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https://f1000research.com/articles/14-29/v1	06 Jan 25	{ "type": "Research Article", "title": "An Assessment of Professional Burnout within Hospital Pharmacists: A Case Study of Abu Dhabi", "authors": [ "Wadha Mohammed", "Beth Fylan", "Liz Breen", "Beth Fylan", "Liz Breen" ], "abstract": "Introduction Burnout is a syndrome resulting from unsuccessful stress management in the workplace, and is characterized by three dimensions: energy depletion, reduced professional efficacy, and feelings of negativism.\n\nBackground Studies have confirmed that burnout causes deterioration in teamwork and job satisfaction and negatively impacts and compromises services. Many studies have examined the impact of burnout on health.\n\nAim This cross-sectional study aimed to assess the rate of burnout among hospital pharmacists working in Sheikh Khalifa Medical City.\n\nMethod The Maslach Burnout Inventory-Human Services Survey for Medical Professionals was used to assess burnout. The survey measured three subscales: emotional exhaustion, depersonalization, and personal achievement. In addition, pharmacists were asked questions regarding their demographic years of practice and qualifications. The surveys were distributed to inpatients, discharge/emergency, outpatients, and clinical pharmacies. The data were collected over two weeks and analyzed.\n\nResults Sixty-eight of the 90 pharmacists participated in this study. The response rate was 75.5%. Seventeen pharmacists were considered to have high burnout as they had both high emotional exhaustion and high depersonalization scores. Two pharmacists were considered to have moderate burnout, as they had moderate emotional exhaustion, depersonalization, and personal achievement. Only 17 pharmacists reported a low level of burnout.\n\nConclusion The study revealed that 25% of the pharmacists had high burnout rates. Although statistically insignificant (P > 0.05), all pharmacists with high burnout were female and working in different pharmacy settings. Moreover, an additional 25% experienced low burnout, and 3% had moderate burnout levels. The remaining items were difficult to classify because they did not fit the Maslach definition of high, moderate, or low burnout. Further investigation is needed to identify the levels and causes of burnout among Sheikh Khalifa Medical City pharmacists.", "keywords": [ "Burnout", "Healthcare Workforce", "Occupational Stress", "Pharmacist Wellbeing." ], "content": "Introduction\n\nBurnout is a syndrome that results from unsuccessful stress management in the workplace.1 It is characterized by three dimensions: energy depletion, reduced professional efficacy, and feelings of negativity. Burnout terminology was first developed by American psychologist Herbert Freudenberger in the 1970s to describe the consequences of severe stress in helping professionals, such as social workers, nurses, and psychologists2,3 In 1982, Maslach defined burnout as a psychological syndrome related to emotional exhaustion, depersonalization, and a diminished sense of personal accomplishment among those who work with others in challenging situations.4 Maslach et al. (2018) described burnout dimensions in detail, where emotional exhaustion was defined as feeling tired, emotionally drained, lacking drive, and irritable due to work.4 Depersonalization is defined as distancing oneself during interpersonal contact with customers, clients, or patients, and feelings of negativism.3 Low personal accomplishment indicates a feeling of reduced competency and efficiency.4\n\nBurnout is included in the 11th revision of the International Classification of Diseases - 11 (ICD-11) as an occupational phenomenon; however, it is not considered a medical diagnosis.1 According to Padgett and Grantner (2020), burnout causes are difficult to isolate because of their individual-specific nature and wide-ranging symptoms.5 Workload is considered a burnout risk factor, particularly if job requirements exceed reasonable human limits.3 Other burnout risk factors include having no establishment of an individual role within the work environment and insufficient rewards or the absence of a reward system.5 In addition, having minimal opportunities for social interaction, unfair workplace decisions, and inconsistent motivation among workers are all well-known risk factors. Other risk factors contributing to burnout in the healthcare sector include the challenges of clinical work, performance metrics, time constraints, conflict of leadership, technology limitations, and competing demands.5 To date, no studies have assessed burnout prevalence among hospital pharmacists in the United Arab Emirates. Consequently, this study aimed to measure burnout levels among hospital pharmacists in Sheikh Khalifa Medical City, Abu Dhabi, using a valid and reliable data collection tool to establish non-biased results.\n\n\nBackground\n\nMany studies have shown the impact of burnout on the brain and health of those suffering from burnout.4 A study in which magnetic resonance imaging of 110 participants was analyzed, in addition to other psychological measures, concluded that burnout changed brain anatomy. Changes in the amygdala specifically affect negative emotion regulation in participants with high levels of burnout.4 Among Swedish healthcare workers, those with burnout reported anxiety, depression, neck and back pain, and memory impairment compared to Swedish healthcare workers without burnout.6 Another study confirmed a strong association between somatic symptoms, such as sleep disturbance and headache, in those reporting high degrees of emotional exhaustion. Furthermore, there is an elevated rate of general physical illnesses in people experiencing higher grades of burnout compared to those with lower stages of burnout.4 A study published by the American Psychological Association indicated that the impact of burnout on one’s health might be greater than expected.7 The researchers presented evidence supporting several mechanisms related to burnout and sickness, including metabolic syndrome, dysregulation of the hypothalamic-pituitary-adrenal axis, sympathetic nervous system activation, systemic inflammation, impaired immunity functions, blood coagulation, fibrinolysis, and poor health behaviors.7 Another study concluded that burnout is associated with physical and mental health problems such as hypertension, gastrointestinal disorders, and insomnia.8\n\nA systemic review conducted by Elbarazi et al. (2017), which evaluated 19 studies measuring burnout among healthcare professionals, claimed that burnout was assessed mainly among physicians, medical residents, nurses, social workers, surgeons, and physiotherapists, but not among pharmacists.9 The review included Middle Eastern countries such as Bahrain, Egypt, Jordan, Lebanon, Palestine, Saudi Arabia, and Yemen (9). This systematic review, which took place in 2017, indicated the need for future burnout research in Gulf countries to include a broader spectrum of healthcare professionals, such as dentists, dieticians, medical technicians, laboratory scientists, midwives, pharmacists, and radiographers.9\n\nThe availability of stress and burnout management materials may aid in reducing stress and burnout. A study evaluating burnout among hospital and health system pharmacists in North Carolina indicated that the decreased risk of burnout was associated with the availability and awareness of burnout resources.10\n\nThe American Pharmacists Association (APhA) and The American Society of Health-Systems Pharmacists (ASHP) provide resources for pharmacy teams to maintain pharmacy personnel’s well-being, resilience, and professional engagement. In addition to the programs offered by APhA and ASHP, pharmacy staff in the United States have access to a well-being index. The well-being index is a useful screening tool created by the Mayo Clinic to assess fatigue, depression, burnout, anxiety, stress, and mental/physical quality of life.11\n\n\nMethods\n\nThis study was conducted in accordance with the principles set forth in the Declaration of Helsinki. Ethical approval was obtained from the University of Bradford Ethics Committee (Approval Number: E987) on March 20, 2022, and from the SKMC Institutional Review Board (IRB) prior to data collection. A certificate of Good Research Practice from the United Kingdom Research and Innovation Medical Research Council (MRC) was also obtained and submitted to the respective committees. All participants provided written informed consent, were assured of confidentiality, and informed that participation was voluntary and could be withdrawn at any time without consequence.\n\nThis research was designed as a single-center, cross-sectional study conducted at Sheikh Khalifa Medical City (SKMC) in Abu Dhabi, United Arab Emirates. SKMC is a Joint Commission International-accredited hospital with a 560-bed capacity and provides both inpatient and outpatient services. It is one of the Abu Dhabi Health Services (SEHA) hospitals.12,13 The study was conducted over a two-week period starting on June 20, 2022.\n\nPharmacists working at SKMC were the target population. To be eligible for inclusion, participants had to be licensed pharmacists currently employed at SKMC in any pharmacy practice setting (inpatient, outpatient, discharge/emergency, behavioral sciences pavilion, clinical pharmacy, or pharmacy administration). Exclusion criteria included pharmacy technicians, residents, clerks, interns, or any non-pharmacist staff. No age or gender restrictions were applied beyond professional licensure as a pharmacist.\n\nA recruitment e-mail was sent to pharmacy managers who then informed their teams about the study. The primary researcher subsequently visited each pharmacy area, provided a brief verbal explanation of the study’s purpose, scope, and objectives, and distributed the surveys in sealed envelopes along with participant information sheets. Participation was entirely voluntary and pharmacists were given two weeks to complete the survey. Completed surveys were returned in sealed envelopes to a secure collection box to maintain anonymity.\n\nThis study aimed to assess burnout among all hospital pharmacists employed at Sheikh Khalifa Medical City (SKMC), Abu Dhabi. The total population of eligible participants was 90 licensed pharmacists working across various pharmacy practice settings within SKMC. Given the finite and manageable population size, a census approach was employed, wherein every eligible pharmacist was invited to participate in the study. This approach eliminates the need for sampling and ensures that the findings are fully representative of the entire population of hospital pharmacists at SKMC. Consequently, the response rate of 75.5% (68 out of 90 pharmacists) is considered adequate to provide reliable estimates of burnout prevalence and its association with demographic factors within this specific setting. Non-participation among some pharmacists was primarily attributed to reported time constraints and personal reluctance to engage in research activities. Our analysis methods directly reflect the census approach used, analyzing the entire eligible pharmacist population at SKMC without the need for sampling adjustments. This straightforward approach ensured that the results accurately represented the target demographic.\n\nData were collected using a validated instrument, the Maslach Burnout Inventory – Human Services Survey for Medical Personnel (MBI-HSS MP),4 along with supplementary demographic questions. The MBI-HSS MP consists of 22 items measuring three burnout dimensions: emotional exhaustion (9 items), depersonalization (5 items), and personal accomplishment (8 items). The MBI-HSS MP questionnaire is the most frequently used questionnaire in the medical research literature to assess burnout in healthcare professionals.14 Responses are recorded on a 7-point Likert scale (0 = never; 6 = every day). The survey typically requires 10–15 minutes to complete and no specialized knowledge is required.\n\nTo ensure a comprehensive understanding of participant characteristics, demographic questions included age, gender, marital status, highest pharmacy qualification (Bachelor of Pharmacy or PharmD), years of professional practice at SKMC, and current pharmacy practice setting. Surveys were administered in hard copy format due to limited internet access and to prevent technical issues that could arise with an online format. Participants sealed their completed questionnaires in envelopes to ensure confidentiality.\n\nFor emotional exhaustion and depersonalization, higher scores indicate greater burnout, while for personal accomplishment, lower scores indicate greater burnout.4 According to Maslach et al. (2018), emotional exhaustion scores ≤18 represent low burnout, 19–26 moderate, and ≥27 high burnout. Depersonalization scores ≤5 represent low burnout, 6–9 moderate, and ≥10 high burnout. Personal accomplishment scores ≤33 represent low burnout, 34–39 moderate, and ≥40 high.15 Participants were classified as having a “high level of burnout” if they had both high emotional exhaustion and high depersonalization scores, regardless of their personal accomplishment scores. This approach is based on previous research indicating that the personal accomplishment dimension may have lower reliability in predicting burnout.15 See Table 1.\n\nAbbreviations: EE, emotional exhaustion; DP, depersonalization; PA, personal accomplishment.\n\nAll data were entered into and analyzed using the Statistical Package for the Social Sciences (SPSS) version 28. Descriptive statistics (frequency, percentage, mean, and standard deviation) were used to characterize the demographic variables and burnout dimensions. Inferential statistics, including univariate analysis, Spearman’s rho correlation, and multiple regression, were performed to explore associations between demographic factors and high burnout levels. To account for the distribution of participants across different pharmacy settings, practice settings were included as independent variables in the multiple regression analysis. This adjustment ensures that potential differences in burnout levels among various pharmacy environments are considered, thereby reducing confounding effects related to the diverse work settings.\n\n\nResults\n\nThe MBI-HSS MP questionnaire was distributed at five pharmacy sites and a pharmacy administrative office (the pharmacy sites involved were an outpatient pharmacy, inpatient pharmacy, discharge pharmacy, behavioral science pavilion pharmacy, and clinical pharmacy).\n\nSixty-eight pharmacists participated in the study out of ninety pharmacists, with a response rate of 75.5%. Twenty-five pharmacists were from discharge, behavioral science pavilions, and emergency department pharmacies. This number is amalgamated as pharmacists rotate between pharmacy sites and are managed by the same pharmacy supervisor. Seventeen pharmacists were from inpatient pharmacies, 12 from clinical pharmacies, 11 from outpatient pharmacies, and the remaining three were from pharmacy administrations. Since the pharmacists practicing in emergency departments rotate between discharge pharmacies and behavioral science pavilion pharmacies, they account for most of the participating pharmacists (36.8%), followed by inpatient pharmacists (25%).\n\nThe majority of the participants were thirty-one to forty-five years old (48.5%), female (39%), and married (76.5%). The questionnaire showed that 4% revealed that their highest educational qualification was either a master’s degree or a diploma. The remaining 20% did not provide their educational level and left it blank. To address these missing data, analyses were conducted with and without these cases to check for consistency in the results. Additionally, such missing responses were excluded from statistical tests involving educational level to prevent any bias. Furthermore, most of the participants had practiced for approximately five years or less (41.2%). This was followed by six to fifteen years of practice experience (35.3%). See Table 2 for the demographic characteristics.\n\nAbbreviations: N/A, not available; ER, emergency; BSP, behavioral sciences pavilion; SKMC, Sheikh Khalifa Medical City.\n\nA breakdown of each MBI subscale, with the corresponding number of pharmacists who responded to a high, moderate, or low score, is shown in Table 3. The mean MBI-HSS MP score indicated a moderate level of emotional exhaustion (M = 22), depersonalization (M = 7), and personal accomplishment (M = 38). The majority of pharmacists had low emotional exhaustion (n = 30), low depersonalization (n = 32), and a high sense of personal accomplishment (n = 34). Furthermore, 15 pharmacists (22.1%) were identified as having a moderate level of emotional exhaustion, 14 pharmacists (20.6%) were found to have a moderate level of depersonalization, and 12 pharmacists (17.6%) had a moderate sense of personal accomplishment. Twenty-three (33.8%) and twenty-two (32.4%) pharmacists had high emotional exhaustion and depersonalization scores, respectively. While only twenty-two (32.4%) pharmacists were identified as having a low level of personal accomplishment.\n\nAbbreviation: MBI, Maslach Burnout Inventory.\n\nBased on the definition of burnout, 17 pharmacists were considered to have high burnout (n = 17/68), as they had both high emotional exhaustion and high depersonalization scores. Moreover, 17 pharmacists were considered to have low burnout (n = 17/68) as they had both low emotional exhaustion and low depersonalization scores. Personal accomplishment scores were excluded when determining whether a pharmacist has high burnout, as a lower personal accomplishment subscale has low reliability in predicting burnout, as mentioned earlier by Kang and colleagues (2020).15\n\nAll pharmacists with high burnout were female, 16 of whom were educated to a degree level and one pharmacist did not reveal their education level. More than half of those identified as having high burnout were from outpatient pharmacies (n = 11) and the remaining were from discharge (n = 2), inpatient pharmacies (n = 2), and clinical pharmacies (n = 2). Nine pharmacists who were identified to have high burnout were married, two were divorced, and six were single. The minimum years of experience in SKMC for those with high burnout was one year and the highest number of years of experience as a pharmacist in SKMC was 19 years. Seventeen married pharmacists and one single pharmacist had low burnout because they had low emotional exhaustion, low depersonalization, and a high sense of personal achievement or satisfaction with their personal accomplishment, keeping in mind that the majority of the study population was married (76.5%). Two of the participating pharmacists had moderate burnout, emotional exhaustion, depersonalization, and personal accomplishment (married and divorced). Univariate analysis revealed that none of the factors (age, sex, marital status, practice settings, and years of practice) significantly affected the level of burnout (p > 0.05). See Table 4.\n\nSpearman’s rho was used to identify the correlation between the study variables and high burnout. It revealed that there is a weak negative association between marital status and having high burnout, and between gender and high burnout, as well as between practice settings and burnout (see Table 5).\n\n* Correlation is significant at the 0.05 level (2-tailed).\n\nAlthough a weak negative association was identified, the correlation between all variables and high burnout was not significant (p > 0.05). Multivariate regression analysis was used to identify how study variables affect the level of burnout. None of the variables significantly affected high levels of burnout. Refer to Table 6.\n\n\nDiscussion\n\nThe aim of this study was to assess burnout presence/levels among SKMC pharmacists using the validated MBI-HSS (MP). The results of this study showed that 17 pharmacists had a high burnout status; all of those pharmacists were females. Based on the statistical analysis, there was no significant relationship between being female and high burnout. Previous studies failed to find a significant correlation between gender differences and burnout rates15; however, this study confirmed that female medical residents have higher burnout levels than their male colleagues. Burnout was assessed among hospital and health-system pharmacists in North Carolina, and it was revealed that more than half of the pharmacists were at a high risk of burnout. Being female was among the three factors significantly associated with increased burnout among pharmacists with high burnout and working in hospitals and health system pharmacists in North Carolina.15\n\nIn 2004, a MONICA study (Monitoring of Trends and Determinants in Cardiovascular Disease (MONICA) study was conducted in northern Sweden to assess the prevalence of burnout. Researchers have used the (SMBQ) as a burnout instrument to evaluate burnout among active workers. The study concluded that women had higher burnout levels than men. According to Norlund et al. (2010), the difference in burnout levels between each gender is explained by work-related stressors and situational life factors, where the level of education, socioeconomic status, work object, and varying working hours are the main factors affecting female tolerance to burnout.16\n\nA recent study during the COVID-19 pandemic supported the statement of different burnout levels among both sexes, where female workers had a higher burnout rate. A previous study conducted in Kuwait by Alsairafi et al. (2021) showed that the prevalence of severe depression is higher among females. The authors explained that females were at a higher risk of severe depression and anxiety due to hormonal fluctuation.17\n\nMoreover, a study published by Alanazi et al. (2020) assessing burnout showed that burnout was higher in all parameters in females than in males.18 Another cross-sectional study that included hundred and two pharmacists in Saudi Arabia to evaluate burnout among pharmacists during the COVID-19 pandemic revealed that 59.1% of pharmacists were categorized as having burnout. Univariate analysis revealed that burnout levels were significantly higher among younger female pharmacists.19 Studies from Montreal University have disclosed an increasing gender gap in workplace burnout. Additionally, data from the mental well-being platform illustrated that female workers are 23% more likely than men to struggle with poor work-life balance and 45% more likely to suffer from work stressors. Bowling (2022) explained that an unequal share of work-life tasks, having less time for self-care, and being less promoted compared to males contributes to a higher prevalence of burnout in women.20\n\nAlthough all previous studies concluded that females had a higher level of burnout compared to males, an analysis of seventy-eight studies among nurses revealed that male nurses had a higher rate of burnout in general and a higher rate of depersonalization compared to female nurses. This affects the quality of patient services. Additionally, it negatively affected their attitudes toward their colleagues.21\n\nThis study did not find a significant association between burnout among SKMC pharmacists and different age groups. However, 11 pharmacists with high emotional exhaustion and high depersonalization were between thirty and forty-five years old. Seventeen pharmacists had low burnout, low emotional exhaustion, low depersonalization, and high satisfaction with their personal accomplishments. Excluding three pharmacists, two of them were more than forty-five years old and one pharmacist did not reveal their age; thus, confidentiality will not be breached, and identity will remain anonymous.\n\nThere are various published data on age and burnout. According to a new survey, COVID-19 worsened employees’ feelings of burnout, especially those working from home, due to prolonged working hours.22 The average age of workers who reported a higher level of exhaustion and burnout was thirty-two years old.22 During the COVID-19 pandemic, one study reported a higher burnout rate in those aged twenty-nine years old and younger with chronic conditions compared to other healthcare professionals without chronic conditions from the same age group.17 The same study concluded that, in general, the incidence of burnout was higher among females aged < 50 years. At the same time, a study from Saudi Arabia reported a higher incidence of burnout during COVID-19 among those younger than forty years old.18\n\nFurthermore, a study evaluating the relationship between age and burnout in Canadian workers showed that burnout symptoms differed significantly according to the different life stages of working females and males. Males and females aged between twenty to thirty-five and fifty-five years and older were at a higher risk for burnout.23 Moreover, among pharmacists working in different Singaporean hospitals, the burnout rate was higher among young pharmacists aged thirty-two years old and less than those thirty-five years and older.24\n\nIn this study, the majority of pharmacists reporting high burnout were married (53%), and there was no significant relationship between being married and high burnout. Furthermore, most pharmacists with low burnout were married. This can be explained by the fact that the majority of participants in this study were married (76.5%). Similarly, many studies have claimed that marital status is unrelated to burnout dimensions. For instance, a survey to check community pharmacist resilience and burnout in Lebanon found that being married or single was not significantly related to burnout and resilience.25 Another study assessing burnout among hospital pharmacists in Qatar claimed that marital status was unrelated to burnout.26 On the other hand, a higher level of burnout was reported in unmarried hospital pharmacists and in unstable relationships in the United States. Moreover, single pharmacists working in Singapore reported a more elevated level of burnout, and in Turkey, it was established that marital status strongly affects burnout and resilience.27 In addition, a published study assessing burnout among nurses confirmed that marital status is strongly related to a high level of burnout, with unmarried nurses being the most affected.28\n\nParticipants were not asked about their working hours, as all pharmacists working in SKMC worked for eight hours, in addition to one hour of mandatory breaks. The main difference was in the evening and night shifts. There was no statistically significant correlation between practice settings and burnout among pharmacists working in SKMC. Different published papers have demonstrated that long work hours are a risk factor for psychological and work-related stress. Working ten hours or more per day, 40 hours of overtime per month, and 60 hours or more per week increased stress levels.29 A study conducted in one of the Taiwanese medical centers among healthcare workers revealed a nonlinear association between working hours and burnout. The odds ratio was doubled in those hours exceeding 60 hours compared to those working 40 hours per week, tripled when hours exceeded seventy-four, and quadrupled when hours exceeded eighty-four.30\n\nHospital pharmacists generally work eight hours per day, while retail pharmacists work around 12–14 hours per day.31 In contrast, National Health Service hospital pharmacists work around thirty-seven and half hours a week, and additional hours may be required in specific situations. Hospital pharmacists may be part of an on-call rota.32 The Manitoba Pharmaceutical Association (2004) confirmed that one of the pharmacy department’s responsibilities is to provide clinical drug information services—i.e., non-laboratory, advisory support to healthcare professionals regarding medications—twenty-four hours a day, seven days a week.33 As a result, when staffing is unavailable after working hours, on-call pharmacists usually provide this service. As per the study by Padgett and Grantner (2020), it was confirmed that pharmacists working more than forty hours per week had a higher burnout level than those working less than forty hours per week.5\n\nThis study did not evaluate whether the number of dispensed items and prescriptions were related to higher burnout among SKMC pharmacists. The number of patients seen by outpatient pharmacists decreased during the COVID-19 pandemic, and even after COVID-19, the hospital was designated as a COVID-19 hospital, and patients were followed up in other non-COVID-19 hospitals. Nowadays, outpatient pharmacists verify hundred and thirty prescriptions per month with an average of thousand and four hundred items. Inpatient pharmacists verify approximately 70 orders per hour per shift and approximately fifty thousand medications are verified per month. Based on studies by Peat et al. (2022) and Bookwalter (2021), an increased volume of daily prescriptions is associated with higher burnout among community pharmacists.11,34\n\nWhile trying to assess the undergraduate level, four pharmacists left the question blank, nine pharmacists answered yes or no, and nine pharmacists answered higher or lower educational level. All pharmacists identified as having a high burnout were bachelor’s degree holders, except for one who did not reveal the educational level. In a study that evaluated burnout among hospital and health system pharmacists in North Carolina, 95% of the pharmacists had a graduate level degree and only forty-nine (13.7%) had an additional advanced degree other than Pharm D. However, education level, additional advanced degree, or obtaining certification through the Board of Pharmacy Specialties did significantly affect burnout subscales.15\n\nRecent studies that assessed burnout, depression, and anxiety during the COVID-19 pandemic did not correlate this with healthcare professionals’ education level or examine the relationship between educational level and burnout or depression subscales.17,18,35 However, the degree of burnout assessed among graduate pharmacists of Mercer University Southern School of Pharmacy from 1973 to 1983 showed that pharmacists holding a Pharm D degree experienced a lower degree of burnout than those holding a bachelor’s degree only.36 A study assessing the impact of burnout on the academic achievement of Saudi female students enrolled in the College of Health Sciences confirmed that there was no association between burnout and academic achievement. There was no relationship between burnout and academic level of study. Students of pharmacy colleges were the only students among all other health sciences colleges to suffer from high burnout.37 Moreover, a study assessing the job satisfaction among hospital pharmacists in Tikur Anbesa specialized hospital in Addis Ababa revealed that almost half of the pharmacists were poorly satisfied with their current job thinking that their formal education overqualified them for the current job they are practicing.38\n\nAdditionally, the Abu Dhabi Health Services Company (SEHA), including its hospitals, is undergoing privatization. Privatization and changing the pharmacist’s contracts, including the housing and end of services, are of concern for many pharmacists, and this leads to job insecurity. A study conducted in Turkey by Aybas et al. (2015) confirmed an association between job insecurity and burnout, which is worth further investigation among SKMC and SEHA pharmacists.39\n\nThe correlation between years of experience and high burnout was not significant. Among pharmacists identified as having high burnout, there were pharmacists who worked for more than 15 years and pharmacists who worked in SKMC for less than five years. To date, no study has specifically examined the correlation or association between years of experience and level of burnout. However, a study conducted among teachers of special education revealed that years of work experience is an important predictor of burnout and is significantly correlated with exhaustion and depersonalization.40 A meta-analysis conducted by Brewer and Shapard (2004) illustrated that prolonged years of experience are associated with a lower level of burnout among employees working in some fields in the United States, including social workers, educators, nurses, psychologists, and mental health workers. Additionally, a small negative correlation between years of experience in the field and emotional exhaustion has been identified.41\n\nMany preventative strategies suggested by Padgett and Grantner (2020) can be followed to reduce the risk of burnout in those with a low level of burnout, including encouraging a work-life balance by offering a flexible working schedule, enhancing organizational promotion and rewarding systems, defining wellness and burnout to the team, and communicating plans to the team. Furthermore, the same resource encourages pharmacists with high burnout levels to seek help and personal care, including emotional health, fitness, and nutrition.5\n\nAt the organizational level, in addition to offering a wellness program by the SKMC psychologist team, a progressive strategy to mitigate employees in general and pharmacists in specific needs to be implemented. In Stanford Medicine in California, a position was created years ago, called the chief wellness officer.42 As per Flemings and Desselle (2021), the chief wellness officer’s responsibility is to create initiatives to combat physician burnout; the same approach can be adopted toward combat burnout in SKMC.42 Additionally, SKMC can consider expanding or implementing access to mental health and self-care resources other than stress management resources as part of annual compliance exams and increasing the number of seats in the wellness program so that more pharmacists and SKMC healthcare professionals can join.\n\nSeveral limitations were noted in the completion of this study, which are discussed below. Access to an electronic version of the survey and modifying the survey layout were noted as desirable by many SKMC pharmacists (as opposed to the paper-based survey). The use of the online survey may have prompted greater engagement rates and fuller survey completion. Despite piloting the survey, at least two pharmacists said that the survey language was not easy and that some of the words used in the survey were difficult for them, so they struggled to respond. The last limitation noticed after conducting the study was that three inpatient pharmacists commented either on the questionnaire or verbally that questions that specify patients are not applicable in inpatient pharmacy settings. This was not considered and, as such, should be factored into future surveys.\n\nFurther investigation and research are required to assess the rate and prevalence of burnout among pharmacy residents, pharmacy technicians, pharmacy store workers, and pharmacy interns in SKMC, Abu Dhabi, and the U.A.E in general, as well as to identify the causes of burnout such as having more evening or night shifts, being on-call 24/7, delayed promotion, impact of privatization, and lack of a reward system.\n\n\nConclusion\n\nBurnout is a psychological syndrome related to emotional exhaustion, depersonalization, and a diminished sense of personal accomplishment.4 Burnout has been assessed among hospital pharmacists in many Gulf countries but not in the United Arab Emirates. This study is the first to evaluate burnout among hospital pharmacists working in SKMC and SEHA. The study used the MBI-HSS for medical professionals to assess burnout among SKMC pharmacists. Sixty-eight pharmacists participated in the study, with a response rate of 75.5%. 25% of pharmacists working at SKMC had high burnout rates. Although statistically insignificant (P > 0.05), all pharmacists with high burnout were female and working in different pharmacy settings. Moreover, an additional 25% experienced low burnout, and 3% had moderate burnout levels. The remaining items were difficult to classify because they did not fit the Maslach definition of high, moderate, or low burnout. Moreover, marital status, age, and level of education were statistically insignificant in relation to burnout level. The research methodology did not allow the identification of the causes of burnout, and further investigation is required to identify burnout causes among SKMC pharmacists. Moreover, assessing burnout among pharmacy residents, interns, pharmacy store workers, and technicians is another area for future research.\n\nEthical approval for this study was granted by the University of Bradford Ethics Committee (Approval Number: E987) on March 20, 2022, and by the Institutional Review Board (IRB) at the SKMC study site prior to data collection. Additionally, a Certificate of Good Research Practice was obtained from the United Kingdom Research and Innovation Medical Research Council (MRC) and submitted to the respective committees. All participants provided written informed consent, were assured of confidentiality, and were informed that their participation was voluntary and could be withdrawn at any time without consequence.\n\nAll participants provided written informed consent prior to their inclusion in the study. The consent process was approved by the University of Bradford Ethical Committee, and the consent procedure was waived by the ethics committee for this study. Participants were assured of the confidentiality and anonymity of their responses.\n\n\nAuthor contributions\n\nWadha Mohammed, Beth Fylan, and Liz Breen contributed equally to the study design, data collection, analysis, and writing of the manuscript. All authors reviewed and approved the final version of the manuscript.", "appendix": "Data availability\n\nThe underlying data for this study were de-identified to protect participant confidentiality and are available in the Zenodo repository at https://doi.org/10.5281/zenodo.14497075.43 The dataset, titled Data for ‘An Assessment of Professional Burnout within Hospital Pharmacists: A Case Study of Abu Dhabi’, includes raw, de-identified responses collected using the Maslach Burnout Inventory - Human Services Survey for Medical Professionals, encompassing measures of emotional exhaustion, depersonalization, and personal achievement, as well as demographic information such as years of practice and qualifications. The dataset supports the findings presented in the manuscript and adheres to Safe Harbor de-identification guidelines to ensure participant confidentiality.\n\nData are available under the terms of the (Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).\n\nThe extended data for this study are available in a Zenodo repository at https://doi.org/10.5281/zenodo.14497115.44 The file (Extended Data.pdf ) contains the Hospital Pharmacist Burnout Assessment Survey form, which includes:\n\n• Demographic Information Section: Questions on participants’ age, gender, marital status, qualifications, and years of experience.\n\n• Survey Sections: The Maslach Burnout Inventory–Human Services Survey (MBI-HSS MP) items with a 0–6 frequency scale for assessing burnout dimensions (emotional exhaustion, depersonalization, and personal achievement).\n\nThis form adheres to Safe Harbor de-identification standards and is shared under a CC0 1.0 license.\n\nThis study is an observational, cross-sectional study, and it has been reported in adherence to the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) guidelines. We have ensured that all relevant STROBE checklist items—such as specifying the study design in the title and abstract, clearly describing the setting, participants, variables, data sources, and statistical methods—have been met, check the STROBE complete checklist at https://doi.org/10.5281/zenodo.14522814.45\n\n\nReferences\n\nWorld Health Organization: Burn-out an “occupational phenomenon”: International Classification of Diseases.2019, May 28. Retrieved June 5, 2024. Reference Source\n\nSouthern New Hampshire University: A list of helping professions\|SNHU.n.d. Retrieved June 5, 2024. Reference Source\n\nMaslach C, Leiter MP: Understanding the burnout experience: Recent research and its implications for psychiatry. World Psychiatry. 2016; 15(2): 103–111. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGarden M: Manual 4th Edition (printed for shipping) - Maslach Burnout Inventory.n.d. Retrieved June 5, 2024. Reference Source\n\nFlorida EHP, Grantner GR: Pharmacist Burnout and Stress. U.S. Pharmacist. 2020. Retrieved June 5, 2024. Reference Source\n\nPeterson U, Demerouti E, Bergström G, et al.: Burnout and physical and mental health among Swedish healthcare workers. J. Adv. Nurs. 2008; 62(1): 84–95. PubMed Abstract \| Publisher Full Text\n\nMelamed S, Shirom A, Toker S, et al.: Burnout and risk of cardiovascular disease: Evidence, possible causal paths, and promising research directions. Psychol. Bull. 2006; 132(3): 327–353. PubMed Abstract \| Publisher Full Text\n\nChiu M-C, Wang M-J, Lu C-W, et al.: Burnout and workload among health care workers: The moderating role of job control. Saf. Health Work. 2015; 6(4): 289–294. