (the data collection spreadsheet used for data extraction for the review)\n\nZenodo: ‘PRISMA-P’ checklist for ‘Protocol for a systematic review and meta-analysis on preoperative risk factors for failure after fixed sling implantation for post-prostatectomy stress urinary incontinence’. https://doi.org/10.5281/zenodo.7396318. 79\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nAcknowledgements\n\nPastorino Roberta, Fondazione Policlinico Universitario Agostino Gemelli IRCCS, Università Cattolica del Sacro Cuore, Dept. of Salute della Donna e del Bambino e di Sanità Pubblica, Rome, Italy\n\n\nReferences\n\nBauer RM, Bastian PJ, Gozzi C, et al.: Postprostatectomy Incontinence: All About Diagnosis and Management. Eur. Urol. 2009; 55: 322–333. Publisher Full Text\n\nSacco E, Prayer-Galetti T, Pinto F, et al.: Urinary Incontinence after Radical Prostatectomy: Incidence by Definition, Risk Factors and Temporal Trend in a Large Series with a Long-Term Follow-Up. BJU Int. 2006; 97: 1234–1241. PubMed Abstract \| Publisher Full Text\n\nHamdy FC, Donovan JL, Lane JA, et al.: 10-Year Outcomes after Monitoring, Surgery, or Radiotherapy for Localized Prostate Cancer. N. Engl. J. Med. 2016; 375: 1415–1424. Publisher Full Text\n\nDonovan JL, Hamdy FC, Lane JA, et al.: Patient-Reported Outcomes after Monitoring, Surgery, or Radiotherapy for Prostate Cancer. N. Engl. J. Med. 2016; 375: 1425–1437. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVenderbos LDF, Deschamps A, Dowling J, et al.: Europa Uomo Patient Reported Outcome Study (EUPROMS): Descriptive Statistics of a Prostate Cancer Survey from Patients for Patients. Eur. Urol. Focus. 2021; 7: 987–994. 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PubMed Abstract \| Publisher Full Text\n\nGuyatt GH, Oxman AD, Schünemann HJ, et al.: GRADE Guidelines: A New Series of Articles in the Journal of Clinical Epidemiology. J. Clin. Epidemiol. 2011; 64: 380–382. PubMed Abstract \| Publisher Full Text\n\nHuguet A, Hayden JA, Stinson J, et al.: Judging the Quality of Evidence in Reviews of Prognostic Factor Research: Adapting the GRADE Framework. Syst. Rev. 2013; 2: 71. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLipsey MW, Wilson DB: Practical Meta-Analysis; Practical meta-analysis. Thousand Oaks, CA, US:Sage Publications, Inc;2001; ix, 247. 0-7619-2167-2.\n\nEMILIO SACCO: Protocol for a systematic review and meta-analysis on preoperative risk factors for failure after fixed sling implantation for post-prostatectomy stress urinary incontinence.2022. Publisher Full Text" }	[ { "id": "159580", "date": "20 Jan 2023", "name": "Frank M. J. Martens", "expertise": [ "Reviewer Expertise Functional and reconstructive urology including male stress urinary incontinence." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nProtocol description for a systematic review/meta-analysis of pre-operative potential risk factors for failure of synthetic fixed perineal slings in males with stress urinary incontinence.\nRelevance of meta-analysis: Predictors of success/failure are of great importance to counsel patients well about all options available for treatment of stress urinary incontinence.\nQuestion to the authors: One of the exclusion criteria is >20% inclusion of patients with non-post-prostatectomy incontinence. I was wondering why this exclusion criterium is chosen and why exactly >20% of patients within a study? As the cause of incontinence could be a potential risk factor of failure I would like to propose to include them all (so no exclusion criterium), or if you only want to focus on post-(radical?)-prostatectomy (because of malignancy) incontinence then leave studies/patients with other causes completely out of the meta-analysis.\n\nIs the rationale for, and objectives of, the study clearly described? Yes\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Not applicable", "responses": [ { "c_id": "10319", "date": "16 Nov 2023", "name": "EMILIO SACCO", "role": "Author Response", "response": "Dear Reviewer, On behalf of all authors, I would like to thank you for the constructive comments. We hope that our replies and accompanying modifications will be considered satisfactory. Sincerely, Emilio Sacco Corresponding author Reviewer 1 Question: One of the exclusion criteria is >20% inclusion of patients with non-post-prostatectomy incontinence. I was wondering why this exclusion criterion is chosen and why exactly >20% of patients within a study? As the cause of incontinence could be a potential risk factor of failure I would like to propose to include them all (so no exclusion criterium), or if you only want to focus on post-(radical?)-prostatectomy (because of malignancy) incontinence then leave studies/patients with other causes completely out of the meta-analysis. Reply: We thank the reviewer for this very appropriate comment. In a meta-analysis like this, a lot of clinical heterogeneity is expected and a major reason is the difference in patient population. This increases the risk of making erroneous inferences based on the results. To reduce this source of heterogeneity we decided to use a more stringent set of selection criteria to determine which studies to be included in the meta-analysis. The 20% cut-off was a convenient one. However, we agree with the reviewer that the etiology of incontinence (non-postprostatectomy vs. postprostatectomy) may be a potential prognostic factor as well. Consequently, we addressed this issue by removing this inclusion criterion, considering that the heterogeneity issue was to some extent accounted for by using the random-effect model with Hartung-Knapp adjustment and calculating prediction intervals. Removing this criterion, some advantages of a larger number of included studies should be also considered, such as increased statistical power, improved precision and validity of estimates, increased possibility of investigating the causes of heterogeneity and publication bias, and providing more general trends on the effects of risk factors on sling failure. Furthermore, this strategy is consistent with the recommendations proposed by the “Meta-Analysis of Observational Studies in Epidemiology Group” which advocated the use of broad inclusion criteria for studies along with regression analyses to relate specific study design characteristics to outcome [Stroup DF, et al. Meta-analysis of observational studies for epidemiology: A proposal for reporting. JAMA. 2000; 283:2008–2012]. This maximizes the amount of data available for review." } ] } ]	1	https://f1000research.com/articles/12-19
https://f1000research.com/articles/12-494/v1	15 May 23	{ "type": "Research Article", "title": "Accident severity prediction modeling for road safety using random forest algorithm: an analysis of Indian highways", "authors": [ "Humera Khanum", "Anshul Garg", "Mir Iqbal Faheem", "Mir Iqbal Faheem" ], "abstract": "Background: Road accidents claim around 1.35 million lives annually, with countries like India facing a significant impact. In 2019, India reported 449,002 road accidents, causing 151,113 deaths and 451,361 injuries. Accident severity modeling helps understand contributing factors and develop preventive strategies. AI models, such as random forest, offer adaptability and higher predictive accuracy compared to traditional statistical models. This study aims to develop a predictive model for traffic accident severity on Indian highways using the random forest algorithm. Methods: A multi-step methodology was employed, involving data collection and preparation, feature selection, training a random forest model, tuning parameters, and evaluating the model using accuracy and F1 score. Data sources included MoRTH and NHAI. Results: The classification model had hyperparameters ’max depth’: 10, ’max features’: ’sqrt’, and ’n estimators’: 100. The model achieved an overall accuracy of 67% and a weighted average F1-score of 0.64 on the training set, with a macro average F1-score of 0.53. Using grid search, a random forest Classifier was fitted with optimal parameters, resulting in 41.47% accuracy on test data. Conclusions: The random forest classifier model predicted traffic accident severity with 67% accuracy on the training set and 41.47% on the test set, suggesting possible bias or imbalance in the dataset. No clear patterns were found between the day of the week and accident occurrence or severity. Performance can be improved by addressing dataset imbalance and refining model hyperparameters. The model often underestimated accident severity, highlighting the influence of external factors. Adopting a sophisticated data recording system in line with MoRTH and IRC guidelines and integrating machine learning techniques can enhance road safety modeling, decision-making, and accident prevention efforts.", "keywords": [ "Traffic Accidents", "Accident Severity", "Road Safety", "Accident Prediction Modeling", "Random Forest" ], "content": "Introduction\n\nRoad accidents are a significant public health concern worldwide, with an estimated 1.35 million deaths caused by road traffic accidents each year.1 Developing countries, such as India, are disproportionately affected, with over 150,000 fatalities reported annually.1 Road safety is a major concern in India, with a large number of accidents and fatalities reported each year. According to the Ministry of Road Transport and Highways, there were 449,002 road accidents in India in 2019, resulting in 151,113 deaths and 451,361 injuries.2 To understand the factors that contribute to accidents and to develop strategies to prevent them, accident severity modelling is a statistical technique used in the field of road safety.\n\nThe modelling process involves analyzing data on past accidents and identifying the factors that contributed to their occurrence and severity. These factors can include road conditions, weather, driver behaviour, and vehicle type, among others. The goal of accident severity modelling is to identify the factors that are most important in contributing to accidents and to develop evidence-based strategies to improve road safety and reduce accidents’ number and severity.3–6\n\nStatistical models, such as logit models and probit models,7 have been widely used for predicting traffic accidents’ severity since the early 1990s. However, if assumptions imposed on these models are violated, results may be inaccurate. Artificial intelligence models, on the other hand, do not make any assumptions and are more adaptable. They are capable of handling intricate nonlinear relationships and generally offer higher predictive accuracy than statistical approaches. random forest (RF) is a powerful and versatile algorithm for accident severity prediction, with several advantages over other machine learning algorithms. It has been successfully applied in various contexts, and its performance can be further improved by tuning the key parameters and carefully pre-processing the data.8–11\n\nThe performance of the random forest algorithm is significantly influenced by the selection of hyperparameters.12\n\nTo optimize its performance, identifying the optimal parameter values is crucial. Previous research has predominantly utilized grid search to explore values within the parameter space. However, alternative approaches may be necessary to overcome the computational challenge of grid search in high-dimensional parameter spaces.13 Random search and Bayesian optimization are effective alternatives to grid search for hyperparameter optimization in random forest, particularly in high-dimensional parameter spaces.13\n\nAccurate prediction of traffic accident severity is essential for improving emergency response, reducing fatalities, and minimizing injuries. To achieve this goal, accurate data, appropriate machine learning algorithms, and regular updates to the predictive models are necessary. Given the advantages of random forest models in terms of prediction accuracy and interpretability, they can be used as the primary predictive model for traffic accident severity on Indian highways. Several factors contribute to accidents on Indian highways, including poorly designed or maintained roads, speeding, and roadside hazards. Identifying these factors through accident severity modelling can help develop evidence-based strategies to improve road safety and reduce accidents’ number and severity.14–16\n\nIndian highways have a high incidence of accidents, and several contributing factors have been identified. These include road design and geometry, speed, roadside hazards, and driver behavior. Poorly designed or maintained roads, such as those with narrow or winding stretches, lack of markings, and poor road surfaces, increase the likelihood of accidents.17 Speeding is a major factor in many accidents on Indian highways, which may result from a lack of enforcement, cultural norms, and driver attitudes.18 Roadside hazards, such as trees, poles, and animals, are prevalent on Indian highways and increase the risk of collision.19 Driver behavior, including drunk, distracted, and reckless driving, is also a significant contributor to accidents on Indian highways.15 Addressing these factors is crucial to improving road safety and reducing accidents on Indian highways.\n\nThe main objective of our study is to develop a predictive model for the severity of traffic accidents on Indian highways. To achieve this goal, we have chosen random forest models due to their ability to provide accurate predictions and interpretability.\n\nThe findings of our study will be used to develop a predictive model for accident severity that can inform road safety policies and interventions. This model can be used to identify high-risk areas and to prioritize resources for accident prevention and mitigation.\n\nThe study areas selected are the National Highways two stretches as mentioned below\n\n1. Pune-Sholapur Section of NH-9 in km.144/400 to Km. 249/000 in the State of Maharashtra (Figure 1).\n\n2. Six-Laning of Barwa-Adda-Panagarh Section of NH-2 from km 398.240 to km 521.120 including Panagarh Bypass in the States of Jharkhand and West Bengal (Figure 2).\n\nThe study areas for this research project were selected based on specific criteria. Firstly, the researchers had prior experience of working on one of the stretches, which is the Pune-Sholapur Section of NH-9 in km.144/400 to Km. 249/000 in the State of Maharashtra. This experience could have provided insights and knowledge that could be useful in conducting the study.\n\nAdditionally, data was also provided by the same concessionaire as of the previous stretch on request for another stretch, which is the Six-Laning of Barwa-Adda-Panagarh Section of NH-2 from km 398.240 to km 521.120 including Panagarh Bypass in the States of and West Bengal. This data could have been relevant to the research objectives and could have assisted in achieving the desired outcomes.\n\n\nMethods\n\nThe proposed methodology for this research involves the following steps for implementing a random forest model machine learning technique for the accident severity prediction.\n\nData Preparation: The first step in implementing a random forest model for accident severity prediction is to collect and prepare data. Raw data of road accidents for the selected stretches of the highway can be obtained from secondary sources such as the Ministry of Road Transport and Highways (MoRTH) and National Highways Authority of India (NHAI).2 Data wrangling and mining techniques can be used to clean and preprocess the data.\n\nFeature Selection: Once the data is prepared, selecting appropriate features for the model becomes crucial. Feature selection plays a vital role in reducing the dimensionality of the data and enhancing the model’s accuracy. There are several techniques available for feature selection, such as statistical tests, correlation analysis, and principal component analysis (PCA).20\n\nModel Training: In the next step, a random forest model can be trained on the preprocessed data. The model can be developed using a machine learning based framework, as described in Breiman’s work on random forest.21 The RF algorithm involves bagging and random feature selection techniques to create multiple decision trees that are aggregated to form a stronger learner.22\n\nParameter Tuning: To improve the performance of a random forest model, it is important to fine-tune its parameters. The three key parameters that significantly impact the tuning performance of the random forest model are the total number of trees (n_estimators), the number of features used for each node segmentation (max_feature), and the maximum depth of a tree (max_depth).23\n\nModel Evaluation: After training the random forest model and optimizing its parameters, it is important to evaluate the model’s performance. Various evaluation metrics can be used, including accuracy, precision, recall, F1 score, and Area Under the Curve - Receiver Operating Characteristis (AUC-ROC) curve.24\n\nModel Implementation: Once the model has been trained and evaluated, it can be deployed for accident severity prediction. The methodology can be designed using python for building the model and forecasting the severity of road traffic accidents on Indian highways.\n\nData on road accidents from selected stretches of highways was obtained from the Concessionaires of the National Highways Authority of India (NHAI) for two projects: Pune-Solapur and Bengal (BAEL) Section. For the Pune-Solapur Section of NH-9, which is located between km. 144/400 and km. 249/000 in the state of Maharashtra, accident dates from 2013 to 2018 were used. For the Six-Laning of Barwa-Adda-Panagarh Section of NH-2, which includes Panagarh Bypass and is located in the States of Jharkhand and West Bengal Stretch, accident dates from 2015 to 2019 were used for the stretch between km 398.240 and km 521.120. The raw data was subject to exploratory data analysis, as detailed in the following section.\n\nIn this stage, data gathering and exploration is performed using secondary source data. The dataset consists of 3257 observations out of which the 1855 observations are of Bengal (BAEL) Section and 1402 observations are of Pune- Solapur and 32 variables, including the target variable “accident severity.” The 32 attributes and their corresponding mappings are presented in Table 1.\n\nThe random forest classification algorithm has been employed in this study to forecast the severity of road traffic accidents in India. This section details the procedure for implementing the model, performance evaluation, and discuss the results obtained. The random forest algorithm is written using python programming language.\n\nThe target variable for the random forest model is selected as the’Accident Severity’ which has classes as Fatal, Grevious Injury, Minor Injury and No Injury and indexed as [1-Fatal, 2-Grevious Injury, 3-Minor Injury, 4-No Injury.\n\nThe dataset is partitioned into training and testing sets with a ratio of 80% and 20%, respectively. The hyperparameters’n_estimators’ and’max_depth’ are specified, and a grid search is conducted with cross-validation (cv=5) to identify the optimal hyperparameters. The best parameters and scores are obtained. The best estimator is fit on the training data. Predictions are made on the test data and the accuracy of the model is obtained.\n\nThe algorithm and programme for Accident Severity Modelling using random forest are written in the Python programming language, and the code is made available to the public for further development. The source code can be accessed via the software availability statement.\n\nAccuracy analysis on test data: Three metrics were employed to evaluate the effectiveness of the algorithms: accuracy, precision, and recall. These metrics are defined as follows:\n\nAccuracy: The formula for a metric that measures the proportion of correctly predicted observations to the total number of observations is represented as:\n\nPrecision is a metric that indicates the ratio of correctly predicted positive observations to the total number of predicted positive observations, and is calculated using the formula:\n\nRecall is a metric that reflects the ratio of correctly predicted positive observations to the total number of actual positive observations, and is determined using the formula:\n\n\nResult and Discussion\n\nThe classification model used three hyperparameters -’max_depth’: 10,’max_features’:’sqrt’, and’n_estimators’: 100, and the results generated a confusion matrix for the training set. The matrix indicated the number of correctly and incorrectly classified instances for each class. The classification report provided precision, recall, and f1-score for each class, along with support. The model showed high precision and recall for class 1 but low precision and recall for classes 2, 3, and 4, with an overall accuracy of 67% and a weighted average f1-score of 0.64 on the training set. The macro average f1-score, which assigns equal weight to each class, was 0.53.\n\nThe optimal parameters for a random forest classifier model were determined through a grid search, with a max depth of 2, n estimators of 5000, and a random state of 0. The model was then applied to the test data, and the predictions were saved in an Excel file called “predicted output3.xls” for further analysis. The accuracy of the model on the test data was determined to be 0.4147, or approximately 41.47%, indicating that it accurately predicted the severity of traffic accidents in about 41.47% of test cases.\n\nPredicted outputs\n\nComparative analysis of observed and predicted accident severity index against dates\n\nThe actual accident severity indices are represented by the observed values, while the predicted values are generated by the random forest model using the input features.\n\nThe following is a summary (Figure 3) of the comparison between the observed and predicted values:\n\nOn dates such as 25-02-2017, 17-04-2017, and 22-04-2017, the random forest model accurately predicts the accident severity index.\n\nIn a number of instances, the model predicts a lower accident severity index value than the observed value. 18-02-2017, 23-02-2017, and 27-03-2017, for example.\n\nOccasionally, the model overestimates the accident severity index by predicting a higher value than the observed value, as on 24-05-2017 and 20-10-2017.\n\nIn general, the model frequently predicts a severity index of 2 for accidents, even when the observed values are distinct. This may indicate a bias in the model, possibly as a result of an imbalance in the training dataset, in which severity index 2 occurs more frequently than other categories.\n\nComparative analysis of observed and predicted accident severity index against time\n\nFigure 4 displays the date, day of the week, and time of the accident, as well as the observed and predicted accident severity indices. The plotted for the 165 rows of predicted data doesn’t fit in A4 sheet hence the data is published and the link is provided in the Tableau graphs visuals availbility [i].\n\nThe dataset contains accident data from February 18, 2017 to December 31, 2017, as determined by Tableau analysis of the plot generated from the provided Excel table.\n\nThe observed accident severity index ranges from 1 to 4, where 1 corresponds to the least severe accident and 4 to the most severe accident.\n\nThe observed severity index for the vast majority of accidents in the dataset is 3, followed by 4. 2 indicates a less severe accident, while 4 indicates a more severe accident.\n\nThe majority of accidents within the dataset have a predicted severity index of 2, followed by an index of 1.\n\nThe analysis of the scatter plot reveals that the predicted severity index is typically lower than the observed severity index. This suggests that the model used to predict the severity of accidents is not always accurate and could be improved.\n\nThe Tableau plot (Figure 5) presents a detailed visual analysis of accident data on the right-hand side of the road. The data is organized by date and day of the week, displaying the accident location, observed accident severity index, and predicted accident severity index for each incident. The plot effectively illustrates the spatial distribution of accidents and their severity over time, enabling the identification of patterns and trends. The Tableau plot doesn’t fit in A4 sheet hence the data is published and the link is provided in the Tableau graphs visuals availability [ii].\n\nIt is evident from the analysis that the majority of accidents have an observed severity index of 2 or 3, indicating a moderate severity. However, the predicted accident severity index largely remains at 2, indicating that the predictions may be somewhat conservative and do not fully capture the observed severity range.\n\nIn addition, there appears to be no correlation between the day of the week and the frequency or severity of accidents across the different days of the week. This may suggest that external factors, such as traffic patterns or weather conditions, have a greater impact on the occurrence and severity of accidents than the day of the week.\n\nThe graph displays (Figure 6) the date, day of the week, and accident location on Left Hand Side (LHS) of the road, as well as the observed and predicted accident severity indices. The plotted of predicted data doesn’t fit in A4 sheet hence the data is published and the link is provided in the Tableau graphs visuals availability [iii].\n\nThe scatterplot reveals that the majority of accidents on the left side of the road had a severity index of 2 or 3, with only a few instances of severity index 1 and 4. This indicates that the majority of collisions on the left side of the road were of moderate severity.\n\nIn the majority of cases, the predicted accident severity index was 2, with only a few instances of values 3 and 4. This suggests that the predictive model may be biased towards predicting less severe accidents.\n\nThere was no discernible pattern or trend between the day of the week and the occurrence of accidents. Accidents appeared to occur every day, indicating that the day of the week may not be a significant predictor of accident severity on the left side of the road.\n\nThe accident locations, as measured by Accident Location A Chainage km, were scattered along the roadway at various distances. This suggests that there may not be a particular accident hotspot or concentration on the left-hand side of the road.\n\nThe recording of road accident data in India must comply with the MoRTH & IRC guidelines, utilizing the Road Accident Recording and Reporting Formats. Despite this, there exists a need for a more advanced data recording system to effectively model road safety. The digital monitoring of road accidents can increase the frequency of data collection and minimize the absence of crucial information. Often, the lack of a system or individual to document the accident leads to the absence of important road accident data. This missing data can be regained through the use of machine learning, thus enhancing the accuracy of road safety modeling.\n\n\nConclusion\n\nThe random forest classifier model predicted the severity of traffic accidents with an overall accuracy of 67% on the training set and approximately 41.47% on the test set. Indicating possible bias or imbalance in the training dataset, the model tended to predict a lower severity index than the observed values. There were no discernible relationships between the day of the week and the occurrence or severity of accidents. The performance of the model can be enhanced by correcting the dataset imbalance and refining the model’s hyperparameters.\n\nThe observed and predicted accident severity indices were compared against a number of variables, including dates, times, and locations on both sides of the road. In some instances, the model accurately predicted the accident severity index, but it frequently underestimated accident severity. No discernible patterns or trends were observed in terms of accident location, indicating that external factors may have a greater influence on the occurrence and severity of accidents.\n\nTo improve road safety modelling, it is essential to adopt a more sophisticated data recording system consistent with MoRTH and IRC recommendations. Digital monitoring of road accidents can increase the frequency of data collection and reduce the loss of vital information. Integrating machine learning techniques can contribute to more effective interventions and decision-making in the field of traffic accident prevention and mitigation.\n\n\nFuture Scope\n\nThe study presented provides a good starting point for future research in the field of road safety modeling and accident prevention for Indian highways. However, with the limitations of the present study there opens potential areas for future research as mentioned below which will be taken up in continuation.\n\nDataset improvement: The study identified the possibility of dataset bias and imbalance affecting model performance. Future research will focus on improving the quality and quantity of data, reducing bias and improving model performance. This will involve exploring alternative data sources, enhancing data collection methods, and addressing data quality issues.\n\nModel improvement: The study used the random forest algorithm to develop a predictive model for traffic accident severity. In future research, other machine learning algorithms or ensemble models to improve model performance will be explored. Additionally, refining hyperparameters and addressing dataset imbalance will be done to improve model accuracy.\n\nExternal factors analysis: The study highlighted the influence of external factors on accident severity prediction. Future research can focus on exploring the impact of external factors such as weather conditions, road infrastructure, and driver behavior on accident severity. This can enhance the accuracy of predictive models and inform decision-making in accident prevention efforts.\n\nReal-time monitoring: The study highlighted the need for a sophisticated data recording system in line with MoRTH and IRC guidelines. Future research can focus on developing a real-time monitoring system that can capture road safety data in real-time and provide insights for accident prevention efforts.", "appendix": "Data availability\n\nZenodo. Data for Accident Severity Prediction Modelling for Indian Highways Case Study, https://doi.org/10.5281/zenodo.7773156. 25\n\nThis project contains the following underlying data:\n\n• Accdataset_hk_PS_BAEL_Combined.csv (The dataset consists of 3257 observations out of which the 1855 observations are of Bengal (BAEL) Section and 1402 observations are of Pune- Solapur.)\n\n• predicted_output_1.xlsx (This is level-2 processed data derived from raw accident data using prediction modeling. The data has been indexed from 1 to 4 for further analysis, and there are a total of 165 rows in the predicted output observations.\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgement\n\nWe are grateful to National Highways Authority of India and IL&FS Engineering and Construction Company for making the raw accident data available.\n\n\nReferences\n\nWorld Health Organization: Global Status Report on Road Safety 2018.Reference Source\n\nMinistry of Road Transport & Highways: Road Accidents in India 2019.Reference Source\n\nMannering F, Bhat C: Statistical Methods for Analyzing Highway Accident Data. John Wiley & Sons; 2014.\n\nBarbosa P, Andrade M, Ferreira S: Machine learning applied to road safety modeling: A systematic literature review. Journal of Traffic and Transportation Engineering (English Edition). 2020; 7(6): 775–790. Publisher Full Text\n\nAl-Mistarehi BW, Alomari AH, Imam R, et al.: Using Machine Learning Models to Forecast Severity Level of Traffic Crashes by R Studio and ArcGIS. Front. Built Environ. 2022; 8(April): 1–14. Publisher Full Text\n\nMoghaddam FR, Afandizadeh S, Ziyadi M: Prediction of accident severity using artificial neural networks. Int. J. Civ. Eng. 2011; 9(1): 41–48.\n\nMohamed AA, Mannering FL: Probabilistic models of traffic accident occurrence and severity. Handbook of traffic psychology. Elsevier; 2010; (pp. 129–143). Publisher Full Text\n\nHoang Long V, Ahmed K, Ma W: A Random Forest Approach to Predicting Traffic Accident Severity. IEEE Access. 2021; 9: 1219–1232. Publisher Full Text\n\nSze NN, Wong SC: Diagnostic analysis of the logistic model for pedestrian injury severity in traffic crashes. Accid. Anal. Prev. 2007; 39(6): 1267–1278. PubMed Abstract \| Publisher Full Text\n\nAbdel-Aty M: Analysis of driver injury severity levels at multiple locations using ordered probit models. J. Saf. Res. 2003; 34: 597–603. PubMed Abstract \| Publisher Full Text\n\nShiran G, Imaninasab R, Khayamim R: Crash Severity Analysis of Highways Based on Multinomial Logistic Regression Model, Decision Tree Techniques, and Artificial Neural Network: A Modeling Comparison. Sustainability. 2021; 13(10): 5670. Publisher Full Text\n\nYan L, Liao W: Evolutionary hyperparameter optimization for random forest. J. Ambient. Intell. Humaniz. Comput. 2019; 10(7): 2801–2810. Publisher Full Text\n\nSnoek J, Larochelle H, Adams RP: Practical Bayesian optimization of machine learning algorithms. Adv. Neural Inf. Proces. Syst. 2012;2951–2959.\n\nSingh G, Kumar A: Random forest-based prediction model for traffic accident severity on Indian highways. Journal of Traffic and Transportation Engineering (English Edition). 2021; 8(6): 693–706. Publisher Full Text\n\nPatel M, Patel R: A study on causes of road accidents in India. Int. J. Eng. Res. Appl. 2013; 3(6): 1386–1391.\n\nYan M, Shen Y: Traffic Accident Severity Prediction Based on Random Forest. Sustainability (Switzerland). 2022; 14(3): 2. Publisher Full Text\n\nIndian Road Congress: Guidelines for the Design of At-Grade Intersections on Rural Highways. New Delhi: 2012.\n\nRamanujam V, Bhalla K: Speeding on Indian roads: A survey of Indian drivers. Accid. Anal. Prev. 2009; 41(3): 527–532. Publisher Full Text\n\nJoshi S, Shaikh T: Animal-related crashes on national highways in India. Traffic Inj. Prev. 2017; 18(2): 120–124. Publisher Full Text\n\nAdele Cutler DRC, Stevens JR: Random Forests.2012. Publisher Full Text\n\nBreiman L: Random forests. Mach. Learn. 2001; 45(1): 5–32. Publisher Full Text\n\nLiaw A, Wiener M: Classification and regression by randomForest. R News. 2002; 2(3): 18–22.\n\nEl-Basyouny K, Sayed T, Abdel-Aty M: Predicting accident occurrence and severity on arterials using random parameter and random effect models. Accid. Anal. Prev. 2010; 42(3): 718–727.\n\nSokolova M, Lapalme G: A systematic analysis of performance measures for classification tasks. Inf. Process. Manag. 2009; 45(4): 427–437. Publisher Full Text\n\nKhanum H, Garg A, Faheem MI: Data for Accident Severity Prediction Modelling for Indian Highways Case Study (Accidentdata_V1). [Data set]. Zenodo. 2023. Publisher Full Text" }	[ { "id": "202786", "date": "18 Sep 2023", "name": "Hamsa Zubaidi", "expertise": [ "Reviewer Expertise Tansportation Safety" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe author mentioned \"accident\" in the paper, \"Accident\" is a broader term that implies an unintentional or unforeseen incident. It can encompass various events, including crashes, collisions, or other incidents resulting from human error, mechanical failure, or environmental factors. \"Crash\" is a more specific term that typically refers to a collision or impact between vehicles, objects, or individuals. It often implies a sudden and forceful event.\nUltimately, the choice between \"accident\" and \"crash\" should be based on the specific details and nature of the event being described in the paper, and relying on that \"accident\" should be replaced by \"crash\".\n\nThe author should add a random forest formula in the methodology.\n\nIt might be better if there is a Gini impurity test in the paper to understand the importance of explanatory variables.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10315", "date": "29 Nov 2023", "name": "Humera Khanum", "role": "Author Response", "response": "Comment:1 The author mentioned \"accident\" in the paper, “Accident\" is a broader term that implies an unintentional or unforeseen incident. It can encompass various events, including crashes, collisions, or other incidents resulting from human error, mechanical failure, or environmental factors. \"Crash\" is a more specific term that typically refers to a collision or impact between vehicles, objects, or individuals. It often implies a sudden and forceful event. Ultimately, the choice between \"accident\" and \"crash\" should be based on the specific details and nature of the event being described in the paper, and relying on that \"accident\" should be replaced by \"crash\". RESPONSE: Thank you for your insightful comments and for bringing up the concern regarding the usage of the term \"accident\" in our paper. Your observation about the broad implication of \"accident\" and the specificity of \"crash\" is indeed valid. I would like to clarify that in the context of our paper, the mention of \"accidents\" is based on Indian data, which is commonly referred to as road traffic accident data in official and legal documents in India. The term \"accident\" here encompasses various events, including crashes, collisions, or other incidents resulting from human error, mechanical failure, or environmental factors, as per the definition used in the Indian context. All the parameters mentioned in the paper have been taken into consideration, and the terminology used is consistent with the data source and regional context. We appreciate the suggestion to use the term \"crash\" for more specificity and will certainly consider this in our future research and writings, ensuring that the terminology aligns with the global standards and is clear to all readers. Comment 2: The author should add a random forest formula in the methodology. RESPONSE: The random forest formula is added to the methods section as below: Random Forest Algorithm Formulation: The Random Forest algorithm can be represented as: [RF(X) = (1/B) * Σ(T_b(X)) from b=1 to B] where X are the input features, B is the number of trees, and T_b(X) is the prediction of the b-th individual decision tree. Comment 3: It might be better if there is a Gini impurity test in the paper to understand the importance of explanatory variables. RESPONSE: The random forest formula is added to the methods section as below: In the construction of the Random Forest model for predicting accident severity, Gini impurity is employed as a criterion to evaluate the significance of different explanatory variables. Gini impurity, a measure utilized within the framework of decision trees (the base learners in a Random Forest), is crucial for the optimal selection of features at each node split. It offers a quantitative metric to discern the effectiveness of a variable in segregating the target classes. Mechanism of Gini Impurity: In the context of binary classification, the Gini impurity for a node is calculated as: IG(p)=k=1np2k where, pk is the proportion of samples classified to class k at that node, and the summation operates over all classes. A lower Gini impurity score suggests a higher purity of the node, indicating an enhanced classification. Gini Importance in Random Forest: In the developed Random Forest model, the Gini impurity plays a dual role: Node Splitting: It aids in the identification of the most significant variable at each node by evaluating the potential reduction in impurity for each split, and Feature Importance: Post model training, the average decrease in impurity caused by each feature across all trees is computed, known as Gini importance. This metric offers insights into the relative significance of different features for the prediction task." } ] }, { "id": "173654", "date": "05 Oct 2023", "name": "Sanjeev Sinha", "expertise": [ "Reviewer Expertise Traffic Engineering", "Transportation Planning", "Highway Materials", "Logistics" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe work is clearly and accurately presented however, the motivation behind the work and its academic contribution needs to be highlighted.\n\nThough relevant literature has been cited, it has not been described for its application in the present study. Further, there is some literature which seems not to be correctly referred and cited. As example the references, 7, 14 and 15 needs to be re-checked.\n\nThe title of the work is development of prediction modeling for accident severity, the choice of factors, their contribution and transferability of the model developed needs to be highlighted.\nThe literature review shows several works related to the topic thus novelty of the work also need to be highlighted.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10380", "date": "30 Nov 2023", "name": "Humera Khanum", "role": "Author Response", "response": "Comment 1: The work is clearly and accurately presented however, the motivation behind the work and its academic contribution needs to be highlighted. Author's Response: Thank you for your constructive feedback. We agree that highlighting the motivation and academic contribution of this work is crucial for a well-rounded understanding of the study. In the revised manuscript, we have included a section in the Introduction where we elaborate on the motivation behind this work, particularly focusing on the rising number of road accidents and the necessity to develop predictive models to understand and mitigate accident severity. Additionally, we have outlined the academic contribution of our study in the Discussion section, emphasizing the novelty and practical implications of our findings. Comment 2: Though relevant literature has been cited, it has not been described for its application in the present study. Further, there is some literature which seems not to be correctly referred and cited. As example the references, 7, 14 and 15 needs to be re-checked. Author's Response: We appreciate your attention to detail. We have revisited the cited literature, especially references 7, 14, and 15, and corrected the citations as suggested. Moreover, we have expanded the section to better explain the relevance and application of the cited works in the context of our study. Comment 3: The title of the work is Development of prediction modeling for accident severity, the choice of factors, their contribution and transferability of the model developed needs to be highlighted. Author's Response: Thank you for your suggestion. In the revised manuscript, we have dedicated a subsection within the Methodology section to discuss the choice of factors, their contribution, and the transferability of the developed model. Comment 4:The literature review shows several works related to the topic thus novelty of the work also need to be highlighted. Author's Response: We value your input on highlighting the novelty of our work. In the revised manuscript, we have included a paragraph at the end in the conclusion section where we delineate the novelty of our study in comparison to existing literature. We have emphasized the innovative aspects of our predictive model and how our work contributes uniquely to the understanding and prediction of accident severity." } ] } ]	1	https://f1000research.com/articles/12-494
https://f1000research.com/articles/10-1033/v1	11 Oct 21	{ "type": "Research Article", "title": "Perceived risk and condomless sex practice with commercial and non-commercial sexual partners of male migrant sex workers in London, UK", "authors": [ "Elisa Ruiz-Burga" ], "abstract": "Background: Since the emergence of HIV and the AIDS pandemic, the majority of risk-reduction interventions have been centred on the use of condoms in sex workers. Methods: This qualitative study recruited 25 male migrant sex workers in London to understand their risk perception and condomless sex experiences within the context of sex work and private life. The data was collected using face-to-face interviews, analysed using thematic analysis, and the findings interpreted through the theory of planned behaviour. Results: The themes explain that condomless sex with clients occurred when participants consciously accepted to perform this service deploying a risk assessment of clients, faulty strategies, and sexual practices to reduce their risk; or when they lost control because of recreational drugs, feeling attraction to clients, in precarious circumstances, or were victims of violence. Conversely, condomless sex with non-commercial partners occurred according to the type of relationship, with formal partners it was rationalised through emotional aspects attached to this kind of relationship, while with casual partners it was connected to sexual arousal and the use of alcohol and drugs. Conclusions: Reinforce educational interventions to deliver STI-HIV information, enhance the use of condoms, and to address specific contextual factors that facilitate condomless practice with commercial and non-commercial sexual partners.", "keywords": [ "HIV", "STI", "male migrants", "sex work", "condomless sex" ], "content": "Introduction\n\nSince the emergence of HIV and the AIDS pandemic, sex workers were considered a highly vulnerable group1 because of their high-risk exposure to acquire these infections compared to adult non-sex workers.2 Across the globe, approximately 8% of the newly HIV cases are reported among sex workers.3 Of serious concern are male sex workers (MSW) who are at more HIV risk than female sex workers (FSW). Receptive anal intercourse and insertive anal intercourse are considerably at higher risk of HIV transmission than vaginal sex.4 Further, MSW are greatly affected by other sexually transmitted infections (STIs).5–7 In response to AIDS and the concurrent increase in HIV prevention research, numerous interventions that aimed at decreasing the risk of infection have been conducted to reduce the practice of condomless sex.8–11 After decades, there is still a debate about the success of these interventions - while some authors claim that new infections are yet associated to an inconsistent use of condoms,12,13 others argue that MSW are using them more regularly, for either insertive anal sex or receptive anal sex.14–16\n\nA large number of sex workers in Europe are migrants who are living and working in disadvantaged circumstances, facing isolation and social exclusion.17–19 Moreover, migrant sex workers are extremely exposed to HIV and STIs due to their overlapping risks3,6,20 and structural inequalities that can create difficulties to use health services for HIV prevention, testing and treatment17,21,22 in some European countries. For example, it has been reported that male street sex workers, in particular illegal migrants in Germany, have lack of access to health care services due to their socio-legal position.23 This is a significant aspect as non-European sex workers who are highly mobile in Europe19 and are under different migration status,24 can be impacted by the legislation and internal policies of each country that determines their access to the health care services.17 In the United Kingdom, an important proportion of the sex work population is represented by male migrant sex workers (MMSW) who mostly work indoors in London.25–27 Reports show that they are mainly from Europe, Latin America and the Caribbean countries.26,28 Epidemiological and qualitative research have demonstrated that these migrants utilize national health care services (NHS), including sexual health clinics. In this manner, they can be tested for HIV and other STIs, receive counselling, adequate information, and a provision of condoms and lubricants.24,29 However, a study using national data demonstrated that although the use of sexual health clinics does not vary between British MSW and MMSW, the latter group seem to be more exposed to HIV and chlamydia infections.29\n\nThis paper explores the risk perception and condomless sex experiences of MMSW with commercial and non-commercial sexual partners, as discrepancies have been reported in the use of condom according to the type of sexual partner,14,16,30–32 and the sexual role performed during the act.27 In this manner, this paper aims to contribute to lessons learnt and recommendations for future educational interventions for this highly vulnerable group. This paper is pertinent in an era when the role of behavioral interventions is evolving with the advent of efficient alternatives of prevention such as pre-exposure prophylaxis (PrEP) and post-exposure prophylaxis (PEP). Despite of the efficacy demonstrated by the PrEP scheme, it requires high adherence to the treatment, strict laboratory monitoring,33 and more importantly, the use of supplementary methods of prevention such as the use of condoms to further reduce the HIV risk, and specifically for protection against other STI; since PrEP only protects against HIV.34\n\n\nMethods\n\nThis qualitative research was carried outed between May 2013 to August 2015. Convenience sampling method was used to recruit men aged 18 and over, who were non-British, lived in the country for at least one year, and who had worked or were still working as sex workers. The main recruitment sites were sexual health clinics and projects that provide health and social services to sex workers in London. Health professionals and health workers took part in the recruitment by providing the participant information sheet (PIS) and flyers to potential participants.\n\nAn interview guide was prepared using pertinent literature and first-hand information obtained from researchers and health professionals working in projects or programmes focused on sex work in London. The questionnaire (Underlying data)35 was tested on three MMSW, however the results were not included in the final analysis. Participants were interviewed in counselling rooms and small meeting rooms located in St. Marys hospital and City, University of London to secure their privacy. Literal transcription of the recordings was printed and revised by each participant to confirm their accuracy. In this manner, the risk of misinterpretation and misunderstanding was minimised. The software ATLAS.ti version 8·0 was used to store the data and thematic analysis was performed as previously described.36 Coding rules were established as well as a clear definition of the themes to avoid ambiguity or inconsistency. For this investigation, 'condomless sex' was defined as any penetrative or receptive sexual intercourse (oral, vaginal, or anal) without using condoms. Conversely, 'safer sex' was specified as the use of condoms for the aforementioned practices. The emergent themes were interpreted using the theory of planned behavior (TPB)37 framework. This theory is based on the significance of attitudes, norms, and perceived control to explicate different forms of risky behaviours,37 and to plan health interventions.38 In this manner, the analysis was focused on 1) attitudes towards the use of condoms: a result of personal beliefs about condoms; 2) subjective norms derived from participants’ perception of what others think about condoms (normative beliefs) and their motivation to comply with norms, and 3) perceived behavior control: participant’s beliefs about the degree of control they have over the use of condoms during the intercourse.\n\nThis study was revised and approved by the Ethics Committee of City, University of London (18 April 2013, Ref: PhD/12-13/18), by the NRES London Central Committee (9 October 2013, Ref:13/LO/1306, IRAS ID: 132947), and by the Research Committee of St. Mary’s Hospital (15 January 2014, Ref:13SM1864). Written informed consent was obtained from all individual participants included in the study. This form was revised and approved by the ethics committee.\n\n\nResults\n\nDue to the explicit nature of the interviews some quotes have been edited for clarity.\n\nIn this study a total of 25 MMSW (n = 25) participated. This sample was almost evenly represented by men from Latin-America and Europe. The socio-demographic characteristics of participants as well as their patterns of migration, and entrance into sex work suggest their diversity (Table 1). The whole group was operating as independent internet-based escorts, providing sexual services to men and women. The latter in the context of sexual services for ‘couples’ (man and woman). The analysis shows two dominant themes and distinct subthemes:\n\nA. Condomless sex with commercial sexual partners\n\nThis theme explains the perspective of the use of condoms within the context of sex work and experiences of condomless sex that contains three sub-themes:\n\n1. Policy on condom use with clients\n\nIt describes the attitude and perceived norm about the use of condoms with clients. The statements indicate that the entire group of participants were aware of the HIV and other STIs, claimed to be risk averse, and more importantly, stated a consistent use of condoms as a perceived norm because of their work. In line with this, many of them have made explicit their rejection to condomless sex in their online adverts:\n\n“Always, always condom, nothing without a condom, never, ever, ever. They can pay me any money, there are some offers, and some people asked me - do you do [condomless sex]? For that reason, in my profile I wrote no [condomless sex], don’t even ask me”\n\nConsistently, the majority of participants expressed a favourable attitude towards safer sex. For some, using condoms with clients was a way to differentiate sexual services from having sex in their personal life. Others thought that condoms were useful devices to avoid poor hygiene, odour, and some bodily features of clients that they find unattractive or dislike (e.g., overweight, excessive corporal hair). Most of the participants said that they especially demand the use of condoms to provide services such as vaginal intercourse, ‘full-service’ for men (usually include anal sex) and performing “passive” sexual role service (receptive anal sex):\n\n“I offer a full service, a complete service but I try to be specific because people always ask you if you do without condom, So I always say condom, I won’t give you my phone, I am a very discrete person and I only do outcalls.”\n\nBy contrast, other participants admitted an unfavourable attitude towards condoms. While the most mentioned concerns of losing clients that reject condoms, few did not like their use as it reduced their sexual arousal, especially with erection which caused difficulties in their sexual performance:\n\n“I know that I won’t have many clients if I insist on having oral sex with a condom, so I prepare to take that risk that is the only one that I prepare to take the chances.”\n\n“I don’t enjoy at all when I use a condom for [oral sex] because it is like you are sucking a rubber and I just get soft [lose the erection] when I wear a condom for a [oral sex], once again it is because of it something like squeeze it is a bit weird.”\n\nThe discrepancy between these two different perspectives suggests that risk awareness and intention of using condoms do not guarantee safer sex.\n\nIn addition, the analysis of the narratives showed that condomless sex with clients occurred in two different scenarios. In the first scenario participants perceived control of the situation and made a risk-taking decision to dismiss the use of condoms, and in the second, they lost control of the situation that ended in a condomless sex event. These are described under the following sub-themes:\n\n2. Decision-making process to accept condomless sex\n\nThis sub-theme explains the decision-making process that participants applied to dismiss the use of condoms with clients. They used two main processes a risk assessment of the clients, and performing sexual practices that participants catalogued as 'low-risk' in contracting HIV and other STIs. The participants assessed the risks of the client based on their physical appearance, followed by a subtle physical examination to identify sores, warts or blisters on genitals or rectum, or presence of penile discharge. Through this practice participants accepted condomless sex with customers that were labelled as ‘healthy’ and ‘clean’ (e.g., absence of warts):\n\n“I [had sex with] him all without condom, but I was not a risk as I could tell that I was probably the second or third person who he has sex with. I think I could tell about everything I do not think he has anything because he probably has very little sexual life.”\n\nThe participants also considered some social and demographic characteristics to rank clients as 'low risk' or 'high risk'. In this instance, participants favoured clients who were 'married men' for condomless sex as they were perceived as ‘straight’ men who ‘only have sex with their wives.’ In the same way, participants considered their ‘regular’ clients with whom they had established a long-term and trustworthy relationship, as ‘low risk’:\n\n“For example, yesterday I had a man from Barbados who looked very healthy, but I know he is from a high-risk region for these diseases. The guy was very clean, he was very nice”\n\n“There are a couple of people that I don’t use a condom because I know them for quite long time. I know it is not a good policy. I know I should use a condom with everybody and that’s it, but there are few people that I do that.”\n\nA second procedure to accept condomless sex with clients was the selection of sexual practices that participants considered as 'low-risk.' By far, condomless oral intercourse (COI) was the most frequent practice. Some participants mentioned that as additional strategies of protection they reduce the time for COI and avoid contact with the client’s semen. Although, condomless insertive oral intercourse (CIOI) and condomless receptive oral intercourse (CROI) were equally reported by participants, some assigned different levels of risk to each:\n\n“I know it is less risky when I suck him than when he sucks me or to kiss him. But it is not for everybody, depend on of the situation”.\n\nAnother recurrent risk reduction practice mentioned was performing condomless anal sex as the active sexual role (penetrative anal intercourse) instead of a passive role (insertive anal intercourse), as it was perceived as less risky:\n\n“Part of me think I am mainly top, I normally do not get people to [have sex with] me, I [have sex with] them, I am mainly top, and that is a very low risk, [censored]! I am not at risk because it is the very little rate to catch something if I am mainly top.”\n\n“I think, I am in this scene, I am earning money, but I am also scared because this is very risky, but I always pray to God please nothing bad happen.”\n\n3. Structural factors determining unplanned condomless anal sex\n\nThis sub-theme describes the role of four main factors that made participants to 'lose control' or to be under pressure to perform condomless anal intercourse. One of the most recurrent conditions was the use of recreational drugs that provoked a strong sexual arousal among participants. Many of these events occurred when they were providing ‘overnight' or 'chemsex’ services:\n\n“With crystal meth your brain is still more there, but with mephedrone you do not even think straight so much, you are so [aroused] that you do not even think, you only want sex, if I take mephedrone I know I am not going to use condoms”\n\nA second recurrent and independent condition was feeling strong attraction to a client:\n\n“Actually, I wasn’t on drugs, this time I wasn’t on drugs, I came to see this guy in the Ritz Hotel, and he was an Arabic, he was about my age, and he was so gorgeous! Sexy, he was like my God! I just wanted to eat him alive, he was so sexy and then, you know what actually I did it without condom”\n\nA third factor driving condomless anal sex was the financial reward offered by clients, which was mostly reported by participants who were in precarious conditions. In these situations, the participants felt that they could not reject the offer:\n\n“I had a client once, the same client three times because that client, he pays very well, much more that what I asked”.\n\n“I am at risk if you ask me how I feel about it, not very safe, not very clever. But I gamble for the best, I need the cash, I need the cash for food, I need the cash for transport, and I need to get out of this hole because I smile when I meet new people and everything, but when I am alone is not easy.”\n\n“Once I was really bad about money and a client called me and he wanted to take drugs also if you don't take drugs, you can last all that you need or you can even cope with the client.”\n\nThe fourth condition describes scenarios in which participants were overpowered by clients who removed or broke condoms or took advantage of the dynamic during the sex session to perform condomless anal intercourse. This condition was usually reported by the participants who offer a ‘passive’ sexual role as part of their sexual services:\n\n“I was having sex with a guy who was doing as active, and you know suddenly I saw him with the condom on his hand and I asking him, ‘Were you [having sex with] me without a condom?!”\n\nIn few cases, participants reported that these events occurred in a context of physical and verbal violence perpetrated by clients, or within a context of drugs use:\n\n“We were taking cocaine and drinking, I drank so much that day and I passed out […] few hours later, I woke up and the reason that I woke up was because something was painful, ok? And the painful thing was that he was [having sex with] me on my sleep, he was [having sex with] me on my sleep and without condom.”\n\nB. Condomless sex with non-commercial sexual partners\n\nThis theme describes experiences of unpaid or non-commercial condomless sex, which was defined as sexual acts without the use of condoms that were performed without any intention of material or economic reward. In general, many participants declared a more inconsistent use of condom with non-commercial sexual partners than with clients:\n\n“Then it happened again, but he wasn’t a client he was a person that I met, a casual partner and again it was three months of waiting for the test and I was - Oh my God, I shouldn’t do it again!”\n\n“I haven’t been in risk. My sexual practices are very low risk in the context of work, and the only person with I have been in high risk is with my ex-partner. During the time when we knew that he got infected we used protection until he completed the treatment”\n\nThis theme contains two sub-themes to differentiate condomless sex practice with formal sexual partners from casual sexual partners. Most of the participants reported having casual partners along with a formal partner in the past year.\n\nThe sub-themes are described below:\n\n1. Condomless anal sex with formal partner\n\nThe category of formal sexual partner was used by participants to describe people with whom they had a romantic, stable/regular or committed relationship. Almost half of the group reported to have male formal sexual partners. Some mentioned that these partners were also working as escorts, even few worked together. Most importantly, majority reported an irregular or complete lack of condom use with these partners. They decided not to use condoms when their partners agreed to just have sex with them, and/or knew both were HIV negative:\n\n“When I am dating someone if we both checked [got tested for HIV] and we both are fine, we do not use condom, like my ex that we split up two months ago, we were together for a year as we never use condoms, but I knew he only was sleeping with me”\n\nFor these participants condomless anal sex represented pleasure, intimacy, and commitment with their partners:\n\n“Have sex without condom with my ex-partner wasn’t good idea, even if that gives me more pleasure and it gives me more intimacy because sex between us hasn’t been the most important part of our relationship”\n\nCoherently, few participants said that they 'always' used condoms with their formal partners because they knew that one of them was HIV positive (serodiscordant couples):\n\n“He found out that he was HIV positive and then at that time I got syphilis from him and at that time I wasn’t working as an escort I was working as a cleaner and I didn’t get the HIV, so I got treated for syphilis, he got treated as well and from then we started to have sex with condom.”\n\n2. Condomless anal sex with casual partners\n\nCasual sexual partner was defined as people with whom participants engaged in sexual intercourse without a sense of commitment or emotional attachment. They mainly met casual partners using dating mobile applications, websites or in places such as clubs, saunas, or clubs. These participants decided not to use condom with these partners to satisfy their pleasure and personal enjoyment. Some admitted that they perceived the use of condom was optional:\n\n“We are humans and sex is the most animal part of us, you know, we are animals completely, so you cannot always control it, you have to accept it, if you always are having sex […] that is why you do without condom and see what happen.”\n\n“If someone carry a condom, then we do it with condom, or we just leave the condom around and try to see how it goes.”\n\nHowever, it is important to mention that many participants also acknowledged the role of recreational drugs and alcohol consumption as well as feeling sexually attracted to casual sexual partners in the practice of condomless anal intercourse:\n\n“When you are in drugs the only thing that you want is to have sex, well it depends, in my case I only wanted to have sex, for free, sex with people that if I could be rational, I wouldn’t like to have sex with, and exposing yourself to catch anything.”\n\n“The very last time was about 6 months ago and that was with a neighbour, a very, very hot Spanish guy who came around and we had some fun and when he started [having sex with] me without condom”\n\n\nDiscussion\n\nThis study has used the theory of planned behaviour to explore the risk perception and condomless sex experiences of 25 male migrant sex workers with commercial and non-commercial sexual partners. Unlike other research,39 the participants of this study knew the risks of HIV and other STIs, self-reported risk adverse, and consistently declared their position against condomless sex in their online adverts. However, despite their perception of safer sex as a norm and their intention of using condoms with clients, they revealed that condomless sex was a frequent practice.40–42 In accordance with this, the participants exposed an unfavourable attitude towards condoms due to displeasure, concerns of losing clients that reject condoms, and issues affecting their sexual response and performance. The latter, not so often acknowledged, highlights the importance of male sexual arousal in this type of work.43 In addressing past events of condomless sex with clients, this study identified a perceived behavioural control among participants that made this high-risk decision based on signs of physical evidence for HIV and other STIs in clients, assessing client’s social and sexual risk characteristics, and performing sexual practices that they considered 'low risk', to lessen the risk of transmission. These practices, that indicate a self-protective behaviour, demonstrate the persistence of inaccurate information about HIV and other STIs.15,44–47 In addition, this study identified factors that made the participants lose their perceived control and drove them to condomless anal sex. One of main factors was the use of recreational drugs with clients.48–50 Authors describe the use of drugs with clients as a social aspect of their occupation,51 while improving their performance.52 Further, some argue that this is a difficult aspect to avoid with clients who are regular drug users.49 Another interlinked factor was feeling sexually attracted to clients that implied opting for personal pleasure over a professional perspective.53,54 A third factor was the precarious situation that made participants to accept the financial reward offered by clients.55–58 Similar to other studies,55,58 physical domination and verbal violence perpetrated by clients was the other aspects facilitating condomless experience, which validates MSW's vulnerability regardless of the situation.59\n\nWithin the context of private life, this study found that differences in the perspective of condomless sex was related to the type of non-commercial sexual partner. As such, most of participants dismissed the use of condoms with formal partners as they were emotionally attached to these relationships. Participants also said that they agreed not to use condoms when they both were HIV negative and keep this practice strictly among themselves. Although condomless sex with formal partners was perceived as a safer practice, some participants had episodes of STIs that were associated to these sexual partners. Besides, it is worth noting that in line with other studies43,60 few participants reported a consistent use of condoms with formal partners when they were HIV-serodiscordant. Regarding condomless sex practice with casual sexual partners, participants said that it was more frequent with these sexual partners than with their clients. They connected this practice to strong sexual arousal and the use of drugs and alcohol. Drug use mainly initiates sexual interaction between gay and bisexuals,40,61 and more importantly,62,63 it facilitates sexual acts.64,65 Of serious concern is that these substances affect the perception and response to risk, directing the behaviour to high-risk sexual practices,66 and consequently, increase the possibility of contracting HIV and other STIs.67,68 This finding suggests the use of recreational drugs and attraction to the client are significant factors that intersected both private and sex work experiences of the study participants, and reinforce claims that condomless sex with casual partners can be a predictor of condomless sex with clients.53 Additionally, these results support the perspective that the type of sexual partner chosen in the MSW’s personal life can also be a risk factor.40,61\n\nOverall, participants that experienced condomless sex visited sexual health clinics to have screening tests for HIV and other STIs, as they felt exposed to contracting these infections. The most concern and anxiety was expressed for HIV, therefore almost all participants had requested for post-exposure prophylaxis (PEP) to reduce their chances of contracting this infection.69,70 Some had received PEP more than twice in the past 12 months. Few admitted to not continuing with their PEP treatment due to the adverse effects. These findings raise concerns of possible seroconversions when considering the poor medication adherence.71 It is relevant to mention that nearly the whole group of participants (22/25) had been diagnosed with one or more STIs including HIV (Table 1).\n\nInterpretation of the findings and the evaluation of their significance should be made considering the limitations of this study. For instance, the study design prevents the generalization of the findings. Also, limiting the recruitment of participants to sexual health clinics and health projects in London due to the recommendation of the research ethics committees, restrict the results only to the perspective and experiences of migrants who attend these services. Even with these limitations, this study is one of the few on male migrant sex workers in the UK that captures their experiences of high-risk sexual behaviour in detail. In addition, the heterogeneity of the sample provided a rich qualitative data on MMSW’s risky sexual behaviour with commercial and non-commercial sexual partners. Furthermore, this study provides in-depth socio-behavioural insights for designing more effective and tailored interventions for MMSW self-identified as homosexuals and bisexuals.\n\n\nConclusions\n\nParticipants experienced condomless sex with commercial sexual partners as risk-taking decision that intuitively triggered a set of risk reduction practices, which may not work effectively as they were based upon misinformation. Condomless sex with clients also occurred in a context of perceived loss of control with recreational drug use, experiencing precarious conditions, physical domination and verbal violence perpetrated by clients. These findings challenge claims that recreational drugs are not problematic among male escorts,72–74 that they work in safer environments, obtain higher earnings, and can control work conditions.73,75,76 In addition, condomless sex with non-commercial sexual partners was also a common practice, but clearly differentiated by the meanings attached to formal and casual partners. More importantly, this study found that the use of recreational drugs and attraction to the client are significant factors that intersected private and sex work experiences.\n\n\nRecommendations\n\nThe findings suggest the need to reinforce educational interventions to deliver appropriate information about the transmissibility of HIV and other STIs, improve skills of self-control, strengthen the risk-reduction counselling for those requesting PEP, and the use of condoms for those who decide to take pre-PEP as the best action to secure its success. Likewise, emphasising the relevance of training healthcare professionals to identify MMSW who use recreational drugs, and to facilitate their referral to programmes of harm reduction in substance use and mental health services. Finally, the identification of other subgroups among MMSW such as those whose partners are also sex workers, have a HIV serodiscordant partner, tend to have condomless sex with casual sex partners, and are experiencing difficult-living or working conditions, can allow for tailoring behavioural interventions.\n\n\nData availability\n\nRepository: Perceived risk and condomless sex practice with commercial and non-commercial sexual partners of male migrant sex workers in London, UK https://figshare.com/s/cbf4a21a9d93d63472c8.35\n\nThis project contains the following underlying data:\n\n• File docx. This file contains the blank interview questionnaire.\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 4.0 Public domain dedication).", "appendix": "Acknowledgments\n\nI would like to thank to my supervisors, student advisor, and health professionals of Working Men’s Project and SWISH project who collaborated with this study.\n\n\nReferences\n\nCDC: Compendium of HIV Prevention Interventions with Evidence of Effectiveness. Atlanta, Georgia:Centers for Disease Control and Prevention;1999 November 1999.\n\nUNAIDS: UNAIDS Data 2019: State of Epidemic. UNAIDS;2019.\n\nUNAIDS: UNAIDS Data 2020. Switzerland:Nations Programme on HIV/AIDS (UNAIDS);2020.\n\nVarghese B, Maher JE, Peterman TA, et al.: Reducing the risk of sexual HIV transmission: quantifying the per-act risk for HIV on the basis of choice of partner, sex act, and condom use. Sex. Transm. Dis. 2002; 29(1): 38–43. 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Loughborough University;2014." }	[ { "id": "96637", "date": "14 Oct 2021", "name": "Prof Victor Minichiello", "expertise": [ "Reviewer Expertise sex work", "qualitative data analysis" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nUnderstanding how male sex workers make decisions about condom usage with clients and in their personal sexual lives is an important question that this study addresses. Recent studies have explored this topic in some detail. This study aims to contribute by interviewing migrant male escorts in London and offering explanations that delve into why sex with a condom occurs or not in their encounters with clients.\nThis is an interesting paper, but there are several gaps:\nFirst, the literature review is not complete. I suggest the author read the recent work by John Scott and his colleagues on male sex work: see The Routledge Handbook of Male Sex Work, Culture and Society published by Routledge this year to better grasp the literature on this topic.\nSecond, could the author expand on new insights for public health campaigns concerning PrEP and HIV prevention that emerge from this study, and better argue how the results of this study further advance knowledge on the topic of condom use and safe sex among male escorts and particularly vulnerable groups like migrant sex workers?\nThird, some methodological issues require addressing. What topics were included on the interview guide? How were the interviews conducted? Can more detail be given about how the data themes were developed and what sort of qualitative data analysis was used?\n\nFinally, what new insights for public health campaigns emerge from this study?\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10320", "date": "19 Oct 2023", "name": "Elisa Ruiz", "role": "Author Response", "response": "Dear Victor, I am very grateful for your comments and suggestions to improve this paper. Please see my answers below: Reviewer comment: \"The literature review is not complete. I suggest the author read the recent work by John Scott and his colleagues on male sex work: see The Routledge Handbook of Male Sex Work, Culture and Society published by Routledge this year to better grasp the literature on this topic\" -Author answer: I have updated the literature including also some significant information from the source suggested by this reviewer. Reviewer comment: \"Second, could the author expand on new insights for public health campaigns concerning PrEP and HIV prevention that emerge from this study, and better argue how the results of this study further advance knowledge on the topic of condom use and safe sex among male escorts and particularly vulnerable groups like migrant sex workers?\" -Author answer: the recommendation section has been updated to highlight the promotion of condom use, as important element of the combine prevention of HIV. Reviewer comment: \"Third, some methodological issues require addressing. What topics were included on the interview guide? How were the interviews conducted? Can more detail be given about how the data themes were developed and what sort of qualitative data analysis was used?\" - Author answer: Additional information required by the reviewer has been added to the methods section. Reviewer comment: \"Finally, what new insights for public health campaigns emerge from this study?\" - Author answer: Recommendation section has been updated to address this suggestion," } ] }, { "id": "169988", "date": "15 May 2023", "name": "Carlos F. Caceres", "expertise": [ "Reviewer Expertise HIV and sexual health" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis is an interesting report about a relevant topic for the UK and worldwide. To make it more suitable for indexing, however, a few issues should be resolved:\nThe most visible one is the slang of the quotations (of migrant male sex workers who don't speak English as a first language). Probably there are guidelines as to how they should be edited.\n\nLikewise, there are some issues with the English language writing of the main text that limit its clarity and should be resolved.\n\nThe MMSW quotations should ideally include a code assigned to each participant, as well as their nationality. That would help interpreting the findings.\n\nThe introduction fails to contextualize the discussions in a framework of combination HIV prevention. Treatment as prevention (and U=U, that is viral suppression as risk-reducing) is never mentioned, PEP is mentioned once, and PrEP is presumably mentioned (\"pre-PEP\") in the recommendations only. It is unclear whether people who are having condomless sex are really out of any form of protection.\n\nIn that sense, the dates of implementation of this study should be specified, for better interpretation of the context.\n\nIn A.1, when describing unfavourable attitudes towards condoms: (a) the authors inappropriately use condom use in oral sex to illustrate the dilemmas of condom use, while it is not that relevant given its very low risk; (b) authors conflate MSW's fear of others rejecting condoms, with their own predisposition against condoms - those should be presented separately, although certainly with regard to anal, not oral sex.\n\nThe theory of planned behaviour should be used at least in the discussion.\n\nThe discussion should address other options of combination HIV prevention. Either at the introduction or at the discussion, current access to treatment, PEP and PrEP should be discussed as alternatives to condom use.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? No\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10321", "date": "19 Oct 2023", "name": "Elisa Ruiz", "role": "Author Response", "response": "Dear Carlos, Thanks very much for your comments and suggestions that helped me to improve the paper. Please see my answers below: Reviewer comment: The most visible one is the slang of the quotations (of migrant male sex workers who don't speak English as a first language). Probably there are guidelines as to how they should be edited. Author answer: the quotes have been edited by the editorial team of this journal according to their internal policies. In this manner, some words have been removed (as state the disclaimer section (in Results) Reviewer comment: Likewise, there are some issues with the English language writing of the main text that limit its clarity and should be resolved. Author answer: The text has been amended to improve their clarity. Reviewer comment: The MMSW quotations should ideally include a code assigned to each participant, as well as their nationality. That would help interpreting the findings. Author answer: variables as age, pseudonym and country of birth were part of the initial version of this manuscript, but they were removed by the editorial team of this journal. I will be happy to restore this information if they agree to this. Reviewer comment: The introduction fails to contextualize the discussions in a framework of combination HIV prevention. Treatment as prevention (and U=U, that is viral suppression as risk-reducing) is never mentioned, PEP is mentioned once, and PrEP is presumably mentioned (\"pre-PEP\") in the recommendations only. It is unclear whether people who are having condomless sex are really out of any form of protection. Author answer: I really appreciate this suggestion that open an interesting scope for further research. However, it is important to mention that the data collection for this study took place between 2013-2015, while PrEp and PEP initiatives were increasingly available since 2016. Participants who had access to them was mostly because were part of trials. Reviewer comment: In that sense, the dates of implementation of this study should be specified, for better interpretation of the context. Author answer: This info has been updated in abstract and methods section Reviewer comment: In A.1, when describing unfavourable attitudes towards condoms: (a) the authors inappropriately use condom use in oral sex to illustrate the dilemmas of condom use, while it is not that relevant given its very low risk; (b) authors conflate MSW's fear of others rejecting condoms, with their own predisposition against condoms - those should be presented separately, although certainly with regard to anal, not oral sex. Author answer: I understand this point, however, condomless sex was examined as an approach for HIV-STI prevention. Reviewer comment: The theory of planned behaviour should be used at least in the discussion. Author answer: Discussion section has been amended to satisfy this suggestion. Reviewer comment: The discussion should address other options of combination HIV prevention. Either at the introduction or at the discussion, current access to treatment, PEP and PrEP should be discussed as alternatives to condom use. Author answer: the introduction section has been amended to satisfy this recommendation." } ] } ]	1	https://f1000research.com/articles/10-1033
https://f1000research.com/articles/12-1376/v1	18 Oct 23	{ "type": "Research Article", "title": "Impact of COVID-19 on Mobile Technology use in adults in the United States", "authors": [ "Yi Lin", "Graham D. Rowles", "Arnold J. Stromberg", "Rola S. Zeidan", "Robert T. Mankowski", "Graham D. Rowles", "Arnold J. Stromberg", "Rola S. Zeidan", "Robert T. Mankowski" ], "abstract": "Background: Mobile technology (MT) has become essential in receiving information and services during the COVID-19 pandemic. Imposed quarantines could have led to varying adaptations of MT use. This study explored how COVID-19 impacted behavior, perception, and attitudes toward MT use in the United States. Methods: We distributed a pilot-tested survey online. Participants were MT users ≥ 35 years old. All participants responded based on their recalled experience of using MT before COVID-19 and their recent experience during COVID-19. Data analysis involved descriptive statistics and the Wilcoxon signed-rank test. Results: The average age of the 1212 participants was 56.1±12.2 years (55% female). Daily use frequency (from ≤3 to ≥4 hours/day) and perceived necessity (from some need to a strong need) during COVID-19 significantly increased (p<0.001) compared to before COVID-19. There was a significant increase (p<0.001) in video calls/meetings, online education, grocery/food delivery, and ordering taxi/car during COVID-19 compared to before. Participants increased (p≤0.001) their attention to the physical, social, and emotional benefits of using MT during the pandemic. COVID-19 increased MT use and acceptance in the United States. Conclusion: The knowledge gained from this study will help remove barriers to using and accepting MT and provide directions for MT development in middle-aged and older populations.", "keywords": [ "behavior", "perception", "perspective", "choice of function", "decision to use", "user need analysis", "technology acceptance model" ], "content": "Introduction\n\nCOVID-19 was a disease caused by the SARS-CoV-2 virus and has become a worldwide pandemic since 2020. The SARS-CoV-2 virus was easily spread from person to person (Rothan & Byrareddy, 2020), which led to long-lasting quarantines and the need for spatial distancing. Federal and local governments and public health professionals advised people to stay home, reduce their physical contact with others, and reduce their frequency of going to public places. The associated change in lifestyles directly increased the use of digital technologies such as next-generation telecommunication networks and artificial intelligence (Ting et al., 2020). Research indicated that using digital technologies could be a solution for pandemic preparedness and response; for instance, healthcare or surveillance technologies could be helpful for pandemic management (Whitelaw et al., 2020). Researchers suggested investments in mobile applications as valuable tools for developing digital health and enhancing communication (Torous et al., 2020). At least 114 mobile applications were developed and used during the COVID-19 pandemic (Collado-Borrell et al., 2020), which reflects the significance of mobile technology (MT) use as a coping strategy.\n\nWith its portable features and convenience, MT has become a primary source of information, knowledge, services, and connections (Bhavya & Sambhav, 2020). MTs (e.g., smartphones, tablets, or smartwatches) differ from traditional ones. Smartphones, for example, are feature-packed MTs that act like pocket-sized computers. They differ from traditional mobile phones that provide the function of making and receiving calls primarily. Internet access and mobile applications are essential elements that make MTs unique. Mobile applications are programs designed to work on a mobile device to provide various functions for specific purposes (Wallace et al., 2012), such as navigation, entertainment, or health care that supports daily life.\n\nDuring the COVID-19 pandemic, engagement in online shopping, education, entertainment, and food delivery has become increasingly important (Donthu & Gustafsson, 2020). MT provides the necessary channels to engage in activities during quarantine. Mobile applications contribute to managing the COVID-19 crisis by providing home monitoring, information sharing, contact tracing, and decision-making (Kondylakis et al., 2020). The COVID-19 pandemic has led to a phenomenon where many people have been forced to use MTs regardless of their preferences. Indeed, there are increasingly negative consequences of not using contemporary technologies in daily living. For instance, a person who does not use MT during COVID-19 will experience difficulties participating in social events, obtaining services, or interacting with others.\n\nThe nature of COVID-19 led to the phenomenon that middle-aged and older adults have a high risk of hospitalization rate and death rate after COVID-19 infection than younger people (Centers for Disease Control and Prevention, 2023), which increases the essentiality of MT use to lower the risk of infection and health management among middle-aged and older adults. In addition, middle-aged and older adults tend to be less tech-savvy than younger. Younger adults aged 18 to 28 are likely to use a wider variety of technologies than adults aged 65 or older (Olson et al., 2011). Use of mobile health applications, for example, research indicated that adults aged <35 in the United States are more likely to use mobile health applications than adults aged ≥35 years old (Bhuyan et al., 2016). The phenomenon of younger adults having higher acceptance and technology usage than middle-aged and older groups has been evidenced in the previously developed Unified Theory of Acceptance and Use of Technology, which used “age” as a moderator for technology use and acceptance (Venkatesh et al., 2003). Hence, this study adopted the concept of potential age difference in MT use experience and focused on MT users aged 35 or older.\n\nThe threat of the SARS-CoV-2 virus undoubtedly has made MT a normal part of life. However, we have limited knowledge of how COVID-19 changed behavior, perception, and perspectives on using MT. Therefore, this cross-sectional study sought to document and compare MT use before and during COVID-19 among middle-aged and older adults in the United States. This study defines MTs as portable products that contain touchscreen functions, Internet access, and mobile applications. We hypothesized that the pandemic would lead to an increase in daily MT use, an increase in the perceived necessity of use, an increase in new MT functions adoptions, differences in the choice of MT functions used compared to before COVID-19, and trigger a shift of values regarding factors that affect MT use decisions and variation of perspectives toward MT.\n\n\nMethods\n\nInclusion criteria for this cross-sectional study were people who are: (1) aged 35 or older; (2) able to understand and communicate in English; (3) living in the United States; and (4) MT users. Participants were recruited via online survey portals, Amazon Mechanical Turk (MTurk), and Prolific (a free alternative could be the SurveyPlanet software). A snowball sampling strategy was used to increase the response rate of participants aged 65 or older by sharing online survey links and electronic flyers with potential participants using email and social media. The previously pilot-tested survey was distributed online on March 17, 2021, and the online survey response window closed on August 02, 2021.\n\nAt the pilot testing phase of survey development, responses from 17 female and 16 male respondents were included in the data analysis. None of the participants reported difficulty in accessing or finishing the survey. Ten participants gave positive feedback on the survey content. One participant had minor suggestions on the wording of the survey and added extra response options on questions regarding education level and reasons for adopting MT for the first time. The survey was refined based on the participants’ feedback.\n\nThe pilot-tested survey contained both closed-ended and open-ended questions. If the participant agreed to join the study and confirmed they met the inclusion criteria, a paragraph introducing MT was displayed to the participant. For questions about MT use before the pandemic, all participants were asked to answer the questions by recalling their experience before COVID-19.\n\nThere were four sections in the survey. Eleven questions collected data on participant characteristics and assessed the impact of COVID-19 on the individual’s life. Twelve questions collected information on participants’ experience with MT use before and during COVID-19; these included questions on the types of MT used, how long the participants had used MT, the main reason for using MTs, the frequency of using MTs each day, the top five most frequently used functions, and the top three factors that influence the decision to engage in MT use. Twenty-two statements (including 16 positive and six negative attitudes measured by a four-point Likert score) collected information about perspectives toward MT; these statements were adopted and modified from the extended Unified Theory of Acceptance and Use of Technology (UTAUT2), and studies aimed to examine MT acceptance and use (Boontarig et al., 2012; Shaw & Sergueeva, 2019; Venkatesh et al., 2012). Finally, two open-ended questions encouraged respondents to comment on their responses in more depth. The list of the questions can be found in the Extended data (Lin, 2023).\n\nA total of 2,035 responses were recorded on the Qualtrics XM database (a free alternative could be the SoGoSurvey platform). Survey responses that met any of the following conditions were excluded from data analysis: Participants who did not meet the inclusion criteria (n=576), incomplete survey responses (participants who failed to read all the questions or skipped 50% of questions) (n=174), and unreliable survey responses (n=73). The strategies to identify unreliable survey responses included 1) participant spent less than three minutes on the survey; 2) duplicates or responses flagged as spam (when multiple identical responses were submitted from the same IP address within 12 hours) by the Qualtrics XM software; and 3) survey completed by a robot (the Qualtrics XM has adopted Google’s invisible reCaptcha technology for robot detection). After data cleaning, 1,212 usable survey responses were kept for analysis.\n\nData analyses were completed using IBM SPSS version 28, which involved descriptive statistics demonstrating participant characteristics and Wilcoxon signed-rank test for the paired data that explored differences before and during COVID-19. Quotes collected from open-ended questions were used to support the findings and enrich the discussion and conclusion.\n\nThis study was approved by the University of Kentucky Medical Institutional Review Board (IRB) on Feb 25, 2021 (IRB number: 65430), and was performed in line with the principles of the Declaration of Helsinki.\n\nAn IRB-approved electronic informed consent form was presented as the online survey cover letter and obtained from all participants included in the study. Each participant had to read the electronic consent and provide consent by ticking the box that represents agreement.\n\n\nResults\n\nThe median time to finish the survey is 13.3 minutes (interquartile range=8.6 minutes). As evidenced by the ZIP code, responses were received from a nationwide sample (20.5% from the West, 18.7% from the Midwest, 40.6% from the South, 17.7% from the Northeast region, and 2.4% did not provide a ZIP code). The average age of participants was 56 (ranging from 35 to 83). More than half (55.4%) were female. Most of the participants were white Caucasians (78.6%), were currently married (59.2%), had a college or higher education level (84.5%), and had annual incomes of USD 50,000 or higher (55.2%). More than three-quarters of the participants (77.9%) indicated good or excellent self-reported health status. With regard to residence, most were community-dwelling: 75.4% lived in a house, 21% in an apartment, and 1% in a retirement community. No participant lived in a skilled nursing facility.\n\nMost of the participants (89.4%) owned two or more MT devices: 96.9% owned a smartphone, 74.8% owned an iPad or tablet, 32.3% owned a smartwatch, and 7.3% owned other MT devices, including a Kindle, laptop, and personal digital assistant. With regard to the length of use experience, 89.1% had used MT for more than three years, 5.4% for one to three years, 3.6% for more than six months but less than a year, and only 1.7% for six months or less. Responding to a question on the circumstances behind their initial decision to use MT, more than two-thirds (67.2%) indicated they needed it to function in today’s society, 39.5% adopted MT for its mobile applications, 38.2% because everyone around them was a user, and 26.8% due to job requirements. Finally, 9.8% stated other reasons that influenced their decision to adopt MT including “curiosity,” “safety,” “fun,” and “make extra money.”\n\nDuring the pandemic, most participants (78.7%) reported having to stay in their residence more than before, 5.6% reported staying less, and 15.5% reported spending the same time. Two-thirds of the participants (66%) stated that they had less chance to go out due to the pandemic, 72.5% indicated having less opportunity to interact with people, and 33.3% spent more time staying with family due to the pandemic.\n\nRegarding physical status, 25.4% reported having less chance to exercise during the pandemic. In contrast, 22.7% reported having more opportunities to exercise. Participants’ self-reported mental status indicated that 23.4% felt depressed, 34.9% felt anxious, and 38.9% felt stressed due to the pandemic. Finally, a few participants (7.9%) thought the pandemic did not affect their lives, and 18.6% felt their health was unaffected by the pandemic.\n\nMT daily use frequency during COVID-19 was significantly increased (z=-19.740, p<0.001) compared to before (Table 1). Less than half of the participants (43.7%) spent more than 4 hours per day using MT before the pandemic, but this number increased to 68.2% during the pandemic.\n\nAll the participants provided information about their five most used MT functions before and during COVID-19 (ranked from most to least used). According to the ranks assigned by the participants to each function, the “text messaging” function was the most used before COVID-19, followed by “emails,” “voice calls/meetings,” “GPS/navigation,” “following breaking news,” and “playing games.” The most used MT functions during COVID-19 were “text messaging,” followed by “emails,” “voice calls/meetings,” “taxi/car service,” “online shopping,” and “video calls/meetings.” There was a significant rise (p<0.001) in the rank of harnessing the following MT functions during COVID-19 compared to before: video calls/meetings (percentage of participants who reported an increased rank: 29.3%, z=-12.160), online education (participants: 9.7%, z=-4.726), grocery/food delivery (participants: 22.0%, z=-11.083), online shopping (participants: 28.9%, z=-5.110), getting information about a health condition (participants: 11.6%, z=-5.652), and ordering taxi/car services (participants: 23.5%, z=-5.531).\n\nIn addition to use frequency and choice of functions, more than half of the participants (54.5%) reported experience using a new MT function during the pandemic. These included: online meetings, shopping (including delivery or pickup), health monitoring, receiving news updates, making reservations and checking-in, fitness programs, entertainment, and banking. In the open-ended section of the questionnaire, participants described their experience:\n\n“I now use curbside pickup and check-in using store apps, and I had never done that prior to the pandemic.” (46-year-old female)\n\n“I have used a lot more the video calls function, either to communicate with my family and friends or for work. Online shopping became a necessity during the pandemic too.” (36-year-old female)\n\n“I used mobile technology for online banking as it was not possible to go to a physical location for a long time.” (65-year-old male)\n\nThere was a significant increase (z=-20.448, p<0.001) in the feeling of necessity to use MT during COVID-19 compared to before (Table 1). Few participants (7.0%) did not feel the need to use MT in their daily life; this number decreased to 2.7% during the pandemic. One-third of the participants (33.2%) felt a strong need to use MT every day before COVID-19, and the number increased to 73.5% during COVID-19.\n\nAll the participants identified the top three factors (ranked from most to least important) that affected their decision to use MT before and during COVID-19. According to the ranks assigned by participants, the most crucial factor affecting the decision to use MT before COVID-19 was the “availability of functions that support daily life,” followed by the “necessity of using MT,” “ease of use,” and “pleasure of using MT.” On the other hand, the most crucial factor that affected the decision to use MT during COVID-19 was the “availability of functions that support daily life,” followed by the “necessity of using MT,” “social benefits of using MT,” and “ease of use.” There was a significant rise (p≤0.001) in considering the following factors essential in deciding to use MT during COVID-19 compared to before: physical benefits (percentage of participants who reported an increased rank: 14.9%, z=-4.440), social benefits (participants: 23.1%, z=-3.238), emotional benefits (participants: 18.3%, z=-6.343), the availability of functions that support daily life (participants: 29.3%, z=-4.265), and necessity (participants: 27.3%, z=-4.902).\n\nParticipants’ agreement level to fifteen statements regarding positive perspectives toward MT significantly increased (p<0.001) during COVID-19 compared to before (Table 2). These statements reflected participants’ views about ease of use, the pleasure to use, and the benefits of using MT. Participants’ agreement level to one statement regarding positive view did not reach a significant difference (p>0.05) compared between the time before and during COVID-19; the statement was “MTs are reasonably priced.”\n\na Based on positive ranks (agreement level during COVID-19 > before COVID-19).\n\nb Based on negative ranks (agreement level during COVID-19 < before COVID-19).\n\nParticipants’ agreement level with two statements regarding negative perspectives toward MT significantly decreased (p<0.001) during COVID-19 compared to before. These statements reflected pleasure to use and emotional benefits, which were 1) “Using the MT is annoying;” and 2) “I have fear of using MT.” Two statements regarding negative views significantly increased (p<0.001) during COVID-19 compared to before. These statements reflected perceived risk and perceived sacrifice of using MT, which were 1) “I am worried that my privacy could be threatened due to the use of MT;” and 2) “Using MT limits my human interaction in daily life.” Two statements regarding negative attitudes did not reach a significant difference (p>0.05) between the time before and during COVID-19. These statements reflected the perceived sacrifice and perceived price value of MT, which were 1) “Using MT prevents me from developing other habits;” and 2) “MTs are too expensive for me to use.”\n\n\nDiscussion\n\nIn this study, we have found a significant increase in daily use frequency and perceived necessity during COVID-19 compared to before. A significant increase was found in using several MT functions such as video calls/meetings, online education, grocery/food delivery, and ordering taxi/car services during COVID-19. More than half participants in this study had experience in adopting new MT functions during COVID-19. Participants increased their attention to the physical, social, and emotional benefits of using MT during COVID-19. Moreover, participants’ perspectives toward MT during COVID-19 positively increased.\n\nPeople in at least 42 states were urged to stay home during the outbreak of COVID-19 (Mervosh et al., 2020). Evidence indicates that daily human mobility was reduced by 5% in the United States due to the stay-at-home policies (Xiong et al., 2020). In addition, previous studies found that the pandemic was associated with increased health anxiety and financial worry (Tull et al., 2020), led to significantly reduced physical activity (Flanagan et al., 2021; Knell et al., 2020) or an increase of health-enhancing behavior due to having more time (Knell et al., 2020). Our study discovered changes in emotional status and time spent on physical activity during COVID-19, reinforcing previous studies’ findings.\n\nOur study identified a significant increase in daily use frequency and perceived necessity to use during COVID-19. Similar results were found in a study in Jeju-si, South Korea (156 participants, age range: 15 to 80 years old), which reported increased MT use time due to COVID-19 (Chae, 2020). Increased MT use frequency and perceived necessity suggested the potential influence of stay-at-home policies and spatial distancing strategies that were commonly applied during the pandemic.\n\nChanges in commonly used functions of MT and the adoption of new functions during COVID-19 were discovered. Although text messaging, email, and voice calls were consistently the top three most used functions before and during COVID-19, people began to use more functions during COVID-19. These changes reflected different human needs for maintaining the quality of life during the pandemic. In-store shopping and meeting in person became very difficult during COVID-19, leading to the use and adoption of MT functions. Increased use of taxi/car services reflected people’s perception of the risk of using public transportation (e.g., bus) during the pandemic. As a result, demand for public transportation declined worldwide during the pandemic (Tirachini & Cats, 2020). The change in behavior in using MT functions reflected adaptation to the situation in fulfilling daily tasks and transportation needs.\n\nThis study identified changes in prioritizing factors that influenced MT use decisions during COVID-19. The availability of functions that support daily life and the necessity of using MT were the top two factors before and during COVID-19, which reflected the common motivation to use MT. When MT becomes a requirement for living, people have no option but to adopt the technology to stay engaged. During COVID-19, people started to weigh the physical, social, and emotional benefits they could gain in deciding whether or not to use mobile devices. The finding suggested the importance of developing MT functions that create apparent benefits and introducing MT use benefits to users, which could be a solution for improving the quality of life in particular areas. For instance, a previous study demonstrated the benefits of telehealth coaching for patients who live in rural areas (Young et al., 2014). Based on our findings, healthcare delivery via a mobile device will be even more widely used in rural areas if the targeted users receive sufficient information regarding the benefits of remote health-promotion services.\n\nUsers’ perspectives toward MT became more positive in almost all dimensions during COVID-19 except the views of perceived risk and sacrifice. As for the price of mobile devices, people were likely to have similar perspectives toward it before and during COVID-19 since there was no remarkable adjustment in salary during the pandemic. The COVID-19 pandemic indeed created a once-in-a-lifetime platform to display the advantages of using MT; the effects were reflected in the changes in perspectives toward MT. However, we should not ignore that frequent use of MT might increase the privacy risk and prevent people from engaging in other in-person activities.\n\nThis study included the responses from participants of each region of the United States, which made our findings representative and reflected the MT use phenomenon in the United States. Our data cleaning process was strict and included multiple criteria to exclude unreliable data. Our study identified a shift of values regarding factors that affect MT use decisions, which offers the potential for developing strategies to increase MT adoption. The increase in MT adoption would provide a vast platform for delivering services to fulfill the variety of needs of MT users. Moreover, the findings of this study indicated a decrease in the variation of perspectives toward MT. We discovered substantial growth of acceptance toward MT during COVID-19; this is thus an excellent time to educate people about technology use and encourage teleservices development. Although the pandemic is less severe compared to the beginning of the outbreak and the COVID-19 restrictions are mostly lifted, the knowledge from this study in MT use during COVID-19 may serve to understand technology needs in this population better and to better plan for implementing new technologies.\n\nOur study had several limitations—first, the lack of diversity in participants and recruitment methods. The data was collected during the severe period of the pandemic; thus, the recruitment method was extremely limited. Second, adding additional questions such as participants’ household arrangements, employment information, and the impact of COVID-19 on participants’ health would enrich the findings. Third, the survey study required participants to recall their experience with MT before COVID-19, which may lead to memory recall bias. Fourth, the survey question design could not consider personal factors (such as personality, intelligence, emotional state, and learning ability) that are more complex to measure. Fifth, this study might have response bias since the data was self-reported. Although the self-reported data had the advantage of revealing participants’ actual thoughts, participants’ concerns with social desirability might influence the data unconsciously.\n\nFinally, it is essential to acknowledge that a shift in MT use behavior might already have occurred prior to the pandemic; COVID-19 might have simply provided a convenient excuse or trigger in accelerating behavior change. The data in this study do not allow us to explore MT use behavior changes before the pandemic, but such changes certainly occurred during the pandemic. More details about participant information and data on behavioral decisions during COVID-19 with respect to MT use would provide deeper insight.\n\nThis study could help design future studies to examine the relationships between health, technology implementations, acceptance, and adoption among middle-aged and older adults in the United States during the continuous waves of pandemic and post-pandemic. Comprehensive knowledge regarding the associations between MT use behavior, physical health, and mental health warrant longitudinal studies in the future. MT use could become even more critical in maintaining the quality of life; thus, future studies could consider recruiting diverse participant groups, including non-technology users, to explore MT use and adoption. Providers working on service delivery via mobile devices should explore strategies for increasing learning motivation. Developing strategies to give people a comfortable experience while embracing technology and making people in diverse situations learn fast will be more critical than ever. A 56-year-old female participant in the study explained, “I worry that as I get older, the world will depend more on mobile technology, and my ability to use it will decline.” A 63-year-old female who described her experience of using MT noted, “The technologies change so quickly, that I gave up on trying to keep up with all of them.” Technology use is supposed to bring convenience and efficiency to human beings; it is essential to reduce the burden of people trying to use it. Following the rapid steps of technology evolution, the question of how to provide a friendly interface or strategy for human-machine interaction demands immediate attention.\n\n\nConclusion\n\nThe environment under the influence of COVID-19 has increased MT use behavior, acceptance, and mobile application adoption in the United States. The knowledge gained from this study will have implications to help remove barriers to using and accepting MT and provide directions for MT development and implementation in middle-aged and older populations.", "appendix": "Data availability statement\n\nAnonymized data from this study are available on request from the corresponding author [YL] from the date of publication. The anonymized data are for academic use only; the request should contain a full research plan. The data are not publicly available as they contain information that could compromise research participant consent.\n\nFigshare: Questionnaire for Mobile Technology Use during COVID-19, https://doi.org/10.6084/m9.figshare.23613828.v1 (Lin, 2023).\n\nThis project contains the following extended data:\n\n- Survey questions for investigating mobile technology use behavior during COVID-19\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgments\n\nThe paper was prepared from part of a published dissertation entitled “Mobile technology use during the COVID-19 Pandemic (Lin, 2021)” available via https://uknowledge.uky.edu/gerontol_etds/19/. We want to express our deep gratitude to all the study participants for their enthusiasm and thoughtful responses.\n\n\nReferences\n\nBhavya R, Sambhav S: Role of mobile communication with emerging technology in Covid-19. International Journal of Advanced Trends in Computer Science and Engineering. 2020; 9(3): 3338–3344. Publisher Full Text\n\nBhuyan SS, Lu N, Chandak A, et al.: Use of mobile health applications for health-seeking behavior among US adults. J. Med. Syst. 2016; 40: 1–8. Publisher Full Text\n\nBoontarig W, Chutimaskul W, Chongsuphajaisiddhi V, et al.: Factors influencing the Thai elderly intention to use smartphone for e-Health services. 2012 IEEE symposium on humanities, science and engineering research. IEEE; 2012, June; pp. 479–483.\n\nCenters For Disease Control And Prevention: Risk for COVID-19 Infection, Hospitalization, and Death By Age Group. COVID-19.2023, April 7. Reference Source\n\nChae S-G: A survey on the use of mobile phones due to COVID-19. International Journal of Internet, Broadcasting and Communication. 2020; 12(3): 233–243.\n\nCollado-Borrell R, Escudero-Vilaplana V, Villanueva-Bueno C, et al.: Features and functionalities of smartphone apps related to COVID-19: systematic search in app stores and content analysis. J. Med. Internet Res. 2020; 22(8): e20334. Publisher Full Text\n\nDonthu N, Gustafsson A: Effects of COVID-19 on business and research. J. Bus. Res. 2020; 117: 284–289. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFlanagan EW, Beyl RA, Fearnbach SN, et al.: The impact of COVID-19 stay-at-home orders on health behaviors in adults. Obesity. 2021; 29(2): 438–445. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKnell G, Robertson MC, Dooley EE, et al.: Health behavior changes during COVID-19 pandemic and subsequent “stay-at-home” orders. Int. J. Environ. Res. Public Health. 2020; 17(17): 6268. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKondylakis H, Katehakis DG, Kouroubali A, et al.: COVID-19 mobile apps: a systematic review of the literature. J. Med. Internet Res. 2020; 22(12): e23170. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLin Y: Mobile technology use during the COVID-19 Pandemic. [Doctoral dissertation, University of Kentucky]. UKnowledge.2021. Reference Source\n\nLin Y: Questionnaire for Mobile Technology Use during COVID-19.pdf. [dataset]. figshare. 2023. Online resource. Publisher Full Text\n\nMervosh S, Lu D, Swales V: See which states and cities have told residents to stay at home.2020, April 20. Reference Source\n\nOlson KE, O’Brien MA, Rogers WA, et al.: Diffusion of technology: frequency of use for younger and older adults. Ageing Int. 2011; 36(1): 123–145. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRothan HA, Byrareddy SN: The epidemiology and pathogenesis of coronavirus disease (COVID-19) outbreak. J. Autoimmun. 2020; 109: 102433. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nShaw N, Sergueeva K: The non-monetary benefits of mobile commerce: Extending UTAUT2 with perceived value. Int. J. Inf. Manag. 2019; 45: 44–55. Publisher Full Text\n\nTing DSW, Carin L, Dzau V, et al.: Digital technology and COVID-19. Nat. Med. 2020; 26(4): 459–461. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTirachini A, Cats O: COVID-19 and public transportation: Current assessment, prospects, and research needs. J. Public Transp. 2020; 22(1): 1–21. PubMed Abstract \| Publisher Full Text\n\nTorous J, Myrick KJ, Rauseo-Ricupero N, et al.: Digital mental health and COVID-19: using technology today to accelerate the curve on access and quality tomorrow. JMIR Ment. Health. 2020; 7(3): e18848. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTull MT, Edmonds KA, Scamaldo KM, et al.: Psychological outcomes associated with stay-at-home orders and the perceived impact of COVID-19 on daily life. Psychiatry Res. 2020; 289: 113098. Publisher Full Text\n\nVenkatesh V, Morris MG, Davis GB, et al.: User acceptance of information technology: Toward a unified view. MIS Q. 2003; 27: 425–478. Publisher Full Text\n\nVenkatesh V, Thong JY, Xu X: Consumer acceptance and use of information technology: extending the unified theory of acceptance and use of technology. MIS Q. 2012; 36: 157–178. Publisher Full Text\n\nWallace S, Clark M, White J: ‘It’s on my iPhone’: attitudes to the use of mobile computing devices in medical education, a mixed-methods study. BMJ Open. 2012; 2(4): e001099. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWhitelaw S, Mamas MA, Topol E, et al.: Applications of digital technology in COVID-19 pandemic planning and response. Lancet Digit. Health. 2020; 2(8): e435–e440. Publisher Full Text\n\nXiong C, Hu S, Yang M, et al.: Mobile device location data reveal human mobility response to state-level stay-at-home orders during the COVID-19 pandemic in the USA. J. R. Soc. Interface. 2020; 17(173): 20200344. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYoung H, Miyamoto S, Ward D, et al.: Sustained effects of a nurse coaching intervention via telehealth to improve health behavior change in diabetes. Telemedicine and e-Health. 2014; 20(9): 828–834. Publisher Full Text" }	[ { "id": "219469", "date": "21 Dec 2023", "name": "Sri Rahayu Natasia", "expertise": [ "Reviewer Expertise Information System Evaluation", "Technology Acceptance", "UI/UX Research" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nI think this paper is superb, I enjoyed reading it. The overall structure is coherent and well-explained. The authors explain the introduction part clearly and refer to the latest publications as they talk about the phenomena of MT use during COVID-19. The method section, instrument, and data cleaning are excellent for producing high-quality and reliable data for further analysis. This paper discovers changes in MT use behavior, MT use perception, and perspective toward MT on users before and during the pandemic COVID-19 in the USA. Strengths and limitations, also future directions of this research are insightful to the reader and potential researcher on a similar topic.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nI cannot comment. A qualified statistician is required.\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1376