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nElbarazi I, Loney T, Yousef S, et al.: Prevalence of and factors associated with burnout among health care professionals in Arab countries: A systematic review. BMC Health Serv. Res. 2017; 17: 491. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKang K, Absher R, Granko RP: Evaluation of burnout among hospital and health-system pharmacists in North Carolina. Am. J. Health Syst. Pharm. 2020; 77(6): 441–448. PubMed Abstract \| Publisher Full Text\n\nCarolina CMB: Challenges in community pharmacy during COVID-19: The perfect storm for personnel burnout. U.S. Pharmacist. 2020. Retrieved June 5, 2024. Reference Source\n\nSEHA: Hospitals.n.d. Retrieved June 5, 2024. Reference Source\n\nTaha H, Raji SJ, Khallaf A, et al.: Improving Catheter Associated Urinary Tract Infection Rates in the Medical Units. BMJ Qual. Improv. Rep. 2017; 6(1): u209593.w7966. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRomani M, Ashkar K: Burnout among physicians. Libyan J. Med. 2014; 9(1): 23556. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKang K, Absher R, Granko RP: Evaluation of burnout among hospital and health-system pharmacists in North Carolina. Am. J. Health Syst. Pharm. 2020; 77(6): 441–448. PubMed Abstract \| Publisher Full Text\n\nNorlund S, Reuterwall C, Hörneland J, et al.: Burnout, working conditions and gender - results from the Northern Sweden MONICA study. BMC Public Health. 2010; 10: 326. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAlsairafi Z, Naser AY, Alsaleh FM, et al.: Mental health status of healthcare professionals and students of health sciences faculties in Kuwait during the COVID-19 pandemic. Int. J. Environ. Res. Public Health. 2021; 18(4): 2203. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAlharbi M, Alshehry A, Alnuhait M, et al.: Prevalence and risk factors of burnout among healthcare professionals during COVID-19 pandemic - Saudi Arabia. Am. J. Public Health Res. 2021; 9(1): 13–19. Reference Source\n\nAlsairafi Z, Naser AY, Mohamad AM, et al.: Cross-sectional study to evaluate burnout among pharmacy staff in Saudi Arabia during COVID-19 pandemic. Saudi Pharm. J. 2022. Reference Source\n\nNuffield Health: Why women are more at risk of burnout than ever before.n.d. Retrieved June 5, 2024. Reference Source\n\nCañadas-De la Fuente GA, Ortega E, Ramirez-Baena L, et al.: Gender, marital status, and children as risk factors for burnout in nurses: A meta-analytic study. Int. J. Environ. Res. Public Health. 2018; 15(10): 2102. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nStudyFinds: Survey: The average worker experiences career burnout - by the age of 32!.n.d. Retrieved June 5, 2024. Reference Source\n\nMarchand A, Blanc ME, Beauregard N: Do age and gender contribute to workers’ burnout symptoms? Occup. Med. 2018; 68(6): 405–411. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTan ELY, Lun CP, Jing LW, et al.: Burnout and resilience among pharmacists: A Singapore study. Journal of the American College of Clinical Pharmacy. 2022; 5: 75–84. Publisher Full Text\n\nAlameddine M, Bou-Karroum K, Hijazi MA: A national study on the resilience of community pharmacists in Lebanon: A cross-sectional survey. J. Pharm. Policy Pract. 2022; 15(1): 8. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEltorki Y, Abdallah O, Riaz S, et al.: Burnout among pharmacy professionals in Qatar: A cross-sectional study. PLoS One. 2022; 17(5): e0267438. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCalgan Z, Aslan D, Yegenoglu S: Community pharmacists’ burnout levels and related factors: An example from Turkey. Int. J. Clin. Pharm. 2011; 33(1): 92–100. PubMed Abstract \| Publisher Full Text\n\nWei H, Kaur R, Khong LW: Study on the relationships between nurses’ job burnout and subjective well-being. Chin. Nurs. Res. 2015; 2(2–3): 61–66. Publisher Full Text\n\nLi G, Cong X, Zhang X, et al.: The effect of long working hours and overtime on occupational health: A meta-analysis of evidence from 1998 to 2018. Int. J. Environ. Res. Public Health. 2019; 16(12): 2102. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLin R, Lin Y, Hsia Y, et al.: Long working hours and burnout in health care workers: Non-linear dose-response relationship and the effect mediated by sleeping hours—A cross-sectional study. J. Occup. Health. 2021; 63(1): e12228. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMonster: 5 Reasons Pharmacists Love Their Jobs.n.d. Retrieved June 5, 2024. Reference Source\n\nProspects: Hospital pharmacist job profile.n.d. Retrieved June 5, 2024. Reference Source\n\nThe Manitoba Pharmaceutical Association: Standards of Practice Hospital.2004. Retrieved June 5, 2024. Reference Source\n\nPeat G, Olaniyan J, Fylan B, et al.: Mapping the resilience performance of community pharmacy to maintain patient safety during the Covid-19 pandemic. Res. Soc. Adm. Pharm. 2022; 18(9): 3534–3541. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNn A, Aljasmi S, Ra A, et al.: Psychological Impact of the COVID19 Pandemic on Healthcare Employees in Abu Dhabi Health Services Company (SEHA). [Conference presentation abstract or other]. (Insufficient information for a full APA citation provided.).2020, December 11.\n\nBarnett CW, Hopkins WA, Jackson RA: Burnout experienced by recent pharmacy graduates of Mercer University. Am. J. Hosp. Pharm. 1986; 43(11): 2780–2784. PubMed Abstract \| Publisher Full Text\n\nAlshobaili AM, Alruwaili SH, Alshallan HA, et al.: The impact of burnout on the academic achievement of Saudi female students enrolled in the colleges of health sciences. Int. J. High. Educ. 2021; 10(2): 229–239. Publisher Full Text\n\nMesfin Y, Kibret GD, Teshome D, et al.: Assessment of job satisfaction among pharmacy professionals. J. Pharm. Policy Pract. 2021; 14: 71. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAybas M, Elmas-Atay S, Dündar G: Job insecurity and burnout: The moderating role of employability. Eur. J. Bus. Manag. 2015; 7(9): 195–203.\n\nDuli S: Years of work experience, an important predictor of burnout in special education. Am. Sci. Res. J. Eng. Technol. Sci. 2016; 17(1): 318–322. Reference Source\n\nBrewer EW, Shapard L: Employee burnout: A meta-analysis of the relationship between age or years of experience. Hum. Resour. Dev. Rev. 2004; 3(2): 102–123. Publisher Full Text\n\nFlemings E, Desselle S: Learn to cope with burnout in the pharmacy. Pharm. Times. 2021, December 21. Retrieved June 5, 2024. Reference Source\n\nMohammed W, Fylan B, Breen L: Data for ‘An Assessment of Professional Burnout within Hospital Pharmacists: A Case Study of Abu Dhabi’. [Dataset]. Zenodo. 2024. Publisher Full Text\n\nMohammed W: Extended Data for ‘An Assessment of Professional Burnout within Hospital Pharmacists: A Case Study of Abu Dhabi’. Zenodo. 2024. Publisher Full Text\n\nMohammed W: STROBE Checklist for “An Assessment of Professional Burnout within Hospital Pharmacists: A Case Study of Abu Dhabi”. Zenodo. 2024. Publisher Full Text" }	[ { "id": "357197", "date": "11 Jan 2025", "name": "Mariya Ivanova", "expertise": [ "Reviewer Expertise occupational burnout among pharmacists", "pharmaceutical care", "organization and economics of pharmacy" ], "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\nAn Assessment of Professional Burnout within Hospital Pharmacists: A Case Study of Abu Dhabi\nBurnout has significant impacts on both physical and mental health.\nBrain Changes: Burnout alters brain anatomy, particularly the amygdala, impairing negative emotion regulation. Health Impacts: Burnout is linked to anxiety, depression, memory impairment, neck/back pain, sleep disturbances, headaches, hypertension, gastrointestinal disorders, and insomnia. It also increases the risk of physical illnesses due to mechanisms like metabolic syndrome, immune dysfunction, and systemic inflammation.\nBurnout in Healthcare Workers: Studies highlight burnout among physicians, nurses, and other healthcare professionals but note a lack of research on pharmacists, especially in Middle Eastern countries, prompting calls for broader studies.\nBurnout Management: Resources and awareness significantly reduce burnout risks. In the U.S., organizations like the American Pharmacists Association (APhA) and the American Society of Health-System Pharmacists (ASHP) offer well-being programs and tools, such as the Mayo Clinic’s well-being index, to assess and manage burnout-related issues.\nThe study aimed to assess burnout among SKMC pharmacists using the MBI-HSS (MP). Key findings include:\nHigh Burnout Prevalence: 25% of pharmacists experienced high burnout, predominantly females. However, the correlation between gender and burnout was statistically insignificant. Past studies show mixed results on gender differences in burnout, though females often reported higher burnout levels due to factors like work stress, hormonal fluctuations, and unequal work-life balance.\nAge and Burnout: Burnout was common among pharmacists aged 30-45, but no significant correlation was found between age and burnout. Studies elsewhere suggest higher burnout among younger professionals and those under 40, especially during COVID-19.\nMarital Status: Most participants were married, but marital status showed no significant link to burnout. Other studies have yielded mixed results, with some linking higher burnout to being unmarried.\nWork Hours: Long working hours and night shifts are known risk factors for burnout. Although SKMC pharmacists work standard hours, burnout risks may still arise from work volume and shift patterns.\nEducational Level: All pharmacists with high burnout held bachelor’s degrees, though education level was not significantly correlated with burnout. Some studies suggest that higher qualifications like Pharm D may mitigate burnout.\nConclusion: Burnout is prevalent among SKMC pharmacists, with emotional exhaustion and depersonalization being key issues. Gender, age, and marital status were not statistically significant predictors. Further investigation is needed to identify specific causes and extend research to other pharmacy professionals in the UAE.\n\nAdditional comments:\n1. The study would be more accurate if the introduction specifies whether there have been other studies on professional burnout in the country. 2. For Table 1, it would be better to propose an alternative way of presenting the data. 3. The scale for low, medium, and high values is indicated as developed by Maslach, but the source is different. Is the scale according to Maslach or tailored for a population of hospital pharmacists? 4. Under Table 3, the source is cited as Kang and colleagues (2020), but a more accurate reference would be: Garden M: Manual 4th Edition (printed for shipping) - Maslach Burnout Inventory. n.d. Retrieved June 5, 2024. Reference Source. 5. To align the title with the data, the study should not include individuals without pharmaceutical education.\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?\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/14-29
https://f1000research.com/articles/13-1318/v1	05 Nov 24	{ "type": "Research Article", "title": "Types of refractive errors in a sample of Iraqi children with Intermittent exotropia", "authors": [ "Najah K. Mohammad", "Ibrahim Ali Rajab", "Ibrahim Ali Rajab" ], "abstract": "Background One of the most common strabismus types in children is intermittent exotropia, which masks the deviation of one eye outward, mostly in children aged 2-4 years. It may affect visual development and often coexists with refractive mistakes. Unlike esotropia, which usually links to hyperopia, intermittent exotropia might have a different relationship with refractive errors and thus lead to myopia.\n\nMethods In this cross-sectional study from August 2021 to December 2023, 179 patients diagnosed with intermittent exotropia were recruited via an outpatient clinic of Najah Al-Quraishi, Baghdad, Iraq. The refractive errors were compared by autorefractometry and retinoscopy after cycloplegic dilation. Data analysis was constructed under the use of IBM SPSS V.26 for the determination of emmetropia, myopia, and hyperopia prevalence.\n\nResults The most expected refractive error among the 179 patients was emmetropia, which occurred in 68 patients (38%). Low hyperopia and low myopia were also common, with 64 and 40, respectively. A limited number of patients had moderate/high myopia and moderate/high hyperopia; in detail, the prevalence was following: 5% of patients had moderate myopia, while 0.5% of patients suffered from high myopia; symmetrically, 5% were moderate to high hyperopia. From the data collected, a trend emerged for a low refractive error and symmetric refractive error in both eyes.\n\nConclusion Contrasted to prior conventional wisdom about refractive error in strabismus, it established the greater incidence of emmetropia in patients with intermittent exotropia. The findings call for specific management strategies to be applied in this population.", "keywords": [ "Intermittent exotropia", "emmetropia", "myopia", "refractive errors", "pediatric strabismus." ], "content": "Introduction\n\nIntermittent exotropia is an active type of strabismus that affects quite a large number of children. However, its prevalence might vary from one ethnic and age group to another.1,2 It is insidious since this outward deviation of an eye happens only once in a while, hence disturbing binocular vision. This has immense implications for visual development and the quality of life. The complicated relationship between intermittent exotropia and refractive errors has been known, though not so much understood; according to some studies, different patterns of refractive anomalies could exist in these patients.\n\nStrabismus prevails in approximately 5%-8% of the general population; among these, exotropia ranks as one of the most common varieties, mainly between ages 2-4 years.1,3 Intermittent exotropia is probably considered the most tricky subset of exotropia in terms of coexistent refractive error and management. Whereas esotropia can be associated with plus, the relationship between exotropia and refractive errors is more complex. Of importance, many children with intermittent exotropia become myopic over time; this has been reported as greater than 90% in some cohorts by some series.4\n\nIn order to better understand the therapy and prognosis of intermittent exotropia, this study will look into the distribution of refractive errors in children and young people with the illness. It will also look for common refractive patterns.\n\n\nMethods\n\nThis is a cross-sectional study that took place in the corresponding author’s private ophthalmology clinic. The clinic serves a diverse population, primarily children from various walks of life suffering from ocular conditions including intermittent exotropia. This being an observational study, there is no control, hence all eligible patients diagnosed with intermittent exotropia within the study period ranging from August 2021 to December 2023 were included.\n\nPatients were enrolled consecutively from the routine clinic visits. All subjects had been referred to the study from the clinics for clinical presentations of intermittent exotropia and, therefore, represented a homogenous population by diagnosis. No external advertisements for patient recruitment were utilized, and consecutive eligible patients presenting at the clinic were included, with further selection based only on exclusion criteria regarding previous ocular surgery or other types of strabismus.\n\nRefractive errors were determined by autorefractometry, using a Topcon KR-800®,™ (Topcon Corporation, Tokyo, Japan) following cycloplegia with two drops of 1% cyclopentolate given 10 minutes apart. Testing was done 30 minutes after the final drop to ensure that complete cycloplegia had been achieved. All patients underwent three successive refractometry examinations on each eye to reduce variability. The amount of exotropia was measured by prism cover test. All equipment was calibrated according to the manufacturer’s guidelines before use and checked regularly during the study for accuracy.\n\nThis study followed the STROBE checklist for cross-sectional studies.5 The study design, setting, participants, variables, data sources, measurement techniques, and statistical methods are those in the list that apply and have been appropriately reported to ensure the reproducibility and transparency of the study. Therefore, a completed STROBE checklist is attached and provided to ensure verification of standards.\n\nThis study only included patients whose inclusion criteria consisted of a proven clinical diagnosis of intermittent exotropia, ages from childhood to early adulthood, and those who gave their assent to participate in this research study. The exclusion criteria for this study include those patients with a history of ophthalmic surgery, all constant forms of strabismus, secondary exotropia ensuing from other eye pathology, and systemic conditions that affect visual function.\n\nSquint division into two age groups, namely childhood below six years and school age six years or above, goes in line with the consideration of refractive errors. Both problems are of great educational and psychosocial concern to these age groups, especially since the starting age for school is six years old in Iraq.6,7\n\nThe total number of participants involved in this study was calculated according to the number of patients who presented with intermittent exotropia in the consultation at the outpatient clinic from August 2021 to December 2023. It is an exhaustive sample of all eligible patients who visited with a diagnosis of intermittent exotropia from August 2021 to December 2023. 179 patients were included to be eligible out of 195 and the rest were excluded mainly for non-sufficient data.\n\nRefractive errors were diagnosed by routine ophthalmic tests, including auto-refractometry and retinoscopy after cycloplegia induced by cyclopentolate. The prism cover test found the amount of exotropia. The data collected were then classified and tabulated according to the refractive status of both eyes – emmetropia and myopia are classified into low (less than three diopters), moderate (3 – 6 diopters), and high (more than six diopters). Still, the divisions of hypermetropia are not distinct in the readings accept that follows the similar rating scale depending on the magnitude of refractive error.8 The Ethics Committee (Iraqi Board of Medical Specializations) further stated that the current research study does not require approval because it is a retrospective and has no intervention. Moreover, verbal consent was obtained from all participants in this study instead of written consent. This approach was chosen for several reasons. First, many participants had low literacy levels, making written consent impractical and potentially exclusionary. Second, cultural sensitivities within the community associated signing documents with legal issues or distrust, so verbal agreements were more culturally appropriate and helped build rapport. Lastly, using verbal consent enhanced anonymity and confidentiality by avoiding written records that could compromise participants’ privacy, especially given the sensitive nature of the research topic. The Ethics Committee approved the verbal consent.\n\nData analysis was done using IBM SPSS V.26, where descriptive statistics summarized data and tabulated results in accounts and percentages of each category of refractive error. Another level summary of analysis was undertaken by calculating the mean for both the spherical equivalence and the cylindrical and spherical correction outcome of either eye. These were also tabulated and interpreted to make sense of the refractive trends in this population of patients.\n\nThe data were checked for completeness and accuracy before statistical analysis; partial refraction records and those values that did not logically fit the established norms, after cross-checking with the patients’ records, were screened out. The outliers were defined as those values that lay beyond three standard deviations from the mean of the distribution and were reviewed for possible recording errors. These were included if clinically relevant, as this reflected extreme cases of refractive errors. Sensitivity analyses excluding these outliers were conducted to see the robustness of the results. Overall analysis was conducted using IBM SPSS V.26, with descriptive statistics conducted to summarize patient demographics and refractive error distributions.\n\n\nResults\n\nA total of 179 patients with intermittent exotropia were included in this study. Of these, 96 (47.5%) were male, while 67 (37.4%) were children, and 112 (62.6%) were young students. The demographics of the study participants are summarized in Table 1.\n\nEmmetropia was the most common refractive error, observed in 68 eyes (38%). Low hyperopia and low myopia followed, affecting 64 (35.7%) and 40 (22.3%) patients, respectively. Moderate myopia was present in 9 patients (5%), high myopia in 1 patient (0.5%), and moderate to high hyperopia in 9 patients (5%) (Figure 1).\n\nA cross-tabulation of the refractive errors between the two eyes showed a high degree of symmetry (Figure 2). Most patients had very similar refractive errors in both eyes. Emmetropic and low myopic patients showed a robust bilateral correlation in refractive errors. All moderate to high refractive errors, although less symmetrical, were still recorded in some minority of patients (Table 2).\n\nEmmetropia showed a mean spherical equivalence of 0 for both eyes. Low myopia had a mean spherical equivalence of -2 for both eyes, while moderate myopia showed a mean spherical equivalence of -4 for OD and -4 for OS. High myopia was rare, with only one case, showing a mean spherical equivalence of -6 for OD and not applicable for OS. Low hyperopia had a mean spherical equivalence of 2 for both eyes, and moderate/high hyperopia had a mean spherical equivalence of 5 for OD and 4 for OS.\n\n\nDiscussion\n\nProbably the most striking fact brought out by this study of intermittent exotropia—an eye misalignment characterized by outward deviation—is the remarkably high prevalence of emmetropia, wherein fully 50% of the patient sample showed no refractive error.9 This rate not only forms the bulk of the refractive statuses but contrasts significantly with the expected rates for myopia and hypermetropia traditionally associated with strabismic conditions. This high quantity of emmetropia strongly suggests that intermittent exotropia does not follow the usual patterns of refractive error seen in most other strabismus.\n\nThis study ranges from 20 to 23% in terms of the prevalence of myopia among the cohort, standing in striking contrast to the 55.5% of myopia prevalence in the cohort studied by Robaei et al.10 This discrepancy may reflect demographic, genetic, or environmental differences between the populations or, perhaps more likely, differing criteria for classifying myopia. Our rates are more comparable to those found in a Pakistani study, which also reported an exotropic population with a 50% rate of emmetropia and 29.4 % myopia.11\n\nOur research further elaborates on the relationship between hyperopia and intermittent exotropia. While this association has been reported constantly, our findings give further sophistication to this understanding in pointing out that hyperopia is not as frequent as emmetropia or myopia within this type of strabismic condition. This makes us think about thought whether the refractive error profile for intermittent exotropia might differ from other strabismic disorders in which hyperopia shows up more frequently.12–14\n\nThe paper also discusses the refractive error as dynamic. In particular, it focuses on the trend of an increase in myopia among children who are diagnosed with intermittent exotropia.15 This can indicate that developing myopia is not the cause for exotropia. Still, rather the condition of intermittent exotropia itself could be one of the causes that contribute to the development of myopia. It has been hypothesized that this increased accommodative demand that accompanies the effort at maintaining binocular alignment in intermittent exotropia could very well explain its progression into myopia.16 This is corroborated by studies showing that treatments meant to lessen accommodation, like wearing bifocal glasses, impede the advancement of intermediate myopia.17\n\nGiven our findings, therapeutic strategies for addressing refractive errors in intermittent exotropia patients have to be tailored accordingly. Since the most significant proportion accounted for was emmetropia and myopia and hyperopia relatively less common, treatments must be directed toward maintaining good visual acuity and possible changes? For those who have myopia, particularly progressive forms, interventions such as orthokeratology may be well worth considering. What should always be put forward in any management for intermittent exotropia is the fact that comprehensive care embodies both correction for strabismus and management for refractive error.18\n\nThese findings are in agreement with reports of previous studies that intermittent exotropia may have a refractive error profile different from other forms of strabismus. For example, the relatively high prevalence of emmetropia and low hyperopia in our cohort, compared to the types usually observed among esotropic patients, underlines this distinct distribution of refractive errors and further justifies the need for diagnostic and therapeutic strategies specific to intermittent exotropia.19,20 This differentiation of treatment strategies is essential for the designation of intermittent exotropia as an independent clinical entity, outlining its characteristics of refraction.21\n\nOur study also opens the window for further understanding of how these refractive error patterns may influence long-term visual outcomes. The relative incidence of high myopia and hyperopia may be low, and the prognosis for intermittent exotropia might turn out to be relatively favorable when compared to those with constant esotropia or exotropia if timely and proper measures are implemented.22 This fact heightens concern for regular ophthalmologic follow-up examination and early corrective measures to avoid the progression of refractive errors and achieve optimum visual development.23 Longitudinal studies in the future must be directed toward understanding the progression of refractive error in intermittent exotropia and setting up evidence-based guidelines for its management.24,25\n\nThis current study, hence, has two significant implications: an alternative view of the distribution of refractive error in intermittent exotropia and a causative relationship between strabismus and myopia development. Herein lies critical insight as to which targeted treatment methods may be tailored to treat not only strabismus itself but also consider the potential of growing refractive errors. However, the limitations of our study merit mention. One major limitation in the drawing of solid conclusions regarding the causality of intermittent exotropia and refractive errors is not having a control group of nootropic subjects. Second, there is still missing longitudinal data defining the temporal development of refractive errors relative to the onset and progression of intermittent exotropia. This paper thus sheds light on the refractive landscape of intermittent exotropia. Still, it makes an even stronger case for other and more comprehensive studies to tease out complicated interactions between refractive errors and this common form of strabismus.\n\n\nConclusions\n\nOur study showed that contrary to the traditionally expected predominance of myopia and hyperopia in strabismus, the most frequent refractive errors in intermittent exotropia patients are emmetropia, followed by hypermetropia. Therefore, this might suggest that rather than the reverse, intermittent exotropia is perhaps one of the underlying predisposing factors for myopia development.\n\n\nEthics and consent\n\nThe Ethics Committee (Iraqi Board of Medical Specializations) further stated that the current research study does not require approval because it is a retrospective and has no intervention. Moreover, verbal consent was obtained from all participants in this study instead of written consent. For the minors, the patents and/or legal guardians’ consents were taken. This verbal consent approach was chosen for several reasons. First, many participants had low literacy levels, making written consent impractical and potentially exclusionary. Second, cultural sensitivities within the community associated signing documents with legal issues or distrust, so verbal agreements were more culturally appropriate and helped build rapport. Lastly, using verbal consent enhanced anonymity and confidentiality by avoiding written records that could compromise participants’ privacy, especially given the sensitive nature of the research topic. The Ethics Committee approved the verbal consent.", "appendix": "Data availability statement\n\nThe raw data is available at:\n\nMohammad, N. (2024). Types of refractive errors in a sample of Iraqi children with Intermittent exotropia [Data set]. Zenodo. https://doi.org/10.5281/zenodo.13980859. 26\n\nThe project contains the following data:\n\n• Types_of_refractive_errors_in_a_sample_of_Iraqi_children_with_Intermittent.Updated 1.3.xlsx.\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nNo extended data is available.\n\n\nReferences\n\nBillson F: Fundamentals of clinical ophthalmology. Malaysia: Elsevier; 1st ed.2003; 3.\n\nTang SM, et al.: Refractive Errors and Concomitant Strabismus: A Systematic Review and Meta-analysis. Sci. Rep. 12 Oct. 2016; 6: 35177. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLouis S: Optometric Clinical Practice Guideline: Care of the patient with strabismus: esotropia and exotropia. American Optometric Association; 1997.\n\nZaidi SR, Sadiq MA, Khan AA, et al.: Association Between Refractive Errors and Heterotropia: A Counter Check. Pak. J. Ophthalmol. 2018; 34(3): 107–113.\n\nVon Elm E, Altman DG, Egger M, et al.: The Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) statement: guidelines for reporting observational studies. Lancet. 2007 Oct 20; 370(9596): 1453–1457. Publisher Full Text\n\nMohammed Dhaiban TS, Ummer FP, Khudadad H, et al.: Types and Presentation of Refractive Error among Individuals Aged 0-30 Years: Hospital-Based Cross-Sectional Study, Yemen. Adv. Med. 2021; 2021: 5557761.\n\nSawers N, Jewsbury H, Ali N: Diagnosis and management of childhood squints: investigation and examination with reference to red flags and referral letters. Br. J. Gen. Pract. 2017; 67(654): 42–43. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNémeth J, Tapasztó B, Aclimandos WA, et al.: Update and guidance on management of myopia. European Society of Ophthalmology in cooperation with International Myopia Institute. Eur. J. Ophthalmol. 2021 May; 31(3): 853–883. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKim M, Kim U, Cho MJ, et al.: Hyperopic refractive errors as a prognostic factor in intermittent exotropia surgery. Eye. 2015; 29: 1555–1560. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRobaei D, Kifley A, Mitchell P: Factors associated with a previous diagnosis of strabismus in a population-based sample of 12-year-old Australian children. Am. J. Ophthalmol. 2006; 142: 1085–1087.e1. PubMed Abstract \| Publisher Full Text\n\nCotter SA, et al.: Risk factors associated with childhood strabismus: the multi-ethnic pediatric eye disease and Baltimore pediatric eye disease studies. Ophthalmology. 2011; 118: 2251–2261. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVon Noorden GK, Campos EC: Binocular Vision and Ocular Motility: Theory and Management of Strabismus. 6th ed.St. Louis: Mosby; 2002. Publisher Full Text\n\nXu J, Yu X, Huang Y, et al.: The psychosocial effects of strabismus before and after surgical correction in Chinese adolescents and adults. J. Pediatr. Ophthalmol. Strabismus. 2012; 49(3): 170–175. PubMed Abstract \| Publisher Full Text\n\nColburn JD, Morrison DG, Estes RL, et al.: Longitudinal follow-up of hypermetropic children identified during preschool vision screening. J. AAPOS. 2010; 14(3): 211–215. PubMed Abstract \| Publisher Full Text\n\nWeakley DR: The association between nonstrabismic anisometropia, amblyopia, and subnormal binocularity. Ophthalmology. 2001; 108(1): 163–171. PubMed Abstract \| Publisher Full Text\n\nHuynh SC, Wang XY, Ip J, et al.: Prevalence and associations of anisometropia and aniso-astigmatism in a population based sample of 6 year old children. Br. J. Ophthalmol. 2006; 90(5): 597–601. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nO’Connor AR, Stephenson TJ, Johnson A, et al.: Strabismus in children of birth weight less than 1701 g. Arch. Ophthalmol. 2002; 120(6): 767–773. PubMed Abstract \| Publisher Full Text\n\nLi T, Zuo X, Zhang T, et al.: Patients with Intermittent Exotropia and Exophoria Exhibit Non-aggravated Lens Decentration After Orthokeratology Application: The Nanjing Strabismus Cohort. Ophthalmol. Ther. 2023 Jun; 12(3): 1535–1545. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMohney BG, Lilley CC: The relationship between refractive error and the onset of intermittent exotropia. Ophthalmology. 2017; 124(10): 1634–1640.\n\nCho YA, Kim JH: Analysis of refractive errors in children with intermittent exotropia. Korean J. Ophthalmol. 2018; 32(2): 75–81.\n\nTan H, Yow AP, Hwang SW: Refractive error changes in children with intermittent exotropia: A longitudinal study. Clin. Exp. Optom. 2019; 102(4): 334–340.\n\nChung SA, Mazow ML: Long-term visual outcomes in patients with intermittent exotropia: The impact of early intervention. J. AAPOS. 2016; 20(5): 389–393.\n\nLim SH, O’Hara MA: Efficacy of early corrective measures for intermittent exotropia in preventing refractive error progression. Ophthalmic Epidemiol. 2020; 27(2): 136–144.\n\nLee SY, Lee YJ: Longitudinal analysis of refractive changes in children with intermittent exotropia. Invest. Ophthalmol. Vis. Sci. 2017; 58(5): 2098–2104.\n\nPatel DE, Davey CJ: Developing evidence-based guidelines for managing refractive errors in intermittent exotropia. Br. J. Ophthalmol. 2018; 102(3): 335–340.\n\nMohammad N: Types of refractive errors in a sample of Iraqi children with Intermittent exotropia. [Data set]. Zenodo. 2024. Publisher Full Text" }	[ { "id": "339230", "date": "28 Nov 2024", "name": "Martin Ming-Leung Ma", "expertise": [ "Reviewer Expertise Binocular vision" ], "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\nEnglish editing is needed for this paper. Citations are lacking in some places. And there are some problems in how the authors analyse the data. The lack of written consent is concerning. I also did not understand how and when the verbal consent is obtained if it is a retrospective study.\n\nAbstract: Background: “mostly in children aged 2-4 year”: clarification needed refractive mistakes: should be “refractive error”\nResults: “The most expected refractive error”?\nManuscript: Introduction: “It is insidious since this outward deviation of an eye happens only once in a while, hence disturbing binocular vision “not true for all patients with IXT\n“The complicated relationship between intermittent exotropia and refractive errors has been known, though not so much understood; according to some studies, different patterns of refractive anomalies could exist in these patients.” Citation needed\nMethod: Data collection: what is the dioptric definition of emmetropia? Author should stated it is a retrospective study in the abstract\nI don’t understand why there is a verbal consent if it is a retrospective study. I cannot comprehend what is meant by the “sensitive nature of the research topic”. If obtaining written consent from the participants is not feasible, why didn't the authors obtain it from their legal guardian or parents?\nIs there any tests for normality?\nDiscussion: The main problem here is that the high prevalence of emmetropia in patients with intermittent exotropia should be viewed in context.\n\nAuthor should also collect data regarding the distribution of refractive error types in patients with normal binocular vision. By comparing the distribution of refractive error in patients with and without IXT, we can then establish whether there is a relationship between these two factors. For instance, authors can take a look at this paper for reference: Ma, Martin Ming-Leung, et al. Ref 1.\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": "12901", "date": "16 Dec 2024", "name": "Najah Mohammad", "role": "Author Response", "response": "Thank you for your valuable feedback. We hope that the following changes address your concerns. We have revised the abstract as follows: Clarified the age group to \"predominantly in children aged 2 to 4 years.\" Replaced \"refractive mistakes\" with \"refractive errors.\" Rephrased \"The most expected refractive error\" to \"Emmetropia was the most commonly observed refractive status.\" We have addressed your concerns in the introduction section as follows: Revised the statement “It is insidious since this outward deviation of an eye happens only once in a while, hence disturbing binocular vision” to reflect that this is not true for all patients with intermittent exotropia (IXT). Added appropriate citations for the statement regarding the relationship between intermittent exotropia and refractive errors. We have addressed the concerns regarding the methodology as follows: Added the dioptric definition of emmetropia (±0.50 diopters). Updated the abstract and methods to explicitly state the study's retrospective nature. Clarified the consent process, specifying verbal consent for additional data collection and parental/guardian consent for minors. Included normality testing using the Shapiro-Wilk test in the statistical analysis section. We have made the following adjustments in the discussion section: Discussed the high prevalence of emmetropia in the context of refractive error distribution in normal binocular vision. We acknowledged the retrospective nature of our study, which limits the inclusion of a control group. We have suggested that future prospective studies include such comparisons to further explore the relationship between refractive errors and intermittent exotropia." } ] }, { "id": "343840", "date": "10 Dec 2024", "name": "Yiqiu Lu", "expertise": [ "Reviewer Expertise Amblyopia", "Binocular vision", "Visual plasticity" ], "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 conducted a two-year cross-sectional study in an outpatient clinic of Najah Al-Quraishi and demonstrated that the most common refractive error among the children recruited was emmetropia. This study contains some interesting findings and are valuable for the understanding of refractive status in Iraqi children with intermittent exotropia. Overall, the article still needs some revisions in the description of its content and presentation of data, and I believe that the overall causal logic and clinical significance of the article is debatable.\nThe purpose of the study is not clearly stated in the abstract, and it is recommended to articulate it explicitly in the background section. Incidence and prevalence are distinct statistical concepts, and it is inconsistent to use prevalence in the abstract’s results section and then switch to incidence in the conclusion. This study merely reports the proportion of existing refractive errors among participants, making it inappropriate to claim a high incidence of emmetropia. Prevalence would be the more accurate term. Furthermore, the classification of emmetropia as an illness is questionable. The quality of the references is average and there seems to be no classic literature in the field of strabismus. The authors mention in the introduction that many children with intermittent exotropia become myopic over time, but the literature cited is not a longitudinal study to draw this causal relationship. The study’s purpose, based on its findings, should focus on the distribution of refractive errors in Iraqi pediatric and adolescent patients, rather than the treatment and prognosis of intermittent exotropia. Additionally, the last paragraph of the background does not flow logically. Please clearly state the gaps or limitations of previous research, as I couldn’t identify them from the text. In the first paragraph of “Statistical Analysis,” the phrase “the mean for both the spherical equivalence and the cylindrical and spherical correction” should use the precise term “spherical equivalent refraction (SER)” if that is what it refers to. If so, the refraction should have two decimal places and units. The content under “Ethics and Consent” is duplicated in “Data Collection.” In the Data collection section, orthopia should not be included in the grouping of low-moderate and high myopia, with low myopia being [-0.75D to -3.00), moderate myopia being [-3.00 to -6.00), and high myopia being -6.00 and above The images in Figure 1 are duplicated in Figure 2; there’s no need to include both. Fig1 should be a histogram distribution instead of heatmap, suggest to change it. The use of punctuation in the key takeaway in Figure 2 should be more uniform and standardized. It is suggested to revise Table 2 by adding “Refractive Error OD” and criteria for classifying refractive errors for improved clarity. Please clarify whether the unit for refractive errors is eye or participant, as there is some confusion in the results section of the paper. The results presented in “Symmetry in Refractive Errors” are not entirely convincing. They come across as too vague and generic, and there is a discrepancy between Figure 2 and this section of the results. Could you please do further analysis to conclude that the correlation is high? The results of the detailed refractive error analysis should be presented in tables or plots for improved clarity. The discussion section contains several instances of misplaced or missing citations. For example, the reference in the final sentence of the second paragraph does not correspond to the Pakistani study mentioned in the text. Revision is recommended. In the fourth paragraph of the discussion section, the author states, “The paper focuses on the trend of an increase in myopia among children diagnosed with intermittent exotropia.” However, the evidence provided is neither sufficient nor convincing. Furthermore, the following conclusion that “This can indicate that developing myopia is not the cause of exotropia” cannot be supported based on this statement. It is recommended to adopt a less definitive tone. In the fifth paragraph of the discussion section, the author states, “Given our findings, therapeutic strategies for addressing refractive errors in intermittent exotropia patients have to be tailored accordingly.” While this is an interesting premise, there is insufficient discussion on how or why such tailoring should be implemented. In the final paragraph of the discussion, the conclusion that there is “a causative relationship between strabismus and myopia development” does not seem supported by the study’s results, and this conclusion cannot be considered to have “significant implications.” It is recommended to soften the tone. Additionally, the later mention of “missing longitudinal data defining the temporal development of refractive errors relative to the onset and progression of intermittent exotropia” contradicts this conclusion.\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? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "12968", "date": "16 Dec 2024", "name": "Najah Mohammad", "role": "Author Response", "response": "We appreciate the time and effort you have taken to review our manuscript and for providing insightful comments and suggestions. We have carefully considered each point raised and have revised the manuscript accordingly. Below is a detailed response to your comments: We have revised the abstract and background sections to explicitly state the purpose of the study. The purpose now highlights our aim to investigate the distribution of refractive errors in children with intermittent exotropia and its clinical implications. We have replaced \"incidence\" with \"prevalence\" throughout the manuscript, as this study reports proportions rather than the emergence of new cases. Additionally, we clarified that emmetropia is not classified as an illness. We carefully selected the cited literature to ensure it aligns with the focus and scope of the manuscript. While I aimed to include relevant and high-quality studies, I acknowledge that perspectives on what constitutes classic literature may vary. I did my best to include the most pertinent and available sources within the context of this work, and I hope the selection adequately supports the study's objectives. The introduction and discussion sections have been revised to adopt a less definitive tone. We now emphasize the need for longitudinal studies to establish causal relationships. We have restructured the background to clarify the study's primary focus on refractive error distribution and removed extraneous emphasis on treatment and prognosis. We have expanded the introduction and discussion sections to explicitly outline the gaps in prior studies, such as the lack of comprehensive data on refractive error profiles specific to intermittent exotropia and the absence of longitudinal analyses. We have replaced \"spherical equivalence\" with \"spherical equivalent refraction (SER)\" where appropriate and included refractive measurements to two decimal places with units. The overlapping content has been streamlined to remove duplication and improve clarity. Orthopia has been excluded from the myopia classification, and we clarified the myopia ranges in the data collection section. We have removed redundancy between Figures 1 and 2. The data is categorical in nature, histogram is not suitable for such issues Figure 2 has been removed Table 2 has been revised to include a column for \"Refractive Error OD\" and a footnote detailing the refractive error classification criteria. We have clarified in the methods and results sections that refractive errors are reported per patient, not per eye, to reflect the nature of the data. Figure 1 has been replaced by a confusion matrix to better visualize the data We have added a table to present the detailed refractive error analysis for better clarity. We have corrected all misplaced citations and ensured they align with the referenced studies. We have revised the statement to reflect the observational nature of the findings and removed unsupported causal implications. We have expanded the discussion to include specific examples of tailored therapeutic strategies based on the study's findings. The final paragraph has been revised to remove unsupported causal claims and emphasize the need for longitudinal studies. We believe these revisions have strengthened the manuscript and addressed your concerns thoroughly. Thank you again for your valuable feedback. Sincerely," } ] } ]	1	https://f1000research.com/articles/13-1318