https://f1000research.com/articles/12-1375/v1	18 Oct 23	{ "type": "Research Article", "title": "A cross-sectional audit and survey of Open Science and Data Sharing practices at The Montreal Neurological Institute-Hospital", "authors": [ "Sanam Ebrahimzadeh", "Kelly D. Cobey", "Justin Presseau", "Mohsen Alayche", "Jessie Virginia Willis", "David Moher", "Sanam Ebrahimzadeh", "Kelly D. Cobey", "Justin Presseau", "Mohsen Alayche", "Jessie Virginia Willis" ], "abstract": "Background: Open science is a movement and set of practices to conduct research more transparently. The adoption of open science has been recognized to support innovation, equity, and transparency. The Montreal Neurological Institute-Hospital (Neuro) has committed to becoming an ‘open science’ institute, the first of its kind in Canada. Here we report on an audit of open data practices in Neuro publications and on a survey of Neuro-based researchers’ barriers and facilitators to data sharing. Methods: In the first study, we retrieved 313 unique publications and collated all Neuro publications from 2019 and extracted information from each article pertaining to data sharing and other open science practices. We included all empirical papers and pre-prints that were reported in English. In the second study, one hundred twenty-four participants (out of 553) completed the survey, with a response rate of 22.42%. We surveyed all Neuro researchers. For the audit, we examined data sharing and open science practices. For the survey, we asked participants questions about their data sharing practices and perceptions. Results: We found that 66.5% of these publications (n=208) included a data sharing statement. Overall, 74.5% (n=155) of articles had data that was publicly available. When examining broader open science practices, rates of compliance tended to be lower. For example, 94.9% (n=297) of publications failed to register a protocol. Among participants who had published a first or last authored paper in the past year, most participants, 53 of 74 (71.62%), reported that they had openly shared their research data. Less than half of the participants, 37.50% (n=45), reported having engaged in training related to data sharing within the last 12 months. Conclusion: We found that half of all publications included in the audit shared data. Participants indicated an appetite for resources for learning about data-sharing signaling a willingness to perform better.", "keywords": [ "Open Science", "Data Sharing", "Audit", "Survey", "Facilitators and barriers" ], "content": "Introduction\n\nOpen science refers to the practices of making research outputs openly available for others to use and build upon. Open science can be seen as an alternative to the typically ‘closed’ model of science which tends to stress concern over intellectual property and controlling access to scientific knowledge and data.1 Benefits to open science include increased transparency, the ability to replicate research, equity in access to research information, reputational gains and increased chances of publication.2 Recognizing the value of open science, many stakeholders have begun to implement policy mandates to see that open science practices are being implemented in the community. This includes the creation of federal roadmaps and policies, organization policies, and funder policies, such as Canada’s Roadmap to Open Science,3 the second French national plan for open science,4 the United Nations Educational, Scientific and Cultural Organization’s open science policy,5 the European Union’s open science policies,6 and others.\n\nBeyond these stakeholders, academic institutions have an important role in implementing open science.1 Despite their seemingly obvious role, academic institutions have tended not to feature in discussions around implementing open science. Indeed, academic institutions have been previously criticized for not playing more of a role in addressing issues of research reproducibility7 which is conceptually related to open science. The reality is that few academic institutions have strong transparent processes in place to encourage open science. Despite this, research institutions are uniquely positioned to help contribute to defining incentives in research and to valuing open science practices. Shifting researchers’ behavior towards openness will require education, training, as well as a culture shift. Research institutions could provide the environment of this training to verify common understanding of open science and to help shift culture. Inaction by research institutions to play their role in implementing open science will have downstream consequences for how research is disseminated.\n\nIn Canada, a notable exception is the Montreal Neurological Institute-Hospital (henceforth called the ‘Neuro’) which has committed to becoming an open science institute. Having made the scientific and cultural decision to become ‘open’ after a significant consultation and buy-in process,8 the Neuro is now positioned to implement open research practices. The Neuro has made a structural decision to run a focused implementation program wherein a small set of open science practices are initially focused on, and additional practices then added in an incremental manner.\n\nAt present there is a focus on data sharing, namely making the data underlying the results reported in a given publication publicly available for others to use and build upon, or to verify the work reported. The rationale of choosing data sharing from the potential open research practices was: 1. Few researchers have the skills required to share their data and most researchers are not trained in data sharing9; 2. Data sharing has the potential to lead to novel discovery and enhances the transparency of disseminated research findings; 3. The largest government health funder in Canada, Canadian Institutes of Health Research, recently announced new polices on data management.10 By adopting data sharing now, the Neuro stands to be ahead of incoming mandates and may offer insight for others to follow. Finally, while some data sharing infrastructure exists, a significant investment is being made at the federal level to build capacity for Canadian researchers.11,12\n\nThis article describes the results of two studies examining data sharing practices at the Neuro. In the first study, we audited all publications produced by Neuro researchers in 2019. In the second study, we surveyed Neuro-based researchers about barriers and facilitators to data sharing. The results will provide the Neuro with a better understanding of barriers and facilitators to data management and sharing and will identify educational needs related to data sharing that can be implemented to address barriers. Further, the audit of publications can be used to benchmark for improvements over time and to monitor change.\n\n\nMethods\n\nThe study received ethical approval from the Ottawa Health Science Research Network Research Ethics Board (OHSN-REB #20210514-01H).\n\nWe used the STROBE guideline to inform our reporting of the audit13 and the CHERRIES guideline to inform reporting of the survey.14\n\nSearch strategy\n\nWe identified the Neuro publications produced in 2019 by searching the Web of Science (WoS)15 and using its meta-data to capture the Neuro’s output including searching two preprint servers MedRxiv16 and bioRxiv.17 The search strategy was developed by trained information specialists and librarians (Sanam Ebrahimzadeh and Alex Amar-Zifkin). For the full search strategy please see Extended Data.\n\nPatient and public involvement\n\nPatients or the public were not involved in the design, or conduct, or reporting, or dissemination plans of our research.\n\nEligibility criteria\n\nWe aimed to include all research papers disseminated by researchers based at the Neuro in 2019. We included research papers within any field. We used the last listed date of publication on each paper to determine if the publication fits within this timeframe. We included all research publications irrespective of the role of the Neuro based author (e.g., including trainees, graduate and postdoctoral students, early career researchers and more established researchers). We included all publications where a Neuro-based author was listed, irrespective of where they were named on the author byline. We included publications in English only. We included publications in traditional peer-reviewed journals as well as those on preprint servers.\n\nScreening and extraction\n\nDistillerSr18 was used to manage study records retrieved by the search, this process could also have been accomplished manually, but DistillerSr helped with organization of files. We obtained all full-text documents, and two reviewers independently screened these records against our inclusion criteria for included articles, we then extracted basic epidemiological information from each included study, including the names of the Neuro based author(s), the outlet of publication, and we classified articles based on their study design and content area. In addition, we extracted information related to data sharing practices. This included information on whether the publication contained a data sharing statement, whether or not data sharing occurred, and if so, what format and tools were used for this sharing. In addition, we extracted information on a range of other open science practices as per Table 1.\n\nData analysis\n\nOnce all extracted data were in agreement (i.e. discrepancies between assessors resolved) the complete dataset for all included articles was exported from DistillerSR into SPSS 2819 where data were cleaned. We presented the total number of included articles, and basic descriptive analyses for all items extracted using count data and percentages. Also, we used Unpaywall20 to check the open access status of the articles captured in our sample. To do so, we inserted the DOIs we extracted from the included articles into the Unpaywall tool.\n\nSampling\n\nThe survey was closed (i.e., only open to those we invited) and was administered using SurveyMonkey software.21\n\nSurvey items\n\nAfter providing informed consent, participants were presented with a 14-item survey (See Extended data). The survey was custom-built but draws on items previously reported by Van Panhuis.22 We presented participants with a series of questions regarding their willingness to share data that were developed using the 14 domains of the Theoretical Domains Framework (TDF) to help structure the survey items pertaining to barriers and facilitators to data management and actual data sharing. The TDF was created to better understand health professional behavior and is an integrated theoretical framework.23 Participants were asked to indicate their level of agreement with each statement on a Likert scale of 1 (strongly disagree) to 5 (strongly agree) was used to allow the participants to express how much they agree or disagree with each item. The Neuro ‘Open Science Grassroots committee’, a group of Neuro-based researchers targeting improvements in open science at the institution and beyond,24 reviewed and provided feedback on the survey which we incorporated. The survey was also piloted by three researchers prior to dissemination for clarity.\n\nSurvey recruitment\n\nWe included all the Neuro’s currently employed graduate students, postdoctoral candidates, research support staff, and independent investigators. The list of researchers was provided to us by the Neuro’s human resources. Participants were invited to complete the survey by the Director of the Neuro, via email, using a standard recruitment script. Participants were asked to complete informed consent online. The consent form described the aims of the study, specify that data collected would be anonymous, and detail our data management plan which includes making data openly available. Completion of the survey was by as implied consent.\n\nParticipants were originally sent the survey on September 20th, 2021, with a standardized reminder sent after one week (September 27th, 2021). Some targeted internal email strategies were also implemented to help maximize the response rate. We closed the survey on November 3rd, 2021.\n\nData analysis\n\nData was analyzed using IBM SPSS 28. We also report the completion rate for individual items, and report frequencies and percentages, or means and standard deviation, for each of the survey items. We report comparative data, using a Chi square test, illustrating findings by researcher category and gender.\n\nFor open-ended survey questions, we conducted a thematic analysis25 of responses provided. Here, two researchers familiarized themselves with the qualitative responses. Then, each of them independently coded responses. Codes were discussed and iteratively reviewed until there was consensus. Then, key themes were identified, and codes were grouped within these. Themes were defined and described in the results section.\n\n\nResults\n\nSample\n\nWe initially identified 623 publications from our search. We then removed 15 duplicates. We screened a total of 608 unique references, 313 publications met our inclusion criteria (See Figure 1). Of the retrieved 623 publications, about half were not research outputs. Epidemiological characteristic of included publications is provided in Table 2.\n\nIn about half of the included publications (52%; n = 162), the Neuro based author was the corresponding author on the paper. Neurology 2.9% (n=9), NeuroImage 2.9% (n=9), PLOS One 2.6% (n=8), Brain 2.6% (n=8) and Proceeding of the National academy of science 2.6% (n=8) were the most prevalent journals that Neuro authors published in (see Table 2). The number of authors in the included papers ranged between 1-374 authors. Studies with more than 100 authors were typically a systematic analysis, such as GBD 2016 Traumatic Brain Injury and Spinal Cord Injury Collaborators. Global, regional, and national burden of traumatic brain injury and spinal cord injury, 1990-2016: a systematic analysis for the Global Burden of Disease Study 2016.26 The median number of authors was eight. In addition, we investigated the number of authors with a Neuro affiliation. We found that 31% (n=104) of the publications had a Neuro affiliated author. The number of authors with the Neuro affiliation ranged between 1-12 in the included publications. The median number of the Neuro affiliated authors on a given publication was two.\n\nSeventy percent of the included papers reported on clinical research (n=220), while 18.2% (n=57) reported non-human data, and 11.5% (n=36) used another form of data, such as literature review, protocol, both clinical research and non-human and simulation analyses.\n\nWhen we examined the type of clinical data shared, we found that biological data (e.g., implementation of an antibody) was used in 18.2% (n=55) of publications, DNA in 11.2% (n=34), brain MRI in 32.2% (n=97), non-MRI imaging in 9.6% (n=29), behavioral data in 18.2% (n=55), sequencing data in 4.0% (n=12) and clinical chart in 3% (n=9) of publication. Sleep studies, web-based surveys, stem cells, brain tissue from autopsies, OCT testing, interviews, EEG recordings were used in the “other” 3.6% (n=11) publications.\n\nWe also examined data sharing of non-human research publication. We found that animal data was being used in 91.2% (n=52) publications, five (8.8%) publications used other data (e.g., radiology technology, model). Mice 48.1% (n=25) and rats 34.6% (n=18) were being used in most of the publications. Sixty-seven percent (n=210) of the publications reported quantitative data and 10.9% (n=34) reported qualitative data. Twenty-two percent (n=69) of publications reported both quantitative and qualitative data. Forty percent of publication (n=138) were observational studies.\n\nFor full details of types of data, please see Table 2.\n\nData sharing audit findings\n\nComplete results of our data sharing audit are reported in Table 3; here we report key findings. More than half of the publications (66.5%; n=208) included a data sharing statement. Two-thirds of the data sharing statements were found at the end of the paper in a ‘declaration’ section of the publication (66.0%; n=139/208). Other statements were found in the manuscript methods section 19.7% (n=41), in the manuscript results section 2.9% (n=6), or in other sections 26.0% (n=54) such as the front page, abstract, Dryad, or the Supplementary Materials section of the publication. Of the papers that had a data sharing statement, we were able to download 74.5% (n=155/208) of data from the included publications directly. Of the entire sample of articles included, 49.5%; 155/313) of the papers had openly available data. When examining publications with data sharing statements that did not have openly available data, 58.7% (n=27 of 46) of them specified to contact the author to gain access to the data. Eleven percent (n=5) of the publications identified other ways to access the data (e.g., registration to a site required, request through the Vivli platform which gives access to anonymized individual participant-level data (IPD) or the cleaned raw data that is collected during a clinical trial.27 Thirty percent (n=14) of these publications did not specify a way to gain access to the data.\n\nWhen data was shared openly it was done so on: journal websites (n=120, 66.0%), the Open Science Framework (n=8, 4.4%), Institutional repository (n=17, 9.3%), Figshare (n=7, 3.8%), or using other platforms (n=30, 16.5%). Data was shared mostly in PDF format (n=61, 30.0%), Word format (n=59, 29.0%), and Excel format (n=29, 14.2%). Most of the data 66.0% (n=103) shared was published without a unique identifier, with just 29.0% publications (n=45) providing a Digital Object Identifier (DOI). It was unclear for 2.6% (n=4) publications if a DOI was used.\n\nOpen scripts and materials\n\nWe found that most publications 75.7% (n=237) indicated that there is no analysis script and/or statistical analysis plan (SAP) available. Twenty percent (n=64) of the publications had analysis/SAPs available. Four (1.3%) publications stated that the analysis scripts and statistics data are available upon request. Two percent (n=8) were other (e.g. available from the corresponding author on reasonable request). The analysis scripts were available via a personal or institutional webpages (n=9, 11.5%), a journal webpage (n=16, 20.5%), supplementary information hosted by the journal or a pre-print server (n=16, 20.5%), or an online third party repository (e.g., OSF, Figshare, etc.) (n=26, 33.4%).\n\nWhen publications shared an analysis script, the vast majority, 93.7% (n=60/64), could be downloaded directly. There was only one publication (n = 64; 1.6%) where we could not download the analysis script.\n\nWe defined “Materials availability” as sharing materials used that were to conduct the stud (e.g., such as video of Cognitive Behavior Therapy for psychological interventions; surveys; cell lines; reporting checklist(s), supplementary files, Gene banks). We found that (n=137, 43.8%) of the publications indicated that study materials were available. Less than one percent (n=2) of the publications indicated that the materials were available upon request. Materials were made available via the journal publication webpage (n=51, 30.9%), supplementary information hosted by the journal (n=51, 30.9%), using personal or institutional webpages (n=15, 9.2%), or via an online third-party repository (e.g., OSF, Figshare) fourteen percent (n=23). A small number of articles indicated materials were available by request (n=6, 4.4%). Six presents of materials (n=9) could not be downloaded directly, 92% (n=126) of materials in the publications could be downloaded, and two (1.5%) publications did not use them in their publications. Both publications were review.28,29\n\nAll of these results are included in Table 4.\n\nTransparency and open science practices\n\nResults from our audit of transparency and open science practices are reported in Table 5. Fifteen percent (n=47) of the publications did not mention a conflict-of-interest statement, 15% (n=47) indicated that there were one or more conflicts of interests and 68.8% (n=215) indicated that there was no conflict of interest. In addition, we investigated whether the publications reported funding and, if so, funding sources. Most of the publications (88%; n=296) reported the funding statements. The primary sources of funding were the federal public funder, the Canadian Institutes of Health Research (CIHR; n=149) and the provincial funder, the Fonds de Recherche du Québec – Santé (FRQS; n=71).\n\nMost publications (94.9%; n=297) did not link to an accessible protocol; only 2.9% (n=9) did. 1.6% (n=5) of the publications reported that a protocol was available upon request, and two (0.6%) publications were actual reports of publication protocols.\n\nMost publications (95.8%; n=300) did not include a study registration statement. Only 2.6% (n=8) of the publications indicated registration, 1.6% (n=5) of them indicated that there was no registration. Most registrations 87.5% (n=7) were for clinical trials referring to registration on ClinicalTrials.gov; 12.5% were located on the International Prospective Registry of Systematic Reviews, the PROSPERO database (n=1).\n\nFor studies in which ethics approval was considered approval, 66.6% (n=208) reported this information. Ethics approval was not relevant or not required for 20.4% (n=64) publications. Most publications (96.5%; n=302) did not report using a reporting guideline to improve the completeness and transparency of the completed research, only 3.5% (n=11) publications explicitly mentioned adherence to reporting guidelines.\n\nWe found that 70.0% (n=219) of the articles were open access, while the 29.0% (n=91) were not openly available. Data were missing for 1.0% (n=3) of the journals. Among the studies that were openly available, 37.9% (n=83 of 219) were in ‘gold’ (i.e., the final published version of your article (or Version of Record) is immediately permanently and freely available online) open access journals. Thirteen percent (n=30 of 219) were published in hybrid journals, 20.1% (n=44 of 219) were in ‘bronze’ (i.e., the final published version of your article is free to read on the publisher page but without a creative commons license) journals, and 28.3% (n=62 of 219) were made available green (i.e., when you place a version of the your manuscript into a repository, often after an embargo, making it freely accessible for everyone) open access via a repository (see Table 5).\n\nParticipants\n\nOf the 553 individuals who were emailed the survey, 22.42% (n=124) completed it. We removed 10 participants for analyzing the data as they had answered only demographics question.\n\nDemographic details of these participants are provided in Table 6. Forty-seven (41.1%) participants were female and 66 (57.9%) were male. one participant did not respond to the question concerning gender (0.9%). Most of the participants were between 35 and 44 years of age, 91.4% (n=70 of 114) had a PhD degree, Of the 114 participants who answered the question about their research role, slightly more than a quarter of them 36.8% (n=42) were staff (manager, associate, assistant). Approximately one in three 32.5% (n=37 of 114) were trainees (e.g., MSc, PhD students, postdoctoral fellow).\n\nData sharing practices and Training in data sharing\n\nWe asked participants two questions about their data sharing practices in the past 12 months (see Table 7). Over half of those who responded 61.4% (n=70 of 114) reported that they published a first or last authored paper in the last 12 months. Of these, 70.0% (n=49) indicated that they openly shared research data related to one of their publications.\n\nWe asked one question about past and future engagement in training about data sharing. The results are presented in Table 7. Approximately 40% of the respondents indicated that they had never engaged in training related to data sharing (e.g., online webinars, workshops, or a course) (n=49 of 113). Approximately a third of the respondents (38%; n=43) reported having engaged in some form of training (e.g., online webinars, workshops, or a course) around data sharing within the last 12 months. Also, twenty (17.8%) respondents reported having engaged in training (online webinars, workshops, or a course) around data sharing within the last three years and few respondents (3.5%; n=4) indicated they engaged in training around data sharing more than three years ago.\n\nParticipants indicated a preference for an online training video that they could return to and a series of several modules, each lasting about 10 minutes and a live webinar were less preferred by participants. These results are reported in Table 8.\n\nParticipants showed a preference for an online handbook walking through the practical steps of data sharing. Next, they indicated they would value having access to a data sharing expert (hired by the Neuro) to directly consult with when questions arise about data sharing. The order of the preference for other resources was: an online interactive learning module, an online video walking through the practical steps of data sharing (why, where, how), a central data sharing expert that facilitates data sharing for projects working directly with the project team, a collection of best practice case study examples (see Table 9).\n\nPerceptions about data sharing\n\nParticipants were most familiar with patient privacy considerations when sharing data (Mean=3.14, SD=1.091), the ethical considerations when sharing their data (Mean=3.13, SD=1.001) and practical steps involved to share their data (Mean= 2.91, SD=1.085). Respondents were less familiar with concepts including First Nations Principles of Ownership, Control, Access, and Possession (OCAP)7 (Mean=1.45, SD=1.710) and new metrics to measure data sharing contributions (Mean=1.86, SD=0.872). See Table 10 for full results.\n\nMost of the respondents expressed their opinion that data sharing helps to stimulate new hypotheses from the study (Mean=4.38, SD=0.671), they want to help others to use their study results (Mean=4.34, SD=0.691), data sharing helps the advancement of their research by allowing additional investigators to access the data for future research (Mean=4.33, SD=0.730), that they want to help others to reproduce their study (Mean=4.31, SD=0.767), that they want to help others to transparently assess their study (Mean=4.28, SD=0.720), and that they are optimistic that efforts to adhere to data sharing best practices will help support greater reproducibility and transparency of research (Mean=4.27, SD=0.862). Most of the respondents think that the benefit of data sharing is delayed and uncertain (Mean=2.51, SD=0.989). Also, most respondents disagreed that there is sufficient financial support to help them to adhere to data sharing best practices in the coming year (Mean=2.57, SD=1.016), that they feel stressed out when they think about how to adhere to best practices regarding data sharing for their studies in the coming year. (Mean=2.72, SD=1.014). For full results, see the Extended data.\n\nThematic analysis\n\nFor the item “What incentives do you think the Montreal Neurological Institute-Hospital could introduce to recognize data sharing?” we classified text-based responses into six themes. The themes were: 1) financial support, 2) recognize and incentivize data sharing, 3) provide infrastructure to support data sharing, 4) enforcement of specific and clear data sharing standards, 5) change research and recognition priorities, and 6) provide educational support for data sharing (see Table 11).\n\nFor the question “Is there anything else you want to share about data sharing?” we classified responses into eight themes. The themes were: 1) lack of resources for data sharing, 2) barriers to data sharing, 3) recommendations for standards surrounding data sharing, 4) relevancy of data sharing, 5) provide training for data sharing, 6) privacy considerations when sharing data, 7) technical requirements for data sharing, 8) data sharing requires more resources (see Table 12).\n\n\nDiscussion\n\nThe aim of the first study was to audit all publications produced by Neuro researchers in 2019. We found that 66.5% (n=208) of publications had a data sharing statements and 61% (n=31) of publications specified to contact the author to gain access to the data. Sharing by request creates burden and barriers for other researchers to access and use data and should only be implemented in instances where open data is not possible. Most of the publications indicated that there is no registration statements and protocol availability. We note that the authors are more eager to share their publications material than analysis scripts. Most publications did not report using a reporting guideline to maximize the transparency and completed of their research.\n\nIn the second study, to identify barriers and facilitators to data sharing, we found that there was a preference that training in the form of an online video they could return to, a series of several modules, each lasting about 10 minutes and a single module lasting about 2 hours be developed. A key finding was that more than a third of the respondents (40.7%), had not engaged in training around data sharing. We identified learning gaps in some key areas (e.g., OCAP Principles, metrics to measure data sharing contributions). Respondents noted that barriers they faced when sharing data included: financial support, training, and technical support.\n\nWe recommend an educational and training intervention devoted to data sharing practices to further normalize and support this practice, including training on sharing statistical analysis plans and sharing of other materials related to data sharing. This training could then be followed by additional sessions on further open science practices. Further research is needed to examine the universities’ data sharing practices to track needs and preferences over time and investigate to provide clear data sharing standards for the staff, researchers, and managers.\n\nAlso, it is important to repeat the audit over time to track changes in the Neuro publications and researcher perceptions and barriers and facilitators. Hence, it is important to repeat the audit biennially with the same questions in survey. It is also should be mentioned that in addition to the same questions, additional questions should be added to gain a better understanding about certain data sharing practices in universities and scientific institutes.\n\nThe results of the two studies will provide the Neuro with a better understanding of open access statues and barriers and facilitators to data management and sharing and will identify educational needs related to data sharing that can be reduce barriers to data sharing. We hope this report also provides other organizations wanting to engage their communities about data sharing with a valid approach to the topic and some comparative data. Further, the audit of publications can be used to benchmark for improvements over time and to monitor change. Also, decision-makers in government and universities will be able to structure future open access by providing training for their researchers. Also, this study can serve as a baseline to benchmark for improvements in data sharing and other open science practices and to measure progress over time.\n\nWe acknowledge certain limitations. First, as less than a quarter of the Montreal Neurological Institute-Hospital’s staff completed this survey, the results of the study cannot be generalized. Also, it is hard to measure changes in the community unless two or more surveys are done in different time periods. Hence, we recommend annual surveys of the Neuro community to track changes in needs and preferences over time.\n\nThis study was registered a priori (i.e., before any data collection) using the Open Science Framework (https://osf.io/3tafc).", "appendix": "Data availability\n\nAll study data and materials have been made publicly available.\n\nOSF: Openness and data sharing at The Neuro (Montreal Neurological Institute-Hospital), https://doi.org/10.17605/OSF.IO/3TAFC. 30\n\nThis project (https://osf.io/mx6rp/) contains the following underlying data:\n\n‐ Raw survey data (individual participant survey responses).\n\n‐ Audit data (information extracted from publications).\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nOSF: Supplementary materials and extended data, https://doi.org/10.17605/OSF.IO/3TAFC. 30\n\nThis project (https://osf.io/mx6rp/) contains the following extended data:\n\n‐ Neuro audit supplement (search strategy for audit; survey items).\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nWe thank Alex Amar-Zifkin for helping us to design the search strategy, Dylan Roskams-Edris for providing information on the Montreal Neurological Institute-Hospital staff lists and for facilitating the circulation of the survey among the Montreal Neurological Institute-Hospital researchers and the open science grassroots committee of the Montreal Neurological Institute-Hospital for providing feedback on our extraction form and the survey. The previous version of this article has been published in a preprint server and can be accessed via https://doi.org/10.1101/2022.08.03.22278384.\n\n\nReferences\n\nGold ER. Should Universities Get Out of the Patent Business?: Centre for International Governance Innovation.[cited 2022 Feb 5]. Reference Source\n\nAllen C, Mehler DMA: Open science challenges, benefits and tips in early career and beyond. PLoS Biol. 2019 May 1; 17(5): e3000246. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGovernment of Canada I: Roadmap for Open Science - Science.gc.ca.[cited 2022 Apr 1]. Reference Source\n\nSecond French Plan for Open Science: [cited 2022 Apr 1]. Reference Source\n\nUNESCO Recommendation on Open Science. UNESCO; 2020 [cited 2022 Apr 1]. Reference SourceReference Source\n\nOpen Science: European Commission - European Commission.[cited 2022 Apr 1]. Reference Source\n\nBegley CG, Buchan AM, Dirnagl U: Robust research: Institutions must do their part for reproducibility. 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BMC Public Health. Publisher Full Text\n\nAtkins L, Francis J, Islam R, et al.: A guide to using the Theoretical Domains Framework of behaviour change to investigate implementation problems. Implement. Sci. 2017 Jun 21; 12(1): 77. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOpen Science Ambassadors: The Neuro.[cited 2021 Apr 22]. Reference Source\n\nNowell LS, Norris JM, White DE, et al.: Thematic Analysis: Striving to Meet the Trustworthiness Criteria. Int. J. Qual. Methods. 2017 Dec 1; 16(1): 160940691773384. Publisher Full Text\n\nGBD 2016 Traumatic Brain Injury and Spinal Cord Injury Collaborators: Global, regional, and national burden of traumatic brain injury and spinal cord injury, 1990-2016: a systematic analysis for the Global Burden of Disease Study 2016. Lancet Neurol. 2019 Jan; 18(1): 24–25. Publisher Full Text\n\nFAQs: Vivli.[cited 2021 Oct 27]. Reference Source\n\nLecrux C, Bourourou M, Hamel E: How reliable is cerebral blood flow to map changes in neuronal activity? Auton. Neurosci-Basic Clin. 2019 Mar; 217: 71–79. PubMed Abstract \| Publisher Full Text\n\nSossin WS, Costa-Mattioli M: Translational Control in the Brain in Health and Disease. Cold Spring Harb. Perspect. Biol. 2019 Aug; 11(8): a032912. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEbrahimzadeh S, Cobey KD, Moher D, et al.: Openness and data sharing at The Neuro (Montreal Neurological Institute-Hospital). [dataset]. 2023, June 28. Publisher Full Text" }	[ { "id": "219459", "date": "03 Nov 2023", "name": "Evgeny Bobrov", "expertise": [ "Reviewer Expertise Open science", "open data", "data sharing", "data reuse", "research data management" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIn the first part of the study (‘audit’), the authors present an analysis of 313 research articles from their institution, published in 2019. The authors determined by manual screening whether for these articles data had been shared, as well as the open access status, and the availability of scripts and materials. In the second part of the study (‘survey’), the authors present results from the survey of researchers at their institution. This survey, which was completed by 114 researchers, addressed questions of practices, views, needs, and knowledge surrounding the topic of data sharing. Free-text answers were analyzed by coding according to specific categories.\nI have most closely looked at the data sharing in the audit section of the study, as this is my primary field of expertise. Unfortunately, I see massive issues with the analysis presented in this part of the manuscript. The authors have applied a definition of data sharing which is so liberal that for me it renders the term nearly useless. Supplementary data were always considered “available data”, irrespective of\nthe type of data (which was mostly summary statistics or patient sample properties, and not what would be needed to reproduce the article’s results), the format of the data (which were sometimes PDFs for tables, rendering them extremely difficult to reuse), and the availability status of the data supplement (a considerable amount of articles were not open access articles, and for these, in a majority of cases I checked, the data supplement was also not available without a subscription to the journal).\nI acknowledge that different definitions of data sharing are possible, but deposing a table with summary statistics as a PDF on the publisher website behind a paywall is not an open science practice, and thus data sharing, defined this way, would not be an open science practice either.\nIn addition, the authors multiple times use the terms “open data” and “openly available data”, prominently in the abstract, which makes things further confusing and in my opinion contradictory. The term “data sharing” is so far much less clearly defined and could be justifiably used liberally, as long there is a clear definition applied. However, the aforementioned examples are, to my understanding, not “open data” under any definition.\nWhat is more, the term “data” as such has been used very liberally as well, further eroding the meaning of this screening. E.g., https://github.com/ripolleslab/Diffusion-Tools was considered open data, even though the authors themselves explicitly mentioned that they shared code, and in study Winkler-Schwartz et al., (2019)1, supplemental figures but no “data” in my understanding were shared.\nI also disagree with including “upon request” as open or available data, after the literature has repeatedly shown that such data are not consistently and persistently available. The authors also do not distinguish between own data and data reuse. In the case of article Granados Moreno et al., (2019)2, which was a review and did not generate any data, it seems that the statement “All relevant data are within the paper” was sufficient for it to be considered an example of open data. It might be a simple mistake, but it could also indicate that there was little checking of the actual article.\nIn addition to the analysis as such, the discussion of these observations is extremely short and does not in any way address the discrepancies to the literature. The authors report that approximately 50% of all articles screened had “open data” which could be downloaded. This might be the case applying a certain definition of data sharing, but it would be critical to put this number into perspective, given that others have reported open data numbers far below that (see Hamilton et al., 20233, for a review; in the case of our own dashboard, numbers for 2019 were approximately 5%).\nOverall, for this analysis to be understandable, useful, and relatable to former work, I would recommend that the authors:\nClearly define what is open data for them, aligning it with definitions applied by others Reassess the publications with this definition Discuss the findings in light of other open data screenings\nIf this is not feasible, it seems inevitable to me to extend both the methodological description and the discussion, and to put the screening into perspective, explicitly mentioning aforementioned limitations.\nI have also checked the underlying data table “Copy of Clean-Data-audit-The Neuro-2022-03-3 – Copy” in OSF. The name of the file itself is not well chosen. More importantly, the structure of the table does not comply with good practices of reusable, machine-readable tabular data; in particular, there are:\ncomments in cells information coded by the color of the cell multiple columns of the same name numbers formatted as text missing values are indicated by empty cells\nAlso, the content has not been properly cleaned. Importantly, there are spurious spaces in DOIs, which can thus not be used without further manual correction. There are further issues with data quality, e.g. apparently the same author is present with different writing (S-M Fereshtehnejad and S Fereshtehnejad), and from what I see article Elliott et al., (2019)4 was published in \"Multiple sclerosis journal\", not in \"Model Systems of Synaptic Transmission\". Of course it is time-consuming to check all the automated extractions manually, and it is understandable that such a large table does contain some mistakes, but the fact that from a few spot-checks I found several such issues suggests to me that the authors should have a careful look at the table and try to fix any obvious issues.\nRegarding other parts of the audit, I did look less closely at the underlying data. However, I would recommend to reconsider some parts of the analysis. E.g., I do not think that a categorization of shared data types as presented in table 2 is applicable. In particular, I think that:\n“biological data” would be a super-category, and at least most of the other categories would fit into “biological data” “DNA” and “sequencing” are not clearly distinct “clinical charts” are not data at all in my understanding, if I get this term correctly\nI think it would be worthwhile to reconsider this classification. However, an option could also be to remove the analysis of data types, which I think is not directly related to the open science questions addressed in this manuscript.\nHaving looked in much detail at open data and other aspects of the audit, I could not look at the tables with individual statements and their analysis in full depth. Also, I am not an expert on qualitative data analysis. However, from the experience I have, my impression is that the authors have not organized and labelled the statements in a precise and sufficiently detailed way. E.g.:\nThe statement \"waive open access fees\" does not refer to data, so seems unrelated to the question The statement \"promotions of shared data [...]\" is coded as \"competitions for data sharing innovations\", which I cannot follow It was stated by one researcher that \"advancing research through transparency\" was an incentive in itself, and I think this would have warranted its own category \"for most of us basic scientists [...]\" states that the researcher would feel unfairly treated if data sharing were rewarded, as for him or her it does not apply - the code in contrast states that there should be benefits for data sharing, which I think is not the same \"support for academic promotions and tenure\" is not clear to me, and I cannot follow the coding as \"provide educational support for data sharing\" the title of the table is confusing to me: \"providing infrastructure\" is a facilitator, but it is not an incentive the same is true for embargo periods - this is something which can be offered (by the repository), but I would not say it can be \"provided\"\nSome of these issues might be subjective, and there is surely not one single way of coding. However, I do see some potential issues here as well, and I think it would be very important to receive feedback on this section from an expert on qualitative research.\nApart from issues of methodology and interpretation, the manuscript is in many instances not precise or easy to follow when it comes to results and numbers. E.g.:\nIn the abstract, 208 seems to refer to the 206 cases of \"yes\", as well as one \"other\" and one \"unclear\". However, this is not explicitly stated, and in my opinion indeed there needs to be a clear yes/no decision in the end. The number 155, both in in the abstract and in table 3, is not clear: there are 160 entries for \"can you directly access, download, and open the data?\" with either \"yes\", \"other\" or empty cells, and of these, 3 are empty and 7 are “other”, so I don't know how n = 155 is calculated. For the 64 studies which shared an analysis script, it could be downloaded in 60 cases and could not be in one case, but the three remaining cases are unclear. There was a direct contradiction between “Less than one percent (n=2) of the publications indicated that the materials were available upon request” and “A small number of articles indicated materials were available by request (n=6, 4.4%).” In table 5 “Does the publication report ethics approval?“ n=8 does not fit the data presented.\nSome terminology and categorization was also imprecise or does not seem useful to me. E.g., there is no distinction made between the identifier of the data and the identifier of the article. Also, I think “analysis script” and “statistical analysis plan” should not be pooled. In my understanding, an analysis script serves to reproduce the analysis, but it does not show, how the authors planned the analysis beforehand, and thus does not contribute to reducing bias.\nAs already pointed out, the study does not sufficiently discuss the results in light of the literature. Many highly relevant studies are not referenced, e.g. Milham et al.5 as well as the aforementioned study by Hamilton et al.3. Overall, the introduction could be expanded, and, most importantly, the discussion should be expanded considerably and supplemented with further references to the literature.\nIn addition, I found the way the results were described unfortunately difficult to follow or imprecise in multiple cases. E.g.:\nIt says that 31% of publications had a Neuro affiliation, which cannot be correct – I assume it was meant to say that 31% had ONE Neuro-affiliated author. The description “Top three journals that publications were published in those journals” is probably understandable in its context, but not quite clear in itself and I would recommend rephrasing. The statement “says attached at bottom of article” in table 3 is not clear in this context. Dryad is a repository, so it is not clear to me, how the data sharing statement can be in Dryad. In the statement “[…] statistics data are available upon request” it is not clear what this refers to, as “statistics data” has not been mentioned before. “Supplementary files” is not a type of material on the same level as “reporting checklists” or “surveys”.\nFurther issues I observed while reading are not critical for understanding, but were quite distracting. E.g. Figure 1 was squeezed, and there were multiple typos, as e.g. for ProceedingS of the National Academy of Sciences, a missing bracket, “stud” instead of “study” etc. Of course this is not what a scientific study is about, but I would recommend the final version of the article is also copy-edited for typos.\nAs a recommendation, I think that some results could be presented much more intuitively as bar charts, instead of tables.\nAs there are several issues with the article in this current form, it is difficult to make concrete recommendations for the discussion, except that it should be more detailed and refer better to the literature. In addition, I would say that the discussion should state more clearly, which research is needed in the future, and not just state it generally. Furthermore, I would find it interesting to know, which actions are or at least could be based on the findings. This is only touched upon in a way which I find quite vague. Important limitations are not discussed, including what researchers themselves understand certain open science practices to be, and the fact that publications from 2019 were screened. As it stands, it's late 2023, and while it is perfectly legitimate to publish such findings, it is necessary to acknowledge rapid changes in the field.\nOverall, the authors address an interesting and timely question, and the sample sizes of both articles screened and researchers surveyed would be sufficient to derive interesting insights. However, unfortunately, the analysis suffers from serious methodological shortcomings. I think some of them could still be fixed and need to be addressed. Others might be more difficult to fix, but can be alleviated by a much more thorough discussion of methods and limitations. In addition to the content as such, it is difficult to follow the manuscript in its current form due to numerous imprecisions and unclarities with regard to numbers, categories, and wording. The data underlying the analysis also need checking and editing by the authors, especially given the topic of the article itself, which in my opinion mandates that shared data are also of high quality. For the manuscript to receive the approval of peer review, it thus needs a thorough overhaul, followed by another round of review.\n\nIs the work clearly and accurately presented and does it cite the current literature? No\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "219458", "date": "28 Nov 2023", "name": "Felix Nikolaus Wirth", "expertise": [ "Reviewer Expertise Medical data sharing" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe presented work is an assessment assessing (1) the data sharing and open science practices of the institution of the authors as well as (2) the perceptions of the institution’s staff regarding data sharing and open science.\nMy first point is that the author’s need to define more strictly how they understand some terms such as 'data sharing' and 'open science', as I think the terms are not always used correctly. A clear definition would also important when it comes to the data items chartered: I did a short review of the attached (not well named) excel file “Copy clean data audit file” and found e.g. that the text which qualified as provided data sharing statement (column BG) differed a lot. For instance, the text \"Supplemental Information includes eighteen figures and three tables and can be found with this article online at\" does not appear as a data sharing statement to me. Moreover, I checked a small sample regarding the question whether data can be accessed (column BO). In my three samples1-3. I found information I would perceive rather as supplementary material than free available data. These last two points undermine some of the core claims of the work. Moreover, when I filter column BO to “yes” I see 153 records, while the table in the paper states a number of 155.\nFinally I more extensive discussion of the results might be worthwhile. Two points which are possible to include are: (1) that most “data sharing” (see also comment above) is conducted with word or pdf files, which e.g. would be violation of FAIR principles. Moreover, the work could reflect, that although authors claim to be willing to share their data, in practice this does not happen4.\nMinor points:\nWhy was the data collected for 2019 and not for 2022 or 2023?\n\nI personally do not understand fully the difference between table 8 and table 9.\n\nSome of the tables make the paper cumbersome to read (e.g. tables 12 and 13). Maybe some tables can be put in the appendix?\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1375
https://f1000research.com/articles/12-1374/v1	18 Oct 23	{ "type": "Systematic Review", "title": "Metastasis to the stomach: a systematic review", "authors": [ "Arturan Ibrahimli", "Altay Aliyev", "Aykhan Majidli", "Aysegul Kahraman", "Aysuna Galandarova", "Emil Khalilzade", "Heydar Mammadli", "Kamran Huseynli", "Karam Assaf", "Cagatay Kilinc", "Nijat Muradov", "Omer F. Alisan", "Sabir Abdullayev", "Yeliz I. Sahin", "Elgun Samadov", "Altay Aliyev", "Aykhan Majidli", "Aysegul Kahraman", "Aysuna Galandarova", "Emil Khalilzade", "Heydar Mammadli", "Kamran Huseynli", "Karam Assaf", "Cagatay Kilinc", "Nijat Muradov", "Omer F. Alisan", "Sabir Abdullayev", "Yeliz I. Sahin", "Elgun Samadov" ], "abstract": "Background: This study reviews the literature on gastric metastases (GM) in terms of diagnosis, treatment, and outcomes. The goal of this study was to provide clinicians with a reliable and beneficial source to understand gastric metastases arising from various primary tumors and to present the growing literature in an easily accessible form. Methods: Articles published in English language from implementation of MEDLINE and Cochrane databases until May 2022 were considered for the systematic review. Articles other than English language, letters to the editor, posters, and clinical images were excluded. Hematogenous and lymphogenic metastases were included whereas direct tumoral invasion and seeding were excluded. Articles and abstracts were analyzed and last selection was done after cross-referencing and by use of defined eligibility criteria. Results: In total 1,521 publications were identified and 170 articles were finally included totaling 186 patients with GM. The median age of patients was 62 years. Gynecologic cancer was the most common cancer type causing GM (67 patients), followed by lung cancer (33 patients), renal cancer (20 patients), and melanoma (19 patients). One of the main treatment methods performed for metastasis was resection surgery (n=62), sometimes combined with chemotherapy (ChT) or immunotherapy. ChT was the other most used treatment method (n=78). Also, immunotherapy was amongst the most preferred treatment options after surgery and ChT (n=10). Conclusions: As 172 case reports were screened in the systematic review from different journals, heterogeneity was inevitable. Some articles missed important information such as complete follow-up or clinical information. Moreover, since all of the included articles were case reports quality assessment could not be performed. Among 172 case reports reviewed, resection surgery was performed the most and was sometimes combined with ChT and immunotherapy. Further research about what type of treatment has the best outcomes for patients with gastric metastases is needed.", "keywords": [ "Metastasis to stomach", "gastric metastases", "gastric metastasis", "gastric cancer", "stomach cancer" ], "content": "Introduction\n\nMetastases to the stomach are rare conditions with poor prognosis that may present with both gastrointestinal and systemic symptoms, such as loss of appetite, abdominal pain, fatigue, nausea, and vomiting, with a reported incidence of 0.2-0.7% based on clinical and autopsy findings.1 Gastrectomy is thought to be the only potentially curative treatment for metastatic gastric cancer but the primary site of the tumor is also considered along with the type and grade of the tumor when planning treatment in gastric metastases. Therefore, chemotherapy is also an option for patients with higher grades and multi-focal cancers. This study reviews the literature on gastric metastases in terms of diagnosis, treatment, and outcomes. The intended goal of this study was to provide clinicians with a reliable and beneficial source to understand gastric metastases arising from various primary tumors and to present the growing literature in an easily accessible form by reviewing the case reports of different primary tumors separately with consideration of diagnosis, treatment, and clinical presentation which may vary from patient to patient depending on primary site of the tumor.\n\n\nMethods\n\nThis systematic review adheres to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines.172 A computerized literature search through MEDLINE/PubMed and Cochrane databases was conducted until May 2022.\n\nThe following combination of keywords was used for the search: ({({gastric (MeSH Terms)} AND {neoplasm metastasis (MeSH Terms)}) OR (gastric metastasis)} OR {gastric metastases}) OR (metastasis to the stomach). The search was limited by filtering for “free full text” and “case reports.” After the decision of inclusion and exclusion criteria by the team, two of the reviewers independently screened and retrieved each report.\n\nHematogenous and lymphogenic metastases were included whereas direct tumoral invasion and seeding were excluded from the study. Articles other than the English language, letters to the editor, posters, and clinical images were excluded. After the studies were screened and separated based on the inclusion and exclusion criteria, reviewers were divided to groups based on primary tumor location. Each group contained two reviewers to collect the data from studies of its specific location for example metastasis from gynecologic cancers or lung cancers.\n\nThe following data were extracted from the databases: first author, number of cases, age, sex, site of the primary tumor, histology and treatment of primary tumor, treatment of metastasis, clinical presentation of gastric metastases (GM), synchronous or metachronous GM, the time between primary and secondary GM, diagnostic procedures, other metastasis, and overall survival.\n\nSince the study only contains screening of case reports, assessment of bias risk was not performed and thus it is mentioned as a limitation of study in discussion section.\n\n\nResults\n\nThe PRISMA flow chart below illustrates details about data collection (Figure 1).\n\nIn total, 1,521 publications were identified and 170 articles were finally included totaling 186 patients with GM (101 female and 85 male). The median age of patients was 62 years (IQR: 55-70.5). Gynecologic cancer (including breast cancer) was the most common cancer type causing GM (66 patients), followed by lung cancer (33 patients), renal cancer (20 patients), and melanoma (19 patients) (Figure 2). Results are presented below according to the origin of the primary tumor. The main treatment method performed for metastasis was resection surgery (n=62, total, subtotal or partial gastrectomy, proximal gastrectomy, radical total gastrectomy with Roux-en-Y, wedge gastrectomy, and laparoscopic resection of gastric metastasis), sometimes combined with chemotherapy (ChT) or immunotherapy. Chemotherapy was the other most used treatment method (n=78). Also, immunotherapy was among the most preferred treatment options after surgery and chemotherapy (n=10).\n\nThe median age of the 66 patients was 57 years. In total, 46 cases had metastases other than GM. Bone was the most common site of metastasis. Five cases had no other metastases. The total number of cases in the breast group was 54, and one of them was a male patient. The median age of the breast group is 56, the youngest patient was 36 years old and the oldest patient was 84 years old. Invasive lobular carcinoma (ILC) had the largest number of patients in comparison to ovarian and uterine groups. A total of 31 patients presented with ILC. The ovarian group had nine patients; the median age was 61 years. The oldest patient was 73 years old; the youngest patient was 47 years old. The uterine group had two patients with ages 49 and 80 years. In most cases, systemic therapy was more effective than surgery. Surgical treatment had a role in palliative treatment. As a systemic treatment, chemotherapy was the most utilized treatment. Overall survival was given in only 25 cases and ranged from a few days to nine years. Six of the total patients are still alive. Table 12–56 summarizes the findings of included studies regarding gynecologic cancers.\n\nMedian age of the 16 patients (11 male, five female) was 69 years, ranging from 22 years to 85 years. Overall survival of the seven patients whose data were given ranged from two months to 16 months. Although, there were six cases who were still alive and the survival of three cases was not reported. Among histological types of gastrointestinal cancers, adenocarcinoma (Adeno Ca) was the most common cancer type (seven patients), followed by hepatocellular cancer (HCC) (four patients) and squamous cell carcinoma (two patients). Endoscopy is the most frequently used method in the diagnosis of metastases. Methods such as computer tomography (CT), positron emission tomography and computed tomography (PET-CT), and endoscopic ultrasound were also used for diagnosis. One patient underwent laparotomy and biopsy. According to this research nine of these patients had surgery. Transcatheter left gastric artery embolization was performed in one patient. On the other hand, seven patients received chemotherapy and one patient had palliative radiotherapy. Nevertheless, one patient is unknown. Findings regarding gastrointestinal cancers are summarized in Table 2.57–72\n\nThe median age of the 33 patients (25 male, eight female) was 62, ranging from 39 years to 78 years. Twenty- seven of the total cases had other metastases in addition to gastric ones. The survival time of the 22 patients whose data were given ranged from two weeks to 30 months. Yet, there were two cases that were still alive four and five years after metastases were found, respectively. Among histological types of primary lung cancers that lead to gastric metastases, adenocarcinoma was the most typical diagnosis (13 patients), followed by small cell lung cancer (SCLC) and squamous cell carcinoma (SCC). Regarding the treatment of GM, different combinations of chemotherapy were the most common choice (15 patients). On the other hand, seven of the total cases received surgical treatment (one esophagogastrostomy, two total, and four partial gastrectomies). However, since one patient’s metastasis was diagnosed after an autopsy, he could not receive any gastric treatment. Moreover, one patient refused any metastasis treatment, while six other cases’ treatments are unknown. Data pertaining to GM originating from primary lung cancers are summarized in Table 3.73–102\n\nThe median age of the 19 patients (seven female, 12 male) was 67, ranging from 28 years to 89 years. In 16 patients, other organ metastases were also discovered in addition to malign melanoma. Overall survival was not mentioned in 10 cases. Two of these cases deceased two and four days after hospital admission respectively, and one patient died after a year. Moreover, one of these patients was alive at five years, and another was alive at six months. Overall survival of three cases is three, 27, and four months, respectively. One of these patients refused treatment, and one of them did not receive treatment. However, immunotherapy was applied to six patients, surgery to five patients, radiotherapy to two patients, and only supportive treatment to three patients. In addition, the treatment of GM was not mentioned in three cases. Table 4 summarizes the findings of included studies regarding malign melanoma.103–121\n\nThe median age of 20 patients (11 male and nine female) with kidney cancer was 68.5 years old. A total of 11 patients had metastases other than GM. Overall survival was mentioned only in four cases and ranged from two months to one year. One of the 20 patients did not receive any therapy for GM, whereas 13 patients underwent surgical treatment (four endoscopic mucosal resections, nine gastrectomies), four patients had chemotherapy and one patient was treated with radiotherapy. Regarding prostate cancer, the median age of the affected individuals was 67 years old. Concerning the GM treatment four patients received chemotherapy, one patient underwent mucosal resection, and one patient refused treatment. Overall survival was mentioned for three patients ranging from four months to 19 months. All four patients with testis cancer had other metastases and two of them received chemotherapy. One study included bladder cancer without other metastases and the patient was referred to palliative care. Data pertaining to gastric metastases originating from primary urogenital cancers are summarized in Table 5.71,122–151\n\nThe median age of the four patients with Merkel cell carcinoma was 73 years old. Two patients had other metastases in addition to GM. Three patients underwent surgery, chemotherapy, and radiotherapy, whereas one patient was treated with chemotherapy and radiotherapy. One patient with squamous cell carcinoma had other metastases in addition to GM and received chemotherapy and radiotherapy for the primary tumor.\n\nRegarding bone cancers (n=3) one of the patients was 14 years old and stood out as the youngest patient in this group. Concerning the GM therapy, one of the patients with a known treatment underwent surgery and chemotherapy the other received only surgery. In all patients, GM was discovered metachronous. Three studies were included for soft tissue cancer. All three patients had metastases in addition to GM and underwent different types of GM treatment (including radiotherapy, chemotherapy, excision with snare, and cautery). For the thyroid cancer group, the median age was 71 years old. Overall survival (OS) was only mentioned for one patient (2.5 months). Regarding diffuse large B-cell lymphoma (DLBCL) (n=2), patients received chemotherapy for primary cancer and for GM. GM was discovered synchronously. Kovecsi et al., described the only case of GM from adrenocortical carcinoma of the adrenal gland.152 The patient underwent adrenalectomy for primary and total gastrectomy with splenectomy and end-to-side Roux-en-Y esophagojejunal anastomosis for GM. One patient with choriocarcinoma from retroperitoneum underwent chemotherapy for primary cancer and GM. Table 6 summarizes the findings of included studies regarding gastrointestinal cancers.151–171\n\n\nDiscussion\n\nGastric metastases are uncommon and give information about the progressed stage of malignant disease, with a reported incidence of 0.2-0.7% based on clinical and autopsy findings.1 Furthermore, metastasis to the stomach frequently indicates short survival. These metastases are observed rarely due to clinical problems regarding their diagnosis and treatment.2 Progressively, with improvements in prognosis for cancer patients, metastatic tumors in the stomach are being detected more frequently.1 There are several symptoms of gastric metastases, such as abdominal pain, diarrhea, nausea, vomiting, weight loss, and dyspepsia. The most preferred treatment method for gastric metastasis is surgical resection of the tumor. Also, chemotherapy is the most applied alternative option.\n\nThis systematic review has a few potential limitations that need to be mentioned. As 172 case reports were screened in the systematic review from different journals the heterogeneity was inevitable. Some articles missed important information such as complete follow-up or clinical information. Moreover, since all of the included articles were case reports, quality or bias assessment could not be performed.\n\nGastric metastasis mainly occurs due to breast cancer. Both ovarian and uterine metastases are distinctly less frequent.38 Invasive lobular carcinoma is the type with the highest affinity to the digestive system with an incidence of 4.5% compared to 0.2% in ductal carcinoma.26 Breast cancer metastases to the gastrointestinal tract are rare, with a median time interval from the diagnosis of the primary tumor to metastasis up to seven years.21 The longest disease-free interval is 22 years after the initial diagnosis 17 of 24.10 Some metastatic tumors may have a similar presentation as primary gastric cancer.38 The detailed immunohistochemical analysis will allow the most accurate diagnosis to differentiate between primary gastric cancer and gastric metastasis from breast cancer.26 Most gastric metastatic breast cancers are estrogen receptor (ER)-positive, progesterone receptor (PR)-positive/negative, and human epidermal growth factor receptor (HER2)-negative. However, in primary gastric adenocarcinoma, ER and PR can be positively expressed in 20-28% of patients.19 In a few cases, metastatic breast cancer is negative for ER and PR, so a diagnosis cannot be made based on these two investigations alone.59 ER and PR can be used as markers; however, they are not always suitable diagnostic markers to confirm if a tumor has originated.11 Treatment of gastrointestinal metastases from breast cancer is discussed frequently in the literature. Systemic therapy is the first option.36 The effective rate of systemic treatment is about 46%.60 Surgical treatment may have a role in palliative treatment.34 Surgical treatment is considered in cases with obstruction or bleeding.36 Metastasis to the gastrointestinal tract can be the first presentation of breast cancer, therefore it is imperative to consider the possibility of breast cancer metastasis to the gastrointestinal metastasis.26,46\n\nAmong cancers that metastasize to the stomach, gastrointestinal system cancers are encountered in a minority. Gastric metastases gave unspecific findings, such as anemia, bleeding, and pain. Pancreas, liver, and colon account for the majority of primary cancers. Nine of the cases had other metastases in addition to gastric ones. Since pancreatic cancers are usually caught at an advanced stage, the chances of surgical treatment and their response to treatment are low. We see that three of five patients died within a year.2,6,8 Among these cases, the prognosis of pancreatic head cancers was worse than body and tail cancers.\n\nIn fact, lung cancer is the most mortal type of all cancers. However, the stomach is not a common site for primary lung cancers’ metastases, especially compared with brain, liver, adrenal glands, and bones.74 Yet, the expected lifespan after diagnosis of metastasis is found to be relatively low. The median survival time was four months (average 6.8 months) among 16 cases who died. On the other hand, data showed that endoscopy is the gold standard in diagnosis. In addition, pathology and immunohistochemistry are considered important factors to differentiate gastric metastases from primary cancers.81 Regarding the treatment of gastric metastases of the pulmonary origin, although non-invasive chemotherapy treatments were the most common choice, patients who received surgery, particularly partial gastrectomy, but also esophagogastrostomy and laparotomy, tended to have relatively much longer survival time.77,90,95 However, this conclusion is not definitive, since in some cases surgeries may be avoided when the patient’s condition is extremely severe and the number of cases with given surgical treatments is scarce. So the potential benefit of surgeries to the expected lifespan of the patients needs further investigation.\n\nAlthough melanoma accounts for only 5% of cutaneous malignancies, it makes up nearly 75% of skin cancer- related deaths.103,107 Malignant melanoma ranks as the most common metastatic tumor of the gastrointestinal (GI) tract.103,110,120 It takes an average of 52 months for a primary cutaneous melanoma to spread to the gastrointestinal tract.107,110 Only 1-4% of patients with malignant melanoma deceased before gastrointestinal metastases are diagnosed. On the other hand, GI tract metastasis was observed in more than 60% of melanoma patients by autopsy.103,110,111,114,121 The most commonly involved sites include the small and large bowels and rectum; however, gastric metastasis is a rare case110,111,117,119,121 due to the non-specificity of its symptoms, such as epigastric discomfort, nausea, vomiting, weight loss, hematemesis, and melena.103,107,110,111,114,117,121 The average survival is four to six months.103,107,121 Endoscopy is an effective method for detecting melanoma metastases due to pigmentation, which can then be confirmed by histology and immunohistochemistry.114,121 Treatment options include surgical resection, immunotherapy, chemotherapy, and targeted therapy. If a patient is symptomatic, surgical excision can be a palliative technique that can also prolong survival.103,121\n\nRegarding urogenital metastases GM is uncommon and the incidence is reported to vary between 0.2% and 0.7%.123 The most common clinical presentations are gastrointestinal bleeding (melena and hematemesis), anemia, and malaise. Whereas two patients had no symptoms associated with the gastrointestinal system.131,140 Esophagogastroduodenoscopy is often necessary for diagnosis and localized treatment.131 The presence of gastric metastases is considered an important indicator of advanced disease.149 Treatment options varied depending on the stage of the metastasis including endoscopic resection, partial or total gastrectomy, chemotherapy, and palliative care. Even though overall survival seems to be longer in patients who underwent surgery, the main reason for this may be that these patients have early-stage diseases suitable for surgery. Therefore, treatment options should be decided upon the stage of the disease and the general well-being of the patient.\n\nThe most common symptoms, in terms of frequency, are melena, abdominal pain, vomiting, weight loss, anemia, fatigue, and loss of appetite. Gastrointestinal endoscopy plays an important role in the diagnosis of GM if suspected.162 Tumor seeding after endoscopic gastrostomy tube replacement was observed in two cases.161,169 Even though surgery is the frequent treatment for solid organ cancer metastasis, chemotherapy is the chosen treatment for DLBCL, skin cancer, and sarcoma. Overall survival was only mentioned for four cases; therefore, it is difficult to comment on which treatment method is more beneficial. Metastasis to the stomach is not reported frequently. Thus, determining the prognosis and planning the treatment based on scientific evidence seems to be problematic for clinicians.\n\nIn conclusion, among 172 case reports reviewed, resection surgery was performed the most for treatment and was sometimes combined with chemotherapy and immunotherapy. However, the literature regarding the management of patients with secondary gastric cancer is limited. Therefore, further multi-centric research to reach a consensus about what type of treatment has the best outcomes for patients with gastric metastases is needed.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\nOSF: Tables for ‘Metastasis to the stomach: a systematic review’. https://doi.org/10.17605/OSF.IO/Y4QD5. 172\n\nOSF: PRISMA checklist for ‘Metastasis to the stomach: a systematic review’. https://doi.org/10.17605/OSF.IO/Y4QD5. 172\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nAcknowledgements\n\nThe abstract of this paper was presented in 41st congress of the European Society of Surgical Oncology and was published by the journal of the same society (EJSO) on February 2023 (https://doi.org/10.1016/j.ejso.2022.11.492).\n\n\nReferences\n\nNamikawa T, Munekage E, Ogawa M, et al.: Clinical presentation and treatment of gastric metastasis from other malignancies of solid organs. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTiwari P, Tiwari A, Vijay M, et al.: Upper gastro-intestinal bleeding - rare presentation of renal cell carcinoma. Urol Ann. 2010; 2: 127–129. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYodonawa S, Ogawa I, Yoshida S, et al.: Gastric metastasis from a primary renal leiomyosarcoma. Case Rep Gastroenterol. 2012; 6: 314–318. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChibbar R, Bacani J, Zepeda-Gómez S: Endoscopic mucosal resection of a large gastric metastasis from renal cell carcinoma. ACG Case Rep J. 2013; 1: 10–12. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSakurai K, Muguruma K, Yamazoe S, et al.: Gastric metastasis from renal cell carcinoma with gastrointestinal bleeding: a case report and review of the literature. Int Surg. 2014; 99: 86–90. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPatel H, Kumar A, Shaaban H, et al.: Synchronous metastasis of prostate adenocarcinoma to the stomach and colon: a case report. N Am J Med Sci. 2014; 6: 152–154. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSharifi Doloui D, Fakharian T, Yahyavi V, et al.: Primitive neuroectodermal tumor with kidney involvement: a case report. Iran J Radiol. 2014; 11: e4661. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGreenwald D, Aljahdli E, Nepomnayshy D, et al.: Synchronous gastric metastasis of renal cell carcinoma with absence of gastrointestinal symptoms. ACG Case Rep J. 2014; 1: 196–198. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCosta TN, Takeda FR, Ribeiro U Jr, et al.: Palliative laparoscopic resection of renal cell carcinoma metastatic to the stomach: report of a case. World J Surg Oncol. 2014; 12. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSoe AM, Bordia S, Xiao PQ, et al.: A rare presentation of metastasis of prostate adenocarcinoma to the stomach and rectum. J Gastric Cancer. 2014; 14: 271–274. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBhandari V, Pant S: Carcinoma prostate with gastric metastasis: a rare case report. J Cancer Res Ther. 2015; 11: 659. Publisher Full Text\n\nLowe K, Paterson J, Armstrong S, et al.: Metastatic testicular choriocarcinoma: a rare cause of upper gi bleeding. ACG Case Rep J. 2015; 3: 36–38. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nInagaki C, Suzuki T, Kitagawa Y, et al.: A case report of prostate cancer metastasis to the stomach resembling undifferentiated-type early gastric cancer. BMC Gastroenterol. 2017; 17: 10–93. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTavukcu HH, Aytac O, Aktepe F, et al.: Ductal adenocarcinoma of the prostate with a rare clinical presentation; late gastric metastasis. Urol Case Rep. 2016; 7: 28–30. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKovecsi A, Jung I, Bara T, et al.: First case report of a sporadic adrenocortical carcinoma with gastric metastasis and a synchronous gastrointestinal stromal tumor of the stomach. Medicine (Baltimore). 2015; 94: e1549. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKoti KA, Backianathan S, Sebastian P, et al.: A rare case of gastric metastasis in ewing’s sarcoma of the femur. Case Rep Oncol Med. 2019; 2019. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDodis LB, Bennett MW, Carr-Locke DL: Ewing’s sarcoma metastasis to the gastric wall in a 72-year-old patient. MedGenMed. 2006; 8. PubMed Abstract\n\nUrakawa H, Tsukushi S, Tsurudome I, et al.: Metastasis of osteosarcoma to stomach made clinically evident by hematemesis: a case report. World J Surg Oncol. 2013; 11. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nShibuya T, Osada T, Kodani T, et al.: Gastrointestinal hemorrhage as the first manifestation of metastatic extragonadal choriocarcinoma. Intern Med. 2009; 48: 551–554. PubMed Abstract \| Publisher Full Text\n\nTarangelo NP, Kistler CA, Daitch Z, et al.: Synchronous gastric and duodenal metastases from head and neck squamous cell carcinoma: a unique presentation of upper gastrointestinal bleeding. Ann Gastroenterol. 2018; 31: 381–383. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKamihara Y, Murai S, Kikuchi S, et al.: Tumor-to-tumor metastasis of diffuse large B cell lymphoma to gastric adenocarcinoma via CXCL12 (SDF-1)/CXCR4 axis: a case report. BMC Gastroenterol. 2021; 21: 270. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZepeda-Gomez S, Camacho J, Oviedo-Cardenas E, et al.: Gastric infiltration of diffuse large B- cell lymphoma: endoscopic diagnosis and improvement of lesions after chemotherapy. World J Gastroenterol. 2008; 14: 4407–4409. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTeh JL, Wong RK, Gowans M, et al.: Gastric metastases of oral carcinoma resulting from percutaneous endoscopic gastrostomy placement via the introducer technique. Gastroenterol Rep (Oxf). 2013; 1: 211–213. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nElkafrawy A, Numan L, Albawaliz A, et al.: A rare case of metastatic merkel cell carcinoma to the stomach and pancreas presenting with upper gastrointestinal bleeding and obstructive jaundice. ACG Case Rep J. 2021; 8: e00523. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHa JY, Park SE, Kim HS, et al.: A case report of recurrent Merkel cell carcinoma with synchronous metastases to the heart and stomach. Medicine (Baltimore). 2018; 97: e13032. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIdowu MO, Contos M, Gill S, et al.: Merkel cell carcinoma: a report of gastrointestinal metastasis and review of the literature. Arch Pathol Lab Med. 2003; 127: 367–369. Publisher Full Text\n\nParikh MP, Samo S, Ganipisetti V, et al.: Gastric metastasis of Merkel cell carcinoma, a rare cause of gastrointestinal bleeding: case report and review of the literature. J Gastrointest Oncol. 2014; 5: E68–E72. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSubramanian S, Kumar M, Thulkar S, et al.: Bowel metastases from primary leiomyosarcoma of the gluteal region. Singap Med J. 2008; 49: 68–70.\n\nDent LL, Cardona CY, Buchholz MC, et al.: Soft tissue sarcoma with metastasis to the stomach: a case report. World J Gastroenterol. 2010; 16: 5130–5134. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSamuel T, Norly S, Ros’aini P: Gastric ulcer that turned out to be metastasis of a synovial sarcoma: a case report and literature review. Med J Malaysia. 2016; 71: 363–365. PubMed Abstract\n\nThorburn D, Karim SN, Soutar DS, et al.: Tumour seeding following percutaneous endoscopic gastrostomy placement in head and neck cancer. Postgrad Med J. 1997; 73: 430–432. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFuladi R, Nagarkar R, Roy S: Metastasis to stomach in a patient with anaplastic thyroid carcinoma: a clinical challenge. Am J Case Rep. 2019; 20: 134–138. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyaz T, Sahin SB, Sahin OZ, et al.: Anaplastic thyroid carcinoma presenting with gastric metastasis: a case report. Hippokratia. 2015; 19: 85–87. PubMed Abstract\n\nKarrasch T, Doppl W, Roller FC, et al.: Unusual gastric mucosal infiltration by a medullary thyroid carcinoma: a case report. J Med Case Rep. 2016; 10: 208–210. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIbrahimli A: Metastasis to the Stomach: A Systematic Review. [Dataset]. OSF. 2023. Publisher Full Text" }	[ { "id": "232517", "date": "02 Jan 2024", "name": "Murat Sarı", "expertise": [ "Reviewer Expertise Medical oncology" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe stomach is a rare organ in terms of metastasis sites. It would be valuable to write a review evaluating this issue. The review is clearly the result of extensive research. Figures and tables are adequate and informative. Therefore, acceptance of this article will provide impact and citations to the journal.\n\nAre the rationale for, and objectives of, the Systematic Review clearly stated? Yes\n\nAre sufficient details of the methods and analysis provided to allow replication by others? Yes\n\nIs the statistical analysis and its interpretation appropriate? Yes\n\nAre the conclusions drawn adequately supported by the results presented in the review? Yes", "responses": [] }, { "id": "261964", "date": "02 May 2024", "name": "Fabio Grizzi", "expertise": [ "Reviewer Expertise My primary area of research lies in investigating cancers originating from unrelated histologies as intricate systems. I am particularly driven by the pursuit of novel prognostic", "predictive", "and therapeutic biomarkers." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nI found Ibrahimii et al.'s manuscript on systematically reviewing stomach metastasis quite compelling. Their study stands out for its clarity and significance within its research domain. While I have some minor suggestions for the authors, such as delving deeper into the heterogeneity noted across the included studies and exploring its implications for managing gastric metastasis, I also recommend addressing the variations in terminology among studies, particularly in distinguishing between the term “studies” and “case reports”. This attention to detail will enhance the coherence and applicability of their findings, advancing our understanding of the underlying biology of this neoplastic process.\n\nAre the rationale for, and objectives of, the Systematic Review clearly stated? Yes\n\nAre sufficient details of the methods and analysis provided to allow replication by others? Yes\n\nIs the statistical analysis and its interpretation appropriate? Yes\n\nAre the conclusions drawn adequately supported by the results presented in the review? Yes\n\nIf this is a Living Systematic Review, is the ‘living’ method appropriate and is the search schedule clearly defined and justified? (‘Living Systematic Review’ or a variation of this term should be included in the title.) Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1374