https://f1000research.com/articles/12-441/v2	08 May 24	{ "type": "Research Article", "title": "The interaction between brain-based learning strategies and patterns of infographics and its influence on the development of information concepts among Saudi undergraduate cybersecurity students", "authors": [ "Yasser Shabaan Abdelaziz Mohammed", "Ayman Sabry Daif-Allah", "Ayman Sabry Daif-Allah" ], "abstract": "The study reported in this paper investigates the impact of the interaction between brain-based learning strategies (spaced repetition and distributed practice - mental models) and patterns of infographics (static/animated) in e-learning environments and its impact on the development of informational concepts among cybersecurity students. The quantitative data were collected by means of Information Concept Test to determine the levels of information concepts according to the Frayer model (1969). The research sample consists of 80 male and female undergraduate students in Qassim University, who were classified into four quasi-experimental groups according to their preferred patterns in the infographic. Quantitative analysis of findings from this study shows an effect for the interaction between the variables of brain-based learning strategies (spaced repetition and distributed practice - mental models) and patterns of infographics (static/animated) in e-learning environments in the test scores of information concepts in the post-administration, where the significance level (.011) is less than the significance level (0.05). Findings also revealed that the repetition of information at increasing intervals of time using the infographics is effective to learning concepts in an organized manner and to improving short memory. Furthermore, the mental model’s strategy also helped to retain information. The study recommends encouraging the use of brain-based learning strategies and patterns of infographics (static/animated) in e-learning environments to explore the relationships between different scientific concepts, to expand students’ knowledge, and to discover inter-relationships between different subject elements. The study provides a platform for further research to employ different brain-based learning strategies within e-learning environments to acquire higher-order thinking skills and improve students’ memory and mental capacity with the use of modern technological techniques that achieve learning quality.", "keywords": [ "Learning", "E-learning", "E-learning environments", "Infographics", "Brain-Based Learning Strategies", "Mental Models Strategy", "Spaced Repetition and Distributed Practice Strategy", "Information Concepts", "Saudi Undergraduate Students", "Cybersecurity" ], "content": "Editorial note\n\nEditorial Note (26th July 2023): The F1000 Editorial Team is currently substantiating that this research was presented at an IIARP conference and should be included in the IIARP Gateway. The Editorial Team is currently requesting further information from the authors regarding this. This Editorial Note will be updated as new information is provided.\n\n\nIntroduction\n\nIn light of the information revolution and technological development, it has become necessary to use pedagogical, mental and cognitive strategies and modern technologies in adaptive e-learning environments. In addition, there is a need to capitalize on research on how the brain works to introduce strategies for learning, education and training. In this regard, the needs, motives, interests and tendencies of students must be taken into account to help them achieve learning outcomes in a better way and achieve the success of the educational process. Teaching strategies based on brain-based learning are new strategies used by teachers and trainers to enhance and improve learning and memory in the classroom and in e-learning environments for learning quality and memory improvement (Lubis, 2017; Yustitia et al., 2019). Brain-based learning has become a necessity for learner education, as it provides multiple patterns and forms of thinking, and allows for the existence of many diverse, supportive and effective training strategies and applications in the field of thinking and methods of improving memory. Brain-based learning theory is a theory that links brain science, neuroscience, behavioral science, and cognitive science. Brain-based learning strategies help enrich teaching and learning processes and provide an adaptive learning environment by focusing on attention, memory, motivation, the way information is presented, and ways of acquiring conceptual knowledge. Instructional environments and strategies based on brain-based learning are also characterized by the diversity of digital learning resources, sharing of information, obtaining immediate feedback that learners need, as well as developing thinking and creativity skills (Sesmiarni, 2015). Brain-based learning strategies allow the learner to reduce memory loss as well as improve basic learning ability by directly enhancing basic learning processes in the brain. The aspects of learning that rely on the brain are varied (Caine & Caine, 1994; Crow, 2011; Jensen & McConchie, 2020). Similarly, Amit-Danhi (2020) discovered that students recalled more of what they read when it was accompanied by visuals, such as pictures or graphs (Amit-Danhi, 2020). Therefore, infographics would increase knowledge retention can and help improve learning outcomes.\n\n\nContext of the problem\n\nThe previous introduction shows brain-based learning strategies that may help improve students’ memory through the operation of the left and right hemispheres and the optimum using activities, e-learning resources such as infographics (static/animated). Research studies that dealt with brain-based learning have proven the effectiveness of brain-based learning on developing e-learning environments and improving memory recall and retention of facts and details (Jensen & Mcconchie, 2020). In addition, the integration of brain-based learning strategies and content presentation methods using graphs and models is significant for the improvement in the performance of the e-learning system based on learning environments (Erol & Karaduman, 2018). The Twelve Steps of Brain-Based Learning allow instructors to reach a more diverse group of learners, confirming the idea that not all students learn the same way and allowing instructors to teach in many ways. In addition, results of a pilot study on a sample of cybersecurity diploma students show positive perception toward brain-based learning strategies due to performing a number of active tasks that help them interact and share digital resources. It also allows students to work together to solve problems, both by synchronous and asynchronous communication.\n\n\nProblem of the study\n\nThere are many previous related studies whose results and recommendations dealt with brain-based learning, its role, importance, and effectiveness in achieving educational and training objectives (Craig, 2003; Green, 2017; Elyusra, 2017; Jensen & Mcconchie, 2020). However, no research has been carried out to investigate the interaction between brain-based learning strategies and infographics for Saudi cybersecurity undergraduate students to improve their low achievement scores in the scientific terminology course, due to the difficulty in understanding the concepts and information of the course and the students’ exposure to memory loss and forgetfulness.\n\n\nResearch questions\n\n\n\n1. What are the informational concepts that Saudi undergraduate cybersecurity students should have?\n\n2. What are the criteria of designing and producing infographics (static/animated) to develop informational concepts for Saudi undergraduate cybersecurity students?\n\n3. What is the educational design of an adaptive e-learning environment-based brain-based learning strategies (spaced repetition and distributed practice - mental models) for developing information concepts among Saudi undergraduate cybersecurity students?\n\n4. What is the effect of the interaction between brain-based learning strategies (spaced repetition and distributed practice-mental models) and infographics (static/animated) and its impact on the development of information concepts for Saudi undergraduate cybersecurity students?\n\n\nHypotheses of the study\n\nThere is a significant interaction between brain-based learning strategies (spaced repetition and distributed practice - mental models) and patterns of infographics (static/animated) in e-learning setting in the post-measurement of Information Concepts Test after adjusting the pre-measurement effect.\n\n\nPurpose of the study\n\n\n\n1. Determine the informational concepts that cybersecurity students should have.\n\n2. Develop acquisition of informational concepts for cybersecurity students.\n\n3. Determine the criteria of designing the adaptive electronic environment based on learning strategies.\n\n4. Introduce a proposed conceptualisation of the e-learning environment based on brain-based learning strategies to develop informational concepts among cybersecurity students.\n\n5. Identify the interaction between both brain-based learning strategies and infographics, and its impact on the development of information concepts among cybersecurity students.\n\n\nSignificance of the study\n\nThree reasons make this research noteworthy. First, it provides insight into a topic that has received a lot of attention from scholars all around the globe. Second, it helps fill a gap in Arab scholarship on the topic of infographics (both static and dynamic) and the role they play in the evolution of conceptual understanding. Third, it keeps pace with global modern trends to develop education and increase the effectiveness of the educational process by making the learner the focus of the educational process. The study would also provide guidelines for presenting a model for designing the adaptive electronic environment based on providing educational situations that remove turmoil, confusion and fear of failure through cooperation with colleagues and participation to take responsibility that can be emulated in the field of e-learning technology applications. Furthermore, the current research may help in providing a framework for developing an educational situation that helps learners enhance their memory and retention abilities.\n\n\nLiterature review and related studies\n\nBrain-based learning is a paradigm of learning which addresses student learning and learning outcomes from the point of view of the human brain. Jensen and Mcconchie (2020) state that brain-based learning refers to teaching methods, lesson designs, and school programs that are based on the latest scientific research about how the brain works, including factors such as cognitive development—how students learn differently as they age, grow, and mature socially, emotionally, and cognitively. The researchers define brain-based learning procedurally as learning compatible or harmonious with the brain according to the way the brain works in order to achieve the best possible excitation of the brain to learn in the best possible way and achieve the best results.\n\nBrain-based learning strategies can be defined procedurally as brain-based learning strategies that include a set of steps, procedures and practices based on the theory of brain-based learning that is compatible with its functions that the instructor uses to improve the concepts of cybersecurity, and it consists of the following five stages: readiness for learning, organized integration, quiet alertness, active processing, increase brain capacity to achieve the best results.\n\nBrain-based learning uses several brain-based strategies, including:\n\n1. With spaced repetition, the time intervals between learning attempts can be increased, and each learning attempt strengthens neural connections, supporting the findings of distributed practice and spaced repetition that distributing learning over time and repeating information over increasing periods strengthens memories and shields against forgetting. As shown in Figure 1, where the student studies a related set of concepts (A), then a set of concepts (B), and so on for a set of concepts (C) and (D), and this spaced repetition is distributed in the week, while the study of concepts is prohibited with these Method (AAAA), (BBBB), (CCCC), (DDDD).\n\nThe current research focuses on spaced repetition strategies and distributed practice. The Ebbinghaus Forgetting Curve may change Figure 2:\n\n2. Activities such as developing art, instructing others, applying knowledge in real-world situations, writing blog entries, participating in online discussion forums, etc. all fall under the umbrella of “generative learning.” In order to rethink and restructure information, it is necessary to combine new knowledge with previously acquired information. This helps to reinforce what was learned and reveals any gaps in understanding. Recall, comprehension, and the ability to acquire new information in-depth and broadly are all enhanced as a result.\n\n3. The production effect, in which knowledge is replicated vocally or graphically, is a frequent strategy for improving retention of that information. In order to quickly override the brain’s natural inclination to consolidate new knowledge, it is helpful to speak, listen to, or teach the material aloud. One’s ability to remember and memorise information is enhanced.\n\n4. Memorising knowledge, practicing, and the testing impact all go under “retrieval practice.” Because forgetting and remembering are not opposites, this may not prevent people from remembering. Retrieval training and testing are effective learning strategies because they strengthen prior knowledge while also providing an opening for new information to be acquired through the investigation of misconceptions and the accumulation of little successes. Memory may be strengthened in the context of remembering by retrieval exercise. The brain’s optimal state for learning has a distinct profile from that for recall and memory. The capacity to recollect information is crucial for making use of new knowledge. As a result, one’s ability to remember material, feel secure in oneself, and do well under testing conditions.\n\n5. Mental models are depictions of concepts that serve as templates for thinking and provide feedback loops; they are schemas of real-world processes. Stories and comparisons are powerful learning tools because people naturally gravitate toward them. When we utilise a metaphor, we “abstract” and grasp the core of the unknown process because of the contrast it establishes between the identified and unidentified processes.\n\n6. What we mean when we talk about “chunking” is breaking down larger chunks of text into smaller, more manageable chunks that all relate to the same topic. Information can be chunked based on superficial attributes such as colour, similarity, category, relationships, etc. The brain uses networks of information, thus it is simpler to recall several chunks than it is to remember several separate parts. As opposed to relying only on rote memorization, cognitive processes may help you retain complex information, make sense of complex concepts, and make connections between ideas. Figure 3 demonstrates the effectiveness of infographics in elucidating ideas by drawing parallels to the mental models approach.\n\nBased on a thorough survey of related literature (Blakemore & Frith, 2016; Medina, 2018; Jensen & Mcconchie, 2020; O’mahony, 2021), the basic characteristics of brain-based learning can be illustrated as follows:\n\n1. The theory of brain-based learning is based on the fact that the human being is able to learn and acquire, if the appropriate conditions are provided for him, and his ability to learn increases by stimulating and activating neurons to form the largest possible number of neural connections with other neurons.\n\n2. The human brain can learn best when the left and right hemispheres are involved in information processing, storage and retrieval and teaching is in accordance with the principles and postulates of brain-based learning.\n\n3. Learning according to traditional curricula based on memorization is no longer valid for teaching now. Therefore, teaching and learning methods and strategies should evolve to suit the new challenges, especially the different thinking skills.\n\n4. Principles underlying Brain-Based Learning Theory involve body physiology since the search for meaning is innate and learning occurs through simulation of emotions and the mind processes parts and words simultaneously.\n\nCohen et al. (2003), Willis (2008), Blakemore and Frith (2016), and Masurkar (2021) illustrate many uses for brain-based learning, including:\n\n1. Brain-based learning enhances students’ academic progress and fosters a positive learning environment.\n\n2. Instructors who apply brain-based learning often observe increases in knowledge retention and academic performance.\n\n3. When students develop an intrinsic love for learning and approach the classroom with the right mindset, the classroom will be motivating and exciting.\n\n4. Brain-based learning creates a learning environment and classroom strategy where all students can thrive.\n\nInfographics can be defined as a representation of information in a graphical format designed to make data easy to understand. Teachers use infographics to communicate a message to students simplify. Infographics include line charts, graphs, charts, and mind maps.\n\nInfographics are of great importance in the teaching and learning processes, and there are many studies that have confirmed the effectiveness of the infographic in the teaching and learning process. On top of these, Rendgen et al. (2020) who emphasized the importance of employing infographics in e-learning environments, and its effective role in supporting educational content and simplifying complex and large information. In addition, infographics are attractive and exciting in presenting information, improving understanding of information, ideas and concepts that improve information retention and retrieve, enhancing the ability to think critically, attracting attention, making images easy and quick to read, and helping generate large amounts of information (Smiciklas & Wiegand, 2012; Krum, 2014).\n\nCriteria for a successful infographic design are illustrated by (Beegel, 2014; Julien et al., 2020) as organizing the content and categorizing it according to the level of importance. Correct and effective visual coding is a guide for students to focus and follow the flow of information; Determine the audience for which the infographic will be presented, the infographic must be educationally sound; It helps learners to remember information easily, divide complex processes into simpler components and represent them visually and sequentially for ease of understanding, and the infographic should be compatible with the content, reduce text and use visual elements, focus on the most important pieces of information so that they are prominent, organize data and know the most important parts. The information to be included in the infographic, and the infographic to be effective and attractive.\n\nThe current study is based on the Constructivist Theory in which students learn better when they participate in learning experiences. Learning is by nature a social process because as it is embedded in a social context where students and teachers work together to build knowledge. The experience can make it easier for students to build their own knowledge. The study is also based on the Information Processing Theory that describes how sensory inputs are received, transformed, stored, retrieved, and used. This theory is based on allowing enough time for the classroom to process information, pay attention to tests, persevere and reduce information (Slate & Charlesworth, 1988; Clark & Paivio, 1991; Steffe & Gale, 1995; Moser & Chen, 2012; Szymanski, 2021).\n\n\nMethods\n\nThe research design used is quasi-experimental. This design gives detailed information about the data gathered.\n\nThe subjects of this study are Saudi Cybersecurity Diploma Students at Qassim University. They graduated from the high school and have joined the Diploma in Cybersecurity for two years to practice the profession of protecting electronic devices and networks from hacking. The research groups consist of 80 cybersecurity students at Qassim University (40 male and 40 female students). These subjects were randomly chosen from 198 level one students enrolled in the cybersecurity program in Qassim University. They were divided into four groups: experimental (G1, G2, G3, G4), and the number of each group was 20 students studying through the Blackboard system (see Figure 4).\n\nTo achieve the objectives of the research and verify its hypothesis, the researchers used an Information Concept Test according to the Frayer model (Frayer et al., 1969) “In the Frayer model, the student begins by writing a term in the centre, and in the four boxes that surround the term write the properties of this term, example Figure 5 and identify antonyms or synonyms”. This test was developed and adapted from different sources (Brooks & Al, 2017; Lowe, 2018; Panek, 2020; Steinberg, 2020). The purpose of the test is to measure the knowledge and concepts acquired by cybersecurity students in the content of scientific terms after exposure to brain-based learning strategies and the use of infographic presentation methods. Many test items are largely derived from the scientific terminology and an introduction to cybersecurity courses, which are presented to Saudi first-year undergraduate cybersecurity students. These courses were selected because of their focus on modelling learning concepts using static and infographics graphs to explain some of the basic aspects related to cybersecurity, and the concepts can be modelled by exploiting similar relationships for concept representations. It consists of four topics: Information security, Network security, Operational security, and Application security.\n\nTo answer the research questions, the following procedures were followed:\n\n1. Preparing the theoretical framework by reviewing the pertinent literature and the related studies aligned with the research variables.\n\n2. Developing the Information Concept Test.\n\n3. Identifying a list of criteria for designing electronic content based on brain-based learning strategies.\n\n4. Choosing a model for designing the e-learning situation compatible with brain-based learning strategies as described in Figure 6.\n\n5. Designing an e-learning situation based on brain-based learning strategies to develop information concepts among cybersecurity students.\n\n6. Linking the e-learning setting that was designed to the Blackboard Learning Management System.\n\n7. Conducting the exploratory experiment to adjust the research instrument.\n\n8. Selecting research groups of cybersecurity students at Qassim University. Eighty students were divided into four groups: experimental (G1, G2, G3, G4), and the number of each group was 20 students, and they were taught through the Blackboard system. These students were divided according to the patterns of infographic to develop the information concepts of the cybersecurity students.\n\n9. Piloting of the Information Concept Test:\n\n10. Administration of the Information Concept Test. Data were obtained by distributing the test to 80 male/female undergraduate cybersecurity students studying at Qassim University during the academic year 2021/2022.\n\n11. Students’ answers were scored; the researchers give a score of 1 for each correct answer of the 60 questions, and 0 for every wrong answer. This can be seen in the raw data of the pre and post-tests.\n\n12. Monitoring, analyzing, statistically processing and interpreting the results.\n\n13. Presenting recommendations and suggestions in light of the research results.\n\n\nDelimitations\n\nResearch is delimited to these strategies: Mental Models, Spaced repetition, Distributed practice, and Spacing Effects. It is also delimited to Saudi undergraduate cybersecurity students at Qassim University.\n\n\nEthics and consent\n\nThe researchers obtained the approval of both the Committee of the Department of Applied Natural Sciences in the Applied College and the Research Ethics Committee of Qassim University (REC) on April 16, 2021 based on the written consent of all participants in the research, and the participants were notified by the researchers that the data will be published while preserving data privacy and they had no objection.\n\n\nResults and discussion\n\nThe study hypothesizes that “there is a significant interaction between brain-based learning strategies (spaced repetition and distributed practice - mental models) and patterns of infographics (static/animated) in e-learning environments in the post-measurement of the Information Concept Test after adjusting the pre-measurement effect”. To verify the validity of this hypothesis, the ANOVA (analysis of covariance) test was used to find out if there is an effect of the interaction between brain-based learning strategies and patterns of infographics in e-learning environments in the post-measurement of Information Concept Test as shown in Tables 1 and 2 below:\n\nThe results in Tables 1 and 2 show that:\n\n1. There are no significant differences between the average scores of students according to the methods of presenting patterns of infographics (static/animated) in e-learning environments, where the significance level (0.740) is greater than the level of significance (0.05), and therefore, it is not a statistically significant value.\n\n2. There are significant differences between the mean scores of students according to brain-based learning strategies in e-learning environments in the post-administration of Information Concept Test since the significance level (0.034) is less than the level of significance (0.05). Therefore, it is a statistically significant value. The differences in the means were in favor of the students who learned the strategy (spaced repetition and distributed practice) (highest mean = 59.10), and the average scores of the students who learned the strategy (mental models) were (58.40).\n\n3. There is an effect for the interaction between the variables of brain-based learning strategies and patterns of infographics in e-learning environments in the test scores of information concepts in the post-administration, since the value of (P = 6.873), where the significance level (0.011) is less than the significance level (0.05), and therefore it is a statistically significant value.\n\nThe results presented in Figures 7 and 8 are consistent with Erol and Karaduman (2018) who found that brain-based learning raised the efficiency of students with difficulties in numerical cognition and that Graphic design has a positive effect on the cognitive achievement and skill performance of secondary school students. Also, Sesmiarni (2015) and Solomon (2020) concluded that brain-based learning helped in acquiring students’ higher-order thinking skills, writing skills, and develop students’ creativity, due to the optimal use of diverse learning materials compatible with the brain, and significantly improved students’ academic participation, and enhanced their ability to critical thinking. The results are in line with Johnson and Mayer (2009) who found that infographics increased academic achievement and helped acquire visual thinking skills and increased willingness to learn on university students, distinguish and perceive visual objects, improve writing, comprehension and retention, and enhance information comprehension speed. In addition, Smiciklas and Wiegand (2012) confirms the research results by concluding that brain-based learning helped increase cooperation, participation and understanding among students, and the results also showed the effect of using infographics on developing creative thinking skills for educational technology students, which helped in enhancing children’s visual-motor memory, and the use of drawings.\n\nThe results of the current study can be attributed and interpreted in light of the following considerations:\n\n• Good design of static and animated infographics, adaptive e-learning environment in light of design and production standards, choosing a model for designing e-learning environment compatible with brain-based learning strategies,\n\n• Effective design of educational tasks and activities, related to the objectives and educational content that help the student to remember,\n\n• Good design of media and electronic resources and employing them within the learning environment in proportion to the learning objectives, and consistent with the learning environment,\n\n• Implementing a strategy of spaced repetition and practice distributed in four steps based on regular planning of study sessions that helped students remember previous information, study and review information constantly,\n\n• Presenting content in infographic patterns (static/animated), helped students to retain information for a longer period.\n\n• Using an Information Concept Test according to the Frayer model.\n\n• On-going assessment of the students through discussions, questions and participation in interactive activities available via the blackboard system,\n\n• The student’s recall of information at spaced intervals according to the schedule based on spaced repetition and distributed practice in retrieval,\n\n• The use of a brain-based learning strategy (mental models), which encouraged the student, through discussion and group work across forums, to think and mentally visualize informational concepts related to the field of cybersecurity.\n\n• the use of the infographic pattern (static/animated) in the formation of mental images about similarities and comparisons between known and unknown concepts to link previous information and new information, which significantly helped the students to retain the information.\n\n\nConclusion and implications\n\nFindings reveal that the use of spaced repetition and distributed practice strategy with static and infographics infographic content presentation style had more impact on learning and information retention in the field of cybersecurity, than mental models strategy with the use of static and infographics infographic content presentation style, Where the strategy of spaced repetition and distributed practice helped students to train the mind to remember information and concepts through the learning schedule and multiple learning sessions extended for three months, the student’s permanent review of information at regular intervals, and the use of training tests a better way to learn by focusing on the salient points; Which helped to remember a large percentage that may reach 100%, the mental models strategy with the use of static and infographics infographic content presentation style had the least impact on learning and retaining informational concepts in the field of cybersecurity in an attempt to link old information with modern information.\n\n\nRecommendations and suggestions\n\n\n\n1. Employing brain-based learning strategies in different teaching situations to address individual differences among students.\n\n2. Employing static or animated infographics in electronic learning environments to develop informational concepts and activate memory for students with poor memory.\n\n4. Holding courses for teachers to train the skills of applying brain-based and infographic learning strategies in the classroom and within e-learning environments.\n\n5. Conducting further research on the effect of brain-based learning strategies on developing types of thinking.\n\n6. Conducting further research on the effect of brain-based learning strategies in e-learning environments to increase students’ mental capacity.", "appendix": "Data availability\n\nFigshare: https://doi.org/10.6084/m9.figshare.20455503.v4 (Yasser Mohammed et al., 2022)\n\nThis project contains the following underlying data:\n\n- Raw data.xlsx\n\n- Pre-Test\n\n- Post-Test\n\n- Strategies Data\n\nThis project contains the following extended data:\n\n- Information Concept Test - (full test items).\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgement\n\nResearchers would like to thank the Deanship of Scientific Research, Qassim University for funding publication of this project.\n\n\nReferences\n\nAmit-Danhi E: TMI: Information Rhetoric Types in Digital Political Infographics. AoIR Selected Papers of Internet Research. 2020. Publisher Full Text\n\nBeegel J: Infographics for dummies. John Wiley & Sons, Inc; 2014.\n\nBrooks CJ, Al E: Cybersecurity Essentials: Website Associated with Book. John Wiley & Sons Inc.; 2017.\n\nCaine R, Caine G: Making connections: Teaching and the human brain. Menlo Park, CA: Addison-Wesley; 1994.\n\nClark JM, Paivio A: Dual coding theory and education. Educ. Psychol. Rev. 1991; 3(3): 149–210. Publisher Full Text\n\nCohen I, Dennison PE, Dennison G, et al.: Hands on: how to use Brain Gym in the classroom. Edu-Kinesthetics; 2003.\n\nCraig DI: Brain-compatible learning: principles and applications in athletic training. J. Athl. Train. 2003; 38(4): 342–349. PubMed Abstract\n\nCrow JT: Learning to write for readers using brain-based strategies. National Council of Teachers of English; 2011.\n\nEagleman D, Downar J: Brain and behavior: a cognitive neuroscience perspective. Oxford University Press; 2016.\n\nElyusra.: A Meaningful Brain-Based Literature Learning Model. JISAE: Journal of Indonesian Student Assessment and Evaluation. 2017; 3(1): 10–23. Publisher Full Text\n\nErol M, Karaduman GB: The Effect of Activities Congruent with Brain Based Learning Model on Students’ Mathematical Achievement. NeuroQuantology. 2018; 16(5). Publisher Full Text\n\nFrayer D, Frederick WC, Klausmeier HJ: A Schema for Testing the Level of Cognitive Mastery. Madison, WI: Wisconsin Center for Education Research; 1969.\n\nGreen O: Top 10 Brain-Based Teaching Strategies.2017. Reference SourceReference Source\n\nJensen E, Mcconchie L: Brain-based learning: the new paradigm of teaching. Corwin Press; 2020.\n\nJohnson CI, Mayer RE: A Testing Effect with Multimedia Learning. J. Educ. Psychol. 2009; 101: 621–629. Publisher Full Text\n\nJulien HE, Gross M, Latham D, et al.: The information literacy framework: case studies of successful implementation. Rowman & Littlefield; 2020.\n\nKrum R: Cool infographics: effective communication with data visualization and design. John Wiley & Sons, Inc; 2014.\n\nLowe D: Networking all-in-one for dummies. John Wiley & Sons, Inc; 2018.\n\nLubis RH: The Effect of Cooperative Learning Model Group Investigation Against Student Learning Outcomes Physics Viewed from The Adversity Quotient. Jurnal Pendidikan Fisika. 2017; 6; 44(1). Publisher Full Text\n\nMasurkar RK: Brain Based Learning. Educational Resurgence Journal. 2021; 145. Reference Source\n\nMedina J: Brain rules for ageing well: 10 principles for staying vital, happy, and sharp. Scribe; 2018.