https://f1000research.com/articles/12-1372/v1	18 Oct 23	{ "type": "Case Report", "title": "Case Report: Whole lung lavage therapy for pulmonary alveolar proteinosis: a double-edged sword", "authors": [ "Nouha Guediri", "Chirine Moussa", "Emna Ben Jemia", "Houda Rouis", "Hamida Kwas", "Ibtihel Khouaja", "Habib Ghedira", "Sonia Maalej", "Chirine Moussa", "Emna Ben Jemia", "Houda Rouis", "Hamida Kwas", "Ibtihel Khouaja", "Habib Ghedira", "Sonia Maalej" ], "abstract": "Pulmonary alveolar proteinosis (PAP) is an uncommon disease characterized by the excessive accumulation of lipoprotein material in the alveoli. No uniform treatment existed. However, whole lung lavage (WLL) remains the main treatment. We report the case of a 33-year-old non-smoking man, with no reported history of medical problems, who complained of dyspnea on exertion and cough for over 5 years. A CT scan revealed diffuse pulmonary interstitial infiltrates. Bronchoalveolar lavage was milky. WLL was planned, and the procedure was repeated 70 times and a total of 3,5 liters of saline serum was instilled and 2,8 liters of milky liquid was removed. The operation procedure was interrupted because of acute pulmonary edema. The patient was transferred to the intensive care unit for acute respiratory distress syndrome. He received 3 days of noninvasive ventilation and regression of supplemental oxygen. Faced with the seriousness of the complication it was decided not to perform a third wash and to monitor the patient functionally and radiologically. Currently, after one year of the lung wash the patient is stable functionally and radiologically. We presented a rare complication of a WLL in a patient with PAP. Physicians should evaluate the patient carefully before suggesting WLL, which is an invasive procedure requiring general anesthesia for a pathology that may evolve spontaneously favorably.", "keywords": [ "Alveolar proteinosis", "Whole lung lavage", "Treatment", "Prognosis" ], "content": "Introduction\n\nPulmonary alveolar proteinosis (PAP) is an uncommon disorder in which excessive lipoproteins are deposited in the alveoli. Respiratory failure is caused by an increased respiratory effort and disturbance of air circulation.1 In 1958, SH Rosen, Benjamin Castleman, and AA Liebow identified PAP, also known as the Rosen-Castleman-Liebow triad syndrome.2\n\nCases of PAP are classified into the following three categories. Idiopathic PAP is the most common type of PAP, representing more than 90% of patients, and includes autoimmune PAP, which is determined by the existence of circulating auto-antibodies against granulocyte-macrophage colony stimulating factor (anti-GM-CSF antibodies). Secondary PAP results from high-dust levels (such as silica) or disease or underlying malignancy, and congenital PAP is caused by defects in surfactant production.1\n\nChest imaging can suggest PAP. However, bronchoalveolar lavage remains the keystone for the diagnosis of PAP, which gives a milky-looking bronchoalveolar reaction indicating extracellular proteinaceous material which leads to the accumulation of amorphous, periodic acid-Schiff (PAS)-positive lipoproteinaceous material in the distal air spaces. PAP is now rarely confirmed by surgical lung biopsy.2 No uniform treatment exists. However, whole lung lavage (WLL) remains the standard therapy of care as it provides long-lasting results in most patients. Inhaled GM-CSF remains as an alternative therapy of choice.3\n\nWLL is considered a safe technique and can provide most patients with durable recovery. However, in some cases of PAP, WLL may be potentially damaging and dangerous with poor results.4\n\nHere we report a rare case of a patient with pulmonary alveolar proteinosis treated with WLL which led to a clinical complication requiring intubation and hospitalization in the intensive care unit.\n\n\nCase report\n\nA 33-year-old male patient, policeman, non-smoker, with no reported history of medical problems, complained of dyspnea on exertion and cough for over 5 years for which he had never consulted a doctor before and had never had any treatment. Three months before hospitalization, he reported progressive worsening of dyspnea. An initial physical exam revealed an acute respiratory distress syndrome (polypnea, PaO2=51mmHg) and bibasal crackles. Chest x-ray showed alveoli-interstitial syndrome. CT scan revealed diffuse pulmonary interstitial infiltrates without any particular diagnostic orientation (Figure 1). Bronchoalveolar lavage was milky with cytology of extracellular proteinaceous material with positive periodic acid-Schiff (PAS)-positive. After the stabilization of the respiratory condition and the clear consent of the patient, an abundant lung wash was programmed in the operating room. The procedure began with the administration of curare and sedation and endotracheal intubation with a simple tube. With a bronchial endoscopy placed in the right bronchial tree, we instilled 50mL of saline serum and removed the milky liquid. The procedure was repeated 70 times and a total of 3,5 liters of saline serum was instilled and 2,8 liters of milky liquid was removed. The operation procedure was interrupted because of acute pulmonary edema. The patient was transferred to the intensive care unit. The diagnosis of acute respiratory failure and pneumonia was retained upon admission. After 3 days, under invasive ventilation (FiO2=100%), the patient’s respiratory state worsened (PaO2/FiO2=76, pH=7,30, PaO2=76mmHg, PaCO2=59mmHg). Thus, a second abundant lung wash was performed under general anesthetic with the instillation of 1,5 liters of saline serum and 500mL of milky liquid was removed, with the patient put in a ventral decubitus position. As a result, the evolution was favorable (FiO2=45%, PaO2/FiO2=240) and extubation was possible on day 7 of hospitalization. He received 3 days of noninvasive ventilation and regression of supplemental oxygen. On the 10th day, the patient was eupneic, the physical exam was normal, and SpO2 was 97% in the ambient air.\n\nFaced with the seriousness of the complication, it was decided not to perform a third wash and to monitor the patient functionally and radiologically.\n\nFive years after the lung wash the patient was stable functionally, assessed via spirometry and radiologically with CT scan control (Figure 2).\n\n\nDiscussion\n\nWhole lung lavage, first reported by Ramirez Rivera, is a well-established approach in the therapy of PAP.5 Nevertheless, there is some debate about the appropriate time to carry out a WLL procedure in patients with PAP. This issue is especially challenging given that some spontaneous remission of PAP has been reported.6 The choice of when to carry out a WLL is still a clinical judgment, although the usual criteria for considering a WLL are the presence of significant respiratory restriction or oxygen desaturation with or without effort.\n\nWLL is an invasive therapy that should be done in the operating room with general anesthesia. The anesthesia and lavage operation risks make it necessary to separate the lungs under general anesthesia and lavage the non-ventilated lung. Patients are intubated with a double-lumen selective intubation tube. One channel of the tube is used for ventilating and the other channel is used for lavage. The supine patient is sedated and curarized and 1-2 L of saline is instilled at 37C. The lavage fluid is discharged by gravity (‘siphoning’).7\n\nAfter washing lung with 15-20 liters of saline water or even more, the effluent fluid becomes clear and the procedure can be stopped.8,9\n\nIn our case, the diagnosis of pulmonary edema induced by the lavage fluid was retained. After the WLL, local hypoxia sets in due to retained fluid which leads to additional edema as a result of activation of inflammatory cytokines and damage to capillaries and alveoli in the corresponding lung fields, as seen in stage 3 of near-drowning pulmonary edema.10\n\nThe use of a continuous positive airway pressure (CPAP) valve with a pressure limit of 5 ~ 10 mmHg can increase the removal of accumulated material, as has been reported in previous research.11,12\n\nIn a survey by Campo et al. that includes 30 centres, an estimated 1,110 WLL procedures were performed. The most common reported complications were fever in 18% followed by hypoxis (14%) pneumonia (5%) and fluid leakage (4%).13\n\nThe decision to perform a WLL remains a clinical decision, although the usual criteria for considering a WLL are the presence of substantial respiratory limitation or oxygen desaturation with or without exercise.3\n\nThe particularity of our case report is the favorable evolution of our patient in spite of an interrupted lavage. The lavage remains the treatment of preference but exposes to a risk of complication which can put in stake the vital prognosis.\n\n\nConclusion\n\nWe reported an acute respiratory distress syndrome caused after a whole lung lavage in a patient with PAP. This uncommon but severe complication allows us to reconsider WLL for each PAP. Considering the evident treatment differences, clinicians must evaluate the patient carefully before suggesting WLL, which is an invasive procedure requiring general anesthesia for a pathology that may evolve spontaneously favorably.\n\nFrom this case we conclude that:\n\n‐ WLL remains the gold standard treatment in pulmonary alveolar proteinosis\n\n‐ An observational time before conducting the WLL must be considered\n\n‐ Indication of WLL should be made on a case-by-case basis to measure the pros and cons.\n\n\nConsent\n\nThe authors declare that appropriate written informed consent was obtained for the publication of this manuscript. The patient consented to the publication of their clinical images.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\n\nAcknowledgements\n\nWe would like to express our sincere gratitude to Dr. Ines Zendah for her invaluable contributions to the patient’s care. Her dedication played a crucial role in the successful management of this case. We are truly grateful for her exceptional commitment and tireless efforts.\n\n\nReferences\n\nKumar A, Abdelmalak B, Inoue Y, et al.: Pulmonary alveolar proteinosis in adults: pathophysiology and clinical approach. Lancet Respir. Med. juill 2018; 6(7): 554–565. PubMed Abstract \| Publisher Full Text\n\nJouneau S, Ménard C, Lederlin M: Pulmonary alveolar proteinosis. Respirol. Carlton Vic. août 2020; 25(8): 816–826. Publisher Full Text\n\nAbdelmalak BB, Khanna AK, Culver DA, et al.: Therapeutic Whole-Lung Lavage for Pulmonary Alveolar Proteinosis: A Procedural Update. J. Bronchol. Interv. Pulmonol. juill 2015; 22(3): 251–258. Publisher Full Text\n\nAwab A, Khan MS, Youness HA: Whole lung lavage—technical details, challenges and management of complications. J. Thorac. Dis. 2017 Jun; 9(6): 1697–1706. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRamirez-Rivera J: The strange beginnings of diagnostic and therapeutic bronchoalveolar lavage. P. R. Health Sci. J. 1992; 11: 27–32. PubMed Abstract\n\nBonella F, Bauer PC, Griese M, et al.: Pulmonary alveolar proteinosis: new insights from a single-center cohort of 70 patients. Respir. Med. déc 2011; 105(12): 1908–1916. PubMed Abstract \| Publisher Full Text\n\nJouneau S, Kerjouan M, Briens E, et al.: La protéinose alvéolaire pulmonaire. Rev. Mal. Respir. déc 2014; 31(10): 975–991. PubMed Abstract \| Publisher Full Text\n\nCampo I, Luisetti M, Griese M, et al.: A Global Survey on Whole Lung Lavage in Pulmonary Alveolar Proteinosis. Chest. juill 2016; 150(1): 251–253. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEfficacy of Whole-Lung Lavage in Pulmonary Alveolar Proteinosis: A Multicenter International Study of GELF - FullText - Respiration 2017. Karger Publishers; [cité 9 févr 2023]; Vol. 93(3). Reference Source\n\nGluecker T, Capasso P, Schnyder P, et al.: Clinical and radiologic features of pulmonary edema. Radiogr. Rev. Publ. Radiol. Soc. N. Am. Inc. 1999; 19(6): 1507–1531. discussion 1532-153. Publisher Full Text\n\nSilva A, Moreto A, Pinho C, et al.: Bilateral whole lung lavage in pulmonary alveolar proteinosis--a retrospective study. Rev. Port. Pneumol. 2014; 20(5): 254–259. PubMed Abstract \| Publisher Full Text\n\nRebelo HM, Guedes L, Veiga D, et al.: Anaesthetic, procedure and complications management of serial whole-lung lavage in an obese patient with pulmonary alveolar proteinosis: case report. Rev. Bras. Anestesiol. 2012; 62(6): 869–877. PubMed Abstract \| Publisher Full Text\n\nCampo I, Luisetti M, Griese M, et al.: Whole lung lavage therapy for pulmonary alveolar proteinosis: a global survey of current practices and procedures. Orphanet J. Rare Dis. 31 août 2016; 11(1): 115. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "252250", "date": "16 Apr 2024", "name": "Effrosyni D Manali", "expertise": [ "Reviewer Expertise PAP", "ILD" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIn the manuscript by Nouha Guediri and co-workers, the authors describe a 33-year-old patient with pulmonary alveolar proteinosis who was managed with WLL due to the severity of the disease and developed the complication of pulmonary oedema. They conclude that WLL remains the gold standard for PAP therapy but necessitates a meticulous evaluation for each patient before its performance. The authors could ameliorate their work by addressing the following comments\nWas anti GMCSF antibodies measurement performed Had the patient any other co-morbidity that could increase the risk of complications? Could the authors report the results of microbiology analysis from BAL, including the examination for Pneumcystis jirovecii? The CT images of Figure 1 and Figure 2 are not comparable because they do not correspond to the same level and the quality of the second image is not good enough to allow evaluation Did the authors have any additional data regarding response to treatment 1 year after the WLL? Based on the complication that they are describing, it is a good opportunity to comment on the shift from WLL to inhaled GMCSF for the treatment of PAP in case it is an autoimmune case. The authors could add some references to support this option like [Refer Ref 1,2] The WLL was rather limited however the authors report that it was effective, could they comment on micro-WLL please? Could the authors provide some explanation about the significant deterioration while the patient was intubated? How did they decide to perform a second WLL? Where other reasons of deterioration excluded? Was a cardiology evaluation performed?\n\nIs the background of the case’s history and progression described in sufficient detail? Partly\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Partly\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Partly\n\nIs the case presented with sufficient detail to be useful for other practitioners? Partly", "responses": [] }, { "id": "241772", "date": "29 Aug 2024", "name": "Brenna C Carey", "expertise": [ "Reviewer Expertise Rare lung diseases", "Pulmonary Alveolar Proteinosis" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe manuscript entitled “Case Report: Whole lung lavage therapy for pulmonary alveolar proteinosis: a double-edged sword” is a case report description about an individual diagnosed with PAP by CT scan and bronchoalveolar lavage. The report does not identify the specific type of PAP lung disease. The report describes the lung lavage procedure as repeated instillations of 50 mls of serum saline 70 times for a total of 3.5 liters with recovery of 2.8 liters. This lavage was followed by a second procedure with the instillation of 1.5 liters and recovery of 500 ml. The team chose not to do a third wash and have concluded that the patient is doing well after the procedure.\n\nMajor Concerns\nThe procedure used for lung lavage is not that which is practiced anywhere in the world known to this reviewer, who is quite familiar with both WLL and segmental lung lavage. The use of infusion aliquots of 50 ml is not only standard of practice anywhere, but also not expected to be at all efficient and subjects the patient to unnecessary repeated low-efficacy procedures, which would be expected to worsen the risk to benefit ratio. Further, a total lavage volume of 3.5 liters is far below the range of values expected to be effective for efficient lavage of a single lung (20 – 30 liters, and sometimes up to 50 liters). The authors can add more information about the topic in the manuscript of PAP syndrome, individual PAP-causing diseases, or the indications for or performance of WLL. Details in the abstract and the text of the paper are not consistent. The authors do not provide sufficient clinical or biochemical data to support the conclusions reached. For example, while they claim the patient developed ARDS, they do not provide any information adequate to justify this diagnosis. Do you have data/information from the clinical lab for the evaluation of the fluid recovered for cell counts, PAS+ material, number or percent of oil-red-O positive cells, infection, etc? If so, please include in the paper. The actual PAP-causing disease was apparently not determined (or data not presented). However, the authors discuss it as if the PAP-causing disease was autoimmune PAP caused by a high concentration of circulating GM-CSF autoantibodies. This data should be considered to be essential in reports published in 2024.\nModerate concerns:\nThe data quality is poor. Specifically, the CT images are poor. In the opinion of this review, they are best presented in grayscale rather that colour. The description of the radiographic findings is poor. The abstract describes the presence of “diffuse pulmonary interstitial infiltrates”. PAP of various etiologies is often (and in this reviewer’s opinion, more correctly) described as diffuse, geographic ground glass opacification with superimposed interlobular thickening” or, more simply, “crazy paving”. Overall, the manuscript is poorly written.\n\nIs the background of the case’s history and progression described in sufficient detail? No\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? No\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? No\n\nIs the case presented with sufficient detail to be useful for other practitioners? No", "responses": [] } ]	1	https://f1000research.com/articles/12-1372
https://f1000research.com/articles/12-1371/v1	18 Oct 23	{ "type": "Case Report", "title": "Case Report: Idiopathic avascular necrosis of the femoral head following pregnancy", "authors": [ "Manasa Suryadevara", "Gaurav Mishra", "P.H. Parihar", "Sheetal Shelar", "Vadlamudi Nagendra", "Gaurav Mishra", "P.H. Parihar", "Sheetal Shelar", "Vadlamudi Nagendra" ], "abstract": "Background: Hip pain during pregnancy is very common, but avascular necrosis (AVN) represents a very rare entity. Osteonecrosis of the head of the femur during the pregnancy, or shortly after the pregnancy, is a rare clinical problem. Little is known about pregnancy as an etiological factor for AVN of the femoral head. Case: In this case report, a documented case of femoral head osteonecrosis, a rare complication after pregnancy is presented. The patient presented with complaints of pain in the left hip that gradually worsened and led to restriction of movement. The patient was advised an X-ray, which showed a fracture of neck of the left femur, and an MRI following the X-ray revealed features of AVN of the hip. The patient was later taken for total hip arthroplasty, which was uneventful. On follow-up, the patient had relief from her symptoms and had no complications. Alcoholism, steroid use, systemic lupus erythematosus, organ transplantation (particularly following kidney or bone marrow transplantation), dyslipidemia (particularly hypertriglyceridemia), Gaucher disease, decompression sickness, and drepanocytosis are known risk factors for osteonecrosis. We can list pro-coagulation abnormalities, chemotherapy, and HIV among the less established factors. Here, we present a case of a patient with postpartum AVN of the femoral head that suffered AVN of the left hip in the post-partum period, her symptom of hip pain was neglected as a benign cause initially, which led to aggressive treatment (total hip arthroplasty) at a young age. Conclusions: Having high suspicion of the diagnosis of AVN and a low threshold for magnetic resonance imaging (MRI) imaging in a pregnant woman with hip pain is a requirement to avoid complications. This case highlights the need for high suspicion of osteonecrosis as a cause of hip pain in the post-partum period.", "keywords": [ "Pregnancy", "femoral head", "avascular necrosis", "hip pain", "hip replacement" ], "content": "Introduction\n\nBone marrow cells and osteocytes die as a consequence of impaired blood supply to the bone affected. This pathological process is osteonecrosis, acknowledged as aseptic necrosis, ischemic necrosis, or avascular necrosis (AVN).1 This impaired blood supply induces demineralization and trabecular thinning, leading to consequent joint surface collapse and subchondral bone fracture. It was first reported in 1957 by Pfeiffer.2 AVN most commonly affects men between the ages of 30 and 50 years old.\n\nHip osteonecrosis has a number of known causes, including trauma, a genetic predisposition, sickle cell disorder, coagulation abnormalities, alcoholism, and high-dose corticosteroid therapy. Pregnancy is not a well-known risk factor for femoral head AVN, and it should be distinguished from one of the more prevalent hip pathologies in pregnancy, the so-called “Pelvic pain syndrome” and transient osteoporosis of the hip (TOH). Other than pregnancy, all other risk factors were ruled out for our patient. Not much is known regarding the development or management of AVN related to pregnancy since there have only been a few cases reported until now. Here, we have a case of neglected hip pain in a pregnant patient that resulted in AVN of the femoral head due to the uncommon association between pregnancy and the condition, which, if properly investigated early on, could have been treated more conservatively. According to the symptoms, examination, and imaging findings, a clinical diagnosis compatible with the disease is made.\n\n\nCase report\n\nIn this study, we present the case of a 25-year-old healthy Asian woman, a homemaker, who had complaints of mechanical pain in the left hip that started shortly postpartum, after caesarean section giving birth to a 4.1 kg female baby (primigravida, G1). This caused discomfort in the left hip when ambulating, which aggravated on movement and was relieved on rest. She consulted her gynecologist about the symptoms and her complaints were neglected as hip pain of a benign cause. Her symptoms eventually worsened, which caused a painful limp and hindered her daily activities. The patient had no previous medical history. There was no history of trauma, alcohol, smoking, coagulation abnormalities, sickle cell disease, or steroid usage.\n\nOn physical examination, the patient was 150 cm in height and her weight was 64 kg. She had a painful limp and a range of painful hip movements, with restrictions of mainly the abduction (30°), internal rotation (20°), and flexion (90°) of the hip.\n\nThe patient, a 25-year-old female came to our hospital in February 2023 for assessment of pain in her left hip. Her pain began a year and a half ago at the end of her first pregnancy following a caesarean section through which she delivered a healthy 4.1 kg female baby. The pain was aggravated on movement and initially relieved on rest, which gradually worsened and radiated to the knee. The patient consulted her gynecologist and was treated conservatively considering it a benign cause of pain. Her symptoms were not relieved and hence the patient came for further management, where she was advised to undergo blood tests, X-ray, and magnetic resonance imaging (MRI) study.\n\nRadiograph showed left femoral neck fracture with superolateral displacement of the greater trochanter and the rest of the shaft of the left femur (Figure 1). T1 and T2 weighted imaging sequences MRI of the femoral heads showed an irregular geographical area in the anterosuperior part of the left femoral head (Figure 2). T2, T1, and PD FatSat MRI images show the double line sign (Figure 3). Radiograph images showed a hip replacement implant prosthesis on the left side post-hip replacement surgery (Figure 4). The laboratory tests—complete blood count (CBC), erythrocyte sedimentation rate, liver function tests, c-reactive protein (CRP), creatinine, lipid panel, thyroid-stimulating hormone (TSH), rheumatoid factor and antinuclear antibody were all within normal limits. There weren’t any abnormities in the abdominal ultrasound. The patient was diagnosed with AVN of the femoral head on the left side and associated secondary osteoarthritis. One other pathology that could be confused with AVN is the TOH, which is self-resolving.\n\nT1WI, T1 weighted image; T2WI, T2 weighted image.\n\nT1WI, T1 weighted image; T2WI, T2 weighted image.\n\nThe blood test results of the patient were normal. The patient had been prescribed pain killers that had not resolved the patient’s symptoms. The patient had her X-ray done, which showed fracture of neck of the left femur with displacement of the shaft. The orthopedic advised for MRI of the hip, which showed the features of osteonecrosis of left hip. The patient then underwent a total hip arthroplasty. The surgery was uneventful.\n\nPost-op follow-ups at four weeks, two months and three months were done, which revealed the resolution of her symptoms with a gait free of pain and the limping.\n\n\nDiscussion\n\nThe most common causes of atraumatic hip osteonecrosis include alcohol intake and corticosteroids.3,4 Other risk factors include genetics, vasculitis, hyper coagulopathy, cocaine use, and micro emboli.5 While idiopathic AVN of the hip accounts for around 20% of cases,3,4 femoral head osteonecrosis is rarely associated with pregnancy.\n\nAVN of the hip associated with pregnancy has been linked to a number of pathophysiological mechanisms, but none of them have been proven to be the sole contributing factor. Pregnancy commonly results in increased coagulability and venous congestion.6–8 Also, some endocrine changes that increase the levels of unbound cortisol during pregnancy have been linked to an increased risk of hip AVN.6,7 Hyperplasia of parathyroid can also occur during pregnancy, which increases the parathyroid hormone levels thus increasing the risk for developing osteonecrosis.9 The endogenous lipoproteins can be destabilized by the placenta, which can eventually lead to fat embolism.10\n\nThe right common iliac artery lies right above left common iliac vein, making the vein more susceptible to compression with excessive weight gain and during pregnancy, especially on the left side. This mechanical factor has also been suggested as a contributing factor to the increased risk of hip osteonecrosis in pregnancy.6,7,11,12 The literature indicates that the left hip is most frequently affected and this involvement mostly occurs in the third trimester or soon after delivery,6,7,12,13 which supports the mechanical theory. In addition, hip AVN more commonly develops in primigravida,7,14 and in older women6,7,13,14 in contrast to our case, where the patient is young. From current studies, another cause may be ovulation induction, which activates both fibrinolytic and coagulation systems.15,16 It has been reported that a difficult delivery or the compression of a growing uterus can lead to injury of the artery in the round ligament or the femoral joint.17 Biochemical, hematological and coagulation parameters were all within the normal limits in our patient.\n\nThe initial complaint is generally unilateral and can either be a gradually worsening or a sudden groin pain that radiates to the back, thigh or knee. Physical examination shows a restriction of active and passive joint movements associated with pain, more pronounced with rotation and less with flexion and abduction.18\n\nTOH is another common hip pathology in pregnancy and is sometimes mistaken for hip AVN.6,14,19–21 It is a rare cause of acute hip pain in pregnancy and most commonly occurs in the third trimester,19 characterized by sudden onset of pain in the hip that can be severe without movement restriction. TOH is a condition that is self-limiting and resolves gradually within seven to eight months. The management comprises of weight bearing restrictions.6,21,22 By contrast, AVN onset is insidious, with pain increasing progressively, leading to movement limitation, and will eventually need early surgical intervention.19,20,21 Hence it is very important to differentiate between these entities since the treatment and prognosis differ greatly.6,12,19,21\n\nSince the changes of osteopenia in TOH are evident only four to eight weeks from the onset of symptoms, the standard X-ray lacks sensitivity in distinguishing AVN and TOH.20,21 With specificity and sensitivity of more than 99%, MRI is the gold standard imaging modality for diagnosing AVN in its early stages.23 The MRI features of TOH are diffuse edema of the head and neck of the femur and, by contrast, MRI features of AVN include diffuse edema with focal defects and subchondral changes.7,19–21 Double line sign is a pathognomonic MRI finding in AVN and refers to two adjacent serpentine lines demarcating the boundary between viable and devitalized bone marrow. The inner line corresponds to reparative granulation tissue, while the outer line corresponds to reactive sclerosis.24\n\nThe best way to treat AVN associated with pregnancy is not very clear. Early diagnosis and advice are significant. Conservative methods such as physical therapy, restricted weight bearing, medication, electrical stimulation, extracorporeal shock-wave treatment, and hyperbaric oxygen treatment are usually started if there is no evidence of collapse.25 The prognosis appears to be good following this conservative therapy, however surgical treatment would eventually be required for the secondary osteoarthritis or degenerative changes at a later age.18\n\n\nConclusions\n\nAlthough AVN developing post-pregnancy is a rare occurrence, AVN of the femur should be one of the top differentials for post-partum hip pain. This could aid medical professionals in identifying the disease earlier before the collapse of the femur head occurs, which may aid in the preservation of the joint as opposed to replacement.\n\n\nConsent\n\nWritten informed consent for publication of their clinical details and clinical images was obtained from the patient.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\n\nReferences\n\nYu-Chun L, Wei-Shiu C, Chiung-Chiung C, et al.: Aseptic necrosis of bilateral femoral heads after pregnancy. Taiwan. J. Obstet. Gynecol. 2011; 50(1): 111–113.\n\nPfeifer W: Eine ungewiinliche Form und Genese von symmetrischen osteonekrosen beider Femur und Humertiskophkappen. Fortsch. Geb. Rontgenstr. Nuklearmed. Ergangzungsband. 1957; 86(3): 346–349. Publisher Full Text\n\nLespasio MJ, Sodhi N, Mont MA: Osteonecrosis of the hip: a primer. Perm. J. 2019; 23: 23. Publisher Full Text\n\nHasegawa Y, Iwase T, Iwasada S, et al.: Osteonecrosis of the femoral head associated with pregnancy. Arch. Orthop. Trauma Surg. 1999 Feb; 119: 112–114. Publisher Full Text\n\nEl-Yahchouchi C, Moussa MK, Khalaf Z, et al.: Simultaneous Bilateral Avascular Necrosis of the Femoral Heads Associated With Cocaine Use. Cureus. 2020 Aug 19; 12(8): e9865. PubMed Abstract \| Publisher Full Text\n\nMontella BJ, Nunley JA, Urbaniak JR: Osteonecrosis of the femoral head associated with pregnancy. A preliminary report. J. Bone Joint Surg. Am. 1999 Jun 1; 81(6): 790–798. PubMed Abstract \| Publisher Full Text\n\nCheng N, Burssens A, Mulier JC: Pregnancy and post-pregnancy avascular necrosis of the femoral head. Arch. Orthop. Trauma. Surg. 1982 Oct; 100: 199–210. Publisher Full Text\n\nVandenbussche E, Madhar M, Nich C, et al.: Bilateral osteonecrosis of the femoral head after pregnancy. Arch. Orthop. Trauma Surg. 2005 Apr; 125(3): 201–203. PubMed Abstract \| Publisher Full Text\n\nSchnatz PF, Curry SL: Primary hyperparathyroidism in pregnancy: evidence-based management. Obstet. Gynecol. Surv. 2002 Jun 1; 57(6): 365–376. PubMed Abstract \| Publisher Full Text\n\nMyllynen P, Mäkelä A, Kontula K: Aseptic necrosis of the femoral head during pregnancy. Obstet. Gynecol. 1988 Mar 1; 71(3 Part 2): 495–498.\n\nCockett FB, Thomas ML: The iliac compression syndrome. Br. J. Surg. 1965 Oct; 52(10): 816–821. PubMed Abstract \| Publisher Full Text\n\nHernigou P, Jammal S, Pariat J, et al.: Hip osteonecrosis and pregnancy in healthy women. Int. Orthop. 2018 Jun; 42: 1203–1211. Publisher Full Text\n\nLausten GS: Osteonecrosis of the femoral head during pregnancy. Arch. Orthop. Trauma Surg. 1991 Jul; 110: 214–215. Publisher Full Text\n\nGribble RK, Berres LE: Idiopathic osteonecrosis of the hip during pregnancy: outcome in a subsequent gestation. Obstet. Gynecol. 2001 Nov 1; 98(5): 911–913. Publisher Full Text\n\nChan WS, Dixon ME: The “ART” of thromboembolism: a review of assisted reproductive technology and thromboembolic complications. Thromb. Res. 2008; 121(6): 713–726. Publisher Full Text\n\nMarios G, Lykissas ID, Gelalis IP, et al.: Korompilias: the role of hypercoagulability in the development of osteonecrosis of the femoral head. Orthop. Rev. 2012; 4(10): 73–78.\n\nMwale F, Wang H, Johnson AJ, et al.: Abnormal vascular endothelial growth factor expression in mesenchymal stem cells from both osteonecrotic and osteoarthritic hips. Bull. NYU Hosp. Jt. Dis. 2011 Apr 2; 69 Suppl 1: S56–S61. PubMed Abstract\n\nNassar K, Rachidi W, Janani S, et al.: Aseptic necrosis of the femoral head after pregnancy: a case report. Pan. Afr. Med. J. 2016; 24: 195. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSteib-Furno S, Mathieu L, Pham T, et al.: Pregnancy-related hip diseases: incidence and diagnoses. Joint Bone Spine. 2007 Jul 1; 74(4): 373–378. PubMed Abstract \| Publisher Full Text\n\nGuerra JJ, Steinberg ME: Distinguishing transient osteoporosis from avascular necrosis of the hip. J. Bone Jt. Surg. 1995 Apr 1; 77(4): 616–624. Publisher Full Text\n\nBalakrishnan A, Schemitsch EH, Pearce D, et al.: Distinguishing transient osteoporosis of the hip from avascular necrosis. Can. J. Surg. 2003 Jun; 46(3): 187.\n\nMens JM, Vleeming A, Stoeckart R, et al.: Understanding peripartum pelvic pain: implications of a patient survey. Spine. 1996 Jun 1; 21(11): 1363–1369. PubMed Abstract \| Publisher Full Text\n\nPierce TP, Jauregui JJ, Cherian JJ, et al.: Imaging evaluation of patients with osteonecrosis of the femoral head. Curr. Rev. Musculoskelet. Med. 2015 Sep; 8: 221–227. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPalmer W, Bancroft L, Bonar F, et al.: Glossary of terms for musculoskeletal radiology. Skelet. Radiol. 2020 Jul; 49: 1–33. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTurgay T, Aydeniz A: Avascular necrosis of the bilateral femoral head with pregnancy: A case report." }	[ { "id": "248359", "date": "23 Apr 2024", "name": "Douglas Dunlop", "expertise": [ "Reviewer Expertise Trauma & Orthopaedic Hip Surgeon" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis is an article that provokes interest by presenting a potential link between pregnancy and AVN of the hip, a previously rare association. The authors outline the normal expected aetiology of the condition and suggest possible mechanisms of pathogenesis to account for the link. Unfortunately the article just mentions, almost incidentally, the femoral neck fracture and fails to address the causation or relationship between the fracture and the AVN. They mention 'no history of trauma' but there are no salient details of possible causes. For example: did the patient have an epidural during delivery; was vaginal delivery abandoned and hence a caesarean; were they in stirrups; at what point did the pain begin, immediately post partum or later; was it sudden or insidious; did they breast feed and/or have normal bone markers etc. It is entirely possible that the fracture predated the AVN and is a potential cause of AVN rather than the pregnancy, although this in itself is a little odd as there is no retained density of the head, more a generalised (probably lack of weightbearing) osteopenia of the left hip head and proximal shaft. The patch of AVN on the MRI is relatively small and does not appear to relate to the fracture. This suggests that AVN is unlikely to be causal to the fracture, but again, this should be part of their conjecture. Regarding the patient treatment, there are alternatives in such a young patient rather than just a total hip replacement. The femoral head is still spherical (pre-collapse) and so fixation, with or without biological augmentation (free or vascularised fibula) could be considered / should be mentioned, even if it is not in the surgeons skill set.\n\nIs the background of the case’s history and progression described in sufficient detail? No\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? No\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? No\n\nIs the case presented with sufficient detail to be useful for other practitioners? No", "responses": [] }, { "id": "269409", "date": "28 May 2024", "name": "Evren Ustuner", "expertise": [ "Reviewer Expertise Radiology", "ultrasound", "Nonvascular interventional Radiology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors are describing a case report of AVN after pregnancy. The topic is interesting and is describing a rarely reported event, the information presented in the paper is valid, addition of detailed information about the case would further improve it. -AVN in idiopathic cases may be related to rapid increase in weight and obesity, as often seen in pregnancy, this association should be mentioned in the report. - Key clinical information is missing about the patient, for instance, -BMI of the patient -Exact time period of the start of symptoms after delivery -Has there been a venous insufficiency during pregnancy, -Did the patient receive any anticoagulation medication? -Did the patient undergo any venous Doppler Ultrasound examination during and after the symptoms? İntroduction section - How many AVN cases related to pregnancy so for? Discussion section - A comprehensive review about the outcomes of AVN's (early detected vases vs late cases) and outcome percentages need to be given in the discussion section. For instance Turgay T et al, Avascular necrosis of the bilateral femoral head in pregnancy, Agri 2020 mentions a case report of AVN in a pregnant patient that has been detected early and treated conservatively and recovered without sequela or femoral head collapse or necrosis. This may be shown as an example of how early diagnosis and management may lead to recovery, whereas outcome of late diagnosis is grim -Please do not use the term neglected but say attributed to benign causes because this is not neglect but misdiagnosis. -Treatment options need to be more thoroughly discussed -Emphasis on the specific vascular anatomy of the femoral neck and the involved artery should be mentioned.\n\nIs the background of the case’s history and progression described in sufficient detail? Partly\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Partly\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? No\n\nIs the case presented with sufficient detail to be useful for other practitioners? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1371
https://f1000research.com/articles/12-1370/v1	18 Oct 23	{ "type": "Study Protocol", "title": "Evaluation of comparative efficacy of cow milk treated Sida Cordifolia oil nasal administration (Ksheerbala Taila Nasya) versus Convolvulus pluricaulis oil nasal administration (Shankapushpi Taila Nasya) in the management of primary insomnia (Anidra)- a trial protocol", "authors": [ "Dr.Annya Gautam", "Dr.Vinod Ade", "Dr.Vinod Ade" ], "abstract": "Background: Ayurveda, formerly known as the \"Science of Life,\" is a distinct science and philosophy that aims to achieve comprehensive wellbeing by balancing the psychological, emotional, spiritual, and physical facets of life. The three basic principles listed by Ayurveda to maintain good health are Food (Ahara), sleep (Nindra), and Celibacy (Bramhacharya). According to Ayurveda, it is a strong rigid support of life. Many people today depend on sleeping pills and have developed a dependence on them. Potentially harmful side effects of sleeping drugs include drowsiness during the day, constipation, dizziness, and problems maintaining equilibrium. It is said that a vitiated Vata Dosha sets off the effects of insomnia, hence balancing Vata Dosha can provide relief in the event of Insomnia. Convolvulus Pluricaulis (Shankhpushpi) herb is a memory enhancer that can also be used as a brain tonic to stimulate brain and brain function. According to Ayurveda, the entrance to the brain is through the nose. Nasal administration refers to the practice of administering medication nasally. It is the most important therapy to treat upper clavicular disease. Objective: To study the evaluation of comparative efficacy of cow milk treated sida cordifolia oil nasal administration (Ksheerbala Taila Nasya) versus Convolvulus Pluricaulis Oil Nasal Administration (Shankapushpi Taila Nasya) in the Management of Primary Insomnia (Anidra)-RCT\n\nProtocol: In this study, a total of 128 patients will be divided into two groups. In group, A (64 patient) cow milk treated with Sida Cordifoila Oil Nasal Administration will be administered eight drops in each nostril for 15 days. The same will be used for the other 64 patients i.e., group B with Convolvulus Pluricaulis Oil Nasal Administration.\n\nExpected result: The result will be assessed on the objective parameter and data will be compared after the treatment.", "keywords": [ "Insomnia", "Nasal Administration", "convolvulus pluricaulis", "sida cordifolia", "oil" ], "content": "Introduction\n\nOne of the most well-known traditional medicinal practices from antiquity that has survived and developed over time is ayurveda. This system will endure for many more millennia because of the extensive knowledge of natural remedies, the linkages between environmental factors and human body composition and performance, and the cosmos's interconnected elements that have an impact on living things.1\n\nPanchkarma (detoxification therapy) is an Ayurvedic detoxifying retreat technique. Panchkarma is a detoxification and relaxing approach that has been used for centuries to help people cope with seasonal and societal changes. Benefits of this therapy include improvements in one's perception of social support, quality of life, and healthy behavior.2\n\nFor daily activities, focus, alertness, mental and physical performance, etc., sleep is crucial. Sleep disruptions can lead to serious health issues and other difficulties.\n\nAn individual who struggles to fall or stay asleep is said to have insomnia. Short-term and longer insomnia are both possible; short-term insomnia can last anywhere from one night to a few weeks, while long-term insomnia develops when sleep problems endure for longer than three months. Primary and secondary insomnia are the two categories of the condition. In contrast to secondary insomnia, which occurs when a patient's sleep problems are brought on by an underlying health problem such as asthma, cancer, heartburn, pain, medications, or other substances such as alcohol whereas primary insomnia is characterized by the absence of any other health concerns in the patient.3\n\nThe three pillars of life in Ayurveda are food (Ahara), sleep (Nindra), celibacy (Bramhacharya), according to a Hindu religious teacher, insomnia is mentioned in the Charaka Samhita, Susruta Samhita, and Astang Hridaya, among other ayurvedic classic texts.4 Insomnia is defined in eight undesirable conditions (Asthonindtiya Adhyaah) by spiritual guide Charak, and insomnia is mentioned in Maharoga Adhyaya as one of the 80 types of Vata disease.5 It is described in the Susruta Samhita in the Sharir Sthana under Garbhavyakaran Sharir.\n\nThe causes of insomnia, according to Acharya Sushruta, are Santapa of vaata, pitta, and mana, as well as physical harm. Insomnia is caused by a variety of factors, the most prevalent of which are listed above.6 In Astang Hridiya spritual guide Vaghbhat has quoted the symptoms of insomnia as following throbbing pain, heaviness in head, yawning, stiffness all over body, exhaustion, giddiness, indigestion, drowsiness it may further causes Vata disease.7 It is said that a vitiated Vata Dosha sets off the effects of insomnia, hence balancing Vata Dosha can provide relief in the event of insomnia.\n\nInsomnia is said to affect 9% of the general population, with 30% of people experiencing it occasionally. 18.6% of the cohort research participants said they had sleeplessness. A north Indian urban population was found to have a greater prevalence of sleep disorders (28%) that affect the onset and maintenance of sleep.8\n\nConvolvulus pluricaulis herb is a memory enhancer that can also be used as a brain tonic to stimulate brain and brain function. It improves focus and learning capacities while also assisting in the treatment of insomnia, stress, depression, and mental stress.\n\nConvolvulus pluricaulis characterizes bitter taste. It shows oily, sticky, and slimy property, cold potency, and sweet metabolic property help in preventing a variety of health problem that disrupt sleep.\n\nThe intellectual effect boosts mental power and hence resists depression and anxiety etiology, resulting in restful sleep. Shankhpushpi is known to lower cortisol (the stress hormone), reducing stress and improving the quality and quality of natural sleep. Gastric problems can disrupt sleep, but this magical herbal remedy aids digestion and prevents reflux and gas accumulation, allowing you to sleep better. Chemical components found in Shankhpushpi, such as Shankhapushpine, convoline, convolidine, confoline, myristic acids, scopoletin, -sitosterol, tetratriacontane acids, flavonoid-kaempferol, and phytosterols, help to boost general health and ward off sickness- resulting in insomnia.9\n\nAccording to Ayurveda, the entrance to the brain is through the nose. Nasal administration refers to the practice of administering medication nasally. It is the most important therapy as it is used for the treatment of upper clavicular disease so in some places it has been given first place in the sequence of detoxification for e.g., in the chapter of Apamarga Tanduliyam of Charaka Samhita Sutrasthana.\n\nThe medication used in nasal administration enters the brain via a pathway known as the Vital point (Shringataka Marma), where it destroys the connection between toxins and brain tissues before transporting those molecules to the nose, where secretions force them out. People with brain illnesses are frequently given large amounts of unwanted medications by mouth. This may become troublesome for a patient's health if they live a long time or if they have a chronic ailment. Because of the brain's blood barrier restriction to oral use, tablets delivered orally are an incredibly poor approach to deliver medications to the brain when it comes to brain illnesses. Sneha can cross the blood-brain barrier because to its lipophilic activity, which releases the active principles of the constituents where they are needed. The likelihood that a medicine will cross the blood-brain barrier increases with its lipophilicity.10\n\n\nNeed of the study\n\nIn present era, Digital media and smart phones have raised productivity demands while blurring the boundaries between work and personal life. All these factors, including reduced free time, a faster pace of life, and stress, contribute to sleep deprivation, which has major medical and emotional consequences, health, job performance, and overall quality of life are all factors to consider.\n\nModern medical research has therapies such as hypnotics, sedatives, anxiolytics, and psychotropics, but there is still no definitive cure for insomnia, and these medications have their own set of limitations due to their side effects.11 Due to their addiction and side effects, current anxiolytic medications have a limited application.12\n\nAs a result, these drugs cannot be used for an extended period. As a result, it is crucial to seek out drugs from natural systems of medicine such as Ayurveda that are both safe and successful in treating insomnia patients. Hence this study is conducted with the aim evaluation of comparative efficacy of cow milk treated Sida Cordifolia oil nasal administration (Ksheerbala Taila Nasya) versus Convolvulus Pluricaulis oil nasal administration (Shankapushpi Taila Nasya) in the management of primary insomnia (Anidra).\n\n\nAim\n\nEvaluation of Comparative Efficacy of cow milk treated sida cordifolia oil nasal administration (Ksheerbala Taila Nasya) versus convolvulus pluricaulis oil nasal administration (Shankapushpi Taila Nasya) in the Management of Primary Insomnia (Anidra).\n\n\n\n1. To assess the efficacy of Shankahpushpi Taila Nasya in the PSQI (PITTSBURG SLEEP QUALITY INDEX), Insomnia Severity Index, WHO Quality of life scale.\n\n2. To assess the efficacy of Ksheerabala Taila Nasya in the PSQI (PITTSBURG SLEEP QUALITY INDEX), Insomnia Severity Index, WHO Quality of life scale.\n\n3. To Compare the efficacy of of Shankhpuspi Taila Nasya with Ksheerbala Taila Nasya in all three scale in the management of Primary Insomnia (Anidra).\n\nTrial design- Randomization standard control double blind superiority clinical trial.\n\nThe allocated intervention will blind participants.\n\n\nProtocol\n\nThe patient of Insomnia will be recruited from the panchakarma OPD and IPD of Mahatma Gandhi Ayurved College, Hospital & Research Centre, Salod(H), and from peripheral camps.\n\nA total of 60 patients will be recruited for the study. They will be randomly divided into two groups. Group A will be given nasal administration with Ksheerbala oil, Group B will be given nasal administration with Shankhapushpi. All the baseline parameters will be recorded at the start of the study. The patients will undergo treatment for 15 days for both groups. After treatment, all the parameters will be recorded at the 16th and 30th day.\n\nGuideline: Used the SPIRIT guideline for the study.\n\nCase definition: Patient of insomnia (having age group between 20-60 year of irrespective gender) diagnosed based on Insomnia Severity Index.\n\nSampling procedure: Randomization computer generated table.\n\nType of Study: Interventional Study\n\nStudy design: Refer to Figure 1.\n\nStudy design: Randomization standard control double blind superiority clinical trial.\n\nRoles and Responsibilities of committees: The study will start after clearance from IEC of Mahatma Gandhi Ayurved College Hospital and Research Centre, Salod(H), Wardha. And after CTRI registration Ref.No. MGACHRC/IEC/June-2023/707\n\nThe committee will decide on the endpoint and oversee the trial as it progresses.\n\nThe researcher will assess any adverse events and will report to the ethics committee.\n\n\n\n1. Patients who are willing to give written inform consent.\n\n2. Patients’ gender age is between 20-60 years.\n\n3. Patients who are fit for Nasal administration procedure.\n\n4. Patient with controlled hypertension\n\n• (Systolic not more than 140 mmHg and diastolic not more than 90 mmHg).\n\n5. ICD10 criteria for primary insomnia – F51.01\n\n• Problems falling asleep or staying awake.\n\n• Sleep varies, i.e., one night of good sleep comes after several nights.\n\n• Daytime sleepiness.\n\n• Problems in performing daily activities.\n\n\n\n1. Patients < 20 years and > 60 years.\n\n2. Pregnant women and lactating mothers.\n\n3. Patient with co-morbidities Ex. uncontrolled Hypertension, DM-2, Thyroids, Cardiac disorders.\n\n4. Patient not fit for nasal administrating procedure, Rhinitis, Thirst, Indigestion, Menstruation etc.\n\n5. Patients with drug dependency like Antihistamines, Narcotics.\n\nOccupational disease.\n\n\n\nThe coarse powder of the kalka Dravya will be taken in khalva yantra and triturated with little quantity of water till we obtain kalka (Paste).\n\n\n\nNow, tila taila will be consumed in a clean stainless-steel vessel with wide opening that is set over low flame.\n\n\n\nThen, shankhapushpi kwatha (decoction) and water followed by kalka will be added and processed with frequent stirring.\n\n\n\nThen as soon as all the Sneha siddhi lakshanas will be attained, the hot taila will be filtered into a clean stainless-steel vessel.\n\n\n\nThen, after the tail cool down by its own, it is packed and preserved in appropriate airtight containers for further therapeutic use.\n\n\n\n1. Pre-procedure\n\n2. Procedure\n\n3. Post-procedure\n\nAll of the necessary tools and supplies should be acquired and organized prior to the procedure. The drugs required for Procedure i.e., Paste, Powder, Milk, Decoction should be available in a separate room free of dust, with direct air flow, sufficient lighting, Nasal administration table, and material required (the drugs required for procedure i.e., Paste, Powder, Milk, Decoction, nasal dropper, cotton swab, oil, medicinal smoke machine. A special smoke machine is required for Pradhamana Nasya. An attendant, as well as dressing tools, a spittoon, napkins, bowls, and towels, should be available.\n\nThe patient should be appropriately prepared by ensuring that they thoroughly comprehend the procedure and that they are willing (permission) to carry it out. Before lying down on the nasal administration table, the patient is recommended to pass his or her natural cravings. Soft and gentle massage should be done on scalp, forehead, face, neck for approximately 10-15 minutes using medicated oil. For the liquefaction of the Dosha, mild sudation (mridu swedan) should be performed, Mild sudation must be administered with caution, as sudation should not be performed on the head region, according to Ayurveda's classical text.\n\nThe patient should be advised to lie down on the nasal administration table in a supine posture with his or her head hanging downward and not overly stretched or flexed. The reason behind this is because if the head is not appropriately lowered, the injected medicine may miss its target, and if it is lowered too much, the drugs may reach the brain and become lodged there. Cotton or cloth should be used to cover the patient's eyes. The physician should next use the left thumb to lift the tip of the nose, and the right hand to pour the medicine into the nostrils. The drug should be taken in the correct amount, neither too much or too little. Depending on the patient's tolerance, it should be lukewarm. The patient should be encouraged to remain cool and composed throughout the process, and to refrain from speaking, laughing, sneezing, or shaking his head. Then keep the patient lying down and observe the symptoms arising in him.\n\n\n\n\nNasal administration complications and its management\n\nThere are two varieties of complication, according to spiritual guide Sushruta: Dosha Vriddhi Janya, Secondly, Dosha Kshaya Janya Vyapad. Shodhana and Shaman Chikitsha should be stated in the case of Dosha Vriddhi Janya Vyapad, while Brihmana Chikitsa is recommended in the case of Dosha Kshaya Janya Vyapa.\n\nThe patient should be asked to lie in a supine position for roughly 1 minute after receiving the nasal medication, and the feet, shoulders, palms, and ears should be massaged during this time. If there is too much of a certain medicine, it should be spit out rather than consumed. The remnant Kapha Dosha that may be lodged in the Kantha and Sringataka Marma is then expelled with medicated Gandusha and medicated smoke. After that, the patient should be encouraged to stay in a room with no breeze, eat a light meal, and drink plenty of water.\n\nAssessment criteria:\n\nObjective criteria:\n\n(A) PSQI (PITTSBURG SLEEP QUALITY INDEX)15 [Table- 1]\n\n(B) INSOMNIA SEVERITY INDEX16 [Table – 2]\n\n(C) QUALITY OF LIFE17 [Table – 3]\n\nInvestigations: NIL\n\nSample size-\n\nSample size by Cohen’s effect size by comparing two means.\n\nPrimary Variable: - Insomnia Assessment by PSQI Scale and Insomnia Severity Index.\n\n(Estimated)\n\nConsidering large effect size difference = 0.8 (Large effect size)\n\nSample size\n\nat 5 % level of significance = 1.96\n\nat 80 % Power = 0.84\n\nRatio allocation (Group2/Group1) = 1\n\nSample size = 26 per group.\n\nConsidering 15 % drop out total = 4\n\nTotal 30 samples required per group. Sample size by Cohen’s effect size by comparing two means.\n\nPrimary Variable: - Insomnia Assessment by PSQI Scale and Insomnia Severity Index.\n\n(Estimated)\n\nConsidering large effect size difference = 0.5 (Medium effect size)\n\nSample size\n\nat 5 % level of significance = 1.96\n\nat 80 % Power = 0.84\n\nRatio allocation (Group2/Group1) = 1\n\nSample size = 64 per group.\n\nConsidering 10 % drop out total = 6\n\nTotal 70 samples required per group.\n\nThe patient of insomnia will be selected from the panchakarma OPD and IPD of Mahatma Gandhi Ayurved College, Hospital & Research centre, Salod(H), and from peripheral camps. A total of 60 patients will be recruited for the study.\n\nAllocation sequence generation- Computer-Generated Random Numbers.\n\nAllocation implementation- The researcher or the first author will generate an allocation sequence, enroll participants, and assign participants to intervention.\n\nBlinding- Double blind superiority clinical trial.\n\n\n\n\n\n1. Ayurveda Texts\n\n2. Modern texts\n\n3. Online search- Google scholar etc.\n\n4. Cow milk treated with Sida Cordifolia oil nasal administration\n\n5. Ksheerbala Taila Nasya\n\n6. Convolvulus Pluricaulis oil nasal administration\n\n7. Shankhapushpi Taila Nasya\n\n8. Case record form\n\n9. Patient Information\n\n10. Written and informed consent form\n\nDrug collection/authentication: The raw drug will be procured from reliable sources and authenticated by Department of Dravyaguna and Rasashastra of Mahatma Gandhi Ayurveda College, Hospital and Research Centre, Wardha.\n\nStatistics outcome: After the study data will be analyzed according to a suitable statistical test.\n\nData Analysis: The collected data will be analyzed with the help of inferential statistical test.\n\n\nDiscussion\n\nThe three Ayurvedic sthambhas—Vata, Pitta, and Kapha—oversee all the body's essential processes. When the three Sthambhas are in an equilibrium state, a person is said to be healthy. There are three Pillars (Upsthambha) to support these Sthambha.\n\nFood (Ahara), Sleep (Nindra) and Celibacy (Brahmacharya) are the three. Upsthambha is one of the main forces that contribute to and support life. Such Pillars (Upsthambha) increase the strength of an individual. Sleep (Nindra) has an impact on both mental and physical elements. Nidra is therefore crucial for both physical and emotional wellness.\n\nWork, ageing, illnesses, constitution, and some Doshas like Vata and Pitta were only a few of the many causes of sleep deprivation that were explained by Ayurveda. These elements have a direct impact on sleep and result in sleep loss. Loss of sleep can occur for a variety of causes, including disease, stress, ageing, pain, mental illness, and others, according to modern scientific theory. Insomnia was included in the 80 Vataja disease by Charaka and Kashyapa.\n\nThe herb Convolvulus Pluricaulis (Shankhpushpi), a memory enhancer utilized as a brain tonic to stimulate cognition and brain function. It improves focus and learning skills and aids in the treatment of conditions such as depression, stress, and mental tiredness.\n\nThe Medhya effect increases strength of mind, which helps one resist the pathophysiology of depression and anxiety and promotes restful sleep. Convolvulus Pluricaulis (Shankhpushpi) is said to improve the quality and length of natural sleep by lowering cortisol (the stress hormone) levels, which in turn reduces stress.\n\nThe chemical components of Shankhpushpi, including flavonoids kaempferol and phytosterols, myristic acids, scopoletin, -sitosterol, tetratriacontane acids, and convoline, aid to promote general health and prevent disease-related sleeplessness.\n\nSince the nose serves as the brain's entrance, this procedure allows for the administration of medications in accordance with the severity of the patient's illness. It is a very effective therapy modality that is included in the arsenal of purification therapies due to the numerous classifications and varieties of nasal procedure that have been stated, as well as the various methods of medication delivery. Following a thorough examination of the patient and the selection of a particular nasal procedure, pre-procedure, procedure, and post procedure can be readily carried out systematically. The method's likely mode of action indicates that it is successful in producing the desired effects in the treatment of insomnia.\n\n\nConclusion\n\nConvolvulus Pluricaulis oil nasal administration may be more effective than cow milk treated with Sida Cordifolia oil nasal administration in treating insomnia while having fewer adverse effects.\n\nThe study will start after clearance from the IEC of Mahatma Gandhi Ayurved College Hospital and Research Centre, Salod(H) Wardha. And after CTRI registration. Ref. No. MGACHRC/IEC/June-2023/707\n\nThe committee will decide on the endpoint and oversee the trial as it progresses.\n\n\nConsent\n\nBoth written and verbal consent of the patient will be taken before conducting the trial in the local language while explaining every aspect of the study. The researcher will take consent from trial participants.\n\n\nDissemination\n\nThis protocol will be further published as a thesis to disseminate the study of Insomnia. The study protocol provides a detailed overview of the study design, methodology, data collection, data analysis plan, and ethical consideration. By disseminating this protocol, we hope to advance knowledge in the field of research.\n\n\nStudy status\n\nWe have started enrolling patients.\n\n\nAyurvedic Terminology and its translation\n\n", "appendix": "Data availability\n\nNo data as this is a protocol.\n\nFigshare: - SPIRIT Checklist for study. Evaluation of Comparative Efficacy of cow milk treated sida cordifolia Oil Nasal Administration (Ksheerbala Taila Nasya) versus Convolvulus Pluricaulis Oil Nasal Administration (Shankapushpi Taila Nasya) in the Management of Primary Insomnia (Anidra). DOI: 10.6084/m9.figshare.24037839\n\nLicense: Creative Commons Attribution 4.0 International.\n\n\nReferences\n\nJaiswal YS, Williams LL: A glimpse of Ayurveda–The forgotten history and principles of Indian traditional medicine. J. Tradit. Complement. Med. 2017 Jan 1; 7(1): 50–53. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPeterson CT, Lucas J, John-Williams LS, et al.: Identification of altered metabolomic profiles following a panchakarma-based ayurvedic intervention in healthy subjects: The self-directed biological transformation initiative (SBTI). Sci. Rep. 2016 Sep 9; 6(1): 1–4. Publisher Full Text\n\nhttp\n\nSamhita C, Shukla AV, Pratishthan CS: delhi. Edition- 2015. Sutra Sthana Chapter 21 Asthoninditiya Adyaah sloka 35.\n\nSamhita C, Shukla AV, Pratishthan CS: delhi. Edition- 2015. Sutra Sthana 20 Asthodariya Adyaah sloka 11.\n\nSamhita S, Ambikadattashashtri A, Prakashan CS: Varanasi. Edition-2007. Sharir Sthana Chapter 4 Garbhavyakaran Sloka 41.\n\nHridayam A, Brahmanandtripathi CSP: delhi, reprint edition- 2017. Sutra Sthana Chapter 7 Annaraksha Adhyaah Sloka 64.\n\nhttp\n\nKumar S, Patel D, Tiwari A, et al.: A Review on Role of Shankhpushpi Kalka and Yashtimadhu Taila Shirodhara towards the Management of Anidra. Journal of Drug Delivery and Therapeutics. 2019 Dec 6; 9(4-A): 884–886.\n\nSingh G: Nasya Therapy-A Pharmacological Route for Drug Delivery to Brain. Journal of Ayurveda Physicians & Surgeons (JAPS) (EISSN 2394-6350). 2018 Nov 6; 5(3).\n\nShilpa SN, Krishnamurthy MS, Ravishankar B, et al.: PRELIMINARY ANALYTICAL STUDY OF SAINDHAVADYA GHRUTA. Int. J. Ayur. Pharma Research. 2014; 2(4): 135–140.\n\nSingh AK: Manasika Bhavas in Anidra -Stress Induced Chronic Insomnia - Mamsyadi Ghrita - Dashamula Kwatha Shirodhara-Kc (Manasa)- 2007-Ipgt&Ra, Gau, Jamnagar.\n\nRavindra A: A text book of Bhaisajya Kalpana Vijnana[Pharmaceutical science], Chaukhamba Surbharati Prakashan- Varanasi, edition-2011.\n\nhttp\n\nhttp\n\nBastien CH, Vallières A, Morin CM: Validation of the Insomnia Severity Index as an outcome measure for insomnia research. Sleep Med. 2001 Jul 1; 2(4): 297–307. Publisher Full Text\n\nFlanagan JC: A research approach to improving our quality of life. Am. Psychol. 1978; 33: 138–147. Publisher Full Text\n\nSamhita S, Ambikadattashashtri A, Prakashan CS: Varanasi. Edition-2007. Chikitsasthan chapter 40/28." }	[ { "id": "216809", "date": "09 Feb 2024", "name": "Manjusha Sunil More", "expertise": [], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe need of the study is explained appropriately. Materials and Methods are found. Selection of drugs are justified with proper classical Ayurvedic References Plan of procedure and study design is formed correctly. The selected disease is really good.\nOverall the work is good and may be beneficial for society.\n\nAdditional information to add: Scope of research, and upcoming research.\n\nMaybe work by increasing the duration of treatment. Maybe work by using the same protocol for other diseases.\n\nIs the rationale for, and objectives of, the study clearly described? Yes\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [] }, { "id": "216808", "date": "09 Feb 2024", "name": "Anup Jain", "expertise": [ "Reviewer Expertise Ayurved", "Panchakarma", "Musculoskeletal", "Neuromuscular and mental health" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe article requires thorough grammatical vetting and needs to be clear upon the status of obtaining the IEC and CTRI numbers if they have started the recruitment of patients. It will also be prudent to provide the basis on which superiority trial is being conducted, if any pilot study has been conducted on the efficacy of the trial drug to arrive at the exact sample size which is required for superiority trial. The author may also elaborate the method of blinding in the final work\n\nIs the rationale for, and objectives of, the study clearly described? Yes\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1370
https://f1000research.com/articles/12-1369/v1	18 Oct 23	{ "type": "Research Article", "title": "Bactericidal activity of newly synthesized antimicrobial peptides against methicillin-resistant staphylococcus aureus and biofilm-forming methicillin-resistant staphylococcus aureus", "authors": [ "Ali Salama" ], "abstract": "Background: The emergence of multidrug-resistant bacteria (MDRB) poses a significant global challenge for healthcare professionals. Methicillin-resistant Staphylococcus aureus (MRSA), a prominent pathogen responsible for both hospital-acquired (nosocomial) and community-acquired infections, is particularly difficult to treat. Existing treatment options, such as vancomycin, linezolid, or clindamycin, have limitations. Therefore, there is a need for innovative approaches to combat infections caused by drug-resistant organisms. Methods: In this study, we synthesized a novel ultra-short antimicrobial peptide composed of three units of tryptophan and three units of lysine. We evaluated the effectiveness of this peptide against MRSA and MRSA that forms biofilms. Results: Encouraging results demonstrated that the peptide effectively killed both MRSA and biofilm-forming MRSA, while exhibiting low toxicity to human red blood cells. Additionally, our novel peptide showed excellent synergistic effects when combined with vancomycin against MRSA. Furthermore, when combined with levofloxacin and clarithromycin, our peptide exhibited synergistic effects against biofilm-forming MRSA. Conclusions: In conclusion, this study presents a novel ultra-short antimicrobial peptide (USAMP) that holds potential as a new generation of antibiotics to combat globally prevalent drug-resistant bacteria.", "keywords": [ "multidrug-resistant bacteria", "community-acquired infections", "resistance", "peptides and biofilm" ], "content": "Introduction\n\nAccording to the world health organization (WHO), Staphylococcus aureus is the major bacterial species causing nosocomial infections globally. S. aureus is a Gram-positive bacterium that causes a wide range of human diseases responsible for many critical hospital-acquired (nosocomial) and community-acquired infections.1 The strain has become a serious problem internationally with the development of methicillin resistant causes invasive infection which led to a 18% increase in mortality rate. For the study, which appears in the Journal of the American Board of Family Medicine, researchers analyzed data from the 2001-2004 National Health and Nutrition Examination Survey, a large, nationally representative study that combines survey questions with laboratory testing, including nasal swabs to test for the presence of MRSA. The researchers linked data on participants ages 40-85 with data from the National Death Index to track deaths over an 11-year period. Researchers adjusted for factors including gender, race and ethnicity, health insurance, poverty-income ratio, hospitalization in the previous 12 months, and doctor diagnosis of heart disease, diabetes and asthma. They found the mortality rate among participants without MRSA was about 18%, but among those with colonized MRSA, the mortality rate was 36%. Participants who carried staph bacteria on their skin, but not MRSA, did not have an increased risk for premature death.2\n\nMethicillin resistant Staphylococcus aureus (MRSA) was induced with a transfer of mecA gene from an ancestral Staphylococcus species, with the gene being mediated by a special mobile genetic element.3 The incidence of staphylococcal infections has increased due to the pathogen's ability to develop resistance to multiple antibiotics and form biofilms.4 The Central of Disease Control (CDC) has reported a 60% rise in MRSA infections in intensive care units, according to the National Nosocomial Infections Surveillance System. Treating these infections has become challenging as they exhibit resistance to traditional antibiotics, including second- and third-line drugs.4,5\n\nSome strains of Staphylococcus aureus not only display resistance to antibiotics such as methicillin but also are prolific producers of biofilm materials. Biofilm is formed when cells stick to each other’s, after adhering to solid surfaces, the process is expatiated with the production of extracellular polymeric substance.6 This matrix composes of DNA, proteins, polysaccharides, water; and microorganism cell. Various microorganism has the ability to form biofilm such as bacteria, fungi and protists.7 The biofilm formation ensures good strategy to microorganism to survive and adapt to living environmental and nutritional conditions.8 The biofilm formation process undergoes important multi-steps. The first step is the attachment of floating microorganisms to a surface. Attachment is followed by a period of growth and formation of micro-colony, creating a complex 3D structure. Followed by development of a small biofilm, maturation and detachment. In fact, when compared to planktonic, those growing as a biofilm can be up to 1,500 times more resistant to antibiotics and other biological and chemical agents.9 To control the biofilm formation, several treatment strategies have been proposed. The intensive and aggressive antibiotic treatment are used to retard their spreading but not to eradicate the whole biofilm community. Due to the great problem caused by these types of bacteria and to increase their immunity to the types of antibiotics currently available.10 A new type of drug was needed to combat it, and one of these drugs is antimicrobial peptides (AMPs). These peptides are characterized by their ability to attack the cell by binding to the cell wall and causing cell lysis.11 In this study, we synthesized an ultra-short antimicrobial peptide (USAMPs) and combined it with a number of antibiotics and measured its effectiveness against planktonic and biofilm forming Staphylococcus aureus.12\n\n\nMethods\n\nWe use methicillin resistance Staphylococcus aureus (ATCC 33591) as a biofilm forming bacteria and MRSA (ATCC BAA-41) as a planktonic strain which were obtained from the American Type Tissue Culture Collection (ATCC, Manassas, VA, USA).\n\nThe synthesized peptide is made up of three tryptophan (w) subunits and three lysine (K) amino acids, and ferulic acid was used to conjugate the peptide. The peptide was created using the solid-phase Fmoc chemistry. The designed peptides used in the present study were synthesized by (GL Biochem Ltd., Shanghai, China) using the solid-phase method and Fmoc chemistry was finally obtained as a lyophilized state. Reverse phase highperformance liquid chromatography (RP-HPLC) was used for purification of the peptide using a C18 internsil® ODS-SP column, the column was eluted with acetonitrile/H2O-TFA gradient at a flow rate of 1.0 mL/min. and validated using mass spectrometry and electrospray ionization mass spectrometry (ESIMS)13 The absorbance was at λ = 214 nm, the solvent which was use is dichloromethane.\n\nFor biofilm formation we used the Calgary biofilm device, as previously reported14 with Staphylococcus aureus (ATCC 33591). Biofilm formation was performed employing the Calgary biofilm device (Innovotech, Edmonton, Canada). The bacteria was left to grow in TSB at 37°C for 20 hours. Then, a concentration of 107 CFU/mL was prepared by diluting the cultures in the same medium. Using 96-peg lids on which the cells of biofilm can build up, 150 μL of that bacteria culture was added to each peg lid to allow the formation of biofilm on the purposed designed pegs, followed by incubating the pegs for 20 hours at 37°C under 125 rpm rotation. Blank lanes were prepared by adding 150 μL TSB to six wells. To discard planktonic cells after biofilm formation, PBS was used to wash pegs three times.\n\nWe used sterile 96-well microtiter plates to determine the minimum inhibition and bactericidal concentrations (MIC/MBC), we used the microbroth dilution method for the later determination. Muller–Hinton broth (MHB) was used as a revival growth medium for the staphylococcus aureus, it was also used as the main broth media in determining the MIC. Briefly, S. aureus were revived from glycerol stock at -70C using MUB, S. aureus cells were grown overnight and diluted to 106 CFU/mL in MHB. We also diluted our peptide in different concentrations and in separate 96-well microtiter plates, we mixed 50 μL of peptide with 50 μL of diluted bacterial suspension, we performed six replica for each peptide concentration. Plates were incubated for 18 h at 37 °C. Bacterial growth were determined via measuring the optical density at λ = 570 nm. MIC was defined as the lowest concentration of peptide which inhibited the growth of S. aureus. We included positive and negative controls with each plate to ensure bacterial growth and MHB sterility. For MBC determination we streaked 10 μL from the clear negative wells on nutrient agar and incubated the plates overnight at 37 °C. MBC value we defined as the lowest concentration that killed 99.9% of S. aureus (<0.1% viable cells). Experiments were performed in triplicate.15\n\nAntibiotic used in this study\n\nLevofloxacin, Chloramphenicol, Rifampicin Amoxicillin, Clarithromycin, Doxycycline, Vancomycin and Cefixime were obtained from sigma Aldrich.\n\nMIC and MBC determination of antibiotics alone\n\nVarious concentrations of each antibiotic (ranging from 0.25 to 250 μM) were prepared to determine the minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) against planktonic Methicillin-Resistant Staphylococcus aureus (MRSA). The antibiotics were dissolved in water and then further diluted in sterile broth for the preparation of each antibiotic solution.16\n\nMIC determination of peptides-antibiotics combinations\n\nThe broth microdilution checkerboard technique was employed to test the MICs of peptide-antibiotic combinations against planktonic MRSA. In this assay, each well of a sterile flat-bottomed 96-well plate contained a mixture of one peptide and one antibiotic in varying concentrations. Specifically, 25 μl of the peptide concentration and 25 μl of each antibiotic concentration (ranging from 0.25 to 200 μM) were added to six wells. These wells also contained 50 μl of the diluted bacterial suspension. The MICs were determined in triplicate, ensuring robustness of the results.17\n\nDetermination of synergism using fractional inhibitory concentration\n\nThe fractional inhibitory concentration (FIC) is the summation of the inhibitory concentration values of each component resulted in the antimicrobial combination divided by the inhibitory concentration alone. The FIC indices were interpreted as ≤ 0.5: synergistic activity, 0.5-1: additive activity, 1-4 indifferent, >4: antagonistic. Interpretation and assessment of the FIC index and antimicrobial activity of peptides-antibiotics combinations were conducted according to the broth microdilution checkerboard technique mention in the section above.18\n\nHemolytic assay of red blood cells was determined according to previous study19 where we used the equation below to determine the RBCs hemolysis due to the use of peptide.\n\nWhere A is Optical density 450 with the peptide solution\n\nA0 is Optical density 450 of the blank.\n\nAnd AX is Optical density 450 of control (0.1% triton X-100).\n\nIn this study, all data-generating experiments were conducted three times. The resulting values from these experiments were compared and analyzed using one-way analysis of variance (ANOVA) with least significant difference (LSD) multiple comparison tests on the means. Any variations observed were reported at a 95% confidence level (P<0.05). The analysis of the data was performed using SPSS software version 21.\n\n\nResults\n\nWhen we synthesised the peptide we used a lysin amino acid to charge the peptide (+3) which has been deemed as a suitable charge that presented effective antimicrobial effect.20 We also used Tryptophan because it has a good lipophilic characteristic and good interaction with bacteria membrane. We also combined the peptide with ferulic acid to increase the peptide hydrophobicity. The structure of the peptide shown in Figure 1.\n\nThe result of the viable cell count was shown in Figure 2, the result indicates that the combination of the peptide with antibiotics lead to dramatically decrease of the viable cell.28\n\nWe also investigated the effect of the peptide alone against MRSA and in combination with the antibiotics panel, then we determined the synergistic effect using FIC equation (Table 1 and 2).\n\nResults in Table 3 show that the conjugation at concentration of 100 μM only causes 1% hemolysis on Human erythrocytes after 30 minutes’ incubation.\n\n\nDiscussion\n\nBiofilm-producing pathogens, such as S. aureus, pose a significant challenge in medical treatment, especially once biofilm formation has occurred. These pathogens, exemplified by MRSA, are known for their ability to produce biofilms and their resistance to antibiotics. Furthermore, they are frequently associated with hospital-acquired infections, presenting a major concern for human health in the field of drug design.21 The coronavirus disease 2019 (COVID-19) pandemic resulted in a great number of patients with pulmonary destress, and great number of elderly patients were put on ventilators. Studies have shown that a large number of covid patients die as a result of pneumonia after using ventilators rather than the viral infection caused by the coronavirus.22\n\nIn this study, we synthesized an ultra-short anti-microbial peptide to study its effect on the biofilm producing Staphylococcus aureus bacteria and MRSA. The results proved a good efficacy of this compound to resist this type of bacteria after measuring the counting the percentage of viable cells. This peptide mechanism of action involved the ability of the peptide to attack the cell wall and make pores in it, which causes cell lysis and death.23 We also studied the degree of toxicity of our synthesized peptide to red blood cells, as toxicity seems to be one of the biggest issues in developing peptide therapeutics in the future, our experiment showed that our synthesized peptide had very low toxicity to red blood cells.\n\nIn order to increase the effectiveness of the peptide, we combined it with eight antibiotics to see the effect against biofilm, and the results showed a significant increase in effectiveness with Levofloxacin, Chloramphenicol, Rifampicin, Clarithromycin and Doxycycline. This can be explained by the fact that these antibiotics work inside the cell and the peptide facilitates their entry into the cell because it makes holes in the cell wall, which promote its entry into the cell and do its work easily.24,25\n\nThe minimum inhibitory concentration (MIC) values of eight conventional antibiotics, namely levofloxacin, chloramphenicol, rifampicin, amoxicillin, clarithromycin, vancomycin, cefixime, and doxycycline, were evaluated against Methicillin-Resistant Staphylococcus aureus (MRSA) strains, specifically ATCC BAA-41. The results showed that rifampicin exhibited the highest potency among these antibiotics against Gram-positive bacteria, including S. aureus (ATCC: 29215 and BAA-41), with MICs of 0.025 and 0.005 μM, respectively.26 Furthermore, vancomycin demonstrated a synergistic effect against Gram-positive bacteria. This could be attributed to vancomycin's role in targeting the cell wall, facilitating the penetration of peptides to their target sites in the cell membrane. This ultimately led to rapid cell lysis and a decrease in the effective concentrations required to inhibit bacterial growth, as observed with the peptide and vancomycin combination.27\n\n\nConclusions\n\nIn conclusion, the results proved that the peptide that we synthesized has a good efficacy against MRSA, with low toxicity to red blood cells. The results of the study indicate that the combination of peptides with available antibiotics could be one of the most significant methods to enhance the efficacy of existing antibiotics today. The peptide mode of action delivers a high concentration of antibiotics with peptides to bacterial species and ensure bacterial killing (bactericidal activities) regardless of bacterial resistance status.", "appendix": "Data availability\n\nFigshare: Peptide raw data. https://doi.org/10.6084/m9.figshare.23895903.v1. 28\n\nThis project contains the following underlying data:\n\n- supplementry file (2).docx\n\n- micS.xlsx\n\n- combination peptide.xlsx\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nAn earlier version of this article can be found on Research Square (doi: https://doi.org/10.21203/rs.3.rs-1549550/v1).\n\n\nReferences\n\nWHO\|WHO’s first global report on antibiotic resistance reveals serious, worldwide threat to public health. WHO; [cited 2017 May 1]. Reference Source\n\nMainous AG, Rooks BJ, Carek PJ: Methicillin-Resistant Staphylococcus Aureus Colonization and Mortality Risk Among Community Adults Aged 40-85. J. Am. Board. Fam. Med. 2021; 34(2): 439–441. Publisher Full Text\n\nChen L, Chavda KD, Solanki M, et al.: Genetic variation among Panton-Valentine leukocidin-encoding bacteriophages in Staphylococcus aureus clonal complex 30 strains. J. Clin. Microbiol. 2013 Mar; 51(3): 914–919. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWoodhead M, Blasi F, Ewig S, et al.: Guidelines for the management of adult lower respiratory tract infections--full version. Clin. Microbiol. Infect. 2011 Nov; 17 Suppl 6: E1–E59. PubMed Abstract \| Publisher Full Text\n\nAlbrich WC, Harbarth S: Health-care workers: source, vector, or victim of MRSA? Lancet Infect. Dis. 2008 May; 8(5): 289–301. Publisher Full Text\n\nSydnor ERM, Perl TM: Hospital Epidemiology and Infection Control in Acute-Care Settings. Clin. Microbiol. Rev. 2011 Jan; 24(1): 141–173. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGill SR, Fouts DE, Archer GL, et al.: Insights on Evolution of Virulence and Resistance from the Complete Genome Analysis of an Early Methicillin-Resistant Staphylococcus aureus Strain and a Biofilm-Producing Methicillin-Resistant Staphylococcus epidermidis Strain. J. Bacteriol. 2005 Apr 1; 187(7): 2426–2438. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRice LB: Antimicrobial resistance in gram-positive bacteria. Am. J. Infect. Control. 2006 Jun; 34(5, Supplement): S11–S19. Publisher Full Text\n\nSelsted ME, Ouellette AJ: Mammalian defensins in the antimicrobial immune response. Nat. Immunol. 2005 Jun; 6(6): 551–557. PubMed Abstract \| Publisher Full Text\n\nHancock REW, Brown KL, Mookherjee N: Host defence peptides from invertebrates--emerging antimicrobial strategies. Immunobiology. 2006; 211(4): 315–322. PubMed Abstract \| Publisher Full Text\n\nWang G, Li X, Wang Z: APD2: the updated antimicrobial peptide database and its application in peptide design. Nucleic Acids Res. 2009 Jan 1; 37(suppl 1): D933–D937. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLai Y, Gallo RL: AMPed up immunity: how antimicrobial peptides have multiple roles in immune defense. Trends Immunol. 2009 Mar; 30(3): 131–141. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSun H, Hong Y, Xi Y, et al.: Synthesis, self-assembly, and biomedical applications of antimicrobial peptide–polymer conjugates. Biomacromolecules. 2018; 19(6): 1701–1720. PubMed Abstract \| Publisher Full Text\n\nYasir M, Willcox MDP, Dutta D: Action of antimicrobial peptides against bacterial biofilms. Materials. 2018; 11(12): 2468. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTincho MB, Morris T, Meyer M, et al.: Antibacterial activity of rationally designed antimicrobial peptides. Int. J. Microbiol. 2020; 2020: 1–9. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJenssen H, Hamill P, Hancock REW: Peptide Antimicrobial Agents. Clin. Microbiol. Rev. 2006 Jul 1; 19(3): 491–511. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWang Q, Hong G, Johnson GR, et al.: Biophysical Properties of Membrane-Active Peptides Based on Micelle Modeling: A Case Study of Cell-Penetrating and Antimicrobial Peptides. J. Phys. Chem. B. 2010 Nov 4; 114(43): 13726–13735. PubMed Abstract \| Publisher Full Text\n\nRotem S, Mor A: Antimicrobial peptide mimics for improved therapeutic properties. Biochim. Biophys. Acta Biomembr. 2009 Aug; 1788(8): 1582–1592. PubMed Abstract \| Publisher Full Text\n\nSalama A, Almaaytah A, Darwish RM: The design of alapropoginine, a novel conjugated ultrashort antimicrobial peptide with potent synergistic antimicrobial activity in combination with conventional antibiotics. Antibiotics. 2021; 10(6): 712. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTorrent M, Andreu D, Nogués VM, et al.: Connecting Peptide Physicochemical and Antimicrobial Properties by a Rational Prediction Model. PLoS One. 2011 Feb 9; 6(2): e16968. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAlbada HB, Prochnow P, Bobersky S, et al.: Short Antibacterial Peptides with Significantly Reduced Hemolytic Activity can be Identified by a Systematic l-to-d Exchange Scan of their Amino Acid Residues. ACS Comb. Sci. 2013 Nov 11; 15(11): 585–592. PubMed Abstract \| Publisher Full Text\n\nDmytriw AA, et al.: Outcomes of acute respiratory distress syndrome in COVID-19 patients compared to the general population: a systematic review and meta-analysis. Expert Rev. Respir. Med. 2021; 15(10): 1347–1354. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRosenfeld Y, Lev N, Shai Y: Effect of the Hydrophobicity to Net Positive Charge Ratio on Antibacterial and Anti-Endotoxin Activities of Structurally Similar Antimicrobial Peptides. Biochemistry (Mosc). 2010 Feb 9; 49(5): 853–861. PubMed Abstract \| Publisher Full Text\n\nHancock REW, Sahl H-G: Antimicrobial and host-defense peptides as new anti-infective therapeutic strategies. Nat. Biotechnol. 2006 Dec; 24(12): 1551–1557. Publisher Full Text\n\nBrogden KA: Antimicrobial peptides: pore formers or metabolic inhibitors in bacteria? Nat. Rev. Microbiol. 2005 Mar; 3(3): 238–250. PubMed Abstract \| Publisher Full Text\n\nNguyen LT, Haney EF, Vogel HJ: The expanding scope of antimicrobial peptide structures and their modes of action. Trends Biotechnol. 2011 Sep; 29(9): 464–472. PubMed Abstract \| Publisher Full Text\n\nChan DI, Prenner EJ, Vogel HJ: Tryptophan- and arginine-rich antimicrobial peptides: Structures and mechanisms of action. Biochim. Biophys. Acta Biomembr. 2006 Sep; 1758(9): 1184–1202. PubMed Abstract \| Publisher Full Text\n\nSalama A: peptide raw data. [Dataset]. figshare. 2023. Publisher Full Text" }	[ { "id": "235412", "date": "25 Jan 2024", "name": "Yingxia Zhang", "expertise": [ "Reviewer Expertise To find and design bioactive peptides with antimicrobial and anti-inflammatory activities." ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIn the manuscript \"Bactericidal activity of newly synthesized antimicrobial peptides against methicillin-resistant staphylococcus aureus and biofilm-forming methicillin-resistant staphylococcus aureus\", the authors created an ultra-short peptide that was characterized for its antibacterial activity. The peptide killed MRSA and exhibited low toxicity to red blood cells. The manuscript is hard to follow at times and would benefit from a thorough review for typos and English grammar. And some issues seriously need to be addressed before it can be recommended for publication.\nIntroduction:\nSome typos should be corrected, for example “Staphylococcus aureus” and “S. aureus” should be italic. The full name of “MRSA” should be given when it mentioned at the first time. Then the abbreviation can be used. So does “Staphylococcus aureus”.\n\nMethods:\nSynthesis of ultra-short antimicrobial peptides (USAMPs): “the column was eluted with acetonitrile/H2O-TFA gradient at a flow rate of 1.0 mL/min. and validated using mass spectrometry and electrospray ionization mass spectrometry (ESIMS)13 The absorbance was at λ = 214 nm, the solvent which was use is dichloromethane.” Typos and English grammar errors were appeared. Determination of the minimum inhibitory & bactericidal concentrations (MIC/MBCs): “We used sterile 96-well microtiter plates to determine the minimum inhibition and bactericidal concentrations (MIC/MBC), we used the microbroth dilution method for the later determination. Muller–Hinton broth (MHB) was used as a revival growth medium for the staphylococcus aureus, it was also used as the main broth media in determining the MIC.” English grammar errors were appeared.\n\nResults: All the results were depicted too simply to understand.\nIs there any evidence that ferulic acid can increase the peptide hydrophobicity? In figure 2, what cells and the concentration of the peptide and antibiotics were used? The unit of Y-axis is CFU/ML, so is it MRSA? But why the peptide and antibiotics didn’t show antibacterial activity. And the method of cell viability was not mentioned in Methods. The author mentioned in Methods that MRSA (ATCC 33591) as a biofilm forming bacteria and MRSA (ATCCBAA-41) as a planktonic strain were used. But in Results, only the antibacterial activity of MRSA (ATCCBAA-41) were described. In Methods, Biofilm activity assay was mentioned. But I didn’t see any results.\n\nDiscussion:\nThe author mentioned “This peptide mechanism of action involved the ability of the peptide to attack the cell wall and make pores in it, which causes cell lysis and death”. There has no experimental evidence in the manuscript to support this conclusion. The FIC≤ 0.5 was interpreted as synergistic activity. In table 2, the FIC of the peptide and vancomycin was the lowest (0.5), why the author claimed “the results showed a significant increase in effectiveness with Levofloxacin, Chloramphenicol, Rifampicin, Clarithromycin and Doxycycline”?\n\nIs the work clearly and accurately presented and does it cite the current literature? No\n\nIs the study design appropriate and is the work technically sound? No\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? No\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [] }, { "id": "250211", "date": "14 Mar 2024", "name": "Hui-min Neoh", "expertise": [ "Reviewer Expertise antimicrobial resistance in MRSA" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nBactericidal activity of newly synthesized antimicrobial peptides against methicillin-resistant staphylococcus aureus and biofilm-forming methicillin-resistant staphylococcus aureus\nAccording to the authors in the manuscript abstract, the manuscript was supposed to describe work performed in synthesizing USAMPS and testing of the peptides' effectiveness in MRSA killing and biofilm eradication via determination of MIC, MBC and FIC. Synergism of the USAMPS and antibiotics in MRSA killing and also biofilm eradication was also supposed to be explored. Toxicity of the USAMPS towards red blood cells was to be investigated.\nThere are interesting data obtained by the authors especially on the apparent reduction in MIC and MBC readings of antibiotics when coupled with the synthesized USAMPs. However, no data about biofilm experiments can be found in the manuscript. There are also concerns in other areas of the manuscript, as stated in below comments:\n1) Authors can expand the introduction and include more information about antimicrobial peptides and USAMPS. Why were tryptophan and lysine chosen for the synthesis - what was the basis? Reference for the methodology used should be provided. Figures depicting the synthesis will be good to help readers understand the synthesis process.\n2) Methodology - biofilm activity assay: was the inoculum 107 CFU/mL? or was it 10*7 CFU/ml? Similar with 106 CFU/mL in MIC/MBC determination.\n3) \"We also diluted our peptide in different concentrations and in separate 96-well microtiter plates...\". USAMPs concentration in MIC/MBC and checkerboard assay should be defined.\n4) concentrations of antibiotics should preferably be in ug/ml or mg/L (as in EUCAST and CLSI) instead of uM.\n5) will be good to give a brief description of the erythrocyte hemolytic assay, even though a reference of the method has been provided. Was the experiment carried out in exactly the same way as the reference?\n6) Statistical analysis – Authors mentioned that “all data-generating experiments were conducted three times” and used ANOVA for analysis. This is not correct, as ANOVA is used to analyse difference between groups. I supposed the authors performed calculations to obtain the means from the experiment triplicates, and not ANOVA.\n7) What is the KWKWKW-NH2 peptide? Was this the name given to the USAMPs? It should be properly defined in the manuscript.\n8) Description about the experiment performed to obtain “viable cell counts” in Figure 2 should be added into the manuscript. What was the rationale of this experiment? What were the concentrations of the antibiotics and USAMP used? What was the condition of the experiment?\n9) No results provided about biofilm assays though the methodology was described.\n10) Will be good to include figures from the erythrocyte hemolytic assay in the manuscript.\n11) Will be helpful for the authors to engage English editing for the manuscript.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNo\n\nAre all the source data underlying the results available to ensure full reproducibility? No source data required\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1369
https://f1000research.com/articles/12-1365/v1	18 Oct 23	{ "type": "Research Article", "title": "New 3-aminorhodanine derivatives: Synthesis, characterization, docking study and biological activates", "authors": [ "Noor Alhuda Dakhel Khalaf", "Hiba Ali Hasan", "Karima Fadhil Ali", "Hiba Ali Hasan", "Karima Fadhil Ali" ], "abstract": "Background: Epidermal growth factor receptor (EGFR) is seen in almost all cases of non-small cell lung cancer (NSCLC). As a result, targeting EGFR has become an attractive subject of research in fields such as antitumor research and the most important strategy for cancer treatment, in particular, in the treatment of non-small-cell lung malignancies using EGFR-targeting therapies.\nMethods: For this purpose, novel N- and 5- disubstituted aminorhodanine derivatives were synthesized by Schiff base and using a Knoevenagel condensation approach over two steps of reactions. Then, rhodanine derivatives were analyzed their cytotoxic effect on A549 lung cancer and Hdfn normal cell and compared the analysis result with erlotinib (anticancer drug) as standard to determine their activities on cancer cell and toxicity or safety on normal cell.\nResults: Synthetic compounds (2a1-2, 2b1-2) showed different effects at 24, 48 and 72h. The higher anticancer effect was seen for 2a2 and 2a1with IC50 10 μg/mL and 32.59 μg/mL, respectively, at 24h and 72 h. Also, they show high anticancer potency at 72h with low effect and high safety on human normal cell.\nConclusions: Developing a new series of rhodanine derivatives and evaluating their anticancer activity is the main goal of the study. In silico and in vitro antitumor investigations for newly synthesized compounds with different properties and functional group in chemical structure revealed different activities against lung cancer cell. Compounds 2a1-2 which contain pyridine ring in their chemical structure showed more potent effect than the derivative that bearing furyl ring (2b1-2).", "keywords": [ "Rhodanine", "lung cancer", "EGFR mutation." ], "content": "Introduction\n\nCancer remains one of the main health issues globally.1 As reported by the World Health Organization (WHO), cancer is the most significant cause of death in the world, accounting for over 10 million fatalities in 2020 and by 2040, 29.5 million additional cancer cases are expected to be reported. Lung cancer can be classified into two types2,3: Non-small cell lung cancer (NSCLC) which accounts for 80% to 85% of all lung cancers,4–7 and SCLC (small cell lung cancer) which accounts for 10–15% of all lung malignancies.8 This kind of lung cancer grows and spreads more rapid than NSCLC. Tobacco smoke exposure causes a high mutations and is associated with frequent DNA duplicaions and deletions.9–11 The existence of gene mutations or rearrangements is determined by molecular analysis; these molecular changes include epidermal growth factor receptor (EGFR),12,13 gene mutations, anaplastic lymphoma kinase (ALK) gene rearrangements.14,15 First-generation EGFR tyrosine kinase inhibitors (TKI) that bind to the kinase domain of EGFR reversibly, such as gefitinib and erlotinib, are frequently used to treat NSCLC patients with activating EGFR mutations.16–23 gefitinib/erlotinib resistance has emerged in more than half of previously treated tumors.24,25\n\nHeterocyclic compounds are often employed in medicinal chemistry and are crucial to the pharmaceutical industry’s search for novel bioactive molecules. Rhodanine and its derivatives 2, 4-thiazolidinedione (TZD), whose structure includes a thiazolidine nucleus and a carbonyl group on the fourth carbon, are examples of five-membered heterocyclic compounds having a variety of biological functions (Figure 1).26,27\n\nRhodanine has been recognized as an important type of heterocyclic chemicals in the development of drugs in the last few years.28–30 Several natural and synthesized biological active substances having a rhodanine core are employed as therapeutic treatments, such as anti-inflammatory,31 anti-bacterial,32 anti-tuberculosis,33 anti-diabetic,34 hypnotic,35 anti-parasitic,36 anti-HIV37 and carbonic anhydrase inhibitor agents.38 Anti-cancer properties of rhodamine-containing derivatives are shown in Figure 2.39 Furthermore, the complex-rich chemistry of rhodanine, as well as the significance of heterocyclic compounds, support the future development of synthetic techniques in the chemical industry.40,41\n\nIn order to offer higher efficacy of treatment, and to pass the resistance to drug resulting from mutations or inappropriate binding, more effective treatments are required. Moreover, little research has been conducted on the synthesis of amino-rhodanine derivatives. For these purposes, in the present study we synthesized and analysed the anticancer activity of novel N-substituted rhodanine derivatives which are of varying type.\n\n\nResults and discussion\n\nPositions 3 and 5 in the rhodanine ring are highly reactive and play an important role in the design as well as the creation of new drug-like compounds.46,47 So, in the synthesis of N-substituted rhodamine, the regioselectivity of reaction is important to first obtain N-substituted compound, which occurs due to varying factors such as temperature, duration, catalysts and solvents to get the best reaction conditions. We started with 3-amino-2-thioxothiazolidin-4-one, which is commercially available and aldehydes, as shown in Figure 1, the reaction is region-selective at position 3 of 3-aminorhodanine and involves Schiff base formation (Scheme 1). The ideal parameter for the reaction to obtain compound 1a is using acetic acid as a catalyst and using methanol as a solvent, while the high yield of compound 1b can be obtained without using any catalyst.1,48,49\n\nCompounds 2a1-2, 2b1-2 were synthesized via Knoevenagel condensation between aldehyde and the final stages of the first step (1a, 1b), which are N–substituted rhodanine, using NH4OH/NH4Cl as a catalyst and reflux for 6-8 hrs. Condensation in alcoholic solution in an environment of NH4OH/NH4Cl always resulted in 5-[(aryl) alkylidene]-3-aminorhodanine50; Petlichnaya and colleagues obtained similar results (1967 and 1970).48,49 In the synthesis procedure, compounds were highly affected by increasing heat, so the used reflux temperature was not more than 50°C- 60°C.\n\nThe structures of the resulting products were examined using spectral data analysis; the Fournier transform infrared (FTIR) spectrum of compounds 1a and 1b showed characteristic absorption bands at 1721 cm-1 and 1737 cm−1, respectively, corresponding to the C=O group of thioxothiazolidin ring. The C=N stretching bands appeared at 1605 cm−1for compound 1a and at 1614 cm−1for 1b. Another two bands appeared at 1418 cm−1 for 1a as well as 1545 cm−1and 1467 cm−1 for 1b, which were attributed to C=C stretching aromatic. Moreover, C=S group appeared at 1104 cm−1 and 1084 cm−1 for 1a and 1b compounds, respectively.\n\nThe 1H-NMR spectrum identified a proton resonating as a singlet at δ = 7.83 ppm for 1a and 8.58 ppm for 1b for azomethine group (CH=N), which confirmed the imine group formation. The most significant singlets at δ = 4.24 ppm and 4.37 ppm for compounds 1a and 1b, respectively, correspond to the 2H proton at C5 of thioxothiazolidine ring, explaining the reaction that occurs in 3-amino position of 3-aminothioxothiazolidine core and formation of Schiff base. In compound 1a doublet signals appeared at 7.59 ppm for H-2, 6 and 8.75 ppm for H-3, 5 of the pyridine ring. 1H-NMR (proton nuclear magnetic resonance) spectrum of 1b showed signals of furan ring as doublate of doublat for H-4, doublate for H-5 and doublate for H-3 at 6.81, 7.39 and 8.10 ppm, respectively.\n\nThe 13C-NMR analysis identified Schiff base carbon which appeared at 158.2 ppm and 147.5 ppm for 1a and 1b compounds, respectively. The thioxothiazolidine ring C5 resonated at 33.8 ppm in 1a and 35.6 in 1b, and the carbonyl groups appeared at 164.0 ppm and 170.1 ppm for 1a and 1b, respectively. In contrast, the C=S group resonated at 188.1 ppm for 1a and at the 197.8 ppm for 1b. The 13C-NMR spectrum demonstrated another distinctive signal such as 124.2 ppm for C-2, 6, 140 ppm for C-1 and 151.1 ppm for C-3, 5 which represent the carbon atoms of pyridine ring in 1a compound, while 113.6 ppm for C-4, 121.8 ppm for C-5, 148.8 ppm for C-3 and 158.7 ppm for C-1 which refer to carbon atoms of furfural in compound 1b.\n\nThe FTIR spectrum of compounds 2a1 and 2a2 showed characteristic absorption bands at 3045 cm−1 and 2933 cm−1 corresponding to =C-H sp2, -C-H sp3 of new compound 2a1 whereas compound 2a2 showed these peaks in the 3174 cm−1 and 3037 cm−1 regions. Two bands at 1737 and 1678 cm−1 corresponded to the C=O of thioxothiazolidin ring in compounds 2a1 and 2a2, respectively. FTIR spectra also showed distinctive peaks in 1593 and 1595 cm−1 regions, corresponding to C=N of new compounds 2a1 and 2a1, respectively. A C=S group appeared at 1097 cm−1 for 2a1 and at 1170 cm−1 for 2a2.\n\nThe 1H-NMR spectrum of compounds displayed the most important singlets at the 8.12 ppm for 2a1 and 8.17 ppm for 2a2 of the azomethine group (-N=CH). Also, signals representing the α-hydrogen appears as singlets at 7.94 and 7.99 ppm for 2a1 and 2a2, respectively.\n\nThe 13C-NMR (carbon nuclear magnetic resonance) spectra for the same derivatives showed the most significant peaks at 143.4 ppm of C-α for 2a1 and 141.2 ppm for 2a2. Also, compound 2a1 showed 153.6 ppm for –N=CH carbon while this signal appeared at 158.2 ppm for 2a2 derivative.\n\nThe FTIR spectrum of compounds 2b1 and 2b2 show peaks in 2951, 2841, 1703, 1591 and 1081 cm−1 for compound 2b1 corresponding to =C-H sp2, -C-H sp3, C=O, C=N and C=S while these signals appeared at 3120, 2918, 1710, 1610 and 1057 cm−1 in 2b2.\n\nThe 1H-NMR spectrum identified protons resonating as singlet at 8.14 ppm for compound 2b1 and at 8.26 ppm for compound 2b2 for the azomethine group (-N=CH) which confirmed the imine formation. Another distinct 1H-NMR signal appeared as a singlet at 7.68 ppm and 7.58 ppm represents the α-hydrogen of 2b1 and 2b2, respectively.\n\nThe 13C-NMR spectra for 2b1 and 2b2 showed the most significant peaks at 143.3 ppm of C-α of methine group for 2b1 and 142.6 ppm for 2b2. Also, compound 2b1 showed a signal at 132.9 ppm for –N=CH carbon, while this signal appeared at 134.5 ppm for 2b2 derivative.\n\nAs a result, all of these distinguished bands and signals identify the functional groups present in each particular derivative and explain the formation of new scaffolds.\n\nDocking of co-crystallized ligands was performed, and the most active ligand was shown to be at the receptors’ expected catalytic active region. As shown by Figure 3, both compounds 2a2 and 2a1 occupied the same binding site as erlotinib.\n\nCompound 2a2 coordinated with THR766 amino acid of receptor with a binding score of 2.856 Å and the estimated docking scores are shown in Table 1. The score showed a high affinity of 2a2 and a binding plp fitness of 72.71 Kcal/mmol. This result made the ligand more active against their receptor compared to standard inhibitor erlotinib. Moreover, ligand 2a1 showed good binding to the receptor compared with the standard ligand as clarified in Figure 3. It showed a 70.29 Kcal/Mol plp fitness and the binding involved two hydrogen bonds with THR766, THR830 with the amino acid of the receptor protein. While the docked poses of compound 2b2 showed a binding affinity slightly greater than the standard at about 68.88 Kcal/Mol plp fitness and 2.947 Å. On the other hand, compound 2b1 exhibited lower values than Erlotinib plp fitness with a binding score 2.696 Å of the receptor. The optimal position of the ligand was determined to be the best-scored ligand into catalytic active sites. Thus, the synthesized rhodanine derivatives appeared to be capable to access the target’s active site and position (through specific connections) into an optimal state for nucleophilic attacks by the amino acid’s (THR766, THR830) residue of the EGFR receptor.\n\nThe anti-tumor efficacies of the synthesized compounds 2a1, 2a2, 2b1 and 2b2 were examined in vitro using an A549 cell line. The response was measured at 24, 48 and 72 h. It is described in Table 2 and Figures 4-6. The responses were different at different times: some compounds showed an optimal effect during the first 48 h, (2a1 and 2a2), while another compound took more time to have a high anti-tumor effect (2b1). All compounds were examined using the same cancer cell line (A549). As shown in Figure 4, compound 2a2 (IC50 10.8 μg/mL) was much more potent in inhibiting lung cancer cells than erlotinib during the first 24 hours. Furthermore, as seen in Figure 5, compound 2a1 exhibited excellent anticancer activity at 48 hours and was more potent than erlotinib with IC50 (32.59 μg/mL), with a higher safety when its cytotoxicity was examined on HdFn normal cell and showed an IC50 of about 154.4 μg/mL, which made it safer than erlotinib with an IC50 of 122.8 μg/mL. Also, after examination on human normal cell line, compound 2a1 showed high antitumor activity at 72 h and excellent safety as displayed in Figure 6.\n\nIn contrast, other compounds such as 2b1 and 2b2 were less efficient on the cancer cell line (A549). Thus, they showed a moderately lower IC50 value (67.57 μg/mL and 72.02 μg/mL, respectively) with their optimal activity starting after 72 hours. In terms of safety, compared with erlotinib, these two compounds exhibited high safety in their cytotoxicity examination on HdFn normal cell Figure 6. These examinations results indicated significant outcomes for rhodanine derivatives, which might become a candidate for lung cancer treatment.\n\n\nMethods\n\nAll chemicals and solvents were commercially available from Sigma-Aldrich or Merck. FT-Infrared spectra were analyzed by 8400 Shimadzu KBr spectrophotometer, 1H-NMR and 13C-NMR (300 and 75 MHz) spectra were analyzed using a Bruker spectrometer. The melting points were measured without correction using an electro-thermal melting point apparatus. The reaction progress was monitored by thin-layer chromatography (TLC, 0.25 mm-thick precoated silica Merck plates).\n\nGeneral synthesis of N-substituted-rhodanine derivatives (1a and 1b)\n\nTo a solution of 3-amino-2-thioxothiazolidin-4-one (1mmol) in 2.5 mL methanol, aldehydes (1.2 mmol) in 1.5 mL methanol were added slowly. The reaction mixtures were stirred at room temperature with glacial acetic acid as a catalyst for compound 1a and without any catalyst for compound 1b for 4 to 6 h, and were monitored by TLC. After that, the two mixture products were recrystallized from methanol. After recrystallization, N-substituted-rhodanine derivatives were obtained as follows:\n\n(E)-3-((pyridin-4-ylmethylene) amino)-2-thioxothiazolidin-4-one (1a)\n\n\n\nThe product was obtained as an off-white powder (200 mg, 84% yield); m.p: 168°C-170°C; Chemical formula: C9H7N3OS2. FTIR (KBr, cm−1) vmax: 3296 (=C-H sp2 stretching), 2814 (-C-H sp3 stretching), 1721 (C=O stretching), 1605 (C=N stretching), 1418 (C=C aromatic stretching), 1293 (C=S stretching), 1208 (=C-N stretching), 1104 (C=S stretching), 1063 (-C-N stretching), 869 (C-S stretching) 784 (C-H aromatic out of plane bending). 1H-NMR (300 MHz, DMSO-d6) δ ppm 4.24 (s, 2H, CH2), 7.59 (d, 2H, H-2, 6), 7.83 (s, 1H, CH=N), 8.75 (d, 2H, H-3, 5). 13C-NMR (75 MHz, DMSO-d6): 47.2 (CH2), 124.2 (C-2, 6), 140.3 (C-1), 151.1(C-3, 5), 158.2 (CH=N), 164.0 (C=O), 188.1 (C=S).\n\n(E)-3-((furan-2-ylmethylene) amino)-2-thioxothiazolidin-4-one (1b)\n\n\n\nThe product obtained was a yellow glittery crystal (165 mg, 73% yield); m.p: 130°C-132°C; Chemical formula: C8H6N2O2S2. FTIR (KBr, cm−1) vmax: 3128 (=C-H sp2 stretching), 2928 (-C-H sp3 stretching), 1737 (C=O stretching), 1614 (C=N stretching), 1545 and 1467 (C=C aromatic stretching), 1390 (C=S stretching), 1228 (C-O stretching), 1084 (C=S stretching), 1016 (C-N stretching), 825 (C-S stretching), 767 (C-H aromatic out of plane bending). 1H-NMR (300MHz, DMSO-d6): δ 4.37 (s, 2H, CH2), 6.81 (dd, 1H, H-4), 7.39 (d, 1H, H-5), 8.10 (d, 1H, H-3) 8.58 (s, 1H, CH=N). 13C-NMR (75 MHz, DMSO-d6): 35.2 (CH2), 113.6 (C-4), 121.8 (C-5), 147.5 (CH=N), 148.8 (C-3), 158.7 (C-1), 170.1 (C=O), 197.0 (C=S).\n\nSynthesis of 5-substituted rhodanine derivatives 2a1, 2a2 and 2b1, 2b2 (final compounds)\n\nTo a warm solution of 1 mmol of compound (1a or 1b) in 3 mL methanol, benzaldehyde derivatives (1.2 mmol) were dissolved in 2 mL methanol mixed with 0.2 mL. NH4OH and 0.1g of NH4Cl dissolved in 0.2 mL water were added slowly and refluxed at 50°C with stirring for 6-9 h.48 The reaction mixture was monitored by TLC. The products of 5-substituted rhodanine derivatives recrystallized using a mixture of water and methanol and were obtained as follows:\n\n(E)-5-(4-hydroxy-3-methoxybenzylidene)-3-((E)-(pyridin-4-ylmethylene)amino)-2-thioxothiazolidin-4-one (2a1)\n\n\n\nThe product obtained was a brown solid (300 mg, 83% yield); m.p: 105°C-107°C; Chemical formula: C17H13N3O3S2. FTIR (KBr, cm−1) vmax: 3045 (=C-H sp2 stretching), 2933 (-C-H sp3 stretching), 1737 (C=O stretching), 1593 (C=N stretching), 1512 (C=C aromatic stretching), 1406 (C-O stretching), 1278 (=C-N stretching), 1097 (-C-N and C=S stretching), 812 (C-S stretching). 1H-NMR (300 MHz, DMSO-d6) δppm 3.82 (s, 3H, OCH3), 4.91 (s, 1H, OH), 7.02 (d, 1H, H-5′), 7.38 (s, 1H, H-2′), 7.42 (d, 1H, H-6′), 7.94 (s, 1H, H-α), 7.97 (d, 2H, H-2, 6), 8.12 (s, 1H CH=N), 8.30 (d, 2H, H-3, 5).13C-NMR (75MHz, DMSO-d6) δppm 56.2 (OCH3), 111.0 (C-2′), 115.6 (C-5′), 115.8 (-C= of thioxothiazolidin ring), 115.9 (C-2, 6), 122.8 (C-6′), 129.7 (C-1′), 143.39 (C-α), 148.4 (C-1), 148.6 (C-3′), 149.3 (C-3, 5), 150.4 (C4′), 153.6 (CH=N), 166.5 (C=O), 191.5 (C=S).\n\nN-(4-((E)-(4-oxo-3-((E)-(pyridin-4-ylmethylene)amino)-2-thioxothiazolidin-5-ylidene)methyl)phenyl) acetamide (2a2)\n\n\n\nThe product obtained was a dark brown solid (300 mg, 78% yield); m.p; 98°C-100°C; Chemical formula: C18H14N4O2S2. FTIR (KBr, cm−1) vmax: 3174 (=C-H sp2 stretching), 3037 (=C-H sp3 stretching), 1678 (C=O stretching), 1595 (C=N stretching), 1523 (C=C aromatic stretching), 1410 (C=S stretching), 1317 (=C-N stretching), 1170 (C=S stretching), 1093 (C-N stretching), 833 (C-S stretching). 1H-NMR (300 MHz, DMSO-d6) δppm 2.08 (s, 3H, CH3), 7.38 (s, 1H, N-H), 7.61 (d, 2H of H-3′, 5′), 7.71 (d, 2H of H-2′, 6′), 7.84 (d, 2H of H-2, 6), 7.99 (s, 1H of H-α), 8.17 (s, 1H, CH=N), 8.43 (d, 2H of H-3,5). 13C-NMR (75 MHz, DMSO-d6) δppm 24.6 (CH3), 119.0 (-C= of thioxothiazolidin ring), 121.6 (C-2, 6), 122.6 (C-3′, 5′), 128.4 (C-2′, 6′), 129.3 (C-1′), 139.8 (C-4′), 141.2 (C-α), 142.5 (C-1), 150.4 (C-3, 5), 158.2 (CH=N), 169.0 (C=O), 169.6 (C=O of acetamid group), 192.0 (C=S).\n\n(E)-3-((E)-(furan-2-ylmethylene)amino)-5-(E)-(4-hydroxy-3-methoxybenzylidene)-2-thioxothiazolidin-4-one (2b1)\n\n\n\nThe product obtained was a brown solid (300 mg, 83% yield); m.p: 85°C-87°C; Chemical formula: C16H12N2O4S2. FTIR (KBr, cm−1) vmax: 2951 (=C-H sp2 stretching), 2841 (-C-H sp3 stretching), 1703 (C=O stretching), 1591 (C=N stretching), 1460 and 1431 (C=C aromatic stretching), 1381 (C=S stretching), 1290 (C-O stretching), 1081 (C=S stretching), 1022 (C-N stretching), 819 (C-S stretching). 1H-NMR (300 MHz, DMSO-d6) δppm 3.83 (s, 3H, OCH3), 5.94 (s, 1H, OH), 6.77 (d, 1H, H-6′), 7.04 (dd, 1H, H-4), 7.27 (d, 1H, H-5), 7.48 (d, 1H, H-5′), 7.60 (s, 1H, H-2′), 7.68 (s, 1H, Hα), 7.94 (d, 1H, H-3), 8.14 (s, 1H, CH=N). 13C-NMR (75 MHz, DMSO-d6) δppm 56.2 (OCH3), 112.8 (C-2′), 113.3 (C-4), 115.6 (-C= of thioxothiazolidin ring), 116.0 (C-5′), 120.2 (C-5), 122.8 (C-1′), 126.0 (-C= of C-6′), 132.9 (CH=N), 143.3 (Cα), 145.4 (C-3), 148.5 (C-4′), 149.2 (C-1), 150.5 (C-3′), 169.6 (C=O), 191.4 (C=S).\n\n(E)-5-(4-(tert-butyl)benzylidene)-3-((E)-(furan-2-ylmethylene)amino)-2-thioxothiazolidin-4-one (2b2)\n\n\n\nThe product obtained was a brown solid (250 mg, 70% yield); m.p: 112°C-114°C; Chemical Formula: C19H18N2O2S2. FTIR (KBr, cm−1) vmax: 3120 (=C-H sp2 stretching), 2918 (-C-H sp3 stretching), 1710 (C=O stretching), 1610 (C=N stretching), 1541(C=C aromatic stretching), 1232 (C-O stretching), 1057 (C=S stretching), 1018 (C-N stretching), 817 (C-S stretching). 1H-NMR (300 MHz, DMSO-d6) δppm 1.33 (s, 9H, 3×CH3), 5.42 (t, 1H, H-4), 7.11 (d, 1H, H-5), 7.47 (d, 2H, H-3′,5′), 7.58 (s, 1H, Hα), 7.67 (d, 2H, H-2′,6′), 7.89 (d, 1H, H-3), 8.26 (s, 1H, CH=N). 13C-NMR (75 MHz, DMSO-d6) δppm 31.3 (3×CH3), 35.2 (C (CH3)3), 113.1 (C-4), 117.4 (C= of thioxothiazolidin ring), 118.1 (C-5), 126.2 (C-3′, 5′), 128.5 (C-2′, 6′), 133.8 (C-1′), 134.5 (CH=N), 142.6 (Cα), 144.4 (C-3), 149.4 (C-1), 150.1 (C-4′), 169.4 (C=O), 196.0 (C=S).\n\nAll synthesized compounds were analyzed for their activity against lung tumor using a MTT(3-(dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide) assay on A549 lung cancer cell line. The cytotoxic effect of the newly synthesized final compounds were also examined on HdFn normal cells to determine their safety on human normal cells. The antitumor activity and safety of these compounds were compared with clinically used antitumor (TKI) drug erlotinib, which is used as a standard reference. The evaluation of 3-aminorhodanine derivatives was carried out using different concentrations from 25 μg/mL to 400 μg/mL.51 The inhibitory concentrations (IC50, μg/mL) were measured at 24, 48, and 72 h after application of our new compounds (2a1-2, 2b1-2) and compare the viability of cancer cells within non treated normal cell.\n\nThe molecular docking study was performed using CCDC (Cambridge Crystallographic Data Centre), fully licensed Genetic Optimization Ligand Docking (GOLD) Suite (v.5.7.1)52,53 The protein (4HJO) of EGFR crystal structure data was obtained from the Protein DataBank.54 The protein structure and ligands were prepared and corrected before the docking process. Preparation was done by removing of all ligands and water molecules that were not involved in the binding site. energy was calculated and minimized using MM2 minimize and MM2 dynamic option. The receptor was compiled before the docking process by adding polar hydrogen atoms to achieve specific ionization and tautomeric positions of amino acid residues. The binding sites were identified using the co-crystallized ligand erlotinib. All of the protein residue characterized within 10 A° of the typical ligands for the docking process. The piecewise linear potential (plp) function was used in the scoring function. Finally, the output was downloaded as a mol.2 file.\n\nIn the docking study site scores were computed to assess whether binding ligands are catalytically activated sites, and calculating the binding energies of novel compounds with the protein (EGFR), including the H-bond, in addition to other bonds that ligands developed with the receptor amino acids in the active site.\n\n\nConclusions\n\nA new series of rhodanine derivatives consisting of significant aromatic groups, 2a1-2 and 2b1-2, were designed, synthesized and investigated for their potential anti-tumor effects. Some of these new rhodanine derivatives showed the maximum anti-tumor effect at 200 μg/mL concentration against A549 lung cancer cell line. However, rhodanine derivatives that contained pyridine were more efficient toward lung cancer cells; this result was confirmed by the in vitro anti-tumor analysis and docking studies, in which the computed scores revealed that anticipated binding sites displayed catalytic active site features and that these compounds had an anti-tumor effect when compared to the reference medication erlotinib, which was classified based on the plp fitness.\n\nCompounds 2a1 and 2a2 were more potent than standard erlotinib and showed excellent activity at different times. Thus, the new synthesized rhodanine compounds showed a high potential for use as innovative therapeutic agents in the treatment of lung cancer.", "appendix": "Data availability\n\nZenodo: Data of compound 2a1-2, 2b1-2: https://doi.org/10.5281/zenodo.8284738\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgments\n\nAll Authors would like to thank Mustansiriyah University (www.uomustansiriyah.edu.iq), college of pharmacy, Baghdad-Iraq for its support in the present work.\n\n\nReferences\n\nFitzmaurice C, Dicker D, Pain A, et al.: The global burden of cancer 2013. JAMA Oncol. 2015; 1(4): 505–527. PubMed Abstract \| Publisher Full Text\n\nDeVita VT, Lawrence TS, Rosenberg SA: Cancer of the skin: cancer: principles & practice of oncology. Lippincott Williams & Wilkins; 2015.\n\nSemenova EA, Nagel R, Berns A: Origins, genetic landscape, and emerging therapies of small cell lung cancer. 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Protein Data Bank. http" }	[ { "id": "218244", "date": "22 Nov 2023", "name": "Qamar Uddin Ahmed", "expertise": [ "Reviewer Expertise Pharmaceutical Organic Chemistry" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors have done interesting research in the field of synthetic organic chemistry and tried to synthesize novel N- and 5- disubstituted aminorhodanine derivatives which were synthesized by Schiff base and Knoevenagel condensation approach. Then, resultant derivatives were analysed for their cytotoxic effect on A549 lung cancer and HDFN normal cell. Results confirmed an in vitro anticancer effect of some derivatives.\nHowever, an in vivo anticancer effect should have also been evaluated to further confirm the anticancer potential of synthesized derivatives. Language is quite weak in some places and typos do exist and must be rectified. All scientific terms must be italicized.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "350727", "date": "20 Jan 2025", "name": "Sinan Bayindir", "expertise": [ "Reviewer Expertise Organic synthesis and biological activity" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis article investigates the derivatization of rhodamine, an important heterocyclic compound, and explores the anticancer properties of these derivatives. However, several aspects require further consideration:\nLanguage and Clarity: The writing style could benefit from a thorough review and refinement to enhance clarity and conciseness. Scope of Derivatization: The current number of derivatives synthesized appears limited. Expanding the scope of derivatization would significantly strengthen the study. Biological Evaluation: Given the limited number of derivatives, the biological studies should be expanded to include in vivo investigations to provide a more comprehensive understanding of the anticancer potential. Literature Review: Two previously published articles (Yararli K,et al., 2023 [Ref-1] and Ozer E, et al., 2022 [Ref-2]) describe innovative syntheses of related derivatives. These articles should be reviewed and appropriately cited within the manuscript. Molecular Representation: All molecular structures should be redrawn in the ACS format using ChemDraw for consistent and professional presentation.\"\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1365