\n\nMoser SM, Chen P-N: A student’s guide to coding and information theory. Cambridge University Press; 2012.\n\nO’mahony K: The brain-based classroom: accessing every child’s potential through educational neuroscience. Routledge, Taylor & Francis Group; 2021.\n\nPanek C: Security fundamentals. John Wiley & Sons, Inc; 2020.\n\nRendgen S, Wiedemann J, Gmbh T: Information Graphics. Taschen; 2020.\n\nBlakemore S-J, Frith U: The learning brain: lessons for education. Blackwell; 2016.\n\nSesmiarni Z: Brain Based Teaching Model as Transformation of Learning Paradigm in Higher Education. Al-Ta Lim. 2015; 22(3): 266–275. Publisher Full Text\n\nSlate JR, Charlesworth JR Jr: Information Processing Theory: Classroom Applications.1988.\n\nSmiciklas M, Wiegand G: The power of infographics: using pictures to communicate and connect with your audience. Que; 2012.\n\nSolomon J: On the Use of Brain-Based Learning Protocols in Fluid Mechanics Instruction. Journal of STEM Education: Innovations and Research. 2020; 21(3). Reference Source\n\nSteffe LP, Gale JE: Constructivism in education. Lawrence Erlbaum; 1995.\n\nSteinberg J: Cybersecurity. John Wiley & Sons, Inc; 2020.\n\nSzymanski J: Coding. Children’s Press, An Imprint of Scholastic Inc; 2021.\n\nWillis J: Teaching the brain to read: strategies for improving fluency, vocabulary, and comprehension. Association for Supervision and Curriculum Development; 2008.\n\nYasser M: Interaction between Brain-based Learning Strategies and Patterns of Infographics and its Impact on the Development of Information Concepts among Saudi Undergraduate Cybersecurity Students. [Dataset]. 2022. Publisher Full Text\n\nYustitia V, Wardani IS, Juniarso T: The Effect of Brain Based Learning Model On Student’s High Order Thinking Skills. EduHumaniora\|Jurnal Pendidikan Dasar Kampus Cibiru. 2019; 11(1): 71. Publisher Full Text" }	[ { "id": "281641", "date": "04 Jun 2024", "name": "Fozia Fatima", "expertise": [ "Reviewer Expertise Educational Psychology", "Educational Planning and Management", "Research Methods", "Health Professions Education", "Social Sciences" ], "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 report was originally published on 4 June 2024, but an edit was made to the first sentence of the report by our Editorial Team to ensure the sentence is in-line with our peer-review guidelines\nThe idea of investigation is good but ... Here are some shortcomings, that have serious revision; 1- The rationale behind the topic is not strong. Could you try to write the reasons behind selecting this topic, especially in the context of your study? Give brief literature and identify present and past gaps in this area of research. 2- Why research questions and hypothesis both are in this study? 3- The theoretical framework is not clear in the context of this study. 4- Research Methodology needs serious revision, especially in design selection, sample, and sample size calculation. Justify your selection. 5- Align objectives, questions, hypotheses, statistical tests. Describe strong evidences that supports to your 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? 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? Partly\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [ { "c_id": "13013", "date": "06 Jan 2025", "name": "Yasser Mohammed", "role": "Author Response", "response": "The authors would like to thank the reviewer for the valuable comments and a number of modifications and clarifications have been made in the revised manuscript as follows: 1- The rational section has been modified to reflect the theoretical and practical significance of the study. The literature review section clearly identifies present and past gaps in literature that raises the need for conducting the research and this is clearly written in the problem of the study part (There are many previous related studies whose results and recommendations dealt with brain-based learning, its role, importance, and effectiveness in achieving educational and training objectives (Craig, 2003; Green, 2017; Elyusra, 2017; Jensen & Mcconchie, 2020). However, no research has been carried out to investigate the interaction between brain-based learning strategies and infographics for Saudi cybersecurity undergraduate students to improve their low achievement scores in the scientific terminology course, due to the difficulty in understanding the concepts and information of the course and the students’ exposure to memory loss and forgetfulness.) 2- Following rules of scientific research, research questions and hypothesis are both in this study. 3- The theoretical framework covers the important points related to the topic as well as the related studies. 4- The research methodology is professionally presented in light of academic research rules. It provides detailed explanation of the research design, participants and selection procedures, instruments and treatment as follows: Research design The research design used is quasi-experimental. This design gives detailed information about the data gathered. Participants The subjects of this study are Saudi Cybersecurity Diploma Students at Qassim University. They graduated from the high school and have joined the Diploma in Cybersecurity for two years to practice the profession of protecting electronic devices and networks from hacking. The research groups consist of 80 cybersecurity students at Qassim University (40 male and 40 female students). These subjects were randomly chosen from 198 level one students enrolled in the cybersecurity program in Qassim University. They were divided into four groups: experimental (G1, G2, G3, G4), and the number of each group was 20 students studying through the Blackboard system (see Figure 4). Figure 4. Experimental research design. Instruments To achieve the objectives of the research and verify its hypothesis, the researchers used an Information Concept Test. According to the Frayer model (Frayer et al., 1969) “In the Frayer model, the student begins by writing a term in the centre, and in the four boxes that surround the term write the properties of this term, Example Figure 5 and identify antonyms or synonyms”. This test was developed and adapted from different sources (Brooks & Al, 2017; Lowe, 2018; Panek, 2020; Steinberg, 2020). The purpose of the test is to measure the knowledge and concepts acquired by cybersecurity students in the content of scientific terms after exposure to brain-based learning strategies and the use of infographic presentation methods. Many test items are largely derived from the scientific terminology and an introduction to cybersecurity courses, which are presented to Saudi first-year undergraduate cybersecurity students. These courses were selected because of their focus on modelling learning concepts using static and infographics graphs to explain some of the basic aspects related to cybersecurity, and the concepts can be modelled by exploiting similar relationships for concept representations. It consists of four topics: Information security, Network security, Operational security, and Application security." } ] } ]	2	https://f1000research.com/articles/12-441
https://f1000research.com/articles/13-1447/v1	29 Nov 24	{ "type": "Research Article", "title": "Strategic priorities and challenges in research software funding: Results from an international survey", "authors": [ "Eric A. Jensen", "Daniel S. Katz", "Daniel S. Katz" ], "abstract": "Background Research software is increasingly recognized as critical infrastructure in contemporary science. It spans a broad spectrum, including source code files, algorithms, scripts, computational workflows, and executables, all created for or during research. While research funders have developed programs, initiatives, and policies to bolster research software’s role, there has been no empirical study of how these funders prioritize support for research software. Understanding their priorities is essential to clarify where current support is concentrated and to identify strategic gaps.\n\nMethods We conducted an online mixed methods survey of international research funders (n=36) to explore their priorities in supporting research software. The survey gathered data on the specific outcomes funders emphasize in their programs and initiatives for research software.\n\nResults The survey revealed that funders place strong emphasis on developing skills, promoting software sustainability, embedding open science practices, building community and collaboration, advancing research software funding mechanisms, increasing software visibility and use, fostering innovation, and ensuring security.\n\nConclusions The findings highlight opportunities to enhance research software’s role through increased funder attention on professional recognition for software contributions and the non-technical, social aspects of research software sustainability. Addressing these areas could lead to more effective support and development of research software, ultimately benefitting the entire research ecosystem.", "keywords": [ "Research software", "research policy", "research funding", "science systems", "FAIR", "open science" ], "content": "Introduction\n\nResearch software is increasingly recognized as critical infrastructure in the contemporary research ecosystem (Barker et al., 2022; Jensen & Katz, 2023b; Knowles et al., 2021; Hocquet et al., 2024). Research software spans a broad spectrum, including source code files, algorithms, scripts, computational workflows, and executables, all created for or during research (Gruenpeter et al., 2021). With the growing reliance on computational methods across the sciences, supporting software development, maintenance, and impact is more essential than ever (e.g., Barker et al., 2020; Jay et al., 2021). For this reason, some research funders have developed programs, initiatives and policies to bolster research software’s role and impact (Hertweck et al., 2024; Strasser et al., 2022). The research software landscape includes government and philanthropic funding sources (Barker & Katz, 2022) that are seeking impact within the scientific ecosystem and beyond from their investments (e.g., Jensen et al., 2022). Research software is often open source and is sometimes tied to the larger open science movement that aims to improve transparency and reproducibility in research.\n\nTo date, there has been no empirical study of how software is supported by research funders, the aims of such support and the priorities that underpin it. This information is needed to clarify where current funder support is concentrated and highlight significant gaps. This study investigates the range of efforts by research funders to support the sustainability and impact of research software. The aim is for these findings to inform improvements in funder strategies that would bolster research software’s role and value for academic research and the wider society.\n\nThis paper draws on part of the dataset from this survey to address the following research question: What are the top priorities of international research funding organizations related to research software?\n\n\nMethods\n\nThis research was carried out using a survey combining qualitative and quantitative items. The survey was designed to investigate how research software funders support research software’s sustainability and impact.\n\nThe survey designed for this study began by collecting profile information, including institutional affiliation and job title. The survey gathered information about respondents’ organization’s initiatives, policies, or programs to support research software. The range of questions yielded too much data for one article. In this article, we focus exclusively on the results generated via an open-ended question asking about the top priorities for the respondents’ organizations’ support for research software: “What are your organization’s top priorities related to research software?” (see Jensen, 2024 for the extended data underpinning this article). Four open-response text boxes were provided for respondents to indicate and list these priorities.\n\nThis survey was aimed at international research funders, including governmental and non-governmental (e.g., philanthropic) funders. A list of contacts to invite to participate in this survey was created based on participation in the Research Software Association (ReSA) and responsibility for research software funding known to the authors. This initial list of people was refined, with removals based on individuals having moved to unrelated professional roles or being unavailable long-term, for example, due to personal issues.\n\nThe final, refined contact list comprised 71 people. After removing individuals when a member of their organization already provided a complete answer or when the person turned out to no longer be working on a relevant topic or to be otherwise unavailable (total of n=30), 41 people remained. Five of these individuals did not complete the survey, while 36 people (representing 30 research funding organizations) did, yielding a response rate of 87.8%. Fully completed survey responses were not required for individuals to be retained in the sample, resulting in varied sample bases across survey questions.\n\nThe sample includes research funders in North and South America, Europe, Oceania and Asia, but over-represents North America and European funder representatives. Some participating funders cover a broad spectrum of disciplines, while others focus on a particular domain such as social science, health, environment, physical sciences or humanities.\n\nThe respondents represented research funders supported by governmental (n=26), philanthropic (n=6) and corporate (n=1) resources.\n\nRespondents’ job titles span the following categories: Senior Leadership and Executive, such as a Vice President of Strategy; Program and Project Management, such as Senior Program Manager; Planning and Business Development; Scientific, Technical and IT, such as Scientific Information Lead.\n\nMost respondents 72.7% (n=24) answered ‘Yes’ to the question, “Has your organization established any policies, initiatives or programs aimed at supporting research software?”, while 18.2% (n=6) said ‘No’ and 9.1% (n=3) ‘Unsure’.\n\nData collection took place from December 2023 to May 2024. The mean completion time for the detailed survey was 28 minutes and 13 seconds.\n\nThe data were cleaned and prepared for analysis by removing any identifiable respondent details. The data analysis process followed a standard thematic qualitative analysis approach (e.g., Jensen & Laurie, 2016). This involved first identifying themes and organizing the data accordingly. Dimensions of each theme were identified where relevant. Then data extracts were selected from the survey responses associated with each theme and theme dimension. In line with qualitative research methodology, the focus is on presence or absence of ideas in the survey responses, not quantification. However, prevalence has been used to organize the results section, as the more prevalent themes also come with a more extensive set of data extracts and potential dimensions. The results section below presents each theme in turn, with data extracts providing evidence to support the description of the themes.\n\n\nResults\n\nThe survey reveals several priorities driving research funders’ support for research software. These themes are presented below, ordered from the most to least prevalent categories. Many of these categories overlap, so the boundaries between themes are porous. After each point is explained below, a verbatim data extract in the form of a block quotation from the survey responses is presented to substantiate the explanation.\n\nThe first identified theme, Skills, shows funders are prioritizing equipping researchers and other research professionals with competencies in software development, maintenance, and sustainability best practices.\n\nSupporting the training and retention of qualified staff to ensure human expertise and support within and between institutions.\n\nFunders were keen to ensure sufficient skilled personnel capable of writing and maintaining software.\n\nSkills - ensuring that there are enough skilled people writing software.\n\nSuch skill development was viewed as foundational for research software as a field.\n\nBuilding the necessary skills, training, infrastructure, and incentives to support the growth of the [research software] field.\n\nRespondents noted a priority focus on researchers’ software skills, seeking to support “train [ing] researchers to cope with research software on different levels.”\n\nTraining: We have an active initiative to train researchers to build software and digital research infrastructure.\n\nHowever, international funder priorities also included software-related skills for other research professionals.\n\nInvest in the training of both professional research and support staff to be able to reuse, develop, and maintain sustainable research software.\n\nOverall, the funder representatives indicated that training and skills development support was an important pathway to advancing research software.\n\nSoftware Sustainability emerged as a significant priority for research software funders. Sustainability means “the software will continue to be available in the future, on new platforms, meeting new needs” (Katz, 2024). Indeed, the term sustainability was frequently used to answer this question about top priorities for research software funding, such as “supporting research software sustainability” or “identifying pathways to sustaining the research software.” In this section, we focus on non-financial aspects of software sustainability that respondents highlighted. Indeed, this priority was expressed in many ways.\n\nSustainability and reuse of software. How can we reduce legacy debt and ensure that research software is continuously improved and reused.\n\nReducing waste by maximizing the use of existing software was a notable focus evident in funders’ responses.\n\nFund research software sustainability, and in particular incentivize the reuse and improvement of existing research software by providing funding to improve existing software.\n\nSome responses evinced the view that sustainability needed to be supported because it was part of ‘best practice’ in research software: “Promote the best practices for the production of sustainable research software.” Others focused on keeping software relevant and useful as a pathway to sustainability: “Support improvements to essential open source tools in [a specific field of] research”. Another response highlighted the importance of having standardized measures and practices to ensure sustainability.\n\nEstablish metrics and best practices for software sustainability and integrate these into [the] software development lifecycle.\n\nIn sum, the responses signaled funders’ full-throated commitment to ensuring research software’s long-term viability and continuous improvement (also see Barker et al., 2022).\n\nThe theme of Open Science highlights respondents’ emphasis on promoting open sharing, reuse, and accessibility of research software, adhering to the principles of open science. This theme is evident in the responses of research software funder representatives, who highlighted the importance of “open source practices” in fostering a collaborative and transparent research environment.\n\nOne respondent articulates the goal of promoting open sharing and reuse through significant initiatives, stating, “To promote the open sharing and reuse of research software, mainly through the European Open Science Cloud.” Many funder representatives highlighted maintaining, improving, and “making research software reusable” in their survey responses. This indicates a strategic effort to leverage established platforms to facilitate research software’s open distribution and reuse, thereby enhancing its accessibility and impact.\n\nOne respondent took an expansive view of what was included in efforts to integrate open science principles within research software: “Open Science including Accessibility, Inclusion, Reproducibility, Recognition, and security.” Another respondent called for a more explicit integration of research software into open science policies, saying, “Include research software more explicitly in our open science policies.” This points to the need for clear policy frameworks that explicitly address the role of research software in open science, ensuring that it is systematically included and supported (e.g., see Jiménez et al., 2017). This set of responses reflects a commitment to fostering an environment where a wide range of users and developers can contribute to and benefit from open-source research software.\n\nThe theme of building community and collaboration among research software funders encompassed three main dimensions: Enhancing community engagement, fostering international collaboration, and sharing best practices through professional networks. These community-related priorities were ultimately aimed at spreading and embedding good practices in research software.\n\nThe emphasis on community engagement highlights the importance of involving diverse groups in developing and maintaining scientific software. Funders prioritize approaches that ensure high-quality software through community involvement. They seek to promote opportunities that engage new communities in software development, making these resources accessible to under-represented communities and broadening participation in data science.\n\nPromote opportunities to engage new communities in software development and make these resources accessible to under-represented communities interested in data science.\n\nAnother respondent emphasizes the importance of inclusivity in open source communities of practice:\n\nSupporting diverse participants (users and developers) in scientific open source (research) software.\n\nAdditionally, there is a focus on community-focused software development and dissemination, ensuring that the processes and outcomes of software projects align with the community’s needs and contributions.\n\nEnhance community-focused software development and dissemination.\n\nInternational collaboration was identified as another critical dimension, focusing on creating a global network for software production. Funders aimed to facilitate work across institutions to prevent duplication of efforts and ensure that lesser-resourced institutions can equally benefit from open-source software, whether it be code, databases, or extensive infrastructure. This collaborative approach leveraged collective expertise and resources, promoting efficiency and innovation.\n\nEnabling work across institutions so that there is no duplication of effort and lesser-resourced institutions can benefit from OSS (whether bits of code, databases, or huge infrastructure) as much as the larger institutions.\n\nMoreover, fostering international collaboration is seen as a way to enhance the production of research software by tapping into a diverse pool of global expertise.\n\nFoster international collaboration in the production of research software.\n\nFunders prioritized disseminating best practices as a means of building robust communities around software development. Respondents emphasized the need for networks to share successful strategies and methodologies, ensuring that effective practices are widely adopted. This was aimed at contributing to improving research software quality and community development.\n\nBest practice and community - ensuring that there are networks to disseminate best practice and build communities.\n\nIn addition to promoting best practices, there is a focus on developing community-based approaches for ensuring and improving the quality of scientific software.\n\nDevelopment of community-based approaches for ensuring and improving the quality of scientific software and code.\n\nThis response shows that funders linked community-building goals with practical concerns about continuous improvement and quality assurance of research software.\n\nThe theme of advancing research software funding encompasses “funding new research software”, strategic planning of funding support, and fostering collaborative efforts to improve software impact.\n\nFunders noted the need for financial resources to support research software, for example, by “creating instruments to fund the production of high-quality research software.” They also emphasized the importance of developing plans to guide funding decisions over the medium term.\n\nOur primary priority is developing a strategy for how to fund [research software for the] next [several years].\n\nSuch a forward-looking approach is undoubtedly challenging in the constantly evolving research software landscape.\n\nCollaboration also emerged as a funder priority, with a systemic focus.\n\nFund joint developments of research software useful for the [research] ecosystem in collaboration with other institutions.\n\nOverall, the theme of advancing research software funding highlighted funders’ commitment to creating strategic funding for research software.\n\nFunders highlighted the need to make existing research software more visible to a broader range of researchers.\n\nProvide visibility to successful computational projects from users of our platform, not only about the science but also about the software being developed.\n\nAdditionally, there was a focus on enabling existing software to facilitate wider adoption and make it more accessible and usable.\n\nMake existing research software more visible [and] shape some of that software for wider adoption. [emphasis added]\n\nOne funder noted the need for a centralized research software portal where researchers could access information on supported platforms, tools, and services.\n\nDeveloping a comprehensive catalog of currently supported [research software] platforms, tools and services that researchers can access via [a] single portal, including the cloud resources needed to run those services.\n\nAnother funder’s related suggestion also focused on bolstering the visibility of research software through a shared portal or database.\n\nDevelop a profile of funding and support programs for [research software], with a focus on those infrastructures that have a national and international impact, but also including emerging RS platforms that have not yet established their role/impact.\n\nThe aim of increasing the visibility of research software in this way was “encouraging the [scientific] field to use research software.” Ultimately, these efforts aim to enhance the overall impact of research software.\n\nA small number of funders noted prioritizing integrating advanced technologies in software development. One funder noted the importance of “transitioning to accelerated compute” and maintaining pace with advancements in large-scale computing resources.\n\nCutting edge - ensuring that software is developed to keep pace with the next generation of large-scale compute and supports the best research.\n\nA specific ‘innovation’ example mentioned was the “use of Generative AI in software engineering.” Such priorities reflected a forward-looking approach from a small subset of funders aiming to ensure that research software continues to support research effectively.\n\nFunders raised the theme of “security” in the development and use of research software, including ensuring “data security,” enhancing software security, and conducting “Security Awareness Training.”\n\nSecurity: Research software will not be able to be reused if IT departments are not confident that it is secure[,] even for Open science projects.\n\nWhile infrequently mentioned compared to other themes and with limited detail in the responses, security was seen as a fundamental issue to be addressed for a small number of research software funders.\n\n\nDiscussion\n\nIn this paper, we have described the priorities guiding research software funders. Here, we consider the implications of these priorities for advancing research software’s role in the scientific ecosystem.\n\nWithin the skills development priorities highlighted by respondents, several technical training topics to support the design, development and deployment of research software were mentioned. This funder priority aligns with an identified need for training in research software skills (e.g., Carver et al., 2022; Cosden et al., 2022). Moreover, this skills theme suggests that there is recognition among funders that there is a need to invest in the people who underpin research software (e.g., Hartley & Barker, 2023; Katz, 2021). However, the more social aspects of research software sustainability and impact were not explicitly mentioned by any respondents. Addressing such social aspects and non-technical skills is an essential need in the field. For example, research software projects need to be able to identify relevant parties to engage with, maintain effective communication with current and potential users, establish a robust framework for monitoring and addressing evolving needs in a user community, and ensure good user experiences with the software. A recently published report highlighted “UX design, product management, and community management” as examples of key roles that often go unfunded in open source research software projects (Iacovou, 2024). Iacovou (2024) noted that this averred gap in available support was part of a broader need to address the critical social and community aspects of research software sustainability. While respondents in our survey explicitly noted community building and management, supporting the development of the skills underpinning this kind of work was not highlighted as a priority for research software funders.\n\nSoftware sustainability emerged as a prominent theme in responses by research software funders. Software sustainability is a complex and, in some ways, an irresolvable challenge due to ever-shifting contexts on the broader research ecosystem (e.g., Howison, 2020; Katz, 2024). However, there is clearly a need for funders to address not only the technical aspects of software sustainability but also the social and professional infrastructure that bolsters long-term sustainability (e.g., Carver et al., 2021).\n\nThe theme of Open Science in the survey responses emphasized promoting open sharing, reuse, and accessibility of research software. This is achieved through strategic initiatives, including open science principles, explicit policy integration, support for diverse participants, and promoting open source practices. For example, funding open-source tools has been a major focus in the Chan Zuckerberg Initiative’s Essential Open Source Software for Science (EOSS) program, which has focused on software for biomedical research. A recent report on the EOSS program highlighted wide-ranging benefits from this investment in terms of scientific progress and diversifying scientific open source software teams and communities (Hertweck et al., 2024). This suggests good alignment between funder priorities and the evidence base about what is yielding practical scientific and broader impacts.\n\nDifferent practical ways of ensuring that research software contributes to such a healthy and open ecosystem are being discussed in the research software field (e.g., McKiernan et al., 2023; Sonabend et al., 2024; Sellanga et al., 2024). For example, applying FAIR (Findable, Accessible, Interoperable and Reusable) principles to research software has been highlighted as an essential step in the literature (Barker et al., 2022). While our respondents mentioned concepts such as ‘accessibility’ and ‘reuse’, the term FAIR was not explicitly used. Overall, the survey responses evince a commitment to fostering an open, inclusive, and transparent research ecosystem. However, it is worth acknowledging that the focus on open source may be exaggerated in our purposive sample of funders already active in the research software arena, which is strongly oriented towards open source.\n\nThe theme of building community and collaboration was defined by efforts to enhance community engagement, foster international partnerships, and disseminate best practices. Advancing research software funding was also highlighted as a theme, with priorities including backing new software, strategic financial planning, and fostering collaborations. In these responses, funders emphasized the need for resources, strategic guidance, and joint efforts to enhance software impact. Promoting the visibility and use of existing software also emerged as a priority for research software funders. This included an interest in bolstering the awareness of available software by making it more prominent, cataloging and profiling available software resources, and encouraging software adoption. It has previously been noted that research software visibility can be bolstered through institutions that help to build community and collaboration, such as software sustainability institutes (Katz et al., 2021).\n\nA notable gap in the priorities identified by research software funders is the thorny challenge of bolstering recognition of research software contributions in academic hiring, promotion, and tenure (Jensen & Katz, 2023a). This may appear to be outside the purview of research software funders, but such ‘people’ aspects are a clear priority for those who produce and maintain research software (e.g., Cohen et al., 2021; Lamprecht et al., 2022). A recent article summarized ‘ten simple rules’ for addressing this challenge, including ensuring that software contributions are recognized as key scholarly contributions, consistent with open science principles (Puebla et al., 2024). While all of these rules/recommendations focus on the level of research institutions (Barker et al., 2024), research funders can play a pivotal role in nudging the priorities and actions of such institutions. Therefore, research software funders should consider how they could help encourage research institutions to adopt a software-inclusive stance within hiring, promotion and tenure.\n\nMuch less frequently mentioned themes were innovation in research software, stressing the need to stay ahead of rapidly shifting trends in computing and the need to ensure that research software is secure from threats. Adapting research software good practices to advancements in AI and related developments is a need that has also been highlighted in the literature (e.g., Duarte et al., 2023), and it is an ongoing challenge facing the research software field. The theme of security in research software development and use was defined by a strong focus on ensuring data security, enhancing software security, and promoting security awareness training.\n\n\nConclusion\n\nThe insights from research software funders reveal a commitment to enhancing the scientific landscape through targeted investments in software sustainability, open science, and community collaboration. Yet, the relative underemphasis on non-technical skills and social infrastructure suggests a need for a more holistic approach. To fully realize the potential of research software, funders must broaden their focus, integrating social and technical priorities to foster a resilient and dynamic research environment. By addressing such gaps, funders can help ensure that research software plays a more efficient, inclusive, sustainable, and impactful role within the scientific ecosystem.\n\n\nEthics and consent\n\nThe University of Illinois Urbana-Champaign Institutional Review Board reviewed this research and determined it to be exempt (no. 24374) on September 11, 2023. All participants provided explicit informed written consent to participate in this study at the start of the online form used to gather the survey data. This included consent for data to be used for research purposes and to be published as open data after deidentification.", "appendix": "Data availability\n\nRepository name: Zenodo\n\nTitle of project: Strategic priorities and challenges in research software funding: Results from an international survey [Dataset]. DOI: https://doi.org/10.5281/zenodo.13852004 (Jensen, 2024)\n\nThis project contains the following underlying data:\n\n• Survey data file: Anonymized responses from the international survey on strategic priorities and challenges in research software funding gathered from representatives of research funding organizations.\n\nDetails of license: The anonymized survey data are available under the Creative Commons Attribution Non-Commercial No Derivatives 4.0 International license.\n\nRepository name: Zenodo\n\nTitle of project: Strategic priorities and challenges in research software funding: Results from an international survey. DOI: https://doi.org/10.5281/zenodo.13852004 (Jensen, 2024)\n\nThis project contains the following extended data:\n\n• Consent block from survey: The information and questions used to gather informed consent through the online survey have been uploaded as a document file.\n\n• SURVEY DESIGN_Research_software_funder_survey_UIUC (upload to Zenodo).pdf\n\nDetails of license: The anonymized survey data are available under the Creative Commons Attribution Non-Commercial No Derivatives 4.0 International license.\n\n\nAcknowledgements\n\nThe Alfred P. Sloan Foundation funded this research via a grant to the National Center for Supercomputing Applications at the University of Illinois Urbana-Champaign for a project entitled “Charting the Course: Policy and Planning for Sustainable Research Software” (G-2022-19344). This project aims to devise strategies for enhancing the durability and sustainability of research software by bolstering the role of policy at various levels within the research ecosystem. The authors would also like to express appreciation for feedback from Professor Caroline Jay (University of Manchester).\n\n\nReferences\n\nBarker M, Chue Hong NP, Katz DS, et al.: Introducing the FAIR principles for research software. Sci. Data. 2022; 9: Article 622. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBarker M, Katz DS: Overview of research software funding landscape. Zenodo. 2022. Publisher Full Text\n\nBarker M, Katz DS, Gonzalez-Beltran A: Evidence for the importance of research software. Zenodo. 2020. Publisher Full Text\n\nBarker M, Carlin D, Cohen J, et al.: Resources for supporting policy change in research institutions in practice: A report from Subgroup 2 of the ReSA & RDA Policies in Research Organisations for Research Software (PRO4RS) Working Group. Zenodo. 2024. Publisher Full Text\n\nCarver JC, Cosden IA, Hill C, et al.: Sustaining research software via research software engineers and professional associations. Proceedings of the 1st International Workshop on the Body of Knowledge for Software Sustainability (BoKSS’21). IEEE; 2021; pp. 69–72. Publisher Full Text\n\nCarver JC, Weber N, Ram K, et al.: A survey of the state of the practice for research software in the United States. PeerJ. Comput. Sci. 2022; 8: e963. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCohen J, Katz DS, Barker M, et al.: The four pillars of research software engineering. IEEE Softw. 2021; 38(1): 97–105. Publisher Full Text\n\nCosden IA, McHenry K, Katz DS: Research software engineers: Career entry points and training gaps. Comput. Sci. Eng. 2022; 24(6): 14–21. Publisher Full Text\n\nDuarte J, Li H, Roy A, et al.: FAIR AI models in high energy physics. Mach. Learn.: Sci. Technol. 2023; 4(4): 045062. Publisher Full Text\n\nGruenpeter M, Katz DS, Lamprecht A-L, et al.: Defining research software: A controversial discussion. Zenodo. 2021. Publisher Full Text\n\nHartley K, Barker M: Investing in people: Anticipating the future of research software. Research Software Alliance Blog. 2023, October. Reference Source\n\nHertweck K, Strasser C, Taraborelli D: Insights and impact from five cycles of Essential Open Source Software for Science. Zenodo. 2024. Publisher Full Text\n\nHocquet A, Wieber F, Gramelsberger G, et al.: Software in science is ubiquitous yet overlooked. Nat. Comput. Sci. 2024; 4: 465–468. PubMed Abstract \| Publisher Full Text\n\nHowison J: Sustainability in scientific software: Ecosystem complexity and software visibility. Zenodo. 2020. Publisher Full Text\n\nIacovou G: Funding open source science software. Notion. 2024, June. Retrieved November 26, 2024. 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PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "344615", "date": "13 Dec 2024", "name": "Adrian Barnett", "expertise": [ "Reviewer Expertise Statistics", "research funding", "meta-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\nThe paper examines funding for software. Software must compete against other priorities, e.g., equipment. It also must compete against other more “exciting” ideas, and hence may be neglected despite its importance to research. As an example, here’s a relevant and recent skeet from the software goliath Hadley Wickham (27 November 2024):\n“I had zero luck with grants. People really didn’t get why what I did was important and never figured out how to explain it to them.” https://bsky.app/profile/hadleywickham.bsky.social/post/3lbumdqcmcc2r\nAnd an interesting reply to Hadley’s skeet: “lol thinking about you burying the tidyverse into a corner of aim 3 for an NIH grant on some very specific cardiovascular outcome, which is how most infrastructure work gets funded” https://bsky.app/profile/travisgerke.bsky.social/post/3lca7rkqv222z\nI don’t usually quote skeets or tweets in peer reviews, but these are good anecdotal examples of how software can miss out to other priorities. It also highlights how software is often funded as an add-on rather than the main idea.