https://f1000research.com/articles/12-1363/v1	18 Oct 23	{ "type": "Systematic Review", "title": "The therapeutic effect and safety profile of extracorporeal shockwave therapy in treatment of chronic prostatitis/chronic pelvic pain syndrome: a systematic review", "authors": [ "Mikha mikha", "Isaac Ardianson Deswanto", "Isaac Ardianson Deswanto" ], "abstract": "Background: Extracorporeal Shockwave Therapy (ESWT) has been indicated to relieve local perineal symptoms caused by Chronic Prostatitis/Chronic Pelvic Pain Syndrome (CP/CPPS). Current research has examined the efficacy of ESWT in CPPS patients. Different types of energy generators for ESWT lead to development of different clinical protocols for treatment of CP/CPPS. Therefore in this review, we aimed to compare the clinical protocol, efficacy and safety profile of all these different ESWT machines in CP/CPPS treatment. Methods: A systematic literature search of 3 search engines (PubMed, Scielo, and Science Directs) was undertaken using the following keywords: Chronic Prostatitis, Chronic Pelvic Pain Syndrome, and Extracorporeal Shockwave Therapy. A systematic review was conducted in accordance with Preferred Reporting Items for Systematic Review and Meta-analysis (PRISMA) guidelines. This review included original studies that evaluated the efficacy and clinical protocol of Extracorporeal Shockwave Therapy and Chronic prostatitis or Chronic Pelvic Pain Syndrome that are fully written in English with full-text articles readily available. This article excluded letters to the editor, reviews, and editorials about prostatitis other than CPPS. Results: The search strategy yielded 8 journals that meet the inclusion and exclusion criteria from all 3 search engines. These 8 studies included 3 different types of energy generators (Piezoelectric, Electropneumatic, and Electromagnetic) with different protocols applied. All 3 types of energy generators of ESWT can effectively decrease all domains of CPSI score within 12 months of follow-up (P-value 0,05). The limitations of this systematic review include the restricted variety of energy generators with the lack of openly registered protocols. Conclusions: In Conclusion, ESWT provides significant improvement in clinical symptoms as compared to oral medications alone. These therapeutic effects are also observed in all different types of energy generators with different clinical protocols with similar safety profiles.", "keywords": [ "Chronic Prostatitis", "Chronic Pelvic Pain Syndrome", "ESWT", "Protocol." ], "content": "Introduction\n\nProstatitis is an inflammation of the prostate gland that may be detrimental to the quality of life, although not life-threatening. The debates about the definition of Chronic Prostatitis (CP)/Chronic Pelvic Pain Syndrome (CPPS) have been ongoing for decades.1 According to the most recent version of the National Institute of Health (NIH) consensus classification, the diagnosis of chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) relies upon the detection of leukocytes in expressed prostatic secretions (EPS), urine post prostatic massage (VB3), or seminal fluid analysis. These criteria doubled as many patients into the category of CP/CPPS as compared to the previous classification system.2 The inherent diversity of the illness gives rise to a significant amount of variation in establishing clinical diagnosis CP/CPPS. According to the most recent NIH classification, CPPS is defined as chronic nonbacterial prostatitis (group III).3 Approximately 5 to 10% of prostatitis have been linked to a bacterial infection. The average prevalence of CPPS varies from 6.9% in North America to 12.2% in Africa, with Europe at 8.6%, Asia at 7.5%, and Australia at 7.5%.2 The remarkably consistent rate across continents suggests that the disease’s development is not influenced by environmental risk factors. Group III of the NIH Classification is subdivided into two subtypes accoeding to the presence of leukocytes in EPS, VB3 and seminal analysis. Lower urinary tract syndrome (LUTS) has frequently been reported as one of the most prominent symptoms in people with CPPS, frequently manifesting as discomfort and difficulties urinating. Therefore, the NIH determined that the scoring system should be based on three large domains (degree of pain, lower urinary tract symptoms, and quality of life) that may be filled out by patients in order to interpret symptom quality into quantitative illness severity.1–4\n\nNumerous etiologies and pathogenic mechanisms for chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) have been proposed. The authors propose a potential involvement of immunological, neurological, endocrine, and psychological aspects. The implications of these findings on the indigenous bacterial flora in the prostate gland, and its role in triggering the inflammatory response, are also considered. Taken as a whole, it is apparent that the prostate cannot be completely attributed as the only origin of symptoms in chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS).4\n\nThere are various medical treatments available for the management of CP/CPPS although unfortunately with inconsistent efficacy. These medications include 5-alpha reductase inhibitor, alpha-receptor blockers, antibiotics, anti-inflammatory medications and analgesics that have been used individually or in combination.3,5 Physiotherapy, botulinum toxin A intraprostatic injection, trigger-point massage, electromagnetic treatment, laser coagulation, invasive neuromodulation, balloon dilation, thermotherapy are examples of alternative therapies.5 Unfortunately, these treatments have not proven universally effective in the treatment of CP/CPPS. Currently, there is no causative or standard treatment for CP/CPPS. As stated in the European Association of Urology’s guideline, individuals with CP/CPPS should be handled in a multidisciplinary and multispecialty settings, considering all of the symptoms. Recently, ESWT has been suggested as a potential treatment for alleviating localized perineal discomfort associated with CP/CPPS.4\n\nESWT has been recommended as an option for alleviating localized perineal discomfort caused by CP/CPPS. Current research has examined the efficacy of perineal ESWT in CPPS patients. ESWT may alleviate pain through multiple pathways. The mechanisms of how ESWT affects pain include hyperstimulating of nociceptors, repairing tissue through revascularization processes, also lowering muscular tone and stiffness.6 Previous research on the application of ESWT to orthopaedic pain disorders proposed the stated processes. Similar mechanisms of action have been hypothesized as pain-altering methods by which ESWT may treat CPPS patients.7 Up until now, there are a number of different ESWT machinery that applies different technology ranging from Focused ESWT (F-ESWT) such as piezoelectric, electrohydraulic as well as electromagnetic and radial ESWT (R-ESWT) such as electropneumatic energy generator.6,7 This different machineries with different types of energy generators leads to different clinical protocols for treatment of CP/CPPS. Therefore, we attempt to evaluate all the protocols for different ESWT machines. Additionally, this review also aimed to compare the clinical efficacy as well as safety profile of all these different ESWT machines in CP/CPPS treatment.\n\n\nMethods\n\nSearch methodology and criterion for selection\n\nThe present systematic review was conducted using the guidelines established by the Preferred Reporting Items for Systematic Review and Meta-analysis (PRISMA). A comprehensive review of the literature was conducted by searching three different databases, which are PubMed, Scielo, and Science Directs. The search was carried out using the terms “Chronic Prostatitis” OR “Chronic Pelvic Pain Syndrome” AND “Extracorporeal Shockwave Therapy”. The keywords used were the same across all databases. The scope of the search was restricted to scholarly articles that were published during the past ten years. The articles included in this review were chosen according to specific criteria for inclusion. These criteria covered articles written in English, articles that examined the effectiveness and clinical procedure of Extracorporeal Shockwave Therapy in relation to Chronic Prostatitis or Chronic Pelvic Pain Syndrome, and original studies that had easily accessible full-text articles. This article excludes other sorts of writings, such as letters to the editor, reviews, and editorials, that discuss prostatitis types other than chronic pelvic pain syndrome (CPPS).\n\nThe two investigators (M and IAD) had conducted the initial screening from the titles and abstracts independently. Subsequently, a comprehensive evaluation was conducted on the whole texts of publications that were considered possibly necessary, as well as those for which a conclusive determination could not be reached just based on the abstract. The search approach employed in this study resulted in a total of 67 findings across the three aforementioned databases. The author screened every abstract to find articles. Consequently, 59 articles were excluded, leaving 8 articles to be included in this review for further investigation.\n\nThe quality of each randomized controlled study was assessed using an instrument by Sterne et al.8 In order to evaluate the potential for bias in experimental research, an assessment was conducted across six domains that pertain to the internal and external validity of these investigations. These domains include the selection of the reported result, measurement of the outcome, missing outcome data, randomization procedure, deviation from intended treatments, and overall bias (Figure 1). A score was assigned to each domain, namely, yes, probably yes, probably no, and no. The studies were categorized into three levels of bias risk, namely low, moderate, and high, according to the overall assessment made by the algorithm for each randomized controlled trial research. In the context of cohort studies, the evaluation encompassed seven distinct categories, namely: participation selection, confounding bias, deviation from intended interventions, intervention classification, outcome measurement, and selection of reported results. A score was assigned to each domain, namely yes, probably yes, probably no, and no. The studies were categorized into four groups based on the algorithm’s comprehensive assessment, indicating varying levels of bias: low, moderate, serious, and critical.\n\nThe data from the chosen articles were extracted by two authors (M and IAD) separately. This extraction process involved utilizing a standardized form to collect information such as the publication year, authors, clinical protocol, efficacy of Extracorporeal Shock Wave Therapy (ESWT), and its impact on chronic prostatitis or chronic pelvic pain syndrome. The collected data was then assessed using the National Institutes of Health Chronic Prostatitis Symptom Index (NIH-CPSI) scoring system. Any inconsistencies in the retrieved data were resolved by discussion and consultation of the original article in order to reach an agreement.\n\n\nResults\n\nOur search strategy gathered 8 journals that meet the inclusion and exclusion criteria from 3 search engines. From the 8 studies that have been included, the article originated from 8 different countries (China, Greece, Iran, Jordan, Montenegro, Saudi Arabia, South Korea, and Taiwan). From the risk of bias assessment, no study was found to have high risk of bias. From the included studies, the main energy generator used in the paper were mostly electromagnetic (EM), 1 study with piezoelectric (PE) and 1 study with electropneumatic (EP). Energy flow density is counted in mJ/mm2 to generalize the measurement. The protocols that had been compared in this review were pulses counted, energy flow density, frequency, duration of treatment and type of energy generator. CPSI score was evaluated as a benchmark for clinical outcome.\n\nAll included studies in this review only recruited men older than 18 year of age that have been diagnosed with CPPS/Group III prostatitis. Despite CPPS varies in 2 subgroups (IIIA and IIIB) most studies only include prostatitis group IIIB and only 2 studies include both IIIA and IIIB group of prostatitis. Baseline CPSI Score for total domain ranges from 25 to 31 points with different follow up timings in each study (ranging from 1 month to 12 months after ESWT treatment).\n\nAs listed in Table 1, the protocol that mostly used was electromagnetic energy generator with 0.25 mJ/mm2 energy flow density, 3000 pulses counted and 3 Hz of frequency once a week. Every study that stated in this review had a significancy in statistic (p<0.05). Electropneumatic protocol used in 1 cohort study that have been published by Ghazi et al. using Roland-Pagani brand from Italy with 2500 pulses and 3 Hz of frequencies that generated 0.25 mJ/mm2 energy in 1 session of ESWT.9 That study conduct 1 session of ESWT per week and lasted for 4 sessions. The result was the ESWT treatment statistically significant to reduce the symptoms of CP/CPPS by decreasing the CPSI score in most domain as the time followed up progressively (p-value<0.05).9 Meanwhile piezoelectric machine was used in only 1 RCT study using Lubisone brand machine from South Korean with 3000 pulses and 3 Hz of frequencies that generate 0.25 mJ/mm2 per session. This study conducted 1 session per week for along 12 sessions and resulted in statistically significant reducing the CPSI score in all domain compared to the placebo (p-value<0.05).10 On the other hand, one paper from Mykoniatis et al. comparing 2 different protocol for ESWT in CP/CPPS. They compared between group of 1 session per week and 2 sessions per week for overall 6 sessions applied and resulted in statistically insignificant for both groups compared (p-value>0.05) but statistically significant compared to the baseline (p-value< 0.05).11\n\nFrom 8 studies that have been yielded, 2 of them are cohort and the rest are clinical trial. 2 cohort studies showed that the clinical improvement were present for all time follow up CPSI scoring regard from the baseline for all domain (p-value<0.05). Otherwise, study from Zhang et al., stated that there was no significancy between the combination of ESWT and oral triple therapy group and oral triple therapy medication group without ESWT intervention.12 All of the study result containing the clinical improvement by CPSI Scoring summarized in Tables 2, 3 and 4.\n\n\nDiscussion\n\nPressure waves are well-established oscillating waves that have The capacity for propagation across various mediums such as gases, liquid and solid. As mentioned before, shockwaves are a form of pressure wave characterized by nonlinearity and a brief duration of rise time, often lasting for a maximum of 10 seconds. The negative and positive phases of shockwaves have distinct outcomes on the interfaces between different tissues characterized by differing densities. During the positive phase, it is possible for high-pressure shockwaves to either impact an interface and undergo reflection or traverse through it and gradually undergo absorption. In the negative phase, commonly mentioned to as the tensile phase, the shockwave induces cavitation at the interfaces of the tissue, leading to the making of air bubbles. Subsequently, the bubbles undergo implosion at a significant velocity, so producing a further series of shockwaves or fluid microjets.13\n\nThere are two distinct categories of ESWT shockwaves: Focused ESWT (FSWT) and Radial ESWT (RSWT). FSWT is distinguished by the creation of a pressure field that converges at certain depths inside particular tissues, resulting in reaching a level of maximal pressure at the adjustable focus. In the context of medical practice, the utilization of both concentrated shock waves and radial pressure waves is seen. Radial pressure waves are sometimes denoted as radial shock waves, however it is important to note that this terminology needs to align with the precise use in the field of physics. Shock waves and pressure waves show differences not only in their generating mechanisms, but also in the physical parameters often employed and the depths of therapeutic tissue penetration they may attain. Meanwhile FSWT treat at certain waves, RSWT apply the linear effect on every layer of the tissue. Therefore, FSWT are proposed to have a role at urologic intervention including CP/CPPS while RSWT are proposed to have a role at musculoskeletal problem.\n\nThe energy generator that utilized to make RSWT are different from FSWT. RSWT utilized electropneumatic (EP) to produce wave meanwhile FSWT utilize 3 type of generators: Electrohydraulic (EH), Electromagnetic (EM), and Piezoelectric (PE)\n\n1. Electropneumatic (EP) sources generate a plasma bubble in close proximity to a para-ellipsoidal reflector with the application of high-energy release between two electrodes immersed in water. The process of plasma expansion initiates the formation of a shock wave, which subsequently undergoes reflection by a reflector and is then directed towards a secondary target tissue focus.\n\n2. The utilization of electromagnetic (EM) sources using either cylindrical or flat coils is also common. In the initial system, a coil is employed to transmit a high-energy pulse in a direction opposite to that of a metallic membrane. The electromagnetic field produced by the coil induces a rapid bending of the membrane, creating pressure waves inside a fluid medium. A lens is capable of concentrating waves, causing them to undergo a steepening process that ultimately results in the formation of a shockwave in close proximity to the focal point. The next electromagnetic producing source comprises a tubular coil and a metallic membrane positioned within a parabolic reflector that is filled with fluid. The presence of a magnetic field induces displacement of the membrane in a direction away from the coil. Following contemplation by the reflector, a radial auditory pulse is generated and directed toward the focal point of the system.\n\n3. Piezoelectric (PE) generators produce shockwaves through the use of a high-energy release, a configuration of PE devices adhered to the interior surface of a cylindrical backing, which is situated within a cavity filled with fluid is utilized. Each component undergoes expansion, generates a propagating pressure pulse that disperses in the direction of the focus area of the given configuration. The concentration of pressure pulses and nonlinear effects in the focal region results in the formation of a shockwave.7\n\nThe production of focused shockwaves in water is facilitated by the comparable acoustic impedance of water and biological tissue. Therefore, the process of reflection is diminished. Electrohydraulic pulse (EP), electromagnetic pulse (EM), and piezoelectric effect (PE) all employ water as a medium for generating intense shockwaves, hence exhibiting a similarity. However, there is a clear distinction in the instant when shockwaves are formed. Electro-pneumatic (EP) generators are capable of producing shockwaves instantly following the spark gap, in contrast to electromagnetic (EM) and piezoelectric (PE) generators which exhibit a delay in the nanosecond range due to wave focalization.13\n\nAlthough the clinical efficacy of the treatment, the underlying mechanism of action of ESWT remains unclear. During the year of 1997, Haupt put up four potential pathways for the stages of tissue response to ESWT.13\n\n1. Physical stage: this stage reveals that the shockwave generates a positive pressure that results in energy absorption, deflection, refraction, and transmission into cells and tissues. Further investigations indicated that ESWT elicits physical phenomena like cavitation, enhanced process of ionization in biological molecules, and its impact on the permeability of cellular membranes by applying a force generated by negative pressure.13\n\n2. The physicochemical stage of ESWT induces the release of biomolecules, including adenosine triphosphate (ATP), which in turn stimulates cellular signaling pathways.13\n\n3. Chemical stage: during this stage, shockwaves induce modifications in the activity of ion channels located in the cellular membrane, as well as the movement of calcium within cells.13\n\n4. Biological stage: Previous research has demonstrated that extracorporeal shock wave therapy (ESWT) has the ability to modulate various biological processes. These include angiogenesis, anti-inflammatory, and promote wound healing through the activation of Wnt3, Wnt5a, and beta-catenin.13\n\nIn CP/CPPS itself, the main therapeutic pathway of ESWT are in both physical and physiochemical phase which hyperstimulate the nociceptor and release the muscle stiffness to alleviate symptoms in all domain of CPSI scoring.7\n\nThe machinery in this systematic review of focused ESWT separated into 3 kind of energy generator (EP, EM, and PE). All of the 3 machine shared the same amount of shockwave energy with different protocols. Thus, every ESWT operator need to know the basic protocol that are usually used in CP/CPPS for each machine regardless the brand of the machine itself. Ghazi et al. utilized Roland Pagani ES.WT four sessions weekly (2500 Pulses, energy flow density of 0.25 mJ/mm2, and 3Hz).9 Meanwhile PE machine that was used by Pajovic et al. used Lubisone machine 12 sessions weekly (3000 pulses, 0.25 mJ/mm2 of energy flow density and 3 Hz).10 And lastly EM machine using Storz and Cenowave machine, have differed protocol in some studies for 4-6 sessions weekly (3000 pulses, 0.096-0.25 mJ/mm2 of energy flow density and 3-10 Hz).12,14–17 Interestingly, one study from Mykoniatis et al., stated that there were no significant differences of CPSI result between undergoing ESWT treatment once a week or twice a week. However all ESWT clinical protocols in this systematic review have shown significant improvement of CPSI scores compared to baseline.11\n\nFrom 3 different type of ESWT machinery (EP, EM, PE), all of them shared the same effect on pain domain from CPSI scoring system. Study from Ghazi et al., using EP Roland Pagani ES.WT machine showed significant improvement of pain symptom. The peak of the effect was reached for 6 months and lasted for 12 months after a month of ESWT treatment under specified protocol.9 Another study by Pajovic et al., utilized PE machine from Lubisone also demonstrated significant improvement compared to placebo 3 months and 6 months after 12 sessions of ESWT treatment.10 EM machine, the most frequently used in the study included, also showed another improvement in pain symptom. Kim et al. stated in their study that their machine (Cenowave medical) had improved their patients pain symptom in 1 month compared to placebo group.17 Pain improvement also recorded by Rayegani et al. with their Storz Medical EM machine 1 month and 3 months after 4 sessions of ESWT. Compared to placebo, the ESWT group had significant improvement in 1 month until 3 months of observations.14 Afterall, from the studies included in this systematic review, ESWT effect the pain relieve effect can last until 12 months. 3 different energy generator (EP, EM, PE) have similar effect regards the differences of the protocol that have been used in the last 10 years studies.\n\nIn this systematic review we found 1 study stated that there was no significant improvement in urinating domain of CPSI. EM Cenowave medical that was utilized by Kim et al. showed no improvement after 1 months of observation in urinating domain. Small sample sized that was observed during the study and the length of follow up that only observed for 1 month may be the cause of this negative result.17 Meanwhile, the others showed statistically significant improvement in urinary symptoms after ESWT therapy. Ghazi et al. EP Roland Pagani ES.WT showed significant improvement even 12 months after 4 sessions of ESWT therapy compared to the.9 Another study from Pajovic et al., using PE Lubisone Machine protocol stated that ESWT enhanced the condition of the related symptom compared to the placebo.10 Ultimately another EM machine by Storz medical also presented significant improvement in 1 month and reached its peak in 3 months after 4 sessions of ESWT therapy.14 Despite single contradictive result from Kim et al., most of study showed significancy in rectify the urinary symptom in CP/CPPS patients. Therefore, bigger sample size and longer observation are needed to be done for further research.\n\nThe Quality of Life is one of three domain that was comprised in CPSI scoring system. It showed the satisfaction of patient with their condition subjectively. All of the study yielded in this review stated that the patient satiety in quality of life domain had been improved by the treatment of ESWT. EP Roland Pagani ESWT Machine showed significant improvement even in 1 month until 12 month after 4 sessions of ESWT treatment compared to the baseline.9 PE Lubisone machine also stated similar outcome that 4 sessions of ESWT treatment enhanced the quality of life of CP/CPPS patients with and last for 6 months.10 Likewise, an improvement to the quality of life had been reported from all EM machine studies, either by Storz or Cenowave medical. Kim et al. Cenowave medical showed improvement compared to placebo, and Rayegani et al. Storz medical showed positive result for the exact 1 month after ESWT protocol treatment14,16,17\n\nDespite the contradictive result from urinating domain by Kim et al., the overall calculation of CPSI domain showed an improvement from the studies included in our review (2 cohort studies and 3 RCT studies) that analysed each domain of CPSI. Another 2 studies from Sakr et al. and Wu et al., had demonstrated similar outcomes. Sakr et al. had found a significant improvement in total CPSI domain after 4 sessions of ESWT treatment compared to placebo within 12 months of observations.16 Meanwhile Wu et al., reported improvement only after 8 sessions of ESWT treatment resulted compared to the placebo.15 Otherwise, 1 study from Zhang et al. did not observe any significant improvement after 8 sessions of ESWT treatment.12 This phenomenon may be caused by the exclusion of 10 relapsing patients in this particular study. Other possible explanations for these findings may also be explained by the different protocol that have been used by Zhang et al. (10 Hz of Frequency with 0.18-0.2 mJ/mm2 of energy 1 session a week vs 3-4 Hz of frequency, 0.25 mJ/mm2 of energy flow density in other EM ESWT studies). Thus, the standardize protocol for ESWT machine need to be established for each energy generator used.12\n\nThese result from 8 studies that have been yielded, all 7 studies demonstrate the efficacy of ESWT treatment with 3 kind of machinery that available worldwide. The efficacy and safety of ESWT treatment for CP/CPPS patients had showing a good outcome with less adverse effect for the CP/CPPS patients in the maximum of 12 months of observation. The limitations of this systematic review is the restricted variety of energy generators with the lack of openly registered protocols. The lack of adherence to protocols and registration may give rise to the potential of biases and compromise the accuracy and transparancy of the review. This presents the risk of selective in the absence of pre-established protocols, which may later affect the validity of the findings.\n\nLonger additional studies with longer observation are needed to be done in the future research in order to evaluate the efficacy and safety protocol of ESWT on CP/CPPS. Even with the benefit of ESWT usage for CP/CPPS, the standardize protocols are needed to enhance the outcome of the patients. The variety of machine and brand are sometime confuse the operator to set the protocol for each machine, and moreover, some of the brands are not displaying the energy flow density on its devices. Therefore, this review is comparing the protocol and the outcome that has been observed for the last 10 years. Hopefully, this review can be a suggestion and a guidance for many clinician or researcher to be applied in daily practice or to be an insight for further research.\n\n\nConclusion\n\nESWT treatment has showed its effect on CP/CPPS patients. Regardless from the type of machinery or the brand that used, the efficacy and safety of ESWT treatment showed a better outcome than the conventional oral medications alone in 12 months of observations. Although have the same efficacy, the protocol for each type of machinery is different. Therefore, this systematic review provide some insight for clinician on how clinical protocol are established for every energy generator that used in daily practice. Additional studies are needed to evaluate the efficacy of ESWT on EP/CPPS for longer duration of observation. Finally, further research must be conducted to establish the standardized protocol for shockwave delivery.\n\nThe evaluation was undertaken without previous registration and without any registered processes.", "appendix": "Data availability\n\nThe data for this article consists of bibliographic references, which are included in the References section.\n\nFigshare: PRISMA FLOW DIAGRAM The Therapeutic Effect and Safety Profile of Extracorporeal Shockwave Therapy in Treatment of Chronic Prostatitis/Chronic Pelvic Pain Syndrome: A Systematic Review, https://doi.org/10.6084/m9.figshare.23884266. 18\n\nFigshare: PRISMA CHECKLIST The Therapeutic Effect and Safety Profile of Extracorporeal Shockwave Therapy in Treatment of Chronic Prostatitis/Chronic Pelvic Pain Syndrome: A Systematic Review, https://doi.org/0.6084/m9.figshare.23884281. 19\n\nFigshare: ABSTRACT PRISMA Checklist The Therapeutic Effect and Safety Profile of Extracorporeal Shockwave Therapy in Treatment of Chronic Prostatitis/Chronic Pelvic Pain Syndrome: A Systematic Review, https://doi.org/10.6084/m9.figshare.23884278. 20\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nZhang J, Liang CZ, Shang X, et al.: Chronic Prostatitis/Chronic Pelvic Pain Syndrome: A Disease or Symptom? Current Perspectives on Diagnosis, Treatment, and Prognosis. Am. J. Mens Health. 2020; 14: 155798832090320. SAGE Publications Inc. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPontari MA, Ruggieri MR: Mechanisms in prostatitis/chronic pelvic pain syndrome. J. Urol. 2004; 172: 839–845. Lippincott Williams and Wilkins. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKrieger JN, Ross SO, Deutsch L, et al.: The NIH Consensus Concept of Chronic Prostatitis/Chronic Pelvic Pain Syndrome Compared With Traditional Concepts of Nonbacterial Prostatitis and Prostatodynia. Curr. Urol. Rep. 2002; 3: 301–306. PubMed Abstract \| Publisher Full Text\n\nGrabe MBM; BJTBHC ek MLB et al.: Chronic Pelvic Pain EAU Guidelines on.2022.\n\nMurphy AB, Nadler RB: Pharmacotherapy strategies in chronic prostatitis/chronic pelvic pain syndrome management. Expert. Opin. Pharmacother. 2010; 11: 1255–1261. PubMed Abstract \| Publisher Full Text\n\nMoya D, Ramón S, Schaden W, et al.: The role of extracorporeal shockwave treatment in musculoskeletal disorders. J. Bone Joint Surg. (Am. Vol.). 2018; 100: 251–263. Lippincott Williams and Wilkins. PubMed Abstract \| Publisher Full Text\n\nChen PY, Cheng JH, Wu ZS, et al.: New Frontiers of Extracorporeal Shock Wave Medicine in Urology from Bench to Clinical Studies. Biomedicine. 2022; 10. MDPI. Publisher Full Text\n\nSterne JAC, Savović J, Page MJ, et al.: RoB 2: A revised tool for assessing risk of bias in randomised trials. BMJ. 2019; l4898. Publisher Full Text\n\nAl Edwan GM, Muheilan MM, Atta ONM: Long term efficacy of extracorporeal shock wave therapy [ESWT] for treatment of refractory chronic abacterial prostatitis. Ann. Med. Surg. 2017 Feb 1; 14: 12–17. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPajovic B, Radojevic N, Dimitrovski A, et al.: Comparison of the efficiency of combined extracorporeal shock-wave therapy and triple therapy versus triple therapy itself in Category III B chronic pelvic pain syndrome (CPPS). Aging Male. 2016 Jul 2; 19(3): 202–207. Publisher Full Text\n\nMykoniatis I, Pyrgidis N, Kalyvianakis D, et al.: Comparing two different low-intensity shockwave therapy frequency protocols for nonbacterial chronic prostatitis/chronic pelvic pain syndrome: A two-arm, parallel-group randomized controlled trial. Prostate. 2021 Jun 1; 81(9): 499–507. PubMed Abstract \| Publisher Full Text\n\nZhang ZX, Zhang D, Yu XT, et al.: Efficacy of Radial Extracorporeal Shock Wave Therapy for Chronic Pelvic Pain Syndrome: A Nonrandomized Controlled Trial. Am. J. Mens Health. 2019 Jan 1; 13(1): 155798831881466. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSimplicio CL, Purita J, Murrell W, et al.: Extracorporeal shock wave therapy mechanisms in musculoskeletal regenerative medicine. J. Clin. Orthop. Trauma. 2020; 11: S309–S318. Elsevier B.V. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRayegani SM, Razzaghi M, Raeissadat SA, et al.: Extracorporeal shockwave therapy combined with drug therapy in chronic pelvic pain syndrome: A randomized clinical trial. Urol. J. 2020 Mar 1; 17(2): 185–191. PubMed Abstract \| Publisher Full Text\n\nWu WL, Adebayo Bamodu O, Wang YH, et al.: Clinical Medicine Extracorporeal Shockwave Therapy (ESWT) Alleviates Pain, Enhances Erectile Function and Improves Quality of Life in Patients with Chronic Prostatitis/Chronic Pelvic Pain Syndrome. J. Clin. Med. 2021; 10: 3602. Publisher Full Text\n\nSakr AM, Fawzi AM, Kamel M, et al.: Outcomes and clinical predictors of extracorporeal shock wave therapy in the treatment of chronic prostatitis/chronic pelvic pain syndrome: a prospective randomized double-blind placebo-controlled clinical trial. Prostate Cancer Prostatic Dis. 2022 Mar 1; 25(1): 93–99. PubMed Abstract \| Publisher Full Text\n\nKim KS, Choi YS, Bae WJ, et al.: Clinical efficacy of multi-focal low-intensity extracorporeal shockwave therapy in the treatment of chronic prostatitis/chronic pelvic pain syndrome: Prospective-randomized, double blind, placebo-controlled study. World J. Men's Health. 2021; 40: 678. Publisher Full Text\n\nMikha M, Deswanto IA: PRISMA FLOW DIAGRAM The Therapeutic Effect and Safety Profile of Extracorporeal Shockwave Therapy in Treatment of Chronic Prostatitis/Chronic Pelvic Pain Syndrome: A Systematic Review.2023 [cited 2023 Aug 5]. Publisher Full Text\n\nMikha M, Deswanto IA: PRISMA CHECKLIST The Therapeutic Effect and Safety Profile of Extracorporeal Shockwave Therapy in Treatment of Chronic Prostatitis:Chronic Pelvic Pain Syndrome- A Systematic Review.2023 [cited 2023 Aug 5]. Publisher Full Text\n\nMikha M, Deswanto IA: ABSTRACT PRISMA CHECKLIST The Therapeutic Effect and Safety Profile of Extracorporeal Shockwave Therapy in Treatment of Chronic Prostatitis:Chronic Pelvic Pain Syndrome - A Systematic Review.docx.2023 [cited 2023 Aug 5]. Publisher Full Text" }	[ { "id": "231990", "date": "14 May 2024", "name": "Biagio Barone", "expertise": [ "Reviewer Expertise urology", "andrology", "urologic oncology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nGeneral comment: The manuscript entitled “The therapeutic effect and safety profile of extracorporeal shockwave therapy in treatment of chronic prostatitis/chronic pelvic pain syndrome: a systematic review” by Mikha and Deswanto, aims to compare the clinical protocol, efficacy and safety profiles of different ESWT machines in CP/CPPS treatment. The manuscript is overall fairly well written and well structured. In particular, the methodology is well reported and easily followed along the work. Few corrections are suggested in order to improve the quality of the work: Intriduction: Expand the epidemiological data about CP/CPPS. Materials and Methods: Add inclusion and exclusion criteria for studies retrieved. Results: The tables could be improved in terms of understandability. Discussion: Add the limitations of your study. About the role of ESWT in chronic prostatitis, also consider special conditions and potential synergic effects with antibiotics and other drugs. To this regard also see: Barone B et al; 2023[Ref 1] and Crocetto F et al; [Ref 2] Briefly add which are the current guidelines about the treatment of CP/CPPS and how ESWT could be integrated in novel therapeutic treatments.\n\nAre the rationale for, and objectives of, the Systematic Review clearly stated? Yes\n\nAre sufficient details of the methods and analysis provided to allow replication by others? Partly\n\nIs the statistical analysis and its interpretation appropriate? I cannot comment. A qualified statistician is required.\n\nAre the conclusions drawn adequately supported by the results presented in the review? Yes\n\nIf this is a Living Systematic Review, is the ‘living’ method appropriate and is the search schedule clearly defined and justified? (‘Living Systematic Review’ or a variation of this term should be included in the title.) Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1363
https://f1000research.com/articles/12-1362/v1	18 Oct 23	{ "type": "Research Article", "title": "Association of the CD28 markers with the disease activity in systemic lupus erythematosus patients", "authors": [ "Mirza Zaka Pratama", "Kusworini Handono", "Handono Kalim", "Hani Susianti", "Kusworini Handono", "Handono Kalim", "Hani Susianti" ], "abstract": "Background: Systemic Lupus Erythematosus (SLE) is a complex autoimmune disease with diverse manifestations and unpredictable activity. CD28 markers, particularly sCD28, is a promising biomarker for evaluating SLE disease activity. This study aimed to investigate the significance of CD28 markers in evaluating disease activity in SLE and the role of sCD28 in various clinical manifestations. Methods: A total of 40 female subjects, aged between 18 and 45 years, who fulfilled the 2019 EULAR/ACR classification criteria for SLE were recruited in this study. Twenty healthy matched individuals were also recruited as control. Comprehensive data on demographic information, clinical manifestations, laboratory test findings, and treatment history were collected from all participants. The Indonesian version of SLEDAI-2K score was utilized to assess disease activity, categorizing patients into active SLE and lupus low disease activity (LLDAS). Collected data were analyzed on SPSS for Windows version 25.0. Results: Patients with SLE in LLDAS category had significantly lower SLEDAI scores (1.8 ± 1.4 vs 11.7 ± 4.9, p<0.001) with mild clinical manifestation. Active SLE patients had the lowest percentages of CD4+CD28+ cells (5.7 ± 4.1%) and the highest sCD28 concentration (26.2 ± 11.3 ng/ml) compared to other groups. Moreover, sCD28 concentration demonstrated a moderate positive correlation with SLE disease activity. In most cases, higher sCD28 concentrations were associated with clinical manifestations, particularly in neuropsychiatric lupus (OR 7.1 [1.8 – 67.9], p=0.047), nephritis (OR 14.5 [1.6 – 131.9], p=0.017), and mucocutaneous manifestations (OR 3.4 [1.9 – 12.8], p=0.035). Conclusions: Our study establishes the link between CD28 markers and disease activity, including certain clinical manifestations in SLE. We suggest that CD28 has a potential role in predicting disease activity. However, further research through longitudinal studies is required to strengthen these findings.", "keywords": [ "Systemic Lupus Erythematosus", "CD28 markers", "disease monitoring" ], "content": "Introduction\n\nSystemic Lupus Erythematosus (SLE) is an autoimmune disease predominantly in women, leading to inflammation and damage in various organs and tissues. The clinical manifestations and disease course of SLE can vary widely among patients, making it challenging to predict disease activity and tailor appropriate treatment strategies.1 In recent years, researchers have been investigating potential biomarkers that could aid in the assessment and management of SLE. One promising biomarker is the Cluster of Differentiation 28 (CD28) marker.2–4 CD28 is a surface co-stimulatory molecule in T-cells, playing a crucial role in regulating T-cell activation and proliferation. It interacts with its ligands on antigen presenting cells, CD80 and CD86, to facilitate effective T-cell responses.5 In certain conditions, CD28 can be cleaved under certain conditions from the T-cell surface, releasing soluble CD28 into the bloodstream.6 The soluble form of CD28, referred to as sCD28, has attracted much interest as a potential biomarker for autoimmune disorders, such as SLE.7\n\nOur previous study observed a reduction in CD28 expression on T-cell surfaces, which was linked to comorbidities in patients with SLE.8 This downregulation of CD28 may impair the normal regulatory functions of T-cells, leading to uncontrolled immune responses and increased disease activity.9,10 The levels of sCD28 in the blood of patients with SLE or other autoimmune disorders are higher than those of healthy individuals as shown in previous studies.11,12 Increased sCD28 levels may contribute to the autoimmune response’s persistence in SLE.13\n\nHowever, the clinical presentation of SLE can vary widely among patients, suggesting the existence of different phenotypes with distinct underlying pathogenic mechanisms.14 Little studies have described the role of the sCD28 markers according to the disease phenotypes in SLE patients. Therefore, a study is needed to identify the mechanisms and substantiate the clinical efficacy of soluble CD28 as a biomarker in SLE. Thus, this study is going to discover the role and performance of CD28 markers in assessing the disease activity in SLE. We also aimed to observe the role of sCD28 markers in different clinical manifestations in SLE patients. The investigation of sCD28 as a biomarker in SLE may hold promise for improving the diagnosis, assessment, and management of this complex autoimmune disease.\n\n\nMethods\n\nThe subjects in this study consisted of forty patients diagnosed with SLE. Patients were recruited with the consecutive sampling from the Rheumatology Clinic at Saiful Anwar General Hospital in Malang, Indonesia, throughout the designated study period (March to December 2020). The study participants consisted exclusively of female individuals aged between 18 and 45 who met the 2019 European League Against Rheumatism/American College of Rheumatology (EULAR/ACR) categorization criteria for SLE. Briefly, patients should have a positive ANA as a mandatory entry criterion, followed by additional criteria that were scored from 2 to 10 (constitutional, hematological, neuropsychiatric, mucocutaneous, serosal, musculoskeletal, renal, antiphospholipid antibodies, complement proteins, and SLE-specific antibodies). Patients with score ≥10 points would be classified as SLE.15 We excluded pregnant or breastfeeding patients with ongoing infection or malignancy. Twenty individuals in healthy condition, matched in age and gender, were chosen as control subjects. Demographic data, clinical symptoms, laboratory test findings, and therapeutic history were reported for all individuals.\n\nThis study followed the Helsinki Declaration and the ethical committee of Saiful Anwar General Hospital in Malang, Indonesia, granted its approval for this research (ethical number approval 400/085/K.3/302/2020, approved on 23rd March 2020). Written informed consent was also obtained from all participants involved.\n\nThe patients’ disease activity was evaluated using the Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K).16 The SLEDAI-2K score was evaluated by a rheumatologist in the Rheumatology Clinics. Routine laboratory tests, such as complete blood count, urinalysis, anti-dsDNA, C3, and C4 were conducted as part of the SLEDAI-2K assessment. The radiologic and biopsy examinations were also performed as indicated according to the disease manifestations. SLE patients were classified according to the disease activity: active and lupus low disease activity state (LLDAS) SLE patients. LLDAS was categorized based on the previous study.17\n\nThe flowcytometry examination was used to measure the percentages of membrane or surface CD28 expression within the CD4 and CD8 T-cell subsets. The investigation involved the evaluation of CD28 membrane expression percentages in peripheral blood mononuclear cells (PBMC) from subjects. A venous blood sample of 10-15 cc was obtained from each participant. PBMCs were extracted from using Lymphoprep (Stemcell Technology) through centrifugation. The produced PBMC layer was systematically eliminated and washed twice using 10 cc of phosphate-buffered saline (PBS). Afterward, the supernatant was eliminated, and the remaining supernatant was centrifuged at room temperature. FITC anti-human CD4, PerCP anti-human CD8, and PE anti-human CD28 (Biolegend) was used as the staining. The percentages of CD4+CD28+ and CD8+CD28+ cells were assessed using flow cytometry (BD FACScalibur). The measurements were performed on 10,000 cells. The results were expressed as percentages (%) of cells.\n\nThe ELISA examination was performed to measure the concentration of serum sCD28 markers. The analysis of serum SCD28 was performed using the ELISA kit provided by Bioassay Technology Laboratory (cat number E4196hu), following the methods recommended by the manufacturer. Briefly, serum collected from the blood of the subjects by centrifugation at 2000-3000 rpm for 30 minutes. Samples and ELISA reagents were added into each well and incubated for 1 hour at 37oC. The plate was washed five times and the substrate solution were added. Samples were incubated for 10 minutes at 37oC, then the stop solution were added. Samples were read the optical density (OD) value within 10 minutes. The sCD28 marker levels were measured in nanograms per milliliter (ng/ml).\n\nNormally distributed data were characterized using mean and standard deviation, while skewed data were represented by median and interquartile range. Categorical data were presented as percentages. Unpaired t-test or Mann-Whitney test was utilized to compare numerical data between groups. The Chi-square or Fisher exact test was employed to conduct a comparative analysis of two sets of categorical data. Pearson’s correlation was used to examine the relationships between each senescence marker and the SLEDAI score. Receiver operating characteristic (ROC) curves were applied to distinguish between active and LLDAS SLE. Logistic regression models were used to evaluate the association between the variables. The statistical significance of the results was determined based on a p-value of <0.05. Data analysis was conducted in SPSS for Windows version 25.0.\n\n\nResults\n\nThere are twenty healthy individuals, as well as forty SLE patients. The patients with SLE were categorized into two groups: active SLE patients (n=20) and SLE patients in low disease activity state (LLDAS) (n=20). The characteristics of the subjects are displayed in Table 1. All subjects were females of similar ages. Patients with Lupus Low Disease Activity State (LLDAS) showed notably lower SLEDAI score than individuals with active SLE (1.8 ± 1.4 vs 11.7 ± 4.9, p<0.001). The clinical manifestations of SLE patients in both groups are presented in Table 1. Patients with LLDAS showed mild manifestations, such as mucocutaneous lesions or hematologic issues, but none had hemolytic anemia. The anti-dsDNA levels of the active SLE patients were markedly higher compared to LLDAS patients (89.2 ± 62.4 vs. 51.8 ± 46.7 IU/ml, p=0.038).\n\n\n\n- Neuropsychiatric\n\n\n\n- Vasculitis\n\n\n\n- Arthritis\n\n\n\n- Myositis\n\n\n\n- Nephritis\n\n\n\n- Mucocutaneous\n\n\n\n- Serositis\n\n\n\n- Hematologic\n\nThe percentages of membrane CD28 seen in both CD4+ and CD8+ T-cells were displayed in Figure 1. Among all groups, active SLE patients showed the lowest percentages of CD4+CD28+ (5.7 ± 4.1%). Notably, the percentages of CD4+CD28+ were significantly higher in LLDAS SLE patients (8.7 ± 4.7%, p=0.033) and healthy individuals (10.8 ± 4.6%, p=0.001) compared to active SLE patients. There was no statistically significant disparity was observed in the percentages of CD4+CD28+ cells between the patients with LLDAS SLE and the control group of healthy individuals (p=0.175). As for the CD8+ T-cell subset, the lowest percentages were found in active SLE patients (6.3 ± 4.6%) and statistically significant compared to LLDAS SLE patients (10.0 ± 5.5%, p=0.030). However, no significant differences were found in the CD8+CD28+ percentages between the SLE patients (both LDAS patients) with healthy individuals (8.0 ± 3.6%). The comparison of sCD28 between groups are also described in Figure 1. Among all groups, it was shown that active SLE patients had the highest concentration of sCD28 at 26.2 ± 11.3 ng/ml, which was found to be considerably higher when compared to both LLDAS patients with a value of 15.0 ± 5.6 ng/ml (p=0.001), as well as healthy individuals with a value of 14.1 ± 9.4 ng/ml (p<0.001).\n\nThe Pearson correlation analysis was conducted to evaluate the correlation between the CD28 markers and disease activity among the patient, as shown in Figure 2. The study revealed a weak negative correlation between the percentages of CD4+CD28+ cells and the SLEDAI score in SLE patients. Furthermore, a comparable correlation was observed between the percentages of CD8+CD28+ cells and the SLEDAI score. Conversely, the concentration of sCD28 exhibited a moderate positive correlation with the SLEDAI score.\n\nThe study conducted ROC curve analysis to evaluate the performance of the CD28 marker in distinguishing between active SLE patients and SLE patients in low disease activity state (LLDAS), as depicted in Figure 3. The CD4+CD28+ had a relatively low AUC score, measuring 0.697 (95% CI 0.531 – 0.864) with a p-value of 0.033. Both the sensitivity and specificity were 70% for the cut-off value of 6.0%. We reported a greater AUC score for CD8+CD28+ percentages in the differentiation of active SLE patients (AUC 0.700 [95% CI 0.534 – 0.866], p=0.030). Using a cut-off value of 4.8%, the percentages of CD8+CD28+ exhibited a sensitivity of 80% and a specificity of 50%. The sCD28 had the highest AUC score of 0.822 (95% CI 0.690 – 0.955, p=0.001), demonstrating a sensitivity of 70% and specificity of 80% for a cut-off value of 18.6 ng/ml. We also conducted a comparison to evaluate the performance of anti-dsDNA as a conventional marker to differentiate between active and LLDAS SLE patients. The AUC score for anti-dsDNA was determined to be 0.740 (95% CI [0.585 – 0.895], p=0.020), along with the sensitivity and specificity of the test at a cut-off value of 85 IU/ml were found to be 40% and 80% respectively.\n\nSLE patients might have different phenotypes with various clinical manifestations. In order to observe the association between CD28 markers and disease phenotypes, a comparison of sCD28 markers among SLE patients based on their clinical manifestations was performed and presented in Table 2. In most cases, the sCD28 concentrations were higher in subjects with clinical manifestations. Nevertheless, notable differences were noted among patients with SLE who exhibited neuropsychiatric manifestations (p=0.031), nephritis (p=0.043), and serositis (p=0.007). The cut-off value was used to analyze the association between the increased sCD28 marker according to the presence of disease manifestations. Significant associations were demonstrated in neuropsychiatric lupus (OR 7.1 [1.8 – 67.9], p=0.047), nephritis (OR 14.5 [1.6 – 131.9], p=0.017), and mucocutaneous manifestations (OR 3.4 [1.9 – 12.8], p=0.035).\n\n\nDiscussion\n\nSystemic lupus erythematosus (SLE) is a complex autoimmune disorder that can impact multiple organs. The finding for markers to assess the disease activity in SLE is crucial for guiding treatment decisions, monitoring disease progression, evaluating treatment response, facilitating clinical trial design, and improving patient outcomes. These markers enable a more precise and personalized approach to the management of SLE, leading to better disease control, prevention of organ damage, and enhanced overall patient care. The role of CD28, a costimulatory molecule expressed on the surface of T-cells, has been an issue in evaluating disease activity in SLE.18–20\n\nThis study showed a significant decrease in the expression of the CD28 surface marker among patients diagnosed with SLE, particularly in those with active SLE, when compared to healthy participants. Based on previous observations in patients with SLE have revealed a reduction in CD28 surface marker expression on T-cells, which has been linked to both disease activity and severity.20,21 Similar to the findings in this study, our previous data indicated a correlation between the loss of CD28 surface markers on T-cells and the occurrence of SLE morbidities.8 Although multiple studies have documented the association of CD28+ T-cells with SLE disease activity, there is a few detailed comparative studies for the CD28+ T-cell subsets in SLE. Minning et al. described that the CD8+CD28+ T-cells subset was reduced and inversely correlated with SLEDAI score.22 Another study also showed that the peripheral CD4+ or CD8+ with CD28+ was negatively correlated with the CD28+ T-cells.20 The functional characteristics of CD28+ T-cells exhibited increased production of Th1 proinflammatory cytokines along with cytotoxic molecules, perforin, and granzymes.23 Therefore, these proinflammatory phenotypes might potentially impact the disease activity in SLE patients.\n\nCompared to the CD28 surface marker, the levels of soluble CD28 (sCD28) were elevated in SLE patients and showed a positive correlation with disease activity. We also observed that the sCD28 had a better performance in predicting the active SLE patients compared to the surface CD28 marker or the conventional anti-dsDNA. Several specific clinical manifestations in SLE are also associated with the increased sCD28 concentration. The increase of sCD28 had been documented in SLE or other autoimmune diseases from previous studies.11,12 Despite that, few studies had been demonstrated the performance of this marker for the monitoring of SLE disease activity.19\n\nThe correlation between the CD28 surface marker and the CD28 soluble marker is complex. Under certain conditions, CD28 can be cleaved from the T-cell surface, releasing sCD28 into the bloodstream. The soluble CD28 can be generated via the membrane form being shed or by alternative mRNA splicing.24 The elevated shedding of sCD28 could indicate high T-cell activity in SLE patients, resulting in higher levels of sCD28 in the serum. In contrast to the soluble form of CD28, the chronic activation and dysregulation of T-cells among SLE patients might lead to T-cell exhaustion.25 The T-cell exhaustion was linked to the disappearance of CD28 expression on the T-cell surface, which was substituted by inhibitory molecules like programmed cell death 1 (PD-1).26 The association between the CD28 molecules with the disease activity might represent the dysregulation of the T-cell response in SLE. This finding was also confirmed by the previous analysis that the T-cell activation dysregulations had the clinical implications in SLE patients.27\n\nThis study has certain limitations, as it employs a cross-sectional design, potentially resulting in a weak casual-effect relationships. Therefore, a longitudinal study in the future will provide a better insight into the association between the CD28 markers with the disease activity, risk for relapse, or therapeutic efficacy. Although the sample of this study was relatively low, we believed that the group of patients selected was the most suitable for conducting this analysis. The patients and healthy individuals were carefully selected, and the identification of disease activity and clinical manifestations were examined accordingly.\n\nIn conclusion, our findings confirm that CD28 markers are indeed linked to disease activity in SLE. Some of the clinical manifestations are also associated with the levels of CD28 markers. In general, this study indicates a potential role of CD28 in predicting disease activity in SLE. However, this current study requires further research through longitudinal studies to provide multiple advantages in assessing disease activity, personalized treatment strategies, prognostic value, mechanistic insights, and potential therapeutic targets. As research in this field advances, these benefits could contribute to improving patient care and a more profound understanding of the complex mechanisms underlying SLE pathogenesis.", "appendix": "Data availability\n\nFigshare: Role of Soluble CD28 as the Predictor of Disease Activity in SLE, https://doi.org/10.6084/m9.figshare.23897154. 28\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nAcknowledgments\n\nThe author thanks to Mr. Wahyudha for helping assist in the flow cytometry analysis at the Biomedical Laboratory, Faculty of Medicine Universitas Brawijaya.\n\n\nReferences\n\nZucchi D, Elefante E, Schilirò D, et al.: One year in review 2022: systemic lupus erythematosus. Clin. Exp. Rheumatol. 2022 Jan; 40(1): 4–14. Publisher Full Text\n\nBrambila-Tapia AJ, Dávalos-Rodríguez IP, Gámez-Nava JI, et al.: CD28 proximal promoter polymorphisms in systemic lupus erythematosus susceptibility. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWong CK, Lit LC, Tam LS, et al.: Aberrant production of soluble costimulatory molecules CTLA-4, CD28, CD80 and CD86 in patients with systemic lupus erythematosus. Rheumatology (Oxford). 2005 Aug; 44(8): 989–994. PubMed Abstract \| Publisher Full Text\n\nKakoulidou M, Wang X, Zhao X, et al.: Soluble costimulatory factors sCD28, sCD80, sCD86 and sCD152 in relation to other markers of immune activation in patients with myasthenia gravis. J. Neuroimmunol. 2007 Apr; 185(1-2): 150–161. Publisher Full Text\n\nAmbrose N, Morgan TA, Galloway J, et al.: Differences in disease phenotype and severity in SLE across age groups. Lupus. 2016 Dec; 25(14): 1542–1550. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAringer M: EULAR/ACR classification criteria for SLE. Semin. Arthritis Rheum. 2019 Dec; 49(3S): S14–S17. Publisher Full Text\n\nUribe AG, Vilá LM, McGwin G Jr, et al.: The Systemic Lupus Activity Measure-revised, the Mexican Systemic Lupus Erythematosus Disease Activity Index (SLEDAI), and a modified SLEDAI-2K are adequate instruments to measure disease activity in systemic lupus erythematosus. J. Rheumatol. 2004 Oct; 31(10): 1934–1940. PubMed Abstract\n\nFranklyn K, Lau CS, Navarra SV, et al.: Definition and initial validation of a Lupus Low Disease Activity State (LLDAS). Ann. Rheum. Dis. 2016 Sep; 75(9): 1615–1621. PubMed Abstract \| Publisher Full Text\n\nVanhove B, Poirier N, Fakhouri F, et al.: Antagonist Anti-CD28 Therapeutics for the Treatment of Autoimmune Disorders. Antibodies (Basel). 2017 Nov 21; 6(4): 19. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBrambila-Tapia AJ, Gámez-Nava JI, Salazar-Páramo M, et al.: Increased CD28 serum levels are not associated with specific clinical activity in systemic lupus erythematosus. Rheumatol. Int. 2011 Oct; 31(10): 1321–1324. Publisher Full Text\n\nKosmaczewska A, Ciszak L, Stosio M, et al.: CD4+CD28+ T-cells are expanded in moderately active systemic lupus erythematosus and secrete pro-inflammatory interferon gamma, depending on the Disease Activity Index. Lupus. 2020 Jun; 29(7): 705–714. PubMed Abstract \| Publisher Full Text\n\nPiantoni S, Regola F, Zanola A, et al.: Effector T-cells are expanded in systemic lupus erythematosus patients with high disease activity and damage indexes. Lupus. 2018 Jan; 27(1): 143–149. PubMed Abstract \| Publisher Full Text\n\nMinning S, Xiaofan Y, Anqi X, et al.: Imbalance between CD8+CD28+ and CD8+CD28- T-cell subsets and its clinical significance in patients with systemic lupus erythematosus. Lupus. 2019 Sep; 28(10): 1214–1223. PubMed Abstract \| Publisher Full Text\n\nWang Y, Bai J, Li F, et al.: Characteristics of expanded CD4+CD28+ T-cells in patients with chronic hepatitis B. Immunol. Investig. 2009; 38(5): 434–446. PubMed Abstract \| Publisher Full Text\n\nMagistrelli G, Jeannin P, Elson G, et al.: Identification of three alternatively spliced variants of human CD28 mRNA. Biochem. Biophys. Res. Commun. 1999; 259: 34–37. PubMed Abstract \| Publisher Full Text\n\nLima G, Treviño-Tello F, Atisha-Fregoso Y, et al.: Exhausted T-cells in systemic lupus erythematosus patients in long-standing remission. Clin. Exp. Immunol. 2021 Jun; 204(3): 285–295. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHuang Y, Zheng H, Zhu Y, et al.: Loss of CD28 expression associates with severe T-cell exhaustion in acute myeloid leukemia. Front. Immunol. 2023 Mar 7; 14: 1139517. Publisher Full Text\n\nZhou H, Li B, Li J, et al.: Dysregulated T-cell Activation and Aberrant Cytokine Expression Profile in Systemic Lupus Erythematosus. Mediat. Inflamm. 2019 Mar 17; 2019: 1–11. Publisher Full Text\n\nPratama MZ, Handono K, Susianti H, et al.: Role of Soluble CD28 as the Predictor of Disease Activity in SLE. Dataset. figshare. 2023. Publisher Full Text" }	[ { "id": "327707", "date": "12 Oct 2024", "name": "Reem Hamdy A. Mohammed", "expertise": [ "Reviewer Expertise Autoimmune disease and rheumatology." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe article presented illustrates the link between the CD28 expression on CD 4 and CD8 T cells and its' relation to disease activity and organ involvement in lupus. This is a case-control study with a cross sectional design though the study design wasn't mentioned yet the methodology illustrated the design clearly. The objective of the study is very interesting especially being displayed for the Indonesian population which increases the body of evidence in another population other than European and American data that are readily available. This study showed a significant decrease in the expression of the CD28 surface marker among patients diagnosed with SLE, particularly in those with active SLE, when compared to healthy participants. The study analysis showed that the levels of soluble CD28 (sCD28) were elevated in SLE patients and showed a positive correlation with disease activity and correlated with neuropsychiatric, mucocutaneous and nephritis in the study group. The authors also observed that the CD28 had a better performance in predicting the active SLE patients compared to the surface CD28 marker or the conventional anti-dsDNA. The authors mentioned their study limitation and declared data availability with links provided.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "12631", "date": "17 Oct 2024", "name": "Mirza Zaka Pratama", "role": "Author Response", "response": "Thank you for your thorough review and constructive feedback on our article. We appreciate your positive remarks about our study's objective, particularly the focus on the Indonesian population, which aims to broaden the understanding of systemic lupus erythematosus (SLE) beyond European and American contexts. We are glad that you found the methodology and statistical analysis appropriate, as well as the clarity in presenting the link between CD28 expression and disease activity in lupus. Additionally, we appreciate your acknowledgment of the significant findings, particularly the predictive performance of CD28 in active SLE cases compared to conventional markers like anti-dsDNA. We also take note of your observation about the study design not being explicitly mentioned. Although we believe the methodology section conveys the cross-sectional case-control nature of the study, we will incorporate an explicit statement about the design to enhance clarity for readers. Your feedback regarding the reproducibility of our data is encouraging, and we are pleased that our efforts to provide transparent access to the source data were well-received. If there are additional details you believe would further enhance the replicability of the study, we are happy to address them. Once again, we appreciate your valuable feedback and thoughtful insights, which have contributed to improving the quality of our work." } ] } ]	1	https://f1000research.com/articles/12-1362
https://f1000research.com/articles/12-1360/v1	18 Oct 23	{ "type": "Case Report", "title": "Case Report: Prostate adenocarcinoma presented with inguinal lymph node metastasis", "authors": [ "Yeshwant Lamture", "Prajwal Lamture", "Varsha Lamture", "Pankaj Gharde", "Prajwal Lamture", "Varsha Lamture", "Pankaj Gharde" ], "abstract": "Prostate cancer has the highest chance of spreading to the bones. After pelvic and bone involvement, pulmonary metastases and extensive lymphadenopathy are expected consequences. Some men with prostate cancer develop symptomatic metastatic lung lesions and lymphadenopathy despite the absence of other signs of distant spread. Prostate cancer is exceedingly common and the leading cause of cancer in males. Prostate cancer patients with metastases account for half of all cases. In some cases, it occurs in the peritoneum but can also invade the surrounding structures, the bladder, seminal vesicles, and urinary sphincters. For example, we report the case of a 62-year-old man who presented to our hospital with inguinal swelling as his primary symptom. The patient has a history of constipation, burning micturition, and weight loss. The patient was advised of ultrasonography (U.S.G.) abdominonopelvic, which revealed enlarged, irregular, heterogeneous prostate and cystitis and multiple hepatic metastases. Also, there was a lobulated mass lesion in the inguinal canal. The most common spread occurs in bone and lymph nodes. It's uncommon to witness inguinal lymph node metastasis without pelvic lymphadenopathy or other metastasis. The spread occurs usually through a hematogenous, lymphatic, or direct route. Hence this one is a rare presentation. One possible explanation might be abnormal or aberrant lymphatic drainage of the prostate to the inguinal lymph node.", "keywords": [ "Adenocarcinoma", "lymphadenopathy", "metastasis", "hormonal therapy", "chemoradiation", "neoadjuvant chemotherapy." ], "content": "Introduction\n\nProstate cancer is a common cancer among older men that frequently spreads to regional lymph nodes and, in rare cases, bone. The inguinal region is rarely involved in the lymphatic dissemination of prostate cancer. Prostatic adenocarcinoma is an asymptomatic disorder and can be a silent killer. The common mode of presentation in symptomatic disease is in the form of obstructive uropathy. It is the leading cause of death in male cancer patients.1 Around one-half of these cancer patients have metastatic disease at presentation. Bone and lymph nodes are the most common targets of cancer cells. However, spreading to inguinal lymph nodes without pelvic lymphadenopathy or other metastases is rare. This cancer adopts root of spread from the obturator lymph nodes, sacral both pre and lateral, internal and external iliac, and lastly the inguinal lymph node.2 We report a rare metastatic prostate adenocarcinoma only to the inguinal lymph nodes without involvement of other nearby reginal nodes.\n\nProviding more information on the symptoms and diagnosis of metastatic prostate adenocarcinoma could be helpful to readers who are interested in learning more about the disease. This could include details on identifying the signs of metastasis and what tests are typically used to confirm a diagnosis. Additionally, the article could benefit from additional information on treatment options for metastatic prostate adenocarcinoma. This would be especially useful for readers who may be dealing with the disease themselves, like physicians and surgeons.\n\n\nCase report\n\nA 62-year-old of male gender having Indo Aryan ethinicity. He was farmer by occupation, came to Acharya Vinobha Bhave Rural Hospital, Sawangi, with the chief complaint of swelling in the left inguinal region, which had been from two months. The swelling was initially minor and progressively grew larger; no pain was associated. The patient has a past medical case of prostate carcinoma for which he underwent Transurethral resection of the prostate (TURP) with orchidectomy 2 years ago, received two cycles of chemotherapy with docetaxel, and ten sessions of radiotherapy. There is no positive family history means no any other family member were affected by similar disease like prostate. Personal history revealed complaints of constipation, burning micturition, and weight loss. He lost 8 kg in the previous month.\n\nHe was well-oriented to time, place, and person. On general examination, including all the components like sex, age, temperature, pulse, respiration, cyanosis, clubbing, pallar, blood pressure, icterus, development and habitus, state of nutrition, consciousness, facial features and expression, position (decubitous), skin, eyes, jugular venous pressure, limb oedema and lymph nodes etc. It was normal except of inguinal lymphadenopathy. Per abdominal examination was done thoroughly following all four basic steps including inspection, palpation, percussion, and auscultation. The bowel sounds were normal. Auscultation was done with bell of stethoscope in supine posture in calm and quite place. No undue pressure on stethoscope was was given. Bowel sounds were heard for 5minutes in all four quadrants. Auscultation was done immediately after inspection before palpation or percussion as it can stimulate the peristalsis. They were heard in all four quadrants. On palpation found to have mild hepatomegaly. There was no tenderness. There was no evidence of guarding, rigidity, or distension of the abdomen.\n\nOn digital rectal examination, there were no external skin tags or hemorrhoids, normal anal tone, and normal anal mucosa. There was grade 3 prostomegaly, a gloved finger stained with stool. The inguinal region assessment showed a firm, non-tender swelling of approx. 3*2cm on left inguinal region. The skin over the swelling appeared to be normal. Other systemic examination was normal. He underwent further investigations.\n\nA prostate-specific antigen (PSA) was elevated. The patient underwent an enhanced computed tomography (C.T.) scan of the abdomen and pelvis. Enlarged left inguinal lymph nodes with a maximum diameter of 3 cm were discovered on a C.T. scan of the abdomen.\n\nAlong with it, U.S.G. abdomen and pelvis were performed, both revealed an enlarged, irregular, and heterogeneous prostate. The bladder showed cystitis with severe bladder wall thickening at the bladder neck. There were multiple hepatic metastases. The lobulated hypoechoic mass lesion in the left inguinal canal suggestive of metastatic lymph nodal mass (see Figure 1).\n\nTranrectal ultrasound guided fine needle aspiration cytology was performed to reconfirm the diagnosis (see Figure 2A). After obtaining physician fitness under general anesthesia, the patient underwent an excision biopsy of the left inguinal lymph node. Incision taken parallel to and 1.25 cm above medial half of left inguinal ligament. Incision deepened. Lymph node visualized. Separated and freed from all sides. Lymph node mass delivered out. Mop count and instrument count checked. Haemostasis achieved. Incision closed in layers. Skin sutured using ethilon. The procedure is uneventful. Histopathological examination confirms the diagnosis of adenocarcinoma metastasis from adenocarcinoma a (see Figure 2B).\n\n\nDiscussion\n\nDespite all advances in understanding the treatment of prostatic cancer, around two and a half lakhs of persons succumb to death every year worldwide. Patients having distant metastasis suggest a poor prognosis with a median survival of up to 2 and half years with considerable inter-patient variations; few may have a short life, and others may survive up to 10 years with hormonal therapy. The cause of this diversity is unknown.\n\nRosa M. et al. report a case of aggressive prostate cancer in a middle-aged man. He initially had nonspecific symptoms like nausea, vomiting, and inguinal lymphadenopathy for a month. A fine needle aspiration biopsy (F.N.A.B.) and immunohistochemical tests confirm the diagnosis. Inguinal lymphadenopathy was similar to the present case.1\n\nKomeya M et al. reported a case of a middle-aged male with urinary symptoms. During the patient's examination, a nodule was found in the left lobe of the prostate. Further testing revealed that the patient had adenocarcinoma. However, a C.T. scan, magnetic resonance imaging, and bone scan showed no metastasis. The patient received two months of neoadjuvant hormonal therapy (leuproreline and bicalutamide) before undergoing retropubic radical prostatectomy and obturator lymph node dissection. Pathological findings showed that the patient had a moderately differentiated adenocarcinoma. It's important to note that this case was not similar to the present case, as the patient also had inguinal lymphadenopathy and involvement of obturator lymph nodes. Treatment options, their effectiveness, and potential risks and side effects should be discussed with a medical professional.2\n\nDoreswamy K et al. reported that based on the patient's medical history and diagnostic tests, it has been determined that he has prostatic cancer that has spread to the bladder base, seminal vesicles, and vertebral region. The transrectal ultrasound biopsy has confirmed the presence of adenocarcinoma, and the inguinal lymph node biopsy has shown that the cancer has metastasized. The patient has been informed of his diagnosis and is currently undergoing treatment to manage his condition. He needs to continue to receive regular medical care and followup appointments to monitor his progress and address any symptoms that may arise.3\n\nKirpana K. et al. had a similar case of a 65-year-old patient with carcinoma prostate with inguinal lymphadenopathy. As P.S.A. levels were increased, the diagnosis was confirmed with a prostatic biopsy. He underwent an inguinal lymph node Biopsy to confirm the diagnosis of metastasis. Hormonal treatment was advised with a good prognosis of up to ten years of the latest followup without distant metastasis.\n\nOne of the peculiarities of this case report was its presentation as an enlarged lymphadenopathy, as very few were reported in the literature.1–4 One possible explanation might be abnormal or aberrant lymphatic drainage of the prostate to the inguinal lymph node.\n\n\nConclusion\n\nAlthough inguinal metastasis is uncommon in the early stages, the physician must always rule it out. Its existence will significantly affect treatment and management, including hormone therapy with or without locoregional radiation.\n\nWritten informed consent was taken from the patient.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nReferences\n\nRosa M, Chopra HK, Sahoo S: Fine needle aspiration biopsy diagnosis of metastatic prostate carcinoma to inguinal lymph node. Diagn. Cytopathol. 2007 Sep; 35(9): 565–567. PubMed Abstract \| Publisher Full Text\n\nKomeya M, Sahoda T, Sugiura S, et al.: A case of metastatic prostate adenocarcinoma to an inguinal lymph node. Cent. European. J. Urol. 2012; 65(2): 96–97. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDoreswamy K, Karthikeyan VS, Nagabhushana M, et al.: Prostatic adenocarcinoma presenting as isolated inguinal lymphadenopathy. BMJ Case Rep. 2015 Jul 7; 2015: bcr2015210825. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKrpina K, Markić D, Rahelić D, et al.: 10-year survival of a patient with metastatic prostate cancer: Case report and literature review. Arch. Ital. Urol. Androl. 2015 Sep 30; 87(3): 252–253. PubMed Abstract \| Publisher Full Text" }	[ { "id": "253168", "date": "29 Mar 2024", "name": "Chandan Krushna Das", "expertise": [ "Reviewer Expertise Genitourinary Medical Oncology :Prostate Cancer" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\n1)“Around one-half of these cancer patients have metastatic disease at presentation.” Missing Reference 2)We report a rare metastatic prostate adenocarcinoma only to the inguinal lymph nodes without involvement of other nearby reginal nodes. Spelling mistake 3) “Providing more information on the symptoms and diagnosis of metastatic prostate adenocarcinoma could be helpful to readers who are interested in learning more about the disease. This could include details on identifying the signs of metastasis and what tests are typically used to confirm a diagnosis. Additionally, the article could benefit from additional information on treatment options for metastatic prostate adenocarcinoma. This would be especially useful for readers who may be dealing with the disease themselves, like physicians and surgeons. This section looks like out of the context and usually seen in you put your text in any AI software like Chat GPT/Bing AI editing software. 4)“The patient has a past medical case of prostate carcinoma for which he underwent Transurethral resection of the prostate (TURP) with orchidectomy 2 years ago, received two cycles of chemotherapy with docetaxel, and ten sessions of radiotherapy.” It is not clear why TURP and orchidectomy done for a prostate cancer . What was the histology, or Gleason score, and stagging . The most important question is why only 2# of docetaxel and 10 sessions of radiation were given. The correct stagging workup and justification of unusual treatment as compared to the standard of care were missing.\n5)There is no positive family history means no any other family member were affected by similar disease like prostate. This need to be rewritten . 6) The description of examination is unnecessary and deviates from main point\n7) The lobulated hypoechoic mass lesion in the left inguinal canal suggestive of metastatic lymph nodal mass (see Figure 1). After the discovery of metastasis to the liver, the thoracic CT , MDP bone scan or PSMA PET work up is missing .The metastatic workup is incomplete. 8) Tranrectal ultrasound guided fine needle aspiration cytology was performed to reconfirm the diagnosis (see Figure 2A). A biopsy of the prostate rather than a FNAC is the standard of care. Why FNAC was done is unclear. Moreover, Fig. 2A depicts the histopathology of the biopsy specimen rather than cytology, as the author said earlier. 9)Despite all advances in understanding the treatment of prostatic cancer, around two and a half lakhs of persons succumb to death every year worldwide. References are missing and factually wrong Ref : https://seer.cancer.gov/statfacts/html/prost.html. 10) Patients having distant metastasis suggest a poor prognosis with a median survival of up to 2 and half years with considerable inter-patient variations; few may have a short life, and others may survive up to 10 years with hormonal therapy. The cause of this diversity is unknown.Reference missing 11)Although inguinal metastasis is uncommon in the early stages, the physician must always rule it out. Its existence will significantly affect treatment and management, including hormone therapy with or without locoregional radiation. Inguinal metastasis is classified as Stage IV prostate cancer . How the presence of inguinal LN significantly affects treatment with hormonal therapy . Hormonal therapy is the standard of care for all stage IV prostate cancers. The discussion fails to explain why there is an isolated inguinal LN in the absence of a pelvic node. It might be that previous Pelvic RT or prior TURP both alter the usual pathway of LN drainage\n\nIs the background of the case’s history and progression described in sufficient detail? No\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? No\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? No\n\nIs the case presented with sufficient detail to be useful for other practitioners? No", "responses": [] }, { "id": "347731", "date": "16 Dec 2024", "name": "Hiroshi Miyamoto", "expertise": [ "Reviewer Expertise Genitourinary Pathology", "Molecular Biology of Steroid Hormone Receptors in Urological Cancers" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\n1) There are critical issues in Figure 2. 1-A) Figure 2A does not depict the cytopathology from fine needle aspiration (as described in the text). More critically, the image does NOT represent prostatic adenocarcinoma and rather shows non-neoplastic prostate tissue. 1-B) Figure 2B does show an adenocarcinoma morphologically compatible with prostate cancer. However, there is no evidence to conclude that it is a metastatic prostate cancer (even if the patient had prostate cancer). Immunostaining for prostate marker(s), such as Nkx3.1, must be performed to confirm that it is a prostate primary.\n2) Further details about patient’s history of prostate cancer should be provided. 2-A) Why did the patient undergo TURP? Histopathology of the TURP specimen should also be shown. 2-B) Based on the prior treatment (i.e. orchiectomy, chemotherapy, radiotherapy), the patient might have already had advanced disease 2 years ago. In particular, he might have had pelvic lymph node metastasis (before the treatment). 2-C) The topic/title of the present case report (i.e. isolated inguinal lymph node metastasis) may then possibly be inaccurate. Inguinal lymph node metastasis may thus represent a recurrence of advanced prostate cancer (with pelvic lymph node metastasis) following systemic treatment.\n3) This would be a Case Report article, but not a clinical note. There appear to be too many unnecessary/unrelated findings from physical examination and other procedures.\n4) Actual PSA value(s) instead of describing as “elevated” should be provided.\n5) How about CT findings in the liver?\n6) Has bone metastasis been assessed/excluded?\n7) Reference #1 appears to be inadequate to state that prostate cancer is a leading cause of death. Instead, an original epidemiological or clinical study should be cited.\n8) Follow-up information (after the diagnosis of inguinal lymph node metastasis) could be provided.\n9) The Discussion section simply consists of the descriptions of prior case reports. Instead, it has to be used to “discuss” the present case.\n\nIs the background of the case’s history and progression described in sufficient detail? No\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? No\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? No\n\nIs the case presented with sufficient detail to be useful for other practitioners? No", "responses": [] } ]	1	https://f1000research.com/articles/12-1360