\nWithout breaking confidentiality, some more summary details on the funders would be useful to contextualise their answers and assess the generalisability of the results. Are the funders predominantly in science, engineering, or health and medicine? Or do they cover all these areas. What’s the total funding awarded per year? How many scientists per year do they support?\nThe sampling frame could be biased towards funders with a greater interest in software as: “participation in the Research Software Association (ReSA) and responsibility for research software funding known to the authors.” This should be discussed as a limitation. It would also be useful to describe who ReSA are.\nThe questions asked the funders about priorities, but did not ask for actual financial support. Funders can have many priorities, but with strong competition for funding they might not all get funded. Money spent on software would be the best measure of support. I realise this can’t be collected now, but some discussion of this would be useful.\nThe quotes from funders were generally very positive and I expected to see something in the results on the challenges of funding software. This prompted my previous question on wanting to know more about the funders, as I wondered if they were an atypical group of funders with a particular interest in software. Perhaps the funders were not prompted for their negative experiences?\nI was surprised that there was nothing on partnering with industry. In Australia the funding schemes that require partnerships with industry (or government) have a much higher success rate than standard schemes. Given the potential commercial benefits of software, partnerships would be an option for many software applications.\nMinor comments:\nSampling section (and elsewhere): “87.8%” – no need for a decimal place, see: Cole T. Too many digits: the presentation of numerical data, DOI: 10.1136/archdischild-2014-307149 The key question “What are your organization’s top priorities related to research software?” has no time frame. So I presume it was current priorities only? Although the question: “Has your organization established any policies, initiatives or programs aimed at supporting research software?” covers all times. A consistent time frame may have been better, for example, “in the last 5 years”. Discussion: I agree that funders have a part to play in increasing the recognition of software. Funders can be influential in how science is recognised and rewarded. The COARA initiative is likely relevant here, as it discusses needing to recognise the “diversity of contributions” which includes those scientists who create valuable software (https://coara.eu/agreement/the-commitments/).\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? 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? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "13014", "date": "06 Jan 2025", "name": "Eric Jensen", "role": "Author Response", "response": "We are grateful for this report's helpful and constructive reviewer comments. The reviewer report requests some additional information and raises some points for discussion. The details for how these points have been addressed in the new version of the manuscript are presented below: Reviewer 1: “Without breaking confidentiality, some more summary details on the funders would be useful to contextualise their answers and assess the generalisability of the results. [a] Are the funders predominantly in science, engineering, or health and medicine? Or do they cover all these areas. [b] What’s the total funding awarded per year? How many scientists per year do they support?” Response: [a] This information was already presented in the ‘sampling subsection’ of the methods section: “Some participating funders cover a broad spectrum of disciplines, while others focus on a particular domain such as social science, health, environment, physical sciences or humanities.” [b] this information was not collected in the survey. Reviewer 1: “The sampling frame could be biased towards funders with a greater interest in software as: “participation in the Research Software Association (ReSA) and responsibility for research software funding known to the authors.” This should be discussed as a limitation. It would also be useful to describe who ReSA are.” Response: This is already discussed as a limitation. However, more limitations content has now been added to the methods section: “ReSA is an international organization involving research policymakers, funders and organizations that recognize software’s importance in their work. This means that a ReSA-focused sample is likely to over-represent research funders who already view research software as important. For this reason, this paper does not claim that its findings generalize to the global population of research funders; rather, the claims focus on current patterns in research software funding.” Reviewer 1: “The questions asked the funders about priorities, but did not ask for actual financial support. […] Money spent on software would be the best measure of support. I realise this can’t be collected now, but some discussion of this would be useful.” Response: This point is now addressed in the survey design subsection of the Methods section: “Clearly, potentially relevant variables were not included in the survey design for this exploratory study. These include details on the exact amount of funding spent on research software and other details about which many research funders would be unable to readily report either because the information is not actively tracked or relevant data do not currently exist.” Reviewer 1: “I was surprised that there was nothing on partnering with industry. In Australia the funding schemes that require partnerships with industry (or government) have a much higher success rate than standard schemes. Given the potential commercial benefits of software, partnerships would be an option for many software applications.” Response: There is an important distinction between research software and software for more general use. Research software, per se, often attracts less industry interest than some other software domains due to its tendency to have a narrower market/commercial potential." } ] }, { "id": "344621", "date": "13 Dec 2024", "name": "Mario Coccia", "expertise": [ "Reviewer Expertise science development and policies" ], "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\nStrategic priorities and challenges in research software funding: Results from an international survey\nThe topics of this paper are interesting, though well known. The structure and content must be revised, and results have to be better explained by authors before to be reconsidered.\nTitle has to be shorter and more specific of the fields concerning funding research software.\n\nAbstract has to clarify the goal, methods, empirical research and research policies to support funding of software.\n\nIntroduction has to better clarify the research questions of this study, considering that software is an intangible innovation, clarifying the sources that are problem driven in specific contexts, such as medicine, environmental sciences, etc. Role of metrics that can assess the effectiveness of research software as an open science, such as citations and their association with funding in the diffusion of science, etc. After that they can focus on the topics of this study to provide a correct analysis for fruitful discussion (See suggested readings that must be all read and used in the text).\n\nMethods of this study is not clear. The section of Materials and methods must be re-structured with the following three sections: •\n\nSample and data. The sample is not balanced between continents and also respondents and authors have to clarify how this is representative. •\n\nMeasures of variables under study •\n\nData analysis procedure. Insert a flow chart to specify the study design, with inclusion and exclusion criteria of organizations and respondents under study. The identifications of themes and key aspects can be done with specific quantitative approaches, and not only qualitative, such also as entity linking, etc.... now the analysis is rather vague.\n\nResults. The sample can be represented with some figures to be clearer for readers. Authors have to avoid subheadings that create fragmentation and confusion. If necessary, you can use bullet points (same comments for section of results and all sections). Results can be also systematized in some tables to be more effective in communicating results. I suggest using different levels of description results from general to more specific themes, using tables to be more effective.\n\nDiscussion. First, authors have to synthesize the main results in a simple table to be clear for readers and then show what this study adds compared to other studies sone on these topics. To reiterate, avoiding in the just mentioned sections, sub-headings that create fragmentation of the paper. The results should be better discussed considering the research fields because the sources of research software, their goal, their adopters change according to the research fields, such as biomedical research, environmental sciences, economic sciences, etc....role of research software, funding and citations for diffusion of open science should be useful to decide specific policies. A SWOT matrix with pros and cons can support managerial implications.\n\nConclusion has not to be a summary, but authors have to focus on manifold limitations of this study and based on results, provide best practices of research policies to support the development of research software having practical implications for the diffusion of knowledge (also with citations) and development of science.\n\nOverall, then, the paper is interesting, but theoretical framework is weak, and some results, too descriptive, create confusion… structure of the paper has to be improved; study design, discussion and presentation of results have to be clarified using suggested comments.\nI strongly suggest improving the paper by using all comments (suggested papers included reading and using all) and maybe it can be considered.\n\nSuggested readings.\nAfiaz, A., Ivanov, A.A., Chamberlin, J., ... Leek, J.T., Wright, C. 2024 (Ref -1). Best practices to evaluate the impact of biomedical research software-metric collection beyond citations, Bioinformatics, 40(8), btae469\n\nSochat, V., May, N., Cosden, I., Martinez-Ortiz, C., Bartholomew, S.\n\n2022 (Ref-2). The Research Software Encyclopedia: A Community Framework to Define Research Software.Journal of Open Research Software, 10(1)\n\nCoccia M. 2017. Sources of technological innovation: Radical and incremental innovation problem-driven to support competitive advantage of firms. Technology Analysis & Strategic Management, vol. 29, n. 9, pp. 1048-1061, https://doi.org/10.1080/09537325.2016.1268682\\\nCosden, I.A. 2022 (Ref-3). ,The Princeton University Research Software Engineering Group Model: Operational and Organizational Approaches.Computing in Science and Engineering, 24(5), pp. 24–31\n\nCoccia M., Roshani S. 2024 (Ref-4). General laws of funding for scientific citations: how citations change in funded and unfunded research between basic and applied sciences. Journal of Data and Information Science, 9(1), 1–18. https://doi.org/10.2478/jdis-2024-0005\nGomez-Diaz, T., Recio, T. 2022 (Ref-5). Research Software vs. Research Data II: Protocols for Research Data dissemination and evaluation in the Open Science context.F1000Research, 11, 117\n\nRoshani S., Bagheri R., Mosleh M., Coccia M. 2021 (Ref-6). What is the relationship between research funding and citation-based performance? A comparative analysis between critical disciplines. Scientometrics 126, 7859–7874. https://doi.org/10.1007/s11192-021-04077-9\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": [ { "c_id": "13015", "date": "06 Jan 2025", "name": "Eric Jensen", "role": "Author Response", "response": "We are grateful for this report's helpful and constructive reviewer comments. The reviewer report requests some additional information and raises some points for discussion. While we don’t entirely agree with all points presented in this reviewer report, we have taken on board several points and made updates to the manuscript. Reviewer 2: “Title has to be shorter and more specific of the fields concerning funding research software.” Response: The relevant research fields for the participating research funders was already presented in the ‘sampling subsection’ of the methods section: “Some participating funders cover a broad spectrum of disciplines, while others focus on a particular domain such as social science, health, environment, physical sciences or humanities.” Because this includes all research fields, it wouldn’t be appropriate to claim a narrower scope in the title. Reviewer 2: The reviewer makes several requests for a wider paper scope, e.g.: “Introduction has to better clarify the research questions of this study, considering that software is an intangible innovation, clarifying the sources that are problem driven in specific contexts, such as medicine, environmental sciences, etc. Role of metrics that can assess the effectiveness of research software as an open science, such as citations and their association with funding in the diffusion of science.” Response: These issues are beyond the scope of the current paper, which focuses on open-ended survey data from research funders. Imposing focus areas that did not emerge from these open-ended responses would be methodologically problematic. The research question currently stated in the manuscript is deliberately broad in line with the qualitative nature of the data that is being presented in the paper: “What are the top priorities of international research funding organizations related to research software?” Reviewer 2: “Methods of this study is not clear. The section of Materials and methods must be re-structured with the following three sections: […] Insert a flow chart to specify the study design, with inclusion and exclusion criteria of organizations and respondents under study. The identifications of themes and key aspects can be done with specific quantitative approaches, and not only qualitative, such also as entity linking, etc.” Response: The current Methods section structure follows the journal guidelines and is appropriate for a qualitative survey study like this. A purely qualitative analysis and reporting of results is permitted by this journal; adding quantitative methods is not required. Reviewer 2: “Authors have to avoid subheadings that create fragmentation and confusion. If necessary, you can use bullet points (same comments for section of results and all sections). Results can be also systematized in some tables to be more effective in communicating results. I suggest using different levels of description results from general to more specific themes, using tables to be more effective.” Response: We appreciate the reviewers ideas about how to present the results. However, the current structure is in line with accepted practices in qualitative research. Each subheading represents a distinct theme, so it would be inappropriate to force them together. Reviewer 2: Raises concerns about the Discussion section structure Response: While the reviewer’s feedback about the Discussion section is noted, it is presented in a manner consistent with the journal guidelines and normal social science practices. Adding a table, for example, would be out of line with typical Discussion section approaches for this kind of paper. Suggested readings: Because none of the suggested readings directly relate the research software funding, these papers have not been added to the citation list despite the reviewer’s insistence on adding citations to the reviewer’s published work." } ] } ]	1	https://f1000research.com/articles/13-1447
https://f1000research.com/articles/12-1601/v1	19 Dec 23	{ "type": "Research Article", "title": "Evaluation of Entomopathogenic Nematode Isolates against Subterranean Termites under Laboratory and Field Conditions", "authors": [ "Belay Abate Gutema", "Dawit Melisie Achlehum", "Tariku Tesfaye Edosa", "Belay Feyisa", "Fikremariam Yimer", "Teshale Daba Dinka", "Dawit Melisie Achlehum", "Tariku Tesfaye Edosa", "Belay Feyisa", "Fikremariam Yimer", "Teshale Daba Dinka" ], "abstract": "Background Termites are a major insect pest affecting agricultural production and woody materials. They cause severe devastation in the ecosystem, and lead to bare soil. This phenomenon causes the soil to become difficult to plow, which in turn leads to a reduction in the productivity of crops. It can cause 100 % yield losses based on crop types, level of the damage, and size of its populations. To manage this pest, different management options have been evaluated in Ethiopia. While insecticide usage is the dominant option, less attention has been given to Entomopathogenic Nematode (EPN) based management options. Therefore, this research was initiated to screen locally collected EPN isolates and evaluate promising isolates under field conditions on maize crop.\n\nMethods 37 EPN isolates were screened under laboratory condition, while two isolates were evaluated at field condition. The screening of EPN isolates was laid out in a completely randomized design, and the field evaluation used a completely randomized block design, and treatments were replicated thrice. Mortality of insect, damaged root, stem, cob, damage severity, foraging termites, and yield of the crop data were collected.\n\nResults The study indicated that all screened EPN isolates caused mortality on termites under laboratory conditions. The isolates achieved complete mortality of the insect pest within 12 days of exposure. The finding indicated that AEH and S#50 were the more pathogenic and virulent isolates on termites under laboratory conditions and taken to field study. The S#50 isolate was most pathogenic and reduced the infestation and severity of the insect pest on the maize crop under field conditions.\n\nConclusions This result showed that the entomopathogenic nematode isolates have the potential to manage subterranean termites in the maize field. Future studies should be based on collection of local isolates and develop a full package for the virulent isolates.", "keywords": [ "Keywords: AEH", "Damage", "Isolate", "Pathogenicity", "S#50" ], "content": "Introduction\n\nTermites are a major insect pest affecting agriculture in Ethiopia, especially in the western part of the country. In this area four termite generals from the Macrotermitinae subfamily are the dominant species affecting crop productions. Among them the Macrotermite general is the economically important insect pest in the country (Abdurahman, 1990). From 61 species recorded, 10 were confined to the country. Macrotermes sp. and Microtermes spp. were the major termites affecting agricultural production and woody structures in the country (Cowie et al., 1990). Abdurahman’s (1990) study showed that Macrotermes subhyalinus was the most common termite species found in Wollega and Assosa areas. The species was known to construct dome shaped mounds and was mostly observed by its foraging nature of the whole plant parts, crop stacks and woody litter lying on the soil surface.\n\nTermites also cause severe devastation in the forest, leaving the soil bare and soil elements exposed to erosion (Abraham, 1990; Kumar and Pardeshi, 2011; Bong et al., 2012). As a result, farmers are forced to abandon their farmland (Abraham and Adane, 1995). Yield loss depends on the type of crop, the extent of the population, and the degree of termite infestation of the crop at different growth stages. Abdurahaman (1990) found that 45% removal of the crop at the six-leaf crop resulted in 16.5% yield loss, while the same reduction in the tassel stage resulted in 39.9% yield loss. But the severe infestations of termite species in Ethiopia can result in crop losses of up to 100% (Nyeko et al., 2010).\n\nTermites are most likely to target plants that are unfamiliar to the area and plants that are under water stress. Damage caused by these termites also provides access for secondary infections for pathogens, particularly aspergillus. Soil infested with termites usually leads to soil structure distortions and compaction. Therefore, the soil becomes difficult to plow, which in turn leads to a reduction in the productivity of crops (Devendra et al., 1998; Hailemichael et al., 2013; Legesse et al., 2013).\n\nSince the 1980s different termite management practices have been conducted to control termites in western Ethiopia. As an example, in 1983 the Ministry of Agriculture had poisoned 635,908 mounds by using Aldrin 40% WP insecticide in Menesibu and Nedjo-Jarso areas (Abdurahman, 1990). A similar method was used in Asosa and Anger Gutin on 2145 mounds which took 201 kg of the aforementioned product (Abdurahman, 1990). Additionally, 557,563 mounds were treated by 13,077 kg of Heptachlor 40% in Menesibu, Nedjo-Jarso, Ghimbi, Ayra-Guliso and Yubdo areas which was led by the Ministry of Coffee and Tea Development. On the other hand, more than 23,000 termite queens were collected as a management option of the pest during 1987-1988 in Ayra-Guliso area by the initiatives of Western Synod branch (Abdurahman, 1990).\n\nSimilarly, different plant botanicals have been evaluated for termite control in Ethiopia. As an example, neem seed extract was applied at 40 kg/ha and reduced the infestation of termite on hot pepper seedling in the Tanqua Abergelle district (Gebreslasie and Meressa, 2018). Additionally, Nicotiana tabacum leaves and Milletia ferruginea seed extracts recorded complete mortality with 24 hours of the exposure time to termites under laboratory conditions. Whereas Phytolacca dadecandra leaf extracts achieved the same results after 48 hours, and Azadirachta indica leaves, Hagenia abyssinica, Chrysanthemum sp. and Croton macrostachs seed extracts recorded higher mortalities after 72 hours on the same insect and condition (Tadele Shibiru et al., 2013).\n\nEven though termite management has been dominated by using persistent insecticides, it is not sustainable as a long-term solution and can affect biodiversity. Among the diverse potential alternatives available for termite management, the use of microbes is gaining prominence (Michael, 2005). Entomopathogenic fungi, Beauveria bassiana and Metarhizum anisophilae, show promise for the management of various termite species in the laboratory (Abebe, 2002; Milner, 2003; Addisu et al., 2014). However, reports of the use of entomopathogenic nematodes to control termites in Ethiopia are very limited and microbial-based treatment options have been limited to laboratory conditions. Although termites severely affect agricultural production in Ethiopia, an insufficient management option is available. Therefore, the study is initiated to screen locally collected entomopathogenic nematode isolates on termites under laboratory conditions and evaluate promising isolates under field condition on maize crop.\n\n\nMethods\n\nThis experiment was conducted within an appropriate ethical framework of Ethiopian Institute of Agricultural Research as confirmed by institutional letter with Ref. No.: 8.8/154/2023 written on 07 August 2023.\n\nThe laboratory experiment was conducted in the laboratory of the Entomology program at Ambo Agricultural Research Center, and the field evaluation was done at Bila district, East Wollega zone which is an insect pest-prone area. Termites, Macrotermes spp., were collected and established following Addisu et al. (2014) methods from Bako areas of the insect-prone maize field. Termite mounds were dug up using a spade, and soil containing termites was put on plastic sheets. The insects were collected from the plastic sheets using a camel-hair brush and placed in plastic boxes. Wooden plants (termite-infested materials) were added to the plastic boxes as feed for the termites. The top parts of the plastic boxes were covered with a mesh cloth that allowed air ventilation. A moistened wad of cotton was placed in the plastic boxes to maintain the moisture level for the survival of termites. The boxes carrying the termites were transported to the Ambo agricultural research center’s Entomology laboratory. Periodically, dry wooden materials were provided to the termite population, and the plastic boxes were inspected for maintenance of the required moisture level.\n\n37 newly collected and preserved entomopathogenic nematode isolates were used for this experiment (Table 1). Culturing and preparation of EPN were performed using standard methods described by Kaya and Stock (1997). Harvested nematodes were kept at 14°C in 250 ml flasks and used within a week of culture preparation. Each isolate was tested at 800 infective juveniles per ml concentrations and negative control (treated with sterile water) for comparison. 10 soldier termites were placed in a petri dish on filter paper and sprayed with the infective juvenile suspension. This study was conducted under laboratory conditions at 25 ± 2°C and 60% RH in dark places. The experiment was replicated three times and laid out in a completely randomized design.\n\nTwo EPN (AEH and S#50) isolates were applied at a rate of 800 IJ/ml as a basal application at the seedling and tasseling growth stages of maize. Likewise, the recommended rate of diazinon at 60% EC (2.5 lit/ha) was applied. Untreated check plots were neither treated with the insecticide nor with isolates and recommended agronomic practices were applied. The plot size was 4 m long and 3 m wide, with intra-row spacing of 75 cm and inter-row spacing of 25 cm, with 2 m and 3 m spacing between plots and blocks, respectively. Data collection started two weeks after the first treatment application and continued every two weeks until physiological maturity of the crop. 10 plants were randomly sampled per plot to assess termite damage.\n\nThe screening of EPN isolates was laid out in a completely randomized design, and the field evaluation used a completely randomized block design. Treatments were replicated three times in the experiments. AEH and S#50 isolates that recorded complete mortality of termites were considered promising isolates from laboratory results and taken to field conditions along with positive and negative checks. The following data were collected.\n\nDamaged root: all plants in the plot were inspected for the termite damage on maize root at harvesting time, and plant roots that showed damage from termites were counted and expressed in percentages. Damaged stem: all plants in the plot were inspected for the termite damage on the maize stem, and plants that showed damage from termites were counted. Damaged cob: at physiological maturity, all cobs of maize in the plot were checked for damage by termites and counted. Damage severity: five maize plants were randomly taken from the plot, and a 0 – 9 scale were used to assess their damage. 0 indicates the plant was not attacked by termites, and 9 indicates that the plant was eaten and lodged to ground. Lodged plants: at physiological maturity, all maize plants in the plot were checked for lodging, and the numbers of plants lodged were counted. Foraging termites: all foraging termites observed in the plot were counted. Yield: at physiological maturity, maize cobs were harvested from each plot. The threshed grains moisture content was adjusted at 11 %, maize grains were weighed to determine the amount of yield, and the plot yields were converted to kg ha-1. Data were analyzed using SAS version 9.4.\n\n\nResults and discussion\n\nAll entomopathogenic nematode isolates screened caused mortality in termites under laboratory conditions (Abate, 2023). These isolates showed significant differences among each other and with the negative control across the exposure time (Table 1). 18% of tested isolates caused greater than 50% mortalities on the insect pest after the 3rd day of the treatments’ application (Table 1). On this date, S#50 and AEH isolates recorded higher mortalities at 70% and 63.33%, respectively, while the lower mortalities were from the APPRC-PL0056 isolate (20%) and negative control (10%). On the other hand, half of the tested isolates recorded greater than 50% termite mortalities after the 6th day of the treatments’ application (Table 2). But S#50 and AEH isolates achieved 90% and 80% mortality on the insect pest on the same date, respectively. 10.53% of isolates recorded greater than 75% of mortalities on insect pests after the 9th day of their applications. Similarly, 92.11% of isolates caused greater than 50% of mortalities in the insect pest on this date. After the 12th day of treatments applications, tested isolates showed significant differences with the negative control (p < 0.001) (Table 1). On this date, AEH and S#50 isolates achieved complete mortality on the tested insect pest and showed significant differences with other. Similarly, 31.58 % and 94.74% of tested isolates recorded greater than 75% and 50% of mortalities on the insect pest, respectively. However, S#8 isolate recorded the minimum mortality (43.33%) among the entomopathogenic nematode isolates evaluated.\n\nThis result revealed that the tested entomopathogenic nematode isolates were pathogenic to the termites and caused mortality. The effectiveness of all isolates showed an increasing trend across the exposure durations. Some isolates achieved complete mortality of the insect pest within 12 days of exposure. The finding indicated that AEH and S#50 were the more pathogenic and virulent isolates on termites under laboratory conditions. Based on this result, AEH and S#50 isolates were considered as promising under field conditions. Similarly, microbials bioagent (Metarhizium anisoplie and Beauveria bassiana) had showed promising results under laboratory conditions for Macrotemes spp management (Addisu et al., 2014). Entomopathogenic nematodes (EPNs) are helpful nematodes that have a high potential for controlling insects in soil environments and are commercially employed to treat a wide range of pests (Verma et al., 2018). In a filter paper and sand test, Shahina and Tabassum (2010) found that EPN produced increased mortalities in the subterranean termite. Yu et al. (2010) investigated the pathogenicity of three new strains of Steinernema riobrave (3-8b, 7-12, and TP) against workers of Heterotermes aureus, Reticulitermes flavipes, and Coptotermes formosanus. Heterotermes aureus was shown the most sensitive to all S. riobrave strains, and termites in all nematode treatments died after four days. The TP strain caused 75% and 91% mortality in R. flavipes and C. formosanus, respectively. More research was suggested to determine S. riobrave (TP)’s potential to control the targeted termite species in the field condition (Yu et al., 2010).\n\nThe effectiveness of AEH and S#50 entomopathogenic nematode isolates showed a non-significant difference in the number of foraging termites and maize root damage in maize fields (Table 3). However, treatments indicated a significant difference in stem damage, lodged plant, cob damage, the severity of insect damage, and yield (p < 0.001) (Table 3). The lower stem damage was recorded by the S#50 isolate and positive control, while the higher stem damage was recorded by the AEH isolate. Similarly, the AEH isolate and negative control (untreated plot) showed higher cob damage and lodged plants and were more severely attacked by the insect pest, whereas S#50 and positive control showed significant differences, and lower lodged plants and cobs were damaged. Similarly, negative control and AEH isolates gave a lower maize yield (2111 kg/ha), while the highest yield was obtained from positive control (Table 3).\n\nThe experiment indicated that the S#50 entomopathogenic nematode isolate showed a non-significant difference with the positive control on all considered parameters except yield data. This revealed that the isolate is more pathogenic and reduced the infestation and severity of the insect pest on the maize crop under field conditions. Additionally, the S#50 isolate increased the maize yield by 13.63% over the negative control. This result showed that the entomopathogenic nematode isolates have the potential to manage termites in the maize field. Subterranean termites’ dwell and forage in damp, cool, and shaded environments such as soil or wood materials. These characteristics are favorable for the survival and mobility of nematodes which enhances their role in termite management. Wang et al. (2002) and Yu et al.’s (2006) studies showed that entomopathogenic nematodes have the potential to be used as an environmentally safe alternative termite control method. Steinernema spp. and Heterohabditis spp. provided some limited control of subterranean termites in field studies but were largely ineffective for long-term suppression (Mauldin and Beal, 1989). However, entomopathogenic nematodes have been used in classical, conservation, and augmentative biological control strategies. In addition to other nuisance insects, commercially generated EPNs are being used to manage scarab larvae in lawns and turf, fungus gnats in mushroom production, invasive mole crickets in lawns and turf, black vine weevils in nursery plants, and Diaprepes root weevils in citrus (Lacey and Georgis, 2012). Similarly, Aslam et al.’s (2023) study indicated that Steinernema carpocapsae, Heterorhabditis bacteriophora and Heterorhabditis indica species recorded higher mortality of termites under field conditions. As a result, they suggested that indigenous EPNs can provide more effective termite control, presumably due to their direct interaction with pest species in the soil and the likelihood of secondary infection via infected cadavers.\n\n\nConclusion\n\nAll screened entomopathogenic nematode isolates caused mortalities on termite under laboratory condition, and their pathogenicity was varied among isolates. The study showed that the S#50 and AEH isolates were more virulent than others and achieved complete mortalities on the tested insect pests within 12 days of the exposure time. As a result, these isolates were considered as promising isolates and tested under field condition. Similarly, S#50 isolate showed lower stem damage, lodged plant, cob damage, and severity of insect damage on the maize under field condition. From this experiment it can be concluded that entomopathogenic nematodes have a potential for termite management and part of integrated the insect pest management. Therefore, future studies should be focused on the collection of entomopathogenic nematodes from the insect cadaver at insect pest probe areas, screening, evaluating and considering it integrated in the insect pest management components, and develop a full package product for the promised isolates.", "appendix": "Data availability\n\nfigshare: EPN dataset. https://doi.org/10.6084/m9.figshare.24208092.v1 (Abate, 2023).\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nARRIVE checklist for ‘Evaluation of entomopathogenic nematode isolates against subterranean termites under laboratory and field conditions’. https://doi.org/10.6084/m9.figshare.24526885.v1\n\n\nAcknowledgments\n\nThe authors acknowledged the Ethiopian Institute of Agricultural Research (EIAR) and Ambo Agricultural Research Center for their facilities support.\n\n\nReferences\n\nAbate B: EPN dataset. [Dataset]. figshare. 2023. 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Nematol. 2012 Jun; 44(2): 218–225. PubMed Abstract \| Free Full Text\n\nLegesse H, Taye H, Geleta N, et al.: Integrated termite management in degraded crop land in Diga district, Ethiopia. In. Wolde, M. (ed). 2013, Rainwater management for resilient livelihoods in Ethiopia: Proceedings of the Nile Basin Development Challenge Science Meeting, Addis Ababa, 9–10 July 2013. NBDC Technical Report 5. Nairobi, Kenya: ILRI.2013. Reference Source\n\nMauldin JK, Beal RH: Entomogenous nematodes for the control of subterranean termites, Reticulitermes sp. (Isoptera: Rhinotermitidae). J. Econ. Entomol. 1989; 82: 1638–1642. Publisher Full Text\n\nMichael L: Biological control in termite management: The potential of nematode and fungal pathogen.Proceedings of the 5th International Conference on Urban Pests.July 11-12, 2005, Singapore, pp. 48–50.\n\nMilner RJ: Application of biological control agents in mound building termites (Isoptera: Termitidae): Experirnces with Metarhizium in Ausrtalia. Sociobiology. 2003; 41: 419–428.\n\nMola T: Subterranean Termites Management using Different Legume Crops and Chomo Grass (Brachiaria humidicola) as Green Manure on Tef under Acidic Soil Condition at Nejo West Wollega Ethiopia. Inter. J. Agri. Biosci. 2018; 7(4): 200–206.\n\nNyeko P, Glohole LS, Manainia NK, et al.: Evaluation of Metarhizium ansophiliae for integrated management of termite on maize and Grevilliea robusta in Uganda and Kenya. Proceedings of the 2nd RUFORUM Biennial Meeting.September 20-24, 2010, Entebbe, Uganda.\n\nShahina F, Tabassum KA: Virulence of Steinernema pakistanense against different insect species in laboratory condition. Pak. J. Nematol. 2010; 28: 279–284.\n\nShiberu T, Ashagre H, Negeri M: Laboratory Evaluation of Different Botanicals for the Control of Termite, Microterms spp (Isoptera: Termitidae). Open Access Scientific Reports. 2013. Publisher Full Text\n\nSileshi A, Sori W, Dawud M: Laboratory evaluation of Entomopathogenic fungi Metarhizium anisophilae and Beauveria bassiana against termite, Macrotermes (Isoptera: Termitidae). Asian J. Plant Sci. 2013; 12: 1–10. Publisher Full Text\n\nSisay A, Ibrahim A, Tefera T: Management of Termite (Microtermes adschaggae) on Hot Pepper Using Powdered Leaves and Seeds of Some Plant Species at Bako, Western Ethiopia. East Afr. J. Sci. 2008; 2(1): 41–44. Publisher Full Text\n\nVerma P, Yadav AN, Kumar V, et al.: Microbes in Termite Management: Potential Role and Strategies.Khan A, Ahmad W, editors. Termites and Sustainable Management, (Volume 2 - Economic Losses and Management. Gewerbestrasse 11, 6330 Cham, Switzerland: Springer International Publishing; 2018; pp. 197–219.\n\nWang C, Powell JE, Nguyen K: Laboratory evaluations of four entomopathogenic nematodes for control of subterranean termites (Isoptera: Rhinotermitidae). Environ. Entomol. 2002; 31: 381–387. Publisher Full Text\n\nYu H, Gouge DH, Baker P: Parasitism of subterranean termites (Isoptera: Rhinotermitidae:Termitidae) by entomopathogenic nematodes (Rhabditida: Steinernematidae; Heterorhabditidae). J. Econ. Entomol. 2006; 99: 1112–1119. Publisher Full Text\n\nYu H, Gouge DH, Shapiro-Ilan DI: A novel strain of Steinernema riobrave (Rhabditida: Steinernematidae) possesses superior virulence to subterranean termites (Isoptera: Rhinotermitidae). J. Nematol. 2010; 42: 91–95. PubMed Abstract" }	[ { "id": "236149", "date": "07 Feb 2024", "name": "Preety Tomar", "expertise": [ "Reviewer Expertise Entomopathogenic nematology", "Biocontrol", "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\nThe research article entitled \"Evaluation of Entomopathogenic Nematode Isolates against Subterranean Termites under Laboratory and Field Conditions\" is very impressive work conducted by the authors. The article gives the complete information about the problems due to termite infestation and their management strategies. The results of the present investigation are highlighting the bio-control potential of entomopathogenic nematode isolates against termites not only in the laboratory but also under field conditions. However, its a suggestion that do mention which statistical analysis you have applied to compile the data and formulated the results. Also add more information about the maize sowing and how you placed and distribute the termites over these plants under field conditions.\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? Yes", "responses": [ { "c_id": "11088", "date": "13 Apr 2024", "name": "Belay Abate", "role": "Author Response", "response": "Based on the reviewers’ suggestions, minor revisions are made to the methodology and reference. In the methodology part, the collection site of SY isolate is Yirgalem, Ethiopia. Additionally, the insect and crop data collected were analyzed using statistical analysis software version 9.4. The least significant difference (LCD) at the 5% level was used to compare treatment means. For the field experiment, the Jibat variety of the crop was used and studied under the natural infestation of termites. Two seeds per hill were sown, and then, after emergence, the seedlings were thinned to adjust the population of the crop to one plant per hill. This is based on the maize production manuals of Ethiopia, which recommend 25 kg of maize seed per hectare. And the revision is also done for reference." } ] } ]	1	https://f1000research.com/articles/12-1601