https://f1000research.com/articles/12-1358/v1	18 Oct 23	{ "type": "Research Article", "title": "Network Pharmacology Analysis Reveals Bioactive Compounds and Potential Targets of Sea cucumber for Cervical Cancer Therapy", "authors": [ "Irena Ujianti", "Bety Semara Lakshmi", "Zahra Nurusshofa", "Wawang Sukarya", "Leli Indriyanti", "Bety Semara Lakshmi", "Zahra Nurusshofa", "Wawang Sukarya", "Leli Indriyanti" ], "abstract": "Cervical cancer is a leading cause of death among women in many countries, and finding effective anticancer treatments for this type of cancer is challenging due to high rates of HPV infection and low vaccination rates among women of childbearing age. Studies have shown that protein oncogenes produced by HPV stimulate cell growth, promoting tumor development and treatment resistance. It explores the potential therapeutic mechanisms of Scitophus hermanii in treating cervical cancer using network pharmacology, identifying PTGS2, EGFR, and NFE2L2 as targets. Bioactive compounds in sea cucumbers, such as Gangliosides, Stichoposide and variegatuside have the potential to prevent cancer cell proliferation by inhibiting the epidermal growth factor receptor expression. The review suggests that targeting pathways could be a promising strategy for the treatment of cervical cancer. SwissADME also predicted the drug-like properties of the active chemicals in sea cucumbers. This discussion sheds new light on the potential use of marine natural products for the treatment of various types of cervical cancers.", "keywords": [ "Drug discovery", "epidermal growth factor receptor", "pharmacokinetics", "physicochemistry", "sea cucumber", "schistopus hermanii." ], "content": "Introduction\n\nCervical cancer is a significant public health issue worldwide, with an estimated 604,000 new cases anticipated in 2020, according to the World Health Organisation (WHO).1 Unfortunately, the majority of cervical cancer-related deaths (approximately 90%) are expected to occur in low- and middle-income countries, with an estimated 342,000 deaths projected for 2020.1,2 This highlights the urgent need for increased awareness, prevention, and treatment strategies to address this problem. In Indonesia, cervical cancer is the third most common cancer in women, with 36,633 new cases reported in 2020.3 Human papillomavirus (HPV) is the main risk factor for cervical cancer, with types 16, 18, and 45 being the most common.4 HPV has several signalling pathways that are involved in signalling pathway transmission via active molecules such as MEK, ERK, and Akt.5 The virus has several oncoproteins, including E6 and E7, that play a significant role in cancer development. In early stages of cancer, the HPV genome suppresses viral oncoproteins E6 and E7, which maintain Akt phosphorylation status. E6 and E7 activate the Akt/mTOR signalling pathway, promoting viral cap-dependent protein synthesis and leading to carcinogenesis. HPV16 oncoprotein E7 increases keratinocyte migration.6 E5 plays a role in HPV-related cancer proliferation by regulating myogenic signalling pathways and stimulating VEGF expression through ERK activation, which is involved in angiogenesis.7 Cancer eventually develops through these pathways.8\n\nWhile various treatment modalities are available, there is still a need for further investigation into the effects of these treatments. The Surveillance, Epidemiology, and End Results (SEER) programme estimates the overall 5-year relative survival rate for cervical cancer to be 67.2%.9 Therefore, there is a need for continued research to develop more effective treatment strategies. Natural chemicals originating from plants and animals that have potential anti-cancer effects are being researched for new cancer treatments.10 Teripang, also known as Sea cucumber, is a marine natural product that has shown efficacy in medical treatments.11 Researchers have demonstrated the anti-cancer properties of seacucumber in previous studies, but its exact mechanism of action remains unclear. Sea cucumber, is a marine natural product that has shown efficacy in medical treatments. According to current research, a specific group of Sea cucumbers has many promising pharmacological properties.12 This substance is composed of a diverse range of compounds, including various types of polysaccharides, such as glycosaminoglycans, neutral glycans, fucosylated chondroitin sulfates, and sulfated fucans. Studies have shown that Sea cucumber-derived compounds can exhibit cytotoxic activity, induce apoptosis, arrest cell cycle, reduce tumor growth, inhibit metastasis, and prevent drug resistance.13 Cytotoxic activity prevents cancer cell growth. Bioactive carbohydrate compounds from Holothuria scabra species, such as holothurine A3 and A4, are cytotoxic in Hep-G2 and KB cell lines.14 Frondanol A5, derived from Cucumaria frondosa extract, induced apoptosis in pancreatic cancer cells S2013 and AsPC.15 Echinoside A and echonoside A from Pearsonothuria graeffei disrupt the G0/G1 cell cycle of Hep-G2 liver carcinoma cells, preventing DNA replication.16 Saponins from Pentacta quadrangulari, particularly Philinopsides E and A, inhibit tumour growth in sarcoma and hepatoma mouse models.17 Pearsonothuria graeffei bioactive’s compound, Ds-echinoside A, inhibits hepatocellular carcinoma (Hep-G2) cell migration, invasion, and adhesion, thereby reducing cancer cell metastasis.18 The bioactive compounds, potential targets, and underlying mechanisms of Sea cucumber in cervical cancer are not well understood.\n\nThe article aims to investigate the potential of natural products, specifically sea cucumber, as a novel therapeutic strategy for the treatment of cervical cancer. A network pharmacology analysis of S. hermanii, will be presented to identify the active ingredients and targets of S. hermani. The findings of this study may provide a better understanding of the bioactive compounds, potential targets, and underlying mechanisms of S. hermanii, which may be useful for the development of novel therapies for cancer treatment.\n\n\nMethods\n\nThe search for bioactive compounds in Sea cucumbers (Sticophus sp.) was conducted using the CMNPD database.19 Each compound was then searched for its SMILE (simplified molecular-input line-entry system) profile and 3D structure using the PubChem database.20\n\nBased on the information provided, the bioactive compounds found in Sea cucumbers or Schistocopus hermani, were analyzed for their potential using the WAY2DRUG PASS prediction tool as an anticancer treatment.21 The WAY2DRUG Pass Prediction tool uses Structure Activity Relationship (SAR) analysis to compare input compounds with known compounds that have specific potential. The greater the similarity of the structure of the compounds, the higher the prediction value obtained. Compounds with similar structures can be predicted to have similar potential. The Pa value (Probability to be Active) is the output prediction value of the WAY2DRUG PASS, which describes the potential of a tested compound. If the Pa value is greater than 0.7, it indicates that the compound is predicted to have high potential as an anti-inflammatory, for example, because it has a high similarity to compounds in the database. A score of 0.5 is recommended as the cut-off score. The Pa value provides the accuracy of the obtained prediction function, the higher the Pa value of a function, the better the accuracy.22\n\nThe toxicity of bioactive compounds of Sea cucumbers can be predicted using the Protox II database.23 The parameters analyzed include Hepatotoxicity, Carcinogenicity, Immunotoxicity, Mutagenicity, and Cytotoxicity, as described by Banerjee et al. (2018).24\n\nThe targets of teripang were obtained from SuperPred (with score accuracy and probability > 80%).25 The target prediction was obtained by entering the SMILES found in step 1. Genes and proteins related to cervical cancer were obtained from the Open Target database (with overall score prediction ≥ 0.1). The Open Target database was chosen because it includes information from other databases and is the most up-to-date (last update February 2023 (Version 23.02)). The targets related to the disease and the teripang target were then mapped using a Venn diagram to determine the intersection target. The function of each target from the best compound (Variegatusdie) was then mapped using the Database for Annotation, Visualization, and Integrated Discovery version 2021. The R package gplot2 was used to visualize the results of functional annotation from DAVID.26\n\nThe protein target of Variegatuside from teripang was further analyzed using Search Tool for the Retrieval of Interacting Genes/Proteins (STRING DB V.12.0).27 The following parameters were used: Organism: Homo sapiens; Network type: Full STRING network; Required core: medium confidence (0.4). The data format TSV from STRING was then further processed using CytoScape V.10.0 for network analysis.\n\n\nResults\n\nBioactive profile of seacucumber from Comprehensive Marine Natural Products Database (CMNPD).19 In general, this study extracted 16 compounds from the CMNPD database, which were obtained from samples of sea cucumbers in Table 1.\n\nBased on SAR analysis using Way2Drug Pass Online,21 it is found that bioactive compounds in Sea cucumber have a good potential as Chemopreventive (Pa Score: 0.758), Anti-inflammatory (0.566), Antineoplastic (0.774), Apoptosis Agonist (0.614), and Proliferative diseases treatment (0.546). Chemopreventive is the use of natural or synthetic compounds to prevent cancer. Compounds included in chemopreventive can be used to prevent the occurrence of cancer, for someone with a high risk, and can be used to prevent relapse in patients undergoing treatment. The parameters of anti-inflammatory, antineoplastic, apoptosis agonist, and proliferative diseases treatment are used to see the potential as anticancer based on Hallmarks of cancer. Sea cucumber has the highest potential as Chemopreventive and Antineoplastic toxicity in Figure 1.\n\nToxicity analysis of each Sea cucumber sample using the Protox II webserver demonstrated that all compounds analyzed were predicted to exhibit immunotoxicity, with some also displaying cytotoxic effects. The Protox II immunotoxicity model assesses their ability to inhibit B cell growth. Additionally, mutagens are compounds with the potential to induce changes in an organism’s genetic material, while carcinogens can cause cells to become cancerous by altering their genetic structure, leading to uncontrolled cell proliferation. Hepatotoxicity refers to kidney dysfunction or damage associated with an overload of drugs or xenobiotics. Based on the comparison with QSAR data, Variegatuside C and Variegatuside D were identified as potential candidate compounds falling into the toxicity class 4 (range 1 – 6, with lower values indicating higher toxicity), signifying the need for further investigation and evaluation of these compounds as presented in Table 2. Meanwhile, Table 3 displays the profile of the most promising predicted compounds.\n\nFigure 2 shows the Functional Analysis Target of Variegatuside in Sea cucumber, while in Figure 3, it is described that there are 11 overlapping targets between cervical cancer and Sea cucumber, namely MTOR, PDGFRA, PIK3R1, KLF5, NTRK3, HIF1A, CCNE1, AR, TRIM24, HSP90AB1, and TOP2A. MTOR is an oncogene that plays a role in promoting proliferative signalling. It is also involved in triggering invasion, metastasis, angiogenesis, evasion of programmed cell death, and altering cellular energetics (Hallmarks of Cancer: Cosmic Database).\n\nMTOR is considered as a potential target because it has the highest values in terms of betweenness centrality, closeness centrality, degree, and cervical cancer overall score compared to other targets (Table 4, Figure 4). Classic centrality calculations such as degree, closeness, and betweenness centrality are used to identify influential nodes (proteins) in biological networks. Degree provides an indication of how many proteins interact with a protein node. Closeness centrality is useful for estimating how quickly information flows through a node, or in other words, how short the fastest path is from node x to all other nodes. Meanwhile, betweenness centrality is based on communication flow. Nodes with high betweenness centrality values play a role in controlling information flow (Scardoni & Laudanna, 2009) Based on these analyses, it is predicted that Sea cucumber, specifically through the MTOR pathway, may be effective in targeting cervical cancer.\n\nThe blue nodes represent Sea cucumber targets, while the red nodes represent targets of both Sea cucumber and cervical cancer. The diameter of each node indicates its betweenness centrality score, with larger diameters indicating higher scores.\n\n\nDiscussion\n\nOur investigation revealed that Variegatuside C and Variegatuside D were the crucial components responsible for its anti-cancer effects. Recent study findings have identified 11 overlapping targets between cervical cancer and Seacucumber. mammalian Target of Rapamycin (MTOR), Platelet-Derived Growth Factor Receptor Alpha (PDGFRA), Phosphoinositide-3-Kinase Regulatory Subunit 1 (PIK3R1), Krüppel-like Factor 5 (KLF5), Neurotrophic Receptor Tyrosine Kinase 3 (NTRK3), Hypoxia-Inducible Factor 1 Alpha (HIF1A), Cyclin E1 (CCNE1), Androgen Receptor (AR), Tripartite Motif Containing 24 (TRIM24), Heat Shock Protein 90 Alpha Family Class B Member 1 (HSP90AB1), and Topoisomerase II Alpha (TOP2A), are crucial proteins with unique roles in promoting cancer growth and progression. PDGFRA is a protein that promotes cell proliferation and survival in several types of cancer.28 The study conducted by Chang et al. regarding miRNA-487a and its role in promoting proliferation and metastasis in hepatocellular carcinoma, demonstrated that PIK3R1 is a fundamental factor in facilitating cellular survival, growth, and proliferation.29 According to the review conducted by Luo et al., the transcription factor protein KLF5, which is implicated in various cancer types, plays a role in promoting cellular proliferation, growth, and survival.30 Similarly, NTRK3, a receptor tyrosine kinase protein involved in cell survival, proliferation, and differentiation, plays a significant role in cancer development.31 HIF1A is a protein that plays a crucial role in cancer cells’ adaptation to low-oxygen environments, promoting the survival and growth of cancer cells even in challenging conditions.32 CCNE1 is a regulatory protein that contributes to cell cycle progression and is frequently overexpressed in various cancers, leading to the uncontrolled growth of cancer cells.33 Meanwhile, AR is a transcription factor protein that is crucial in the development and progression of prostate cancer, promoting cell growth and survival of androgen-dependent cancer cells. TRIM24 is an oncogenic transcriptional activator that regulates gene expression and promotes cancer cell growth.34 Recent studies have shown that HSP90AB1 is a chaperone protein that plays a critical role in cell signalling pathways related to the growth and survival of cancer cells.35 This is consistent with the findings of Ujianti et al., which suggest that endoplasmic reticulum stress conditions involving HSP90 as a marker for UPR may contribute to the development of liver carcinoma.36 These insights help us better understand the complex role that HSP90AB1 plays in cancer biology and could lead to new potential therapeutic targets for the treatment of liver cancer and other cancers in which this protein is involved. Finally, TOP2A is an enzyme that is often overexpressed in cancer and plays a role in DNA replication and repair, contributing to the proliferation of cancer cells.37 Understanding the functions of these proteins can help researchers develop targeted therapies for cancer treatment develop targeted therapies for cancer treatment.\n\nThe discussion brings attention to the significant potential of MTOR (mammalian target of rapamycin) as a target for cervical cancer treatment. Network analysis, which includes parameters like betweenness centrality, closeness centrality, degree, and cervical cancer overall score, reveals that MTOR possesses the highest values among other targets. These network properties are fundamental in identifying influential nodes or proteins within biological networks. These findings align with the study conducted by Ji et al., which found that MTOR is one of the factors influencing the growth of various cancers, particularly cervical cancer.38 Degree centrality defines the number of proteins interacting with a protein node, while closeness centrality estimates the efficiency of information flow through a node. On the other hand, betweenness centrality is based on controlling the flow of information.39 Considering these network properties, the analysis indicates that targeting MTOR may prove effective in the treatment of cervical cancer. MTOR, a serine/threonine kinase, plays a critical role in cell growth, metabolism, and proliferation. It is well-established that mTOR signalling is involved in multiple cancer characteristics, such as cell growth, survival, metabolism, angiogenesis, and metastasis.40 The activation of mTOR is frequently observed in cancer cells, contributing to their uncontrolled growth. Activation of mTOR signalling can enhance mRNA translation and increase the production of proteins involved in cell cycle progression, thus promoting cancer cell proliferation.41 Additionally, mTOR activation induces metabolic reprogramming in cancer cells, enabling them to adapt to nutrient-deprived and hypoxic conditions.42 The mTOR signalling pathway is also involved in regulating essential cellular processes for cancer progression, including angiogenesis and metastasis. By stimulating the production of vascular endothelial growth factor (VEGF), mTOR signalling promotes the formation of new blood vessels (angiogenesis). Furthermore, mTOR signalling facilitates cancer cell migration and invasion, facilitating the spread of cancer throughout the body.43 Consequently, targeting the mTOR pathway has emerged as a potential therapeutic strategy for cancer treatment. Several mTOR inhibitors, such as rapamycin have been developed and assessed in preclinical and clinical studies. The results are consistent with the SAR analysis conducted in this study using Way2Drug Pass Online, which demonstrated that bioactive compounds found in Sea cucumbers have promising potential as agents for cancer prevention, anti-inflammatory, antineoplastic, apoptosis agonist, and treatment of proliferative diseases. Toxicity analysis using the Protox II webserver showed that Variegatuside C and Variegatuside D are potential candidates, as they belong to toxicity class 4. The use of Protox II as a tool in toxicity analysis is explained in a review study conducted by Benarjee et al.24\n\nThe study suggests that Variegatuside C and Variegatuside D, active ingredients found in seacucumber, show potential as treatment options for cervical cancer. These compounds have the ability to regulate targets associated with the disease, opening up avenues for further exploration and the development of novel therapeutic interventions. However, it is important to consider the limitations of the study. The findings were based on network pharmacology analysis, which relies on computational predictions and may not fully capture the complexities of biological systems. Therefore, further experimental studies, such as in vitro and in vivo experiments on cervical cancer cells and animal models, are needed to validate these findings and provide more concrete evidence. This will help strengthen the evidence presented in the study and provide a more robust understanding of the therapeutic efficacy of Sea cucumber and its active ingredients.", "appendix": "Data availability\n\nFigshare: cmnpd_teripang_final.csv, https://doi.org/10.6084/m9.figshare.23589966. 44\n\nThis project contains the following underlying data:\n\n• CMNPD export.xlsx\n\n• Disgenet.xlsx\n\n• OMIM.xlsx\n\n• QSAR.xlsx\n\n• Target.xlsx\n\n• Target Raw.xlsx\n\n• Venn.xlsx\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nWHO: WHO Mortality Database.2023.\n\nMahendra INB, Prayudi PKA, Dwija IBNP, et al.: HPV16-E6/E7 Oncogene Mutation and p53 Expression among Indonesian Women with Cervical Cancer. Asian Pac. J. Cancer Prev. 2022; 23(8): 2705–2711. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWHO: Cervical Cancer Profile. World Health Organization; 2021.\n\nLiu G-J, Wang Y-J, Yue M, et al.: High expression of TCN1 is a negative prognostic biomarker and can predict neoadjuvant chemosensitivity of colon cancer. Sci. Rep. 2020; 10(1). Publisher Full Text\n\nZhang L, Wu J, Ling MT, et al.: The role of the PI3K/Akt/mTOR signalling pathway in human cancers induced by infection with human papillomaviruses. Mol. Cancer. 2015; 14(1): 13–87. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nStrickland SW, Vande Pol S: The Human Papillomavirus 16 E7 Oncoprotein Attenuates AKT Signaling To Promote Internal Ribosome Entry Site-Dependent Translation and Expression of c-MYC. J. Virol. 2016; 90(12): 5611–5621. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIlahi NE, Bhatti A: Impact of HPV E5 on viral life cycle via EGFR signaling. Microb. Pathog. 2020; 139: 103923. PubMed Abstract \| Publisher Full Text Reference Source\n\nBonab FR, Baghbanzadeh A, Ghaseminia M, et al.: Molecular pathways in the development of hpv-induced cervical cancer. EXCLI J. 2021; 20: 320–337.\n\nCastanon A, Tataru D, Sasieni P: Survival from cervical cancer diagnosed aged 20–29 years by age at first invitation to screening in England: Population-based study. Cancers (Basel). 2020; 12(8): 1–9. Publisher Full Text\n\nLachs L, Oñate-Casado J: Fisheries and Tourism: Social, Economic, and Ecological Trade-offs in Coral Reef Systems. YOUMARES 9 - The Oceans: Our Research, Our Future. 2020; pp. 243–260. Publisher Full Text\n\nWargasetia TL, Widodo.: Mechanisms of cancer cell killing by sea cucumber-derived compounds. Investig. New Drugs. 2017; 35(6): 820–826. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWulandari DA, Gustini N, Murniasih T: Nutritional Value and Biological Activities of Sea Cucumber Holothuria scabra Cultured in the Open Pond System. J. Aquat. 2022; 31: 599–614. Publisher Full Text\n\nRoginsky A, Ding X-Z, Singh B, et al.: Frondanol-A5 from cucumaria frondosa induces cell cycle arrest and apoptosis in pancreatic cancer cells. J. Am. Coll. Surg. - J. AMER. COLL. Surg. 2004 Sep 1; 199: 91. Publisher Full Text\n\nSalindeho N, Nurkolis F, Ben GW, et al.: Anticancer and anticholesterol attributes of sea cucumbers: An opinion in terms of functional food applications. Front. Nutr. 2022; 9(1). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHossain A, Dave D, Shahidi F: Northern sea cucumber (Cucumaria frondosa): A potential candidate for functional food, nutraceutical, and pharmaceutical sector. Mar. Drugs. 2020; 18(5). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZhao Q, Xue Y, Liu ZD, et al.: Differential Effects of Sulfated Triterpene Glycosides, Holothurin A1, and 24-Dehydroechinoside A, on Antimetastasic Activity via Regulation of the MMP-9 Signal Pathway. J. Food Sci. 2010; 75(9): H280–H288. Publisher Full Text\n\nIrena U: CMNPD-database.2023 [cited 2023 Jul 19]. Reference Source\n\nIrena U: PubChem.2023 Jul 22 [cited 2023 Jul 22]. Reference Source\n\nIrena U: WAY2DRUG PASS.2023 Jul [cited 2023 Jul 21]. Reference Source\n\nIrena U: Protox-II.2023 Jul 21 [cited 2023 Jul 21]. Reference Source\n\nIrena U: superPred.2023 Jul 21 [cited 2023 Jul 21]. https://zenodo.org/record/8170980\n\nFilimonov DA, Laqunin AA, Gloriozova TA, et al.: Prediction of the Biological Activity Spectra of Organic Compounds Using the PASS online Web Resource. Chem. Heterocycl. Comp. 2014; 50: 444–457. Publisher Full Text\n\nIrena U: DAVID.2023 Jul 21 [cited 2023 Jul 21]. Reference Source\n\nBanerjee P, Eckert AO, Schrey AK, et al.: ProTox-II: A webserver for the prediction of toxicity of chemicals. Nucleic Acids Res. 2018; 46(W1): W257–W263. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIrena U: String_DB.2023 Jul 21 [cited 2023 Jul 21]. Reference Source\n\nChoudhari AS, Mandave PC, Deshpande M, et al.: Phytochemicals in cancer treatment: From preclinical studies to clinical practice. Front. Pharmacol. 2020; 10(January): 1–17. Publisher Full Text\n\nMazlan NB, Abd Rahman NNB, Shukhairi SSB, et al.: Sea Cucumbers: Source of Nutritional, Medicinal, and Cosmeceutical Products BT - Marine Biotechnology: Applications in Food, Drugs and Energy. Shah MD, Ransangan J, Venmathi Maran BA, editors. Singapore: Springer Nature Singapore; 2023; pp. 171–188. Publisher Full Text\n\nPapadopoulos N, Lennartsson J: The PDGF/PDGFR pathway as a drug target. Mol. Asp. Med. 2018; 62: 75–88. PubMed Abstract \| Publisher Full Text Reference Source\n\nChang R-M, Xiao S, Lei X, et al.: miRNA-487a Promotes Proliferation and Metastasis in Hepatocellular Carcinoma. Clin. Cancer Res. 2017 May 14; 23(10): 2593–2604. PubMed Abstract \| Publisher Full Text\n\nLuo Y, Chen C: The roles and regulation of the KLF5 transcription factor in cancers. Cancer Sci. 2021; 112(6): 2097–2117. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChen Z, Huang Z, Luo Y, et al.: Genome-wide analysis identifies critical DNA methylations within NTRKs genes in colorectal cancer. J. Transl. Med. 2021; 19(1): 13–73. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMasoud GN, Li W: HIF-1α pathway: Role, regulation and intervention for cancer therapy. Acta Pharm. Sin. B. 2015; 5(5): 378–389. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGorski JW, Ueland FR, Kolesar JM: CCNE1 amplification as a predictive biomarker of chemotherapy resistance in epithelial ovarian cancer. Diagnostics. 2020; 10(5): 1–14. Publisher Full Text\n\nHoang DT, Iczkowski KA, Kilari D, et al.: Androgen receptor-dependent and -independent mechanisms driving prostate cancer progression: Opportunities for therapeutic targeting from multiple angles. Oncotarget. 2017; 8(2): 3724–3745. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAlbakova Z, Mangasarova Y, Albakov A, et al.: HSP70 and HSP90 in Cancer: Cytosolic, Endoplasmic Reticulum and Mitochondrial Chaperones of Tumorigenesis. Front. Oncol. 2022; 12(January): 1–14. Publisher Full Text\n\nUjianti I, Sianipar IR, Prijanti AR, et al.: Effect of Roselle Flower Extract (Hibiscus sabdariffa Linn.) on Reducing Steatosis and Steatohepatitis in Vitamin B12 Deficiency Rat Model. Med. 2023; 59: 1044.\n\nPommier Y, Nussenzweig A, Takeda S, et al.: Human topoisomerases and their roles in genome stability and organization. Nat. Rev. Mol. Cell Biol. 2022; 23(6): 407–427. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJi J, Zheng PS: Activation of mTOR signaling pathway contributes to survival of cervical cancer cells. Gynecol. Oncol. 2010; 117(1): 103–108. PubMed Abstract \| Publisher Full Text\n\nScardoni G, Tosadori G, Faizan M, et al.: Biological network analysis with CentiScaPe: centralities and experimental dataset integration. F1000Res. 2015; 3(139). PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source\n\nPapadopoli D, Boulay K, Kazak L, et al.: mTOR as a central regulator of lifespan and aging. F1000Res. 2019; 8(998). PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source\n\nYang M, Lu Y, Piao W, et al.: The Translational Regulation in mTOR Pathway. Biomolecules. 2022; 12(6). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMagaway C, Kim E, Jacinto E: Targeting mTOR and metabolism in cancer: Lessons and innovations. Cells. 2019; 8(12). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLotfimehr H, Mardi N, Narimani S, et al.: mTOR signalling pathway in stem cell bioactivities and angiogenesis potential. Cell Prolif. 2023; e13499. PubMed Abstract \| Publisher Full Text\n\nUjianti I, Semara Lakshmi B, Nurushoffa Z, et al.: cmnpd_teripang_final.csv. Dataset. figshare. 2023. Publisher Full Text" }	[ { "id": "216762", "date": "14 Nov 2023", "name": "Mohd Rehan", "expertise": [ "Reviewer Expertise Computational Biology" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis research focuses on uncovering the anticancer mechanisms of sea cucumbers, resulting in the identification of two critical bioactive compounds with mTOR as their target. However, I have a few concerns that, if addressed, can significantly enhance the quality of the manuscript:\nA comprehensive flow diagram depicting the entire process, along with a clear representation of the number of compounds and targets filtered at each step.\n\nThe two proposed compounds exhibit hepatotoxic and immunotoxic properties. To reconcile this, it is imperative to discuss how these adverse effects can be managed or minimized in a clinical context and provide recommendations for their potential use in cancer therapy.\n\nIncluding the chemical structures of the two proposed compounds is essential. Also, perform a Lipinski Rule of Five prediction for these compounds to assess their drug-likeness and potential for further development as therapeutic agents.\n\nConsider conducting molecular docking studies of these compounds with mTOR targets to evaluate their binding potential. This would provide valuable insights into their interaction and affinity with the target, further supporting their role in anticancer mechanisms.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1358
https://f1000research.com/articles/12-1357/v1	18 Oct 23	{ "type": "Case Report", "title": "Case Report: Management of self-injurious habit in a pediatric patient using soft splint", "authors": [ "Sakshi Kabra", "Nilima Thosar", "Monika Khubchandani", "Shriya Singi", "Sakshi Kabra", "Nilima Thosar", "Monika Khubchandani" ], "abstract": "Mouth ulcer is a common clinical manifestation of the oral cavity that occurs in the oral mucosa and is usually associated with a variety of causes and diseases. This type of habitual injury in children is transient and tends to worsen over time. Various treatment options have been implemented, therefore in this case, the treatment of cheek biting was performed using palliative therapy in conjunction with a soft occlusal splint in a 9-year-old child. The treatment plan involved a soft occlusal splint along with antibiotic coverage and multivitamin therapy. After 1 month follow-up complete healing of the ulcer was seen. This case presented a conservative treatment approach for repeated cheek biting without harm to the tooth structure. Early detection of lesions and identification of risk factors allows for a more conservative clinical approach. It is preferable to rule out the cause behind the lesion and accordingly plan the treatment which fits the patient's needs.", "keywords": [ "Traumatic Ulcer", "soft splint", "self-injuries habits", "Antibiotics", "cheek biting", "oral ulcers." ], "content": "Introduction\n\nMouth ulcer is a recurrent oral cavity clinical manifestation in the oral mucosa and is usually related to various causes and diseases.1 The causes of check biting are Tooth deflection in the dental arch, depression-related cheek biting, Biting the cheek accidentally, Psychological, biting the cheek while sleeping during the night, malignant lesions, etc.2 As a result, determining the exact cause of this oral lesion is challenging. Morsicato mucosae oris, or chronic biting of the oral mucosa is a type of accidental or intentional injury that commonly affects the buccal and labial mucosa and the lateral surface of the tongue. Such transient habitual injuries in children tend to become severe over time; howbeit, stressful situations including examinations, sports events, and other activities may intensify the condition.3\n\nPedodontists often encounter such behavioural patterns in children and are the ones who are consulted about it. Various treatment options, such as reconditioning the patient through strategic counselling, relaxation, and calmative, focusing the habit, and protecting the oral mucous membrane from injury by means of prosthesis such as a variety of removable appliances that guard the oral structures including buccal mucosa, tongue, and lips from chronic lesions, have been implemented and documented in the past to prevent and control repetitive accidental trauma to the oral mucosa.4–6 The treatment procedure should be carefully considered based on the clinical stage and form of the ulcer. Therefore, the current clinical study outlines the treatment of chronic cheek biting with pain management and fabricating a specifically designed soft occlusal splint.\n\n\nCase report\n\nA nine-year-old male reported to the Department of Pedodontics and Preventive Dentistry at Datta Meghe Institute of Higher Education and Research, Wardha, with the chief complaint of mouth ulcers in the lower right and left back teeth region for four weeks. Medical, genetic, and family history was non-contributary. He had been examined and given systemic antibiotics for two weeks at another clinic, but no improvement had occurred. His parents decided to transfer him to our facility after becoming concerned when a clinic doctor requested a biopsy to make an initial diagnosis.\n\nWhen the patient reported to the department, he was afebrile and unaware of the daytime cheek-biting habit. The patient had a swollen face with a 2 x 2 cm size of the submandibular lymph node. The patient had no crowded teeth and could bite usually. While observing his behaviour, he kept sucking his cheeks. Therefore, traumatic mucosal ulcers on both cheeks were made as the final diagnosis. (Figure 1 and 2) Differential Diagnosis can be aphthous ulcer and early squamous cell carcinoma or may be a result of infectious disease.\n\nIn the treatment plan, initially, the patient's parents informed consent was recorded and oral prophylaxis was performed, and the patient started his antibiotic coverage. The medication included amoxiclav (375mg maximum two times a day), ibuprofen (200mg maximum two times a day), and Metronidazole (200mg maximum two times a day) for three days. In the same appointment, an alginate impression was taken of both the arch, and a soft occlusal splint was made using a soft polyvinyl sheet of 2mm thick resilient material. The sheet was placed in a vacuum-formed pressure molding with a thermally controlled infrared heater. After one week, the occlusal splint was placed over the maxillary arch and extended laterally from molar to molar by releasing the area where the premolar erupted. (Figure 3) The patient was instructed to wear the appliance during the day after school and at night while sleeping. 5 ml of 2% chlorohexidine mouthwash was prescribed to the patient for seven days.\n\nAfter complete treatment patient was recalled after ten days, the lesion on both sides was almost healed.\n\nAfter 30 days of treatment, the lesions on the left cheek were completely healed (Figure 4), and the one on the right cheek was almost healed. (Figure 5)\n\n\nDiscussion\n\nWith an average of 21.7 cases per 1000 patients, cheek biting was the fifth most frequent cause of mouth lesions.7 In which women are more likely than men to develop this habit.8 Biting repeatedly causes a severely traumatized lesion that is sometimes scarred, thickened, and paler than the neighbouring mucosa, or it can present as white wrinkled to dry surfaces that are tender or not. It can also present as swelling, purpura, and erosions.3 In the present case lesion was white frayed which was associated with swelling.\n\nVarious dental appliances for controlling oral mucosa biting have been reported in the literature. Examples include lip bumpers, mouth guards, and silicone soft relining material for tongue protection.9 In this case, an occlusal soft splint was used to provide total coverage of the functional cusps of the molars and to prevent repeated traumatic injury. Although an oral prosthesis does not address the underlying cause of oral mucosa biting, it effectively controls this self-mutilation.\n\nIn this case, a biopsy was not performed, as it was a pediatric patient. Biopsy tests performed during acute lesions may show incorrect results, promote further injury, and aggravate patients' pain and fear. According to Ngoc et al., a biopsy should be performed for malignant lesions, specifically for pediatric patients.2\n\nChlorhexidine has manifested activity against some enveloped viruses such as CMV, HSV, and Influenza.10 In the present case, 2% chlorohexidine mouthwash with antibiotics and multivitamin therapy was prescribed, which had an essential role in the healing of the ulcer in conjunction with the soft splint. To avoid any adverse events such as staining of tooth using 2% chlorhexidine, it was prescribed for a week.\n\nThe only limitation of this case was that the treatment plan was only custom suited for a case of habitual cheek biting.\n\n\nConclusion\n\nEarly detection of lesions and identification of risk factors allows for a more conservative clinical approach. As a result, before planning the treatment course for any oral soft tissue lesions, identification of the primary cause such as a repeated habit or other factors is a prime requisite. This is because neglecting the cause of such lesions and treating the same with a surgical approach like excision may lead to the recurrence of the lesion. The soft occlusal splint, fabricated using polyvinyl sheets, is an easy-to-wear simple prosthetic device that can be customized according to the patient.\n\nWhen the child was nine years old, we noticed mouth ulcer in right and left side of cheek region, it lead to pain and discomfort to the child. We had shown this to private practitioner who suggest biopsy for the same which causes fear and anxiety. So, we decided to take our child to this institute, after splint therapy our child was free from pain and ulcers were almost healed.\n\n\nConsent\n\nWritten informed consent for publication of their clinical details and the clinical image was obtained from the patient’s parent.", "appendix": "Data availability\n\nZenodo. CARE checklist. DOI: 10.5281/zenodo.8313857\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC BY 4.0 Public domain dedication).\n\n\nReferences\n\nBruce AJ, Dabade TS, Burkemper NM: Diagnosing oral ulcers. JAAPA. 2015 Feb 1; 28(2): 1–10. Publisher Full Text\n\nNgoc VT, Hang LM, Van Bach H, et al.: On-site treatment of oral ulcers caused by cheek biting: A minimally invasive treatment approach in a pediatric patient. Clinical Case Reports. 2019 Mar; 7(3): 426–430. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFlaitz CM, Felefli S: Complications of an unrecognized cheek biting habit following a dental visit. Pediatr. Dent. 2000 Nov 1; 22(6): 511–512. PubMed Abstract\n\nPeters TE, Blair AE, Freeman RG: Prevention of self-inflicted trauma in comatose patients. Oral Surg. Oral Med. Oral Pathol. 1984 Apr 1; 57(4): 367–370. Publisher Full Text\n\nWillette JC: Lip-chewing: another treatment option. Spec. Care Dentist. 1992 Jul; 12(4): 174–176. Publisher Full Text\n\nRomero M, Vicente A, Bravo LA: Prevention of habitual cheek biting: a case report. Spec. Care Dentist. 2005 Aug; 25(4): 214–216. PubMed Abstract \| Publisher Full Text\n\nCastellanos JL, Díaz-Guzmán L: Lesions of the oral mucosa: an epidemiological study of 23785 Mexican patients. Oral Surg. Oral Med. Oral Pathol. Oral Radiol. Endod. 2008; 105: 79–85. PubMed Abstract \| Publisher Full Text\n\nSaemundsson SR, Roberts MW: Oral self injurious behavior in the developmentally disabled: review and a case. ASDC J. Dent. Child. 1997; 64: 205–9, 228. PubMed Abstract\n\nBhatia SK, Goyal A, Kapur A: Habitual biting of oral mucosa: A conservative treatment approach. Contemporary clinical dentistry. 2013 Jul; 4(3): 386–389. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBelenguer-Guallar I, Jiménez-Soriano Y, Claramunt-Lozano A: Treatment of recurrent aphthous stomatitis. A literature review. J. Clin. Exp. Dent. 2014 Apr; 6(2): e168–e174. PubMed Abstract \| Publisher Full Text" }	[ { "id": "271654", "date": "07 May 2024", "name": "Eduardo David Piemonte", "expertise": [ "Reviewer Expertise Oral cancer risk factors", "with special emphasis in intraoral factors such as chronic mechanical irritation of the oral mucosa" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe article needs major revisions before being accepted, although it proposes a simple and non-aggressive technique to resolve clinical situations difficult to resolve with other techniques, and therefore could be useful in the dental field beyond the specific clinical case.\nThe introduction is confusing and somewhat disorganized, since it mentions mouth ulcers, but immediately refers to chewing of the cheek, which does not always manifest as an ulcer. I suggest that the introduction be developed from the general to the particular, following the following order, although not strictly: What are the most common lesions in the oral mucosa in pediatric patients? What are the lesions of the oral mucosa associated with accidental or chronic trauma in pediatric patients? What are the habits (dysfunctional or parafunctional factors) that facilitate the traumatizing effect of teeth in pediatric patients? What is the clinical challenge that patients with these pathologies represent?\nIn the description of the clinical case it is necessary to improve the description of the causal dental factor: which teeth were involved, if the buccal cusps of the upper molars were very sharp, if when the molars were occluded they did so in a normal position or if they had some transverse alteration of the occlusion. If these data demonstrated that the dental factor was relevant, the presence of the cheek-biting habit shows how the dental factor interacts with the functional factor, since there are many children who may have acute cusps but not all of them develop injuries due to trauma, even if they have some alteration in its position. If there were no relevant dental factor, this would show that the functional factor has the power for an apparently non-traumatizing dental element to produce traumatic ulcers. That will be defined according to the reworking of the description by the authors. Regarding differential diagnoses, there are other options to consider in pediatric patients. Due to the bilateral nature of the ulcers, differential diagnoses should be more oriented towards systemic diseases such as infections (which must be specified) or malignant diseases such as leukemias, since ulcers are observed without much erythema, and with an acute evolution. The presence of squamous cell carcinoma in a 9-year-old child is an extremely exceptional fact, and even more so is the fact that it is bilateral.\nThe discussion must be reworked so as not to repeat concepts already included in the introduction. Suggestions for organizing the discussion, although not strictly speaking, may be the following: What were the bases for defining the diagnosis of traumatic injury compared to other differential diagnoses? What were the reasons for using antibiotics and chlorhexidine in this patient? What evidence did you find to support the use of soft plates for the treatment of traumatic injuries, not only in children but also in patients of any age? Is there another way to manage the trauma generated by a chewing habit? Does the use of soft plates constitute a reversible treatment? What advantages would that have? Does the use of these soft plates really have the limitation of only being able to be used in cases with a similar diagnosis? Or are there other types of trauma injuries, in the buccal mucosa and edges of the tongue, that could be treated with this technique?\nThe conclusion could be reworked, if the authors consider it necessary taking into account the modifications in the rest of the manuscript.\n\nIs the background of the case’s history and progression described in sufficient detail? Partly\n\nAre enough details provided of any physical examination and diagnostic tests, treatment given and outcomes? Partly\n\nIs sufficient discussion included of the importance of the findings and their relevance to future understanding of disease processes, diagnosis or treatment? Partly\n\nIs the case presented with sufficient detail to be useful for other practitioners? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1357
https://f1000research.com/articles/10-1199/v1	25 Nov 21	{ "type": "Research Article", "title": "Effect of Nigella sativa on general health and immune system in young healthy volunteers; a randomized, placebo-controlled, double-blinded clinical trial", "authors": [ "Ayad Salem", "Abdullah Bamosa", "Mohammed Alam", "Saleh Alshuraim", "Hamad Alyalak", "Abdulrahman Alagga", "Faisal Tarabzouni", "Omar Alisa", "Hussein Sabit", "Ashfaq Mohsin", "Mohammad Shaikh", "Abdulaziz Farea", "Thamer Alshammari", "Obeid Obeid", "Ayad Salem", "Mohammed Alam", "Saleh Alshuraim", "Hamad Alyalak", "Abdulrahman Alagga", "Faisal Tarabzouni", "Omar Alisa", "Hussein Sabit", "Ashfaq Mohsin", "Mohammad Shaikh", "Abdulaziz Farea", "Thamer Alshammari", "Obeid Obeid" ], "abstract": "Nigella sativa (N. sativa) is traditionally used as an immune enhancer in different communities. The aim of this study was to evaluate the effect of N. sativa on immunity related parameters in young healthy subjects. This study was a double blind, randomized, placebo controlled clinical trial. Fifty-two healthy subjects (48 male and 4 female) 18-25 years old were enrolled in the study. They were randomly divided into four groups; the first received charcoal capsules and served as controls and the other three received 0.5, 1 g, and 2 g of powdered N. sativa capsules, respectively. Two blood samples were obtained from all participant, before initiation of the trial and at the end of the four weeks intervention. One sample was used for routine health screening by evaluating liver and renal functions as well as complete blood count and differential. The second sample was used to measure certain cytokines including; IL-1, IL-4, IL-6, IL-10, and TNF. A third and fourth samples were obtained from the last cohort of subjects before and after treatment; the third was used for measuring immunoglobulins and CD profile and the fourth for evaluating certain gene expressions (INF-γ, NF-κ-B, TNF-α, IL-1β, IL-13, IL-8, and IL-6). Only 1 g dose of N. sativa produced a significant elevation in total lymphocyte count, CD3+ and CD4+ counts. One gram N. sativa increased the absolute lymphocyte count from 1850±0.24 to 2170±0.26 (p=0.008), CD3+ from 1184.4±75.60 to 1424±114.51 (p=0.009), and CD4+ from 665.6±141.66 to 841±143.36 (p=0.002). This elevation in T cells was lost by increasing the dose of N. sativa to 2g. The rest of the parameters were not changed significantly in all doses. The results show a promising immunopotentiation effect of N. sativa by elevating helper T cells and the optimum dose for young age group seems to be 1 g.", "keywords": [ "Nigella sativa", "black seed", "immunoglobulins", "CD", "immune system", "immunity", "gene expression" ], "content": "Introduction\n\nNigella sativa is one of the most used herbal medicines worldwide nowadays. It has been used for more than 2000 years as a natural remedy for various illnesses. Research has documented its therapeutic potential as an antimicrobial, anti-inflammatory, antioxidant, antidiabetic, antihypertensive, antitumor, and immunomodulatory agent.1,2\n\nThe immune system consists of multiple linked networks of cells, proteins, and lymphoid organs to provide protection against millions of microbes and infections. The immune system includes the innate immunity and the adaptive immunity, where the innate immunity provides an immediate protection to the body, and its effect is merely the same in different individuals, while the adaptive immunity, takes more time to develop, but it is more specific and fatal to invasive pathogens.3 The immunomodulatory effect of N. sativa has been studied extensively on both innate and adaptive immunity as well as related messengers and mediators.4–6\n\nSeveral in vivo studies showed significant effects of N. sativa on immunity, autoimmune diseases, and toxicity. A study demonstrated that aqueous extract of N. sativa showed a significant increase (62.3% ± 6.4%) in the natural killer (NK) cytotoxic cell activity against YAC-1 cells after 1 week of oral administration in a 10-week old BALB/c female mice.7 Another in vitro study on the effect of thymoquinone (TQ) on immunity showed that TQ injected directly to cells in low concentrations (10, 2.5 or 0.62 μg mL−1), increased the survival of activated T-cells, CD8+ T-cells ability to generate IFN-γ, which indicates that TQ may be beneficial against infectious diseases and enhancing immunity.8 N. sativa supplementation to newly evolved crossbred laying hens at levels of 4% or 5% positively enhanced immunity against Newcastle disease virus.9 In another study on Newcastle virus vaccinated broilers, N. sativa supplementation in three doses (5, 10 and 20 g kg−1) for 42 days, increased anti-bodies against Newcastle virus significantly on day 35.10 A diet supplemented with 40 g kg−1 N. sativa fed to broiler chicks improved anti-body production against both Newcastle virus and infectious bursal disease.11 The phagocytic index and rate were significantly higher in STZ-diabetic hamsters treated with N. sativa oil (NSO) at a dose of 400 mg·kg−1 for 4 weeks compared with untreated diabetic animals, as demonstrated by fluorescence microscopy.12 N. sativa extract has stimulatory effect on cellular immunity, in vitro, by increasing the proliferative capacity of T lymphocytes and splenocytes and response to a different mitogens of the human peripheral blood mononuclear cells (PBMC).13 NSO was shown to possess a protective role against vitamin A hypervitaminosis. Rats treated with 800 mg·kg−1 NSO orally showed higher serum levels of IgG and IgM than either the control or those with high doses of vitamin A.14 Gestational diabetes rats showed improvement in their offspring immune status, after oral antenatal feeding with 20 mg kg−1 TQ, by reversing the decreased levels of IL-2, T-cell reproduction, and improving both circulating and thymus homing T-cells proliferation.15\n\nThe above literature shows a very promising immunomodulating effect of N. sativa. However, the immunopotentiation effect of this miracle plant has not been investigated in normal humans. Hence this study was designed to evaluate the impact of different doses of N. sativa on the immune system in young healthy humans.\n\n\nMethods\n\nThis is a placebo-controlled, double blinded, randomized clinical trial. The study was conducted on healthy male and female students studying in Imam Abdulrahman bin Faisal University (IAU), Dammam, Saudi Arabia. Blood extraction was carried out in the main campus University Family Medicine Center. Students received the intervention for one month and were divided into four groups; three were given different doses of black seed and the fourth was given charcoal and served as control.\n\nThe participants were students enrolled in different colleges in IAU. Subjects were randomly divided into four groups; 30 participants were allocated to each group through computer generated randomization table. The sample size was determined based on our previous clinical trials using N. sativa.16,17 The first group was the control group (placebo) and they were given 162 mg of activated charcoal capsules, second group received 500 mg N. sativa capsules, third and fourth group received 1 and 2 g N. sativa capsules, respectively.\n\n\n\n1. Healthy IAU students\n\n2. Age between 18 and 25 years\n\n3. BMI = 18.5–29.9 kg m−2\n\n\n\n1. Subjects with any acute or chronic illness (unless acute illness occurred during the study)\n\n2. Subjects with abnormalities in the basic laboratory investigations\n\n3. Participants with less than 90% compliance\n\n\nMaterial\n\nEthiopian N. sativa, bought from the local market, was cleaned, ground and assembled into 500-mg capsules, in the pharmaceutics laboratory in the College of Clinical Pharmacy at IAU. Activated charcoal capsules (162 mg) similar in size and color to the capsules of N. sativa (Arkopharma Pharmaceutical Laboratories Carros, France) were used as placebo. The placebo capsules were given in the same bottles as the N. sativa capsules. Each participant was given enough capsules for the period of 4 weeks. Bottles in each group were coded by the technical staff in the laboratory to achieve the double blindness in the study. The code was unmasked at the end of the study after statistical analysis of all data.\n\nAfter applying the inclusion and exclusion criteria mentioned above, recruited participants were given a full explanation of the study and required procedures and those who agreed to join, signed written consent. Subjects were recruited in three cohorts on Sundays from 08:00–10:00 h, in the period of February and March, 2020. Full history and physical examination were obtained from each participant to rule out any acute or chronic illnesses. Two blood samples were collected from all participants, in the Family and Community Medicine (FAMCO) Center in the IAU campus, before initiation of the study and at the end of the four-week study duration. The first sample was assessed in the center’s laboratory for basic tests which included complete blood count (CBC), renal function test (RFT) and liver function test (LFT) to assure the general health of the participant. The second sample was assessed in the microbiology laboratory in the College of Medicine at IAU to determine baseline cytokines level by enzyme-linked immunosorbent assay (ELISA). A third and fourth blood sample was collected from the last cohort of participants, before and after intervention. The third sample was used to measure CD profile by flow cytometric kits as well as immunoglobulins (IgG, IgM). The fourth blood sample was collected in heparin tubes to evaluate the gene expression profile of IFN-γ, NF-κB, TNF-α, IL-1β, IL-13, IL-8, and IL-6. Participants were followed daily by telephone calls for the whole study period (4 weeks) to ensure taking the capsules. ELISA kits for IL-1, IL-4, IL-6, IL-10, and TNF were bought from Origin company, USA, and the cytokines levels were measured according to the manufacturer recommendation. CD profile was measured using flow cytometric kits (TBNK kit, BD biosciences, USA).\n\nThe study has been approved by the university ethical board under the reference (IRB-2020-UGS-01-032) and was registered in ISRCTN registry (ISRCTN14150499, 16/11/2020, https://doi.org/10.1186/ISRCTN14150499).\n\nWe have evaluated the changes in the expression of different immunity-related genes listed in the table below, before and after intervention.\n\nStepOne Plus thermal cycler was used in this study, and the thermal profile was as follows: 94°C for 2 min as pre-PCR and 95°C for 30 sec, 62°C for 45 sec, and 72°C for 45 sec for 35 cycles. Followed by 72°C for 10 min as post-PCR step. The 2−ΔΔCt equation was used to analyze the fold change.\n\nStatistical analysis was performed based on intension to treat protocol using the Statistical Package of Social Science (SPSS) version 16 (RRID:SCR_019096); JASP (RRID:SCR_015823) is an open-access alternative. Data is presented as mean ± SD (standard deviation). In each group, readings were compared to their corresponding baseline values using Student’s t-test for paired data. Results in the four groups were compared using ANOVA. A P-value <0.05 was considered as significant.\n\n\nResults\n\nParticipants were invited to join the study through a web page which included the consent form. The total number screened was 137 participants, 43 participants were excluded according to inclusion/exclusion criteria, and 18 refused to participate. Those fulfilling the criteria and agreed to participate were given appointments in the Family Medicine Center of Imam Abdulrahman bin Faisal University. 76 participants were enrolled in this study over three Sundays before intervention and three Sundays after intervention in the period February to March, 2020. 10 participants were excluded due to poor compliance (<90%), while 14 have withdrawn or lost to follow up. All participants tolerated the intervention, and no side effects were reported throughout the four weeks of treatment. Furthermore, all basic investigations including renal and liver function tests, and CBC were within normal limits. The study flow chart is shown in Figure 1. All groups were well matched in age, sex, BMI and other baseline characteristics as shown in Table 1.\n\nVital signs were obtained from the participants before and after intervention; heart rate (HR), blood pressure (BP), and the mean arterial pressure (MAP). Group 2 participants showed a significant decrease in HR and systolic blood pressure from baselines (86.43 ± 18.48 beats/min versus 76.14 ± 11.35 beats/min, P < 0.05; and 130.15 ± 13.42 mmHg versus 119.69 ± 12.83 mmHg, P < 0.05), respectively. Moreover, group 3 showed a significant decrease in the diastolic blood pressure from 79.09 ± 7.46 to 66.09 ± 10.08 (P < 0.05). No other significant changes were shown in HR, SBP, DBP, and the calculated MAP among groups (Table 2).\n\nChanges in lymphocyte profile are presented in Table 3. Lymphocyte absolute count and CD3+ lymphocytes were significantly increased in group 3, receiving 1 g N. sativa, compared with the baseline (P < 0.05). T-helper cells (CD4+) were also significantly increased in group 3 (P < 0.05). There was no significant change in other types of lymphocytes among all intervention groups.\n\nImmunoglobulins IgG and IgM showed non-significant changes in all intervention groups Table 4. The value of IL-4 and IL-10 were below the detection limits of the ELISA kits, the other cytokines (IL-1β, IL-6) did not show any significant changes in all groups (Table 4).\n\nThe expression profile of inflammation- and cancer-related genes (IL-1β, IL-8, TNF-α, IFN-γ, IL-6, NF-κB, and IL-13) were measured using qPCR. Box blotting (Figure 2 and Table 5) showed no significant changes in the expression level of the studied genes in pre-intervention compared to post-intervention readings (P-values of 0.15, 0.48, 0.15, 0.48, 0.37, 0.12, and 0.28 for IFN-γ, NF-κB, TNF-α, IL-1β, IL-13, IL-8, and IL-6, respectively).\n\nQuestionnaire: Online questionnaire was distributed among the participants after the end of the study, the questionnaire contained seven questions, four of which were multiple choice questions, three answers were pre-set to: yes-positive effect, yes-negative effect and no change, and an option to add other observations. The control group had more infections during the period of the study in comparison to all test groups. More subjects reported a positive change in their daily activity in groups 3 and 4.\n\nNo participants of the control group noticed any change in their concentration (n = 15). On the other hand, 27.8% of the third group (n = 5) noted a positive change in their concentration. Most of the subjects in the control group (n = 14) did not notice any change in their sleep, while in Group 2, 12.5% (n = 2) noted a negative change in their sleep pattern. In groups with higher doses (Groups 3 and 4), more participants noted a positive change in their sleep pattern. The questionnaire results are showed in (Table 6).\n\n\nDiscussion\n\nThe use of natural products and herbs as medicines is the practice of humans for centuries. One of the main mechanisms by which such herbs produce their beneficial effect on health is immunomodulation which involves stimulation or inhibition of the immune system.18 Cardamom and black pepper are good examples of such herbs which possess a potent immunomodulatory effects.19\n\nN. sativa, commonly known as black seed or black cumin, is a very common herb which belongs to the Ranunculaceae family.20,21 Several therapeutic effects have been attributed to N. sativa and its active ingredient, thymoquinone, including anti-histaminic, anti-inflammatory, anti-hypertensive, hypoglycemic, anti-cancer, and immunity-boosting effects21,22 The immunoregulatory effect of N. sativa has been studied in animals and several positive potential effects in enhancing immunity have been reported. However, the current clinical trial is the first to study the immunomodulatory effect of N. sativa on healthy humans, and the first to investigate its optimal dose.\n\nOur results showed a significant increase of T-helper cells (CD4+), this is in agreement with a study conducted on beta-thalassemia major children where N. sativa enhanced the cell-mediated immunity significantly via increasing CD4 counts (from 1319.88 ± 74.56 to 2007.64 ± 90.34 cells μL−1) (P < 0.001).23 This study also reported a significant increase in T-suppressor cells from 727.09 ± 42.81 cells μL−1 to 1145.31 ± 77.58 cells μL−1 after N. sativa intervention, (P < 0.001). However, this parameter was non-significantly increased in our study from 425.8 ±77.62 cells μL−1 to 492.4 ± 59.19 cells μL−1 after giving 1 g of N. sativa for 1 month (P = 0.061).23 Furthermore, CD4+ (helper) T lymphocytes has been reported to be stimulated by N. sativa oil in a murine cytomegalovirus (CMV) model using BALB/c mice.24\n\nIncrease in absolute lymphocyte count was also shown in our study, this is supported by a study conducted on diabetic hamsters, where oral administration of N. sativa oil improved lymphocyte count in streptozotocin (STZ)-induced diabetic hamsters.12 Additionally, oral administration of N. sativa oil significantly increased peripheral lymphocyte and monocyte counts in antigen-challenged rats.25\n\nOral administration of N. sativa oil (90 mg·kg−1 per day) for 30–60 days elevated neutrophils and lymphocytes back to normal levels in chloramphenicol treated Albino rats.26 N. sativa seeds extract induced a stimulatory effect on unactivated lymphocytes cell culture.13\n\nIn our study, the increase in the previously discussed cells has been lost when the dose was increased from 1 g N. sativa to 2 g, which is consistent with the results observed in previous two different studies. One study was on the effect of N. sativa on Helicobacter pylori eradication, where 2 g N. sativa shown to be more effective than 1 g N. sativa and 3 g N. sativa in comparison to triple therapy.17 The other study was conducted on the effect of N. sativa on the glycemic control in type 2 diabetic patients which showed that 2 g N. sativa was more effective than the 3-g dose in reducing fasting blood glucose, 2-hour post-load glucose and hemoglobin A1C.16\n\nThe current study showed a non-significant effect of N. sativa on IgG and IgM. Previous studies on the effect of N. sativa on humoral immunity showed inconsistent results. Sapmaz et al., reported that N. sativa oil produced a significant decrease in serum complement component 3 (C3), IgM and IgA levels with no significant change IgG level in rats, treated with formaldehyde inhalation.27 Moreover, a study of the effect of thymoquinone (TQ), which is the active ingredient of N. sativa, was conducted on rats and showed increase in total levels of immunoglobulins (IgG and IgM) and antibody hemagglutination in TQ-supplemented group.28 In contrast, oral administration of N. sativa extract and TQ in mice with allergic diarrhea did not produce a significant effect on total IgE level or ovalbumin-specific IgE.29 These discrepancies in the N. sativa effect on immunoglobulins may be better explained by the differences in species and/or condition of the species among various studies.\n\nThe current study did not show any significant effect of N. sativa on the level of IL-1β and IL-6. Similar findings were reported by Duncker et al., where no significant change was shown in IL-4, IL-5, IL-10 or IFN-γ secretion by mesenteric lymphocytes in mice treated with both oral extract of N. sativa, and intra-gastric TQ.29 Another study showed that N. sativa extracts had no effect on the secretion of IL-4 and IL-2 from lymphocytes, both in presence and absence of PWM.13 In contrast, significant increase in IL-10, but not TNFα was observed after 8 weeks of oral administration of N. sativa oil (1 g per patient per day) in rheumatoid arthritis patients.30 Furthermore, N. sativa extracts increased the secretion of IL-3 from PBMCs cultured in presence or absence of pooled allogeneic cells.13 Theses discrepancies in the effect of N. sativa on various cytokines could be explained by differences in the preparations (i.e., extracts, oil, etc.), doses of N. sativa, study designs (i.e., in vitro or in vivo), species used (other animals versus human), and difference in the studied cell/animal condition.\n\nOur results showed that the second group (0.5 g N. sativa) and the third group (1 g N. sativa) had a significant reduction in in the systolic BP for group 2 and the diastolic BP for group 3 (P < 0.05), respectively. This result is supported by a randomized double-blinded placebo-controlled trial conducted by Huseini and others in 2013 on healthy adult volunteers, where 5 mL of N. sativa oil was administered to 70 participants for an 8-week period. The study resulted in, average of 8.17% decrease in the systolic BP and 12.46% decrease in the diastolic BP.31\n\nIn the present investigation, the expression profiles of seven genes were evaluated before and after intervention with N. sativa. Box blotting of the data showed no significant variation in the expression profiles of these genes either amongst them or before and after treatment within the same gene (P-values of 0.15, 0.48, 0.15, 0.48, 0.37, 0.12, and 0.28 for IFN-γ, NF-κB, TNF-α, IL-1β, IL-13, IL-8, and IL-6, respectively).\n\nThe action of N. sativa and its ingredients on cytokines and inflammatory markers varies depending on the cell or animal tested and its condition. For example, herbal melanin, extracted from N. sativa, enhanced the production of m-RNA expression of TNF-α, and IL-6 in normal human peripheral blood mononuclear cells.32 While, it has been suggested that the treatment with TQ inhibited the production of TNF-α-induced IL-6 and IL-8 in rheumatoid arthritis synovial fibroblasts.33\n\nFurthermore, the therapeutic effects of alpha-hederin extracted from N. sativa has been extensively studied in terms of lung inflammation in rats. Treated animals showed elevated levels of IL-13 compared with the control group. These data indicated that alpha-hederin, like TQ, can indirectly intervene with IL-13 to reduce the inflammatory response.34\n\nOn the other hand, oral administration of N. sativa oil reduced the level of different cytokines including IL-13.35 Furthermore, TQ was found to inhibit NF-κB, TNF-α, IL-1β, and IL-6 induced by CLP.36\n\nThe dual action of N. sativa has been suggested by other research groups; it upregulates IL-8 in non-activated PBMC cells and downregulates it in PWM-activated PBMC cells.13 TQ also can ameliorate the toxic effects of arsenic via downregulating TNF-α and IFN-γ when administered three days before exposure to arsenic,37 and reduced levels of NF-κB and TNF-α.38\n\nNonetheless, this study shows no effect of a one-month N. sativa supplementation in young healthy human on the studied gene expressions and calls for further investigation on other immunity related genes and molecular mechanisms.\n\n\nConclusion\n\nOur results suggest that N. sativa has an immunopotentiation effect; the optimum dose seems to be 1 g for young healthy subjects. We recommend more future clinical trials to explore the use of N. sativa as an immune enhancer for various age groups in different health conditions.\n\n\n\n1. Participants’ withdrawal due to the coronavirus disease 2019 (COVID-19) crisis.\n\n2. Post-intervention results of certain interleukins and CD profile could not be obtained for many participants due to the COVID-19 crisis.\n\n\nData availability\n\nDryad: Underlying data for ‘Effect of Nigella sativa on general health and immune system in young healthy volunteers; a randomized, placebo-controlled, double-blinded clinical trial’. https://doi.org/10.5061/dryad.00000004b39\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).", "appendix": "References\n\nAhmad MF, Ahmad FA, Ashraf SA, et al.: An updated knowledge of Black seed (Nigella sativa Linn.): Review of phytochemical constituents and pharmacological properties. J Herb Med. 2021; 25(100404): 19.\n\nMaideen NMP: Prophetic Medicine-Nigella Sativa (Black cumin seeds) - Potential herb for COVID-19?. J. Pharm. 2020; 23(2): 62–70.\n\nMarshall JS, Warrington R, Watson W, et al.: An introduction to immunology and immunopathology. Allergy, Asthma Clin. Immunol. 2018; 14(2): 49. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSalem ML: Immunomodulatory and therapeutic properties of the Nigella sativa L. seed. Int. Immunopharmacol. 2005; 5(13-14): 1749–1770. PubMed Abstract \| Publisher Full Text\n\nKulyar MF-EA, Li R, Mehmood K, et al.: Potential influence of Nagella sativa (Black cumin) in reinforcing immune system: A hope to decelerate the COVID-19 pandemic. Phytomedicine. 2021; 85: 153277. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKhazdair MR, Gholamnezhad Z, Rezaee R, et al.: A qualitative and quantitative comparison of Crocus sativus and Nigella sativa immunomodulatory effects. Biomed. Pharmacother. 2021; 140: 111774. PubMed Abstract \| Publisher Full Text\n\nAbuharfeil NM, Salim M, Von Kleist S: Augmentation of natural killer cell activity in vivo against tumour cells by some wild plants from Jordan. Phytother. Res. 2001; 15(2): 109–113. PubMed Abstract \| Publisher Full Text\n\nSalem ML, Alenzi FQ, Attia WY: Thymoquinone, the active ingredient of Nigella sativa seeds, enhances survival and activity of antigen-specific CD8-positive T cells in vitro. Br. J. Biomed. Sci. 2011; 68(3): 131–137. PubMed Abstract \| Publisher Full Text\n\nKhan SH, Anjum MA, Parveen A, et al.: Effects of black cumin seed (Nigella sativa L.) on performance and immune system in newly evolved crossbred laying hens. Vet. Q. 2013; 33(1): 13–19. Publisher Full Text\n\nKumar P, Patra AK, Mandal GP, et al.: Effect of black cumin seeds on growth performance, nutrient utilization, immunity, gut health and nitrogen excretion in broiler chickens. J. Sci. Food Agric. 2017; 97(11): 3742–3751. PubMed Abstract \| Publisher Full Text\n\nDurrani FR, Chand N, Zaka K, et al.: Effect of different levels of feed added black seed (Nigella sativa L.) on the performance of broiler chicks. Pak. J. Biol. Sci. 2007; 10(22): 4164–4167. PubMed Abstract \| Publisher Full Text\n\nFararh KM, Atoji Y, Shimizu Y, et al.: Mechanisms of the hypoglycaemic and immunopotentiating effects of Nigella sativa L. oil in streptozotocin-induced diabetic hamsters. Res. Vet. Sci. 2004; 77(2): 123–129. PubMed Abstract \| Publisher Full Text\n\nHaq A, Lobo PI, Al-Tufail M, et al.: Immunomodulatory effect of Nigella sativa proteins fractionated by ion exchange chromatography. Int. J. Immunopharmacol. 1999; 21(4): 283–295. PubMed Abstract \| Publisher Full Text\n\nAl-Suhaimi EA: Hepatoprotective and immunological functions of Nigella sativa seed oil against hypervitaminosis A in adult male rats. Int. J. Vitam. Nutr. Res. 2012; 82(4): 288–297. PubMed Abstract \| Publisher Full Text\n\nBadr G, Alwasel S, Ebaid H, et al.: Perinatal supplementation with thymoquinone improves diabetic complications and T cell immune responses in rat offspring. Cell. Immunol. 2011; 267(2): 133–140. PubMed Abstract \| Publisher Full Text\n\nBamosa AO, Kaatabi H, Lebdaa FM, et al.: Effect of Nigella sativa seeds on the glycemic control of patients with type 2 diabetes mellitus. Indian J. Physiol. Pharmacol. 2010; 54(4): 344–354. PubMed Abstract\n\nSalem EM, Yar T, Bamosa AO, et al.: Comparative study of Nigella Sativa and triple therapy in eradication of Helicobacter Pylori in patients with non-ulcer dyspepsia. Saudi Journal of Gastroenterology: Official Journal of the Saudi Gastroenterology Association. 2010; 16(3): 207–214. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPlaeger SF: Clinical immunology and traditional herbal medicines. Clin. Diagn. Lab. Immunol. 2003; 10(3): 337–338. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMajdalawieh AF, Carr RI: In vitro investigation of the potential immunomodulatory and anti-cancer activities of black pepper (Piper nigrum) and cardamom (Elettaria cardamomum). J. Med. Food. 2010; 13(2): 371–381. PubMed Abstract \| Publisher Full Text\n\nAhmad A, Husain A, Mujeeb M, et al.: A review on therapeutic potential of Nigella sativa: A miracle herb. Asian Pac. J. Trop. Biomed. 2013; 3(5): 337–352. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKooti W, Hasanzadeh-Noohi Z, Sharafi-Ahvazi N, et al.: Phytochemistry, pharmacology, and therapeutic uses of black seed (Nigella sativa). Chin. J. Nat. Med. 2016; 14(10): 732–745. PubMed Abstract \| Publisher Full Text\n\nGholamnezhad Z, Havakhah S, Boskabady MH: Preclinical and clinical effects of Nigella sativa and its constituent, thymoquinone: A review. J. Ethnopharmacol. 2016; 190: 372–386. PubMed Abstract \| Publisher Full Text\n\nEl-Shanshory M, Hablas NM, Aboonq MS, et al.: Nigella sativa improves anemia, enhances immunity and relieves iron overload-induced oxidative stress as a novel promising treatment in children having beta-thalassemia major. J. Herb. Med. 2019; 16: 100245. Publisher Full Text\n\nSalem ML, Hossain MS: Protective effect of black seed oil from Nigella sativa against murine cytomegalovirus infection. Int. J. Immunopharmacol. 2000; 22(9): 729–740. PubMed Abstract \| Publisher Full Text\n\nIslam SN, Begum P, Ahsan T, et al.: Immunosuppressive and cytotoxic properties of Nigella sativa. Phytother. Res. 2004; 18(5): 395–398. Publisher Full Text\n\nEbaid H, Dkhil M, Zahran WS, Feki ME, et al., editors. Role of Nigella sativa in ameliorating chloramphenicol induced tissue damage in rats.2011.\n\nSapmaz HI, Sarsılmaz M, Gödekmerdan A, et al.: Effects of formaldehyde inhalation on humoral immunity and protective effect of Nigella sativa oil: An experimental study. Toxicol. Ind. Health. 2015; 32(9): 1564–1569. PubMed Abstract \| Publisher Full Text\n\nMohany M, El-Feki M, Refaat I, et al.: Thymoquinone ameliorates the immunological and histological changes induced by exposure to imidacloprid insecticide. J. Toxicol. Sci. 2012; 37(1): 1–11. PubMed Abstract \| Publisher Full Text\n\nDuncker SC, Philippe D, Martin-Paschoud C, et al.: Nigella sativa (black cumin) seed extract alleviates symptoms of allergic diarrhea in mice, involving opioid receptors. PLoS One. 2012; 7(6): 29. Publisher Full Text\n\nHadi V, Kheirouri S, Alizadeh M, et al.: Effects of Nigella sativa oil extract on inflammatory cytokine response and oxidative stress status in patients with rheumatoid arthritis: a randomized, double-blind, placebo-controlled clinical trial. Avicenna J Phytomed. 2016; 6(1): 34–43. PubMed Abstract \| Free Full Text\n\nFallah Huseini H, Amini M, Mohtashami R, et al.: Blood pressure lowering effect of Nigella sativa L. seed oil in healthy volunteers: a randomized, double-blind, placebo-controlled clinical trial. Phytother. Res. 2013; 27(12): 1849–1853. PubMed Abstract \| Publisher Full Text\n\nEl-Obeid A, Al-Harbi S, Al-Jomah N, et al.: Herbal melanin modulates tumor necrosis factor alpha (TNF-alpha), interleukin 6 (IL-6) and vascular endothelial growth factor (VEGF) production. Phytomedicine. 2006; 13(5): 324–333. PubMed Abstract \| Publisher Full Text\n\nUmar S, Hedaya O, Singh AK, et al.: Thymoquinone inhibits TNF-α-induced inflammation and cell adhesion in rheumatoid arthritis synovial fibroblasts by ASK1 regulation. Toxicol. Appl. Pharmacol. 2015; 287(3): 299–305. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFallahi M, Keyhanmanesh R, Khamaneh AM, et al.: Effect of Alpha-Hederin, the active constituent of Nigella sativa, on miRNA-126, IL-13 mRNA levels and inflammation of lungs in ovalbumin-sensitized male rats. Avicenna J. Phytomedicine. 2016; 6(1): 77–85. PubMed Abstract \| Free Full Text\n\nBalaha MF, Tanaka H, Yamashita H, et al.: Oral Nigella sativa oil ameliorates ovalbumin-induced bronchial asthma in mice. Int. Immunopharmacol. 2012; 14(2): 224–231. PubMed Abstract \| Publisher Full Text\n\nGuo L-P, Liu S-X, Yang Q, et al.: Effect of Thymoquinone on Acute Kidney Injury Induced by Sepsis in BALB/c Mice. Biomed. Res. Int. 2020; 2020: 1594726.\n\nFirdaus F, Zafeer MF, Ahmad M, et al.: Anxiolytic and anti-inflammatory role of thymoquinone in arsenic-induced hippocampal toxicity in Wistar rats. Heliyon. 2018; 4(6): e00650. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSeflek HN, Kalkan S, Cuce G, et al.: Effects of Nigella sativa oil on ovarian volume, oxidant systems, XIAP and NF-kB expression in an experimental model of diabetes. Biotech. Histochem. 2019; 94(5): 325–333. PubMed Abstract \| Publisher Full Text\n\nSalem AM, Bamosa AO, Alam MH, et al.: Underlying data for ‘Effect of Nigella sativa on general health and immune system in young healthy volunteers; a randomized, placebo-controlled, double-blinded clinical trial’.2021." }	[ { "id": "101149", "date": "18 Jan 2022", "name": "Mohammad Akram Randhawa", "expertise": [], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe study is well designed and conducted a placebo controlled double blind clinical trial that confirms the immunopotentiation effect of Nigella sativa powder, manifested as elevation of helper T cells, in young healthy volunteers. Surprisingly, 2 g N. sativa once daily for one month failed to show a similar increase in helper T-cells than with 1g. To make the article scientifically sound the authors should discuss the possible reasons for that.\nPerhaps, the higher doses trigger some inhibitory mechanisms. The evidence in favour is that the N. sativa & thymoquinone have been shown in the literature to possess beneficial effects in rheumatoid arthritis and asthma by suppressing some immune mechanisms. A careful search of the literature would reveal more such examples and if we calculate dose/concentration used in these cases per 70 kg (Adult human dose), perhaps, would be equal to or higher than 2g N. sativa powder per day.\nIt is desired that the authors find such examples (Human or animals studies) and describe the possible reasons for the absence of elevation in helper T cells by increasing the dose of N. sativa to 2g.\nSome other minor alterations have been suggested in the text (see attached and annotated report).\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "7740", "date": "26 Jan 2022", "name": "Abdullah Bamosa", "role": "Author Response", "response": "We would like to thank Professor Akram for his valuable comments. We have actually discussed the point of losing the significant increase in helper T cells when Nigella sativa dose was increased from 1 to 2g as we have encountered such decrease in the effect of Nigella sativa when increasing the dose in previous clinical trials on diabetic patients and H pylori eradication in dyspeptic patients. We agree that the most possible cause is the presence of some ingredients that possess negative effect at higher doses. Nigella sativa has several active ingredients which makes such explanation sound and logical. Our trial was on young healthy subjects expected to have higher response to lower doses than old or ill subjects. This fact supports our result of higher effect by 1 g than 2 g dose of Nigella sativa and based on that we concluded that optimum dose for enhancing immunity in this age group seems to be 1 g of Nigella sativa." } ] }, { "id": "208614", "date": "11 Oct 2023", "name": "Siddig Ibrahim Abdelwahab", "expertise": [ "Reviewer Expertise Pharmacology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nLimited sample size and gender imbalance: The study includes only 52 healthy subjects, with a significant gender imbalance (48 males and 4 females). This small sample size and gender disparity may limit the generalizability of the findings and raise concerns about the representativeness of the population. Lack of long-term follow-up: The trial's duration is only four weeks, which may not be sufficient to capture the full range of effects on immune parameters. Longer-term follow-up would provide a better understanding of the sustained effects and potential adverse reactions associated with N. sativa supplementation.\nIn the introduction, kindly correct according to following suggestions:\n\"Nigella sativa is one of the most commonly used herbal medicines worldwide nowadays.\" \"It has been used for more than 2000 years as a natural remedy for various illnesses.\" \"Research has documented its therapeutic potential as an antimicrobial, anti-inflammatory, antioxidant, antidiabetic, antihypertensive, antitumor, and immunomodulatory agent [1,2].\" \"The immune system consists of multiple interconnected networks of cells, proteins, and lymphoid organs that provide protection against millions of microbes and infections.\" \"The immune system includes innate immunity and adaptive immunity. Innate immunity provides immediate protection to the body, and its effect is similar in different individuals, while adaptive immunity takes more time to develop and is more specific and effective against invasive pathogens [3].\" \"The immunomodulatory effect of N. sativa has been extensively studied on both innate and adaptive immunity, as well as related messengers and mediators [4–6].\" \"Several in vivo studies have shown significant effects of N. sativa on immunity, autoimmune diseases, and toxicity.\" \"A study demonstrated that the aqueous extract of N. sativa resulted in a significant increase (62.3% ± 6.4%) in natural killer (NK) cytotoxic cell activity against YAC-1 cells after 1 week of oral administration in 10-week-old BALB/c female mice [7].\" \"Another in vitro study on the effect of thymoquinone (TQ) on immunity showed that TQ injected directly into cells at low concentrations (10, 2.5, or 0.62 μg mL−1) increased the survival of activated T-cells and the ability of CD8+ T-cells to generate IFN-γ, indicating the potential benefits of TQ against infectious diseases and immune enhancement [8].\" \"N. sativa supplementation at levels of 4% or 5% positively enhanced immunity against Newcastle disease virus in newly evolved crossbred laying hens [9].\" \"In another study on Newcastle virus vaccinated broilers, N. sativa supplementation at three doses (5, 10, and 20 g kg−1) for 42 days significantly increased antibodies against Newcastle virus on day 35 [10].\" \"A diet supplemented with 40 g kg−1 N. sativa fed to broiler chicks improved antibody production against both Newcastle virus and infectious bursal disease [11].\" \"The phagocytic index and rate were significantly higher in STZ-diabetic hamsters treated with N. sativa oil (NSO) at a dose of 400 mg·kg−1 for 4 weeks compared to untreated diabetic animals, as demonstrated by fluorescence microscopy [12].\" \"N. sativa extract has a stimulatory effect on cellular immunity in vitro by increasing the proliferative capacity of T lymphocytes and splenocytes, as well as the response to different mitogens of human peripheral blood mononuclear cells (PBMC) [13].\" \"NSO was shown to possess a protective role against vitamin A hypervitaminosis. Rats treated orally with 800 mg·kg−1 NSO showed higher serum levels of IgG and IgM than the control group or those receiving high doses of vitamin A [14].\" \"Gestational diabetes rats showed improvement in the immune status of their offspring after oral antenatal feeding with 20 mg kg−1 TQ, reversing the decreased levels of IL-2, T-cell reproduction, and improving both circulating and thymus-homing T-cell proliferation [15].\" \"The literature above demonstrates a very promising immunomodulating effect of N. sativa.\" \"However, the immunopotentiation effect of this remarkable plant has not been investigated in normal humans.\" \"Hence, this study was designed to evaluate the impact of different doses of N. sativa on the immune system in young, healthy humans.\"\n\nThe conclusion could be improved by providing more specific details, addressing the limitations of the study, suggesting specific research questions for future trials, considering the generalizability of the findings, and discussing potential risks or side effects associated with the use of N. sativa.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/10-1199
https://f1000research.com/articles/11-1013/v1	07 Sep 22	{ "type": "Research Article", "title": "Cortical auditory potentials and cognitive potentials in individuals with and without vestibular dysfunction", "authors": [ "Kaushlendra Kumar", "Krishnapriya S", "Anupriya Ebenezer", "Mohan Kumar Kalaiah", "Deviprasad D", "Kaushlendra Kumar", "Krishnapriya S", "Mohan Kumar Kalaiah", "Deviprasad D" ], "abstract": "Background: Among individuals with vestibular dysfunction, the loss of vestibular sensory information is found to alter cognitive abilities that coordinate spatial and non-spatial information. P300 is an event-related potential commonly used to assess cognitive processing. The aim of the present study was to compare the latency and amplitude of cortical auditory evoked potential and P300 between individuals with vestibular dysfunction and individuals with no vestibular dysfunction. Methods: Forty adults with a mean age of 40.5 ± 13.07 participated in the study. Group I included 20 adults diagnosed with vestibular dysfunction and group II included 20 age-matched adults with no vestibular dysfunction. The P300 was recorded from the electrode site Cz and Pz. It was elicited using pure-tones in odd-ball paradigm. The latency and amplitude of peaks P1, N1, P2, and N2 of the cortical auditory evoked potential and the P300 were measured. Results: Significant amplitude difference was observed in cortical potentials at Cz and Pz. The P300 was present only in 70% of individuals with vestibular dysfunction compared to 100% among individuals with no vestibular dysfunction. The mean amplitude of the P300 was slightly larger in group 1 compared to group 2 and the mean latency of the P300 was similar in both groups. However, the difference in amplitude of the P300 between groups was not statistically significant. Conclusions: Knowing the cognitive function of individuals with vestibular dysfunction enables planning vestibular rehabilitation therapy, which enhances the quality of life in these individuals by improving their vestibular and cognitive functions.", "keywords": [ "cognition", "vestibular dysfunction", "vertigo", "P300", "dizziness", "event related potentials", "cortical auditory evoked potentials", "VEMP" ], "content": "Introduction\n\nVestibular dysfunction is caused by pathologies in the peripheral and central vestibular system. The peripheral pathologies constitute 90% of cases with vertigo.1 It involves lesion in the end organs of the inner ear and/or the eighth cranial nerve. The central vestibular pathologies involve lesion in the cortical and sub-cortical pathways of the vestibular system. Vestibular dysfunction results in several adverse physical outcomes such as postural instability, abnormal gait and falls. Further, the majority of individuals with vestibular dysfunction are also found to have anxiety and depression. In addition, the loss of vestibular sensory information is shown to alter cognitive abilities related to the processing of spatial and non-spatial information.2\n\nSeveral studies have investigated the cognitive abilities of individuals with vestibular dysfunction. According to literature, parabrachial nucleus and the hippocampus are the anatomically two regions that account for the relation between the vestibular system and neural networks involved in cognitive and emotional processing.3 The different cognitive skills associated with vestibular function include attention, visuospatial orientation, executive function, memory, metacognition and self-control.4 Research on cognition assessment pertaining to vestibular function has mainly been based on spatial orientation, attention, memory and executive function.5 Smith (2017) reported that cognitive impairment is usually seen in any vestibular dysfunction such as either peripheral or central vestibular dysfunction.\n\nThe P300 is an event-related potential, elicited when the target stimuli in the odd-ball paradigm is identified by the participant. It serves as an index for the assessment of cognitive ability to assess cerebral information processing in the context of various neurological diseases.6 Several studies have documented abnormal P300 in individuals with cognitive dysfunctions such as autism spectrum disorder,7 attention deficit hyperactivity disorder,8 schizophrenia,9 migraine,10 etc. It is usually performed with minimum attention to the stimuli without secondary tasks. The P300 is used to evaluate age-related cognitive dysfunction, reflecting attention and memory processes and overlapping function in cognitive deficit.11 Different areas of the brain that provide the generation of P300 response include subcortical structures, auditory regions in the cortex and frontal lobe and various association areas neo cortex.12 The subcomponents for P300, P3a is generated from the frontal working memory which helps in early attention whereas P3b is attention-driven stimulus generated from the temporal and parietal structures.6 The P300 amplitude response mainly depends on stimulus probability, stimulus significance, task effort, motivation and attentiveness.13 P300 amplitude is directly related to the amount of attention paid to perform a particular task associated with superior memory performance.14 P300 latency reflects stimulus processing time in contrast to response processing time, which corresponds to stimulus evaluation time and is independent of the response section.15\n\nIn the case of individuals with vertigo, the literature has reported reduced behavioural cognitive abilities.16 A national health and nutrition examination survey done in the US revealed an association between vestibular and cognitive function in the adult population.5 There is a steady accumulation of evidence that the vestibular lesion leads to cognitive deficit. It is not essentially directly related to reflexive signs and perceptual disturbances associated with vestibular dysfunction.17 And there is minimal scientific evidence on cortical potentials in individuals with vestibular dysfunction. Hence, the current study aims to compare the findings of cortical potentials (P1, N1, P2, and N2) and cortical potentials (P300 and N4) between individuals with and without vestibular dysfunction. The objective was to investigate the relationship of cortical and cognitive potentials peak latency and peak amplitude in individuals with and without vestibular dysfunction and the correlation between DHI score with P300 findings among individuals with vestibular dysfunction.\n\n\nMethods\n\nA total of 40 adults aged between 20 and 60 years (mean = 40.5, SD = 13.1) participated in this study. Group I included 20 adults with vestibular dysfunction. All participants in group I had abnormal findings on oculomotor examination or vestibular evoked myogenic potentials (VEMP) assessment. The oculomotor examination was performed using videonystagmography (VNG), the subtests included were saccade test, tracking test, and optokinetic test. Group II included 20 age-matched adults with no vestibular dysfunction. Twenty individuals with vestibular dysfunction in group I included 12 individuals diagnosed with peripheral vestibular lesion and eight individuals with central vestibular lesion. All participants in the study had hearing sensitivity within normal limits. Individuals under medication for vertigo and individuals diagnosed as having an autoimmune disease, systemic illness or neurodegenerative disorders were excluded from the study. The study was approved by the institutional ethics committee of Kasturba Medical College, Mangalore (IECKMCMLR11-18/456) and written informed consent was obtained from all participants before they participated in the study.\n\nAll participants in group I completed a Dizziness Handicap Inventory (DHI) questionnaire.18 It assesses quality of life of participants on three domains: functional (nine questions), emotional (nine questions) and physical (seven questions). The participants were instructed to provide responses such as “Yes” when the symptom is present always, “Sometimes” when the symptoms is present sometimes, and “No” when it is absent. Item scores were summed, and the maximum score was 100 and the minimum score was 0. Answers were graded according to 0 for a “No” response, 2 for a “Sometimes” response and 4 for a “Yes” response.\n\nThe P300 was recorded using the IHS Smart EP version 3.92 evoked potential system (Intelligent Hearing Systems, USA). During the recording of the P300, participants were made to sit comfortably on a reclining chair in a sound-treated room. The electrode sites were cleaned using Nu-prep Skin Prep Gel (Weaver and Company, USA). Gold plated disc electrodes were placed on the electrode sites using conduction paste and it was secured using adhesive tape. Two non-inverting electrodes were placed on the scalp, one on the vertex (Cz) and the other on the parietal lobe (Pz). Inverting electrodes were placed on both ear mastoid (linked mastoid), and the ground electrode was placed on low forehead (Fpz). The electrode impedance was maintained below 5 kΩ for each electrode and the inter-electrode impedance was less than 2 kΩ. The P300 was elicited using pure-tones of 1000 Hz and 2000 Hz in an odd-ball paradigm. The 1000 Hz pure-tone served as standard stimuli and the 2000 Hz pure-tone served as deviant stimuli. The standard and target stimuli were presented at a ratio of 4:1. The pure-tones were presented to both ears of participants at 80 dB SPL using ER-3A insert earphones (Intelligent Hearing Systems, USA). A total of 300 stimuli were presented at a repetition rate of 1.1 stimuli/sec and the ongoing EEG was differentially recorded from the scalp. The EEG was amplified 50,000 times and filtered using a bandpass filter of 1 to 30 Hz. The duration of the analysis window was 600 msec with a pre-stimulus duration of 100 msec. The participants were instructed to count the target stimuli and report at the end of the recording.\n\nThe waveforms obtained from all participants for standard and target stimuli were grand averaged separately to identify various components or peaks of event related potentials. The averaged waveforms included peaks P1, N1, P2, and N2 between 50 msec and 250 msec. The broad positive peak after 250 msec from the stimulus onset in the waveform of target stimuli was referred to as P300. And the negativity peak following the P300 was considered as N4 (late negativity). The latency (in msec) and peak amplitude (in μV) of peaks P1, N1, P2, N2, P300 and N4 was measured at the electrode sites Cz and Pz. The peak amplitude was measured relative to the pre-stimulus baseline and the latency was measured from the stimulus onset. The dataset is published as underlying data in Mendeley Data.19\n\nStatistical analysis was conducted using IBM SPSS software version 26 (RRID:SCR_002865). Initially, descriptive analysis and the Shapiro-Wilk test were carried out to check the normality of the data. The latency and peak amplitude of peaks P1, N1, P2, N2, P300, and N4 were normally distributed. Thus, the independent t-test was administered to investigate if the mean latency and amplitude of peaks were significantly different between groups. Correlation analysis was carried out to investigate the relationship between the DHI score and latency and amplitude of P300 in group I.\n\n\nResults\n\nThe grand averaged waveforms for standard and deviant stimuli for both groups are shown in Figure 1. In the figure, it is evident that both pure-tones elicited obligatory P1-N1-P2 response for the stimulus onset among participants in both groups. Further, the waveform for target stimuli showed a large positive peak at a latency of 280 ms following the P1-N1-P2 response in both groups of participants, referred to as P300. The identification rate of peak responses varied across groups at Cz and Pz positions, which is depicted in Figures 2 and 3 respectively.\n\nTable 1 shows the mean latency of peaks P1, N1, P2, and N2 at electrode sites Cz and Pz for both groups. The mean latency of peaks P1, N1, P2, and N2 were similar between groups at the electrode sites Cz and Pz. To investigate if the mean latency of peaks were significantly different between groups, the independent t-test was carried out. It showed no significant difference for the latency of peaks P1 [t(37) = -1.083, p = 0.286)], N1 [t(37) = -1.008, p = 0.320)], P2 [t(36) = 1.007, p = 0.321)], and N2 [t(32) = 1.822, p = 0.078)] at the electrode site Cz. Similarly, at Pz the latency of peaks P1 [t(28) = -0.029, p = 0.977)], N1 [t(31) = -0.029, p = 0.977)], P2 [t(30) = -1.059, p = 0.298)], and N2 [t(27) = 0.56, p = 0.956)] were not significantly different between groups.\n\nTable 2 shows the mean amplitude of peaks P1, N1, P2, and N2 at electrode sites Cz and Pz for both groups. The mean amplitude of peaks was larger in individuals with no vestibular dysfunction (group II) at both Cz and Pz. To investigate if the mean amplitudes of peaks are significantly different between groups, independent t-test was carried out separately for Cz and Pz. It revealed no significant difference for the amplitude of peaks P1 [t(30) = -2.733, p = 0.10)], N1 [t(37) = -0.614, p = 0.543)], P2 [t(36) = -1.061, p = 0.296)] and N2 [t(32) = -0.268, p = 0.790)] at the electrode site Cz between groups. Similarly, the amplitude of peaks P1 [t(28) = -1.742, p = 0.093)], N1 [t(31) = -1.326, p = 0.194)], P2 [t(28) = -1.742, p = 0.093)] and N2 [t(27) = -0.318, p = 0.756)] at the electrode site Pz showed no significant difference between the groups. On peak-to-peak analysis between groups showed no significant difference in P1-N1 and N1-P2 peak to peak amplitude at Cz and Pz.\n\nTable 3 shows the mean latency and amplitude of peaks P300 and N4 at Cz and Pz for both groups. A noticeable difference was observed for the mean amplitude of N4 between groups at both the electrode sites. The mean amplitude of P300 was larger in group I compared to group II, this finding is controversial when compared to the grand average waveform of both groups shown in Figure 1. The mean amplitude of P300 was found to be largest in group 1 compared to group II. To investigate if the mean difference for latency and amplitude were significantly different between groups, an independent samples t-test was carried out. The results showed no significant difference in the latency of P300 [Cz: t(32) = -0.173, p = 0.866); Pz: t(30) = -0.357, p = 0.724)] and amplitude of P300 [Cz: t(32) = 0.218, p = 0.829); Pz: t(29) = 0.797, p = 0.432)] at both Cz and Pz. Whereas no statistical significant difference was found for the N4 latency [Cz: t(16) = 0.415, p = 0.684); Pz: t(16) = 0.786, p = 0.443)] and significant difference was observed in N4 amplitude [Cz: t(14) = -2.178, p = 0.047); Pz: t(15) = -2.107, p = 0.052)] at both positions.\n\nTo investigate the relationship between the latency of P300 and DHI score, Pearson’s correlation analysis was carried out. The results showed a weak negative correlation between the latency of P300 and the DHI score; however, the correlation was not significant at both electrode sites [Cz: r = -0.201, p = 0.490; Pz: r = -0.401, p = 0.196]. Further, no correlation was found between the amplitude of the P300 and DHI score at both electrode sites [Cz: r = -0.167, p = 0.569; Pz: r = 0.087, p = 0.787].\n\nThe mean latency of late latency response and P300 of individuals diagnosed with peripheral vestibular lesion and central vestibular lesion are depicted in Tables 4 and 5 respectively.\n\n\nDiscussion\n\nThe present study compared the latency and amplitude of cortical (P1, N1, P2 and N2) and cognitive potentials (P300) among individuals with and without vestibular dysfunction. The cortical potential includes auditory late latency responses P1, N1, P2, and N2. The cortical and cognitive potentials mainly involve the primary and non-primary auditory cortex of the temporal lobe, especially the primary pathway being more auditory sensitive. As per the literature, the generation of middle latency response is an interplay of primary and non-primary areas in the auditory thalamo-cortical pathway.20 Whereas late latency response is from non-primary cortical areas which measure the integrity of the auditory system beyond the brainstem.21 The P300 is a positive peak that comes after approximately 300 ms has extended importance in the study of cognition and executive functions. The waveform is obtained with parieto-central distribution and shows typical P300 topography for processing oddball stimuli via any sensory modality.22 As per a review, it is clear that cognitive impairment is observed in individuals with vestibular dysfunction, with respect to attention, spatial orientation, executive function and memory.5\n\nThe results of the present study showed no significant difference in the latency and amplitude of peaks P1, N1, P2, and N2 of the cortical auditory potentials. These findings are not in agreement with results of earlier investigation.23 However, findings of the present study could be explained based on the hearing sensitivity of participants in both groups and the characteristics of the P1-N1-P2 response. The P1-N1-P2 response is elicited for the onset of stimuli, therefore, the characteristics of the response are dependent on the onset of the stimuli. Further, the participants in both groups had hearing sensitivity within normal limits. Thus, the latency and amplitude of the peaks are expected to be similar in both groups.\n\nThe P300 was found to be absent in a greater number of individuals with vestibular dysfunction compared to the control group. It was absent in 30–40% of the individuals with vestibular dysfunction; this finding is consistent with results of the previous study.24 Further, when the P300 was present, the mean latency and amplitude of the P300 in both groups were similar. In contrast to the findings of the present study, earlier investigations have reported prolonged latency for P300 in individuals with vestibular dysfunction compared to the control group.19,20 The contrasting findings observed in the present study and earlier investigations could be differences in the site of vestibular lesion across studies. Studies in the literature have included individuals with peripheral vestibular lesions where the site of the lesion was localized to the lateral semicircular canal.23,24 The majority of the participants were reported to have unilateral caloric hypofunction. The findings of the above investigations showed prolongation of P300 latency in individuals with unilateral peripheral vestibular lesions (lateral semicircular canal) compared to the control group. In contrast, participants in the present study had peripheral vestibular lesion with abnormal findings on cVEMP and oVEMP indicating a lesion in the saccule and utricle. Therefore, the contrasting findings observed in the present study could be a consequence of differences in the site of the lesion. In addition, contrasting findings observed in the present study could be due to the degree of severity of dizziness. The majority of the participants with peripheral vestibular lesion in the present study were found to have mild severity/handicap based on DHI scores. The severity of dizziness might have an influence on the latency and amplitude of P300.\n\nStudies on cognitive function assessment using a cognitive failure questionnaire in individuals with vestibular dysfunction revealed that cognitive dysfunction is prevalent in individuals with central and peripheral vestibular pathologies.25 The literature also showed a positive correlation between cognitive dysfunction and dizziness severity in terms of a self-rated questionnaire. DHI helps in evaluating the severity of dizziness based on its impact physically, functionally and emotionally with limited profile on cognition.26 In the current study, the correlation between DHI scores and P300 showed no significant correlation. The lack of correlation between the two measures could be due to the different areas of assessment. DHI is a measure of self-help obtained primarily based on daily activities, whereas the P300 is an electrophysiological measure that assesses cognitive functioning.12 Because of this direct correlation between DHI and P300 was found to be inconclusive in the present study. Similarly, no significant correlation was observed between P300 and vertigo symptoms, whereas another study stated that severity of vestibular symptoms seems to correlate with P300 responses.27 In support of the current study, a randomized controlled trial showed cognitive behaviour therapy influenced patients with chronic subjective dizziness with a significant reduction in DHI and no changes in psychological outcome measures.28 Similarly, other literature has reported that the functional and physical parameter of DHI showed a negative correlation, and the emotional parameter showed a weak significant positive correlation in 369 participants evaluated for functional tests such as electronystagmography, rotational testing, and platform posturography.29 The findings of various studies by several investigators have emphasized the role of the vestibular system’s role on cognition, such as perceptual/visuospatial ability, memory, attention, and executive function. Knowing the cognitive function of individuals with vestibular dysfunction facilitates the setting of vestibular rehabilitation therapy goals. Evidence reveals that in patients with intractable dizziness following vestibular rehabilitation there is a significant improvement in vestibular function and cognitive function including attention, visuospatial ability and executive function with coincidental improvement in DHI.30 The findings of this study are circumscribed to oddball auditory tasks only, which might be a limitation.\n\n\nConclusions\n\nThe present study, which includes a review of the literature, divulges that cortical and cognitive potentials might help us in assessing and understanding the cognitive function in individuals with vestibular dysfunction; however, a lot of research is required in this field. For individuals with vestibular dysfunction, cognitive assessment is necessary to understand the dizziness impact on daily activities. Knowing the cognitive function of individuals with vestibular dysfunction enables the planning of vestibular rehabilitation therapy, which enhances the quality of life in these individuals by improving the vestibular and cognitive function.\n\n\nData availability\n\nMendeley Data: Underlying data for ‘Cortical auditory potentials and cognitive potentials in individuals with and without vestibular dysfunction’ https://www.doi.org/10.17632/hn6z8x5vkk.119\n\nThis project contains the following underlying data:\n\n• Data file 1. Description.txt\n\n• Data file 2. Event related potentials in individuals with vestibular dysfunction.xlsx\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nConsent\n\nWritten informed consent for publication of the participants’ details was obtained from the participants.", "appendix": "References\n\nNeuhauser HK: Epidemiology of vertigo. Curr. Opin. Neurol. 2007 Feb [cited 2022 Jun 1]; 20(1): 40–46. Publisher Full Text Reference Source\n\nSmith PF: Vestibular – Hippocampal Interactions.1997; 471: 465–471.\n\nBrandt T, Schautzer F, Hamilton DA, et al.: Vestibular loss causes hippocampal atrophy and impaired spatial memory in humansVestibular loss causes hippocampal atrophy and impaired spatial memory in humans. Brain. 2005; 128(11): 2732–2741. PubMed Abstract \| Publisher Full Text\n\nGurvich C, Maller JJ, Lithgow B, et al.: Vestibular insights into cognition and psychiatry. Brain Res. 2013; 1537: 244–259. PubMed Abstract \| Publisher Full Text\n\nSemenov YR, Bigelow RT, Xue QL, et al.: Association between Vestibular and Cognitive Function in U.S. Adults: Data from the National Health and Nutrition Examination Survey. J. Gerontol. - Ser. A Biol. Sci. Med. Sci. 2016; 71(2): 243–250. PubMed Abstract \| Publisher Full Text\n\nPolich J: Updating P300: An integrative theory of P3a and P3b.2007; 118: 2128–2148.\n\nSheela P, Puthankattil SD: Event related potential analysis techniques for autism spectrum disorders: A review. Int. J. Dev. Neurosci. 2018 Aug 1 [cited 2022 Apr 6]; 68: 72–82. PubMed Abstract \| Publisher Full Text\n\nArjona Valladares A, Gómez CM, Rodríguez-Martínez EI, et al.: Attention-deficit/hyperactivity disorder in children and adolescents: An event-related potential study of working memory. Eur. J. Neurosci. 2020 Nov 1; 52(10): 4356–4369. PubMed Abstract \| Publisher Full Text\n\nPritchard WS: Cognitive event-related potential correlates of schizophrenia. Psychol. Bull. 1986 Jul [cited 2022 Apr 6]; 100(1): 43–66. Publisher Full Text Reference Source\n\nTitlic M, Ivica N, Pintaric I, et al.: The Event-related Potential P300 in Patients with Migraine.2015; 23(13): 339–342.\n\nPolich J, Kokb A: Cognitive and biological determinants of P300: an integrative review.1995; 41: 103–146.\n\nPicton TE: The P300 Wave of the Human Event-Related Potential. J. Clin. Neurophysiol. 1992; 9(4): 456–479. Publisher Full Text\n\nJohnson R: A Triarchic Model of P300 Amplitude. Psychophysiology. 1986; 23: 367–384. PubMed Abstract \| Publisher Full Text\n\nJohnson R: On the neural generators of the P300 component of the event-related potential. Psychophysiology. 1993; 30(1): 90–97. PubMed Abstract \| Publisher Full Text\n\nKutas M, Mccarthy G, Donchin E, et al.: Augmenting Mental Chronometry: The P300 as a Measure of Stimulus Evaluation Time. Science (80-). 2015; 197(4305): 792–795. Publisher Full Text\n\nBalci B, Naziye Senyuva GA: Definition of Balance and Cognition Related To Disability Levels in Vestibular Migraine Patients. Arch. Neuropsychiatry. 2018; 9–14.\n\nSmith PF, Zheng Y, Horii A, et al.: Does vestibular damage cause cognitive dysfunction in humans? J. Vestib. Res. Equilib. Orientat. 2005; 15(1): 1–9. PubMed Abstract \| Publisher Full Text\n\nJacobson N: The development of the Dizziness Handicap Inventory. Arch. Otolaryngol. Head Neck Surg. 1990; 116(4): 424–427. Publisher Full Text\n\nEbenezer A, Kalaiah MK, Kumar K: Event Related Potentials in Individuals with Vestibular Dysfunction - Mendeley Data.2022 [cited 2022 Aug 3].Reference Source\n\nKraus N, McGee T: The middle latency response generating system. Electroencephalogr. Clin. Neurophysiol. Suppl. 1995 [cited 2022 Mar 11]; 44: 93–101. PubMed Abstract\n\nKonadath S, Manjula P: Auditory brainstem response and late latency response in individuals with tinnitus having normal hearing. Intractable Rare Dis. Res. 2016 [cited 2022 Mar 13]; 5(4): 262. /pmc/articles/PMC5116861/–268. PubMed Abstract \| Publisher Full Text\n\nPolich J, Herbst KL: P300 as a clinical assay: Rationale, evaluation, and findings. Int. J. Psychophysiol. 2000; 38(1): 3–19. PubMed Abstract \| Publisher Full Text\n\nFilha VAVdS, Bruckmann M, Garcia MV: Short- and long-latency auditory evoked potentials in individuals with vestibular dysfunction. Codas. 2018; 30(2): 1–10.\n\nEl-Gharib A, Nada E, Lasheen R: Auditory P300 and mismatch negativity (MMN) in patients with peripheral vestibular hypofunction. Hear Balanc. Commun. 2018; 16(1): 36–40. Publisher Full Text\n\nRizk HG, Sharon JD, Lee JA, et al.: Cross-Sectional Analysis of Cognitive Dysfunction in Patients with Vestibular Disorders. Ear Hear. 2020; 41: 1020–1027. PubMed Abstract \| Publisher Full Text\n\nDonaldson LB, Yan F, Liu YF, et al.: Does cognitive dysfunction correlate with dizziness severity in patients with vestibular migraine? Am. J. Otolaryngol. - Head Neck Med. Surg. 2021; 42(6): 103124. PubMed Abstract \| Publisher Full Text\n\nAgarwal A, Barua N, Walia BS, et al.: Title p-300 wave in patients with post concussion vertigo: A prospective study of 15 cases. Indian J. Otolaryngol. Head Neck Surg. 1996 [cited 2019 Aug 8]; 48(2): 121–124. Publisher Full Text\n\nEdelman S, Mahoney AEJ, Cremer PD: Cognitive behavior therapy for chronic subjective dizziness: a randomized, controlled trial. Am. J. Otolaryngol. 2012 Jul 1 [cited 2019 Aug 8]; 33(4): 395–401. PubMed Abstract \| Publisher Full Text Reference Source\n\nJacobson GP, Newman CW, Hunter L, et al.: Balance function test correlates of the Dizziness Handicap Inventory. J. Am. Acad. Audiol. 1991; 2(4): 253–260. PubMed Abstract\n\nGoto F, Sugaya N, Arai M, et al.: Psychiatric disorders in patients with intractable dizziness in the department of otolaryngology. Acta Otolaryngol. 2018; 138(7): 646–647. PubMed Abstract \| Publisher Full Text" }	[ { "id": "149811", "date": "13 Sep 2022", "name": "Sangamanatha Ankmnal Veeranna", "expertise": [ "Reviewer Expertise Auditory Evoked Potentials", "Auditory Processing Disorders", "Psychoacoustics" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nAbstract: The whole abstract should be rewritten so that the readers can follow. It is not clear whether P300 should be used to understand cognitive deficits in individuals with vestibular dysfunction.\nThe Results section should be rewritten:\nIn the manuscript, the authors have mentioned that there are no statistical differences in the amplitude between Cz and Pz but in the abstract authors have mentioned that there is a significant difference.\n\nIf there are no statistically significant differences in the P300 amplitude, why are the authors discussing this?\n\nIntroduction:\nI would not use “etc.” in a manuscript.\n\nThere are sections where authors should provide references. For example, “Further, the majority of individuals with vestibular dysfunction are also found to have anxiety and depression.”\n\nMethods: A lot of key information is missing in the Methods section.\nFor Group I, did the authors assess oculomotor function and VEMP or both?\n\nThe age range of Group I and Group II? How many male and female participants were included?\n\nAuthors have vaguely mentioned that participants in Group 2 had peripheral and central vestibular lesions. Authors should provide more details on peripheral vs central lesions.\n\nThe authors reported that all participants' hearing thresholds were within normal limits. Do the authors mean < 15 dB HL or 20 dB HL? Some participants were 60 years old, there could be some age-related hearing loss in these participants. The authors should provide hearing thresholds for all the participants (control and experimental groups).\n\nWhat was the artifact rejection?\n\nThe authors stated that “The participants were instructed to count the target stimuli and report at the end of the recording”, did participants correctly count the target stimuli? The authors should add this information.\n\nWas all the testing carried out in one session or multiple sessions?\n\nData analysis:\nWho marked the peaks?\n\nThe authors carried out the statistical analysis using multiple t-tests. But the authors did not include age and hearing thresholds in their analysis. The authors should use different statistical analyses.\n\nDiscussion: I was not happy with the discussion section; I feel it is all over the place.\nParagraph 1: Most of this content should be in the introduction not in the discussion.\n\nParagraph 2: The authors did not report hearing thresholds, or they did not include hearing thresholds in the statistical analysis.\n\nParagraph 3: The authors reported that 30-40% of individuals with vestibular dysfunction demonstrated an absence of P300, is this statistically significant? The Chi-square test may help in determining whether it is statistically significant or not.\n\nThe authors are talking about cVEMP and oVEMP in the discussion, but they did not talk about these measures in the method.\n\nThe authors are also discussing the severity of dizziness, which was not mentioned in the Methods section.\n\nLimitations: Authors should list limitations.\nConclusions: The authors should write the conclusion based on findings from this study. I do not see that conclusion here. This section should be rewritten.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [ { "c_id": "8990", "date": "16 Nov 2022", "name": "Anupriya Ebenezer", "role": "Author Response", "response": "Abstract: Thank you for the suggestions. As per the suggestion, the abstract was rewritten. In the manuscript, the authors have mentioned that there are no statistical differences in the amplitude between Cz and Pz but in the abstract authors have mentioned that there is a significant difference. Appropriate changes were made. If there are no statistically significant differences in the P300 amplitude, why are the authors discussing this? The abstract was rewritten with appropriate changes. Introduction: I would not use “etc.” in a manuscript. Appropriate changes incorporated. There are sections where authors should provide references. For example, “Further, the majority of individuals with vestibular dysfunction are also found to have anxiety and depression.” Respective citation added, and appropriate changes were made in the Reference section. Methods: A lot of key information is missing in the Methods section. For Group I, did the authors assess oculomotor function and VEMP or both? Yes, both the oculomotor function test (VNG) and VEMP (cVEMP and oVEMP) were performed to rule out vestibular dysfunction in Group I. The age range of Group I and Group II? How many male and female participants were included? Details are added in the Method section first paragraph. Authors have vaguely mentioned that participants in Group 2 had peripheral and central vestibular lesions. Authors should provide more details on peripheral vs central lesions. Group II included individuals with no vestibular dysfunction. The authors reported that all participants' hearing thresholds were within normal limits. Do the authors mean < 15 dB HL or 20 dB HL? Some participants were 60 years old, there could be some age-related hearing loss in these participants. The authors should provide hearing thresholds for all the participants (control and experimental groups). The mean and standard deviation of the hearing threshold of the participants are added in the Method section under the heading 'Participants'. What was the artifact rejection? Sweeps with amplitude greater than ±50µV were rejected from averaging. The same has been added to the manuscript. The authors stated that “The participants were instructed to count the target stimuli and report at the end of the recording”, did participants correctly count the target stimuli? The authors should add this information. Yes, all subjects had a practice trial on the task before ERPs were recorded. The same has been added to the manuscript under the Method section. Was all the testing carried out in one session or multiple sessions? Yes, all the testing was carried out in one session. Data analysis: Who marked the peaks? Peaks were identified separately by 3 investigators. Peaks were considered to be present when the markings of 2 investigators were in agreement The authors carried out the statistical analysis using multiple t-tests. But the authors did not include age and hearing thresholds in their analysis. The authors should use different statistical analyses. Descriptive statistical analysis (mean and SD) was done for both age and hearing thresholds. Descriptive statistics of age, as well as the hearing threshold, have been added to the Method section. Discussion: I was not happy with the discussion section; I feel it is all over the place. Thank you for the suggestion. The Discussion section is modified to improve the flow of information. Paragraph 1: Most of this content should be in the introduction not in the discussion. Appropriate changes incorporated. Paragraph 2: The authors did not report hearing thresholds, or they did not include hearing thresholds in the statistical analysis. Statistical analysis of the hearing threshold is added to the Method section first paragraph. Paragraph 3: The authors reported that 30-40% of individuals with vestibular dysfunction demonstrated an absence of P300, is this statistically significant? The Chi-square test may help in determining whether it is statistically significant or not. Thank you for the suggestion. We performed a chi-square test and a significant difference was observed (X2(1) = 8.53, p= 0.003) with the odd ratio 15.83. The same has been added to the manuscript under the Result section. The authors are talking about cVEMP and oVEMP in the discussion, but they did not talk about these measures in the method. cVEMP and oVEMP were administered for the Group I participant selection. The same is mentioned in the Method. The authors are also discussing the severity of dizziness, which was not mentioned in the Methods section. By severity, we were speaking about the dizziness handicap. The word 'severity' is replaced with the word 'handicap'. Appropriate changes were incorporated for better understanding. Limitations: Authors should list limitations. Some of the limitations were added to the Discussion in the last paragraph. Conclusions: The authors should write the conclusion based on findings from this study. I do not see that conclusion here. This section should be rewritten. The conclusion is rewritten with appropriate changes." } ] }, { "id": "149808", "date": "18 Oct 2022", "name": "Prawin Kumar", "expertise": [ "Reviewer Expertise Diagnostic Audiology including Electrophysiology" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nAuthors formulated the research very well except for a few observations.\nIn Abstract, authors can add N4 potential.\n\nIn last paragraph of Introduction section, it should be mentioned as Cognitive potentials (P300 and N4) instead of cortical potential.\n\nIn Method, the rationale for considering only two electrode sites i.e. Cz and Pz should be mentioned.\n\nThe two groups are considered based on abnormal finding of the VNG & VEMP. No gold standard tests were used to differentiate the peripheral and central lesions. That could also be one of the reasons for minimal differences in the performance between two groups for ALLR and P300.\n\nOverall, preliminary finding of the present study throws light for researchers to explore more comprehensive behavioral cognitive assessment along with electrophysiological measures such as multi-channel ERP recording.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "8989", "date": "16 Nov 2022", "name": "Anupriya Ebenezer", "role": "Author Response", "response": "Appropriate changes incorporated. Appropriate changes incorporated. The clinical AEP instrument which we used for the study has the facility to perform only 2 channel recording. So, EEG has been recorded for 2 electrode sites. Literature studies have shown greater amplitude for P300 at Cz and Pz electrode sites. Therefore, Cz and Pz electrode sites were selected to record P300. Group I was selected based on the vestibular test findings. Group II was normal healthy individuals with no complaint of vertigo. The vestibular assessment test battery included subjective vestibular assessment, VEMP, and VNG. Thank you for the encouraging words." } ] } ]	1	https://f1000research.com/articles/11-1013
https://f1000research.com/articles/12-818/v1	12 Jul 23	{ "type": "Research Article", "title": "Game on: Can gamification enhance productivity?", "authors": [ "Habeeb Ur Rahiman", "Rashmi Kodikal", "Sucharitha Suresh", "Rashmi Kodikal", "Sucharitha Suresh" ], "abstract": "Background: Research suggests that gamification can increase work engagement by providing employees with a sense of autonomy, competence, and relatedness, and by creating a fun and engaging work environment. Gamification is designed to increase consumer and employee engagement and see that they holistically collaborate to achieve a shared vision. The concept of gamification is as old as learning itself, just that the use of the terminology “Gamification” is of a recent origin. Methods: This article focuses on the impact of gamification in various organizations and simultaneously sees its relationship with job engagement and productivity. A primary investigation was done to determine the nexus between the various variables and data collection from 400 respondents working in various fraternities of the economy from both public and private domains from countries in the Gulf region. The structural equation model and SPSS has been inferred to analyse the results. Results: The study results show that variable such as perceived adoption and usefulness in the gamified system is significantly associated with job engagement. Similarly, employee’s recognition and perceived motivation have a positive impact on productivity. The study identified job engagement mediating factor to enhance organisational productivity in a gamified system. Conclusion: The effectiveness of gamification in enhancing work engagement may depend on factors such as the design of the gamification system, the preferences and motivations of individual employees, and the organizational culture and goals. The findings have significant implications for insight into how employees in the service sector are aware of the gamified working environment and react to the system through work engagement and productivity.", "keywords": [ "Gamification", "motivation", "adoption", "usefulness", "productivity", "Job Engagement" ], "content": "Introduction\n\nThe precipitously changing workforce dynamics paired with digitization and switching work preferences compel the constant evolvement of practices and procedures (Ashley et al., 2022). In an era of high-speed digitalized technology and a multi-generational capacity team, the scenario has resulted in a struggle for the smarted of the people (Palmquist, 2020). The critical need for firms to entice talent becomes a fascinating case to embrace innovative operational and recruitment methods, such as gamification (Morschheuser et al., 2018). Gamification can increase employee engagement by incorporating game-like elements into non-game contexts such as work, to create a more interactive and engaging experience (Kapp, 2012). By using features such as points, badges, and leader boards, gamification can provide employees with a sense of accomplishment, feedback, and recognition for their work, which can increase their motivation and engagement. The concept of gamification is as old as learning itself, just that the use of the terminology “Gamification” is of a recent origin (Prince, 2013). To make learning and work interesting, use of non-computer-aided techniques has been seen for centuries. However, in the early 1970s and 80s with video games becoming popular, the gamification concept got coined a new label. The term gamification is the most trending and widely applied concept in a non-game context (Buckley et al., 2019). Globally, approximately 40% of the fortune 1000 companies successfully applied gamification in the workplace (Keepers et al., 2022). This progress makes gamification one of the highly important innovative developments in the administration of organizations (Grünewald et al., 2019; Thompson & Irvine, 2014). Technical progress has led to numerous alterations in areas of human resources management (HRM) to connect with the needs of globalization (Woodcock & Johnson, 2018). The acceleration of artificial intelligence, the internet of thinking and machine learning, And other advancements in the field of technology. have bought revolutionary changes in industrial routine operations (Dessureault, 2019; Jia et al., 2017). This digital enhancement affects various stages of organizations, and it needs to adapt to advancements as new methods of working. Human resource (HR) and people management in organizations demand this technology adoption in various modes which has influenced HR operations (Medeiros et al., 2015). In recent years, several methods have been created to encourage and help the workforce accomplish desired behaviors. Since a human being has an instinctive pleasure of playing, progress in this space that is gradually obtaining consideration is gamification (Warmelink et al., 2020).\n\nGamification is applied in several organizational disciplines from manufacturing, operations, and recruitment, staff development activities (Deif, 2019). Artificial intelligence, machine learning, and the internet of thinking are mostly adopted by organizational routine operations. There is a strong correlation between artificial intelligence (AI), machine learning (ML), and internet of things (IoT) with gamification with various tools which helps to enhance better performance (Bahadoran et al., 2023; Jacob et al., 2022). Currently, companies are developing frameworks and tools considering AI technology as a form of the game to identify candidate skills and also utilize gamification to avoid recruitment bias. Human resource specialists had reacted to global digitalized transformation all through advanced platform methods known as gamification (Ramallo-González et al., 2022). Gamification can be started by shifting organization strategy and facilitating the organization in several areas such as operation, recruiting training, and development. Notable organizations like European Central Bank (Donovan, 2011), SAP (Kotsis et al., 2021), Samsung (West & Lockley, 2016), and Apple (Favorskaya et al., 2015), have applied gamification in their organizational activities and practices. In the human resource system, gamification comprises incorporating gaming elements and motivating techniques for example through leader boards, and points, into HR practices to design routine tasks and procedures that are identified by operators as game-like practices (Scurati et al., 2020). In addition to companies promoting the enactment of gamification for their workplaces to improve efficiency, productivity, and enthusiasm, some companies also use it for pre-emptive purposes, such as a vendor or competitive analysis) (Schlömmer et al., 2021). The outcomes of gamification in companies eventually depend on whether the workforce is encouraged to apply it, and whether gamification improves their optimistic opinions related to their employment (Metwally et al., 2021). If the workforce feels more pride, gratification, and engagement in their task as a cause of gamification, that will reflect in organizational productivity (Guven & Sakamoto, 2016; Kaur et al., 2021). With the growth of the E-Human resource management system, the digital revolution has transformed the traditional business process by applying game-thinking in the organizational work process (Behl, Sampat, et al., 2021). To meet organizational goals, one needs to have strong gamification functions that will be utilized during the employment practice and create an advantage for the company and workforce (Zhang et al., 2021).\n\nOur research instrument was used to understand how industrialists and professionals in the Gulf cooperation council (GCC) perceive the concept and practice of gamification. Therefore, our research was taken up to examine the impact of gamification in various organizations and simultaneously see its relationship with job engagement and productivity. The primary objective of this research is to identify the role of perceived usefulness, motivation, adoption, and recognition in the organization while adopting gamified tasks in an HR system.\n\nIn the first section of the manuscript, the paper reviewed flow theory and gamification, and the components and mechanisms of gamification that exist in this domain. The methodology has been explained in the second section, followed by results, and a discussion was presented. The paper ends with a conclusion and suitable theoretical implications and direction for limitations and future research.\n\nNumerous studies have been conducted by researchers globally have discussed that the digital revolution appears to have a significant relationship between efficiency, motivation, work engagement, and productivity. From the review of previously published literature, we can infer the impact gamification will have on organizational operations in various areas.\n\nGamification is a mechanism or element of the game to make available affordance for game events in non-game environments (Brandstätter & Sommerer, 2016; Müller et al., 2016). It is difficult to claim when the idea of gamification surfaced. Several individuals opined that gamification drew its origins in the 20th century when the boy scouts organization was established (Gatautis et al., 2021). Since then, gamification has remained trail blazing in web-related fields and elsewhere. Organizations use gamification to maintain workforce engagement (Prasad et al., 2019), incentivize users (Naeem et al., 2017), recruit, lead, and enhance productivity (Silic & Back, 2017). Flow theory suggests that individuals experience a state of optimal experience and engagement when they are fully immersed and focused on an activity that is challenging but within their skill level. Gamification can be designed to enhance the experience of flow by providing users with clear goals, immediate feedback, and a sense of progress and accomplishment (Hammedi et al., 2021; Murray, 2018). Gamification has in recent times been offered as a favorable prospect to enhance human resource management (HRM) systems and instruments. The purpose of game design components, such as badges (Lee et al., 2016), leader boards, and points have become a recognized exercise all over society (Donnermann et al., 2021). Integration of corporate environmental responsibility and pro-environmental action into the gamified system or game design is an essential element of the gamification approach (Morganti et al., 2017). To enhance environmental performance, it is important to encourage pro-environmental and pro-social behaviour (Marculescu et al., 2020). Therefore, incorporating pro-social aspects in the gamification process by focusing on the benefits of the elements of the gamified system can increase the likelihood of user engagement in pro-environmental and social aspects. Over the years, an immense quantity of research has been conducted to create structures and categorizations for gamification and game model components (Patrício et al., 2018; Zhang et al., 2021). Researchers have discovered methods, layouts, design, styles, and, most recently, the effects of gamified organizations (Beard-Gunter et al., 2019; Patrício et al., 2018) in order to popularise the concept of gamification in the field of HRM. Of recently, the research on gamification focuses on involving methods to examine potential consequences of gamifying the HRM system and tools (Makanawala et al., 2013).\n\nThe engaged workforce tends to be energetic, passionate, and determined to stage their roles. New methods can enable such engagement by establishing resource exchanges, communications, and mutual well-being (Silic et al., 2020). However, such engagement necessitates careful supervision (Hammedi et al., 2021). Globally notable companies have implemented and experienced designing fun events as a method for boosting workforce engagement, with the perception that fun can improve employee’s work satisfaction and commitment and eventually enhance their well-being and productivity (Gatautis et al., 2021).\n\nSeveral studies believe that an effective work environment is the major indicator of workforce well-being, However, such a reflexive form of well-being can be accompanied by more effective methods, for example, job engagement (Gerdenitsch et al., 2020). Well-engaged employees have optimistic appraisals of their employment condition, and beyond mere gratification, they are encouraged to disburse strength to achieve a task; they also recognize their inspiration (Rapp, 2020). Therefore, both work environment and job engagement appear vital for the well-being of the workforce (Behl, Sheorey, et al., 2021; Patrício et al., 2018). Advanced creative technologies, as well as the attractiveness of digital games, approach a unique opportunity for generating a fun or positive environment in the workplace (Pierce, 2019). That is, executives can take advantage of game-based project standards and accept the configuration, appearance, and sense of a game to get employee’s skills more constructive, pleasant, and exciting for members of the organisation, which may well improve managerial objectives (Bouzidi et al., 2019; Browne et al., 2018; Sanmorino et al., 2021).\n\nIn gamification, the entertaining and engaging environments usually observed in games are implemented to improve employees’ dedication to and engagement in real-life creative activities (Mitchell et al., 2020). For example, to enhance the effective learning process, the training and development unit implements certain gaming mechanisms and components to gain potential results (Leon & Peña, 2022). Team building, communication, logical reasoning, situation analysis, body language theory, and brainstorming activities. are the major components of gamification in a certain organisation (Silic et al., 2020). Companies utilise gamification in many aspects of the operation aligning with AI to enhance job engagement. Moreover, AI, ML, and IoT provide more opportunities for employees to concentrate on core issues and provide automation in many operational processes (Marculescu et al., 2020). The combination of gamification with AI, IoT, and ML could create an improved outcome in various tasks. This combination for example in recruitment, many companies use AI and gamification tools to shortlist candidates. Companies may use tools like ‘Scoutible’ a short game developed to determine the ability of the candidates to a particular task. Similarly, tools like Kahoot and Mentimeter are used to improve engagement and enhance interpersonal skills (Moorhouse & Kohnke, 2020). AL or ML combined games can also predict candidates’ thoughts, personalities, and decision-making capabilities by mapping the metrics required for a particular task and identifying the overlapping skills and qualities (Kranthi Kumar et al., 2020; Marculescu et al., 2020).\n\nThe potential use and benefits of the gamified system are described by flow theory. This theory proposes that an individual could achieve a status of flow, distinguished by the comprehensive concentration in the situation when completely engaged in executing a task (Moneta & Csikszentmihalyi, 1996). The concept of flow can be felt in several distinct situations. Artists can achieve flow while acting as a character, and athletes might feel it while playing at the boundaries of their physical ability (Patrício et al., 2018). Likewise, workers can achieve flow by feeling concentration or inclusion, enthusiasm, and enjoyment of the assignment they execute (Duggal & Gupta, 2020). Thus, we postulated that the beginning of gamification in the human resource management approach may influence employee engagement and productivity through perceived adoption, recognition, usefulness, and motivation (Brangier & Marache-Francisco, 2020; Whitson, 2013).\n\nOur research proposes that gamified workplace practice could enhance job engagement and productivity of employees through four factors namely perceived adoption, recognition, usefulness, and motivation (Figure 1). In this research, we examined how these aspects are related to better job engagement and productivity at the workplace and how they help in establishing potential mediating methods through which they correlate to the results. The entire paper was vested around testing the interrelationship between these variables and they are pictorially represented as follows:\n\nSource: Authors’ development.\n\nJob engagement has been defined as an ‘individual’s enthusiasm and involvement in a task assigned to them (Metwally et al., 2021). The highly involved individuals are usually motivated since they are identified in their jobs (de la Peña et al., 2021). They tend to work more efficiently and productively (Behl, Jayawardena et al., 2021). Adoption of a gamified task in an employee’s role and applying those features to involve more efficiently in their organizational task (Moneta & Csikszentmihalyi, 1996) will likely transform the work environment into a productive one, which supports the interconnection between experiencing the game and appreciating work. Therefore, we affirm that if an individual appreciates the gamified HRM or operational method and realizes it is enjoyable, thrilling, or fascinating, applying gamification quickly to their organizational task is likely to improve job engagement. Therefore, Hypothesis 1, states as follows:\n\nH1: Employees’ perceived adoption in the gamified HRM system is significantly associated with job engagement.\n\nRecognition indicates the opinion collected from the society, which causes by kinds of employee engagement that can take up the type of online engagement or accomplishment (Murray, 2018). A well-structured recognition system helps organizations to enhance productivity (Kour et al., 2019). Recognition can be merely explained as the explicit opinion collected through an HRM system on job-related activities (Jeske et al., 2021), and it is designed through the actions that employees examine (Zitars et al., 2021). Recognition often establishes reciprocal behavior whereby a worker could either obtain or offer feedback (Magni et al., 2021), which results in generating more respect and advantages for the whole HRM system as additional relationships and community interactions are made (Ángeles López-Cabarcos et al., 2021). Further gaining positive recognition from colleagues or line managers encourages a worker’s enthusiasm to identify others mutually and reciprocally while utilizing a service, indicating that obtaining recognition generates productivity (Sadick et al., 2020). The performance of employees is often associated with effective performance appraisal and reward systems (Adin, 2021). Gamification or gamified HRM systems can positively create an automated recognition environment that results in productive workforce Considering the importance of recognition in the human resource system, Hypothesis 2 is proposed.\n\nH2: Employee recognition in the gamified HRM system has a significant impact on productivity.\n\nEmployees’ perceived usefulness of gaming signifies the degree of confidence in job engagement after applying a gamified HRM system (Küpper et al., 2021). Job engagement is associated with developing the performance and efficiency in job-related responsibilities (Aubert & Lienert, 2019). Prior findings recognized a relationship between usefulness and job engagement (Golrang & Safari, 2021). If workers realize that the gamified technique is beneficial to them in their job, it will be possible to also realize their engagement in tasks and productivity to increase as a consequence of the executed approach (Sanchez et al., 2020). In a gamified system, individual engagement is through internal communication, familiarity with tasks, and enthusiasm for their assignment (Höllig et al., 2020). For instance, the gamified HRM approach could enable more efficient communication over the shared interaction among the workforce, which relates to the development of the game features (Mullins & Cronan, 2021). Therefore, due to advanced interaction and usefulness, it is feasible for the workforce to perform their job more effectively and ambitiously, thus developing engagement in the organizational tasks (Diefenbach & Müssig, 2019). From an organizational view, we are further likely to see a significant influence on job engagement if the organisational system with the gamified mode is employee-friendly to use and beneficial in performing tasks (Treiblmaier & Putz, 2020). Hence, if gamification is helping the workforce to improve their performance and efficiency in outcomes, it must have the advantage to improve their efficacy and overall enthusiasm. Subsequently, perceived usefulness must be positively impacted by an employee’s job engagement. Accordingly, hypothesis 3 can be proposed as follows.\n\nH3: Employees perceived usefulness in the gamified system is positively associated with job engagement.\n\nMotivation is a vital factor for an individual to convert their energy into a productive outcome (Jeha et al., 2022). Motivation encourages workforce commitment and engagement in tasks more effectively than unsatisfied or demotivated manpower (Albro & McElfresh, 2021; Heyns et al., 2021; Owen et al., 2018; Stirpe et al., 2021). Motivation is the intensity of a person’s motivational feel while engaged in an occupation. Prior findings have proved a positive association between motivation and productivity through job engagement (Tziner & Tanami, 2013). Individuals’ self-motivation often leads to better efficiency and performance in tasks (Saks, 2021). Therefore, we affirm that when workers are well interested to apply the gamified HRM and operational approach, it provides greater amounts of job engagement and productivity due to the significant impacts of the motivational aspects that enhanced the employee’s well-being and emotional association with the organization. Gamification can be a useful tool for motivating employees inside an organisation, and motivating employees is positively correlated with productivity, a crucial result that companies are interested in. Therefore, hypotheses 4 and 5 are proposed in the following way.\n\nH4: Employees perceived motivation in the gamified system is significantly associated with productivity.\n\nH5: Employees’ Job engagement is mediating factor to enhance productivity.\n\n\nMethods\n\nThe adopted questionnaire was submitted to the ‘Academic Integrity and Ethics Committee’ of the College of Business Administration, Kingdom University, and got approved as per the research policy and procedure on 5th June 2022 with ref. no. (CBA/30/22). The data collected will be used only for academic research purposes. All the respondents who participated in this survey have given their written informed consent to participate in the study and use their feedback to publish in our research publications. The consent of the participants has been asked at the commencement of the questionnaire and the participants responded by accepting the statement in the instrument link “I am willing to participate in this survey”. After expressing their consent, rest of the questionnaire appears for the respondents and data has been considered for analysis. To protect the participants’ interest, personal information is kept confidential.\n\nThe current quantitative research explains the casual relation between role of gamification towards productivity and job engagement. The study was conducted from June 2022 to first weeks of February 2023 in Gulf Cooperation Council countries. The entire research framework was based on the primary investigation, and an online survey method was utilized to achieve the objectives of the research work. Data was chosen from various pools of countries located in the gulf region, and the service sector was predominantly chosen for sampling. The multistage sampling method was used for the choice of respondents from public and private domains. The respondents were rendering services in IT, banking, education, and the telecom sector. These respondents were chosen because of their proximity to work with information technology and with gadgets where gamification could be easily made accessible. The countries chosen for sampling were UAE, Saudi Arabia, Oman, Bahrain, Kuwait, and Qatar. Proportional samples have been drawn from countries in the Gulf region. The researcher’s familiarity with regions and their access to organizations enabled a judgment-based sampling. As judgment-based sampling was used to exert discretion in the choice of respondents, only those companies that used gamification concepts in the workplace participated as respondents. Meanwhile the missing data were addressed by deleting the incomplete cases.\n\nThe respondents were chosen from the organizations that adopted the gamification approach in their operations in the Gulf region. The researcher approached the human resource department of the selected companies both private and government ownership and distributed a questionnaire (online link) to respondents in various departments. A total of 600 respondents from all these countries were invited to participate in the survey from June to December 2022, and the researcher managed to receive 400 complete responses based on judgemental sampling methods. The number of samples was determined based on (Burmeister & Aitken, 2012), considering the number of organizations that adopted gamification in their routine operations.\n\nThe selection of participants initiated the administering cluster approach since samples were collected from different parts of the gulf countries. To avoid bias in sample selection, we used a multi-stage sampling approach, so that can ensure that the sample is more representative of the population by including a range of different groups or clusters.\n\nThe data has been analyzed applying SPSS (version 26) and identified descriptive statistics to summarize the characteristics of a quantitative variable, such as the mean, median, standard deviation, and range. Similarly, inferential statistics such as t-tests, ANOVA, regression analysis, and correlation analysis, are used to test hypotheses or make predictions about a population based on a sample of data. Similarly, SPSS Amos version 26 software is used to test complex relationships among variables. The open access alternative for SPSS Amos is Microsoft Excel. Amos also testified the model against the observed data and estimated the model parameters (factor loadings, regression coefficients, and error terms), examined the goodness-of-fit statistics (chi-square, RMSEA, CFI) to determine if the model fits the data.\n\nThe research frame comprised a sample set of 400 respondents drawn based on the judgemental sampling method. The questionnaire was distributed to 600 people out of which only 400 responses could be utilized due to faulty filling or lack of response. After removing the invalid responses, statistical analysis was conducted and a pilot study was conducted before reaching out to them. The sample size of 400 demonstrated sufficient statistical control. A 5-point Likert scale has been utilized to understand the opinion of the respondents regarding the variables. The instrument was predominantly adopted from the past literature and each construct was determined with three or four items using a Likert scale in the design ranging from one to five. The research instrument was categorized into four parts: Section 1: Demographic information (six items); Section 2: Gamification questions (three items) adopted from (Seaborn & Fels, 2015); Section 3: Details about perceived adoption (for example, application of gamification will accomplish tasks, improve my performance, enhance my effectiveness, make my job easy, four items) adopted from (Hu et al., 2023); Section 4: Details of perceived recognition (for example, colleagues must recognize my performance in newly adopted jobs, considering my feedback in new system etc., four items) adopted from (Ortiz-Rojas et al., 2019); Section 5: Details of Perceived Usefulness (For example: Using gamified system improves performance, increases productivity, enhances effectiveness in job etc., four items) adopted from (York & deHaan, 2018); Section 6: Motivation (For example; classified into intrinsic motivation, identified regulation, external regulation, amotivation, four items) adopted from (Zimmerling et al., 2019); Section 7: Productivity (for example; developed team spirit and believe in achieving business goals; ability to market product; contributing profit generation; ability to make quick and qualitative business decision, four items) adopted from (Zainuddin et al., 2020); Section 8: Job engagement (for example; exerting full effort into my job; hardest to perform well on job; absorbed by my task job, concentrated on my task etc., eight items) adopted from (Patricio et al., 2022).\n\nThe details of the results, SEM, and the statistical relevance of the tests conducted are summarised in the next few sections.\n\n\nResults and findings\n\nA glimpse of the descriptive variables in terms of demographic aspects and usage of gamification is listed below in Table 1. As depicted in Table 1, the target respondents mainly comprised of women (60.5%) in the age group of 31 to 40 years old which accounted for the majority of the population. In terms of their educational qualification, 49% had acquired a graduation degree (bachelor’s and post bachelor’s degree) or higher than the graduation degree. It was noted that 61.5% who responded were rendering services in the private sector and were working in the position of a team member and not in a managerial position. The survey-based study also included questions about their opinion about gamification and its usage in their daily life. 82% of the respondents were aware of gamification and 74% of them used gamification in their normal life. But only 9.5% of the respondents used it daily and 14.5% were occasional users and the rest of them hardly used gamification for their daily entertainment. Hence it can be inferred that most of the respondents did not depend upon gamification or any gamification-related mechanism for perceived enjoyment or interactions.\n\nTo find out the relationship that exists between demographic variables (Independent factors) and the factors chosen for the study namely Perceived adoption, Recognition, Usefulness, Motivation, and Productivity (Dependent factors), the Anova test was conducted. Based on the significant P values and the Partial Eta Squared, the results are summarised as follows.\n\nOnly the above given demographic variables (gender, education, and age) had a significant influence on Adoption, Recognition, and Usefulness. Most of the demographic variables did not influence the dependent variables and hence it can be inferred that the variables chosen are not significantly impacted by demographic constructs that are similar irrespective of age, gender, and qualifications.\n\nThe model was analyzed to measure the convergent validity, discriminant validity, and reliability of the constructs. Table 2 illustrates that the loading of each constructed item has outstripped 0.7 (Esposito et al., 2021), for each construct Cronbach’s alpha was found to be above 0.7 and the aggregate reliability is more than the standard of 0.7, signifying acceptable internal consistency and reliability of the items. Moreover, the average variance extracted (AVE) from each construct is higher than 0.5, indicating an acceptable convergent validity of the manuscript measurement model. The mean and standard deviation values of each item indicate that data are clustered around the mean.\n\nThe model was analyzed to measure the convergent validity, discriminant validity, and reliability of the constructs. Table 3 illustrates that the loading of each constructed item has outstripped 0.7 (Esposito et al., 2021), for each construct Cronbach’s alpha was found to be above 0.7 and the aggregate reliability is more than the standard of 0.7, signifying acceptable internal consistency and reliability of the items. Moreover, the average variance extracted (AVE) from each construct is higher than 0.5, indicating an acceptable convergent validity of the manuscript measurement model. The mean and standard deviation values of each item indicate data are clustered around the mean.\n\nAmos-SEM was administered in the study to understand gamification’s influence on productivity and job engagement. Figure 2 illustrates the path coefficient for the research model and all the coefficients relate to the gamified model were significant. The recommended p values for average path co-efficient and average-square must be significant at 0.05 level (Cameron, 2013). Results indicate adequate model fit since the p-value for both these is lower than 0.05 (0.01 and 0.00) respectively. The association between perceived adoption and job engagement (H1) (β=0.017; p=0.005), perceived recognition and productivity (H2) (β=0.639; p=0.000), perceived usefulness, and job engagement (H3) (β=-0.782; p=0.000) and perceived motivation and job engagement (H4) (β=0.834; p=0.000) found to be a significant association and all four hypotheses accepted. On the other hand, mediating variables are vital factors to enhance productivity and results show in perceived adoption (H5a), job engagement mediates to enhance productivity, in contrast, is perceived recognition (H5b), perceived usefulness (H5c), and perceived motivation (H5d) job engagement does not mediate to enhance productivity. Table 4 summarizes the results of the hypothesis.\n\nSource: Data analysis.\n\nThe percentage of variance covered by the calculated population covariance is known as the (Adjusted) Goodness of Fit. Comparable to R2, the recommended GFI and AGFI values are >0.95 and >0.90, respectively (Marsh et al., 2019). A parsimony-adjusted index is the Root Mean Square Error of Approximation. Values that are nearer to 0 indicate a good fit. It should be either 0.05 or 0.08. The NFI has been updated to become the Comparative Fit Index. Unresponsive to sample size (Althoff & Neiva, 2021). evaluates how well a target model fits in comparison to a null, or independent, model. It should be > 0.90 (Marcoulides et al., 2020). The model exhibits that the CMIN/DF: is 4.580 and the p-value is <0.001. It indicates that the model is fit. The other parameters of the model are Goodness-of-Fit Index (GFI)=0.686, Adjusted Goodness-of-Fit Index (AGFI)=0.648, and Comparative Fit Index (CFI)=0.776. These parameters are above the threshold level. Based on the parameters of model results, the study concluded that the factors, perceived adoption, perceived recognition, perceived usefulness, and perceived motivation significantly influence the enhancement of job engagement and organizational productivity.\n\nThe major finding of the study notifies that 82% of the respondents are aware of gamification and are working in the private sector with minimum graduation as their qualification and in team personnel positions. Similarly, the demographic variables do not influence the dependent variables and are similar in terms of their influence. The employees perceived adoption of the gamified human resource management system is significantly associated with work engagement. In the outcome, employee recognition in the gamified HRM system also has a significant impact on productivity. The result further notifies that employees’ perceived usefulness in a gamified system has been significantly correlated with job engagement. The employees’ perceived motivation in the gamified system is also significantly associated with productivity. The outcome of the dependent variable also amplifies the similar outcome by revealing that employee job engagement is a mediating factor to enhance productivity. Overall, the factors, perceived adoption, perceived recognition, perceived usefulness, and perceived motivation significantly influence the enhancement of job engagement and organizational productivity. The underlying data and questionnaire are publicly available (Rahiman et al., 2023a, 2023b).\n\n\nDiscussion\n\nPast literature emphasizes the efforts of industries to enhance employee engagement and productivity with innovative operational practices. In this framework, studies report gamification as a predominant theoretical framework that identifies and conceptualize significant mechanism to influence organizational performance and workforce engagement through various methodologies.\n\nThis study identified the role of gamification to enhance productivity and individual job engagement in various organizations. Although gamification gained popularity in various countries and notable organizations like Google, Apple, Microsoft, Cisco, and Samsung, in their operation, training and development, and human resource activities like recruitment and selection, comprehensive implementation and adoption of gamification in organizational supply chain system remain challenging (Alhammad & Moreno, 2020). It would be challenging for the organizations in the Gulf region to implement gamification in organizational operational activities due to its low popularity. This challenge remains at the industry level where the Gulf region is dominated by energy and power sectors, and transportation and logistics in which gamification isn’t much entertained in operational level (Mekler et al., 2017; Mitchell et al., 2017). Another vital justification for this challenge was that individuals who work in knowledge-intensive industries or companies have tasks that are naturally fun or creative where gamified systems are considered to have lesser scope. As per past studies, at the institutional level, it is challenging to integrate gamification in large firms due to the regulations (Sailer et al., 2017). On the other hand, in small firms, gamification is an unwanted overhead since the outcome of post-implementation is not attractive. Our study drew a difference between government and private organizations in which respondents believe that it is exceedingly challenging to apply gamification in ministry or government organizations since they do not understand this process. Meanwhile, previous studies have revealed that at a strategic level, leadership-concerned challenging aspects had mostly to do with the perception of higher management of an organization (Hammedi et al., 2021). Most of these studies opine that the top management must be tolerant enough to expect outcomes that usually do not happen in most organizations.\n\n\n\na) 82% of the respondents are aware of gamification and are working in the private sector with minimum graduation as their qualification and in team personnel positions.\n\nb) The demographic variables do not influence the dependent variables and are similar in terms of their influence.\n\nc) Employees perceived adoption in the gamified HRM system is significantly associated with job engagement.\n\nd) Employee recognition in the gamified HRM system has a significant impact on productivity.\n\ne) Employees perceived usefulness in the gamified system is positively associated with job engagement.\n\nf) Employees perceived motivation in the gamified system is significantly associated with productivity.\n\ng) Employee Job engagement is a mediating factor to enhance productivity.\n\nh) The factors, perceived adoption, perceived recognition, perceived usefulness, and perceived motivation significantly influence the enhancement of job engagement and organizational productivity.\n\nGamification is a popular concept in the current new trends in HR. based on empirical evidence it has been proven that gamification contributes to increasing in productivity and job engagement. This research paper has shown that gamification has an impact on work engagement and productivity through pursued adoption, recognition, usefulness, and Amotivation in the organization system. Depending on the size, type, and nature of the organization the impact of gamification varies.\n\nThe findings have significant implications for insight into how employees in the service sector are aware of the gamified working environment and react to the system through work engagement and productivity. These findings could be beneficial to industrial practitioners particularly human resources to consider certain key aspects while adopting gamified working practices. Gamification is an emerging trend in the field of human resources. Most organizations are opting for the implementation of gamification with the intention to enhance employee engagement. From application to enhance employee wellness to performance tracking gamification has been applied in every arena of HR. This research article substantiates that gamification has an impact on work engagement and productivity. However, to be effective implementation top management support and cooperation are essential. The implication of this system also differs in terms of the size and needs and resources of the organisation.\n\nIt is essential to recognize that managers should recognise the requirements of the workforce and needs to devote time to know which form of engagement procedures would be appropriate and most excellent for employee productivity. A suitable need-based execution strategy should be constructed by the organisation and pretesting of gamified sections must be performed on smaller events to commence with feasibility and effectiveness to enhance engagement and performance. These requirements and plans to change generation-wise and industry-wise, as already considered in the earlier parts of this manuscript. Finally, as gamification approaches vary among companies, personnel, and their employment profile, it is important for companies to put these elements on a canvas to assist the organizations in vigorously choosing strategies to apply the proper game mechanics. This will improve and benefit them accomplish sustainability by applying gamification for employee productivity and job engagement.\n\n\nConclusion\n\nPeople are coded genetically, just like computers and various permutations and combinations play an important role in improvising their productivity in terms of work. Gamification no doubt has been a recently sought-out tool to improve the job engagement of employees that further enhances productivity, but nevertheless, it is not the only relevant tool. In an IT-enabled work environment, gamification has shown a positive impact, but in the case of pure services like academics, and hospitals it is yet to make a mark. In order to check its relevance, the psychological perspective of individuals has to be tapped on. The study shows that there are people who have performed well though they have not used gamification. Newer methodologies or technological innovations may be needed to use gamification for different industries. That necessitates a new breed of gamification technologies that are tailor-made to suit the needs of different classes of employees. Further research may be warranted in this perspective of widening the scope of gamification.\n\nThe research limitations should be taken into account when interpreting the results. The research was conducted via an electronic survey, which is susceptible to frequent method bias. Data were only collected in Gulf countries only, which limits the generalizability of the findings. The study was based on a sample survey and was not experimental, hence it does not fall under the gamut of causality. Further, the empirical study is based upon only those institutions and organizations which included gamification as a part of their work. Respondents who were working in an organization without gamification would have increased their knowledge with regard to the impact of gamification on productivity.\n\nIn conclusion, it can be written that gamification is quite a new terminology and hence it warrants full-fledged research in this domain. This research paper is an attempt to showcase the usage of gamification in a single sector and can prove to be a vital point for further research.\n\n\nAuthors’ contributions\n\nHabeeb Ur Rahiman: Conceptualization, Formal Analysis, Funding Acquisition, Investigation, Methodology, Resources, Supervision, Writing – Original Draft Preparation\n\nRashmi Kodikal: Conceptualization, Data Curation, Formal Analysis, Investigation, Methodology, Supervision, Validation, Writing – Original Draft Preparation, Writing – Review & Editing\n\nDr. Sucharitha Suresh: Software, Validation, Visualization, Writing – Review & Editing", "appendix": "Data availability\n\nFigshare: Game on: Can gamification enhance productivity?, https://doi.org/10.6084/m9.figshare.22083158 (Rahiman et al., 2023a).\n\nThe project contains the following underlying data:\n\n- Gamification and Productivity Data.sav (Questionnaire responses)\n\nFigshare: Questionnaire. https://doi.org/10.6084/m9.figshare.22262974 (Rahiman et al., 2023b).\n\nThe project contains the following extended data:\n\n- Questionnair Gamification.pdf\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\n\nAcknowledgments\n\nWe hereby acknowledge Kingdom University for their support to carried out research and all the institutions that provided data as open-source and experts who shared their views.\n\n\nReferences\n\nAdin CA: Addressing Unsatisfactory Performance in Employees. 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Publisher Full Text" }	[ { "id": "186391", "date": "31 Jul 2023", "name": "Easwaramoorthy Rangaswamy", "expertise": [ "Reviewer Expertise Management" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe manuscript focuses on one of the contemporary issues, which I think is very interesting and a complex issue too. The authors have managed to articulate the issue reasonably well in the introduction.\nThe introduction part can have a subsection about the research problem and can also show the research questions clearly.\nThe conceptual model can be aligned with the Literature review and the proposed hypotheses. Also, ensure that the latest literature is provided. There are not many 2022/2023 reviews provided.\nMethodologically, the paper does not lack rigour, but it needs more clarity on the sample selected, demographics and also potential limitations due to the methodology. It does not mention the “representativeness of the sample”. The methodology section need also to include details about the research design, research philosophy, research approach, etc. Provide a clear and detailed explanation of the research design and methodology employed in the study. Clearly state the research approach and justify its suitability for addressing the research questions.\nThe section “Ethical Considerations” mentioned, “The data collected will be used only for academic research purposes”. It is suggested to write in the past tense, as the data collected is not in the report stage.\nThe results and findings section should go beyond a mere summary and interpretation of the findings. Provide an in-depth analysis of the data, relating it back to the research questions and objectives. Clearly explain the implications of the findings and their significance in the context of the prevailing literature available. Additionally, discuss any unexpected or contradictory findings and offer potential explanations. I see that certain findings are discussed but I also see a sub-section named “key findings” in section Discussion. Looks duplicated. Also, avoid point-wise writing in “key findings”.\nThe discussion section should include a comprehensive review and synthesis of relevant literature. Compare the findings of the study with existing research in the research area. Identify areas of agreement or divergence and discuss the potential reasons behind these differences. This will strengthen the validity and generalizability of the study's findings.\nReview the manuscript for coherence and flow, ensuring the content is well-organized and logically structured.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] }, { "id": "189162", "date": "08 Sep 2023", "name": "Ola Olayinka", "expertise": [ "Reviewer Expertise Gamification", "information systems", "research methods and qualitative research." ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nInteresting paper covering gamification and its application in organisations in the gulf region. The paper was mostly well written and clear to read with some recent papers in the field cited.\nHowever, there are a number of issues that need to be clarified or better presented.\n\".. our research was taken up to examine the impact of gamification in various organizations and simultaneously see its relationship with job engagement and productivity. The primary objective of this research is to identify the role of perceived usefulness, motivation, adoption, and recognition in the organization while adopting gamified tasks in an HR system.\" It is not entirely clear what the focus of the paper is, examining impact of gamification in organizations, examining its relationship with engagement and productivity or identifying the role of perceived usefulness, motivation, adoption, and recognition in when gamification is used, or a combination of three of them.\nFrom the hypothesis, it is clear that you are trying to understand how perceived usefulness and perceived adoption of gamification in HRM systems influences engagement and motivation. I will suggest that it is made clear in the above quote.\n\nI will suggest general proof-reading of the article as there are some grammatical errors that can make reading the article a bit difficult to understand.\n\nI will suggest the inclusion of a detailed literature review section that critically assesses and discusses other similar research.\n\nA bit more needs to be done in the methodology section. There needs to be more information provided about the research sample and its appropriateness. How many respondents were from each country? The lack of clarity allows for the generalisability of the results to be questioned even in the gulf region.\n\nThere are a number of sentences (or claims) in the discussion section that you need to justify with appropriate references.\ne.g. \"In this framework, studies report gamification as a predominant theoretical framework that identifies and conceptualizes a significant mechanism to influence organisational performance and workforce engagement through various methodologies.\" What studies are these?\n\"It would be challenging for the organisations in the Gulf region to implement gamification in organisational operational activities due to its low popularity. \" Please provide a reference.\n\n\"On the other hand, in small firms, gamification is an unwanted overhead since the outcome of post-implementation is not attractive. \". Please provide a reference.\n\nSome parts of the discussion were not clear. e.g. \"Our study drew a difference between government and private organizations in which respondents believe that it is exceedingly challenging to apply gamification in ministry or government organizations since they do not understand this process.\" How was this done? It was not clear from the methodology or objectives that this was a goal.\n\nThere needs to be more discussed about the findings in the discussion section. This section seems quite weak and omits key findings that the reader would like to know.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [ { "c_id": "10362", "date": "18 Oct 2023", "name": "Habeeb Ur Rahiman", "role": "Author Response", "response": "Dear Reviewer, We would like to express our deep gratitude for your invaluable feedback and the dedicated time you invested in reviewing our research. Your insightful comments have played a pivotal role in guiding our efforts to enhance the overall quality and rigor of our study. Below, we provide comprehensive responses to each of the points you raised. Section Enhancement: We have diligently implemented the changes recommended for the specified section, ensuring that your valuable suggestions have been incorporated effectively. Proofreading and Quality Assurance: In response to your suggestions, we conducted a thorough proofreading of our work, addressing any grammatical and language-related concerns to ensure the utmost clarity and readability. Literature review: While the journal requested a title labeled as \"Background,\" we have adjusted it to align with your preference, now designating it as \"Literature Review\" to accurately represent its content and purpose. Methodology Refinement: Following your guidance, we have made the necessary revisions in the methodology section. Specifically, we have provided a detailed breakdown of the sample respondents, highlighting the distribution by country for enhanced clarity and transparency. Discussion Restructuring: We have undertaken a comprehensive restructuring of the discussion section, addressing instances where references were missing and ensuring that all relevant points are now properly cited. Additionally, we have enriched the section by incorporating a more extensive set of references to further elucidate our results. We sincerely hope that these adjustments meet your expectations and align with the standards expected for publication. If you find the need for any further modifications or have additional suggestions, please do not hesitate to communicate them to us. Your continued guidance is highly valued and greatly appreciated. Warm regards," } ] } ]	1	https://f1000research.com/articles/12-818
https://f1000research.com/articles/12-164/v1	13 Feb 23	{ "type": "Study Protocol", "title": "Characteristics of bibliometric analyses of the complementary, alternative, and integrative medicine literature: A scoping review protocol", "authors": [ "Jeremy Y. Ng", "Henry Liu", "Aimun Qadeer Shah", "L. Susan Wieland", "David Moher", "Henry Liu", "Aimun Qadeer Shah", "L. Susan Wieland", "David Moher" ], "abstract": "Background: There is a growing body of literature on complementary, alternative, and integrative medicine (CAIM), which offers a holistic approach to health and the maintenance of social and cultural values. Bibliometric analyses are an increasingly commonly used method employing quantitative statistical techniques to understand trends in a particular scientific field. The objective of this scoping review is to investigate the quantity and characteristics of evidence in relation to bibliometric analyses of CAIM literature. Methods: The following bibliographic databases will be searched MEDLINE, EMBASE, PsycINFO, AMED, CINAHL, Scopus and Web of Science. Studies published in English, conducting any type of bibliometric analysis involving any CAIM therapies, as detailed by an operational definition of CAIM adopted by Cochrane Complementary Medicine, will be included. Conference abstracts and study protocols will be excluded. The following variables will be extracted from included studies: title, author, year, country, study objective, type of CAIM, health condition targeted, databases searched in the bibliometric analysis, the type of bibliometric variables assessed, how bibliometric information was reported, main findings, conclusions, and limitations. Findings will be summarized narratively, as well as in tabular and graphical format. Conclusions: To the best of our knowledge, this scoping review will be the first to investigate the characteristics of evidence in relation to bibliometric analyses on CAIM literature. The findings of this review may be useful to identify variations in the objectives, methods, and results of bibliometric analyses of CAIM research literature.", "keywords": [ "bibliometric analysis", "complementary and alternative medicine", "integrative medicine", "scientometric analysis", "scoping review" ], "content": "Introduction\n\nComplementary, alternative, and integrative medicine (CAIM) is a complex term referring to three distinct concepts related to the use of non-conventional medicine.1,2 “Complementary medicine” describes non-conventional therapeutic approaches that are used together with conventional therapies.1 “Alternative medicine” describes non-conventional therapeutic approaches used in replacement of conventional therapies.1 “Integrative medicine” describes the combined use of both conventional and non-conventional therapies in a coordinated manner.1,2 For the purpose of this study, each of these approaches may also incorporate elements of “traditional medicine” which is the “knowledge, skills and practices based on the theories, beliefs and experiences indigenous to different cultures, used in the maintenance of health and in the prevention, diagnosis, improvement or treatment of physical and mental illness”.3 All these concepts will be collectively referred to as “complementary, alternative, and integrative medicine” (abbreviated as CAIM).\n\nCAIM practitioners often emphasize a holistic approach to health, including the consideration of cultural and social values.2,4 Clients often perceive CAIM as better at providing individualized, person-centred care, compared to mainstream health approaches.5,6 Prevalence of CAIM use is increasing worldwide, and, accordingly, the body of literature on CAIM research has grown immensely, with the steepest increase in CAIM publications observed between the mid-2000s and mid-2010s.7,8 It is of interest to determine broad research trends of CAIM research literature, and identify specific CAIM topics explored (e.g., acupuncture, aromatherapy). While some CAIM therapies (e.g., yoga for depressive symptoms,9 exercise therapy for reducing falls in older people10) have been shown to be safe and effective, many other therapies have insufficient evidence to demonstrate their effectiveness or safety.11,12 Furthermore, even when basic effectiveness and safety are established, questions often remain about key characteristics such as intervention dose and implementation or applicability to different patient populations and settings. Bibliometric analyses can be used to detect knowledge gaps and to identify research trends that help predict whether such knowledge gaps are likely to be met.\n\nBibliometric analysis involves the application of quantitative statistical techniques to bibliometric data (e.g., total number of citations, total number of publications) and can be used for a variety of purposes, such as identifying patterns in a given field of research.13 Bibliometric analysis techniques can broadly belong to categories of performance analysis (i.e., techniques measuring contributions of research constituents) or science mapping (i.e., techniques measuring relationships between research constituents).13 Examples of research constituents include authors, countries, institutions, and topics.13 Performance analysis techniques can further be divided into publication-related metrics (e.g., total number of publications), citation-related metrics (e.g., average citations, total number of citations), and citation-and-publication-related metrics (e.g., h-index, g-index, proportion of cited publications).13 Science mapping techniques can include methods such as citation analysis, co-citation analysis, bibliographic coupling, co-word analysis, and co-authorship analysis.13 For instance, co-citation analysis examines the frequency of publications being cited together, which may reveal thematic clusters.13 Enrichment techniques of network metrics (i.e., quantitative measures of research constituents’ relative importance), clustering (i.e., grouping of similar objects using clustering algorithms), and visualization (i.e., graphical visualizations of research constituents’ connections) can be employed to enhance understanding of science mapping techniques.13 For instance, software like VOSviewer can be used to graphically visualize thematic clusters in co-citation analysis.13\n\nAdvantages of bibliometric analyses include facilitating the examination of large datasets that are not feasible for investigation by manual review (e.g., literature reviews).13 Further, the relatively low cost and rapidity of conducting bibliometric analyses allow for replicable methods.13,14 Use of bibliometric techniques across different scientific fields is becoming increasingly popular.15,16\n\nA scoping review, which involves mapping the current literature and identifying gaps in research,17 would be appropriate to summarize literature on CAIM bibliometric analyses. To the best of our knowledge, no systematic or scoping reviews have been conducted on bibliometric analyses of CAIM therapies. A preliminary search of the Cochrane Database of Systematic Reviews and the Scopus database revealed no existing systematic or scoping reviews on the topic. Synthesizing bibliometric analyses on CAIM will provide insight into trends, such as the types of CAIM literature typically analysed from a bibliometric lens, statistical techniques that bibliometric analyses on CAIM utilize, and more broadly, where the field of CAIM is headed. Considering the lack of standardized procedures for bibliometric analyses, this review will also improve understanding of how bibliometric analyses are currently conducted on this topic. Thus, the purpose of this review will be to understand the characteristics of bibliometric analyses of CAIM research literature, which can inform future work within the field.\n\n\nProtocol\n\nApproach and eligibility criteria\n\nThe present scoping review’s research question is: “What are the characteristics of bibliometric analyses of the CAIM literature?”. The proposed scoping review will be conducted in accordance with the Joanna Briggs Institute (JBI) methodology for scoping reviews, which recommends quantitative and descriptive analyses of main findings.17 This protocol’s associated files (Appendix A – Search Strategies, and the PRISMA ScR Checklist) has been made openly available on Open Science Framework (see Extended data and Reporting guidelines33).\n\nThe search strategy will aim to locate studies published in journals only (excluding grey literature), with no date restrictions (i.e., from inception to date of search execution). The only eligible study design will be bibliometric analyses (encompassing terms for “bibliometric analysis”, “scientometric analysis”, and “citation analysis”), or articles that include both a bibliometric analysis and another study design (e.g., bibliometric analysis and systematic review). All included bibliometric analyses will be focused on one or more CAIM therapies, as defined by a recently published operational definition of CAIM.18 This operational definition was created using a systematic search of four peer-reviewed or other quality-assessed resource types: 1) peer-reviewed articles from seven major bibliographic databases, 2) “Aims and Scope” webpages of peer-reviewed CAIM journals, 3) entries containing CAIM therapies in highly accessed online encyclopedias, and 4) highly ranked websites resulting from Health On the Net Code of Conduct (HONcode) searches.18 To date, this operational definition includes the greatest number of evidence sources and is the only one that captures the concept of “integrative medicine”, alongside “complementary medicine” and “alternative medicine”.18\n\nGrey literature sources will be excluded as bibliometric analysis studies are unlikely to be found outside of traditional academic publishing channels. Conference abstracts and study protocols will also be excluded as they likely will not contain adequate information required to describe the characteristics of bibliometric analyses on CAIM literature. Finally, all non-English publications will be excluded, due to language constraints of the authors.\n\nSearch strategy\n\nThe following electronic databases will be searched: MEDLINE, EMBASE, PsycINFO, and AMED (accessed via the OVID research platform), as well as CINAHL (accessed via EBSCOhost), Scopus, and Web of Science. The search strategy will include a comprehensive search string of CAIM terms19 encompassing 604 distinct therapies described previously in an operational definition of CAIM.18 This search string of CAIM terms was created for OVID (e.g., MEDLINE, EMBASE, PsycINFO, AMED) and EBSCO platform (e.g., CINAHL) databases, as well as Scopus and Web of Science databases.19 Relevant scientific names and/or synonyms were added as a term (i.e., keyword, phrase), alongside relevant boolean operators. The comprehensive search string of CAIM19 will be combined with search terms for bibliometric analyses (e.g., bibliometric analysis, statistical bibliography, citation analysis). The search strategy, including all identified keywords and equivalent index terms, will be adapted for each included database. All search strategies that will be run are provided in Extended data,33 informed by PRISMA-S guidelines for reporting literature searches.20,33\n\nStudy and source of evidence selection\n\nFollowing the search, all identified citations will be collated and exported into Covidence, and duplicates will be removed. All titles/abstracts, followed by full texts, will be screened by reviewers independently and in duplicate. First, pilot title/abstract screening of a sample of twenty articles will be conducted by AQS and HL. A meeting will be held between AQS, HL, and JYN to discuss any challenges and resolve discrepancies. Following the pilot test, all titles/abstracts will be screened for inclusion by AQS and HL. Then, pilot full-text screening will be conducted, in which AQS and HL will screen ten full texts. A meeting will be held between AQS, HL, and JYN to discuss challenges and resolve discrepancies. After this pilot step, full-text screening will be completed by AQS and HL. Reasons for exclusion of full texts will be recorded. Any disagreements that arise between reviewers throughout the selection process will be resolved on a weekly basis through discussion with HL, AQS, and JYN, if disagreements still cannot be resolved. The results of the search strategy and the study screening process will be presented in a PRISMA-ScR flow diagram in the final scoping review.21\n\nData extraction\n\nData extraction will be conducted using Excel software. The data extraction form that will be used in this scoping review is informed by Donthu et al.,13 which provides an overview of how to conduct a bibliometric analysis. The extraction form will be developed in two stages. In stage one, AQS and HL will select ten articles at random that met the inclusion criteria from a preliminary search of CAIM and bibliometric analysis search terms on Scopus. AQS and HL will independently extract information from five articles each and meet to discuss discrepancies. A meeting will be held among AQS, HL, and JYN, to discuss changes needed to improve the form. In stage two, AQS and HL will identify the ten most highly cited articles that met inclusion criteria after running a search of CAIM literature and bibliometric analyses terms on Scopus, before independently extracting information from five articles each using the latest version of the data extraction form. Another meeting will be held between AQS and HL, and then with JYN, to approve the latest version of the extraction form.\n\nThe following information will be extracted from eligible bibliometric analyses: title, author, year, country, aim of the study, secondary study design (if applicable), the type of CAIM(s), the health condition or population targeted, main findings, conclusions, and limitations. Also, the bibliometric information described will be summarized, including the databases searched, type of bibliometric methodology (i.e., performance analysis [such as citation-metrics or publication metrics] versus science mapping [such as co-word analysis, co-authorship analysis, bibliographic coupling, or enrichment techniques]), the number of studies included in the analysis, the number of metrics used, how information was reported (e.g., narrative summary, figures, tables, visualization software used), and how all variable measures align with the Donthu et al.13 guideline for conducting bibliometric analyses.\n\nTo ensure consistency and quality of the extraction, an initial pilot test of data extraction will be conducted by all participating independent reviewers, assessing five articles. All reviewers will then meet with JYN, HL, and AQS to resolve discrepancies and disagreements. Based on the initial pilot testing, revisions to the data extraction form can be proposed and implemented. Upon completion of the pilot extraction step, reviewers will be divided into two teams led by HL and AQS. Teams will be further divided into pairs of two reviewers for duplicate data extraction of the same set of bibliometric full texts. These duplicate extractions will be reviewed by AQS and HL to ensure consistency. Weekly meetings will be held between reviewers and HL and AQS to standardize the data extraction process and resolve any issues identified. Any conflicts that cannot be resolved will be discussed with JYN.\n\nWe seek to conduct a scoping review. As we are going to use the JBI methodology for scoping reviews, we have elected not to conduct a risk of bias assessment.\n\nThe data will be summarized descriptively (e.g., frequencies of the number of studies, country, types of CAIM, outcomes reported in the bibliometric analyses) with full results presented in tabular format. Frequency of CAIM bibliometric analysis publications over time will be presented in graphical format. An additional figure will be created highlighting the types of bibliometric information each study reported.\n\nDissemination\n\nThe findings of this review will be disseminated in scientific journals.\n\nThe literature search is ongoing.\n\n\nDiscussion\n\nWe anticipate this project will advance the understanding of topics and trends in CAIM research, including what types of CAIMs are most commonly explored and what health conditions these CAIMs target. Accordingly, it can help CAIM researchers locate bibliometric sources to inform future research directions or identify gaps that warrant further investigation. It is anticipated that this review will also provide unique insights on how bibliometric analyses of CAIM literature are conducted, including the type of methodology used (such as performance analysis metrics or science mapping techniques), the number and types of outcomes reported, and how bibliometric information is presented (e.g., narratively, graphically).\n\nAs the number of scientific publications has been growing exponentially in the last fifty years, bibliometrics has become useful to quantitatively analyse publications on a particular topic.22 Generally, there is large variability in how bibliometric analyses are conducted, as there is no authoritative guideline on bibliometric methodology.13,16 Findings from the completed review can help identify whether there are any inconsistencies in the way that bibliometric analyses specifically on CAIM literature are conducted. This may be useful to inform future, standardized reporting guidelines for bibliometric analyses, generally, or with a potential focus on CAIM topics. Further, it may be expected that performance analysis metrics used to measure research constituents’ scientific impact (e.g., h-index, g-index, total citations) differ between bibliometric analyses.23 Given the varying capability of different performance analysis metrics in capturing scientific impact of a given research constituent,24 this review could reveal the extent to which bibliometric analyses of CAIM literature are effectively measuring scientific impact. Comparisons of different research constituents’ scientific impact are further complicated by how average values of bibliometric indicators often differ between disciplines (e.g., molecular biology, nursing).22 This is particularly pertinent to CAIM literature, given the interdisciplinary applications of many CAIM therapies.25–27\n\nBibliometric methodology can be influenced by database changes such as the indexing of new journals or articles.28 Due to limitations of visualization softwares like VOSviewer, often only either Scopus or Web of Science databases can be searched, which cover most but not all databases.29 If the findings of this scoping review reveal similar limitations of bibliometric analyses conducted on CAIM literature, these limitations could potentially drive investigation into techniques that will improve analysis of bibliometric results. Additionally, there are no known critical appraisal tools for bibliometric analyses. Concerns have been expressed over a lack of knowledge of good practices when conducting bibliometric analyses.30 While this is outside the scope of this present review, future research may investigate assessment tools to evaluate the quality of published bibliometric analyses. The identification of a group of bibliometric analyses in this review may potentially serve as a test set for the development and investigation of quality indicators.\n\nStrengths of this study will include adherence to the JBI methodology for scoping reviews17 and use of a comprehensive systematic search strategy19 across several databases to identify eligible articles. Another strength is that screening and data extraction will be conducted in duplicate, significantly reducing bias. There are some limitations expected in this review. By only including studies written in English, we could be missing important international work. For example, Chinese databases may contain a higher volume of CAIM articles but are unable to be searched in this study. This is especially relevant as some forms of CAIM may be practiced more frequently in non-English speaking regions of the world, such as traditional Chinese medicine in China.31,32 Additionally, reported findings are expected to be descriptive in nature, such as the frequencies of the types of CAIMs examined in bibliometric studies or the frequencies of studies that engaged in science mapping versus performance analysis techniques. This makes it difficult to extrapolate themes or correlates, like which CAIMs are effective, or which bibliometric techniques are preferable.", "appendix": "Data availability\n\nNo underlying data are associated with this article.\n\nOpen Science Framework: Characteristics of Bibliometric Analyses of Complementary, Alternative, and Integrative Medicine Literature: A Scoping Review. https://doi.org/10.17605/OSF.IO/JSQWY. 33\n\nThis project contains the following extended data:\n\n- Appendix A - Search Strategies_Jan0923.docx (search strategies for MEDLINE, EMBASE, PsycINFO, AMED, CINAHL, Scopus and Web of Science databases).\n\nOpen Science Framework: PRISMA-ScR checklist for ‘Characteristics of bibliometric analyses of the complementary, alternative, and integrative medicine literature: A scoping review protocol.’ https://doi.org/10.17605/OSF.IO/JSQWY. 33\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nNational Center for Complementary and Integrative Health: Complementary, alternative, or integrative health: what’s in a name?2021.Reference Source\n\nNg JY, Boon HS, Thompson AK, et al.: Making sense of “alternative”, “complementary”, “unconventional” and “integrative” medicine: exploring the terms and meanings through a textual analysis. BMC Complement. Altern. Med. 2016; 16: 134. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWorld Health Organization: Traditional Medicine.2021.Reference Source\n\nGillett J, Andrews GJ, Savelli M: Health & Society: Critical Perspectives. Oxford University Press;2016.\n\nBishop FL, Yardley L, Lewith GT: Treat or treatment: a qualitative study analyzing patients’ use of complementary and alternative medicine. Am. J. Public Health. 2008; 98(9): 1700–1705. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDodds S, Bulmer S, Murphy A: Consumer value in complementary and alternative medicine (CAM) health care services. Australas. Mark. J. AMJ. 2014; 22(3): 218–229. Publisher Full Text\n\nNg JY: Insight into the characteristics of research published in traditional, complementary, alternative, and integrative medicine journals: a bibliometric analysis. BMC Complement. Med. Ther. 2021; 21(1): 185. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTangkiatkumjai M, Boardman H, Walker DM: Potential factors that influence usage of complementary and alternative medicine worldwide: a systematic review. BMC Complement. Med. Ther. 2020; 20(1): 363. Publisher Full Text\n\nCramer H, Lauche R, Langhorst J, et al.: Yoga for depression: a systematic review and meta-analysis. Depress. Anxiety. 2013; 30(11): 1068–1083. Publisher Full Text\n\nSherrington C, Fairhall NJ, Wallbank GK, et al.: Exercise for preventing falls in older people living in the community. Cochrane Database Syst. Rev. 2019; 2019. Publisher Full Text\n\nBrinsley J, Schuch F, Lederman O, et al.: Effects of yoga on depressive symptoms in people with mental disorders: a systematic review and meta-analysis. Br. J. Sports Med. 2021; 55(17): 992–1000. PubMed Abstract \| Publisher Full Text\n\nPratt M, Wieland S, Ahmadzai N, et al.: A scoping review of network meta-analyses assessing the efficacy and safety of complementary and alternative medicine interventions. Syst. Rev. 2020; 9(1): 97. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDonthu N, Kumar S, Mukherjee D, et al.: How to conduct a bibliometric analysis: An overview and guidelines. J. Bus. Res. 2021; 133: 285–296. Publisher Full Text\n\nRowlands I: What are we measuring? Refocusing on some fundamentals in the age of desktop bibliometrics. FEMS Microbiol. Lett. 2018; 365(8). PubMed Abstract \| Publisher Full Text\n\nEllegaard O, Wallin JA: The bibliometric analysis of scholarly production: How great is the impact? Scientometrics. 2015; 105(3): 1809–1831. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGonzález-Alcaide G: Bibliometric studies outside the information science and library science field: uncontainable or uncontrollable? Scientometrics. 2021; 126(8): 6837–6870. Publisher Full Text\n\nPeters MDJ, Godfrey C, McInerney P, et al.:Chapter 11: Scoping Reviews (2020 version).Aromataris E, Munn Z, editors. JBI Manual for Evidence Synthesis. JBI; 2020. Publisher Full Text\n\nNg JY, Dhawan T, Dogadova E, et al.: Operational definition of complementary, alternative, and integrative medicine derived from a systematic search. BMC Complement. Med. Ther. 2022; 22(1): 104. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNg JY, Dhawan T, Dogadova E, et al.: A comprehensive search string informed by an operational definition of complementary, alternative, and integrative medicine for systematic bibliographic database search strategies. BMC Complement. Med. Ther. 2022; 22(1): 200. Publisher Full Text\n\nRethlefsen ML, Kirtley S, Waffenschmidt S, et al.: PRISMA-S: an extension to the PRISMA Statement for Reporting Literature Searches in Systematic Reviews. Syst. Rev. 2021; 10(1): 39. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTricco AC, Lillie E, Zarin W, et al.: PRISMA Extension for Scoping Reviews (PRISMA-ScR): Checklist and Explanation. Ann. Intern. Med. 2018; 169(7): 467–473. Publisher Full Text\n\nBelter CW: Bibliometric indicators: opportunities and limits. J. Med. Libr. Assoc. 2015; 103(4): 219–221. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKumar M, George RJ, Anisha PS: Bibliometric Analysis for Medical Research. Indian J. Psychol. Med. 2022; 02537176221103617.\n\nKoltun V, Hafner D: The h-index is no longer an effective correlate of scientific reputation. PLoS One. 2021; 16(6): e0253397. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFrisch NC, Rabinowitsch D: What's in a Definition? Holistic Nursing, Integrative Health Care, and Integrative Nursing: Report of an Integrated Literature Review. J. Holist. Nurs. 2019; 37(3): 260–272. Publisher Full Text\n\nGupta C, Prakash D: Nutraceuticals for geriatrics. J. Tradit. Complement. Med. 2015; 5(1): 5–14. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nThomson-Casey C, Adams J, McIntyre E: Complementary medicine in psychology practice: an analysis of Australian psychology guidelines and a comparison with other psychology associations from English speaking countries. BMC Complement. Med. Ther. 2022; 22(1): 171. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWang Q: A bibliometric model for identifying emerging research topics. J. Assoc. Inf. Sci. Technol. 2018; 69(2): 290–304. Publisher Full Text\n\nEchchakoui S: Why and how to merge Scopus and Web of Science during bibliometric analysis: the case of sales force literature from 1912 to 2019. J. Mark. Anal. 2020; 8(3): 165–184. Publisher Full Text\n\nMejia C, Wu M, Zhang Y, et al.: Exploring Topics in Bibliometric Research Through Citation Networks and Semantic Analysis. Front. Res. Metr. Anal. 2021; 6: 742311. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCheung F: TCM: Made in China. Nature. 2011; 480(7378): S82–S83. Publisher Full Text\n\nFjaer EL, Landet ER, McNamara CL, et al.: The use of complementary and alternative medicine (CAM) in Europe. BMC Complement. Med. Ther. 2020; 20(1): 108. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNg JY, Liu H, Shah A, et al.:Characteristics of Bibliometric Analyses of the Complementary, Alternative, and Integrative Medicine Literature: A Scoping Review. [dataset].2023, January 19. Publisher Full Text" }	[ { "id": "181379", "date": "14 Aug 2023", "name": "Jenny-Ann Brodin Danell", "expertise": [ "Reviewer Expertise Sociology", "bibliometrics", "medical sociology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThanks for the protocol! The introduction is relevant and ok, both regarding definition of central concepts, relevance of analyzing CAIM literature, and what/how bibliometric methods can be helpful for that.\nHowever, I would prefer a more elaborated motivation of the study. Why is it relevant to analyze the characteristics of bibliometric studies of CAIM? Are there reasons to expect them to be different from other medical/health sub-fields? Are there particular pitfalls? And isn’t it more or less necessary to compare these studies with other areas to be able to say something relevant about the patterns (at least indirectly? I have not checked the details, but there seem to be a few similar scoping reviews on other medical sub-fields.)\nThe search procedure/extraction seems appropriate, as well as the choice of databases. Characteristics/information that will be extracted is also relevant in relation to the scope of the study.\nA minor detail; the formulations about lack of standardized procedures in bibliometric analysis just seem blunt. Bibliometrics is, as you know, a diverse field of research methods and there are never ending discussions about limitations, potential developments (etc.), but there are certainly standardized procedures.\nLooking forward to the results!\n\nIs the rationale for, and objectives of, the study clearly described? Partly\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Not applicable", "responses": [ { "c_id": "10350", "date": "16 Nov 2023", "name": "Jeremy Ng", "role": "Author Response", "response": "We kindly thank the reviewer for their feedback. Regarding our motivations for conducting this study, we are particularly interested in mapping bibliometric analysis studies of CAIM, as, to our knowledge, no systematic or scoping reviews have been completed on this topic. The prevalence of CAIM use is increasing worldwide, and the body of CAIM literature has grown immensely in the past two decades, as we mentioned in our introduction section. Bibliometric analyses of CAIM would capture interesting trends in the field such as the publications over time of different CAIM therapies, or the countries that contribute most to particular research topics. This may be useful to CAIM researchers to identify gaps in the CAIM field and where research is headed. We have stated these points in the last paragraph in the introduction section. Further, analyzing the characteristics of bibliometric analyses of CAIM literature can allow us to identify commonly and less commonly used bibliometric analysis metrics across studies. This could be of interest to inform the development of reporting guidelines for bibliometric analyses. As stated in the introduction section, while there are guidelines for how to conduct bibliometric analyses, there is no consensus in the literature on what should be included. We anticipate that articles may have diverse or unique bibliometric variables, or may even range in the number of variables used. As stated above, this could be useful to improve our understanding of how bibliometric analyses are conducted, and can help to inform future reporting guidelines on this methodology. Regarding your suggestion on comparing characteristics of CAIM bibliometric analyses to other fields, we agree this would be interesting. We have not been able to locate systematic or scoping reviews that report the characteristics of bibliometric analyses (e.g., performance analysis or science mapping techniques used, main findings, health conditions managed) of CAIM literature or of other medical/health sub-fields. Accordingly, it is difficult to ascertain how our findings would compare to other medical/health sub-fields. We would be interested if the reviewer is able to share any identified review articles exploring the characteristics of bibliometric analyses in other medical fields. Regarding the standardized procedures of bibliometrics, we acknowledge that bibliometric analyses can be conducted in diverse ways, and there are some guidelines on the types of metrics that can be used (e.g., Linnenleucke et al., 20191; Donthu et al., 20212). For example, we plan to use the paper by Donthu et al.2 to inform our scoping review. However, there is a lack of consensus in the literature on what baseline information should be required as part of a bibliometric analysis, unlike other research designs (e.g., PRISMA guidelines and JBI methods have been widely adopted for reporting and conduct of systematic reviews). We have rephrased the sentence in the 5th paragraph of the Introduction section to reflect that standardized procedures exist, but that there is no universal standard or consensus in the literature on what a bibliometric analysis should at minimum entail. References Linnenluecke MK, Marrone M, Singh AK. Conducting systematic literature reviews and bibliometric analyses. Aust. J. Manag. 2020;45(2):175-194. doi: 10.1177/0312896219877678 Donthu N, Kumar S, Mukherjee D, Pandey N, Lim WM. How to conduct a bibliometric analysis: An overview and guidelines. J. Bus. Res. 2021;133:285-296. doi: 10.1016/j.jbusres.2021.04.070" } ] }, { "id": "163736", "date": "14 Aug 2023", "name": "Robert J Trager", "expertise": [ "Reviewer Expertise chiropractic", "CAIM" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nOverview I congratulate the authors on an extremely well-written, thorough, and clear protocol for a scoping review of bibliometric studies on CAIM. They adequately justify the rationale for the study and describe how their findings may impact the CAIM literature. The methodology is concise yet descriptive. I also appreciated that open-access files were included on OSF, along with an impressive search strategy. I have no major comments regarding the protocol, and only a couple of minor comments.\nMinor comments\nThe authors note that their study will help identify “what health conditions these CAIMs target,” and in the abstract state “health condition targeted.” While this is certainly OK, I believe that the language could be slightly altered in these sections of text. I expect that the authors may encounter bibliometric studies of CAIM which describe not only the treatment of certain conditions, but also allude to a diagnostic, clinical reasoning, case management process, or the use of CAIM preventative purposes among individuals who are already healthy (e.g., “wellness”). For example, in our bibliometric study of chiropractic case reports, we found an increasing trend in studies describing the diagnosis of vascular disorders. While I am less familiar with other fields, I imagine a similar phenomenon may be noted for CAIM-related professions wherein providers have a broad scope of practice, requisite on diagnosis, such as osteopathy or physical therapy. I think the protocol could be therefore altered slightly to change “target(ed)” to “manage(ed)” or some other language that is broader to reflect more than just treatment, but rather an overall management of the patient.\n\nThe authors describe CAIM as “therapies” throughout the manuscript. I think they have the liberty to describe it this way, yet I would caution them that in some instances the CAIM therapy is distinct from the branch of providers that often use that therapy. For example, in my field of chiropractic, chiropractors often use spinal manipulation, yet chiropractors also perform diagnosis and referral in their management of health conditions and sometimes omit spinal manipulation altogether. One might also consider that a single CAIM therapy could be provided by several types of practitioners. To continue with the above example, osteopaths and physical therapists also use spinal manipulation. The authors could have an a priori method for handling how they categorize CAIM therapies versus practitioner types. However, given that this may be confusing and/or unnecessary to establish before seeing the results, the authors could also describe the categorization of CAIM therapies versus provider types as an iterative process, subject to change if there is overlap between the two.\nComments for clarity\nData extraction – The phrase “We seek to conduct a scoping review” is redundant and could be deleted.\nThis is totally optional as it’s not directly related to the manuscript but may be helpful for other readers – on OSF I could not preview the two Appendix files and had to download them to be able to see them. Consider uploading the files as a PDF in OSF so they can be viewed within an internet browser. However, I recommend also keeping the original Word documents on OSF.\n\nIs the rationale for, and objectives of, the study clearly described? Yes\n\nIs the study design appropriate for the research question? Yes\n\nAre sufficient details of the methods provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Not applicable", "responses": [ { "c_id": "10351", "date": "16 Nov 2023", "name": "Jeremy Ng", "role": "Author Response", "response": "We kindly thank the reviewer for their feedback. Regarding minor comment 1, we agree that the language can be changed from “target(ed)” to “manage(d)” to account for the fact that not all bibliometric analyses may specifically discuss treatment, but rather provide a broad overview of the field in regards to diagnostics, clinical reasoning, or condition management. We have implemented these changes to the wording in the Abstract, 2nd paragraph of Data Extraction, and 1st paragraph of Discussion. Additionally, we will also be extracting main findings, conclusions and limitations of each bibliometric analyses, which is another way that we are able to capture some of the interesting findings the reviewer mentioned that goes beyond the “treatment” of conditions. Regarding minor comment 2 and the terminology of CAIM as a “therapy”, to define the types of complementary, alternative, and integrative therapies (CAIM) eligible for inclusion in our review, we are using a Cochrane Complementary Medicine-adopted operationalized list of CAIMs1, which are described as “CAIM therapies”. For consistency, we are using the same terminology. However, the operationalized list is inclusive of many provider types/branches, including chiropractic and osteopathic fields. The data extraction of “CAIM therapies” will be reported in accordance with terminology used by the operationalized list. We also find the reviewer’s point interesting and acknowledge that different CAIM practices or techniques (e.g., spinal manipulation) may be employed by different types of practitioners. However, we anticipate that not all bibliometric analyses included in our review would include the same types of metrics. Accordingly, not all bibliometric analyses may report the types of providers that are providing these CAIM therapies in the literature. In our data extraction form, we will report whether a paper has included the “types of providers” as a metric in their bibliometric analysis. However, we will unfortunately be unable to extract extensive details on which providers would be providing these therapies. This is for the sake of consistency across all the studies, and is in line with JBI scoping review methods which aims to map and provide a broad overview of the literature. To your point, it would be difficult to devise methods for categorizing CAIM therapies versus provider types as described. Regarding the comments for clarity, we agree that the phrase “We seek to conduct a scoping review” is redundant and have removed it from the protocol. Additionally, we will upload PDF versions of the two Appendix files, alongside the Microsoft Word document files, in Open Science Framework. References Ng JY, Dhawan T, Dogadova E, Taghi-Zada Z, Vacca A, Wieland LS, Moher D. Operational definition of complementary, alternative, and integrative medicine derived from a systematic search. BMC. Complement. Altern. Med. 2022;22(1):104. doi: 10.1186/s12906-022-03556-7" } ] } ]	1	https://f1000research.com/articles/12-164