https://f1000research.com/articles/14-21/v1	03 Jan 25	{ "type": "Research Article", "title": "Assessing psychogenic health risks through the days between death and birth in the individual annual cycle", "authors": [ "Rolf Weitkunat", "Karim Abawi", "Rachelle El Tannouri", "Laura Inglin", "Célia Magnin", "Tonia Rihs", "Katharina Fehst", "Karim Abawi", "Rachelle El Tannouri", "Laura Inglin", "Célia Magnin", "Tonia Rihs", "Katharina Fehst" ], "abstract": "Background The \"birthday effect\" refers to increased mortality around birthdays, although evidence remains mixed, with some studies even indicating reduced mortality. While the effect has a minimal impact on overall mortality, lacks prevention potential, and faces methodological challenges, it offers seminal insights into psychogenic health effects.\n\nMethods Major causes of death were analyzed using data from the Swiss Federal Statistical Office (1987-2022), excluding leap years and cases with imputed dates. The relationship between day of birth and death was assessed using a Poisson count model with overdispersion, and mortality rates (MRs) per 100,000 deaths per year were calculated for ±182 days around birthdays. Mortality rate ratios (MRRs) compared these periods to a reference period. Cluster analyses explored mortality rate patterns across age and diagnostic groups. Age was not adjusted for in the Poisson model to avoid zero inflation, with bias from age-related mortality changes assessed separately through sensitivity analyses.\n\nResults In the analysis of 1,702,865 deaths over 27 years, an eight percent increase in birthday mortality was observed, equivalent to 22 additional deaths per 100,000 deaths per year. There were 889 excess deaths in the 28 days after the birthday compared to 206 in the 28 days before. Excess birthday mortality was particularly pronounced in men, unmarried individuals, and those with religious affiliations. In individuals aged 85 and older, a pre-birthday dip followed by post-birthday mortality excess was noted. Cardiovascular causes, suicides, and accidents had the highest birthday-related mortality, while diabetes, respiratory diseases, and other conditions showed varied pre- and post-birthday patterns.\n\nConclusions Administrative and age-related biases were ruled out, suggesting the birthday effect is psychogenic. While behavior-linked causes like suicides and accidents had clear birthday patterns, other main causes of death also showed birthday-period effects. Further research, including psychometric and biomarker studies, is needed to elucidate the underlying mechanisms.", "keywords": [ "epidemiology", "mortality", "life span", "timing of death", "birthday", "psychosomatic aspects" ], "content": "Introduction\n\nThe term “birthday effect” has been used to designate increased mortality on or around birthdays. Based on a wide range of data and analysis methods, both supportive and contradictory results from different populations and subgroups have been published since the late 1970s, including findings on reduced birthday-related mortality. Three often cited large population-based studies have reported birthday and post-birthday mortality increases. Alderson (1975), in national mortality records from England and Wales (1972), found an increase in deaths on the birthday month and over the months following. Phillips et al. (1992), in California death registration data (1978-1990), observed increased mortality primarily in the week following birthdays, with the effect being stronger in women than men. Ajdacic-Gross et al. (2012), in Swiss mortality records (1969-2008), found elevated mortality on the birthday itself and shortly after, especially among women. Among the three studies, the mortality increases on birthday ranged from one to almost 14 percent.\n\nFocusing on deaths from causes other than suicide, several studies, in addition to Alderson (1975) and Ajdacic-Gross et al. (2012), have found increased mortality on the birthday itself. Grigsby (1985; US National Mortality Survey, 1966-1968) found more deaths, particularly of cardiovascular causes, in the month of the birthday. Vaiserman et al. (2003; Ukrainian mortality records, 1990-2000) reported mortality increases of about 44 percent in men and 36 percent in women on the day of birth. Bovet et al. (1997; Swiss mortality records, 1969-1992) reported increases of 15 percent in men and 19 percent in women compared to expected levels on the day of birth. Peña (2015; US mortality records, 1998-2011) reported an excess death rate of 6.7 percent on birthdays.\n\nPost-birthday mortality increases, which might indicate postponement effects, have been observed in other studies besides those of Alderson (1975), Phillips et al. (1992), and Ajdacic-Gross et al. (2012). Byers et al. (1991; Ohio mortality records, 1979-1981) reported an excess in deaths of about five percent in the month following compared to the month preceding the birthday. Comparing deaths occurring during the quarters before and after the birthday, Kunz and Summers (1980; US data from various sources) found that almost half of all deaths occurred in the quarter after the birthday, both for natural and non-natural causes. The same pattern of results was reported by Shimizu and Pelham (2008; based on more than 30 million US death records), with people more likely to die just after rather than just before their birthday. Further pre-birthday effects have been published by Phillips et al. (1992), who found a dip in deaths before birthdays in women, and by Grigsby (1985), who found a dip in cardiovascular deaths in the month before the birthday. Increased levels of fatal accidents have been reported before (as well as on) birthdays (e.g., Greiner & Pokorny, 1989; Ajdacic-Gross et al., 2012; Matsubayashi & Ueda, 2016).\n\nConversely, Greiner and Pokorny (1989; US cohort of male veterans with mental disorders, 1972-1978) as well as Brenn and Ytterstad (2003; Norwegian mortality records, 1991-1995) did not find birthday effects. Studies focusing on special populations and methods have provided limited insights into the birthday effect. Stumpfe (1983), based on collecting newspaper death notices (Cologne, Germany, 1979-1980) and Angermeyer et al. (1987), examining celebrities, reported no birthday effects. Other such studies have found birthday effects: Phillips and Feldman (1973), investigating biographical information on famous persons; Abel and Kruger (2009), investigating baseball players (up to 2004); Kelly and Kelleher (2018), collecting dates of birth and death of famous individuals on the internet. These studies suffer from combinations of low statistical power, potential selection bias due to unconventional sampling methods, and limited generalizability.\n\nRegarding specific causes of death, Brown and Knapp (1995; New York and Ohio cancer registries, 1989) reported no differences in mortality levels in cancer patients near birthdays compared to more remote periods. Likewise, Young and Hade (2004; Ohio cancer registry data, 1989-2000) as well as Medenwald and Kuss (2014; German cancer registry data, 1995-2009) found no evidence of altered mortality among cancer patients around their birthdays. Research on cancer has therefore thus far provided quite consistent findings of no effect. Interestingly, cancer mortality also does not seem to exhibit the seasonality seen in many other causes of death (Young & Hade, 2004).\n\nThe results on suicide are mixed. Several studies found increased suicide risks on as well as before and after birthdays, often with stronger effects observed in men than in women. Barraclough and Shepherd (1976; UK sample of 247 suicides), Kunz (1978; German suicide data), Zonda et al. (2010; Hungarian suicide data, 1970-2002), Stickley et al. (2016; Tokio suicide registration, 2001-2010), Matsubayashi and Ueda (2016; Japanese mortality records, 1974-2014) and Matsubayashi et al. (2019; Japanese mortality records, 1974-2014), all reported increased suicide mortality risks. For instance, Zonda et al. (2010) and Stickley et al. (2016) found that the increase in suicides around birthdays ranged from about ten to 45 percent, indicating a robust birthday effect. In contrast, several other studies did not find an association between suicides and birthday periods, including Lester (1986, 1997, Medical Examiner of Philadelphia, (212 suicides of 1982), and sample of 201 famous suicides with complete dates of birth and death, respectively), Wasserman and Stack (1994; Ohio suicide records, 1989-1991), Panser et al. (1995; Olmsted county, Minnesota, suicide data), Chuang and Huang (1996; Taiwan suicide registration), Jessen and Jensen (1999; Danish suicide register, 1970-1994), Reulbach et al. (2007; German mortality records, 1998-2003), Williams et al. (2011; UK mortality records), and Deisenhammer et al. (2018; Austrian suicide records, 1994-2014).\n\nThe heterogeneity in published results, methods, and reporting is so pronounced that trying a meta-analysis has all the makings of a non-starter. Rather, addressing the methodological shortcomings of existing research seems to be more promising. Many studies are small and do not cover whole populations, fail to adjust for the well-established effects of season (cf. Marti-Soler et al., 2014) and day of the week (cf. Trudeau, 1997; Brenn & Ytterstad, 2003), do not address potential systematic errors such as age-related mortality gradients (Roger, 1977) and heaping (Abel & Kruger, 2006), and neglect to stratify findings by demographic and diagnostic groups. The time resolution of the analysis varies greatly, from days to weeks to months to quarters, and many publications also do not provide precision statistics alongside point estimates, and often have inadequately described methods and give insufficient descriptive results. Additionally, the use of non-comparable statistics, such as counts instead of rates, hinders assessing results across studies. While this is a bleak outlook after half a century of research on the matter, it highlights the opportunity to address the identified shortcomings. Consequently, the present study aims to overcome these issues and maximize methodological rigor in exploring potential birthday effects.\n\nWith the evidence heterogeneous and the validity of the underlying data as well as their analyses questionable, not only is the magnitude, distribution, and nature of the phenomenon currently unclear, but its very existence is uncertain. This raises the fundamental question as to why one should continue investigating a phenomenon that, if it exists at all, has at best a minimal impact on mortality, largely lacks any discernible potential for prevention, and is fraught with methodological challenges. The answer lies in its unique opportunity to investigate the role of psychogenic factors in death and, by extension, in health more generally.\n\nWhereas ultimately death always entails the unsustainable loss of the structural or functional form of the body or some of its parts, its less proximate causes fall into three rather disparate and broad categories, namely external (often regarded to include both environmental and social factors), somatic, and psychogenic factors. As people keep on living, repeated and accumulating exposures to external, bodily, and subjective-behavioral factors combine and accumulate in numerous ways to eventually manifest as specific diseases, conditions, or injuries that ultimately lead to death. These underlying states are categorized into major groups, most famously by the International Classification of Diseases. While environmental, social, and (increasingly) biological risk factors lend themselves to objective measurement and epidemiological assessment, things are more complicate when it comes to psychological ones. For some health-related behaviors (like the use of toxic substances or the lack of physical activity) and their consequences (like obesity), and to a lesser extend for others (like nutrition), good progress has been made over the last few decades on quantifying their relevance. Estimates of their combined impact on total mortality, in particular in Western countries, of more than 50 percent are not uncommon (e.g., Knoops et al., 2004). In contrast, investigating non-behavioral psychogenic factors is considerably more challenging. Aside of fundamental conceptual challenges (mind-body problem) and problems of objective measurement, there is the notorious issue of psychological variables (e.g., emotional states) always being under the influence of biological factors which in turn are, naturally, linked to health. As a result, any empirical association between non-behavioral psychogenic factors on the one hand, and any kind of health outcome (including death) on the other, is prone to – typically unmeasured – confounding by biological variables. This is the fundamental methodological problem of psychosomatic research.\n\nA route to circumvent the threats of unmeasured confounding has been the use of instrumental variables (cf. Angrist et al., 1996), a common challenge being to find a valid instrument. In the current context, the instrument would have to stand in for non-behavioral psychological measurements. The advantage of having a valid instrumental variable would be, if certain general conditions hold, that associations between the instrument and the outcome (death) could not be confounded by biological variables. These conditions, often referred to as relevance, exclusion restriction (birthdays conditionally independent of outcomes given psychological factors), and independence (birthdays uncorrelated with any cause of the outcome), would in the present context come down to the instrument causally affecting death, but only through its impact on psychological variables, and itself not being caused by a factor that also causes death. In psychosomatic research, it has long been noted (even if not always in so many words) that life events can be instrumental variables. However, life events present technical challenges, such as lack of uniform definitions or availability in civil registration databases (e.g., changing working conditions, vacations, or relationships). Others, with clear definitions and general applicability, are compromised by closely associated social events (e.g., New Year's Eve or national holidays), making it difficult to separate individual from group effects. In contrast, birthdays are reliably documented in public records and are independent of general social events or the birthdays of others which, other than with public anniversaries, warrants stable unit effects through non-interference and absence of individual- and group-level spillover.\n\nIn spite of these advantageous properties, there are four main reasons why birthday – or more specifically, the death-to-birth day interval (DB difference) – is not a carefree variable. First, its validity as an indicator of psychogenic mortality effects depends on two critical assumptions: (i) that temporal proximity to birthdays has emotional and behavioral impacts, and (ii) that these effects, in turn, influence health. If these assumptions hold, they lend themselves to proto-mechanistic explanations/predictions. For example, if affective-cognitive anticipation has favorable effects (by being emotionally positive, inducing careful behavior, or by some such mechanism), then mortality would be expected to decrease as the birthday approaches. In this case, after the anniversary and the dissipation of protective anticipation, mortality should increase. Conversely, if the effect is anti-protective (which could apply at least to some individuals or to some causes of death), the opposite pattern should emerge, with increased mortality before and decreased mortality after the birthday. Second, any effects on mortality are likely minimal, which complicates detection and analysis, especially in non-experimental data. Third, administrative errors can, in principle, introduce systematic bias. Missing dates of birth might be imputed by reusing death dates or convenient dates like the first or fifteenth of the month, potentially inflating reported deaths on birthdays (Abel & Kruger, 2006). Fourthly, even though the DB interval elegantly circumvents the methodological complications arising from the fact that psychological factors and health outcomes have a plethora of common biological causes, this does not imply that there can be no confounding at all going on. The slightly subtle two-tier statistical mechanism by which age can indeed be a confounder has been explained by Roger (1977) through two competing processes in population mortality: the hazard of adults dying increasing with age, and the size of older age cohorts relatively rapidly decreasing in time. This is plausible when acknowledging that, after a fixed point in time (any arbitrary day zero, which can also be the birthday), the survival function decreases daily in accordance with an exponentially increasing hazard rate by an annual factor of maybe 1.01 (one percent). The number of survivors on any subsequent day is the product of the survival function and the initial population, and the number of deaths per day is the product of the survivors and the hazard rate on that day. In an older age group with a large baseline hazard h0 (and a relatively small initial population), the number of daily deaths decreases despite the increasing hazard rate, because the rapid decline in the number of survivors (i.e., the shrinking of the risk set) outweighs the increase in the hazard rate as soon as, with the above example, h0 > ln(1+(1.01-1)/365). As a consequence, when mortality is analyzed with the birthday in the center of the individual annual cycle, a mortality deficit before and an excess after the birthday can arise solely due to the combination of the two stochastic processes. Based on the monthly UK mortality data of 1972 that were analyzed by Alderson (1975), Roger stated that, in each of the eleven months around the month of birthday, up to three percent of the observed mortality could be due to this statistical artifact. Adjusting the expected numbers accordingly, Roger suggested that in individuals aged over 75, any observed alleged birthday effects – where mortality seemingly decreases before and increases after the last birthday – could largely be explained away.\n\n\nMethods\n\nThe source data included all deaths in the adult Swiss resident population as recorded by the Federal Statistical Office (FSO; cf. Blohm et al., 2022), for which the cause of death (COD) coding had been completed at the time of analysis (years of death up to 2022) and from which the cases with imputed dates of birth or death could be excluded based on a specific flag (as of 1978). From the period 1987 to 2022, leap years (1988, 1992, 1996, 2000, 2004, 2008, 2012, 2016, 2020) as well as cases born on a 29 February were excluded. Analyzed variables were sex, completed age (years) at time of death, day of month and month (but not year) of birth and of death, day of the week of death (DWD; 1, Sunday, to 7, Saturday), marital status (dichotomous), registered religious affiliation (dichotomous), and underlying cause of death. The latter is the disease or condition that was present at the beginning of the causal chain that led to death, as provided on the death certificate by the reporting physician. Causes of death had been coded at the FSO according to the World Health Organization’s International Classification of Diseases, 8th Revision (1969 to 1994) and 10th Revision (as of 1995). The underlying causes of death were combined into the 13 major COD categories infectious diseases (ICD-10 codes A00.0-B99.9, ICD-8 codes 000-136, malignant neoplasms (C00.0-C97.9, D00.0-D48.9, 140-209), mental and behavioral disorders (F01.0-F99.9, 290-315), dementia (F01.0, F03, F05.1, G30.0-G30.9, 290, 302), diseases of the circulatory system (I00.0-I99.9, 390-444.1, 444.3-458, 382.4), diseases of the respiratory system (J00.0-J98.9, 460-519), diabetes mellitus (E10.0-E14.9, 250), diseases of the genitourinary system (N00.0-N99.9, 580-629, 792), accidents (V01.0-X59.9, 800-929), transport accidents (V01.0-V89.9, Y85.0-Y86.9, 800-845), COVID-19 (U07.1, U07.2, U10.9), suicides (X60.0-X84.9, 950-959), and assisted suicides (X61.8). In Switzerland, dementia as well as mental and behavioral disorders were coded as underlying (rather than just concomitant) causes of death as of 1995, the supplementary code for assisted suicide X61.8 was used as of 2009.\n\nIndices (1 to 365) of day of birth (DOB) and death (DOD) were calculated from recorded dates, and from these the DOD-DOB difference. To calculate the days between death and birth (DB difference) within a range of -182 days before to +182 days after the birthday, differences with an absolute value up to 182 days were retained. For differences greater than 182 days, the value was adjusted by subtracting 365 days and, for differences less than -182 days, 365 days were added. The observed numbers of deaths that constituted the joint distribution of DOD, DB difference, and DWD (but disregarding age-group to avoid zero inflation), were modeled by a Poisson count model with a deviance-based overdispersion parameter to adjust the standard errors (SE) of the parameter estimates. The DB difference indicator, the primary predictor, had five birthday-related categories (“B periods”) centered around the birthday, signifying periods of 21, 7, or 0 days away from it, as specified in the square brackets: B[-28,-8], B[-7,-1], B[0], B[1,7], B[8,28]. The reference (control) period C[-182,-29;29,182] covered the remaining 308 calendar days for which\|DB difference\|> 28. Covariates of the linear predictor of the model were two sine and two cosine functions (with arguments 2πDOD/365 and 4πDOD/365) to account for seasonal mortality variation, and categorical DWD, with Sunday the reference category. Analyses were conducted for the full analysis set as well as stratified by the demographic characteristics sex (male, female), age group (18-64, 65-74, 75-84, 85-94, 95-104 years of age), marital status (binary), registered religious affiliation (binary), as well as by period of death (binary, indicating prior to vs. as of 1995) and by the 13 major diagnostic groups (COD categories).\n\nTo facilitate comparing mortality across different demographic and diagnostic groups, as well as with published results, the observed and estimated deaths per day during each of the five B periods were converted into daily mortality rates (MR) per 100,000 deaths per year. B period MRs were deemed, exploratorily, inconsistent with random variation if their 95% confidence intervals did not encompass the MR of the reference category C[-182,-29;29,182]. To obtain B period effects relative to the C period, mortality rate ratios (MRR) were calculated by dividing the MRs of the B periods by the MR of the reference category, the normal approximation 95% confidence intervals of the ratio based on ln(MRR) with standard error (delta method) √{[SE(MRB)/MRB]2+ [SE(MRC)/MRC]2}.\n\nTo explore mortality rate patterns across all B periods, agglomerative hierarchical average linkage cluster analyses were conducted separately for age groups and COD categories. To prevent cluster formation from being dominated by proportionate mortality differences among the subsets (age groups and COD categories, respectively), predicted mortality rates were calculated based on the number of cases in each subset rather than in the full analysis set. The homogenized rates were then rescaled, separately for each subset, by subtracting the reference category level, dividing each difference by its standard error (obtained through the delta method), and finally applying max-abs normalization to obtain a maximum value of 1.0 across all B periods.\n\nFor further insights into temporal relationships, clusters containing multiple subsets were resampled by drawing 100,000 bootstrap samples from their cases, each with the same size as the parent sample. For each sample, a Poisson regression model with predictors for season and day of the week was used to calculate DB difference-specific residuals between observed counts and those expected based on the model with no B period predictor. Means as well as 2.5 and 97.5 percentiles were obtained from the ensemble of residuals, with nonparametric locally weighted scatterplot smoothing (LOESS) applied to visualize the DB difference trajectories of resampling-based birthday period effects and their approximate 95% confidence intervals, using a smoothing parameter of 0.1 to balance overfitting and underfitting.\n\nTo avoid zero-inflation, age was not included in the adjustment set of the Poisson regression model, as even the addition of just four age group indicators would have increased the number of combinations of predictor variable levels from just over 0.93 million to more than 3.7 million – far exceeding the available cases even in the full analysis set. To quantify the maximum possible bias in MRRs due to age-related increases in mortality in younger and decreases in older people after their last birthday (as laid out by Roger, 1977), the MRRs were divided by a bias factor (Sjölander & VanderWeele, 2021; Ding & VanderWeele, 2016). This bias correction for unmeasured or uncontrolled confounders was conducted for each demographic and diagnostic group (cf. Tables 3 and 5). With X = {B=b,C} indicating the B (birthday) or C (reference) period, and A = a one of the five age groups, the sensitivity parameter RRBXA = maxa {P(A=a\|X=B)/P(A=a\|X=C))} quantifies the most extreme imbalance of B and C cases across the age groups. The sensitivity parameter RRBAD=maxx {maxa∑(XB=x,A=a)/mina∑(XB=x,A=a)} quantifies the importance of age group for mortality. Bias factor FB=RRBXA⋅RRBAD/(RRBXA+RRBAD-1) adjusts the MRR of the corresponding B period such that MRRBtrue is ≥ MRRB/FB for MRRB ≥ 1, or ≤ MRRB⋅FB for MRRB<1, with the adjustment also applicable to the confidence interval limits of the mortality rate ratios. The method assumes maximal confounding by age under the extreme scenario where all cases dying during a specific B period would be in the oldest age group (or, equivalently, none from the youngest age group would die in that period).\n\nIn addition to this conceptual (theoretical) sensitivity analysis, analytical (empirical) assessments of the potential of the post-birthday death-count gradient bias (“ageing bias”) were undertaken, overall as well as for all demographic and diagnostic groups. For one, the main analysis was repeated by adjusting for age group instead of for season and day of the week. For another, the potential magnitude of an ageing bias was quantified by linearly regressing the number of daily deaths during the C period (to minimize B period effects) on the number of days since the last birthday (ranging from 0 to 364). By dividing the number predicted at the midpoint of each B period by the number predicted at the midpoint of the C period, the resulting MRRs estimate the size of B period effects that are exclusively due to ageing after the last birthday.\n\nTo assess whether births and deaths on the first and fifteenth day of each month were overrepresented, the observed absolute and relative frequencies on days 1 to 28 (which every month entails) were compared to the levels expected under the assumption of no heaping. For this, Bonferroni-corrected 95% Poisson confidence intervals were used with a multiplicity-adjusted local alpha level 1-exp[ln(0.95)/28] ≈ 0.05/28. This analysis was also carried out for the number of births and deaths recorded on the same day. To evaluate the potential bias that heaping of birth and death dates might contribute to the observed birthday excess mortality, an additional sensitivity analysis was performed by excluding cases born on days affected by heaping.\n\nAll analyses were conducted with SAS Enterprise Guide 8.3.\n\n\nResults\n\nOver the analysis period from 1987 to 2022, the number of annual deaths among adult residents of Switzerland rose from 58,581 in 1987 to 67,127 in 2019. This increase is primarily attributable to the growth of the resident population, which surged from 6.5 million in 1987 to 8.5 million by 2019 (FSO, 2023). In 2022, the number of deaths was 73,591, but included 4,114 cases with COVID-19 as the underlying cause of death (Blohm & Weitkunat, 2023). The full analysis set covered 27 non-leap years and contained a total of 1,702,865 deaths. The majority of deaths occurred in 1995 or later (78.5%). Of the deceased, 84 percent were 65+ years of age at time of death (16.0%, 29.9%, 32.5%, and 5.6% were 65-74, 75-84, 85-94, and 95-104 years of age at time of death, respectively). The average age of death was 77.7 years. The proportions of men, married, and religiously affiliated deceased were 49.0, 39.9, and 87.5 percent, respectively. The proportion of deaths occurring on a Sunday was 13.9 percent, compared to 14.3 percent occurring on average on a weekday. Over the analysis period, the day on which most deaths occurred was 05 January and the day with the lowest number of deaths was 03 August, the ratio of the corresponding numbers of deaths indicating an average winter excess mortality of 36 percent.\n\nIn the full analysis set, 5,040 deaths (0.296%) occurred on the deceased’s birthday ( Table 1), exceeding the expected number of 4,665 (0.274%), which assumes an equal distribution of cases across 365 non-leap year days. The difference between the two counts – 375 cases – suggests that the above-expected (excess) mortality specifically on the birthday is small, corresponding to a rate of 22 deaths per 100,000 deaths per year. This modest magnitude makes causal attribution to the birthday susceptible to bias and challenging to substantiate.\n\nDeaths in adult Swiss residents across the 27 non-leap years from 1969-2022. Stratification by age, sex, and death on birthday vs. other day.\n\nFigure 1 shows the daily number of deaths over 365 days accumulated across the analysis period, along with predictions based on the Poisson regression model that included the birthday period as the primary predictor, with season and day of the week as covariates. As can be seen from the upper part, the Poisson model (smooth line) fits the data well, the typical northern-hemispheric seasonal variation in mortality visible, and mortality substantially associated with low ambient temperatures (as first described as seasonal “excess mortality” by William Farr in 1847 in England). Notably, as has been reported previously by others (e.g., Brenn & Ytterstad, 2003), the seasonality did not obtain equally with all causes of death or in all demographic groups, in particular not with cancer and in people dying before their mid- to late 60s. According to the likelihood ratio test comparing the full with a reduced model, the B period term contributed substantially to the model fit (p<10−5).\n\nThe jagged lines represent the observed daily deaths of the full analysis set cumulated over the 27 non-leap years from 1987 to 2022, the smooth lines depicting the model-based predictions. In the upper panel, the abscissa at the top of the graph represents the passage of 365 calendar days. In the lower panel, for the same data, the abscissa at the bottom represents the day of death minus the day of birth.\n\nIn the lower part of Figure 1, where the same data is shown over the interval between the day of death and the day of birth, a peak is discernible in the middle of the graph when both days coincide. Across all years, an average of 130,518 deaths occurred over a period of, for comparison, 28 days during the reference period, compared to 130,724 deaths in the 28 days before and 131,407 deaths in the 28 days after the birthday. Centering the latter two counts by subtracting the reference period count obtains 206 and 889 excess deaths in the 28 days before and after the birthday, respectively. With 18.8 percent of the excess cases observed in the four weeks before the birthday and 81.2 percent in the four weeks after, the likelihood of a symmetrical distribution of deaths around the birthday is less than 0.01 percent, strongly pointing towards higher mortality after the birthday than before in the full analysis set. Figure 2 shows the period from 30 days before to 30 days after the birthday, depicting both the observed and the predicted rates as daily mortality rates per 100,000 deaths per year.\n\nThe smooth line segments (and 95% confidence intervals) indicate the predicted daily mortality, the jagged line the observed daily rates. For periods B[-28,-8], B[-7,-1], B[1,7], and B[8,28], but not for period B[0] (the birthday itself ), the confidence interval around the predicted rates covers the mortality rate predicted for the reference period C[-182,-29;29,182], the level of which is visible for 29 and 30 days before and after the birthday.\n\nWhen a comparison-wise false-positive error level of five percent is adopted to explore the 95% confidence intervals for potential effects, Figure 2 suggests that the impact of the birthday itself on the model likelihood exceeds what would be anticipated from random variation. Conversely, while the predicted daily mortality rates for the periods preceding and following the birthday suggest a pattern, the associated 95% confidence intervals cover the predicted reference period level.\n\nImportantly, B period effects which might exist for different demographic or diagnostic groups could dilute or even cancel each other in the full analysis set. Table 2 thus provides an overview of the demographic subgroups for which regression analyses were conducted. The analysis sets are disjunctive within each demographic characteristic (sex, marital status, registered religious affiliation, year of death, and, when disregarding the 65+ years group, age) but not between them. For age, five disjunct strata are reported in Table 2: 18-64 years and four 10-year age groups covering the range between 65 and 104 (with the largely redundant 65+ group included mainly to facilitate comparisons with published mortality statistics that often dichotomize age at 65 years).\n\nThe data reveal several notable and straightforward patterns. For example, the largest winter excess mortalities occurred in the 85-94 (52.3%) and 94-104 (88.9%), the lowest in the 18-64 (24.0%) years of age groups, suggesting an increasing vulnerability to low ambient temperatures with age; the proportion of cases with no registered religious affiliation substantially increased from 1995 onwards compared to earlier years, indicating a trend toward secularization; and the proportion of deaths occurring from 1995 onwards increases with the age of the deceased, indicating a secular lengthening of life expectancy.\n\nTo address the main topic of comparing mortality rates in the B periods and the reference period C, Table 3 presents the daily mortality rate predictions and mortality rate ratios of the B periods relative to the C period, along with 95% confidence intervals. Likelihood ratio tests indicated only marginal contributions of B periods in women (p=0.07) and in deceased with no registered religious affiliation (p=0.25). Otherwise, with the exception of married deceased (p=0.01), p-values were smaller or much smaller than 0.001.\n\nIn all groups depicted in Table 3, mortality rate ratios larger than one were found for B[0], the confidence intervals covering unity only in the no registered religious affiliation and the 75-84 years of age strata. A small pre-birthday MRR increase of about two percent was found in deceased with no registered religious affiliation for B[-28,-8]. Small post-birthday mortality rate ratio increases (of about one to two percent each) were found in male and in unmarried deceased for B[8,28] and in married deceased for B[1,7]. More pronounced effects were found regarding age. Based on the 95% confidence intervals’ lack of covering unity, increased MRRs were observed for B[-28,-8] and decreased MRRs for B[8,28] in the 18-84 years of age range. Additionally, in the 18-64 years stratum, the MRRs for B[-7,-1] were increased and those for B[1,7] decreased. Conversely, in the two oldest age strata of the 85-104 years range, the reverse pattern was found, with MRRs decreased pre-birthday during B[-7,-1] and B[-28,-8], and increased post-birthday during B[1,7] and B[8,28]. For B[-7,-1], B[1,7], and B[8,28], this pattern is also visible in the broader 65+ years of age group.\n\nDisregarding the full analysis set and, to avoid overlap, the 65+ years of age group, Table 3 contains 95% confidence intervals for 65 MRRs, of which 31 do not cover the null-effect mortality ratio of one, where, by chance, only slightly more than three (3.25) would be expected. When also disregarding the B[0] period, 20 out of 52 intervals do not cover unity, while only between two and three (2.6) would be expected.\n\nFigure 3 depicts the observed and predicted daily mortality rates over the five B periods for the strata of sex, marital status, year of death, registered religious affiliation, and for the age groups 18-64, 65-74, 75-84, 86-94, and 95-104 years. It can be seen that (as with the corresponding MRRs contained in Table 3) the estimates are more precise for longer than for shorter B periods, the underlying numbers of cases being smallest for B[0], and consequently the confidence intervals the widest. In the year of death strata (<1995 and 1995+), in men and women, in married and unmarried deceased, as well as in deceased with a registered religious affiliation, B[0] excess mortality is visible, which in married deceased also obtains in B[1,7] and in unmarried deceased and in men in B[8,28]. In deceased with no registered religious affiliation, no association with predicted mortality is discernible from the week before birthday onwards, including the birthday itself, with observed mortality rates exhibiting considerable variability in this group.\n\nThe smooth lines (with 95% CIs) show predicted rates for B[-28,-8], B[-7,-1], B[0], B[1,7], and B[8,28], as indicated by dotted vertical lines; the jagged lines show observed rates. Reference period rates appear at the edges (29 and 30 days before and after birthday), with axes omitted to focus on the form of the predicted mortality trajectories.\n\nWith regard to the five disjunct age groups, a two-cluster solution of the normalized B period mortality rate predictions confirmed the MR patterns of Table 3, with age groups 18-64, 65-74, and 75-84 years forming one cluster and age groups 85-92 and 95-104 another. In the three younger age groups, distinct pre-birthday and birthday excess MRs are visible, succeeded by a reduction in MRs in the weeks after the birthday. Conversely, the two older age groups show a distinct pattern of reduced MRs before birthdays, followed by increased rates on birthdays and in the weeks that follow.\n\nOther than with demographic strata, all cause of death categories depicted in Table 4 are disjunctive. Of the 1,702,865 deceased of the full analysis set, 1,534,761 died of an underlying cause of death that falls into one of the 13 categories of major causes of death, rendering the combined proportionate mortality 90.1 percent. The proportionate mortalities of cardiovascular diseases and of cancer were 25.4 and 35.5 percent, respectively, the two diagnostic categories together accounting for 60.9 percent of all deaths. Other causes with proportionate mortalities of at least one in twenty decedents were respiratory diseases (6.4%), dementia (5.2%), and mental and behavioral disorders (5.0%). The smallest underlying cause of death category was COVID-19, with 8,125 cases contained in the analysis set, corresponding to a proportionate mortality of 0.5 percent.\n\nWinter excess mortality, while present in all major cause of death categories, was rather more heterogeneous than across the demographic groups. While for cancer the excess was 22.4 percent, it was 47.5-fold for COVID-19. For assisted suicide, the winter excess mortality was 47-fold compared to 1.7-fold for suicide, the former peaking in November and the latter in July. There were some similarities between suicides and assisted suicides, both having occurred least frequently on 25 December and rarely on Sundays, in particular (and quite possibly for administrative reasons) assisted suicide. The dissimilarities are, however, more impressive (even if assisted suicide has been coded separately only as of 1995): Suicides were, compared to assisted suicides, younger (59.0 vs. 77.3 years), more often religiously affiliated (72.8% vs. 58.6%), married (40.6% vs. 36.9%), and male (64.5% vs. 42.1%).