https://f1000research.com/articles/12-1354/v1	18 Oct 23	{ "type": "Research Article", "title": "Cerebrospinal fluid cytochemical analysis from COVID-19 patients with neurological disorders", "authors": [ "David Quispe-Aranda", "Gloria Cruz-Gonzales", "Víctor Rojas-Zumaran", "Arístides Hurtado-Concha", "William Cruz-Gonzales", "Jeel Moya-Salazar", "Eder Walttuoni-Picón", "David Quispe-Aranda", "Víctor Rojas-Zumaran", "Arístides Hurtado-Concha", "William Cruz-Gonzales", "Jeel Moya-Salazar" ], "abstract": "Background: The COVID-19 pandemic caused by SARS-CoV-2 has affected millions of people around the world. Most cytochemical studies of cerebrospinal fluid (CSF) in patients infected with SARS-CoV-2 have shown abnormal results. The objective of the present investigation was to determine the physical, cytological, and chemical alterations of the CSF cytochemical examination of COVID-19 patients with neurological disorders in Peru. Methods: An observational and cross-sectional study was carried out at the Edgardo Rebagliati Martins Hospital. The study population consisted of 94 CSF samples obtained by lumbar puncture from inpatient patients. Likewise, the paired T-test and one-way ANOVA with the Bonferroni post-hoc test was used to determine the differences in the values of CSF biochemical markers. Results: The most frequent neurological disorders were encephalopathy (43%) and brain tumor (23%). The most relevant physical characteristics were cloudy and reddish fluids in the brain tumor and intracerebral hemorrhage; however, in encephalopathies, transparent-looking liquids were observed. CSF glucose from patients with encephalopathy (30%) and intracerebral hemorrhage (13%) had concentrations >70 mg/dL. Proteins >45mg/dl corresponded to 20% of patients with encephalopathy and 17% of patients with intracerebral hemorrhage and brain tumor. Likewise, no differences were found in glucose concentration between neurological disorders (p>0.05); however, differences in protein concentration were observed (p=0.001). Finally, among the cytological characteristics, it was found that patients with encephalopathy (33%) and brain tumor (20%) presented a leukocyte count <5 cells/ul. Conclusion: These findings suggest that the characteristics of CSF may differ depending on the type of neurological complication experienced by patients with COVID-19.", "keywords": [ "COVID-19", "cytology", "neurological disorders", "leukocytes", "glucose", "protein", "SARS-CoV-2", "Peru" ], "content": "Introduction\n\nThe COVID-19 pandemic caused by SARS-CoV-2 has affected millions of people around the world, resulting in an unprecedented health and economic crisis.1 The virus mainly affects the respiratory system; however, there is increasing evidence to suggest that it can also affect the nervous system, resulting in symptoms and neurological comprlications.2 The interaction between SARS-CoV-2 and the nervous system is complex and not yet fully understood.3\n\nNeurological symptoms associated with COVID-19 range from mild symptoms, such as headache and anosmia, to severe symptoms, such as encephalitis and stroke.4 Additionally, COVID-19 can exacerbate pre-existing neurological conditions, such as Parkinson’s disease and multiple sclerosis.5,6 The mechanisms underlying the neurological effects of COVID-19 are thought to involve direct viral invasion of the nervous system, immune-mediated damage, and hypercoagulability. However, the precise mechanisms and long-term effects of COVID-19 on the nervous system remain unclear.7\n\nRecognizing and managing neurological cases related to COVID-19 presents a significant challenge that requires a comprehensive understanding of the underlying pathophysiological mechanisms.4 COVID-19 has been associated with a wide range of neurological syndromes, including encephalopathies, cerebrovascular events, and neuropathies such as Guillain-Barré Syndrome, which affects the entire neuraxis, including the cerebral vasculature.8 In patients with neurological manifestations related to COVID-19, brain MRI abnormalities such as leptomeningeal enhancement and increased inflammatory markers in the cerebrospinal fluid are frequently observed. Furthermore, acute disseminated encephalomyelitis with hemorrhagic changes is highly prevalent in severe cases of COVID-19.9\n\nMost cytochemical studies of cerebrospinal fluid (CSF) in patients infected with SARS-CoV-2 have shown abnormal results, including elevated leukocyte and protein counts.10 Given the importance of timely management of neurological complications in critically ill patients, it is crucial to assess the impact of cerebrospinal fluid analysis. Wu et al.11 have emphasized that requesting a CSF analysis in a timely manner can significantly improve patient outcomes, but further studies are needed to fully characterize the range of neurological conditions associated with COVID-19 and to identify CSF profiles and biomarkers that may help in the diagnosis of this disease.10\n\nThe objective of this study was to determine the physical, cytological, and chemical alterations of the CSF cytochemical examination of COVID-19 patients with neurological symptoms in Peru.\n\n\nMethods\n\nThis study has followed the guidelines of the Helsinki Guide12 and has been approved by the Ethics Committee of the Edgardo Rebagliati Martins Hospital (N°881-GRPR-ESSALUD-2021). Based on a retrospective approach, the reference records were used to obtain only the data corresponding to the results, excluding any other information from the clinical record (for example, the names of the patients), to preserve effective confidentiality and protection of the participants involved.\n\nThis cross-sectional observational study was conducted during the first outbreak of COVID-19. The study was developed in the Clinical Laboratory of the Emergency Center of Metropolitan Lima (CELIM) of the Edgardo Rebagliati Martins Hospital in Lima, Peru. This hospital belongs to the Social Security (EsSalud) and annually receives 350 thousand patients. During the pandemic, it has become a reference hospital for the care of patients with COVID-19 with isolation centers and 24-hour care. This study followed the guidelines of the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) guide.13\n\nThe study population consisted of 94 CSF samples obtained by lumbar puncture from inpatient patients. The inclusion criteria were samples of patients with COVID-19 diagnosed by polymerase chain reaction (PCR) from patients of both sexes and over 18 years of age with neurological symptoms and who had a complete cytochemical analysis (physical, cytological, and chemical). Samples from pregnant patients, with cancer or HIV were excluded. Only 30 samples met the criteria, which is why they were included in the study (Figure 1).\n\nThe age and sex of the participants were obtained by analyzing the medical records, the SAGER guidelines were considered to report information about sex and gender.14 The CSF cytochemical analysis variable included physical (color and appearance), cytological (cellular characteristics), and chemical (glucose and protein) characteristics. Taking into account the reference values in CSF, for glucose from 40 to 70 mg/dl and for proteins from 15 to 45 mg/dl. The other variable was neurological disorders defined according to clinical neurological guidelines and DSM-V.15,16 CSF samples were sent to the laboratory for cytochemical examination following the recommendations of the Clinical and Laboratory Standards Institute (CLSI) H-56A and C49-A guidelines.17,18 The clinical records were obtained from the digital database in MS-Access (Microsoft, Redmond, US) of the emergency laboratory – CELIM.\n\nThe data analysis has been done in SPSS v24.0 (Armonk, USA). Data analysis was initially descriptive to estimate the frequencies of the continuous variables and the mean and the standard deviation of the categorical variables. We used paired T-test and ANOVA one-way with Bonferroni post-hoc test to determine the differences in the values of biochemical markers of CSF considering the p-value <0.05 and the confidence interval of 95% as the significant. The database containing the original data as it was collected, without having been processed or analyzed is available under Underlying data (Database.sav).19\n\n\nResults\n\nThe age range of the 30 patients with COVID-19 was 22 to 77 years (mean 52.2±14.2 years) and 17 (57%) were male. The most frequent neurological disorders were encephalopathy and brain tumor in 13 (43%) and 7 (23%) patients, respectively. Table 1 describes the demographic and clinical characteristics of the patients with COVID-19, whose cerebrospinal fluid was analyzed.\n\nAmong the most outstanding physical characteristics, 14 (47%) liquids were colorless and 21 (70%) had a transparent appearance, corresponding to encephalopathies as the most frequent neurological complication in both cases (Table 2). Brain tumors and intracerebral hemorrhage were neurological complications that frequently presented a cloudy appearance and reddish color in the macroscopic evaluation.\n\nGlucose concentration in COVID-19 patients with encephalopathy, brain tumor, intracerebral hemorrhage, and others was 70.9±20 mg/dl (95%CI 60 to 81.8), 67.6±38.5 mg/dl (95%CI 39.1 to 96.1), 76.8±42.3 mg/dl (95%CI 43 to 110.7), and 56.5±26.0 mg/dl (95%CI 31 to 82), respectively. Among the biochemical parameters, we found that glucose in the CSF of 9 (30%) patients with encephalopathy had values greater than 70 mg/dl, 4 (13%) patients with brain tumors had glucose concentrations between 40 and 70 mg/d. Finally, only 4 (13%) patients with intracerebral hemorrhage had glucose >70 mg/dl.\n\nProtein concentration in COVID-19 patients with encephalopathy, brain tumor, intracerebral hemorrhage, and others was 68.1±74.1 mg/dl (95%CI 27.8 to 108.3), 80.4±96.3 mg/dl (95%CI 9 to 151.7), 221.5±326.3 mg/dl (95%CI -39.6 to 482.6), and 273±271.6 mg/dl (95%CI 6.8 to 539.2), respectively. Proteins >45 mg/dl corresponded to 6 (20%) patients with encephalopathy and 5 (17%) patients with hemorrhage and brain tumor. No differences were found in glucose concentration between neurological disorders (Figure 2); however, differences were observed in protein concentration (p=0.001).\n\np>0.05, p<0.001 and **p<0.05 (significant).\n\nAmong the cytological characteristics, the leukocyte count <5 cells/ul was found in 10 (33%) patients with encephalopathy and in 6 (20%) patients with brain tumors. Patients with encephalopathy and Intracerebral Hemorrhage had a leukocyte count >5 cells/ul which was significant (p>0.05).\n\n\nDiscussion\n\nThis study demonstrated in a cohort of Peruvian patients with COVID-19 an increase in leukocyte count in patients with encephalopathy or intracerebral hemorrhage. In addition, the brain tumor and intracerebral hemorrhage had very marked cytochemical changes and the protein concentration varied significantly according to the neurological condition.\n\nThis is the first comprehensive Peruvian study describing the physical, cytological, and biochemical characteristics of CSF in COVID-19 patients with neurological complications. Our study addresses a critical gap in understanding the unique characteristics of CSF in this patient population and contributes to advancing the field of neurological health.20 By characterizing the CSF in patients with COVID-19, we provide valuable information that can inform and improve care processes for neurological complications associated with COVID-19. Our study also offers significant implications for laboratory professionals, providing critical information on the biochemical and cytological features of these disorders. This knowledge is essential to recognize the key cellular and biochemical changes observed in the CSF of COVID-19 patients and ensure the quality of clinical laboratory results. By providing a detailed understanding of the biochemical and cytological profile of CSF, we hope to contribute to more effective diagnosis and management of neurological complications in patients with COVID-19.\n\nThe study found that the most frequent neurological disorders were encephalopathy (43%) and brain tumor (23%). Likewise, the age group with the highest frequency of patients was between the ages of 50 to 60 years (40%) with a mean ±SD of 52.2±14.2 years and 57% were male. Some investigations such as the studies by Neumann et al.,4 Poyiadji et al.,21 and Ghosh et al.,22 indicate an association between SARS-CoV-2 infection with encephalopathies as one of the neuropathological manifestations with greater relevance in this disease. A systematic review identified 430 patients with COVID-19 presenting with neurological symptoms, with the majority (56%) having encephalopathy.\n\nIn the present investigation, no statistically significant differences were found in glucose concentration between neurological disorders (p>0.05); however, differences in protein concentration were observed (p=0.001) and the highest levels were observed in patients with intracerebral hemorrhage and brain tumors. An investigation in patients with and without neurological disease found that elevated total protein concentrations were associated with the presence of neurological disease, mental disorders, and central nervous system infections.23 On the other hand, Espindola et al.9 observed that patients with encephalopathy had a mean glucose concentration of 64.5 mg/dl in CSF and total protein of 35.5 mg/dl, finding an elevation of total protein in 29.2% of patients; however, the analysis of these analytes did not present significant differences with other neurological manifestations.\n\nEspindola et al.9 and Keller et al.,24 described a high incidence of acute disseminated encephalomyelitis with hemorrhagic changes in these patients, in this work, Intracerebral Hemorrhage was detected as one of the three main diagnoses, this limits the possibility that a patient without any risk factor could develop the formation and rupture of an aneurysm as described in their work Al Saiegh et al.25\n\nWhen comparing the results with what Jaijakul et al.,26 in their work where they indicate that 25% of patients with central nervous system (CNS) viral infections are typically characterized by lymphocytic or neutrophilic pleocytosis of the cerebrospinal fluid (CSF), we could not determine this because, in most Of the study cases, the COVID-19 patients with neurological alterations did not have more than 5 cells/ul, so the cytomorphological differential study was not performed.\n\nMiller et al.,10 in their work mentioned that the few cytochemical studies of the CSF in patients infected with SARS-CoV-2 are mostly abnormal, results with which this work is not related, because, unlike the present work This author found elevated leukocyte counts, in terms of proteins if we had similarity in the study, finding hyperproteinorrachia in both investigations.\n\nThe study has some limitations. First, because it is a cross-sectional study, it was not possible to establish a causality criterion between the independent variables and the dependent variable. Second, the small sample size is a limitation that may limit the generalizability of the findings; because 30 samples of cerebrospinal fluid were obtained using a non-probability sampling for convenience, rigorously classified according to the eligibility criteria, now although the sample may not be very representative, it must be taken into account that it was carried out in full swing of the COVID-19 pandemic, where studies were carried out day by day so that the scientific community began to detail a wealth of knowledge that is available to date about the virus. In addition, the study does not provide information on the long-term outcomes of the patients. Despite these limitations, the study provides important information on the neurological complications of COVID-19 and highlights the need for further research in this area of cytology applied to neurology.\n\n\nConclusions\n\nThese findings suggest that the characteristics of CSF may differ depending on the type of neurological complication experienced by patients with COVID-19. These differences may be helpful in diagnostic and treatment decisions for these patients. However, more research is needed to fully understand the relationship between CSF characteristics and neurological complications of COVID-19.\n\nEarly recognition, investigation, and treatment of neurological diseases related to COVID-19 are crucial to improve patient outcomes and reduce disease burden. A better understanding of the neurological effects of COVID-19 is essential to optimize patient care and develop effective preventive and therapeutic strategies.", "appendix": "Data availability\n\nFigshare: Cerebrospinal fluid cytochemical analysis from COVID-19 patients with neurological disorders, DOI: https://doi.org/10.6084/m9.figshare.23620983. 19\n\nThis project contains the following underlying data:\n\n- Database.sav 19\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC BY 4.0 Public domain dedication).\n\nFigshare: STROBE checklist for Cerebrospinal fluid cytochemical analysis from COVID-19 patients with neurological disorders: a cross-sectional study’, https://doi.org/10.6084/m9.figshare.23621217.\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nMoya-Salazar J, Cañari B, Zuñiga N, et al.: Deaths, infections, and herd immunity in the COVID-19 pandemic: A comparative study of the pandemic control strategies implemented in Peru and the United Kingdom. Rev. Fac. Med. 2022; 70: e92823. Publisher Full Text\n\nPortela-Sánchez S, Sánchez-Soblechero A, Melgarejo P, et al.: Neurological complications of COVID-19 in hospitalized patients: The registry of a neurology department in the first wave of the pandemic. Eur. J. Neurol. 2021; 28: 3339–3347. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRomoli M, Jelcic I, Bernard-Valnet R, et al.: Infectious Disease Panel of the European Academy of Neurology. A systematic review of neurological manifestations of SARS-CoV-2 infection: the devil is hidden in the details. Eur. J. Neurol. 2020; 27: 1712–1726. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNeumann B, Schmidbauer ML, Dimitriadis K, et al.: Cerebrospinal fluid findings in COVID-19 patients with neurological symptoms. J. Neurol. Sci. 2020; 418: 117090. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGarjani A, Middleton RM, Hunter R, et al.: COVID-19 is associated with new symptoms of multiple sclerosis that are prevented by disease modifying therapies. Mult. Scler. Relat. Disord. 2021; 52: 102939. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLeta V, Boura I, van Wamelen DJ , et al.: COVID-19 and Parkinson’s disease: Acute clinical implications, long-COVID and post-COVID-19 parkinsonism. Int. Rev. Neurobiol. 2022; 165: 63–89. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPaterson RW, Brown RL, Benjamin L, et al.: The emerging spectrum of COVID-19 neurology: clinical, radiological and laboratory findings. Brain. 2020; 143: 3104–3120. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLersy F, Benotmane I, Helms J, et al.: Cerebrospinal Fluid Features in Patients with Coronavirus Disease 2019 and Neurological Manifestations: Correlation with Brain Magnetic Resonance Imaging Findings in 58 Patients. J. Infect. Dis. 2021; 223: 600–609. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEspíndola OM, Brandão CO, Gomes Y, et al.: Cerebrospinal fluid findings in neurological diseases associated with COVID-19 and insights into mechanisms of disease development. Int. J. Infect. Dis. 2021; 102: 155–162. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMiller EH, Namale VS, Kim C, et al.: Cerebrospinal Analysis in Patients With COVID-19. Open Forum Infect. Dis. 2020; 7: 7. Publisher Full Text\n\nWu Y, Xu X, Chen Z, et al.: Nervous system involvement after infection with COVID-19 and other coronaviruses. Brain Behav. Immun. 2020; 87: 18–22. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWorld Medical Association: World Medical Association Declaration of Helsinki: ethical principles for medical research involving human subjects. JAMA. 2013; 310(20): 2191–2194. Publisher Full Text\n\nVandenbroucke JP, von Elm E , Altman DG, et al.: Strengthening the reporting of observational studies in epidemiology (STROBE): Explanation and elaboration. PLoS Med. 2007; 4: e297. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHeidari S, Babor TF, De Castro P, et al.: Sex and Gender Equity in Research: rationale for the SAGER guidelines and recommended use. Res. Integr. Peer Rev. 2016; 1: 1–9.\n\nde Salud M : NTS N° 172-MINSA/2021/DGAIN Norma Técnica de Salud para la atención de salud ambulatoria, quirúrgica electiva, en hospitalización y servicios médicos de apoyo, frente a la pandemia por COVID-19. Lima: MINSA; 2021.\n\nDiagnostic and Statistical Manual of Mental Disorders: DSM-5. 5th ed.American Psychiatric Association; 2013. Publisher Full Text\n\nClinical and Laboratory Standards Institute: Body Fluid Analysis for Cellular Composition; Approved Guideline. CLSI document H56-A. Wayne, PA: Clinical and Laboratory Standards Institute; 2006.\n\nClinical and Laboratory Standards Institute: Analysis of Body Fluids in Clinical Chemistry; Approved Guideline. CLSI document C49-A. Wayne, PA: Clinical and Laboratory Standards Institute; 2007.\n\nWalttuoni-Picón E: Cerebrospinal fluid cytochemical analysis from COVID-19 patients with neurological disorders. [Dataset]. Figshare. 2023. Publisher Full Text\n\nCornejo-Olivas M, Custodio N, Mazzetti P: Salud neurológica en tiempos de COVID. Rev. Neuropsiquiatr. 2020; 83: 69–71. Publisher Full Text\n\nPoyiadji N, Shahin G, Noujaim D, et al.: COVID-19–associated Acute Hemorrhagic Necrotizing Encephalopathy: CT and MRI Features. Radiology. 2020; 296: E119–E120. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGhosh R, Dubey S, Finsterer J, et al.: SARS-CoV-2-Associated Acute Hemorrhagic, Necrotizing Encephalitis (AHNE) Presenting with Cognitive Impairment in a 44-Year-Old Woman without Comorbidities: A Case Report. Am. J. Case Rep. 2020; 21: 21. Publisher Full Text\n\nOrlovska-Waast S, Vogdrup L, Gasse C, et al.: Cerebrospinal fluid test results and associations with subsequent mental disorders, neurological diseases, and CNS infections: A population-based cohort study. Brain Behav. Immun. 2021; 98: 210–218. PubMed Abstract \| Publisher Full Text\n\nKeller E, Brandi G, Winklhofer S, et al.: Large and small cerebral vessel involvement in severe COVID-19: detailed clinical workup of a case series. Stroke. 51: 3719–3722. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAl Saieg F, Ghosh R, Leibold A, et al.: Status of SARS-CoV-2 in cerebrospinal fluid of patients with COVID-19 and stroke. J. Neurol. Neurosurg. Psychiatry. 2020; 91: 846–848. PubMed Abstract \| Publisher Full Text\n\nJaijakul S, Salazar L, Wootton S, et al.: The clinical significance of neutrophilic pleocytosis in cerebrospinal fluid in patients with viral central nervous system infections. Int. J. Infect. Dis. 2017; 59: 77–81. PubMed Abstract \| Publisher Full Text" }	[ { "id": "290326", "date": "18 Jun 2024", "name": "Daniel van Wamelen", "expertise": [ "Reviewer Expertise Movement disorders", "particularly Huntington's and Parkinson's disease", "COVID-19 related neurological disorders", "Neuroimaging." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe authors have determined changes occurring in the cerebrospinal fluid of patients with COVID-19. The study addresses an interesting topic, but I feel there are a few issues around study design and inclusion criteria that would need to be better clarified. In addition, I have several other comments and suggestions.\n\n1. I feel the rationale for performing these analyses should be clarified better. A mere description of changes that occur in the CSF of patients with COVID-19 does not add much to the existing literature in my opinion. Related to this the authors state in the introduction ''...further studies are needed...to identify CSF profiles and biomarkers that may help in the diagnosis of this disease''. What would be the additional benefit of CSF analyses when patients have developed (sub)acute neurological symptoms and have also been diagnosed with COVID-19? 2. In the abstract, the authors suggest that they included 94 participants in whom CSF analyses were performed. However, in the results it becomes clear they only included 30 participants in their final analyses. This should be made explicit. In addition, I wonder why they excluded participants without a diagnosis of COVID-19; would this not make a good comparison group to determine the changes in CSF profile between patients with neurological conditions and COVID-19 and those with neurological conditions without COVID-19? Could the authors clarify the reason for not including a ''control'' group? 3. From figure 1 it becomes clear that the authors excluded patients without cytochemical analyses of the CSF. Could they provide an explanation why these analyses where not performed and what the reasons were for these patients to undergo lumbar puncture? 4. Do the authors feel that the participant cohort might represent only a selection of patients with neurological problems and COVID-19? I.e. could the selection criteria have introduced a bias and skewed the cohort towards those with e.g. uncertainty about neurological diagnosis or inflammatory presentations? 5. The methods are rather brief and would benefit from a more detailed description of procedures and whether analyses of CSF had been standardised. In relation to statistical analyses, I would suggest mentioned whether the data were normally distributed justifying the use of parametric statistical tests such as a t-test. If non-normally distributed, non-parametric tests would be more appropriate. 6. In the results (and abstracts) the authors mention ''brain tumor'' as a common neurological diagnosis. Could they specify what is meant by this and what type of brain tumors were included in this diagnostic category? Moreover, what was the evidence on which they based the conclusion that these tumors were related to COVID-19 to exclude the possibility that these tumors were mere coincidental findings? 7. In the results: ''Brain tumors and intracerebral hemorrhage were neurological complications that frequently presented a cloudy appearance and reddish color in the macroscopic evaluation''. What is the relevance in relation to COVID-19? Would this be any different in patients presenting with the same diagnoses but without COVID-19? 8. Table 1: ''Schizophrenia'' is mentioned as a neurological diagnosis in two cases. How was this diagnosed and what evidence is available to assume these diagnoses were related to COVID-19? 9. In the discussion, the authors mention that ''This study demonstrated in a cohort of Peruvian patients with COVID-19 an increase in leukocyte count in patients with encephalopathy or intracerebral hemorrhage''. However, from the results and figures the increase in leukocyte count in patients with encephalopathy is not present.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "11961", "date": "05 Jul 2024", "name": "Eder Walttuoni Picón", "role": "Author Response", "response": "Please go through the Authors' point by point response respectively to the Reviewer Comments: In the current scientific literature, one mainly finds descriptions and case reports regarding the study of cerebrospinal fluid (CSF) in COVID-19 patients. Within this context, the present study provides a detailed analysis of the physical, chemical, and cytological parameters of CSF in a specific group of patients. At the outset of the research, there were 94 participants who had undergone a lumbar puncture for the corresponding cytological study requested by the attending physician. In the initial phase, 44 samples were excluded due to incomplete or inconsistent data in the physical, chemical, and cellular indicators of the cytological examination, presence of clots, therapeutic lumbar puncture (such as in cases of intrathecal chemotherapy), or studies for lymphoma or other hematologic neoplasms. Additionally, 20 more patients were excluded for lacking a confirmed molecular diagnosis (PCR-RT), resulting in a final cohort of 30 eligible patients for the study. A control group was not included because the research was conducted at a hospital exclusively serving patients diagnosed with COVID-19 based on clinical criteria or molecular tests, where cerebrospinal fluid samples were obtained. Patients were excluded if they lacked complete cerebrospinal fluid (CSF) cytological analysis due to shortages of reagents and supplies in the laboratory. In Peru, as in many countries worldwide, most reagents and supplies used by automated equipment are imported by multinational healthcare companies. During the first wave of the pandemic, we faced shortages of these reagents, resulting in incomplete results in these and other tests. The reason these patients underwent lumbar puncture was due to the attending physician's suspicion of a pathology necessitating cytological examination as part of a rapid and precise evaluation for potential neurological disorders of the central nervous system. Yes, we believe that the cohort of participants represents only a selection of patients with neurological issues and COVID-19. We do not believe there is selection bias. The procedures for cerebrospinal fluid cytological analysis were adapted according to our laboratory's internal procedure manual, following recommendations from the Clinical and Laboratory Standards Institute (CLSI), as outlined in their guidelines CLSI C49A \"Analysis of Body Fluids in Clinical Chemistry\" and CLSI H56 \"Body Fluid Analysis for Cellular Composition\". Regarding statistical analysis, it was noted that the data did not follow a normal distribution. Among the clinical characteristics of the patients included in the study, it was observed that 7 (23%) had a diagnosis of brain tumor, distributed as follows: malignant tumor of the temporal lobe (1), malignant brain tumor (3), malignant brain tumor unspecified (2), and unspecified brain tumor (1). We did not conclude that these tumors were related to COVID-19 but rather that these were patients diagnosed with COVID-19. This aligns with literature indicating that some cancer patients and those undergoing cancer treatment have a higher risk of developing severe forms of COVID-19. Unfortunately, as mentioned earlier, a control group was not included in this study. This prevented comparisons and the assessment of the relevance of the findings in relation to patients who had similar diagnoses but without COVID-19. The diagnoses were obtained from the ESSI system (EsSalud Intelligent Health Service), the information management system for medical records. Access to this system allowed us to view the patients' diagnoses, with the appropriate authorization from the institution's ethics and research committee. Among the cytological characteristics of patients infected with COVID-19, it was observed that 10 (33%) of the patients with encephalopathy and 6 (20%) of the patients with brain tumors had a leukocyte count of less than 5 cells/µL." } ] } ]	1	https://f1000research.com/articles/12-1354
https://f1000research.com/articles/12-1350/v1	17 Oct 23	{ "type": "Research Article", "title": "Umbilical cord mesenchymal stem cell; a potential therapy on reducing intimal hyperplasia in rabbit arteriovenous fistula (AVF) model, analysis the expression of HIF-1a, eNOS, and MMP-2", "authors": [ "Yopie Afriandi Habibie", "Dessy Rakhmawati Emril", "Azharuddin Azharuddin", "Dedy Syahrizal", "Maimun Syukri", "Jufriady Ismy", "Cynthia Retna Sartika", "Yopie Afriandi Habibie", "Azharuddin Azharuddin", "Dedy Syahrizal", "Maimun Syukri", "Jufriady Ismy", "Cynthia Retna Sartika" ], "abstract": "Introduction: AVF is the best option for hemodialysis access, but its patency rate drops after one year. UC-MSCs were used to reduce inflammation and promote vascular tissue repair in AVF rabbit models.\nMethods: In this study, 28 male domestic rabbits (Lepus Domestica) were divided into four groups: KN as a negative control, KP as a positive control with placebo therapy, P1 as the treatment group with in situ UC-MSCs, and P2 as the treatment group with intravenous UC-MSCs. The UC-MSCs dose administered was 1,000,000 cells per kilogram of body weight. After 28 days, all groups of rabbit models with AVF were sacrificed. HIF-1α, eNOS, and MMP-2 levels were measured using ELISA Sandwich methods and analyzed using a one-way ANOVA test followed by post hoc Duncan test.\nResults: The study found significant differences in HIF-1α, eNOS, and MMP-2 levels among the treatment groups. P3 and P4 treatments did not significantly differ in HIF-1α levels, but P3 had a lower average HIF-1α level than P4. The KP group had the highest concentration of eNOS, significantly higher than P1, P2, and KN. ENOs concentration decreased in P1 and P2 and was significantly lower than KP. The level of MMP-2 in AVF rabbits that received intravenous UC-MSCs was significantly higher than that of healthy rabbits (KN), but significantly lower than the AVF rabbit group that received a placebo. The MMP-2 level in AVF rabbits receiving in situ UC-MSCs was significantly lower than in the placebo and intravenous UC-MSC groups.\nConclusion. This study suggests that local delivery of in situ UC-MSCs targeting HIF-1α, eNOS, and MMP-2 levels can effectively reduce intimal hyperplasia (IH) in rabbit models of AVF, potentially preventing early AVF failure and serving as a promising therapy to prevent and reduce IH in AVF.", "keywords": [ "Rabbit models AVF", "Intimal Hyperplasia", "Umbilical Cord Mesenchymal Stem Cells", "HIF-1α", "eNOS", "and MMP-2" ], "content": "Introduction\n\nAn arteriovenous fistula is a medical condition in which an abnormal connection exists between an artery and a vein. This connection causes blood to bypass the capillaries, increasing blood flow and blood pressure in the veins. Intimal hyperplasia is a common complication of AVF, characterized by the thickening of the inner layer of the blood vessel wall.1 Intimal hyperplasia is a major cause of arteriovenous fistula failure, contributing to the constriction of the AVF lumen and impaired outflow tract function. Studies have found that AVF hyperplasia can be attributed to factors such as HIF-1 alpha, MMP-2, and eNOS. A research conducted by Kim and colleagues analyzed the association between AVF hyperplasia and these factors.2\n\nHypoxia-inducible factor-1 alpha (HIF-1α) is a transcription factor that plays a crucial role in cellular responses to low oxygen levels. During hypoxic conditions, HIF-1α stabilizes and binds to other cofactors, activating the hypoxia response element of certain gene promoters.3\n\nEndothelial nitric oxidase synthase (eNOS) is also important in the regulation of vascular tone and blood flow through the production of nitric oxide. Nitric oxide acts as a vasodilator, relaxing the smooth muscles in blood vessel walls and promoting increased blood flow. However, in conditions such as arteriovenous fistula, there is a dysregulation of eNOS activity, leading to impaired nitric oxide production and endothelial dysfunction.4 Matrix metalloprotease-2 (MMP-2) is a zinc-dependent enzyme that plays a crucial role in various biological processes, such as tissue remodelling, wound healing, and immune response regulation. However, increased MMP-2 activity can contribute to the development of intimal hyperplasia, inflammation, and plaque rupture in the context of arteriovenous fistulas.5\n\nMesenchymal stem cell (MSC) therapy involves the use of mesenchymal stem cells to treat vascular injuries, specifically in the context of arteriovenous fistula, and reduce the incidence of intimal hyperplasia. These stem cells have the ability to differentiate into specialized cells and promote tissue regeneration, making them a potential therapeutic option for AVF.6–8 Previous preclinical studies have shown that the topical administration of human adipose-derived MSCs to the adventitial surface of the outflow vein can attenuate the formation of venous neointimal hyperplasia and improve AVF patency.7 While surgery is typically recommended for AVF treatment, MSC therapy offers a promising alternative. Recent research has shown that mesenchymal stem cells can play a beneficial role in reducing intimal hyperplasia and promoting AVF maturation.8\n\nThis study aims to evaluate the effect of UC-MSCs administration on HIF-1α, eNOS, and MMP-2 levels in rabbit models with AVF. Once fistulas are established, aggressive anti-inflammatory therapy is important, but surgery is typically necessary.\n\n\nMethods\n\nAn experimental study using local male rabbits (Lepus Domestica) using a post-test design approach (randomized control trial post-test only design), conducted at Faculty of Veterinary Medicine experimental animal hospital Universitas Syiah Kuala, from January to March 2023. The research group was divided into 4, KN as the negative control group, KP as the positive control group of rabbit models AVF with placebo therapy, P1 as the treatment group of rabbit models AVF by giving in situ UC-MSCs, and P2 as the treatment group of rabbit models AVF by giving intra venous UC-MSCs.\n\nThe sample size was calculated using the sample formula from Federer. The minimum number of samples is rounded up to 6 subjects. The estimated number of subjects who dropped out was 10%, so the number of samples from the above formula was added by 10%; the minimum number of research subjects for each group is 7 samples. The total sample to be examined in this study was 28 subjects. The local male rabbits AVF were randomly divided into treatment or control groups. Ethical clearance approval was given by the Research Ethics Committee Faculty of Veterinary Medicine Universitas Syiah Kuala, date 5 December 2022, No.190/KEPH/XII/.\n\nThe research subjects were local male rabbit model AVF. The inclusion criteria were age 8-12 weeks, weight 2000-2500 grams, no visible anatomical abnormalities, no signs of previous infection, and no other diseases. Exclusion criteria were AVF model rabbits males note, drop out before 2 weeks post AVF period, sick or died since treatment and behavior changed during the study (limp and not agile).\n\nThe surgeon, who specialized in thoracic, cardiac, and vascular surgery, as well as a veterinarian, performed all of the anastomoses. This procedure was conducted at the experimental animal laboratory of the Faculty of Veterinary Medicine at Universitas Syiah Kuala. To ensure proper anesthesia, the animals were given 50 mg/kg of intramuscular ketamine and 5 mg/kg of intramuscular xylazine. The surgeon shaved the incision areas to improve visibility during surgery and disinfected them with povidone-iodine. To access the right carotid communis artery, a vertical incision was made in the neck. A dose of 100 IU/kg of intravenous heparinization was given before the right carotid communis artery was anastomosed end-to-side to the right internal jugular vein. The surgeon completed the anastomosis by suturing with a 7-0 polypropylene suture, one by one, and closed the tissues in an anatomical layer. The left internal jugular vein was left untouched.\n\nThe UC-MSCs were administered at a dosage of 1,000,000 cells per kilogram of body weight. The treatment group of rabbits with AVF received UC-MSCs in situ (P1) and intravenous (P2) injections. The UC-MSCs were obtained in Prodia Stemcell syringes, each containing 2,500,000 cells. The positive control group of rabbits with AVF were given a placebo solution (KP) consisting of 2.5 ml of normal saline solution. The negative control group consisted of healthy male rabbits without AVF procedure. After a period of 28 days, all groups of rabbit models with AVF were sacrificed. All groups of rabbits were evaluated for animal care and monitoring at the experimental animal laboratory, Faculty of Veterinary Medicine, Universitas Syiah Kuala, by a veterinarian.\n\nEuthanasia is performed by slowly injecting an anesthetic dose of 150 mg/kgBW of ketamine and 15 mg/kg of xylazine through the intravenous route while monitoring the heart rate. The heart rate will gradually weaken until no heartbeat is audible, and the pupils will slowly dilate, and the eyes will lose their twinkle. Once this happens, the rabbit is considered dead.\n\nThis study assessed HIF-1α, eNOS and MMP-2 level after 28 days of intervention using ELISA sandwich methods.\n\nMeasurement of HIF-1α concentration in rabbits using a specific ELISA kit for rabbits, namely Rabbit Hypoxia Inducible Factor 1 Alpha (HIF-1α) (Cat. No. BZ-22179294-EB, Bioenzy). For eNOS concentration in rabbits using rabbit-specific ELISA kit Rabbit Endothelial Nitric Oxide (eNOS) (Cat. No. BZ-08172420-EB, Bioenzy), and MMP-2 in rabbits using the rabbit specific ELISA kit Rabbit Matrix Metalloproteinase 2 (Cat. No. BZ-22170591-EB, Bioenzy). The procedure for measuring HIF-1α, eNOS, MMP-2 levels was carried out following the procedure contained in the ELISA kit guide. The absorbance value and concentration of HIF-1α, eNOS, MMP-2 were measured using an ELISA reader at 450 nm wavelength (Biolegend, USA).\n\nHIF-1α, eNOS, MMP-2 level data obtained were tested for normality using the Shapiro-Wilk test and obtained HIF-1α, eNOS, MMP-2 data were distributed normally (homogeneous) (P>0.05). To determine differences in HIF-1α, eNOS, MMP-2 concentrations between treatments, a test of variance (One Way ANOVA) was conducted and continued with Duncan’s post hoc test.\n\n\nResults\n\nAccording to the study, the average level of HIF 1-α in healthy rabbits (KN) was 0.09±0.02 ng/mg. In AVF rabbits with placebo (KP), the level was 1.31±0.45 ng/mg, in AVF rabbits with in situ MSC (P1) it was 0.21±0.08 ng/mg, and in AVF rabbits with intravenous UC-MSC (P2) it was 0.33±0.12 ng/mg. The KN group had the lowest HIF 1-α level, followed by P1 and P2. KP had the highest level. P2 had a 15 times higher HIF 1-α level (1429%) than KN. KP and P1 had 3 times higher (286%) and 2 times higher (148%) levels than KN, respectively.\n\nThe statistical analysis showed a significant difference in the level of HIF 1-α between healthy rabbits and rabbits that received AVF treatment with placebo, in situ UC-MSC, and intravenous UC-MSC (Table 1). The P-value was <0.01.\n\n = Significantly different (P<0.01).\n\nThe mean±standard deviation concentration/level of HIF 1-α in healthy rabbits, AVF, and AVF rabbits with UC-MSC addition (N=7) is presented in Table 1.\n\nThe study showed that rabbits with AVF who received UC-MSC treatment, either in situ or intravenously, had similar levels of HIF 1-α as healthy rabbits (KN) with no significant difference (P>0.05). However, rabbits with AVF who received a placebo had 15 times higher levels of HIF 1-α than healthy rabbits (KN) and the difference was statistically significant (P<0.05).\n\nThese findings indicate that both in situ and intravenous UC-MSC treatments can effectively decrease HIF 1-α levels to the same level as healthy rabbits. Moreover, in situ administration of UC-MSC was more effective at reducing HIF 1-α levels compared to intravenous administration. Refer to Figure 2 for a comparison of the mean (±standard deviation) HIF 1-α levels between treatments.\n\nDifferent superscripts a,b above the histogram indicate significant differences (P<0.05).\n\nThe eNOS levels in healthy rabbits (KN), AVF rabbits with placebo (KP), AVF rabbits with UC-MSC in situ (P1), and AVF rabbits with intra-venous UC-MSC (P2) were recorded as 3.73±0.71 ng/mg, 24.77±4.93 ng/mg, 9.21±4.11 ng/mg and 13.79±3.36 ng/mg respectively. The lowest eNOS level was observed in the healthy rabbit group (KN), while the highest was seen in AVF rabbits with a placebo (KP). The eNOS levels in AVF rabbits with in situ UC-MSC (P1) and intra-venous UC-MSC (P2) were lower than AVF rabbits with placebo (KP), but higher than healthy rabbits (KN). The eNOS level in KP, P1, and P2 was higher by 554.07%, 146.91%, and 269.71%, respectively compared to healthy rabbits (KN).\n\nStatistical analysis showed that there was a significant difference in eNOS levels between healthy rabbits, rabbits with AVF treatment followed by placebo, UC-MSC in situ, and UC-MSC intra venous (Table 5.10). The p-value was found to be less than 0.01.\n\nThe results of further tests (post hoc) showed that the concentration of eNOS in the KN group (healthy rabbits) was significantly lower (P<0.05) compared to other treatment groups (KP, P1, and P2). The highest eNOS concentration was obtained in treatment group 2 (KP) and was significantly higher than P1, P2, and KN (P<0.05). The eNOS concentration in treatment groups P1 and P2 decreased and was significantly lower than that in KP (P<0.05), while the eNOS concentration in P1 was significantly lower than that in P2 (P<0.05). These results show that stem cell treatment can reduce the concentration of eNOS in rabbits experiencing AVF. A comparison of the mean (±standard deviation) eNOS between treatments can be seen in Figure 3.\n\nDifferent superscripts a,b,c d above the histogram indicate significant differences (P<0.05).\n\nWe collected data on the levels of MMP-2 in healthy rabbits (KN), AVF rabbits with placebo (KP), AVF rabbits with UC-MSC in situ (P1), and AVF rabbits with UC-MSC intra vein (P2). The mean levels (± standard deviation) were: KN=8.24±1.08 pg/mg, KP=78.57±8.18 pg/mg, P1=6.98±1.63 pg/mg, and P2=15.84±3.74 pg/mg. The lowest MMP-2 level was found in the P1 group, while the highest was found in the KP group. Compared to the KN group, the MMP-2 level was 15.29% lower in P1, and higher by 853.52% and 92.23% in KP and P2 respectively.\n\nStatistical analysis revealed a significant difference in MMP-2 levels between healthy rabbits and those treated with AVF followed by placebo, UC-MSC in situ, or UC-MSC intra venous (Table 3). The p-value was <0.01, indicating a strong correlation.\n\nThe results of a recent test showed that the MMP-2 level in rabbits with AVF and UC-MSC in situ (P1) was not significantly different from the MMP-2 level in healthy rabbits (KN) (P>0.05). However, the MMP-2 level in AVF rabbits with intravenous UC-MSC was significantly higher than that in healthy rabbits (KN) (<0.05), but much lower than that in the AVF rabbit group with placebo (P<0.05).\n\nAdditionally, the MMP-2 level in AVF rabbits with UC-MSC in situ was significantly lower than the MMP-2 level in AVF rabbits with intravenous UC-MSC (P<0.05). These results suggest that administering UC-MSC in situ is more effective in reducing MMP-2 levels, resulting in no significant difference from healthy rabbits. Figure 4 provides a comparison of the mean (±standard deviation) MMP-2 levels between treatments.\n\nDifferent superscripts a,b,c above the histogram indicate significant differences (P<0.05).\n\n\nDiscussion\n\nIn this research as shown in Table 1 and Figure 2 both in the AVF group, those given placebo therapy had a higher level of HIF-1α compared to those given in situ MSC intervention and intra-venous MSC. The latter groups had HIF-1α levels closer to those of healthy rabbits. UC-MSCs may be useful in reducing HIF-1α levels in AVF-related hypoxia by modulating HIF-1α expression and activity through hypoxic preconditioning, which can enhance their angiogenic properties and contribute to the reduction of HIF-1α levels.9\n\nVenous neointimal hyperplasia, which causes stenosis in arteriovenous grafts and late AV fistulas, has a well-described pathogenesis involving both upstream and downstream events. Initial insults lead to endothelial injury, triggering a cascade of mediators that regulate oxidative stress, endothelial dysfunction, and inflammation. The response to this injury results in the migration of smooth muscle cells from the media to the intima, eventually forming neointimal hyperplasia.10\n\nHIF-1α, also called Hypoxia-Induced Factor 1 Alpha, is a transcription factor that regulates the adaptive response to low oxygen levels (hypoxia) in cells, also play an important transcription factor that responds to a lack of oxygen in tissues. It regulates the expression of genes that are involved in various cellular processes, such as angiogenesis, metabolism, and cell survival. In the context of AVF-related hypoxia, HIF-1α has been found to play a crucial role in vascular remodelling and angiogenesis.11 During hypoxia, HIF-1α is stabilized and translocated to the nucleus, which regulates the expression of genes involved in angiogenesis, cell survival, and inflammation.12 Recent studies have shown that the stabilization of HIF-1α can improve the functions of mesenchymal stem cells, including cell adhesion, migration, and proliferation. Moreover, HIF-1α can enhance the regenerative potential of adipose-derived stem cells under hypoxic conditions by increasing the secretion of vascular endothelial growth factor.13 HIF-1α is a biomarker that has been shown to play a crucial role in preventing intimal hyperplasia in arteriovenous fistula.\n\nOne way in which HIF-1α could be used as a biomarker for preventing intimal hyperplasia is by monitoring its expression levels. Studies have shown that increased expression of HIF-1α occurs early during the maturation of arteriovenous fistulas and that this expression is localized to the venous endothelium. By measuring and monitoring HIF-1α expression, clinicians can potentially identify patients who are at a higher risk of developing intimal hyperplasia. This could allow for early intervention and the implementation of preventative measures to minimize the occurrence and severity of intimal hyperplasia.14\n\nThe mechanism through which umbilical cord mesenchymal stem cells exert their anti-intimal hyperplasia effects is multi-faceted. Firstly, these stem cells have the ability to differentiate into various cell types involved in vascular remodeling and repair.15 For example, they can differentiate into smooth muscle cells, endothelial cells, and fibroblasts, which are crucial for maintaining the structural integrity of the vessel wall. By introducing umbilical cord mesenchymal stem cells into the site of arteriovenous fistula creation, it is possible to enhance when introduced into the vessel wall, umbilical cord mesenchymal stem cells can differentiate into smooth muscle cells, endothelial cells, and fibroblasts, which can promote tissue repair and inhibit the development of intimal hyperplasia.16 Furthermore, umbilical cord mesenchymal stem cells have been found to possess immunomodulatory properties. These cells can modulate the immune response and reduce inflammation, which are important factors in intimal hyperplasia development,17 and it may promote vascular repair and inhibit the development of intimal hyperplasia.18\n\nThe combination of umbilical cord mesenchymal stem cells and HIF-1α can enhance these angiogenic properties and improve the outcomes of arteriovenous fistula creation. Zhang et al. conducted a study that found human umbilical cord mesenchymal stem cells produced exosomes promote angiogenesis through HIF-1α, which is beneficial to fracture healing.19\n\nThe role of umbilical cord mesenchymal stem cells in preventing intimal hyperplasia is a promising area of research. Umbilical cord mesenchymal stem cells have shown potential in the prevention of intimal hyperplasia. In animal models, umbilical cord mesenchymal stem cell transplantation has been shown to reduce intimal hyperplasia formation in arterial prosthetic grafts.20\n\nAccording to the data presented in Table 2 and Figure 3, the placebo group had the highest level of eNOS concentration, which was significantly greater than the healthy rabbit and treatment groups. The concentration of eNOS decreased in the treatment groups and was significantly lower than in the placebo group. Moreover, the concentration of eNOS was much lower in the in situ UC-MSC treatment group than in the intra-venous group. These findings indicate that UC-MSC therapy can effectively decrease the concentration of eNOS in rabbits suffering intimal hyperplasia from AVF. This is somewhat contradictory to existing theories because eNOS is expected to cause vasodilation of blood vessels. When intimal hyperplasia occurs, the vascular endothelium should secrete enough eNOS to prevent stenosis. Therefore, the high eNOS levels in the AVF group with placebo can be compared to the lower levels in the groups that received MSC interventions.\n\n = Significantly different (P<0.01).\n\n** = Significantly different (P<0.01).\n\nEndothelial Nitric Oxide Synthase (eNOS) plays a crucial role in maintaining vascular homeostasis by producing nitric oxide, a key regulator of vascular function. Nitric oxide is involved in vasodilation, platelet aggregation inhibition, and smooth muscle cell proliferation suppression. Studies have shown that MSC therapy can enhance the expression and activity of eNOS in vascular endothelial cells, thus promoting nitric oxide production and potentially improving vascular function. In the context of AVF therapy, enhancing eNOS activity through MSC therapy may have beneficial effects on preventing venous neointimal hyperplasia and improving AVF patency.21\n\nMSC therapy has been shown to increase the production of nitric oxide synthase in endothelial cells. This is likely due to the paracrine effects of MSCs, which can secrete various growth factors and cytokines that promote endothelial cell function, including the production of nitric oxide synthase. Specifically, MSCs have been shown to secrete factors such as vascular endothelial growth factor and hepatocyte growth factor, which can stimulate the expression of eNOS in endothelial cells.22 Furthermore, MSC therapy has been associated with the recruitment and activation of endogenous endothelial progenitor cells, which also contribute to the production of nitric oxide synthase. The specific mechanisms by which MSC therapy influences nitric oxide synthase production in endothelial cells are still being studied. However, it is believed that MSCs exert their effects through paracrine signalling, direct cell-to-cell contact, and modulation of the immune system.23\n\nTo investigate the communication between MSCs and endothelial cells in the context of nitric oxide synthase production, researchers have analyzed the vascular endothelial growth factor release by MSCs in cell culture supernatants. This is because vascular endothelial growth factor is known to stimulate nitric oxide production by eNOS through an increase in cytosolic calcium concentrations, which leads to enhanced vascular permeability and vasodilation.24\n\nIn the context of arteriovenous fistula formation and developing intimal hyperplasia, the enhanced production of endothelial nitric oxide synthase (eNOS) through MSC therapy has the potential to play a crucial role. MSC therapy has shown promising potential in improving the outcomes of arteriovenous fistula formation by increasing eNOS activity. Studies have revealed that MSC therapy can increase the ventricular protein expression of eNOS, an enzyme crucial for proper endothelial function.25 This increase in eNOS expression is associated with improved endothelium-dependent vasodilation and enhanced myocardial neovascularization, which are essential for the maturation and success of arteriovenous fistula formation.26\n\nThe mechanism by which MSC therapy enhances eNOS activity in the context of arteriovenous fistula formation involves several pathways. One such pathway is the recruitment and activation of endothelial progenitor cells by MSC therapy. This recruitment and activation of endothelial progenitor cells leads to increased production of eNOS and subsequent release of nitric oxide, promoting angiogenesis and vessel formation.27\n\nOur recent test results demonstrate that rabbits with AVF and UC-MSC present similar MMP-2 levels to healthy rabbits when the UC-MSC is in situ. However, when AVF rabbits are given UC-MSC intravenously, their MMP-2 levels are higher than healthy rabbits but lower than AVF rabbits who received a placebo. Additionally, AVF rabbits with UC-MSC in situ exhibit significantly lower MMP-2 levels compared to those who received UC-MSC intravenously. These findings suggest that administering UC-MSC in situ is more effective in reducing MMP-2 levels, resulting in no significant difference from healthy rabbits. Figure 4 illustrates a comparison of the mean MMP-2 levels (±standard deviation) between treatments.\n\nThe role of matrix metalloprotease 2 (MMP-2) in developing and maturing arteriovenous fistulae (AVFs) has been studied extensively. Research has shown that increased levels of MMP-2 in the serum of adult human patients are associated with AVF maturation, indicating that MMP-2 plays a crucial role in the remodeling and maturation process of AVFs. Additionally, MMP-2 has been linked to the severity of intimal thickening, a biomarker of vascular remodeling underlying AVF stenosis.28 Recent studies have also examined the correlation between MMP-2 and AVF loss. One study found that MMP-2 levels were associated with AVF loss, suggesting that the protein may play a role in the occurrence of AVF disturbances that lead to AVF loss. However, this study did not consider MMP-2 inhibitors, which have been shown to influence MMP-2 activity. Therefore, more research is needed to understand the relationship between MMP-2 levels and AVF disturbances or surgical trauma of the vessels.29\n\nIt has been observed that surgical procedures and shear stress, both of which are characteristics of AVF creation, can upregulate MMP-2. This finding suggests that MMP-2 may be involved in the pathogenesis of AVF failure. MSC therapy has shown potential in treating AVF by modulating the levels of MMP-2. Further studies are necessary to fully understand the relationship between MMP-2 and AVF disturbances and the potential benefits of MSC therapy in treating AVF.30\n\nRegenerative medicine has found a promising approach in MSC therapy, which makes use of mesenchymal stem cells to repair and regenerate damaged tissues and organs.31 One area where MSC therapy shows potential is in treating vascular injuries, particularly arteriovenous fistulae. These fistulae, which enable hemodialysis treatment in patients with end-stage renal disease, often experience complications such as stenosis and neointimal hyperplasia, leading to access failure and repeated interventions.32 Studies have found that matrix metalloprotease 2 (MMP-2) plays a crucial role in the pathogenesis of AVF failure by promoting vascular remodeling, fibrous connective tissue hyperplasia, and lumen stenosis.33 This has led to investigations on the potential correlation between MSC therapy and MMP-2 levels. Research has shown that surgical procedures and shear stress, both characteristics of AVF creation, upregulate MMP-2 expression. Studies conducted in experimental settings have demonstrated that the administration of tissue matrix metalloproteinase inhibitors or the overexpression of such inhibitors can attenuate vascular remodeling and neointimal hyperplasia in arteriovenous grafts.34\n\nOne potential approach for preventing intimal hyperplasia is the use of umbilical cord mesenchymal stem cells. Umbilical cord mesenchymal stem cells have shown promise in various regenerative medicine applications, including tissue repair and angiogenesis. Studies have demonstrated that umbilical cord mesenchymal stem cells can effectively reduce intimal hyperplasia in animal models.35\n\nThis can enable early intervention and the implementation of preventive measures to reduce the incidence and severity of intimal hyperplasia. For instance, by closely monitoring HIF-1α, eNOS and MMP-2 expression levels in patients undergoing arteriovenous fistula creation, clinicians can identify individuals at higher risk for intimal hyperplasia development and initiate targeted interventions.36\n\nThe limitation of this research is the absence of AVF treatment in animal models at the Experimental Animal Lab, Faculty of Veterinary Medicine, Universitas Syiah Kuala was a limitation in this research. To overcome this, researchers created several AVF animal models using rats and local rabbits.\n\n\nConclusion\n\nOur studies have shown that the use of umbilical cord mesenchymal stem cells can prevent intimal hyperplasia in an arteriovenous fistula by controlling the conditions and decreasing hypoxia. We can measure the effectiveness by using HIF-1α, eNOS and MMP-2 as a biomarker and develop better ways to prevent intimal hyperplasia and promote angiogenesis in AVF. However, more research is needed in humans to understand how HIF-1α, eNOS and MMP-2 and umbilical cord mesenchymal stem cells work together, and to optimize their use in preventing intimal hyperplasia.\n\n\nEthics approval\n\nEthical clearance approval was given by the Research Ethics Committee Faculty of Veterinary Medicine Universitas Syiah Kuala, date 5 December 2022, No.190/KEPH/XII/2022.\n\n\nRequirements\n\nEMAIL OF ALL AUTHORS\n\nYopie Afriandi Habibie: yopie@usk.ac.id\n\nDessy Rakhmawati Emril: dessyemril@usk.ac.id\n\nAzharuddin Azharuddin: azharuddin@usk.ac.id\n\nDedy Syahrizal: dedysyahrizal@usk.ac.id", "appendix": "Data availability\n\nFigshare. Data. DOI: https://doi.org/10.6084/m9.figshare.24062265.v1. 37\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nFigshare. ARRIVE. DOI: https://doi.org/10.6084/m9.figshare.24084894.v1\n\n\nReferences\n\nFouquet O, Blossier JD, Dang Van S, et al.: Author’s reply (in reference to letter to editor proposed by Etem Caliskan, Catherine J. Pachuk, Louis P. Perrault, Maximilian Y Emmert and entitled: preservation solutions to improve graft patency: The devil is in the detail). J. Cardiothorac. Surg. 2021; 16: 14. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHu K, Li Y, Guo Y, et al.: Sex Differences in Arteriovenous Fistula Failure: Insights from Bioinformatics Analysis. J. Cardiovasc. Dev. Dis. 2022; 10. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWu D-H, Liang H, Lu S-N, et al.: miR-124 Suppresses Pancreatic Ductal Adenocarcinoma Growth by Regulating Monocarboxylate Transporter 1-Mediated Cancer Lactate Metabolism. Cell. Physiol. Biochem. 2018; 50: 924–935. 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Publisher Full Text\n\nJayasinghe M, Prathiraja O, Perera PB, et al.: The Role of Mesenchymal Stem Cells in the Treatment of Type 1 Diabetes. Cureus. 2022; 14: e27337. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHuang Y, Suguro R, Hu W, et al.: Nitric oxide and thyroid carcinoma: A review. Front. Endocrinol. 2023; 13. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHoeeg C, Frljak S, Qayyum AA, et al.: Efficacy and Mode of Action of Mesenchymal Stem Cells in Non-Ischemic Dilated Cardiomyopathy: A Systematic Review. Biomedics. 2020; 8. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nShiu Y-T, Jaimes EA: Transcription Factor ETS-1 and Reactive Oxygen Species: Role in Vascular and Renal Injury. Antioxidants. 2018; 7. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEndrinaldi E, Darwin E, Zubir N, et al.: The Effect of Mesenchymal Stem Cell Wharton’s Jelly on Matrix Metalloproteinase-1 and Interleukin-4 Levels in Osteoarthritis Rat Model the Creative Commons Attribution-NonCommercial 4.0 International License (CC BY-NC 4.0). Macedonian. J. Med. Sci. 2019; 7: 529–535.\n\nWu JH, Wang DY, Sheng L, et al.: Human umbilical cord Wharton’s jelly-derived mesenchymal stem cell transplantation could improve diabetic intracavernosal pressure. Asian J. Androl. 2022; 24: 171–175. PubMed Abstract \| Publisher Full Text\n\nWong MY, Tseng Y-H, Huang T-Y, et al.: Molecular Characteristics and Distribution of Virulence Genes among Staphylococcus aureus Complex Isolates Derived from Vascular Access Infections. Can. J. Infect. Dis. Med. Microbiol. 2022; 2022: 1–9. 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Publisher Full Text\n\nFikri E: The Role of Vascular Endothelial Growth Factor as a Predictor of Complicated Appendicitis in Animal Model Oryctolagus cuniculus. Open Access Maced J. Med. Sci. 2020; 8: 261–265. Publisher Full Text\n\nHabibie YA: Data. Dataset. figshare. 2023. Publisher Full Text" }	[ { "id": "233515", "date": "11 Jan 2024", "name": "Roberto I Vazquez-Padron", "expertise": [ "Reviewer Expertise arteriovenous fistula failure" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe study evaluates the benefits of Mesenchymal Stem Cells (MSC) on venous remodeling using rabbit arteriovenous fistulas. The design included three groups of four rabbits that had controls and treatments. The paper intends to establish a correlation between a few molecular markers and the benefits of MSC for fistula maturation. However, the study contains no evidence that MSC promote maturation. In addition, the following limitations ended with the quality of the study:\n\nThe role of intimal hyperplasia in AVF failure has become controversial due to the lack of direct evidence supporting the role of intimal hyperplasia in venous stenosis. Can the author provide evidence that intimal hyperplasia causes AVF failure in humans? Please avoid opinion materials like review papers.\n\nThe description of the experimental groups in material and methods needs to be clarified. Define placebo and control.\n\nHistology data is not presented.\n\nAVF functional data such as blood flow and diameter are not presented.\n\nIs the work clearly and accurately presented and does it cite the current literature? No\n\nIs the study design appropriate and is the work technically sound? No\n\nAre sufficient details of methods and analysis provided to allow replication by others? No\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? No\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [ { "c_id": "11134", "date": "13 Apr 2024", "name": "Yopie Afriandi Habibie", "role": "Author Response", "response": "The study evaluates the benefits of Mesenchymal Stem Cells (MSC) on venous remodeling using rabbit arteriovenous fistulas. The design included three groups of four rabbits that had controls and treatments. The paper intends to establish a correlation between a few molecular markers and the benefits of MSC for fistula maturation. However, the study contains no evidence that MSC promote maturation. In addition, the following limitations ended with the quality of the study: The role of intimal hyperplasia in AVF failure has become controversial due to the lack of direct evidence supporting the role of intimal hyperplasia in venous stenosis. Can the author provide evidence that intimal hyperplasia causes AVF failure in humans? Please avoid opinion materials like review papers. --> Thank you for the very constructive comments. In humans, 30-40% of AVF failures occur due to intimal hyperplasia, which is a condition that causes a narrowing of the draining vein's lumen. When the lumen is narrowed, blood clots are more likely to form inside the vein, which can have a negative impact on the maturation of the AVF. The description of the experimental groups in material and methods needs to be clarified. Define placebo and control --> Thank you for the very constructive comments. For the experimental group, a placebo was administered in the form of the normal saline solution via IV (sham group). The control group did not receive any treatment other than AVF surgery. Histology data is not presented.--> Thank you for your brief comment. Our histopathology data was not presented in this study. Instead, we focused on highlighting the importance of molecular markers in identifying the molecular pathway. The preliminary study of histology data has already been published in another journal. Please follow the provided link to access it. https://www.balimedicaljournal.org/index.php/bmj/article/view/4459/2780 AVF functional data such as blood flow and diameter are not presented. --> Thank you for your brief comment. Our AVF functional data, such as blood flow and diameter, are not presented as well in this study. Instead, we focused on highlighting the importance of molecular markers in identifying the molecular pathway. The data is still undergoing the review process in another journal." } ] }, { "id": "248980", "date": "06 Mar 2024", "name": "Ismail Hadisoebroto Dilogo", "expertise": [ "Reviewer Expertise regenerative medicine" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nGeneral comment:\nThis manuscript discusses the potential therapy of UC-MSC to reduce intimal hyperplasia in rabbit models with AVF. The study's main strength is analyzing HIF-1a, eNOS, and MMP-2 expression. However, it is also expected to analyze the microscopic evaluation of the AVF, which needs to be included in the study. Hence, this manuscript should enhance its discussion to be more interesting. Discussing the difference between in situ and intravenous intervention would be interesting for this study. Abbreviations need to be explained or expanded in the first mention of the text. For example, AVF (arteriovenous fistula) is mentioned in the abstract and the introduction sections. Kindly insert the p-value into Figures 2-4.\nAbstract:\nIn the introduction subsection, please explain why the author chose to analyze HIF-1α, eNOS, and MMP-2 levels as the markers. In the methods subsection, how did the author obtain the HIF-1α, eNOS, and MMP-2 levels? Was it from the blood, plasma, or tissue? In the methods and results subsection, please address which groups are P3 and P4. In the results section, please mention the p-value of the significant results. The significant results on HIF-1 alpha levels should have been mentioned.\nIntroduction:\nPlease add how AVF could create a hypoxic environment and its correlation with increasing HIF-1 alpha.\nMethods:\nFrom which source were the UC-MSCs used in the study? Was it a human stem cell or a rabbit's stem cell? How did the author obtain the HIF-1α, eNOS, and MMP-2 levels? Was it from the blood, plasma, or tissue?\nResults:\nRepeating the results in the tables is unnecessary; kindly make reading more effective. Table 1 and Figure 1 present the same results; it is better to present the data effectively. If the numbers of Table 1 was narrated in the text, the table would be unnecessary. The same cases happened between Table 2, Figure 2, Table 3, and Figure 3.”\nDiscussion:\nThe discussion is well written and comprehensive; however, the mechanism behind the significant difference between the effects after in situ and intravenous intervention needs to be discussed. The author should mention the absence of microscopic evaluation of the AVF after intervention as one of the study's limitations. Other limitations should be explored, for example, the short time of evaluation.\nOverall:\nThis manuscript requires minor revision.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1350
https://f1000research.com/articles/12-1348/v1	17 Oct 23	{ "type": "Research Article", "title": "Relationship between eating disorders and pain levels before and after implantation with personality type ", "authors": [ "Halil Bakkaloğlu" ], "abstract": "Background In this study, it was aimed to examine the relationship between eating disorders and pain levels before and after implantation in dental patients.\n\nMethods A pre-implant and post-implant questionnaire was applied to 223 patients who applied to Cyprus Health and Social Sciences University and underwent implant application. Personal information form, Rezz Eating Disorders Scale and Five Factor Personality Types Scale were applied to the patients in the survey application.\n\nResults Extroversion and neuroticism personality type levels of patients were significantly higher after implant (p<0.05). Pain, eating disorder and other personality type differences between before and after implant were statistically insignificant (p>0.05). Before implant, pain level was positively correlated with eating disorder (r=0.190; p<0.01) and negatively correlated with self-control (r=-0.169; p<0.01). Eating disorder level was positively correlated with pain (r=0.190; p<0.01); negatively correlated with extroversion (r=-0.187; p<0.01) and self-control (r=-0.178; p<0.01). After implant, pain level was negatively correlated with neuroticism (r=-0.140; p<0.05) and openness to experience (r=-0.136; p<0.05). Eating disorder level was negatively correlated with extroversion (r=-0.237; p<0.01), self-control (r=-0.151; p<0.05) and neuroticism (r=-0.187; p<0.01). Extroversion personality type level had significant and negative effect on eating disorder after implant at multivariate level (B=-0.43; p<0.01).\n\nConclusions After the implant, psychological support can be given to improve the extrovert personality structures in order to reduce the eating disorder levels of the patients and to prevent the possibility of eating disorders.", "keywords": [ "Implant", "eating disorders", "pain", "personality types" ], "content": "Introduction\n\nDental implant treatment is an increasingly used method in oral and dental health in recent years. Thanks to dental implants, patients regain the functional part of their oral and dental health that they have lost permanently. Thanks to dental implant treatment, important gains are provided in terms of both the quality of life and aesthetic appearance of the patients, especially in nutrition.1–3 For this reason, patients are using more and more implant options every day and thus regain their oral and dental health.\n\nAnother issue where oral and dental health is important is nutrition. Nutrition begins in the mouth and is activated by the grinding of solid foods by the teeth. Therefore, oral and dental health is of great importance, especially in solid food intake. However, in studies on oral health, there are studies reporting that nutritional disorders are seen in individuals due to problems such as dental caries or tissue in the mouth.4–6 The common point of these studies is that there is a relationship between oral and dental health and eating disorders and those eating disorders affect both physically and psychologically.\n\nEating disorders are not only related to oral and dental health in general, but also have psychological roots. In the absence of a physical problem, individuals’ eating disorders are related to their psychological state and mental health.7–10 Based on the cognitive side of every mental health issue, it is possible to state that the personality structures of individuals are also related to eating disorders. Therefore, personality types, which are indicators of personality structure, have a significant effect on eating disorders, as well as other cognitive factors.\n\nAlthough eating disorders are examined according to various variables and personality structure or mental health parameters in the literature, there are not enough studies related to eating disorders and oral health. Therefore, in this study, it was aimed to examine the relationships between personality types, eating disorders and pain perception levels of patients before and after implantation.\n\n\nMethods\n\nA pre-implant and post-implant questionnaire was applied to 223 patients who applied to Cyprus Health and Social Sciences University and underwent implant application. A personal information form (including gender, age, working status, BMI, eating disorder history, implant history, dental fear and dental care), Rezz Eating Disorders Scale and Five Factor Personality Types Scale were applied to the patients in the survey application. Research questionnaire was applied paper between June 2023 and August 2023. Before the application of the questionnaire, permission was obtained from the Scientific Ethics Committee of Cyprus University of Health and Social Sciences.\n\nA total of 223 patients were voluntarily subjected to the study among reached 248 patients during the research period, between June 2023 and August 2023. Since the research is a pioneer, and there have not been reference study for power analysis, sample size calculation was not suitable for the research. By conducting full census sampling in the relevant period, everyone who had an implant within three months and volunteered to participate in the study was included in the study. Time-based sampling method was used.\n\nThe inclusion criteria of the patients were determined as follows:\n\n– 18 years and over\n\n– Not having a chronic disease that prevents the research,\n\n– Not having a psychiatric or psychological diagnosis,\n\n– Volunteer to participate in the research.\n\nThe scale, which was validated in Turkish by Aydemir et al.,11 measures eating disorder in a five-point Likert type and one dimension. Scoring of the scale is from 1 to 5, and total score shows eating disorder levels of participants. The Cronbach Alpha internal consistency coefficient of the scale was reported as 0.74. In this research, we found Cronbach Alpha as 0.754.\n\nThe scale, which was validated in Turkish by Horzum et al.,12 examines personality types in a five-point Likert type and five dimensions. These are extroversion, agreeableness, self-control, neuroticism, and openness to experience. The Cronbach alpha internal consistency value of the scale was reported as 0.88 for extraversion, 0.81 for agreeableness, 0.90 for self-controlling, 0.85 for neuroticism, and 0.84 for openness to experience. In this research, we found Cronbach Alpha 0.728 for extraversion, 0.779 for agreeableness, 0.818 for self-controlling, 0.815 for neuroticism, and 0.886 for openness to experience.\n\nA high score on the scale, which measures the pain threshold on a scale from 1 to 10, indicates a high level of pain.\n\nIn the study, patients who had implants and met the inclusion criteria given above were included in the study on a voluntary basis, without any preliminary evaluation. Participants were asked to fill out the survey by choosing the answers that best suited them.\n\nEthical approval was taken from Scientific Ethics Committee of Cyprus University of Health and Social Sciences with KSTU/2023/202 grant number on 13.05.2023. Written consent from all participants was taken according to ethical approval.\n\nFrequency analysis was used for description of nominal and ordinal parameters. Means and standard deviations were used for description of scale parameters. Cronbach Alpha was used for reliability analysis. Kolmogorov Smirnov test was used for normality of scale parameters. Since all parameter distributions were not normal, nonparametric tests were used. Wilcoxon Signed Rank test was used for differences between before and after implant. Spearman’s rho correlation and Generalized Linear Model (Logit) was used for relationship analysis.13 SPSS 25.0 for windows was used at 95% Confidence Interval and 0.05 significance level.\n\n\nResults\n\nIn total, 248 patients were eligible for the research between June 2023 and August 2023, and 223 of them were respondents.26 Before implant, voluntary method was used for participation. Similarly, patients were asked to participate after implant by voluntary method. For gender distribution, 49.8% of participants were female and 50.2% were male. According to age distribution, 31.1% had 25 and under, 32.0% had 26-35, 36.9% had 36 and over ages. 61.9% of patients were working. 73.1% had normal, 23.3% had overweight and 3.6 had obese BMI level. 12.1% had eating disorder in the family, 3.6% had implant history, 22.0% had oral and maxillofacial surgery history, 19.7% had dentist fear and 83.4% had regular oral care (Table 1).\n\nDifference analysis results showed that extroversion and neuroticism levels of patients were significantly higher after implant (p<0.05). Pain, eating disorder and other personality type differences between before and after implant were statistically insignificant (p>0.05) (Table 2).\n\na Wilcoxon Signed Rank Test.\n\nBefore implant, pain level was positively correlated with eating disorder (r=0.190; p<0.01) and negatively correlated with self-control (r=-0.169; p<0.01). After implant, pain level was negatively correlated with neuroticism (r=-0.140; p<0.05) and openness to experience (r=-0.136; p<0.05) (Table 3).\n\n* p<0.05.\n\n** p<0.01.\n\nBefore implant, eating disorder level was positively correlated with pain (r=0.190; p<0.01); negatively correlated with extroversion (r=-0.187; p<0.01) and self-control (r=-0.178; p<0.01). After implant, eating disorder level was negatively correlated with extroversion (r=-0.237; p<0.01), self-control (r=-0.151; p<0.05) and neuroticism (r=-0.187; p<0.01) (Table 3).\n\nGeneralized Linear (Logit) Model analysis results showed that only extroversion personality type level had significant and negative effect on eating disorder after implant at multivariate level (B=-0.43; p<0.01) (Table 4). Before Generalized Linear (Logit) Model, correlation between demographic and baseline characteristics of patients were also analyzed. Since their correlations were insignificant (p<0.05), they were not included in the regression model.\n\n\nDiscussion\n\nIn this study, the relationship between the personality types of individuals before and after implantation with pain and eating disorders was examined. For this purpose, the effects of personality types on eating disorders were analyzed before and after implantation on 223 patients, and both unidimensional and multidimensional analyzes of factors that may affect eating disorders were performed.\n\nAlthough implant applications are very important applications for oral and dental health today, clinical studies and research generally focus on concrete properties of implants such as biological properties, osseointegration, and permanence.14–18 Studies on the relationship between eating disorders and personality structures of individuals after dental implants are quite limited. However, the relationship between oral and dental health and eating disorders has been the subject of many studies, and it can be stated that there is both a physical and psychological relationship between oral health and eating disorders. Therefore, it can be argued that personality types that affect the psychological structures of individuals are also effective on eating disorders and pain perception.\n\nIn studies on oral and dental health and eating disorders in the literature, eating disorders are also observed in people with impaired oral health.19–22 Patterson Norrie et al.5 reported in their study that individuals with oral health problems have eating disorders. In another study, Brandt et al.21 reported that the relationship between individuals’ oral health and eating disorders was significant in terms of care, and those with eating disorders were obsessed with oral health. In our study, the differences between pre- and post-implant pain levels and eating disorder levels were not statistically significant. This may be related to the fact that implant applications are now much more effective, in a shorter time, and that patients have a high level of trust in both the surgeon and the institution. Thanks to the physician and institution informing the patient at a sufficient level, the patient is informed about what they will live and as a result, he has less anxiety. This situation positively affects the mental well-being of the patient. Therefore, an increase in the level of eating disorder and pain may not have been observed after the implant. The fact that the study was single-centered may also have an effect on this result. Research findings can be further deepened with larger samples and multicenter studies.\n\nStudies on personality types and the characters of individuals report that the character tends to remain unchanged in general. However, possible changes require trauma, unusual events, or prolonged exposure. On the other hand, personality types, on the other hand, change more easily than the character, while they show the attitude that people take according to the event or situation.23–25 In our study, a statistically significant increase was observed in extraversion and neuroticism levels in patients compared to pre-implantation. However, the difference between other personality types before and after implant was not statistically significant. It is possible to state that the symptomatic personality type scale has an effect on this difference. The Five Factor Personality Scale shows the degrees of these personality types, not the exact personality types of individuals. For this reason, it can be stated that post-implantation individuals do not turn into extroverted and neurotic individuals, only their weight on these personality types increases. Another possible reason for this result is that implant application is a traumatizing phenomenon to some extent. Although today, implant applications are very successful and with minimum pain, as a result, an invasive procedure is performed on individuals with implants. This situation can be partially evaluated as trauma.\n\nIn our study, the meaning and positive effect of pain on pre-implant eating disorder is an expected situation that shows the reliability of the data of the research scales. In addition, extroversion and self-control before implant have a decreasing effect on eating disorders. This situation is also significant in terms of its relationship with mental well-being, especially in terms of self-control. The fact that extroverts are more social and social individuals have a higher level of mental well-being supports this situation.\n\nAfter the implant, extraversion, self-control and neuroticism have a decreasing effect on eating disorder. However, this effect, which is significant in one dimension, gives a result in which only the level of extraversion is significant in multivariate analysis. In other words, when all personality types, pain and personal characteristics are evaluated together, eating disorders are less common in extroverted individuals. This finding shows that as the level of extraversion of individuals increases, the level of post-implant eating disorder development will be lower, and the reverse is also true. In other words, more introverted individuals will be more likely to have an eating disorder.\n\nThe most important limitation of the study is that it was conducted as a single center and the demographic characteristics of the patients were close to each other. More comprehensive findings on the effect of implant application, personality types and pain on eating disorders can be obtained in multicenter studies. Most importantly, since the level of informing patients will differ in different institutions, it may be possible to see the effect of psychological effects and personality structures more clearly. However, the process of getting permission from different institutions in the field of health and collecting data includes very serious and tiring processes.\n\nAnother important limitation of the study is the lack of more original and specific measurement tools for the field of oral and dental health. Today, although scales such as fear of the dentist, trust in the doctor, and fear of tooth extraction are being developed, these are the scales that are mostly associated with children’s fear of the dentist. There is a need for more comprehensive and site-specific scales for adults.\n\nThe most important contribution of the research to the literature is that it is a pioneering study in terms of its subject. Studies and studies in the literature mostly focus on biological data related to implants. This research proposes to focus on psychological factors, which are also important in the field of dental implants. In this respect, the research can be shown among the pioneering studies in the field.\n\nAnother contribution of the research to the literature is that, with a simple session to be given to extroverted and introverted individuals after the implant, it may be possible to pre-treat or prevent eating disorders that may develop in these individuals and may be much more difficult to compensate in the future. In this respect, the research gives useful results in terms of clinical applications and original results in terms of preventive medicine.\n\nAccording to the results of the research, it is possible to prevent eating disorders that are likely to develop after dental implants by increasing the degree of extraversion of individuals or by giving short and effective sessions to introverted individuals. Although it is not absolute that every individual with low extraversion will develop an eating disorder, eating disorders can be prevented it, even infrequently, with partially cost-effective eating disorder informative sessions in a large population. This situation can make a significant contribution both in terms of individuals’ quality of life, public resources and public health, and oral and dental health.", "appendix": "Data availability\n\nOpen Science Framework: eating disorders and pain levels before and after implantation with personality type. https://doi.org/10.17605/OSF.IO/BSV28. 26\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nThanks Kadir YILMAZ for his valuable statistical support.\n\n\nReferences\n\nDuong HY, Roccuzzo A, Stähli A, et al.: Oral health-related quality of life of patients rehabilitated with fixed and removable implant-supported dental prostheses. Periodontology. 2022; 88(1): 201–237. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHo K, Bahammam S, Chen CY, et al.: A cross-sectional survey of patient’s perception and knowledge of dental implants in Japan. Int. J. Implant. Dent. 2022; 8(1): 14. Published 2022 Apr 4. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nElani HW, Starr JR, Da Silva JD, et al.: Trends in Dental Implant Use in the U.S., 1999-2016, and Projections to 2026. J. Dent. Res. 2018; 97(13): 1424–1430. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPresskreischer R, Prado MA, Kuraner SE, et al.: Eating disorders and oral health: a scoping review. J. Eat. Disord. 2023; 11(1): 55. Published 2023 Apr 4. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPatterson-Norrie T, Ramjan L, Sousa MS, et al.: Dietitians’ Experiences of Providing Oral Health Promotion to Clients with an Eating Disorder: A Qualitative Study. Int. J. Environ. Res. Public Health. 2022; 19(21): 14193. Published 2022 Oct 30. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPatterson-Norrie T, Ramjan L, Sousa MS, et al.: Oral health and individuals with a lived experience of an eating disorder: a qualitative study. J. Eat. Disord. 2023; 11(1): 121. Published 2023 Jul 17. Publisher Full Text\n\nHay P, Aouad P, Le A, et al.: Epidemiology of eating disorders: population, prevalence, disease burden and quality of life informing public policy in Australia-a rapid review. J. Eat. Disord. 2023; 11(1): 23. Published 2023 Feb 15. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDavey E, Bennett SD, Bryant-Waugh R, et al.: Low intensity psychological interventions for the treatment of feeding and eating disorders: a systematic review and meta-analysis. J. Eat. Disord. 2023; 11(1): 56. Published 2023 Apr 4. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMiskovic-Wheatley J, Bryant E, Ong SH, et al.: Eating disorder outcomes: findings from a rapid review of over a decade of research. J. Eat. Disord. 2023; 11(1): 85. Published 2023 May 30. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMaher AL, Allen A, Mason J, et al.: Exploring the association between early adaptive schemas and self-reported eating disorder symptomatology. Clin. Psychol. Psychother. 2023; 30(1): 152–165. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAydemir Ö, Köksal B, Yalın Sapmaz Ş, et al.: Reliability and validity of Turkish form of SCOFF Eating Disorders Scale. Anadolu Psikiyatri Derg. 2015; 16(Özel sayı.1): 31–35. Publisher Full Text\n\nHorzum MB, Ayas T, Padır M: Adaptation of Big Five Personality Traits Scale to Turkish Culture. Sakarya University Journal of Education. 7(2): 398–408.\n\nYılmaz K, Turanlı M: A Multi-disciplinary Investigation of Linearization Deviations in Different Regression Models. Asian J. Probab. Stat. 2023; 22(3): 15–19. Publisher Full Text\n\nCoelho PG, Pippenger B, Tovar N, et al.: Effect of Obesity or Metabolic Syndrome and Diabetes on Osseointegration of Dental Implants in a Miniature Swine Model: A Pilot Study. J. Oral Maxillofac. Surg. 2018; 76(8): 1677–1687. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNaemi R, Barikani HR, Shahmoradi L: Dental implant quality registries and databases: A systematic review. J. Educ. Health Promot. 2021; 10: 214. Published 2021 Jun 30. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDas S, Dholam K, Gurav S, et al.: Accentuated osseointegration in osteogenic nanofibrous coated titanium implants. Sci. Rep. 2019; 9(1): 17638. Published 2019 Dec 9. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPandey C, Rokaya D, Bhattarai BP: Contemporary Concepts in Osseointegration of Dental Implants: A Review. Biomed. Res. Int. 2022; 2022: 6170411–6170452. Published 2022 Jun 14. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLiang C, Liu X, Yan Y, et al.: Effectiveness and Mechanisms of Low-Intensity Pulsed Ultrasound on Osseointegration of Dental Implants and Biological Functions of Bone Marrow Mesenchymal Stem Cells. Stem Cells Int. 2022; 2022: 7397316–7397335. Published 2022 Sep 26. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRangé H, Colon P, Godart N, et al.: Eating disorders through the periodontal lens. Periodontology. 2021; 87(1): 17–31. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJohansson AK, Johansson A, Nohlert E, et al.: Eating disorders - knowledge, attitudes, management and clinical experience of Norwegian dentists. BMC Oral Health. 2015; 15(1): 124. Published 2015 Oct 14. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBrandt LMT, Fernandes LHF, Aragão AS, et al.: Relationship between Risk Behavior for Eating Disorders and Dental Caries and Dental Erosion. ScientificWorldJournal. 2017; 2017: 1656417. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHambleton A, Pepin G, Le A, et al.: Psychiatric and medical comorbidities of eating disorders: findings from a rapid review of the literature. J. Eat. Disord. 2022; 10(1): 132. Published 2022 Sep 5. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKerber A, Roth M, Herzberg PY: Personality types revisited-a literature-informed and data-driven approach to an integration of prototypical and dimensional constructs of personality description. PLoS One. 2021; 16(1): e0244849. Published 2021 Jan 7. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChilicka K, Rogowska AM, Szyguła R, et al.: Association between Satisfaction with Life and Personality Types A and D in Young Women with Acne Vulgaris. Int. J. Environ. Res. Public Health. 2020; 17(22): 8524. Published 2020 Nov 17. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYavagal PC, Singla H: Prevalence of dental caries based on personality types of 35-44 years old residents in Davangere city. J. Oral Biol. Craniofac. Res. 2017; 7(1): 32–35. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBakkaloğlu H: eating disorders and pain levels before and after implantation with personality type. [Dataset]. 2023, October 4. Publisher Full Text" }	[ { "id": "252849", "date": "09 Mar 2024", "name": "Lut Tamam", "expertise": [ "Reviewer Expertise psychiatry", "psychopharmacology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe study, conducted on 223 dental implant patients, examines the relationships between personality types, eating disorders, and pain perception levels before and after implantation. The findings reveal an increase in extroversion and neuroticism after implantation and a significant negative correlation between extroversion and post-implant eating disorders. The study suggests that dental implantation may impact personality types and emphasizes the importance of psychological factors in oral health research. It proposes interventions, such as sessions for extroverted and introverted individuals, to potentially prevent post-implant eating disorders and contribute to overall public health. This original study needs significant revisions in many parts of the article. My comments are as follows;\nIn the introduction, the study proposes a link between oral and dental health, eating disorders, and personality types; however, it falls short in clearly defining the research gap and specific study objectives until later in the text. To enhance clarity, the authors should provide a more focused and explicit statement regarding the research goals and the unique contribution of the study to existing literature. The concept of personality types or structures introduced in the introduction requires clarification, including their relevance to the study aims. The authors should provide a clear definition and explanation to ensure a better understanding of these terms in the context of their research. While the authors state that sample size calculation was deemed unsuitable due to the pioneering nature of the study, a post-hoc power analysis could have been conducted to ensure statistical robustness. This information should be added to the relevant section. Additionally, the study procedure section needs more details on participant recruitment and should address potential biases introduced by the voluntary participation method. The inclusion criteria lack explicit discussion on their selection and relevance to the research objectives. The authors should provide details on how these criteria, such as the absence of chronic disorders or psychiatric diseases, were evaluated or confirmed. The statement in the procedure that all cases were included \"without any preliminary evaluation\" contradicts the inclusion and exclusion criteria. The validity and reliability of the measures used in the study are inadequately discussed. Relying mainly on Cronbach Alpha values is insufficient to assess the overall psychometric properties of the scales. The authors should provide information on the psychometric properties of both original and adapted measures, including citations for the original scales. The statistical methods section lacks a detailed rationale for the chosen methods, particularly for the composition of the model and the inclusion of variables in the Generalized Linear Model (Logit). A more comprehensive explanation of the chosen statistical methods is necessary. The discussion suggests personality changes after dental implantation, specifically increased extroversion and neuroticism, without sufficient evidence or references. The authors should soften their statements and explore potential mechanisms behind the observed changes to strengthen the discussion. The discussion disproportionately focuses on extraversion as the significant personality factor affecting post-implant eating disorders, neglecting the exploration of other personality factors. A more balanced discussion considering the study's limitations is essential. The discussion makes broad generalizations, such as suggesting interventions for extroverted and introverted individuals to prevent eating disorders after dental implants. However, these statements lack validation and overlook potential individual variations and the complexity of psychological factors. The authors should revise these sections.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-1348