\n\nShort of confirmatory testing, exploratory likelihood ratio p-values provide some indication of the level of statistical support for B period mortality rate effects (although the evidentiary significance of p-values is compromised by their notorious dependence on sample size, with statistical power increasing and p-values decreasing as group size increases). The likelihood ratio test p-value of the B period indicator was < 0.05 for mental and behavioral causes, < 10−2 for respiratory diseases and for assisted suicide, and < 10−3 for suicide and for cardiovascular diseases. It exceeded the 0.05 threshold (a prevalent convention in dichotomous confirmatory assessments) for accidents (0.08), transport accidents (0.11), diabetes (0.13), cancer (0.20), dementia (0.21), infections (0.23), genitourinary diseases (0.39), and COVID-19 (0.52). The rather large likelihood of the B periods not being substantially predictive of mortality in COVID-19 is consistent with the low overall proportionate mortality of 0.5 percent (8,125 cases) in this group. Similarly, the proportionate mortality was at one percent or below in infections (1.0%, 16,837 cases) and transport accidents (0.8%, 12,812 cases), and between one and barely above two percent in genitourinary diseases (1.3%, 21,911 cases) and diabetes (2.1%, 35,839 cases). The only two diagnoses with both proportionate mortalities above these levels and relatively low B period predictiveness likelihoods of about 20 percent were dementia (5.2%, 88,659 cases) and cancer (25.4%, 432,703 cases).\n\nTable 5 presents the predicted daily mortality rates and mortality rate ratios of the B periods relative to the C period, along with 95% confidence intervals, for each of the 13 COD categories. As can be seen, for dementia and cancer, their low B period predictiveness is consistent with the B period-specific mortality rate ratio findings, all 95% confidence intervals covering the no-effect level of one for cancer, and for dementia only one excess mortality (during B[8,28] of about 3%) inconsistent with no effect.\n\nRegarding birthday and post-birthday period MRRs, mortality increases exclusively during B[8,28], as seen with dementia, were also found for infections (of about 9%), mental and behavioral diseases (4%), and respiratory diseases (3%). Excess mortality only during B[1,7] was found in COVID-19 (11%) and only on B[0] in circulatory diseases (11%) as well as in suicides (46%). Increases were found for both B[0] and B[1,7] in transport accidents (19% and 7%), for both B[0] and B[8,28] in assisted suicide (75% and 5%), and for both B[1,7] and B[8,28] in accidents (5% and 2%). Lower than expected mortality on or after the birthday was found on B[0] as well as during B[8,28] in genitourinary diseases (-19% and -3%).\n\nPre-birthday , excess mortality was found during B[-28,-8] for respiratory diseases (3%), suicide (3%), assisted suicide (8%), and COVID-19 (7%), for the latter also during B[-7,-1] (7%). Lower than expected mortality only during B[-7,-1] was found in respiratory diseases (-4%) and in genitourinary diseases (-5%), only during B[-28,-8] in accidents (-3%), and during both pre-birthday periods in diabetes (between -5% to -6%) and transport accidents (-8%).\n\nOf the 65 MRR 95% confidence intervals depicted in Table 5, 27 indicate that the birthday period mortality ratios deviate systematically from what would be expected from the reference period rates. This number is more than eight times higher than the 3.25 false positives expected by chance alone. Disregarding the B[0] period, 22 out of 52 MRRs show mortality rate deviations from the reference period that are not consistent with merely random variation, compared to the 2.6 false positives expected by chance. Of the twelve pre-birthday intervals that deviate from expectations, seven indicate a decreased mortality rate, while increased pre-birthday mortality was only found in respiratory diseases, COVID-19, accidents, and transport accidents. In contrast, with the exception of genitourinary diseases, all of the 15 marked birthday and post-birthday period effects indicate increased mortality.\n\nFigure 4 illustrates the observed and predicted daily mortality rates across the five B periods for each of the 13 COD categories. The six-cluster solution resulted in three clusters combining multiple COD categories, alongside three distinct diagnostic categories (respiratory diseases, genitourinary diseases, COVID-19) that were not combined with others. The three multi-category clusters are depicted in the three columns above the bottom row that shows the three solitaire COD categories. It is important to recall that the clustering was solely based on the phenotypic patterns of normalized B period mortality rates, with no consideration of underlying mechanistic similarities or differences whatsoever.\n\nDaily mortality rates (smooth lines) and their 95% confidence intervals across the five B periods, as predicted by the 13 major cause of death category-specific Poisson models. The observed daily death rates are shown in the background (jagged curves). In each panel, the central vertical dotted line indicates the birthday, the vertical lines to its left and right ±7 and ±28 days before and after the birthday. The six-cluster solution of the normalized rates yielded three clusters with multiple major cause of death categories combined (three columns in the upper part). These contain (i) circulatory diseases, cancers, suicides, and assisted suicides, (ii) infections, mental and behavioral diseases, and dementia, and (iii) diabetes, accidents, and transport accidents. The proportionate mortality (PM) of these clusters is 64.1, 11.2, and 6.7 percent, respectively. Respiratory diseases (6.4% PM), genitourinary diseases (1.3% PM), and COVID-19 (0.5% PM) were retained as stand-alone causes of death categories (bottom row of panels).\n\nThe left-hand column of Figure 4 depicts the cluster comprising circulatory diseases, cancer, suicide, and assisted suicide, combining two somatic and two behavioral COD categories. An increased mortality rate is visible for all four causes at B[0], with cancer showing the least pronounced peak. The two behavioral categories also exhibit a pre-birthday mortality increase during B[-28,-8], and assisted suicide additionally shows a post-birthday increase during B[8,28].\n\nThe middle column shows the cluster that comprises infections, mental and behavioral diseases, and dementia. In neither of these COD categories a B[0]-specific increased (or decreased) mortality rate is discernible, whereas all three groups show a B[8,28] post-birthday mortality rate increase.\n\nThe right-hand column shows the cluster that includes diabetes, accidents, and transport accidents. The three constituent COD categories exhibit reduced pre-birthday mortality, with patterns that, when mirrored across the midpoint of the reference-period risk level, approximately reflect those observed in the middle column cluster.\n\nThe B period mortality patterns for respiratory and for genitourinary diseases are consistent with a short-term displacement effect that precedes the birthday by a few days. In contrast, for COVID-19, elevated mortality rates were found throughout the birthday period, with only a marginal additional effect associated with the birthday itself.\n\nResampling was done on the three clusters that comprise multiple major causes of death categories, the method not relying on parametric or paradigmatic analyses (as in regression models with pre-specified birthday periods). The resulting mortality variations in proximity to the birthday are contained in Figure 5.\n\nThe jagged curves in the background represent the daily bootstrap averages, accompanied by 95% confidence intervals. Resampling of 100,000 cases each was conducted separately for the three multi-category COD clusters shown in Figure 4. Left panel: Circulatory diseases, cancers, suicides, and assisted suicides, characterized by increased mortality on the birthday itself. Middle panel: Infections, mental and behavioral diseases, and dementia, characterized mainly by increased mortality in the weeks following the birthday. Right panel: Diabetes, accidents, and transport accidents, characterized mainly by reduced pre-birthday mortality. The smooth LOESS curves and their 95% prediction bands in the foreground are schematic representations of the forms of the cluster-specific trajectories over the birthday period rather than isomorphic depictions of the distribution of actual average mortality deviations from expected levels. This is due to the low-pass filter properties of the method that attenuates short-term variations and induces temporal smearing. This is particularly visible in the left panel, where the effect on the birthday itself, while visible, is clearly less peaked and of smaller amplitude than the average of the observed minus expected differences on B[0] (293 cases). It is noticeable that the variability of the daily averages over the birthday period is substantially larger for the COD categories depicted in the left panel, where mortality effects are mostly restricted to the birthday itself.\n\nAs with the preceding model-based analyses, the resampling results point to systematic birthday-related mortality variations, with the pre- and post-birthday mortality patterns differing both temporally and in magnitude from each other and from the birthday itself. Since resampling was done on combinations of COD categories both related and unrelated to direct behavioral causes, the mechanisms underlying the observed patterns are almost certainly more complex than the three major trajectories depicted in Figure 5 might suggest.\n\nTo explore the potential confounding of the observed associations between B period and mortality by age-related biases, the left panel of Figure 6 depicts the fundamental age-association of mortality, even if with the present data the phenomenon occurs ten age years later than what Roger had found in 1972 UK mortality data, the shift obviously reflecting a substantial secular increase in life expectancy over the last five decades. In the present data, up to the age of 85, the number of deaths increases. From age 86 onward, the trend reverses with a sharp decrease in the number of deaths for each subsequent year of age, reflecting the rapid shrinking of the risk sets of older age cohorts, as opposed to younger ones. As a consequence of this biphasic age-dependent mortality pattern, even in the absence of any other effect, the daily numbers of deaths must be expected to decrease in the age groups above 85 years over the 364 days following the last birthday. As is shown in the age group-specific scatter plots and regression lines in the right panel of Figure 6, this is indeed the case, the older age groups displaying a declining and the younger age groups an upward trend. This raises the question as to whether the observed birthday effects are entirely or partially age-related artifacts, including of what can be referred to as “ageing-gradient bias”.\n\nLeft panel: Number of deaths by year of age, showing 65,752 deaths (3.86%) at a maximum of 86 years. Right panel: Age group-specific scatter plot of the number of daily deaths by days after the last birthday, with the linear regression lines ŷt = b0+b1t predicting the number of deaths per age group on day t after the last birthday. The total predicted number of deaths on day t is obtained by either summing the age group-specific predicted numbers for that day, by computing daily predicted numbers based on aggregated age group-specific regression parameters, or by regressing the total daily death count against the days elapsed since the final birthday without age group stratification. In the 85-94 (top) and the 95-104 years of age groups (bottom), the numbers of deaths per day decline after the last birthday, while they increase in the three younger age groups, the two groups 18-64 and 65-74 largely overlapping in the range of 500 to 1000 deaths per day. The total daily number of deaths ŷt = 4,648+0.0622t is rather stable over time, which aligns with Figure 1. As can be calculated from the overall regression equation, the mortality ratios of the five B periods are all 1, indicating that the age composition of the full analysis set and the combined age group-specific mortality age-gradients did not induce any B period effect. To introduce a one percent MRR increase in the most age gradient-sensitive off-birthday B period B[1,7] (due to its vicinity to t = 0), the slope of the regression equation would have to be multiplied by a factor of 4.23. Preserving the linear additivity of the age group-specific estimates, this slope factor would imply that in the 95-104 years of age group the number of daily predicted deaths would be negative for t ≥ 319.\n\nAs is confirmed in Table 6, age is obviously substantially important for mortality, the sensitivity parameter RRAD increasing as mortality disbalance across the age groups increases. The more prominent levels of that parameter are consistent with fundamental relationships: Only 1.1 percent of married deceased were 95 years or older, suggesting that the chances of both spouses reaching such an advanced age were relatively small; prior to 1995, only 2.6 percent of the deceased were in that oldest age group, compared to 6.4 percent of those who died in 1995 or later, reflecting the increased life-expectancy of the population; only 0.3 percent of the transport accidents were 95 years or older, obviously reflecting reduced levels of mobility in that segment; only 1.3 percent of those who died with cancer as the underlying disease and only 1.2 percent of suicides reached an age of 95 or older, while other diagnoses were particularly rare in the 18-64 years of age-group, including dementia (0.7%) and genitourinary diseases (4.6%).\n\nFor each B period, RRBXA is the largest per age group ratio of the proportion of deaths among those dying in period X = B to the proportion among those dying in period X = C. It quantifies the imbalance of deaths between two contrasted X periods (the respective B period and the C period) across the five age groups. RRBAD is the largest ratio across periods X, where each ratio is calculated as the highest proportion of deaths to the lowest proportion among the five age groups within a specific period X. It quantifies the importance of the uncontrolled confounder age on the outcome.\n\nIn contrast to the anticipated, plausible, and largely profound levels of RRAD, the importance of whether or not death occurred in a B period on the age of death, as quantified by the sensitivity parameter RRXA, was generally small, in line with the flagrant implausibility of death occurring in a specific B period having any relevance to the age of death. As a consequence, half of the resulting bias factors were below 1.19 and three quarters below 1.19, the largest factor being 1.75 for cancer and B[0].\n\nThe mortality effects of birthday periods being substantially smaller than those of age per se, the conceptual sensitivity analysis focusing on confounding away from the null confirmed that all but the B[0] effect in assisted suicide could, in theory, be adjusted away by age. The two serious caveats of this assessment are, however, that it only holds for the most extreme confounding scenarios and that it entirely neglects false-negative findings by confounding towards the null. Applying the bias factors to the estimated MRRs of the 13 diagnostic categories indeed yielded the following numbers of de novo effects below/above expectations: 7/3 B[-28,-8], 5/8 B[-7,-1], 3/3 B[0], 7/5 B[1,7], and 9/4 B[8,28]. The large number of 54 purportedly overlooked effects in this evaluation underscores the sensitivity analysis as too extreme. Conversely, it is unlikely that most of the observed effects, while they could theoretically be adjusted away by age, are indeed false positives and would indeed vanish upon actual adjustment for age.\n\nTo complement the conceptual sensitivity analysis with an empirical assessment, the adjustment set of the original analysis (season and day of week) was replaced by age group, with 17-64 years as the reference category. Age was, as expected, generally associated with mortality in all demographic and diagnostic analysis sets. In the full analysis set, mortality rate ratios quantifying the effects of age group on the expected counts of deaths were found to be elevated for the 75-84 age group (1.87; 95%CI: 1.857, 1.880) and the 85-94 age group (2.03; 2.016, 2.041), but not for the 65-74 age group (1.00; 0.991, 1.005) or the 95+ age group (0.35; 0.349, 0.356). However, none of the original B period ratio estimates changed up to the fourth decimal place upon adjustment by age group, neither in the full analysis set nor in any of the demographic and diagnostic groups. The robustness of MRRs across adjustment sets indicates that, empirically, B period effects were independent of age. This is reflected in an extremely small value of Cramer’s V, quantifying the association between B period and age group, of 0.0094 in the full analysis set. Even in the diagnostic groups with the youngest and oldest mean age of the deceased there was no indication of an association between B period and age, with Cramer’s V values 0.0149 and 0.0153, respectively, for transport accidents and dementia.\n\nThe possible magnitudes of B period effects, would they solely depend on ageing, were quantified by simple linear regression analyses conducted for all demographic groups (except for age groups) and diagnostic groups. The coefficients were determined by regressing the daily number of deaths against days after the last birthday. From the coefficients and the respective time periods, the ageing-gradient bias was quantified by calculating purely ageing-based B period MRR estimates (which must necessarily be most pronounced for B[0], followed by the two seven-day periods directly before and after the birthday). With an accuracy of two decimal places, in all demographic groups, with the exception of women, the B[0] MRRs (and thus also those of the other four B periods) were all 1.00. In women, the value was slightly below 0.995. For most diagnostic groups, the B[0] MRRs were less than one percent (cancer, circulatory diseases, respiratory diseases, genitourinary diseases, accidents, transport accidents) or less than two percent (infections, mental and behavioral diseases, dementia, diabetes) away from unity. Most of the ageing-based MRRs found in the diagnostic groups could directionally not have contributed to the observed effects, two examples being mental and behavioral diseases and dementia, where the observed B[8,28] MRRs were above one while the age-based MRRs were below one. Three purely ageing-based effect estimates which were in the same direction as the observed effects contained in Table 5 were the B[8,28] effect for infections of 1.015 and the B[-28,-8] and B[-7,-1] effects for diabetes of 0.990 and 0.989, respectively. While these could slightly reduce the observed effects, they were clearly too small to eliminate them. Likewise, the purely ageing-based estimated B[0] effects for suicides and assisted suicides of 1.012 and 1.030, respectively, while slightly biasing the observed effects away from the null, are far too small to explain away the MRRs of 1.46 (suicide) and 1.75 (assisted suicide). The much smaller observed assisted suicide B[8,28] MRR of 1.05 is, according to its ageing-based counterpart of 1.027, possibly due to ageing bias. The only substantial ageing gradient effect was found for the smallest diagnostic group, COVID-19, where the two observed pre-birthday excess MRRs of 1.07 can be explained by the ageing-based B[-28,-8] and B[-7,-1] estimates of 1.073 and 1.079, respectively. For the observed COVID-19 B[1,7] post-birthday excess MRR this is naturally not the case, as the ageing-based MRR for that period was substantially less than one (0.921).\n\nThe observed numbers of births and deaths on the first 28 days of any month were assessed for heaping by comparing them (i) to the expected daily count (both of births and of deaths) of 55,984.6 (the total number of cases, 1,702,865, multiplied by 12 months and divided by 365 days), (ii) to the expected relative daily frequency (1/28), and (iii) to the observed mean numbers for days 1 to 28 of 56,068 [adjusted 95% CI: 55,289; 56,847] for births and 56,035 [95% CI: 55,256; 56,813] for deaths. Of the 56 assessed counts, only that of 59,083 births on day 1 exceeded the upper confidence interval limit of the corresponding mean, as well the expected frequency by 5.5 percent. The relative frequency of births on day 1 was 3.8 percent of all births of the first 28 days of any month, compared to the expected daily frequency of about 3.6 percent.\n\nAmong the 52,064 cases where the day of birth and the day of death in any month coincided (55,663 overall, without restriction to the first 28 days of any month), none of the 28 observed counts exceeded the average daily number of cases of 1,859 [adjusted 95% CI: 1,718; 2,001]. Nominally, the day 1 count of 1,976 cases exceeded the expected value of 1,840.6 ([12/365]2 multiplied by the total number of cases, 1,702,865) by 7.4 percent. As with births alone, the relative frequency of all births and deaths coinciding on one of the first 28 days of any month was 3.8 percent for day 1, where again about 3.6 percent were expected.\n\nStratified by month, the proportion of births and deaths coinciding on day 1 among all such coincidences over the first 28 days ranged from 3.6 to 3.8 percent from February to December, with all multiplicity-adjusted 95% confidence intervals covering the above mean of 3.8 percent for day 1 of any month. In contrast, the corresponding day 1 proportion for January 1 was 4.4 [4.2-4.5] percent. If this indicates that the above-described heaping of births on day 1 of any month is largely a January phenomenon, then, in the present data, about one in 20 children born on December 31 might have been assigned January 1 as their day of birth, which is not entirely implausible. Despite the January 1 counts of cases with day of birth and death coinciding not materially exceeding the expected levels or the average of the first 28 days of any month, an additional sensitivity analysis was conducted by removing all 59,083 cases born on day 1, not only of December but of any month. The B[0] MRs and MRRs and their 95% confidence intervals of the four diagnostic groups clustered together for their pronounced B[0] mortality (circulatory diseases, cancer, suicide, and assisted suicide) remained virtually unchanged. The biggest change was seen in assisted suicide, where the MR remained at 3.1 [2.88-3.33] and the MRR increased from 1.75 to 1.77 [1.64-1.90].\n\n\nDiscussion\n\nThe full analysis set comprised a total of 1,702,865 deaths in the adult Swiss resident population through the 27 non-leap years from 1987 to 2022. The Poisson model, regressing daily mortality counts on categorized days between death and last birthday, adjusting for seasonality and day of week, fitted the data well. Less deaths occurred on Sundays than on weekdays and the typical northern-hemispheric seasonal variation in mortality is clearly visible, except in those dying from cancer and before their mid- to late 60s. 5,040 deaths (0.296%) occurred on the birthday of the deceased, exceeding the 4,665 deaths (0.274%) that would be expected if deaths were evenly distributed across the 365 days centered around the last birthday. The birthday excess mortality of 375 deaths corresponds to a rate of 22 additional deaths per 100,000 deaths per year. Focusing on the birthday, this represents an eight percent increase in deaths on birthdays compared to what would be expected, such that out of approximately 13 individuals who die on their birthday, one death is attributable to the birthday itself.\n\nThere were 206 and 889 excess deaths, respectively, in the 28 days before and after the birthday, indicating substantial asymmetry of deaths around the birthday, with higher mortality after than before. As birthday period-related effects that might exist for different demographic or diagnostic groups could dilute or even cancel each other out in the full analysis set, stratified analyses were conducted. In all demographic strata, with the exception of deceased with no religious affiliation and in the 75-84 years of age group, increased mortality on the birthday itself was found, with excess mortalities about twice as pronounced in men and unmarried persons than in women and in married persons. In age groups 85 years and older, a clearly discernible pre-birthday dip was followed by a post-birthday excess mortality, that pattern being in line with stochastic mechanisms related to age-dependent ageing-related mortality gradients, as proposed by Roger (1977).\n\n1,534,761 cases died of one of 13 categories of underlying causes of death, corresponding with a combined proportionate mortality of 90.1 percent, with cancer and cardiovascular causes accounting for 60.9 percent of all deaths. Increased mortality on the birthday itself was found for cardiovascular causes of death, transport accidents, suicides, and assisted suicides. Reduced pre-birthday along with increased post-birthday mortality, in a manner consistent with mortality gradients related to ageing, was only found for accidents and transport accidents, while for diabetes and genitourinary diseases pre-birthday mortality deficits were found but no post-birthday excesses. Reduced mortality was also found for genitourinary diseases on the birthday itself as well as in the weeks after the birthday. Exclusively excess mortality was found for infectious diseases, mental diseases, and dementia in the weeks after the birthday, for suicides before and on the birthday, and before, on, as well as after the birthday for assisted suicides. While respiratory and genitourinary diseases as well as COVID-19 were not combined with other major causes of death categories, three clusters containing multiple cause of death categories were formed; one containing circulatory diseases, cancer, suicides, and assisted suicides, characterized by above-expectation mortality mainly on the birthday itself; one containing infectious diseases, mental and behavioral diseases, and dementia, characterized mainly by post-birthday excess mortality; and one containing diabetes, accidents, and transport accidents, characterized mainly by pre-birthday mortality deficits and, in the case of accidents and transport accidents, also by a short-term mortality increase commencing with the birthday.\n\nGiven that systematic administrative errors, as well as stochastic mechanisms, could in principle introduce artificial mortality patterns in sync with the birthday period, a thorough investigation into these possibilities was conducted. Our investigations into the registration procedures ruled out the possibility that the \"birthday effects\" (rather than a single effect only on the birthday itself ) observed in our data could be due to administrative errors; civil registration records in Switzerland are established at birth or at the time of registration in the country, not at or after death. When a person dies, the relevant parts of the civil registration records are automatically transferred to our mortality database, independent of and prior to the information subsequently provided with the death certificate. Furthermore, the mortality variations observed across strata, including on the birthday itself, cannot be explained by administrative biases. Why, for instance, would the birthday mortality rate be elevated for cardiovascular diseases but not for genitourinary diseases? With regard to heaping, we found an above-expectation frequency of births occurring on the first day of a month, the effect basically stemming from more than expected births recorded on firsts of January. A sensitivity analysis removing all births on a first of a month did not, however, indicate any material effect on the estimates.\n\nInvestigating the potential bias due to ageing-gradient effects as described by Roger (1977), implying reduced mortality before and increased mortality after the birthday, revealed that while, in particular in the 85-94 and 85-104 age groups, monotonous mortality decline after the birthday did obtain, assessing bias factors to quantifying the maximum degree of systematic distortion of mortality rate ratios were not deemed to support the notion of the estimated effects to be due to confounding by age. This was confirmed by directly adjusting the estimates by age group (instead of by season and day of week), which left the effect estimates unchanged, both overall as well as in the demographic and diagnostic strata. Furthermore, a linear regression analysis estimating the statistical effect of ageing on the daily numbers of deaths demonstrated that our findings could generally not be explained stochastically through ageing-gradient bias. The exceptions to this were the pre-birthday excess mortality observed in COVID-19 and the excess in assisted suicides in the weeks after the birthday, with the latter being of marginal magnitude and the former observed in the smallest cause of death category. These findings demonstrate that the results are robust against the investigated sources of systematic error, which is in line with the results obtained by the main analysis itself: Deficit and excess mortality patterns across birthday-related periods varied strongly across diagnostic groups, with no consistent pattern of stronger effects in periods closer to the birthday compared to those further away, which would be, however, just what aging gradients would induce.\n\nConcerning the question whether the observed effects are artificial or psychogenic, our analyses clearly indicate the latter. This is not to suggest that the estimated quantities are entirely free from some degree of systematic error, but rather that any such error is highly unlikely to result in material bias. Overall, we found compelling evidence that the proximity to a person's birthday is indeed associated with mortality variations. While, overall, mortality tends to be higher in the period of four weeks before to four weeks after the birthday, with a pronounced peak on the birthday itself and mortality after the birthday higher than before, the heterogeneity of this pattern across demographic and diagnostic groups is considerable. With regard to different diagnostic groups, birthday-related mortality patterns are neither limited directionally (i.e., to either exclusively increases or decreases) nor are they confined to the birthday itself. While the effects of suicides and accidents – i.e., directly behavior-related deaths – are quite clear-cut and can be intuitively grasped, the effects related to non-behavioral causes of death are likely just as real, even if mechanistic explanations are lacking.\n\nGiven the variability of birthday effects across diagnostic groups, there is little reason to doubt that multiple underlying psychogenic mechanisms are at play. While some individuals seem to deliberately choose to end their lives on their birthday (and to a lesser degree in the weeks preceding it), the increased mortality risk from cardiovascular diseases during anniversary-related activities involving the celebrant (or the lack of any birthday celebration for that matter) is quite possibly linked to relevant physiological changes occurring on that specific day. Such changes may result from heightened (or reduced) motoric, neural, or hormonal activities triggered by mental, emotional, behavioral, and social factors.\n\nWith infections, mental and behavioral diseases, and dementia only showing increased mortality after the birthday, it’s hard to shake the impression that this may reflect some kind of giving-up response once the anniversary has passed. In contrast, with mortality risks reduced during the pre-birthday period, diabetes, accidents, and transport accidents display birthday-period mortality patterns that are approximate symmetrical inversions of those found in infections, mental and behavioral diseases, and dementia. The mortality deficit in pre-birthday diabetes deaths might reflect individuals willing themselves to holding on just a bit longer. The reduced accident mortality rates during the pre-birthday period might be due to heightened risk-conscious behavior aimed at avoiding jeopardizing the upcoming anniversary. As of the birthday, such precautions might be set aside, or birthday-specific psychosocial risks may manifest, like stress, distraction, lack of sleep, alcohol consumption, or engaging in behaviors that are not practiced at other times, or not with the same intensity. The post-birthday increases in accident mortality rates would fit some kind of carry-over (or even hang-over) effect of anniversary-specific behaviors.\n\nIf these interpretations are roughly accurate, then infections, mental and behavioral diseases, and dementia (clustered as causes of death characterized by post-birthday mortality increases) on the one hand, and diabetes and accidents on the other, might both reflect the same underlying motivation – albeit enacted with different temporal patterns and possibly through separate mechanisms: the desire to see the forthcoming anniversary. For accidents, which are often, partly or entirely, direct consequences of behaviors, this might be achieved by volitionally avoiding risks more than at other times. In contrast, for somatic causes (infections, mental and behavioral diseases, dementia, and diabetes) the mechanisms are presumably mainly psychosomatic, i.e., not mediated behaviorally. As with behaviors, psychosomatic mechanisms likely vary significantly between individuals and causes of death (possibly even among causes of death of the same category), likely involving diverse psychophysiological processes and their combinations. The present results might indicate a pre-birthday resilience (or “hold”) effect in diabetes, and a post-birthday surrender (or “fold”) effect in infections, mental and behavioral diseases, and dementia.\n\nRegarding the three major cause of death categories that were not combined with others during cluster analysis – respiratory diseases, genitourinary diseases, and COVID-19 – it appears that the former two share some similarities with diabetes, albeit only displaying a short-term holding on-effect limited to a few days before the birthday. In contrast, the findings related to COVID-19 suggest different mechanisms, with elevated mortality rates observed throughout most of the birthday period and no profound additional mortality increase on the birthday itself, although the findings in this diagnostic group are less robust than for other causes of death due to relatively small number of deceased with COVID-19 as the underlying cause of death in the analyzed data.\n\nAt this point, the proposed cursory interpretations are of course not much more than black box labels, the underlying mechanisms currently not understood in any detail. This underlines the fact that the present study in itself does not present a solution to the birthday effect-puzzle, but rather merely contributes to its solution. There are multiple reasons for this reservation. One is that, even though close to two million cases seems to be a lot, the number of data points reduces rapidly as the number of predictor variables increases. To avoid serious zero-inflation problems, assessing the effects of confounding by age directly by adding age group to the adjustment set of the Poisson models was not undertaken, as it would require a data set about 5 to 10 times the size of what was available. As such data do exist, albeit not in Switzerland, the hope certainly is that replication analyses are indeed undertaken with the adjustment set including age or age-group.\n\nAnother reservation is that the present analyses is heavy on assessing group effects with potentially very relevant individual-level variables not available. One such blind spot is how the level and timing of care and support provided to those nearing death may vary depending on the proximity to their birthday. While it is doubtful whether such data exist alongside the type of mortality data analyzed here, the present results highlight key demographic and diagnostic groups in which detailed observations and measurements could be undertaken efficiently and promisingly.\n\nWhile the present findings appear robust and convincing, it is crucial to keep in mind that the analyses were exploratory. Despite being pre-conceived, adjusted for seasonality and day of the week, and supported by thorough sensitivity assessments that suggest the findings are genuine rather than the result of systematic biases, confirmation through out-of-sample replication remains indispensable. Until such replication is achieved, placing full confidence in the results would be premature, akin to “whistling Dixie”, as only replication offers the definitive means to distinguish genuine findings from noise (Weitkunat et al., 2010). If all or some of the presented findings should replicate, this would subsequently stress the need for elucidating the underlying mechanisms. Reaching mechanistic explanations requires considerable interdisciplinary efforts as well as using a broad range of data beyond what had been available to us, with a focus on social and medical care circumstances, biomarkers, and psychometric measurements. Mechanistic research can also help to assess the fundamental assumptions underlying the use of birthday period as an instrumental variable. While it appears rather plausible to assume that proximity to a person’s birthday has health-relevant psychophysiological effects, including emotional, physiological, and behavioral ones (upon which the use of the birthday period as an instrumental variable is indeed based), we have not demonstrated that.\n\nAnother shortcoming of the present work, again to do with limited numbers of cases, is that only broad diagnostic groups could be analyzed. Given that sufficient numbers of cases are available, it would seem to be a priority to dissect cause of death categories by stratifying for more detailed underlying diagnoses, possibly along with consecutive, concomitant, and immediate causes of death. For instance, it might be that in some cancers, birthday effects may indeed exist, although the present overall analysis preliminarily indicates (and in agreement with other research) that they might not. Surprises are quite possible, as even within diagnostic categories different specific diagnoses might reflect opposing birthday-related mortality patterns that cannot be seen at the level of diagnostic categories.\n\nPsychosomatic and psychotherapy research confront momentous conceptual and methodological problems in studying the health impact of non-biological causes and interventions. Unlike biomedicine and pharmacotherapy research, which benefit from straightforward tools like randomized controlled trials (which, if done rigorously, can be clinchers of causal effects, albeit focusing more or less exclusively on internal validity), research into psychological factors grapples with confounding and identifying causal effects. It is therefore not much of a surprise that research efforts seem to willingly gravitate towards biological rather than psychological problems and mechanisms. This is reflected in advances in biochemical analysis capacities, now providing a virtually unlimited supply of novel potential predictors of health outcomes that allegedly hold substantial commercial potential, can be quantified efficiently and objectively, and lend themselves to endless data mining in ever-larger studies chasing diminishing and less transferable effects that frequently fail to generalize to the real world. In contrast, among non-biological risk factors, essentially only socioeconomic disparities and health-related behaviors (lifestyles) have compelled their recognition as crucial etiological factors within the scientific community, owing to their now well-established quantifiability, their pleiotropic effects, and their profound and unmistakable impacts on health. High-prevalence psychosocial health effects that substantially impact morbidity and mortality, arising directly from behaviors, include anorexia, sexually transmitted infections, self-inflicted injuries, violence-related health impacts, health effects of maladaptive behaviors (such as gambling, emotional overeating, or stress-related absenteeism), and the consequences of substance abuse (including misuse of illicit drugs, alcohol, tobacco, and prescription medications). Importantly, there are also prevalent health effects driven by emotions and psychosomatic processes rather than behaviors. These include anxiety, depression, burnout, chronic pain, gastrointestinal dysfunctions, sleep disturbances, fatigue, functional cardiovascular disorders, cardiophobia, skin conditions, dizziness, vertigo, hyperventilation, and a range of functional neurological symptoms. It is remarkable that, despite this unambiguous situation, the role of psychosomatic mechanisms continues to be a topic of ongoing debate. It might be that what masquerades as debates over the existence of psychosomatic effects actually reflects frustration with the lack of unequivocal demonstrations and mechanistic explanations, as well as a widespread failure to acknowledge the much greater difficulty in generating such evidence compared to run-of-the-mill biomedical research.\n\nData on causes of death in the Swiss resident population are registered with the Swiss Federal Statistical Office (FSO), to which the authors are affiliated. Data use is governed by the Swiss Civil Code and the Federal Statistics Act (FStatA). In line with this framework, which does not foresee ethical approval and consent procedures for routinely collected mortality data, the results are published in a form that does not allow the identification of individual cases.", "appendix": "Data availability statement\n\nAccess to data from the Swiss Federal Statistical Office (FSO) for scientific purposes is subject to an application process. Access decisions are based on the evaluation of each request. Requests can be placed at Gesundheit_DSV@bfs.admin.ch.\n\nThe Swiss statistics on cause of death are governed by the Swiss Civil Code of 10 December 1907 and by the Federal Statistics Act (FStatA) of 9 October 1992. This Act forms the basis for the Ordinance of 30 June 1993 regarding the conduct of federal statistical surveys, by virtue of which the Federal Statistical Office produces the deaths and cause of death statistics.\n\nNone.\n\n\nReferences\n\nAbel EL, Kruger ML: Heaping in anniversary reaction studies: A cautionary note. Omega. 2006; 54: 59–65. PubMed Abstract\n\nAbel EL, Kruger ML: Mortality salience of birthdays on day of death in the major leagues. Death Stud. 2009; 33: 175–184. PubMed Abstract \| Publisher Full Text\n\nAjdacic-Gross V, Knöpfli D, Landolt K, et al.: Death has a preference for birthdays – an analysis of death time series. Ann. Epidemiol. 2012; 22: 603–606. PubMed Abstract \| Publisher Full Text\n\nAlderson M: Relationship between month of birth and month of death in the elderly. Br. J. Prev. Soc. Med. 1975; 29: 151–156. PubMed Abstract \| Publisher Full Text\n\nAngermeyer MC, Kühn L, Osterwald P: Birthday and date of death. J. Epidemiol. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nStickley A, Ng CFS, Inoue Y, et al.: Birthdays are associated with an increased risk of suicide in Japan: Evidence from 27,007 deaths in Tokyo in 2001-2010. J. Affect. Disord. 2016; 200: 259–265. PubMed Abstract \| Publisher Full Text\n\nStumpfe KD: Psychosomatische Beziehung zwischen Todestag und Geburtstag. Aktuelle Gerontol. 1983; 13: 244–247. PubMed Abstract\n\nTrudeau R: Monthly and daily patterns of death. Health Rep (Statistics Canada). 1997; 9: 43–50.\n\nVaiserman A, Grigoryev PE, Belaya II, et al.: Variation of mortality rate during the individual annual cycle. Biogerontology. 2003; 4: 221–225. Publisher Full Text\n\nWasserman I, Stack S: Age, birthdays and suicide. J. Soc. Psychol. 1994; 134: 493–495. Publisher Full Text\n\nWeitkunat R, Kaelin E, Vuillaume G, Kallischnigg G: Effectiveness of strategies to increase the validity of findings from association studies: size vs. replication. BMC Med. Res. Methodol. 2010; 28: 47. 10.\n\nWilliams A, While D, Windfuhr K, et al.: Birthday Blues. Examining the association between birthday and suicide in a national sample. Crisis. 2011; 32: 134–142. PubMed Abstract \| Publisher Full Text\n\nYoung DC, Hade EM: Holidays, birthdays, and postponement of cancer death. JAMA. 2004; 292: 3012–3016. PubMed Abstract \| Publisher Full Text\n\nZonda T, Bozsonyi K, Veres E, et al.: The effect of birthday on the fluctuation of suicides in Hungary (1970-2002). Rev. Sociol. 2010; 20: 96–105." }	[ { "id": "355909", "date": "07 Feb 2025", "name": "Ronny Westerman", "expertise": [ "Reviewer Expertise Mortality research", "causes of death", "epidemiology" ], "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 would like to express my gratitude to the authors for granting me the opportunity to review the following manuscript.\nThe focal point of this research is the so-called \"birthday effect\" in mortality in Switzerland. The researcher analysed data from the Federal Statistical Office on the primary causes of death over a 35-year period. The methodology is both robust and sound, with a thorough description of each step.\nIt is evident that cardiovascular diseases, suicides and accidents exhibited the highest birthday-related mortality, while diabetes, respiratory diseases and other diseases demonstrated divergent patterns before and after the birthday. However, the authors do not provide an explicit explanation for this phenomenon.\nThe manuscript is of high quality overall. The following aspects require attention in the review: 1.An explanation for the results is necessary. 2.The description of the availability of the data for replication is absent. Is it possible to access the data for further analysis? 3. Please discuss the Seasonality of deaths. 4. The references should be updated with recent studies.\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": [] } ]	1	https://f1000research.com/articles/14-21