https://f1000research.com/articles/11-1370/v1	23 Nov 22	{ "type": "Software Tool Article", "title": "Inglämnlagare - a tool for restructuring Swedish site data for statistical analysis", "authors": [ "Daniel Löwenborg", "Filipp Antomonov", "Filipp Antomonov" ], "abstract": "Background: This paper presents a new software tool, Inglämnlagare, developed to be open-source, that restructures information about ancient remains in Sweden for analysis. The background is a new version of the ancient sites database, the Historic Environment Record, curated by the Swedish National Heritage Board, that was launched in 2018 with a new database model that structures the information differently compared to previous versions. Methods: The program, written in Python programming language, has multicore support in order to improve performance for large files and uses regular expressions to extract information about individual features of composite sites. Such features, together with their summed amount, are written as new individual fields to a comma-separated value file. The program is delivered as a source script file that can be executed in any Python environment. Use cases: As an example of use, a case study of exploring graves of rectangular shape found within Sweden is provided. The use case also describes the different steps involved in preparing the data in QGIS to run the program, as well as some methods to efficiently analyse and visualize the output. Conclusions: Inglämnlagare will make more information from the Swedish record of ancient sites accessible for research and can be used to explore different content of the record more efficiently than previously possible. While the tool is written specifically for this dataset it also provides an example of how open-source tools can be used for data wrangling making information designed for a specific purpose, such as online dissemination, appropriate for analysis.", "keywords": [ "GIS", "Landscape Analysis", "Sweden", "Historic Environment Record", "data restructuring", "python", "regular expressions" ], "content": "Introduction\n\nMany fields of archaeological research are revitalised as digital technology enables the use of large volumes of data for analysis (Huggett 2020). Increased access to information from the internet and online databases enables both completely new approaches to interpret prehistory, and also makes it possible to revisit previous results with new computational methodologies to verify or challenge previous theories (Cooper & Green 2016, Gillings, Hacıgüzeller & Lock 2022). However, access to data is not enough, and it is necessary that data structures are appropriate for the analytical tools used. Especially reusing data from external sources, that may have been collected for other purposes, often means that the data will need more or less extensive preparation before analysis (Gupta 2020). Sweden has good access to geodata about archaeological sites since the Swedish National Heritage Board (NHB, Swedish: Riksantikvarieämbetet, RAÄ) has provided a digital record of sites as open access since 2006 when the Archaeological Sites and Monuments Information System (FMIS) were launched (Larsson & Löwenborg 2020). In 2018 this was replaced by the updated new Historic Environment Record (HER, Swedish: Kulturmiljöregistret KMR), which in addition to protected sites also has information about archaeological investigations, reports and to some extent also documentation (Larsson, Smith, Sohlenius & Klafver 2017). HER currently holds about 800 000 sites, with different levels of legal protection. Since the purpose of this database was to record the antiquarian status of each site, the data structure was not primarily designed to facilitate analysis (Löwenborg 2007, Sohlenius 2014). However, the information in the record still holds much data that will be of interest for research purposes, to understand regional differences through spatial analysis and approaches from landscape archaeology.\n\nOne aspect that makes it more difficult to work with the data is that several sites are composites that contain information about individual features that are part of a larger site. Typically, a burial ground has information about how many graves have been observed during the survey and the classification of their external features. This is described as “grave type”, “shape” and “construction details”, as defined in the Lämningstyplista. Some graves have two variables for construction, to describe different aspects, and there are also a few occurrences of a variable type “fyllning” that only has one value: “Rödockrafärgad sand” i.e. a grave that contains red ochre sand. Thus, to describe a grave there can be up to four values plus the number of graves of each type. The most common grave type, the mode value, is “stone settings” (stensättning) with shape “round” (rund) and construction “earth filled” (övertorvad), as seen in Table 1 below. These represent almost 50% of all graves in HER. In this paper, we will present a new open source python program that uses regular expressions to restructure Swedish HER data to a format that is more suitable for statistical analysis. While the program is specifically designed for this particular dataset, the principles for restructuring data would be relevant in other cases where a compact data structure, suitable for easy dissemination of data online, needs to be reorganised for analysis. This will be illustrated with an example of extracting all the graves in Sweden and looking at some patterns in this material. We will also present a use case of how to prepare the data to facilitate analysis.\n\nThe former FMIS system had a different solution for how information about grave features were included in the data compared to the new HER record. In FMIS, information about individual features (graves) that are part of a composite site (burial ground) was in a separate table, called “nil”, that was included with the exported shapefiles. The nil table had an identification key (samsatt_id) that related to the geometries in the shapefile so that information about the individual graves could be related to the sites. The information about features was divided into separate rows and fields, which together provided information about the number of different graves at a burial ground. Since the information was split up into different fields and rows, it was necessary to rebuild the data in the record in order to perform statistical analysis. This could be done in different ways with different software tools (Löwenborg 2007, Ma 2020). With the transition to the HER system, a different solution for presenting the data was used, which make use of the new data format for geodata; GeoPackage (GPKG) developed by the Open Geospatial Consortium. One of the benefits of the GPKG format is the possibility to include long string values in the attribute table. Rather than keeping the information about individual features in a separate table, and in multiple rows and fields, the HER format makes an array of all this information, and includes that in a field in the GPKG dataset. Information about the transition from FMIS to HER, the quality work involved in this, and some information about how data was restructured are described on the NHB website. The new variable with information about features at a site is called “ing_lamn” (Swe: ingående lämningar) and would look like this for the mode grave type described above:\n\n‘X’ here is the number of this particular type of graves at a site. Since a burial ground often has graves of different types and kinds, each type will be represented by a similar string, and burial grounds could thus have a string of considerable length to describe all the types of graves there. An example from HER, L2014:3046, with one of the larger and more complex sites with many different grave types looks like this:\n\nFor graves registered as individual, single graves, the structure is different. These would generally be solitary graves, or in sparse groups, where each grave is recorded as a unique object in the register. These are usually a point but could also have a polygon geometry. For these objects, the information about grave type is in the variable “lamningtyp” (i.e. stensättning) and information about shape and construction details of these, if any, are recorded in the variable “egenskap” (i.e. Form: Rund, Konstruktion: Stenfylld). If no features are used to describe the grave, the “egenskap” variable will be ‘NULL’. There is no number associated with these objects since they all represent one single grave.\n\nHaving all information about burial types collected in one file is a great benefit compared to the previous version with the “nil” layer. The nature of the information where the data is split up into different variables for individual graves, and the complex structure of the “ing_lamn” string, still means that it cannot be analysed statistically without prior remodelling. A fairly simple question as to how many burials there are at a burial ground site means that all the “antal” (number) values would need to be summed up for each site. More complex queries, such as how many stone settings, mounds or other types of graves there are, mean that the “antal” needs to be summed separately. To also account for the different shapes and construction types makes it quite complex to extract the information for anything more than a few individual sites. In order to calculate this for regions, or even at a national level, it will first be necessary to restructure the data completely. The program Inglämnlagare, written by Filipp Antomonov, does this by extracting all different categories of burial types and adding the number of these into separate fields for each type.\n\n\nMethods\n\nIn order for Inglämnlagare (Löwenborg 2022b) to work correctly, the data has to be supplied in a comma-separated value file and include a fid attribute to each object in the database, needed for sorting the data. The program, provided as a single script file Ing.py, is written in Python programming language (RRID:SCR_008394) and has multicore support in order to improve processing performance of large files. The script can be run in any Python environment (including directly in QGIS) or compiled as a standalone executable for Linux, Windows or Macintosh operating systems to be used without an installed Python environment. A general outline of the process of extracting information from HER data using Inglämnlagare is described in the use case below.\n\nTo get the most out of the program Inglämnlagare there are several steps needed both before and after, to ensure reliable results that are suited for further analysis. Here we will present a short use case of the steps involved, working with all graves from HER Lämningar, and extracting a subset for analysis. For preparation of data beforehand, and for analysing the results, we will use the Open-Source GIS (Geographical Information System) program QGIS (RRID:SCR_018507) version 3.22.11. The HER Lämningar is available as open data from the Swedish NHB at their open data portal. To have all the data, it is the GPKG version that should be used, since the Shapefile version is missing the ing_lamn variable. In the version that was used here, downloaded on August 16, 2022, there are 799,272 objects available, divided into 507,188 point features, 77,242 lines and 214,842 polygons. For regional analysis where the individual geometries are not needed, it is recommended to first convert all features to points (centroids) and merge them to one layer so that all analysis can be done on the same dataset. Converting to points might create duplicates from multipart features, which should be removed before analysis.\n\nDepending on the questions at hand, it will usually be relevant to make a selection of the data in order to limit the number of objects before continuing. In this example we are only interested in actual archaeological sites. The register holds other categories too in the variable “antikv_bed”, describing the legal status of each site. In this case, we want ‘Fornlämning’ (ancient remains) and ‘Ingen antikvarisk bedömning’ that includes sites that have been excavated and removed, thus losing their legal protection as ancient sites. We might want to include ‘Möjlig fornlämning’ (possible ancient site) but the remaining categories would not be relevant in this case (see the NHB website for information about the classification of sites).\n\nThe key variable for selecting relevant sites is “lamningtyp” as this holds information about the general category of each site. Full descriptions of these categories are in Lämningstyplistan, available from NHB. This list holds both the formal definition of each category, and also explains how graves can be either as simple categories, where each object is one type of grave, or be part of composite sites, where graves can be included in either burial grounds (Gravfält) or burial and settlement sites (Grav- och boplatsområde). For this example, graves that would not be prehistoric were excluded. The following query was used in QGIS to select all categories of sites in the HER database that would contain graves according to the criteria above:\n\nThis query returned 127,641 objects. Before proceeding, it might be practical to delete fields with data not needed later, such as antiquarian and meta information about data creation. If working with a large selection of sites, it might also be advisable to write XY coordinates to each site in the attribute table. This can simplify the process of bringing the data back into GIS software after analysis, by adding points based on coordinates. Another option would be to join the output of the analysis to the original point layer, but that can be difficult with large volumes of data. Following this, the data can be exported to a CSV file and parsed by the program.\n\nAs an example, below, Inglämnlagare is executed in QGIS under Windows, via the built-in Python console, located in the Plugins menu. Start by defining the variable ing, by entering the path to the script file Ing.py:\n\nAfter replacing the path above with a correct one for the script’s location on the machine, the process is then initiated by entering the following string in the console, as illustrated in Figure 1:\n\nHowever, if the machine, regardless of the operating system, already has an installed Python environment, Inglämnlagare can simply be started in the shell via the command python Ing.py. And in the case of a compiled executable, just by running it. The program begins by asking for an existing input file and a desired output file. If only a filename is entered for the input, the location is assumed to be the one of the program file; in case of the same for the output – that of the input file. A list of all possible combinations of site types and attributes is then created. First, all the combinations of lamningtyp and egenskap are collected. After this, the list continues to be expanded with the information in the variable ing_lamn so that all unique combinations of objects are found. To identify relevant segments of information within the strings, regular expressions are used throughout the program. Regular expressions are a set of powerful functions within computational linguistics for working with patterns in text, to manipulate or extract information. For example, the command \\d+ finds one or more numerical values – from the string below, the value “6” would be returned:\n\nThe same principle is used to identify grave type, shape and construction detail, using the structure of the “ing_lamn” variable. When all possible types of graves have been identified, these are added as new fields, so that the example above would be a new variable called: “Stensättning – Konstruktion: Övertorvad; Form: Rund”. For each row in the file, the number of features (“antal”) is then extracted and written as a value for the object, identified by the fid. For the single graves where no number is provided, the value is set to “1”.\n\n\nResults\n\nThe output of Inglämnlagare is written to a new CSV, with all the input values intact, and with the addition of fields for all possible combinations of values. This can be a very long list, depending on what selection of sites was used as input, and the complexity of these in terms of possible values. Extracting all the graves contained in the HER database gives a total of 507,078 individual graves from 127 641 sites. The graves are classified into 135 different grave types. Stone settings are both the most common type of grave, and are also represented by the most variables for different combinations, with 53 different types in total.\n\nMany graves are of the same type. The top five categories hold 83,5 % of all the graves, and the most common types are presented in Table 1. At the other end of the scale, there are 14 types of graves that are represented by only one object each, such as one: Stensättning – Konstruktion: Övertorvad; Fyllning: Rödockrafärgad sand; Form: Oval. Some of the less frequent types of graves might be interesting to explore closer as they could represent specific burial practices that can inform us of cultural and social prehistoric phenomena.\n\nFor further spatial analysis the data can be imported back into QGIS, and for an efficient workflow there are a few suggested steps to prepare the data still. Including more general composite categories, such as 'Grav- och boplatsområde' as in the example query provided above, means that when all objects in “ing_lamn” will be written as a new variable in a field, the number of fields might become very large, and will probably contain information that is not needed. In this case, the resulting table holds 356 fields, where a large number are features from settlements, such as hearths, post-holes, houses etc. Deleting unwanted fields from a table might be difficult, especially in GIS software, so a faster approach might be to export the result to a new layer, and only include the desired fields in the export. This can be achieved during the setup of the export to a new GPKG, by excluding all fields except those that should be kept for further analysis, making the data more manageable. When all fields that contained non-grave information had been removed, only the 135 variables with fields containing graves remain. As this still is a fairly large number of variables, that might contain more information than needed for the analysis, a useful technique would be to combine the categories of interest into a new field. For example, we might only be interested in graves of a particular shape, but not all the different descriptions of construction of these.\n\nOne example of grave type that might be out of interest for further analysis are rectangular stone settings, which often have been associated with the early phases of Christianity at the end of the Viking period (Therus 2019, 42). If we would single out stone settings of rectangular shape, we would still be left with five categories, depending on the construction types. It might further be of benefit to combine these with other grave types, such as rectangular graves constructed with erected stones. These antiquarian classifications may or may not be relevant to the archaeological questions we have, so it is up to the research questions to define analytical units. Assuming we would like to explore rectangular graves, it might be relevant to include both kinds of rectangular graves for a total of 8,004 individual graves, distributed on 3,578 different sites, listed in Table 2.\n\nThe differences in the construction types are “boulder stones”, “erected stones”, “supported stones”, “no fill”, “stone filled”, “earth-filled”, “other” or without value.\n\nUnless analysis will be made of all the characteristics of the nine different fields containing information about rectangular stone settings and rectangular graves of erected stones, further analysis would be more efficient if all the rectangular graves are added up to a single new field, that has the sum of all graves of this type. A useful function for doing this is the possibility to filter variables by parts of field names in the QGIS Field Calculator as illustrated in the central section of Figure 2.\n\nThis simplifies the identification of fields that should be included for extracting the values to the new field “rectangular graves”. Note that rectangular mounds and cairns were not included here, as they are more likely to represent a different cultural expression. Screenshot from the program QGIS version 3.22.11.\n\nDepending on what software is used for further analysis, it is also important to be aware of how NULL values are handled. Adding up fields that include any NULL in QGIS values would give a NULL output for the total. To avoid this, NULL values could be changed to ‘0’, or as in the example in Figure 2, the SQL expression COALESCE could be used, in order to regard NULL values as 0 for the calculation and add up any real numbers. Summed values such as the new field “Rectangular graves” would make further aggregation into areal units fast and efficient.\n\nFor a small number of sites, it might well be enough to plot them on a map to see the distribution, as in Figure 3. The version of rectangular graves that have been classified as erected stones are only present in the southern part of Sweden, with a few concentrations for example around the island of Öland.\n\nData from NHB/RAÄ Kulturmiljöregister (KMR), 2022-08-16. Map tiles by Stamen Design, under CC BY 3.0. Data by OpenStreetMap, under ODbL.\n\nWith a larger number of sites, it is better to aggregate the results to visualise spatial differences. Here, the choice was made to use watersheds as the unit of spatial aggregation. Watersheds have been suggested as a geographical unit that might have relevance in this area during the Late Iron Age, as a possible natural background to the early medieval hundred district organisation (Löwenborg 2008). Looking at how the rectangular graves are distributed on the watersheds, there is a concentration in the central parts around Lake Mälaren and on Öland, as seen in Figure 4. However, looking at the ratio of rectangular graves compared to the total number of graves within each watershed displays a different pattern, in Figure 5. A few small areas on the coast in Uppland and Blekinge show high concentrations, and also some areas in the far north. Comparing with Figure 3, we can assume that rectangular graves primarily are a coastal phenomenon also in the north, with a few exceptions in the inland. While the substantial differences in the size of the watersheds might be obscuring some relevant patterns, it illustrates the potential in exploring the data in different ways. With the large amount of data that restructuring the original dataset of sites provides, there are great opportunities to explore different patterns that otherwise would be difficult to observe.\n\nData from NHB/RAÄ Kulturmiljöregister (KMR), 2022-08-16 and SMHI, the Swedish Meteorological and Hydrological Institute, Huvudavrinningsområden 2016. Map tiles by Stamen Design, under CC BY 3.0. Data by OpenStreetMap, under ODbL.\n\nData from NHB/RAÄ Kulturmiljöregister (KMR), 2022-08-16 and SMHI, the Swedish Meteorological and Hydrological Institute, Huvudavrinningsområden 2016. Map tiles by Stamen Design, under CC BY 3.0. Data by OpenStreetMap, under ODbL.\n\n\nDiscussion\n\nThe restructuring of data that Inglämnlagare provides makes the full range of information of the HER variable “ing_lamn” available for analysis. While the information in the array variable can be queried and extracted for limited questions already as it is, the possibility to work with quantitative methods on the whole dataset opens new ways of approaching the material. There is much specific information that might be useful here, such as the characterisation of features that are part of settlements, and thus not visible from the site level information by itself. As the origin of the record is the systematics surveys that were carried out in Sweden during the 20th century, sites that have been recorded are primarily those that are visible above ground (Jensen 1997). With half a million graves in the database, this material can lend itself to intra-regional analysis, something that was an influential branch of Swedish archaeology, especially in the 1960s and 1970s (Ambrosiani 1964, Hyenstrand 1974). It is, however, important to remember that there are substantial quality issues with the record. The information was collected over a long time, and policies and guidelines changed considerably during this period. In some parts of Sweden there has been specific rounds of survey in later time, such as within the project Skog & Historia (Hermodsson 2003). This means that some areas have been much more thoroughly surveyed compared to others, and different definitions of what should be recorded mean that there are differences in which sites were recorded in different areas, depending on at what point in time they were surveyed (Jensen 2006). The record is also incomplete since much of the archaeological remains are not visible above ground. When archaeological investigations are carried out there are often new sites found, including graves and burial grounds (Löwenborg 2010). Any analysis of the record must thus take into consideration these limitations.\n\nThe HER record is now updated with the results of excavations at a general level, with information about the features found during excavations, such as graves, hearths, houses etc. However, the full documentation from excavations is still only partially collected, with find lists as a start, but the full documentation with GIS data and other databases is currently not made available in a systematic way. With the ongoing development of a digital archive at the NHB, and the newly founded national infrastructure for digital archaeology in Sweden, SweDigArch, this will probably change in the future. More data will thus be collected and made available for Swedish archaeology in the future. With rich data from excavations, both the volume and complexity of data will increase considerably, and there will be a need for new tools to make the most of this information, to lay out the ever-growing puzzle of information about the past.", "appendix": "Data availability\n\nThe original data used is available as Open Data from the Swedish National Heritage Board from https://pub.raa.se/ (accessed on August 16, 2022).\n\nZenodo: KMR_Gravar_20220816. https://doi.org/10.5281/zenodo.7229877 (Löwenborg 2022a).\n\nThis project contains the following underlying data:\n\n- KMR_Gravar_20220816.gpkg (dataset of graves for all of Sweden, as a subset of the HER data above, and restructured with the Inglämnlagare program).\n\n\nReferences\n\nAmbrosiani B: Fornlämningar och be-byggelse: studier i Attundalands och Södertörns förhistoria. Uppsala:Almqvist & Wiksell;1964.\n\nCooper A, Green C: Embracing the complexities of “Big Data” in archaeology: The case of the English landscape and identities project. J. Archaeol. Method Theory. 2016; 23(1): 271–304. Publisher Full Text\n\nGillings M, Hacıgüzeller P, Lock G: Archaeological spatial analysis: a methodological guide. Abingdon, Oxon:Routledge;2022.\n\nGupta N:Preparing archaeological data for spatial analysis. Archaeological Spatial Analysis: a methodological guide. Routledge;2020; (pp. 17–40).\n\nHermodsson Ö: 2002 års fornminnesinventering i Uppsala län: Tierps kommun, Tegelsmora socken’, Riksantikvarieämbetet, 2003.2003.Reference Source\n\nHuggett J: Is Big Digital Data Different? Towards a New Archaeological Paradigm. J. Field Archaeol. 2020; 45(sup1): S8–S17. Publisher Full Text\n\nHyenstrand Å: Centralbygd – randbygd. Strukturella, ekonomiska och administrativa hu-vudlinjer i mellansvensk yngre järnålder. Stockholm:Almqvist & Wiksell;1974.\n\nJensen OW: Fornlämningsbegreppets historia. En exposé över 400 år. Stockholm:Riksantikvarieämbetet;2006.\n\nJensen R: Fornminnesinventeringen–nuläge och kompletteringsbehov. En riksöversikt. Stockholm:Riksantikvarieämbetet;1997.\n\nLarsson Å, Smith M, Sohlenius R, et al.: Digitising the Archaeological Process at the Swedish National Heritage Board: producing, managing and sharing archaeological information. Internet Archaeol. 2017; 43. Publisher Full Text\n\nLarsson Å, Löwenborg D:The digital future of the past – Research potential with increasingly FAIR archaeological data. Re-imagining Periphery: Archaeology and Text in Northern Europe from Iron Age to Viking and Early Medieval Periods. Oxford, Philadelphia:Oxbow Books;2020; (pp. 61–70). Publisher Full Text\n\nLöwenborg D:Flexibility instead of standards? – How to make digital databases on cultural heritage useable to large audiences – a researchers perspective. Communicating Cultural Heritage in the 21st Century. The Chiron Project and its Research Opportunities. Budapest:Archeolingua;2007; (pp. 13–18).\n\nLöwenborg D: Watersheds as a Method for Reconstructing Regions and Territories in GIS. Digital Discovery: Exploring New Frontiers in Human Heritage. CAA 2006. Computer Applications and Quantitative Methods in Archaeology. Proceedings of the 34th Conference, Fargo, United States, April 2006. Budapest: Archaeolingua;2008; (pp. 128–134).\n\nLöwenborg D: Using Geographically Weighted Regression to Predict Site Representativity. Making History Interactive: CAA 2009. Computer Applications and Quantitative Methods in Archaeology. Proceedings of the 34th Conference, Williamsburg, United States. Oxford: Archaeolopress;2010; (pp. 203–215).\n\nLöwenborg D: KMR_Gravar_20220816. [Data set]. Zenodo. 2022a. Publisher Full Text\n\nLöwenborg: lowenborg/Inglamnlagare: Inglämnlagare (Archaeology). [Source code] Zenodo.2022b. Publisher Full Text\n\nMa Y: Multiple expressions of the wheel cross motif in South Scandinavian rock carvings: case studies of Tanum and Enköping in Sweden. MA dissertation.Uppsala University;2020.\n\nSohlenius R: Förstudie FMIS-processen, Rapport 2014:36. Stockholm:Riksantikvarieämbetet;2014.Reference Source\n\nTherus J: Den yngre järnålderns gravskick i Uppland: Framväxten av den arkeologiska bilden och en materialitet i förändring. Doctoral dissertation.Uppsala University;2019." }	[ { "id": "156392", "date": "13 Mar 2023", "name": "Daniella Vos", "expertise": [ "Reviewer Expertise Digital archaeology" ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThis is a very useful and practical example of how Inglämnlagare can be used to improve the use of an openly accessible dataset in Sweden. However, this is also a very specific case study, and the potential application of this software and approach in other contexts is not addressed (which is OK if this was the original intension, but it might be good to be explicit about the scope of the paper and the use of this software).\nThe title of this article sounds like an introduction to Inglämnlagare. However, I see the focus of this paper more as an example for the application of this tool. In any case, it would be good to include a short description of the software, simply referring to another article describing the tool seems lacking in an article devoted to the use of Inglämnlagare. It might also be a good idea to adjust the title to better match the content of this paper. Being explicit about the aim of this paper, and about the intended use of the tool / methodology, can help the reader understand this study.\nCertain statements and sentences are a bit unclear, it might be worthwhile to carefully re-read everything to ensure that there are no vague descriptions or unclear formulations.\nFor example, the introduction includes the following statement: \"However, access to data is not enough, and it is necessary that data structures are appropriate for the analytical tools used\".\nThis sentence is a bit unclear. Why is access to data not sufficient? In which context, why? Perhaps you could explain more and relate this statement better to the information that will follow (being explicit about the main aim of this paper / approach would also help tie everything together).\nThe discussion does not elaborate a lot about the software and methodology / approach here, which in a way makes sense as this is quite evident and the benefits are pretty straightforward. However, it could have included some information about the choices made throughout the process, alternatives to these, placing this method in a broader context, or addressing the durability (how long do you expect this tool and the site offering the data to remain the same / updated?) and the applicability of the software to other cases.\n\nIs the rationale for developing the new software tool clearly explained? Yes\n\nIs the description of the software tool technically sound? Partly\n\nAre sufficient details of the code, methods and analysis (if applicable) provided to allow replication of the software development and its use by others? Yes\n\nIs sufficient information provided to allow interpretation of the expected output datasets and any results generated using the tool? Yes\n\nAre the conclusions about the tool and its performance adequately supported by the findings presented in the article? Yes", "responses": [ { "c_id": "10385", "date": "17 Oct 2023", "name": "Daniel Löwenborg", "role": "Author Response", "response": "The authors wish to thank the reviewer for valuable comments and suggestions. This is indeed a very specific case study, and to make this clearer we suggest to change the title to include the name of the specific database, HER. We have added an outlook at the end to discuss how the specific case could serve as a more general example. The software has also been described in more detail in the Methods section. Note that this is the only article describing the tool and we are not referring to any other article describing the software, only linking to the software itself and the data used." } ] }, { "id": "178708", "date": "07 Aug 2023", "name": "Francesco Carrer", "expertise": [ "Reviewer Expertise Spatial Analysis", "Digital Archaeology" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe new analytical tool described in this paper is of great relevance for data science in archaeology, as it provides a quick and accurate method to extract information from complex sets of HER data. Although the software is specifically designed for the Swedish HER, its applicability to GPKG formats will extend its use to other digital archives.\n\nOn the other hand, I believe that more technical details should be included in the description of the software and the dataset it was designed for. For example, the organisation of the entries in the Swedish HER might require additional explanation, possibly supported by figure(s) describing the information associated to each entry. This will facilitate the comprehension of the software application.\nThe description of the tool is quite synthetic, and it might be useful to provide some details about the algorithm and its implementation in Python, before moving to the case study.\n\nThe application of the tool and the results produced are well explained and described, but as I mentioned before, too much space is given to the application of the software compared to its description. Under \"Methods\", more space is given to the preparation of the datasets in QGIS than to the description of scope and structure of Ing.py.\nThe conclusions show that the software has great potential, but more technical information is required to foster its use beyond the Swedish HER.\n\nIs the rationale for developing the new software tool clearly explained? Yes\n\nIs the description of the software tool technically sound? Partly\n\nAre sufficient details of the code, methods and analysis (if applicable) provided to allow replication of the software development and its use by others? No\n\nIs sufficient information provided to allow interpretation of the expected output datasets and any results generated using the tool? Yes\n\nAre the conclusions about the tool and its performance adequately supported by the findings presented in the article? Yes", "responses": [ { "c_id": "10386", "date": "17 Oct 2023", "name": "Daniel Löwenborg", "role": "Author Response", "response": "The authors wish to thank the reviewer for valuable comments and suggestions. We have added an outlook at the end to discuss how the specific case could serve as a more general example. The structure of the software in Python has also been described in more detail in the Methods section. We have contacted the NHB to ask about a technical description of the Swedish HER, but they have no publicly available description that we can use, only a submitted article, in Swedish only. There is some information about HER available from the links in the introduction, but unfortunately, this is also mostly in Swedish." } ] } ]	1	https://f1000research.com/articles/11-1370
https://f1000research.com/articles/12-1347/v1	17 Oct 23	{ "type": "Research Article", "title": "A novel homomorphic polynomial public key encapsulation algorithm", "authors": [ "Randy Kuang", "Maria Perepechaenko", "Maria Perepechaenko" ], "abstract": "Background: One of the primary drivers in development of novel quantum-safe cryptography techniques is the ongoing National Institute of Standards and Technology (NIST) Post-Quantum Cryptography (PQC) competition, which aims to identify quantum-safe algorithms for standardization. Although NIST has recently announced candidates to be standardized, the development of novel PQC algorithms remains desirable to address the challenges of quantum computing. Furthermore, to enhance security and improve performance. Methods: This paper introduces a novel public key encapsulation algorithm that incorporates an additional layer of encryption during key construction procedure, through a hidden ring. This encryption involves modular multiplication over the hidden ring using a homomorphism operator that is closed under addition and scalar multiplication. The homomorphic encryption key is comprised of two values - one used to create the hidden ring and the other to form an encryption operator. This homomorphic encryption can be applied to any polynomials during key construction over a finite field with their coefficients considered private. Particularly, the proposed homomorphic encryption operator can be applied to the public key of the Multivariate Public Key Cryptography schemes (MPKC) to hide the structure of its central map construction. Results: This paper presents a new variant of the MPKC with its public key encrypted using the proposed homomorphic operator. This novel scheme is called the Homomorphic Polynomial Public Key (HPPK) algorithm, which simplifies MPKC central map to two multivariate polynomials constructed from polynomial multiplications. The HPPK algorithm employs a single polynomial vector for the plaintext and a multi-variate noise vector associated with the central map. In contrast, in MPKC, a single multivariate vector is created by segmenting the secret plaintext over a small finite field. The HPPK algorithm is Indistinguishability Under Chosen-Plaintext Attack (IND-CPA) secure, and its classical complexity for cracking is exponential in the size of the prime field GF(p).", "keywords": [ "Post-Quantum Cryptography", "PQC", "Public-Key Cryptography", "Multivariate Polynomial Public Key", "MPPK", "Key Encapsulation Mechanism", "KEM", "Multivariate Cryptography" ], "content": "Introduction\n\nIn 1978, Rivest et al. proposed homomorphic encryption, a technique for performing computations on encrypted data without knowledge of the decryption key or procedure.1 This was just one year after filing the patent for Rivest-Shamir-Adleman (RSA) public key cryptography.2 Homomorphic encryption is distinct from the cryptographic algorithms used for secure communication or storage with public key mechanisms like RSA,2 Diffie-Hellman,3 and elliptic curve cryptography,4,5 which are used to establish shared keys for symmetric encryption with algorithms like Advanced Encryption Standard (AES).\n\nHomomorphic encryption can be divided into two categories: partially homomorphic and fully homomorphic. Partially homomorphic encryption supports either multiplicative or additive homomorphic operations. Examples of multiplicatively homomorphic systems include RSA6 and ElGamal cryptosystems,7 while additively homomorphic systems include Goldwasser-Micali,8 Benaloh,9 and Paillier.10 The first milestone for fully homomorphic encryption was achieved by Gentry in 2009 using lattice-based cryptography11 to support both addition and multiplication operators in the encrypted mode. Additionally, Chan proposed a symmetric homomorphic scheme based on an improved Hill Cipher12 in 2009, while Kipnis and Hibshoosh proposed their symmetric homomorphic scheme in 201213 with a randomization function for non-deterministic encryption. Gupta and Sharma proposed their symmetric homomorphic scheme based on linear algebraic computation in 2013.14 More recently, Li et al. proposed a new symmetric homomorphic scheme in 2016, called Li-Scheme, for outsourcing databases.15 Their scheme involves two finite fields: a secret small field Fq and a big public field Fp. It uses modular exponentiation with its secret base s followed by modular multiplication with plaintext message m. Li-Scheme supports both additive and multiplicative operations, making it a fully homomorphic encryption. However, in 2018, Wang et al. performed a cryptoanalysis of the Li-Scheme16 and broke the scheme with certain known plaintext-ciphertext pairs. They further improved their cryptoanalysis in 2019 and successfully recovered the secret key with the ciphertext-only attack using lattice reduction algorithm.17\n\nHomomorphic encryption is a technique that enables performing mathematical operations on encrypted data without decrypting it first, thus providing data privacy. However, it would be interesting to extend this technique from data privacy to public key schemes. Specifically, it would be beneficial to explore cases where the public keys can provide privacy with variables that can take user inputs in a public key scheme. Some research in this area has yielded fruitful results,18,19 but there is still room for further research. Motivated by this need, we introduce a new asymmetric key encapsulation scheme called Homomorphic Polynomial Public Key (HPPK), which uses partial homomorphic encryption to encrypt public keys. The HPPK algorithm leverages multivariate polynomials to not only take advantage of the homomorphic properties of addition and scalar multiplication, but also to allow for the encryption of the input of the encrypting party during the creation of the ciphertext. In other words, the coefficients of the public key polynomial are encrypted using a homomorphic function to ensure that they are not truly public and to hide the structure of the public key. At the same time, treating the variables of the public key polynomials as user input allows for greater flexibility during the encryption process.\n\nThe HPPK cryptosystem possesses two distinct features. Firstly, it employs homomorphic encryption of the public key, which enables the inclusion of user input during ciphertext creation. Secondly, it utilizes a hidden ring. However, HPPK is not the first cryptosystem to employ hidden structure. For instance, Li et al. developed a cryptosystem that utilizes a hidden prime ring.15 Hidden Field Equations (HFE) cryptosystems are another notable example. Several asymmetric multivariate encryption schemes based on HFE have been proposed.20–23 Additionally, various signature schemes based on HFE have been developed.24–28 In HFE, both the private polynomials and the structure they are defined over, a field extension, are concealed using affine transformations.\n\nThe aforementioned algorithms based on HFE fall under the category of quantum-safe algorithms, specifically multivariate quantum-safe algorithms. With the advent of quantum computing, quantum-resistant cryptography has gained significant attention from both academia and industry leaders. The National Institution of Standards and Technology (NIST) played a pivotal role in this arena by initiating the post-quantum cryptography (PQC) standardization process in November 2017.29–31 Recently, the NIST announced the third-round finalists, which include four key encapsulation mechanism schemes (KEM) and three digital signature finalists.29 The four KEM finalists include Classic McEliece, which is code-based32; CRYSTALS-KYBER, which is lattice-based33; NTRU, which is based on lattices34,35; and Saber.36 NIST has selected CRYSTALS-KYBER as the standardized algorithm for KEM. Besides the aforementioned finalists, Multivariate Public Key Cryptosystems (MPKC) are noteworthy for their simplicity and efficiency.37,38,27,28 Algorithms based on multivariate polynomial problems are considered quantum-safe and can also serve as excellent candidates for homomorphic encryption due to the use of multivariate polynomials.\n\nThe framework of MPKC is based on a system of quadratic polynomials, with the public key represented by a central map P:Fpm→Fpℓ, where m variables and ℓ polynomials are used.39 Since its introduction by Matsumoto and Imai in 1988,40 numerous variants of MPKC central map constructions have been proposed, including single field systems and mixed field systems.41 Single field MPKC comprises several triangular systems and the Oil and Vinegar system, which were first introduced by Patarin and Goubin in 1997,42 as well as the unbalanced Oil and Vinegar scheme proposed by Kipnis et al. in 1999.20 Mixed field MPKC refers to the Matsumoto-Imai system40 and the HFE.43 Moreover, Wang et al. proposed a Medium-Field MPKC scheme in 2006,44 which was improved upon in 2008.45 Additionally, Ding and Schmidt proposed Rainbow as a MPKC digital signature scheme in 2005.27\n\nAttacks on MPKC cryptosystems are mainly classified into two categories: algebraic solving attacks and linear algebra attacks. Algebra solving attacks attempt to solve the MPKC multivariate equation system from the public key with ciphertext z1z2…zℓ to recover the pre-image x1x2…xm. Faugére reported his first attack on MPKC in 199946 and in 200247 using Gröber bases (F4), later in 2003 Faugére and Antoine reported their attack on HFE Gröber bases (F5). Ding et al. proposed their new Zhuang-Zi algorithm to solve the multivariate system in 2006.48 In linear algebra attacks, Courtois et al. reported their attack on MPKC using the relinearization technique, aclled XL in 2000.49 The Minrank attack has been successfully applied by Goubin and Courtois on the single field system in 200050 and by Kipnis and Patarin on the mixed field system in 1999.51\n\nA novel type of polynomial public key has recently been proposed by Kuang in 2021,52 based on univariate polynomial multiplications. Additionally, Kuang and Barbeau have further proposed a related public key in 2021,53,54,55 which utilizes multivariate polynomial multiplications with two noise functions to improve public key security against potential attacks. Recently, Kuang, Perepechaenko, and Barbeau have proposed a majorly improved Key Encapsulation Mechanism (KEM),56 where modular multiplication encryption over a hidden ring was applied for both noise functions. Building on top of this previous work,56 this paper investigates the potential benefits of combining a new homomorphic encryption with key construction to further enhance the security of MPPK cryptography for KEM and reduce the cipher size. Furthermore, a digital signature scheme based on the multivariate polynomial public key, or MPPK, has been introduced by Kuang, Perepechaenko, and Barbeau in 2022.57\n\n\nHomomorphic encryption operator designed from a hidden ring\n\nIn contrast to conventional homomorphic cryptography used for data privacy, in this paper we propose homomorphic encryption to be applied to the public key in the framework of multivariate asymmetric cryptography. This will allow for an asymmetric scheme with encrypted public keys. Moreover, by construction, homomorphic encryption allows for user’s input during the ciphertext generation procedure. That is, the ciphertext can be created with the input of the encrypting party, however, the public key used for encryption is itself encrypted using a homomorphic operator. The decrypting party is the only party that has knowledge of the private key associated with the homomorphic encryption operator as well as the asymmetric scheme private key. Essentially, the homomorphic encryption defined in this paper provides a round-trip envelope for a public key encryption.\n\nIn a way, such approach combines three main areas of cryptography, namely, asymmetric cryptography, homomorphic cryptography, and symmetric cryptography with the self-shared key. This phenomenon is illustrated in Figure 1. In the figure, the traditional public key derived from a given asymmetric algorithm is called plain public key (PPK), the homomorphically encrypted PPK is called cipher public key or CPK. The cipher is produced by evaluating the public key polynomial values using a user-selected secret together with the random noise variables. The decryption would perform in two stages: homomorphic decryption and then secret extraction.\n\nWe begin by introducing the homomorphic encryption operator. In order to allow for the user’s input during ciphertext creation, and leverage additive and scalar multiplicative homomorphic features, the homomorphic encryption is applied to polynomials. We discuss the reason for this further. Hence, when introducing the said operator we assume that it will be applied to polynomials.\n\nLet S be a positive integer, and R be a randomly chosen value such that R∈ℤS and gcdRS=1. We propose a homomorphic encryption operator ℰ̂RS, with a secret homomorphic key being a tuple RS. The values S and R are never shared. In its general form, the encryption operator is defined as a multiplicative operation modulo a hidden value S as\n\nSuch operator decrypts the coefficients of the polynomial h. That is, it successfully decrypts the cipher coefficients back to the plain coefficients. More precisely,\n\n• ℰ̂RS is additively homomorphic: if a and b are two plain constants, then ℰ̂RSa+b=Ra+bmodS=Ra+RbmodS=ℰ̂RSa+ℰ̂RSb;\n\n• ℰ̂RS is scalar multiplicatively homomorphic: if a is a plain constant and x is a variable, then ℰ̂RSax=Rax=Rax=ℰ̂RSax.\n\nThus, the operator ℰ̂RS offers partially homomorphic encryption. We leave it to the reader to verify that the same properties hold true for the proposed homomorphic decryption operator ℰ̂R−1S. Note that these homomorphic properties come from linearity, and thus are natural to polynomials. Indeed, polynomials hold additive and scalar multiplicative properties through their coefficients. Moreover, polynomials can be defined and evaluated with coefficients in a field or a ring, different from a field or a ring for variables. We leverage this property, and thus, apply the homomorphic encryption to public key cryptosystems with polynomial public keys.\n\nAs we have previously stated, the proposed homomorphic encryption is applicable to all polynomials over a ring ℤq or finite field Fq characterized by a prime q. In this work, when we refer to polynomials, we imply that the plain polynomials, to be encrypted, are considered modulo q, unless stated otherwise. A generic multivariate polynomial has the following form\n\nAlternatively, let Xj=x1j1⋯xmjm denote the monomials of such polynomial, then\n\n• The monomials Xj are to be computed as Xj=Xjmodq. The values of monomials reduced modulo q are used to compute the value of the polynomial px1…xm.\n\n• The homomorphic secret key value S should satisfy the bit length condition: S2>2p2+L2.\n\nThe first condition ensures that polynomial px1…xm is evaluated as if the monomials Xj are new variables over Fq, Indeed, the operator ℰ̂RS is applied to the polynomial px1…xm in the following way\n\nSuch encrypted polynomial can be computed as\n\nNote that the computed value was not reduced modulo any integer, nor is the arithmetic performed modulo any integer. Thus, the user’s input through monomials Xj remains intact and can be decrypted correctly. Let the plain value of the polynomial with user’s input, that is, if the polynomial was not encrypted with ℰ̂RS, be\n\nTo ensure successful decryption, the second condition must be met. If the size of S is sufficiently large, the values of coefficients and variables remains the same after decryption, and it is possible to recover p̂. Indeed,\n\nLinear polynomials\n\nRecall, that we encrypt the coefficients of the polynomials defined over Fq, which successfully maps polynomials from Fqx1…xm to ℤSx1…xm, leaving x1,…,xm∈Fq. A generic linear multivariate polynomial over a finite field Fq has form\n\nConventionally, in the asymmetric encryption schemes, the public key inherits mathematical logic from the private key, making it vulnerable. Hence, if public key consists of polynomials, we wish to encrypt the coefficients of the said polynomials using homomorphic operator, to hide the mathematical logic. In this case we share the cipher public key, encrypted using homomorphic operator. To ensure that the ciphertext can be still created in the framework of asymmetric public key scheme, the variables in the public key polynomials are used for user’s input. They are not encrypted using homomorphic encryption, but only using the encryption procedure from the asymmetric scheme. Such variable values can consist of the plaintext only, or plaintext and noise used for obscurity.\n\nApplying homomorphic encryption operator to the above linear polynomial, defined in Equation (6), produces a cipher linear polynomial with coefficients in a hidden ring ℤS, and variables in Fq:\n\nWhile the plain coefficients, pj, are encrypted into cipher coefficients, Pj, the cipher polynomial Px1x2…xm can still be evaluated with a set of chosen values r1,…,rm∈Fq to produce value P¯\n\nLet the value p¯=∑i=1mpjrjmodq, be the original ciphertext of the asymmetric scheme. However, it is encrypted into the value P¯ using homomorphic encryption. To recover the plain polynomial value, that is, decrypt the cipher coefficients, into the plain coefficients and evaluate polynomial modulo q, we first apply the homomorphic decryption operator ℰ̂R−1S to get ℰ̂R−1SP¯, and then reduce this value modulo q. More precisely,\n\nIn a framework of asymmetric scheme with homomorphic encryption element, polynomials such as in Equation (6) are associated with plain coefficients, that is, the original public keys. The cipher polynomials have form as in Equation (7), with coefficients being encrypted from the plain public keys, using homomorphic encryption. Such cipher public keys are shared, and the plain public keys are stored securely and never shared. The ciphertext in this combined algorithm is of the form as in Equation (8). The decrypting party first needs to decrypt the ciphertext to nullify the homomorphic encryption of the public key, as shown in Equation (8). Afterwards, the decryption party can perform decryption procedure that corresponds to the given asymmetric scheme.\n\nQuadratic polynomials\n\nMultivariate quadratic polynomials serve as the foundation of Multivariate Public Key Cryptosystem or MPKC.39,58–60 Thus, we want to pay special attention on applications of homomorphic encryption on multivariate quadratic polynomials. A general quadratic multivariate polynomial px1x2…xn over a finite field Fq has the following form\n\nHere, the encrypted coefficients are defined over the hidden ring ℤS, however, all the variables x1,…,xm are still elements of the field Fq. As we have previously mentioned, we refer to the coefficients pij as plain coefficients, and Pij are referred to as cipher coefficients. Similarly, Px1x2…xm and px1x2…xm are referred to as cipher and plain polynomials respectively. While coefficients are encrypted with homomorphic encryption operator, the polynomial Px1x2…xm still accepts user’s input. That is, the cipher polynomial value P¯ can be still calculated with a chosen set of r1,…,rm from the field Fq as follows\n\nNote that the computed value P¯ is an integer. The arithmetic to compute such value was not performed modulo any integer. The plain polynomial values are securely hidden through the hidden ring ℤS. To recover the plain polynomial equation, decryption operator ℰ̂R−1S can be applied to the cipher polynomial value P¯, followed by reduction modq:\n\nThe value p¯ is the plain polynomial value for the chosen values of variables x1,…,xm by the encrypting party.\n\nSimilar to the linear case, the public key of the asymmetric scheme consist of quadratic polynomials of the form (9), to be encrypted using homomorphic encryption operators. The cipher public keys are of the form (10). Such cipher public keys are the ones shared, while the plain public keys are not. The ciphertext in the combined scheme is of the form (13), which needs to be decrypted back to the plain value. For that a homomorphic decryption operator is applied, as in Eq. (14), and the plain ciphertext value is recovered.\n\n\nHomomorphic Polynomial Public Key Cryptosystem\n\nThe Homomorphic Polynomial Public Key Cryptosystem or HPPK for short is a variant of a MPKC scheme with public keys encrypted using homomorphic encryption operator. We feel it is valuable to provide the reader with a brief summary of MPKC to facilitate better understanding of the HPPK scheme.\n\nAn interested reader can find the detail description of MPKC schemes by Ding and Yang.39 In this section, we briefly outline the basic mechanism of MPKC algorithms. The framework mainly consists of ℓ quadratic multivariate polynomials\n\n• Public Key: P¯=T∘P∘S.\n\n• Private Key: TPS.\n\nThe MPKC encryption procedure simply evaluates ℓ polynomials over the field Fq as\n\nThe major step to use MPKC is to construct the invertible central map P over a finite field Fq to perform a map: Fqm→Fqℓ.\n\nThere may be a potential way to enhance the security of MPKC cryptosystem by applying the proposed homomorphic encryption on its map: Fqm→Fqℓ. The homomorphic encryption effectively hides the public key construction logic over a hidden ring ℤS. In this case, an encryption key Rk is required for each quadratic polynomial pkx1…xm, with value Rk chosen over the hidden ring ℤS for all k. Hence, there are a total of ℓ encryption keys for MPKC. The MPKC encryption in this case is almost the same as the original MPKC encryption. The ciphertext z1z2…zℓ is to be homomorphically decrypted to create original multivariate equation system, as illustrated in Equation (18). This means, Equation (18) is hidden under the hidden ring ℤS. On one hand, applying the homomorphic encryption would increase the public key size for MPKC, however, the number of variables can be reduced due to the homomorphic encryption.\n\nIn this paper, we are not going to further explore this variant of MPKC schemes but we will focus on another variant of MPKC, called HPPK which we propose in the new section.\n\nWe propose a new variant of an MPKC scheme, called the Homomorphic Polynomial Public Key or HPPK, with the following considerations:\n\n• The vector on the left hand side of the map P is treated as x→l and the vector on the right hand side as x→r;\n\n• The vector x→l is replaced with x→l=x0x1x2…xn, considering x→l as a message vector in a polynomial vector space represented by a basis x0x1x2…xn for a message variable x and x→r=x1…xm as a noise vector for noise variables x1,…,xm;\n\n• The proposed homomorphic encryption is applied to the central map P, mapping the elements from Fq→ℤS: P¯=ℰ̂RSP\n\nand the decryption is de-mapping from ℤS→Fq: P=ℰ̂R−1SP¯modq\n\n• The number of polynomials is reduced to ℓ=2;\n\n• The decryption mechanism is changed from inverting maps to modular division, which automatically cancels the noise used for obscurity.\n\nKey construction\n\nWithout loss of generality, we change the notation of the unencrypted central map to P. Under the above considerations, the central map P consists of two multivariate polynomials\n\nNote that the matrix maps P1 and P2 are of size n+1×m, thus, no longer square. The construction of p1xx1…xm and p2xx1…xm can alternatively be achieved with polynomial multiplications\n\nHere, nb and λ are orders of base multivariate polynomial and univariate polynomials with respect to message variable x respectively. Without loss of generality, we assume that the univariate polynomials f1x and f2x are solvable, in other words λ<5. Using Equations (21) and (22), we can express\n\nWe set public key to be the cipher central map P, while private key consists of the homomorphic operators, the hidden ring, together with univariate polynomials:\n\nSecurity parameters: nb,λ, and the prime finite field Fq which is agreed on before the key generation procedure.\n\nPrivate key:\n\n• hidden ring ℤS with a randomly selected S for the required bit length;\n\n• homomorphic encryption key values R1 and R2 chosen from ℤS;\n\n• univariate polynomials f1x and f2x with coefficients randomly selected from Fq;\n\nPublic key: the map P, consisting of\n\n• polynomial P1x→lx→r=x→l⋅P1⋅x→r\n\n• polynomial P2x→lx→r=x→l⋅P2⋅x→r\n\nEncryption\n\nEncryption is straightforward, by determining the value for the secret x and randomly choosing values for the noise variables x1,…,xm over the field Fq and evaluating ciphertext integer values P¯1 and P¯2. That is, the ciphertext consists of two integer values C=P¯1P¯2, where\n\nHere, Pij1 and Pij2 denote the cipher coefficients encrypted with the homomorphic encryption operators. Note that the cipher polynomials have coefficients in the hidden ring ℤS, and all monomial calculations are performed modq, the rest of the arithmetic is performed over integers. The values P¯1, and P¯2 are integers forming the ciphertext C=P1P2.\n\nDecryption\n\nIt is easy to verify that the HPPK map as in Equation (19) and Equation (20), under construction as shown in Equation (21), holds a division invariant property on the multiplicand or the base multivariate polynomial Bxx1x2…xm. Indeed,\n\nThe first step in the decryption process is to apply the homomorphic decryption operator to the ciphertext to recover plain polynomial values p¯1 and p¯2, which are evaluation results of plain multivariate polynomials p1xx1…xm and p2xx1…xm at the chosen message and noises respectively. This can be done as\n\nThese values are used to compute the ratio K modulo q of the form\n\nNote that the noise vector x→r is automatically eliminated through the division. The secret x can then be found from Equation (27) by radicals if f1x and f2x are solvable such as linear or quadratic polynomials. The optimal choice of λ is 1 in the framework of the HPPK algorithm. This division invariant property is the foundation for the HPPK encapsulation to be indistinguishable under chosen plaintext attacks.\n\n\nA toy example\n\nWe demonstrate how HPPK works with a toy example.\n\nKey pair generation\n\nConsidering a prime field F13 with the prime q=13 and two noise variables x1,x2 for the simplicity of the demonstration purpose only, we can choose the hidden ring characterized by an integer of length >12 bits. The private key consists of the following values:\n\n• S=6798,R1=4267,R2=6475\n\n• f1x=4+9x\n\n• f2x=10+7x\n\n• Bxx1x2=8+7xx1+5+11xx2 (note: just for key pair construction procedure; this polynomial is not stored in the memory)\n\nThe PPK is simply constructed as\n\nThe PPK polynomials are encrypted with the self-shared key R1,R2 over the ring ℤS\n\nEncryption\n\nWe randomly choose variables from F13: x=8,x1=3,x2=6. We, then, pre-calculate values\n\nNow we can calculate the ciphertext C=198082192229 as follows\n\nDecryption\n\nWe first perform the homomorphic decryption to rebuild the plain polynomial equations\n\n\nHPPK security analysis\n\nIn this section, we analyze the security of the proposed HPPK algorithm. The security of HPPK relies on the computational hardness of the Modular Diophantine Equation, introduced in Definition 0.1, and Hilbert’s tenth Problem, introduced in Definition 0.7. We begin by proving that HPPK satisfies the IND-CPA indistinguishability property. These results are then extended to prove that the task of recovering plaintext from the ciphertext in the framework of HPPK is NP-complete, and state its classical and quantum complexity. Afterward, we focus on the private key attack and prove that the problem of obtaining the private key from the public key is NP-complete. Here we also provide classical and quantum complexities of obtaining private key from the public key.\n\nAn attentive reader will notice that the evaluated ciphertext as illustrated in Equation (26) has not been reduced modulo any integer. Thus, an adversary looking to perpetrate an attack to recover the plaintext can treat the coefficients of the polynomials in Equation (26) and evaluated ciphertext as integers. The plaintext values, sought after by the adversary, are elements of the field Fq, thus the malicious party can reduce the public values of the ciphertext modulo q to solve for plaintext variables in the Equation (26). We formally phrase it in the following remark.\n\nFor the purpose of obtaining the plaintext, the ciphertext and cipher coefficients as illustrated in the Equation (26) can be considered modulo q as follows\n\n(Modular Diophantine Equation). The Modular Diophantine Equation asks whether an integer solution exists to the equation\n\nA positive answer to this question would include a solution.\n\nLet m+1>2. Note that the system in the Equation (28) can be normalized as\n\nA naive way of solving such a normalized system is to solve each equation and find a common solution. Each such equation is an instance of a Modular Diophantine Equation. The more obvious way to solve the system in Equation (28) would be to use Gaussian elimination and transform the system into a single equation. Indeed, since the coefficients of the ciphertext are publicly known, and the noise variables are linear in the ciphertext, the adversary can express any noise variable using the remaining terms of the equation and reduce the system to a single equation of the form\n\nLet m+1>2. The ciphertext in its normalized reduced form is a single equation\n\nNote that even in its normalized reduced form the ciphertext is an instance of a Modular Diophantine Equation. Since m+1>2 we can argue that the adversary does not benefit much by reducing the system in Equation (29) to a single equation (31), and eliminating one variable. The number of expected solutions to the Equation (31) remains qm−1, and the adversary is facing with the problem of deciding which solution is the correct one. That is, a brute-force search algorithm can find a list of solutions to the Equation (31) by trying all the possible m−1 variables values over Fq. The adversary is interested in a particular solution from the list.\n\nOne might argue that the attacker is interested only in the plaintext variable x∈Fq. Thus, the adversary can simply guess the value x. The complexity of this guess is Oq. This is a probabilistic attack. For a deterministic result, this guess has to be tested for correctness. This will require coming up with values for the noise variables and testing whether the guess is correct. Moreover, NIST requires the size of the actually communicated secret to be 32 bytes.61 Thus, the secret that is transferred between two parties consists of v=32/q8 blocks, where each block is q2 bits. Each block corresponds to the HPPK secret x. The secret message is then v different values x concatenated together to form a 32 byte secret. Each such block x is encrypted separately using HPPK. The complexity of correctly guessing the transferred secret message is then Oqv, where v=32/q, comparing with the complexity Oqvm−1 of attacking all blocks.\n\nThe Modular Diophantine Equation Problem is NP-complete.\n\nThe proof, using the Boolean Satisfiability Problem, is given by Moore and Meterns [,62 Section 5.4.4]. □\n\nTheorem 0.1 states that a brute-force search algorithm can find a solution to the Modular Diophantine Equation by trying all the possible solutions. Thus, without loss of generality, we treat the ciphertext-only attack on a ciphertext in its normal reduced form as a Modular Diophantine Problem. Indeed, by Theorem 0.1 the algorithm to find a solution to a Modular Diophantine Equation does not simply terminate to give a solution, it is a brute-force search algorithm that considers every possible solution before producing a result. In other words, it goes through all the possibilities to choose the correct one.\n\nWe suppose that the adversary will choose to perpetrate the attack on the ciphertext in its normal reduced form as in Equation (31), for its easier to attack. In the framework of HPPK, the public key elements are the coefficients of the ciphertext polynomials.\n\n(HPPK has IND-CPA property). Let m>1, where m is the total number of variables in the normalized reduced form of the ciphertext as in Equation (31). If the Modular Diophantine Equation is NP-complete, the HPPK encryption system is provably secure in the IND-CPA security model with a reduction loss of qm−2.\n\nAssume that there exists an adversary A that tε-breaks the HPPK encryption system in the IND-CPA security model. We construct a simulator ℬ that solves the Modular Diophantine Equation. Given as input, a Modular Diophantine Equation instance qHxx1…xm−1, where Hxx1…xm−1 is of the form Equation (31) and m>1, the simulator ℬ runs A as follows. The simulator sets the normalized public key over Fp to the coefficients of the polynomial Hxx1…xm−1. The challenge consists of the following game. The adversary A generates two distinct messages m0 and m1∈Fq, and submits them to the simulator. The simulator ℬ randomly chooses b in 01 as well as random values r1,…,rm−1 for the noise variables, and sets the ciphertext to be the value\n\nThe challenge for the adversary then consists of the following equation to be solved for x:\n\nThis equation remains to meet the definition of the Modular Diophantine Equation Hxx1…xm−1−1=0, since the value 1H¯ can be pushed to the noise variables, which are random and do not influence the plaintext. Indeed, let hij be the coefficients of the polynomial Hxx1…xm−1 for any i∈0…n and j∈1…m−1, then\n\nThe coefficients of the challenge equation come from the submitted Diophantine equation, and thus, from the point of view of the adversary are random. The values r1,…,rm−1 are selected at random. The adversary does not have knowledge of the values x1′…xm−1′ and they can not be calculated from the other parameters given to the adversary. So the noise variables xj′ for all j∈1…m−1 are random. Hence, the simulation holds randomness property. By construction, the simulation is indistinguishable from a real attack. That is, the adversary is challenged with solving the equation as in the Equation (30), which is HPPK ciphertext in its normalized reduced form.\n\nThe simulator does not abort in the simulation while interacting with the adversary. The adversary outputs a random guess b′ of b. When b′ is equal to b, the adversary wins. Otherwise, the adversary looses. The probability of simply guessing the value for x is Pr=12. We will calculate the probability of solving the IND-CPA challenge with the advantage of the adversary, that is Pr=12+α. The advantage comes from the assumption that the adversary can break the HPPK cryptosystem.\n\nThe challenge has a general form as in the Equation (31), thus, the equation is expected to have qm−1 distinct solutions, considering all m variables. On the other hand, it is known that the variable x∈m0m1. Assuming x=m0, there are now qm−2 possible solutions to choose the correct solution from. The same is true for x=m1. That is, the probability of finding correct solution of the equation Hxx1…xm−1−1=0 is\n\nAccounting for the advantage that the adversary has, the probability α is Prcorrect solution=εqm−2. The total probability of solving the IND-CPA challenge is then 12+εqm−2.\n\nThe simulation is indistinguishable from a real attack. So the adversary who can break the challenge ciphertext will uncover the solution to the given Modular Diophantine Equation problem. The probability of breaking the ciphertext is εqm−2.\n\nThe advantage of solving the Diophantive Equation problem is then εqm−2. Let Ts denote the time cost of the simulation. We have Ts=O1. The simulator ℬ solves the Modular Diophantine Equation with time cost and advantage t+Tsε/qm−2=tε/qm−2. Thus, contradicting the Theorem 0.1 so the initial assumption is wrong. □\n\nThe framework of the IND-CPA challenge entails known plaintext, in other words, the adversary knows that the secret x∈m0m1. We now state the complexity of the unknown plaintext ciphertext-only attack.\n\n(Ciphertext-only attack). Let m+1>2. The classical complexity of finding the plaintext from the ciphertext is Oqm−1.\n\nLet the adversary favour the ciphertext in its reduced normal form (31). Without any knowledge about the plaintext, the adversary will need to solve the Equation (31) to obtain the plaintext along with the noise variables. A single equation over Fq with m variables is expected to have qm−1 possible solutions over Fq. The correct one is among them. That is, the plaintext encapsulated in a single variable x is not the sole variable in the ciphertext equation. However, it is the only unknown of interest. The adversary can try and simply guess x, the complexity of the guess is Oq. However, they have to test whether their guess is correct. Moreover, the secret transferred between the communicating parties consist of 32 bytes as required by NIST. Thus, the adversary will have to guess K many values for x, where K=32×8q. In this case, the complexity is OqK. We expect K>m−1. Quantum complexity of the described attack due to Grover’s search algorithm is Oqm−12. □\n\n\n\nLet λ≤2. There exists a polynomial time algorithm to find coefficients of univariate polynomials f1x0 and f2x0 given the plain central maps P1 and P2.\n\nNote that all the plain coefficients of the polynomials p1xx1…xm and p2xx1…xm as defined in Equation (22) are defined over the prime field Fq. Thus, for any fixed j, it is possible to use Gaussian elimination to reduce the system of equations formed by the plain coefficients of p1xx1…xm and p2xx1…xm of the form\n\nLet λ≤2. Finding private key from the cipher public key in the framework of HPPK reduces to finding the homomorphic encryption key S,R1, and R2.\n\nThe private key consists of the coefficients of the univariate polynomials f1x,f2x as well as values S,R1,R2 used to encrypt the plain public key to the cipher public key. By Lemma 0.4 once the values R1,R2 and S are known, the coefficients of f1x,f2x can be found in polynomial time. □\n\n(Diophantine set). The Diophantine set is a set S⊂ℕ associated with a Diophantine equation Pba1…ak∈ℤba1…ak, where k>0 such that\n\n(MRDP Theorem). The Matiyasevich–Robinson–Davis–Putnam (MRDP) theorem states that every computably enumerable set is Diophantine, and every Diophantine set is computably enumerable.\n\nThe result has been proven in various works, for instance.65 □\n\n(Hilbert's tenth problem) Hilbert’s tenth problem asks whether the general Diophantine Problem is solvable. Due to MRDP, Hilbert’s tenth problem is undecidable.\n\nFor proof see.65 □\n\nPrivate key attack is non-deterministic and has complexity of at least OT3, where T is the largest number with n+λ+1q2+\|(\|q2)2 bit-length.\n\nBy Lemma 0.4 and 0.5 the attack on public key reduces to finding the values S,R1, and R2. From the perspective of the attacker, the values S,R1, and R2 could be treated as a one-time pad keys as they have been chosen at random, and can not be calculated from other parameters given to the attacker. An obvious attack would be a brute force search for all the three values, S,R1, and R2. The direct brute force search classical complexity would be greater than OT3 for the three values together, where T is the largest n+λ+1q2+\|(\|q2)2-bit number. Due to Grover’s algorithm, the quantum complexity is greater than OT32. Note however, because of the condition gcdSR1=gcdSR2=1, once S is found the search span for R1 and R2 reduces. Brute force search entails a non-deterministic result, however, we provide a more formal argument below.\n\nFor each fixed chose of j, each public key coefficient can be written in the integer domain as follows\n\nSolving such equation by Theorem 0.6 and 0.7 is an NP-complete task. For each j, we can generate one such equation. Considering them all together, the adversary will arrive at an underdetermined system as the variables in the system depend on j. Each equation in such a system is a multivariate Diophantine equation. One way to solve this system is to solve each equation separately and search for common solutions. However, by Theorem 0.6 and 0.7 this is an NP-complete problem. Reducing the system to a single polynomial still produces a multivariate Diophantine equation, solving which is an NP-complete problem by Theorem 0.6 and 0.7. □\n\nRecall, that the homomorphic operator ℰ̂S,R is defined by the two secret values S,R. These values are known only to the decrypting party and are never shared. Thus, we assume that the adversary can not simply access the homomorphic encryption oracle. Indeed, if the adversary were to have access to such oracle, they could pass 1 to be encrypted, resulting in the output value R. Thus, the adversary would learn half of the homomorphic secret key. In order to find the value S, the adversary can pass values R−1k to be encrypted, producing k mod S. The values k can be chosen to determine S. For instance, with public knowledge of the bit-size of S, the adversary can consider an interval between the smallest number of that bit-size and the largest number of that bit-size, and divide that interval in half, where k is the mark at the half. If kmodS≠k, then S must be smaller than k. If kmodS=k, then S is larger than k. In the first case, take the interval between the smallest number of bit-size 2p2+L2 and k, and consider the halfway mark. Repeat the experiment, each time decreasing the interval until the value S is “trapped” in a small interval and can be determined. Similarly for the latter case, take the interval between k and the largest number of bit-size 2p2+L2, mark a halfway and use that mark value to repeat the experiment. Based on whether or not the value changes, decrease the interval and repeat the experiment. This is only but one way to find S, while R is known.\n\nSince the operation is deterministic, after the values of R and S are fixed, it is important that they remain secret and the adversary does not have access to the homomorphic encryption oracle. Note that in the framework of HPPK, the adversary has access to the public key polynomials which are essentially randomly chosen values encrypted using the homomorphic operator with secret values R,S. Without the knowledge of the values before they were acted on with ℰR,S, finding R,S can be considered a brute force search problem.\n\nAt large, the security of the HPPK cryptosystem relies on the problem of solving undetermined system of equations over Fq. Such system is expected to have qv−w possible solutions, where v is the number of variables and w is the number of equations in the system. The attacker can solve this system to find all possible solutions, however, it is the problem of determining the correct solution from all the possible solutions that makes HPPK secure.\n\nThe ciphertext attack requires the adversary to solve an underdetermined system of equations over Fq, which can be reduced to a single Modular Diophantine equation. Solving this equation is an NP-complete problem.\n\nThe public key attack aimed to unveil the plaintext reduces to a brute force search for three unknown values S,R1,R2. To find these values, the attacker can either use brute-force search or solve an underdetermined system of equations over the integers. The former yields non-deterministic results and the latter is an NP-complete problem.\n\nWe conclude that from the point of view of the adversary, the following is true.\n\nThe best classical complexity to attack HPPK is Oqm−1.\n\nWe assume that the malicious party will take the most advantageous path for them. Thus, by Lemma 0.3, Lemma 0.4, Lemma 0.5, and Theorem 0.8 we can conclude that the best attack is to obtain the plaintext from the ciphertext. Such attack is non-deterministic with classical complexity of Oqm−1. □\n\n\nConclusions\n\nIn this paper, we introduce a new homomorphic encryption scheme intended to secure public keys of multivariate asymmetric cryptosystems. Unlike conventional homomorphic encryption, our scheme uses encrypted plain public keys or CPK that can be published for anyone to use to establish a secret sharing between two parties. Our homomorphic encryption is applied to polynomials, leveraging homomorphic properties and allowing for user input through variables. The homomorphic encryption and decryption operators are modular multiplication operators modulo a hidden value S, with values R1 and R2 chosen uniformly at random from the hidden ring ℤS under certain conditions. We propose using this homomorphic encryption in conjunction with Multivariate Polynomial Public-key Cryptosystem (MPKC) to secure polynomial public keys, called Homomorphic Polynomial Public Key (HPPK). We describe the HPPK algorithm in detail, drawing from the framework of MPKC. HPPK public keys are product polynomials of a multivariate and univariate polynomials, encrypted with a homomorphic encryption operator. The ciphertext is created by the encrypting party through the input of plaintext and random noise as public polynomial variables. The decryption procedure involves first homomorphic decryption of the ciphertexts to produce plain polynomial values for a division of two plain polynomials. By construction, such division cancels the base multiplicand polynomial with the noise variable and retains a single equation in one variable. This variable is the plaintext, which can be found by radicals. We provide a thorough security analysis of the HPPK cryptosystem, proving that the hardness of breaking the HPPK algorithm comes from the NP-completeness problem of the Modular Diophantine Equation. We also show that HPPK holds the IND-CPA property. In the future work, we will perform a detailed benchmarking study with variety of configurations as well as a more extensive security analysis, considering attacks that have not been described in the work.\n\n\nAuthor contributions\n\nR.K. provided the core ideas. M.P. developed the security analysis. All authors reviewed the manuscript and approved its publication.", "appendix": "Data availability\n\nNo data is associated with this article.\n\n\nAcknowledgements\n\nAll authors acknowledge Professor Michel Barbeau for his contributions to the MPPK algorithm and discussions regarding security reductions included in this work.\n\n\nReferences\n\nRivest RL, Adleman L, Dertouzos ML: On data banks and privacy homomorphisms. 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Springer; 1997; pages 356–368.\n\nPatarin J: Hidden fields equations (hfe) and isomorphisms of polynomials (ip): Two new families of asymmetric algorithms.Maurer U, editor. Advances in Cryptology — EUROCRYPT’96. Berlin, Heidelberg: Springer Berlin Heidelberg; 1996; pages 33–48. 978-3-540-68339-1.\n\nWang LC, Yang BY, Hu YH, et al.: A medium field multivariate public-key encryption scheme. In CT-RSA 2006, volume 3860 of LNCS. 132–149.\n\nWang X, Wang X: An improved medium field multivariate public key cryptosystem. 2008 Third International Conference on Convergence and Hybrid Information Technology. 2008; volume 2: pages 1120–1124.\n\nFaugére J-C: A new efficient algorithm for computing gröbner bases (f4). Journal of Pure and Applied Algebra. 1999; 139(1): 61–88. 0022-4049. Publisher Full Text\n\nFaugère JC: A new efficient algorithm for computing gröbner bases (f4). 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Advances in Cryptology — EUROCRYPT’99. Berlin, Heidelberg: Springer Berlin Heidelberg; 1999; pages 206–222.978-3-540-48910-8.\n\nKuang R: A deterministic polynomial public key algorithm over a prime galois field gf(p). 2021 2nd Asia Conference on Computers and Communications (ACCC). 2021; pages 79–88.\n\nKuang R, Barbeau M: Indistinguishability and non-deterministic encryption of the quantum safe multivariate polynomial public key cryptographic system. 2021 IEEE Canadian Conference on Electrical and Computer Engineering (CCECE). 2021; pages 1–5.\n\nKuang R, Barbeau M: Performance analysis of the quantum safe multivariate polynomial public key algorithm. 2021 IEEE International Conference on Quantum Computing and Engineering (QCE). 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Translated from the 1993 Russian original by the author, with a foreword by Martin Davis." }	[ { "id": "216597", "date": "01 Nov 2023", "name": "Gui-Lu Long", "expertise": [ "Reviewer Expertise Quantum information and quantum algorithm." ], "suggestion": "Approved", "report": "Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIn this work, the authors presents a variant of the MPKC. It simplifies MPKC central map to two multivariate polynomials constructed from polynomial multiplications. The HPPK algorithm employs a single polynomial vector for the plaintext and a multi-variate noise vector associated with the central map. It is Indistinguishability Under Chosen-Plaintext Attack (IND-CPA) secure, and its classical complexity for cracking is exponential in the size of the prime field GF(p). It is an important PQC algorithm and has great potential in the near future applications. I recommend its acceptance in its present form.\n\nIs the work clearly and accurately presented and does it cite the current literature? Yes\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "242797", "date": "22 Mar 2024", "name": "Taniya Hasija", "expertise": [ "Reviewer Expertise Cryptography", "Post quantum cryptography" ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nA novel homomorphic polynomial public key encapsulation algorithm The author has presented a proposed method named as homomorphic polynomial public key encapsulation algorithm that incorporates an additional layer of encryption during key construction procedure, through a hidden ring. The proposed algorithm is applied to Multivariate Public Key Cryptography Schemes (MPKC) to hide the structure of their central map construction. A homomorphic operator is performed to encrypt the public key, called as new variant of MPKC. There is a need for some improvemeAczxnts before acceptance of the manuscript. 1. The abstract should begin by outlining the background information or context of your field, followed by the precise subject of your study, a problem statement, and the course of action for this article. Finally, take into account the significance of your work, particularly how the community will benefit from your article. There are several errors in the abstract, which is not up to par. Certain lines lack context, while others are incomplete. There is not enough clarity. 2. Title of the article is general; it should be specific. 3. The meaning of homomorphic world in context to cryptography is not clear in introduction. Why is there a need to work on homomorphic cryptography? 4. The description of the algorithm is useful, but much better would be to provide the code as supplementary material, which would facilitate the use of this algorithm to those who are not experts on recommender systems. 5. This paper's citations are not current; it would be beneficial to incorporate some more recent work. 6. Literature work should be improved in the paper, as it is very limited. Summarize the related works in the form of a table. Add one or two tables of comparison along with line diagram of progress in existing work. What are the limitations of existing algorithms that motivated you to do the current research?\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nYes\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1347
https://f1000research.com/articles/11-886/v1	02 Aug 22	{ "type": "Research Article", "title": "Observation of SARS-CoV-2 genome characteristics and clinical manifestations within eight family clusters from GH and GK clades in Jakarta, Indonesia", "authors": [ "Fera Ibrahim", "Augustine Natasha", "Andi Yasmon", "Fithriyah Fithriyah", "Anis Karuniawati", "Rivia Gina Rahmawaty", "Yulia Saharman", "Pratiwi Sudarmono", "Augustine Natasha", "Andi Yasmon", "Fithriyah Fithriyah", "Anis Karuniawati", "Rivia Gina Rahmawaty", "Yulia Saharman", "Pratiwi Sudarmono" ], "abstract": "Background: SARS-CoV-2 rapid mutation generates many concerning new strains. Although lockdown had been applied to contain the disease, the household remains a critical place for its transmission. This study aimed to assess the variation of SARS-CoV-2 strains and their clinical manifestations within family clusters in Jakarta, Indonesia. Method: Naso-oropharyngeal swab specimens from family clusters positive for SARS-CoV-2 were collected for whole-genome sequencing. Their baseline data, symptoms, and source of infection were recorded. The whole-genome data was then analyzed with the bioinformatics program to evaluate the SARS-CoV-2 genome characteristic and submitted to GISAID for strain identification. The phylogenetic tree was built to observe the relationship between virus strain within the family cluster and its clinical manifestation. Result: This study obtained eight family clusters from twenty-two patients. Half of the cluster's source of infection was a family member who had to work at the office. The infection rate ranged from 37.5% to 100%. The phylogenetic tree showed that the same virus within a cluster could generate various clinical outcomes. Five clusters had one family member with pneumonia, while others had mild to no symptoms. Four breakthrough cases were detected in this study, which were infected by the virus from GH and GK clade. Conclusion: Our study observed the SARS-CoV-2 transmission to the household setting through the workplace, which might become a common pathway if the pandemic status is removed. Although vaccination is expected to reduce the burden of COVID-19, adequate control measures remain essential, given that breakthrough infections are evident.", "keywords": [ "SARS-Cov-2", "Family Cluster", "Whole-Genome Sequence", "Next Generation Sequencing", "Breakthrough infection." ], "content": "Introduction\n\nSARS-CoV-2 was first detected in late 2019; however, its rapid mutations have resulted in several dominant strains that threaten the efficacy of current treatment and control protocols. D614G was the first reported mutation to rapidly emerge among the dominant viral strains worldwide.1 This mutation then increased and generated the emergence of the variants of concern, such as Alpha, Beta, Gamma, and Delta, which alternately became the dominant strain in many countries.1 These variants had been reported to have higher transmissibility, more severe clinical manifestations, and the ability to potentially escape the immunity conferred by the current vaccines.2\n\nLockdown and quarantine have been the main actions applied to contain the spread of SARS-CoV-2 in the population. Nonetheless, the household setting has been a critical SARS-CoV-2 transmission pathway.3,4 The rate of secondary transmission among household contacts has varied from 16.6% to 30%.3,5 Household transmission is more challenging to control, given the frequent unavoidable contacts among family members sharing the same home.6 In addition, transmission within households has been reported to be increased in families of greater size.7 The urban areas of Indonesia are less likely to have a family size of fewer than four individuals.8 The basic reproduction number in Jakarta during the early spread of COVID-19 was 1.75,9 which has made the household setting one of the vital arenas for COVID-19 transmission in Indonesia.\n\nSARS-CoV-2 surveillance has been implemented in many referral laboratories in Indonesia since the middle of 2020, and it has supported researchers in examining the dynamics of SARS-CoV-2 variants throughout the year. This study was performed to assess the variation of SARS-CoV-2 strains and their clinical manifestations within family clusters in Jakarta, Indonesia.\n\n\nMethods\n\nThis study was conducted after obtaining ethical clearance from the Research Ethics Committee of Universitas Indonesia (protocol number 20-05-0516). Patients were informed before providing written consent. All the patients involved in this study agreed to have their medical information and specimens used for analysis.\n\nWe selected naso-oropharyngeal swab specimens from family clusters positive for SARS-CoV-2 for whole-genome sequencing, choosing at least two patients to represent each family cluster. The specimens were stored in -800C from the date of detection to the sequencing process. The 200-μl of specimen extraction was performed according to the manufacturer’s instructions (PureLink™ Viral RNA/DNA Mini Kit, Thermo Fisher Scientific, USA). The real-time reverse transcription polymerase chain reaction (PCR) US Centers for Disease Control and Prevention validation test targeting N1 and N2 was performed according to the manufacturer’s instructions (SensiFAST™, Meridian Bioscience, USA).\n\nThe total RNA from positive extraction was then cleaned with DNase I and concentrated (RNA Clean & ConcentratorTM-5, Zymo Research, USA). The libraries were prepared according to the ARTIC multiplex PCR method, using an nCoV-19 V3 panel.10 The libraries were then processed with Oxford Nanopore’s GridION sequencer, employing MinKNOW version 20.06.9 and MinKNOW core version 4.0.11.10 High accuracy basecalling was conducted using Guppy version 4.0.11.11 All generated reads were assembled using EPI2ME Labs platform employing ARTIC workflow.10\n\nThe complete genome sequences were uploaded to GISAID for identification of clade, lineage, and mutations. The sequences were further annotated using SnpEff v4.3.12 The phylogenetic tree was built using Geneious Prime 2021.1.1, using UPGMA as the tree build method and Tamura-Nei as the genetic distance model, with NC_045512 as the reference sequence.\n\n\nResults\n\nThis study included 22 patients from eight family clusters, performed from January 2021 to August 2021. The patient characteristics are shown in Table 1. The phylogenetic tree is shown in Figure 1.\n\nThe source of infection in clusters A and D was unknown, and the infections were detected due to the screening program; the source of infection in clusters B, C, E, and F was a family member who had a history of working from the office during the pandemic; the source of infection in cluster G was a family member who had a history of traveling to another province via airplane for business assignments; and the source of infection in cluster H was the nanny who worked in the house during the day. Five patients had been vaccinated, with four of them completing the second dose.\n\nIn clusters C (4 of 4), D (5 of 5), E (5 of 5), F (3 of 3), G (4 of 4), and H (4 of 4), the whole family had become infected with SARS-CoV-2. For cluster A, only three of eight family members became infected with COVID-19. For cluster B, 4 of 5 family members caught COVID-19. The infection rate ranged from 37.5% to 100%.\n\nNineteen of the 22 patients isolated at home, with patients six, seven, 11, and 20 having pneumonia during their isolation, but who were not able to be admitted to the hospital because of the lack of available beds. However, these patients had access to the outpatient COVID-19 clinics to obtain doctor assessment, symptomatic medications, and radiology tests. The only patient with pneumonia who was able to be admitted to the hospital was patient three. Most patients did not have their blood tested during their early days after being detected positive due to difficulties reaching the healthcare service at the peak of the first and second wave. The Pango lineages circulating within the clusters were B.1.459, B.1., B.1.466.2, B.1.1.526, B.1.617.2 + AY23, and AY24. Most clusters had the same strain within each patient, except cluster F. For cluster F, patient 16 was detected positive after regular screening because he took care of patient 15, which explains the distance between the dates of the SARS-CoV-2 positive results between the patients.\n\nThis study had 2 dominant clades, GH and GK. The evaluation of the variation rates, transition-transversion (ti/tv) ratio, total single nucleotide polymorphism (SNP), total deletion, total variation, missense-silent ratio, spike (S) and nucleocapsid (N) gene nonsynonymous mutations, and total nonsynonymous mutation are shown in Table 2.\n\nIn addition, given that the spike gene had the highest number of mutations, the distribution of its nonsynonymous mutations within the patients is shown in Table 3.\n\n* Patients with pneumonia.\n\n\nDiscussion\n\nFour clusters in our study had an index patient with a history of working in an office. Cases of SARS-CoV-2 infection from office workers outside the healthcare setting were scarcely reported.13 Six of eight family clusters had a 100% infection rate, showing the efficacy of SARS-CoV-2 transmission within the family group. As reported by several studies, the household infection rate ranged between 75% and 100%.3,14 The high infection rate within households combined with poor preventive measures in the workplace could generate a spike in COVID-19 cases.\n\nThis study also found SARS-CoV-2 infections in patients who had been vaccinated (breakthrough cases). Infection after complete vaccination was possible, no matter which type of vaccine had been administered.15,16 Duarte et al. reported the breakthrough cases mainly were mild, and that other factors might have influenced the symptomatic infections, given that they were not correlated with lack of vaccine-induced immunity.15 Although our study could not represent the general population, the concern of vaccine efficacy in the face of new variants with more mutations has become relevant since Christensen et al. reported an increase in breakthrough cases caused by the Delta variant (as part of the GK clade).17 Therefore, preventive measures such as wearing a mask are still necessary, even after full vaccination.\n\nThe clinical manifestation varied within the patients in each family cluster. This situation was in concordance with several case reports, which have mentioned variations in disease severity within a family cluster.18 However, seven clusters were monophyletic, which showed that the same virus could generate a different outcome. Furthermore, we found the virus genome of the patients with pneumonia was similar to the other patients without pneumonia (Table 1, Figure 1). No specific variation or combination in the genome mutation could generate a particular clinical outcome (Table 3). The D614G spike mutation was also distributed evenly within the patients. This finding was in concordance with Korber et al. and Lorenzo-Redondo et al., who noted that the D614G mutations did not correlate with more severe outcomes.19,20\n\nOur study showed a ti/tv ratio decline between the GH and GK clades. This decline was in concordance with the study by Duchêne et al., which revealed the ti/tv scaled negatively over time, especially for rapidly evolving RNA viruses.21 We also found an increase in the GK total spike gene mutations compared with the GH clade. Given that the GK clade appeared later in our study, it was expected to have more mutations because the RNA virus would utilize various mechanisms of genetic variation to guarantee its survival.22 The evolving behavior can facilitate the virus to evade the current vaccine-generated immunity, resulting in breakthrough infections.\n\nOne limitation of our study was the small number of patients and family clusters. This study could not provide serological test results, especially from patients with vaccination history, given that there were no guidelines to confirm an immune response after vaccination. Our study showed that the same virus within a cluster could generate various clinical outcomes. Our observation showed the SARS-CoV-2 transmission in the household setting through the workplace, which could be a common pathway in the future of COVID-19 once the pandemic status is lifted. Although vaccination is expected to reduce the burden of COVID-19, control measures are crucial, given that breakthrough infections due to a new variant are evident.\n\n\nData availability\n\nThe virus genome included in this study was published at GISAID with accession ID:\n\nEPI_ISL_2692986, EPI_ISL_2692987, EPI_ISL_2692989, EPI_ISL_2993633, EPI_ISL_2993659, EPI_ISL_2993632, EPI_ISL_2692990, EPI_ISL_2692991, EPI_ISL_2692992, EPI_ISL_2993434, EPI_ISL_2993539, EPI_ISL_2993658, EPI_ISL_2692909, EPI_ISL_2692981, EPI_ISL_2692983, EPI_ISL_3000155, EPI_ISL_3456060, EPI_ISL_4847411, EPI_ISL_4847413, EPI_ISL_5054899, EPI_ISL_5054915, EPI_ISL_5054918.", "appendix": "Acknowledgements\n\nThe authors would like to thank Universitas Indonesia, through PUTI Grant with contract number NKB-1493/UN2.RST/HKP.05.00.2020.\n\n\nReferences\n\nChoi JY, Smith DM: Sars-cov-2 variants of concern. Yonsei Med. J. 2021; 62(11): 961–968. 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Publisher Full Text\n\nMartin CA, Jenkins DR, Minhas JS, et al.: Socio-demographic heterogeneity in the prevalence of covid-19 during lockdown is associated with ethnicity and household size: Results from an observational cohort study. EClinicalMedicine. 2020; 25: 100466. PubMed Abstract \| Publisher Full Text\n\nLaksono AD, Wulandari RD: The factors correlate to family size in indonesia. Aspirasi: Jurnal Masalah-masalah Sosial. 2021; 12(1): 1–13.\n\nAldila D, Khoshnaw SHA, Safitri E, et al.: A mathematical study on the spread of covid-19 considering social distancing and rapid assessment: The case of jakarta, indonesia. Chaos, Solitons Fractals. 2020; 139: 110042. PubMed Abstract \| Publisher Full Text\n\nTyson J, James P, Stoddart D, et al.: Improvements to the artic multiplex pcr method for sars-cov-2 genome sequencing using nanopore. bioRxiv. 2020.\n\nWick RR, Judd LM, Holt KE: Performance of neural network basecalling tools for oxford nanopore sequencing. Genome Biol. 2019; 20(1): 129. PubMed Abstract \| Publisher Full Text\n\nCingolani P, Platts A, Wang le L, et al.: A program for annotating and predicting the effects of single nucleotide polymorphisms, snpeff: Snps in the genome of drosophila melanogaster strain w1118; iso-2; iso-3. Fly. 2012; 6(2): 80–92. PubMed Abstract \| Publisher Full Text\n\nAgius RM, Robertson JFR, Kendrick D, et al.: Covid-19 in the workplace. BMJ (Clinical research ed). 2020; 370: m3577. Publisher Full Text\n\nMaltezou HC, Vorou R, Papadima K, et al.: Transmission dynamics of sars-cov-2 within families with children in greece: A study of 23 clusters. J. Med. Virol. 2021; 93(3): 1414–1420. PubMed Abstract \| Publisher Full Text\n\nDuarte LF, Gálvez NMS, Iturriaga C, et al.: Immune profile and clinical outcome of breakthrough cases after vaccination with an inactivated sars-cov-2 vaccine. Front. Immunol. 2021; 12: 742914. PubMed Abstract \| Publisher Full Text\n\nTenforde MW, Self WH, Naioti EA, et al.: Sustained effectiveness of pfizer-biontech and moderna vaccines against covid-19 associated hospitalizations among adults - United States, March-July 2021. MMWR Morb. Mortal. Wkly Rep. 2021; 70(34): 1156–1162. PubMed Abstract \| Publisher Full Text\n\nChristensen PA, Olsen RJ, Long SW, et al.: Delta variants of sars-cov-2 cause significantly increased vaccine breakthrough covid-19 cases in houston, texas. Am. J. Pathol. 2021.\n\nChen D, Li Y, Deng X, et al.: Four cases from a family cluster were diagnosed as covid-19 after 14-day of quarantine period. J. Med. Virol. 2020; 92(10): 1748–1752. PubMed Abstract \| Publisher Full Text\n\nKorber B, Fischer WM, Gnanakaran S, et al.: Tracking changes in sars-cov-2 spike: Evidence that d614g increases infectivity of the covid-19 virus. Cell. 2020; 182(4): 812–827.e19. PubMed Abstract \| Publisher Full Text\n\nLorenzo-Redondo R, Nam HH, Roberts SC, et al.: A unique clade of sars-cov-2 viruses is associated with lower viral loads in patient upper airways. medRxiv. 2020.\n\nDuchêne S, Ho SY, Holmes EC: Declining transition/transversion ratios through time reveal limitations to the accuracy of nucleotide substitution models. BMC Evol. Biol. 2015; 15: 36. Publisher Full Text\n\nDomingo E, Holland JJ: Rna virus mutations and fitness for survival. Annu. Rev. Microbiol. 1997; 51: 151–178. Publisher Full Text" }	[ { "id": "146395", "date": "15 Aug 2022", "name": "So Nakagawa", "expertise": [ "Reviewer Expertise Genomics." ], "suggestion": "Approved With Reservations", "report": "Approved With Reservations\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nIbrahim et al. sequenced SARS-CoV-2 genomes that were obtained from 22 COVID-19 patients in 8 family clusters. Except for one family, the results of SARS-CoV-2 genome analysis strongly suggested that the disease was transmitted by household infection.\nThe study itself is small but solid, and their observation is still reasonable for the current infection dynamics of COVID-19.\nMajor comments The details of each nanopore sequencing should be shown in the manuscript. For nanopore sequencing, the quality is known to be relatively low compared to Illumina sequencing. Therefore, if the depth is insufficient, it is impossible to distinguish whether those are mutations or sequencing errors. Please provide more information on the sequencing status (including depth, coverage, and any “shallow” regions if they exist) for each sample.\nMinor comments\nIntroduction: “D614G” should be clearly explained when it first appeared.\n\nTable 1: Are PANGO IDs isolated for Patients 4 to 6 “B.1”? The B.1 lineage is not common in January 2021 in the world. Please verify it.\n\nFigure 1: The legend of the figure is missing, and I could not understand the meaning of the values shown for each branch. In addition, clades and PANGO IDs should be shown, which could be informative for readers.\n\nTable 3: “Total S mutation” and “Total N mutation” are ambiguous. Do these include synonymous mutations as well?\n\nReferences: Citations should be checked. For example, Reference #1 is not suitable for mentioning SARS-CoV-2 VOCs.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and analysis provided to allow replication by others? Yes\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nPartly\n\nAre all the source data underlying the results available to ensure full reproducibility? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "10088", "date": "16 Oct 2023", "name": "Fera Ibrahim", "role": "Author Response", "response": "Dear So Nakagawa, Thank you very much for your input and effort to review our paper. Major comments The details of each nanopore sequencing should be shown in the manuscript. For nanopore sequencing, the quality is known to be relatively low compared to Illumina sequencing. Therefore, if the depth is insufficient, it is impossible to distinguish whether those are mutations or sequencing errors. Please provide more information on the sequencing status (including depth, coverage, and any “shallow” regions if they exist) for each sample.[P1] Response: Thank you very much for the inputs. We already use the cutoff 100x depth of coverage for all regions before we proceed for analysis. That’s also why we only included this small number of specimens, since only these specimens could pass such cutoff. In addition, sequencing using illumina platform was not feasible and too expensive in Indonesia for small project like ours during the period of our study, therefore we relied on Nanopore technology to support our country’s surveillance initiatives. Your message on Illumina is well noted, we will try them if we got more fundings in the future. Minor comments Introduction: “D614G” should be clearly explained when it first appeared. Response: thank you for the input, it’s been revised. Table 1: Are PANGO IDs isolated for Patients 4 to 6 “B.1”? The B.1 lineage is not common in January 2021 in the world. Please verify it. Response : According to pango lineage page (Cov-Lineages, B.1 earliest date of detection was January 1st,2020. Therefore, we’re sure that the B.1 lineage had arrived to Indonesia in 2021. We will add O’Toole’s study as part our methods in terms of evaluating the pangolins lineage. Figure 1: The legend of the figure is missing, and I could not understand the meaning of the values shown for each branch. In addition, clades and PANGO IDs should be shown, which could be informative for readers. Response : Thankyou for the input, the figure is revised. Table 3: “Total S mutation” and “Total N mutation” are ambiguous. Do these include synonymous mutations as well? Response : the mutations included only the non-synonymous one, as resulted from the GISAID analysis reports. References: Citations should be checked. For example, Reference #1 is not suitable for mentioning SARS-CoV-2 VOCs. Response: Thank you for the concerns. We consider your input and we had replaced them with different citations." } ] }, { "id": "146400", "date": "23 Aug 2022", "name": "Serafeim C. Chaintoutis", "expertise": [ "Reviewer Expertise Clinical microbiology", "infectious diseases", "immunology", "virology" ], "suggestion": "Not Approved", "report": "Not Approved\n\ninfo_outline\nAlongside their report, reviewers assign a status to the article:\n\nApproved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested\n\nApproved with reservations\nA number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.\n\nNot approved Fundamental flaws in the paper seriously undermine the findings and conclusions\n\nThe article \"Observation of SARS-CoV-2 genome characteristics and clinical manifestations within eight family clusters from GH and GK clades in Jakarta, Indonesia\" by F. Ibrahim et al., provides basically the genetic findings (based on NGS analysis) on the genetic characteristics of SARS-CoV-2 strains retrieved from 8 infected families (“family clusters”) in Jakarta as of Jan. 2021. In my opinion, the present manuscript has only geographically limited interest and brings limited novel findings to the international scientific community. Main concern I am concerned about whether the findings are representative of the situation in Jakarta, at the time of sampling, as there is no mention of how the 8 families that were included in the analysis were selected, geographical characteristics, and so on. The authors should present how those family clusters were selected and investigated.\nSpecific comments:\nAbstract:\n\n\"Although lockdown had been applied to contain the disease, the household remains a critical place for its transmission.\" The two parts of the sentence are not combinable in a scientific manner. Several measures were applied to contain the disease. And many places remain critical for transmission. This also applies to the introduction.\n\n\"Naso-oropharyngeal swab...\" Do authors mean both nasopharyngeal and oropharyngeal? Please elaborate.\n\n\"The infection rate ranged from 37.5 to 100%\". You mean within families?\n\n\"Four breakthrough cases were detected...\" Do authors mean vaccine breakthrough?\n\nIntroduction:\n\"D614G was the first reported mutation...\" This is technically an amino-acid substitution. Please, be precise.\n\n\"This mutation then increased and generated the emergence of the variants of concern\". How is this supported? This is scientifically unjustified.\n\n\"This study was performed to assess the variation of SARS-CoV-2 strains and their clinical manifestations within family clusters in Jakarta, Indonesia\". By this, the impression that of large-scale study is being given. However, 8 families were only investigated. This is not representative. How were these family clusters chosen? Was there any reason? There might be a bias in the selection of families/research material, with potential false community representation results.\n\nMaterials and Methods:\nDetails on the sequencing methodology and analysis are missing. Herein, sequencing was performed using \"Oxford Nanopore’s GridION sequencer\", with read depth lower than Illumina or Ion Torrent systems usings critical to verify that the identified mutations are not an \"artifact\" of the sequencing process.\n\nReferences:\nNeed to be checked and verified.\n\nFig. 1:\nThe relevant caption is missing several important pieces of information. What type of tree is this? Many trees are characterized as \"phylogenetic trees\" Details are needed on how this analysis was performed and the tree was constructed. This information needs to be added to the figure caption. The information provided on the branches is not easy to read and understand.\n\nTables:\nTable 1. This table presents Pango lineages (i.e., B.1) that were not anticipated as of January 2021.\n\nTable 2. It is not clear whether the authors mean in fact mutations, or substitutions by \"total N mutations\", \"total S mutations\", etc.\n\nTable 3. This table also presents amino-acid substitutions, but they are named \"mutations\". Authors need to be precise.\n\nIs the work clearly and accurately presented and does it cite the current literature? Partly\n\nIs the study design appropriate and is the work technically sound? Partly\n\nAre sufficient details of methods and analysis provided to allow replication by others? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate?\nNot applicable\n\nAre all the source data underlying the results available to ensure full reproducibility? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "10087", "date": "16 Oct 2023", "name": "Fera Ibrahim", "role": "Author Response", "response": "Dear Serafeim C. Chaintoutis, Thank you very much for your time and effort to review our study. Here's our response for each input: Abstract: \"Although lockdown had been applied to contain the disease, the household remains a critical place for its transmission.\" The two parts of the sentence are not combinable in a scientific manner. Several measures were applied to contain the disease. And many places remain critical for transmission. This also applies to the introduction. Response: Revised sentence: The lockdown had been applied to prevent transmission of virus, even the transmission in household remains a critical problem especially in poor and developing countries with more burden family. \"Naso-oropharyngeal swab...\" Do authors mean both nasopharyngeal and oropharyngeal? Please elaborate. Response: yes, it refer to nasopharyngeal and oropharyngeal. \"The infection rate ranged from 37.5 to 100%\". You mean within families? Response: yes, this rate resembles the infection rate in the families. Revised sentence: Therefore, the infection rate in family ranged from 37.5% to 100%. \"Four breakthrough cases were detected...\" Do authors mean vaccine breakthrough? Response: it means COVID19 occur even after the individual received a vaccine. Revised sentence: Four vaccine breakthrough cases were detected in this study, which were infected by the virus from GH and GK clade. Introduction: \"D614G was the first reported mutation...\" This is technically an amino-acid substitution. Please, be precise. Response: it is an amino-acid substitution, thank you very much for the corrective comment; we will change this “mutation” in to “amino acid substitution” on whole manuscript. \"This mutation then increased and generated the emergence of the variants of concern\". How is this supported? This is scientifically unjustified. Response: we agree that mutation not always drive to the increased or generated novel voc, thus this sentence will be change in to “The mutation may increase and generate the emergence of the variants of concern” \"This study was performed to assess the variation of SARS-CoV-2 strains and their clinical manifestations within family clusters in Jakarta, Indonesia\". By this, the impression that of large-scale study is being given. However, 8 families were only investigated. This is not representative. How were these family clusters chosen? Was there any reason? There might be a bias in the selection of families/research material, with potential false community representation results. Response: samples on this study are from government program of COVID-19 tracking which RT-PCR diagnosis only taken place when symptoms are appears, thus it was limitations in this study. Materials and Methods: Details on the sequencing methodology and analysis are missing. Herein, sequencing was performed using \"Oxford Nanopore’s GridION sequencer\", with read depth lower than Illumina or Ion Torrent systems usings critical to verify that the identified mutations are not an \"artifact\" of the sequencing process.[P1] Response: The methods for nanopore sequencing were done exactly as the protocols described in the citations (Citations number 10 and 11) and according to manufacturers instructions. For the depth, as manufacturers suggestions and based on the citations, the sequencing was set to achieve 100x depth of coverage across all targets. Therefore, all the sequencing results only can be proceeded for further analysis if the 100x depth of coverage was achieved. We determined the sequence was good enough for further analysis because we used more depth than required (50x) and run only 10 specimens per nanopore’s flowcells to achieve our desired coverage. We will add this information to the manuscript for the confirmation. In addition, sequencing using illumina platform was not feasible in Indonesia for small project like ours during the period of our study, therefore we relied on Nanopore technology to support our country’s surveillance initiatives. Fig. 1: The relevant caption is missing several important pieces of information. What type of tree is this? Many trees are characterized as \"phylogenetic trees\" Details are needed on how this analysis was performed and the tree was constructed. This information needs to be added to the figure caption. The information provided on the branches is not easy to read and understand.[P2] Response: The phylogenetic tree was built using Geneious Prime 2021.1.1, using UPGMA as the tree build method and Tamura-Nei as the genetic distance model, with NC_045512 as the reference sequence. We had mentioned it into our manuscript. Tables: Table 1. This table presents Pango lineages (i.e., B.1) that were not anticipated as of January 2021. Table 2. It is not clear whether the authors mean in fact mutations, or substitutions by \"total N mutations\", \"total S mutations\", etc. Response: They refer to amino acid substitution within N or S protein. Table 3. This table also presents amino-acid substitutions, but they are named \"mutations\". Authors need to be precise. Response: Thank you very much for the corrective comments, the data presented in Table 3 are refer to amino acid substitutions, thus we will revise its “mutation” in to “amino acid substitution”." } ] } ]	1	https://f1000research.com/articles/11-886

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