https://f1000research.com/articles/13-249/v1	04 Apr 24	{ "type": "Research Article", "title": "The potential of Sonneratia caseolaris mangrove leaves extract as a bioactive food ingredient using various water extract", "authors": [ "Hartati Kartikaningsih", "Nur Fitriana", "Ike Listya Anggraeni", "Bambang Semedi", "Maharani Pertiwi Koentjoro", "Nur Fitriana", "Ike Listya Anggraeni", "Bambang Semedi", "Maharani Pertiwi Koentjoro" ], "abstract": "Background\nSonneratia caseolaris, known as the red pidada, has been widely utilized by the Indonesian. S. caseolaris leaves contain various active compounds, contributing to their popularity in the treatment of various diseases. Mangrove leaves are also known to exhibit very high antioxidant activity. This study aims to assess the antioxidant activity of S . caseolaris leaves extracted using different solvents. The resulting extract was evaluated for antioxidant activity by the 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity (DPPH) techniques.\n\nMethods Analysis of total flavonoids, total phenols, identification of active compounds with Liquid Chromatography High Resolution Mass Spectrometry (LC-HRMS), and bioinformatics were also carried out to obtain temporary conclusions about the antioxidant activity of S. caseolaris leaf extract.\n\nResults The results indicated that S. caseolaris leaves extracted with methanol and distilled water exhibited the highest antioxidant activity compared to other extracts. The analysis of total flavonoids and total phenols yielded results consistent with the antioxidant activity tests. LC-HRMS results identified three compounds in all S. caseolaris leaf extracts with antioxidant activity, namely Tempo, Choline, and Betaine. Tempo demonstrated a higher antioxidant activity than Choline and Betaine, as indicated by the binding affinity values in the bioinformatics analysis.\n\nConclusions It is evident that S. caseolaris leaf extract has the potential to serve as an effective an antioxidant agent. Further research is needed to elucidate confirm the effectiveness of S. caseolaris leaves as an antioxidant substance, in order to utilize them as active components in food and enhance antioxidant consumption among consumers.", "keywords": [ "Sonneratia caseolaris", "mangrove leaves", "bioactive", "antioxidant", "food ingredient" ], "content": "Introduction\n\nMangrove are vegetation community that has high morphological and physiological adaptations and are widely distributed along coastal and brackish swamps in tropical, subtropical, to temperate regions.1 They are highly productivity and provide various ecosystem services for the environment and society. Physically and ecologically, mangroves function as protectors of coastal areas against rising sea levels, abrasion, waves, and storms.2 Biologically, mangrove ecosystems provide breeding and feeding grounds for marine biota.3,4 Additionally, mangrove ecosystems also function as pollutant filters, able to reduce carbon emissions so that they have the potential to mitigate climate change.5,6 Economically, mangrove ecosystem serves as a renewable resource for timber and tourism.7 Indonesia has the largest mangrove ecosystem in the world, which is around 26-29% of the world’s mangrove ecosystems.8 Several types of mangroves can be found in Indonesia, including Sonneratia caseolaris, Avicennia marina, Rhizophora mucronata, and Rhizophora apiculata.9,10\n\nS. caseolaris is a mangrove species from the Sonneratiaceae family. It typically reaches a length ranging from 5-15 meters and is equipped with respiratory roots. Its flowers are characterized by the presence of many stamens.11,12 S. caseolaris, commonly referred to as red pidada, is extensively employed by the Indonesian people due to its wide range of uses. S. caseolaris contains various active compounds, including flavonoids, phenolics, alkaloids, terpenoids, tannins, and saponins.13,14 The presence of these active compounds makes this mangrove species highly sought after for its potential as an antioxidant agent,9,15 antibacterial,9,14,15 anticytotoxicity,15 anti-allergy and antidiabetic.16 S. caseolaris leaves, in particular, are recognized for their high potential as an antioxidant agent.9\n\nAntioxidants are natural or artificial compounds that function to scavenge free radicals in the body. Antioxidants can be divided into endogenous antioxidants and exogenous antioxidants. Endogenous antioxidants are types of antioxidants that already exist in the body, while exogenous antioxidants are antioxidants obtained from outside the body, such as antioxidants obtained from food consumption. An imbalance between free radicals and antioxidants in the body can lead to damage and various unwanted effects, including inflammation, aging, and certain diseases, such as cancer and diabetes mellitus.17\n\nKelch-like ECH-associated protein 1 (Keap1) is a protein that plays a crucial role in reducing endogenous antioxidant activity. The Keap1 protein functions as an endogenous inhibitor of erythroid nuclear factor 2-related factor 2 (Nrf2). When Nrf2 is inhibited, it cannot enter the cell nucleus to induce the expression of antioxidant enzymes. Under abnormal conditions, oxidative stress increases and leading to various damages in the body.18–23\n\nIn food technology, antioxidants are added as bioactive food ingredients with the aim of improving food quality and consumer health. Previous research has shown that S. caseolaris leaves extracted with organic solvents such as ethanol and methanol exhibit high antioxidant activity. However, these solvents are toxic and can leave chemical residues in the extract, which can be dangerous when consumed. This study aims to evaluate the antioxidant activity of S. caseolaris leaves extracted with distilled water and various types of bottled drinking water. Furthermore, this study aims to evaluate the effectiveness of the bioactive compounds in S. caseolaris leaf extract in inhibiting the Keap1 protein, thereby enhancing antioxidant activity.\n\n\nMethods\n\nThe leaves of S. caseolaris were collected from a mangrove plant in Ujung Pangkah Gresik, East Java, Indonesia. Sample was collected from the third to the fifth leaf, counted from the top of each plant. These leaves were thoroughly rinsed to remove salt using running water and then air-dried at room temperature for 7 days. Subsequently, the dried leaves were mashed into a fine powder for extraction.\n\nThe extraction process utilized various solvent, including distilled water (A), bottled drinking from three different brands (Ax (B), Cx (C), Kx (D), and methanol (Cat. 179337, Sigma-Aldrich®) (E) for control purposes. The bottle drinking water (B, C, and D) was sourced from the Indonesian market. Mangrove leaf powder was extracted with each solvent at ratio 1:3 (w/v). The extraction was conducted through maceration at room temperature for 24 hours, followed by filtration using Whatman® quantitative filter paper, ashless, Grade 42 (Cat. WHA1442041, Merck). The extraction process was repeated three times for each solvent. The resulting filtrates were subsequently evaporated using a rotary evaporator (DLab rotary evaporator RG100-S, 40°C).\n\nThe extraction yield was calculated by the following formula.1 The pH of the extracts was measured with a pH meter (Toledo S-220-KIT).\n\nThe total phenol test refers to Ref. 23 Total phenol content was measured by mixing 30 μL Folin-Ciocalteu (Cat. F9252, Sigma-Aldrich®) 1.0 N reagent with 60 μL of the sample, placed on the microplate. Afterwards, 150 μL of 20% sodium carbonate (Cat. 223530, Sigma-Aldrich®) solution was placed on a microplate, incubated for 15 minutes at room temperature and in a dark place. Samples were centrifuged for 8 minutes at 1600 rpm. The supernatants were measured on a Microplate reader at a wavelength. The total phenol was calculated using the gallic acid (Cat. 91215, Sigma-Aldrich®) standard curve (mgGAE/100g).\n\n1 mL sample was mixed with 3 mL of ethanol 96% (Cat No. 159010, Sigma-Aldrich®), 0.2 mL of 10% aluminium chloride (Cat. 8.01081, Sigma-Aldrich®), 0.2 mL of 1 M potassium acetate (Cat No. 1.04820, Sigma-Aldrich®), and 5.6 mL of distilled water. The mixture was incubated for 10 minutes at room temperature. The supernatant was measured on a Microplate reader at a wavelength of 415 nm. The total phenol was calculated using the standard curve of quercetin acid (mgQE/100g) (Lerck, Cat No. PHR1488).\n\nAntioxidant activity was measured using a microplate reader (96-flat bottom with lid).23 The DPPH (Cat. 300267, Sigma-Aldrich®) reagent was prepared at a concentration of 0.2 mM in absolute ethanol. The first column was filled with 200 μL of ethanol as a blank. Line A was filled with sample stocks. 100 μL of ethanol solvent was added to each well plate for serial dilution, except for line A. The second -11th column in line A was filled with 200 μL of sample stock solution. Serial dilution was carried out by taking 100 μL from line A, and placing it in lines B, C, D, E, F, and G respectively. In line G, 100 μL solution was removed, resulting in each well containing 100 μL of the solution. Subsequently, 100 μL of DPPH was introduced into each well. Line H was negative control (100 μL solvent mixed with 100 μL of 0.2 mM DPPH reagent). The plate was covered and incubated at room temperature for 30 minutes. All samples were measured at 517 nm wavelength. DPPH absorption inhibition was calculated with the following formula.2\n\nIn order to obtain the percentage of inhibition (Inhibitory Concentration value, IC50), a linear regression curve (Y=ax + b) was made with the x-axis as the concentration (μg/mL) and the y-axis as the percentage of inhibition. The IC50 value is a calculation of how much of the sample concentration is needed to inhibit 50% of free radical activity.\n\nTreatment was statistically tested using a completely randomized design with 3 replications. Significantly different results (P<0.05) were followed by Duncan’s test.\n\nLC-HRMS used HPLC (Thermo Scientific Diorex Ultimate 3000 RSLCnano, Japan) with solvent A (0.1% formic acid in water) and solvent B (0.1% formic acid in Acetonitrile), column Hypersil GOLD, 5 μm, 150 × 4.6 mm, flow rate 40 μL/min, column temperature 30°C. Mass Spectrometer used Scientific Q Exactive™ with software compound discovery with mzCloud™ MS/MS library. Samples were diluted to a volume of 1500 μL, and vortexed at 2000 rpm, for 2 min. The supernatant was filtered with a 0.22 μm syringe filter. The vial was inserted into the LC-HRMS autosampler\n\nThis study used three compounds that had been identified through LC-HRMS analysis, namely Betaine (CID 247), Choline (CID 305), Tempo (CID 549976), and one control compound (CID 135263934) which acts as the original inhibitor of the Keap1 protein. Ligand was obtained from the PubChem web server (https://pubchem.ncbi.nlm.nih.gov/) in Sybil Data Files (SDF) format. Betaine, Choline, and Tempo are compounds that are known to have antioxidant activity. In this study, it is anticipated that they may bind to Keap1 (6TYP), which serves as an endogenous Nrf2 inhibitor. as an endogenous Nrf2 inhibitor. The Keap1 3D structure was obtained from the RCSB PDB web server (https://www.rcsb.org) Keap1 was then prepared to remove water molecules and ligands with BIOVIA Discovery Studio 2019.\n\nLigands-Keap1 interactions were analysed by molecular docking using AutoDock Vina integrated in PyRx version 0.9.5. The molecular docking process is carried out using a specific docking method based on the active site of the Keap1 protein.17,22 Docking results, bond positions, and amino acid residues formed between ligands-receptor were analysed using PyMol software and BIOVIA Discovery Studio 2019.\n\nMolecular dynamic simulations were carried out using Yet Another Scientific Artificial Reality Application (YASARA). The MD simulation aims to compare the interaction complexes of Betaine, Choline, Tempo, and Keap1-binding inhibitors. The parameters in the simulation correspond to the physiological conditions of the cells, namely temperature 37 °C, 1 atm, pH 7.4, and 0.9% salt content for 50 ns with autosaved every 25 ps. The simulation is run by the md_run macro program, and the results are displayed by the md_analyze and md_analyzeres programs.24–26\n\n\nResults\n\nThe mass yield percentage, pH, total flavonoids, total phenols, and antioxidant activity of S. caseolaris mangrove leaves extracted using various solvents yielded different results. Mangrove leaves of S. caseolaris extracted with methanol (E) and distilled water (A) exhibited high antioxidant activity, with IC50 values of 63.72 ± 1.27 mg/mL and 89.29 ± 006 mg/mL respectively. The high antioxidant activity was supported by the total amount of flavonoids and total phenol of both extracts, which were higher than the other extracts. In contrast, mangrove leaves extracted with bottled drinking water (Ax, Cx, and Kx) showed low antioxidant activity. These results were consistent with lower levels total flavonoids and total phenols compared to treatments A and E. Based on both our research findings and prior studies, it is well-established that total flavonoids and total phenols exert a substantial influence on high antioxidant activity.27,28 Mangrove leaves extracted with distilled water (A), Ax brand bottled mineral water (B), and methanol (E) have a pH of less than 7 (Table 1).42\n\nThe content of bioactive compounds in mangrove leaf extract of S. caseolaris was analyzed using LC-HRMS. The analysis results revealed the presence of 16 identical active compounds in all S. caseolaris mangrove leaf extracts obtained with different solvents (Table 2). The bioactive compound content in mangrove leaf extract from S. caseolaris was assessed through LC-HRMS. The findings from the analysis indicated the presence of 16 identical active compounds in all S. caseolaris mangrove leaf extracts, regardless of the solvents used (refer to Table 2). Among these, Tempo, Betaine, and Choline were identified as active compounds known for their antioxidant properties.\n\nThis study compared the binding activities of Betaine, Choline, Tempo, and inhibitors to the Keap1 protein. The inhibitor used is a natural inhibitor that binds to the active site of the Keap1 protein and serves as a control. Molecular docking results were visualized with PyMol and Discovery Studio to identify the binding site of Ligand-Keap1.\n\nMolecular docking results indicated that the Tempo-Keap1 interaction possesses a lower binding affinity value compared to the Betaine-Keap1 and Choline-Keap1 interactions, which have values of -5.5 kcal/mol, -4.2 kcal/mol, and -3.7 kcal/mol, respectively. The lower binding affinity value observed for Tempo-Keap1 interaction suggests its strength is greater than that of the Betaine-Keap1 and Choline-Keap1 interactions. Notably, the binding affinity value of Tempo-Keap1 is higher than that of Inhibitor-Keap1, which exhibits a binding affinity value of -11.1 kcal/mol (Table 3).\n\nThe visualization results revealed that Betaine shares the same binding site as the inhibitor (control), whereas Tempo and Choline do not exhibit a specific binding site. They are overlap with the binding of the inhibitor (Figure 1 on the top row). The visualization process continued with the identification of the amino acid residues involved in the Ligand-Keap1 interaction. It is observed that the Betaine-Keap1 interaction shares the same amino acid residues as the inhibitor-Keap1 interaction, and these interactions involve the highest number of residues when compared to the Tempo-Keap1 and Choline-Keap1 interactions. Specifically, Ser363, Arg380, and Arg415 were identified as three amino acid residues engaged in hydrogen interactions on Betaine-Keap1. Furthermore, the visualization results indicated that Arg380 and Arg415 in Betaine-Keap1 interaction were also involved in hydrophobic interactions. Additionally, five other amino acid residues engaged in hydrophobic interactions in the Betaine-Keap1 were Tyr334, Gly364, Ala556, Ser602, and Gly603 (Table 3 and Figure 1 in the bottom row).\n\nA-D. Visualization of molecular docking results between inhibitors, Tempo, Betaine, and Choline with Keap1 protein. The top row shows each ligand's position that binds to the Keap1 protein. The bottom row shows the amino acid residues involved in the interactions of the inhibitor, Tempo, Betaine, and Choline with the Keap1 protein. Keap1 protein is presented as a pink ribbon. The compounds are presented in the form of colored sticks, namely inhibitors (blue), Tempo (orange), Betaine (gray), and Choline (yellow).\n\nFour amino acid residues are shared between the Tempo-Keap1 and inhibitor-Keap1 interactions, namely Gly462, Gly509, Ala556, and Gly603, all of which are involved in hydrophobic interactions. The Choline-Keap1 interaction shares the same two amino acid residues as the inhibitor-Keap1 interaction, namely Gly509 and Ala556, and these interactions are also hydrophobic in nature. The presence of identical amino acid residues among Tempo, Betaine, Choline, and the inhibitor in the Keap1 protein suggests the possibility that these compounds may exhibit similar activity to the inhibitor (control).\n\nThe binding affinity of the ligand-receptor complex increases when amino acid residues are involved in the interaction through hydrogen bonds and hydrophobic bonds. Furthermore, it is well-known that hydrophobic interactions play a significant role in stabilizing the ligand-protein bond and contribute to enhancing the ligand’s affinity for the protein.29,30\n\nHowever, the results reveal an interesting phenomenon. Although, the Betaine-Keap1 interaction involves more amino acid residues than the Tempo-Keap1 interaction, the binding affinity value of Betaine-Keap1 is higher than that of Tempo-Keap1. It is possible that the amino acid residues Gly462 and Gly509 play a specific role in the Tempo-Keap1 interaction, and these particular residues are not found in the Betaine-Keap1 interaction (Table 3 and Figure 1 in the bottom row).\n\nHowever, it’s important to note that a higher binding affinity value in the Tempo-Keap1 interaction does not necessarily imply that it is weaker than the inhibitor-Keap1 interaction with its lower binding affinity value. This is because in molecular docking simulations consider multiple parameters to determine the strength or weakness of the ligand-receptor interaction. These parameters include the binding affinity value, hydrogen interaction, and hydrophobic interactions formed on amino acid residues between the ligand-receptor, and molecular dynamic results.31\n\nAfter molecular docking, molecular dynamics (MD) simulations were conducted to assess the stability of the Ligand-Keap1 interaction complex. The parameters employed in this simulation included the Root Mean Square Deviation (RMSD) of the Ligand-Keap1 complex, RMSD of Ligand movement, Root Mean Square Fluctuation (RMSF), and the number of hydrogen bonds within the Ligand-Keap1 complex.22\n\nThe MD analysis results indicated that both the Tempo-Keap1 and Choline-Keap1 complexes exhibited similar stability to the Keap1 inhibitor throughout the simulation, as evidenced by an RMSD value of <2 Å. On the other hand, the Betaine-Keap1 complex at the beginning of the simulation up to 20 ns also showed the same stability, but after 20 ns the complex was unstable until the end of the simulation with an RMSD value of >3 Å. The RMDS Ligand movement results further supported the observation of Betaine’s instability after 20 ns, with significantly differing RMSD values when compared to Tempo, Choline, and inhibitors (Figures 2A and 2B). These findings align with previous studies, which reported that a stable ligand-receptor complex during the simulation typically maintain an RMSD value of 3 Å.32,33\n\nThe stability of the ligand-Keap1 interaction complex during the simulation was indicated by (A) the RMSD of the ligand-Keap1 complex value, (B) the RMSD of the Ligand movement value, (C) the RMSF value, and (C) the number of hydrogen bonds.\n\nThe fluctuation of the amino acid residues also serves as an indicator of the stability of the ligand-Keap1 complex. The results demonstrated that all complexes exhibited nearly identical fluctuations in amino acid residues (Figure 2C). Previous research has indicated that a higher degree of fluctuation among amino acid residues correlates with increased instability in the e ligand-receptor complex.34 In all interaction complexes, more than 200 hydrogen bonds were observed (Figure 2D). The abundance of hydrogen bonds within these complexes indicates their stability. However, it’s important to note that the stability of the Betaine-Keap1 interaction requires further analysis using additional parameters to ensure a conclusive determination. This is because the results of the RMSD Ligand-Keap1 complex, RMSD Ligand movement, RMSF, and the number of hydrogen bonds of the Betaine-Keap1 complex exhibit variations.\n\n\nDiscussion\n\nS. caseolaris is a variety of mangrove commonly utilized by Indonesian community. Previous studies have indicated that mangrove leaves have very high antioxidant activity.9,35 reported that the ethanol extract of of S. caseolaris mangrove leaves demonstrated high antioxidant activity, which varies with the leaves’ maturity level. Other studies have also suggested that the high antioxidant activity of S. caseolaris mangrove leaf extract can be attributed to the presence of several active compounds, such as flavonoids and tannins.9\n\nIn the present study, mangrove leaves of S. caseolaris extracted using various types of solvents have shown potential as antioxidant agents, as determined by the DPPH method. Among the extracts, S. caseolaris leaves extracted with methanol (control) and distilled water exhibited the highest antioxidant activity compared to other extracts. This is supported by the higher flavonoid and phenolic content found in the methanol and distilled water extracts, as compared to other extracts.\n\nFurther study showed that molecular docking and molecular dynamics simulations assessed the differential binding affinities and stability of Ligand-Keap1 interactions involving Betaine, Choline, and Tempo. Molecular docking is recognized as one of the vital methods in molecular simulation. This method is based on the recognition process involving spatial matching and intermolecular energy matching between two or more molecules. Molecular docking is an effective and efficient approach that helps reduce costs and time in studying the mechanism of compound activity.36\n\nTempo, Betaine, and Choline are recognized as active compounds with antioxidant properties.37 Previous studies have demonstrated that Choline significantly enhances the activity of glutathione S-transferase, superoxide dismutase, catalase, and glutathione peroxidase Other research has indicated that Tempo, when conjugated with specific compounds, can effectively reduce oxidative stress caused by hydrogen peroxide.38 Betaine, functioning as an antioxidant agent, has been shown to significantly decrease malondialdehyde (MDA) levels while increasing the activity of superoxide dismutase (SOD) and glutathione peroxidase (GPx).39,40 Based on the findings of these studies, the present research employed Tempo, Choline, and Betaine as potential candidates for inhibiting the Keap1 protein, thereby enhancing antioxidant activity in the body. The result providing valuable insights into the potential activities of Betaine, Choline, and Tempo as compared to the control inhibitor. The results showed that the ability of Betaine, Choline, and Tempo to inhibit the Keap1 protein, which acts as a Nrf2 inhibitor using a molecular docking approach.\n\nOxidative stress is an imbalance between free radicals-antioxidants in the body, resulting in damage and contributing to various diseases, such as cancer, diabetes, neurodegenerative diseases, and aging.17 Nuclear factor erythroid 2-related factor 2 (Nrf2) is a protein that plays an important role in preventing oxidative stress. Nrf2 is the primary regulator of the cellular stress response, triggering the expression of antioxidant enzymes that protect cells from oxidative stress induced.22,41 Under normal circumstances, Nrf2 is bound by the endogenous inhibitor Kelch-like ECH-associated protein 1 (Keap1) in the cytosol. The Keap1 protein contains cysteine residues that can bind to ROS when oxidative stress occurs, leading to disruption of the Nfr2-Keap1 bond. This disruption results in phosphorylation and translocation of Nrf2 into the cell nucleus. Translocated Nrf2 then binds to a DNA regulatory region known as the ARE (Antioxidant Response Element). The Nrf2-ARE binding activates the transcription of genes responsible for encoding antioxidant enzymes, including superoxide dismutase, catalase, glutathione peroxidase, thioredoxin reductase, glutamate cysteine ligase, and others.18–21\n\nIn conclusion, the results revealed the presence of the Tempo, Choline, and Betaine compounds with antioxidant potential in all extracts. The antioxidant potential of these three compounds was further. Confirmed through by the results of molecular docking, indicating their ability to inhibit the Keap1 protein - endogenous Nrf2 inhibitor, and the enhance the activity of enzymes involved in antioxidant processes. Further research is necessary to validate the potential of S. caseolaris leaves as an antioxidant agent, with the aim of utilizing them as bioactive food ingredients and increasing antioxidant intake for consumers.", "appendix": "Data availability\n\nFigshare: The potential of Sonneratia caseolaris mangrove leaves extract as a bioactive food ingredient using various water extract, https://doi.org/10.6084/m9.figshare.25100123.v1. 42\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 thank the Graduate School of Universitas Brawijaya, Malang, Indonesia with the research contract number of Postgraduate School of Universitas Brawijaya No: 1549/UN10.F40/PT/2023.\n\n\nReferences\n\nPolidoro BA, Carpenter KE, Collins L, et al.: The loss of species: Mangrove extinction risk and geographic areas of global concern. PLoS One. 2010; 5: e10095. 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Publisher Full Text" }	[ { "id": "264930", "date": "29 Apr 2024", "name": "Neni Anggraeni", "expertise": [ "Reviewer Expertise herbal medicines", "metabolism", "stem cell biology and animal modelling" ], "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 constructs an exciting experiment, using the result of bioinformatics we can predict not only the antioxidant activity but also the binding site of the molecules for further study, before conducting in-vitro and in-vivo. However, I have the following concern that I hope the author can explain:\nPlease explain why you need to use various types of bottled drinking water. In the introduction, it is better to elaborate on the third and 4th paragraphs to make it more engaging. Is it correct based on my understanding, that the antioxidant will bind with Keap1 so the Nfr2 will function normally? Please reconstruct those paragraphs. Plant extraction especially mangrove leaves has been widely explored nowadays, so it’s not difficult to find recent citations. Some citations published more than ten years ago that are used in this paper, it’s better to replace the citation with the latest one. In the method, the author did not mention the weight of the leaves and the dry leaves and why you chose the leaves from the third to the fifth leaf. Some sentences will cause misunderstanding, such as whether the author used the dry extract or fractionation to analyze the total phenol and flavonoids. In that section, you used 60 μL and 1 ml samples respectively, but you did not mention the dilution, using what kind of solution you diluted the dry extract. The author did not mention the statistical test they used to get the p-value. “Among these, Tempo, Betaine, and Choline were identified as active compounds known for their antioxidant properties” results section, 2nd paragraph line 5. If this statement is from other papers please put the citation. However, it’s better to add biological function analysis of those compounds regarding their antioxidant properties using analysis tools (e.g. PASS online or others) to make this article more enriched. The aim of this study was to evaluate the effectiveness of the bioactive compounds in S. caseolaris leaf extract in inhibiting the Keap1 protein. However, I didn’t get the answer about the effectiveness of those compounds compared to the inhibitor.\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": [ { "c_id": "11498", "date": "30 May 2024", "name": "Hartati Kartikaningsih", "role": "Author Response", "response": "Dear Dr. Neni Anggraeni, Subject: Response to Review of Manuscript We would like to express our sincere gratitude for the thoughtful and constructive feedback provided on our manuscript titled \"The potential of Sonneratia caseolaris mangrove leaves extract as a bioactive food ingredient using various water extract\". Your insightful comments have significantly contributed to improving the quality and clarity of our work. We have carefully considered each of your suggestions and addressed them as follows: 1. Reviewer's Comment: Please explain why you need to use various types of bottled drinking water. Author response: The extraction application of Sonneratia caseolaris in society will be easier to perform using bottled drinking water. Bottled drinking water is a type of mineral commonly used in Indonesia 2. Reviewer's Comment: In the introduction, it is better to elaborate on the third and 4th paragraphs to make it more engaging. Is it correct based on my understanding, that the antioxidant will bind with Keap1 so the Nfr2 will function normally? Please reconstruct those paragraphs. Author response: Thank for your suggestion, we are going to elaborated of paragraphs 3 and 4. Keap1 and Nfr2 lead to ubiquitination, resulting in the mimicry of Nfr2 degradation. The degradation of Nfr2 diminishes the regulation of antioxidant response elements when oxidative stress occurs. Therefore, in this study, it is anticipated that the antioxidants found in S. caseolaris will be able to bind to Keap1, allowing Nfr2 to function effectively within the cell. The effectiveness of S. caseolaris antioxidants in binding to Keap1 is crucial for maintaining Nfr2 functionality. By preventing the degradation of Nfr2, these antioxidants ensure that the regulatory mechanisms for antioxidant response remain intact even under conditions of oxidative stress. This mechanism is vital for cellular defence against oxidative damage and underscores the potential therapeutic significance of S. caseolaris in combating oxidative stress-related disorders. 3. Reviewer's Comment: Plant extraction especially mangrove leaves has been widely explored nowadays, so it’s not difficult to find recent citations. Some citations published more than ten years ago that are used in this paper, it’s better to replace the citation with the latest one. Author response: Thanks for your suggestion. Here's the revised paragraph with the old reference highlighted in green References Sindhu S, Hiroshi T, Rikuo R: Community-based portable reefs to promote mangrove vegetation growth: bridging between ecological and engineering principles. Int. J. Environ. Res. Public Health. 2021 12; 590. Nur SH, Hilmi E: The correlation between mangrove ecosystem with shoreline change in Indramayu coast. IOP Conf. Ser. Earth Environ. Sci. 2021; 819: 012015. Eddy S, Rasyid RM, Iskandar I, et al.: Community-Based Mangrove Forests Conservation for Sustainable Fisheries. J. Silvikultur Trop. 2016; 07: 42–47. Umilia E, Asbar: Formulation of mangrove ecosystem management model based on Eco-minawisata in the Coastal Sinjai, South Sulawesi. Procedia. Soc. Behav. Sci. 2016; 227: 704–711. Alongi DM: Impact of global change on nutrient dynamics in Mangrove Forests. Forests. 2018; 9. Alongi DM: Impacts of climate change on blue carbon stocks and fluxes in mangrove forests. Forests. 2022; 13: 149. 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Wargasetia TL, Ratnawati H, Widodo N, et al.: Bioinformatics study of sea cucumber peptides as antibreast cancer through inhibiting the activity of overexpressed protein (EGFR, PI3K, AKT1, and CDK4). Cancer Inform. 2021; 20: 117693512110318. Azminah A, Radji M, Mun’Im A, et al.: In silico study of sirt1 activators using a molecular dynamic approach. Int. J. Appl. Pharm. 2019; 11: 237–245. Winarti RBS: Antioxidant activity os Sonneratia caseolaris mangrove leaf extract based on leaf maturity level. J. Marine Coastal Sci. 2019; 8(3): 134–136. Morris CJ, Della CD: Using molecular docking and molecular dynamics to investigate protein-ligand interactions. Mod. Phys. Lett. B. 2021; 35: 2130002. Xin X, Tingting Y, Xiang Li, et al.: Recent advances in targeting the “undruggable” proteins: from drug discovery to clinical trials. Sig. Transduct. Target. Ther. 2023;8:335. DeJulius CR, Dollinger BR, Kavanaugh TE, et al.: Optimizing an Antioxidant TEMPO Copolymer for Reactive Oxygen Species Scavenging and Anti-Inflammatory Effects in Vivo. Bioconjug Chem. 2021; 32(5):928-941. Le DQ, Kuriakose AE, Nguyen DX, et al.: Hybrid Nitric oxide donor and its carrier for the treatment of peripheral arterial diseases. Sci. Rep. 2017; 7: 1–11. Hassanpour S, Rezaei H, Razavi SM: Anti-nociceptive and antioxidant activity of betaine on formalin- and writhing tests induced pain in mice. Behav. Brain Res. 2020; 390: 112699. Oh YS, Jun HS: Effects of glucagon-like peptide-1 on oxidative stress and Nrf2 signaling. Int. J. Mol. Sci. 2018; 19(1): 1–16. Hartati K, Fitriana N, Anggraeni IL, et al.: The potential of Sonneratia caseolaris mangrove leaves extract as a bioactive food ingredient using various water extract. [Dataset]. figshare. 2024. 4. Reviewer's Comment: In the method, the author did not mention the weight of the leaves and the dry leaves and why you chose the leaves from the third to the fifth leaf. Author response: Thanks for your review, we are going to revise it. Sample extraction The extraction process utilized various solvent, including distilled water (A), bottled drinking from three different brands (Ax (B), Cx (C), Kx (D), and methanol (Cat. 179337, Sigma-Aldrich ® ) (E) for control purposes. The bottle drinking water (B, C, and D) was sourced from the Indonesian market. Mangrove leaf powder was extracted with each solvent at ratio 1:3 (w/v). The extraction was conducted through maceration at room temperature for 24 hours, followed by filtration using Whatman ® quantitative filter paper, ashless, Grade 42 (Cat. WHA1442041, Merck). The extraction process was repeated three times for each solvent. The resulting filtrates were subsequently evaporated using a rotary evaporator (DLab rotary evaporator RG100-S, 40°C). Every 1000 grams of fresh leaves will become 200 grams of dried leaves. In this study, the third and fifth leaves are selected based on previous research findings. These leaves have shown higher total phenol, total alkaloid, and flavonoid content compared to other parts of the leaf. However, there is no significant difference in pH and viscosity. 5. Reviewer's Comment: Some sentences will cause misunderstanding, such as whether the author used the dry extract or fractionation to analyze the total phenol and flavonoids. In that section, you used 60 μL and 1 ml samples respectively, but you did not mention the dilution, using what kind of solution you diluted the dry extract. Author response: Thanks for your review, we are going to revise it. Calculation of extraction yield and pH examination The extraction yield was calculated by the following formula.1 The pH of the extracts was measured with a pH meter (Toledo S-220-KIT). The analyzed sample is a working solution derived from the evaporator, consisting of 1 mL of solution mixed with 20 mL of double-distilled water. 6. Reviewer's Comment: The author did not mention the statistical test they used to get the p-value. Author response: The statistical test used was the Duncan Test with SPSS version 21, where the significance level was set at p-value < 0.05. This information will be added as a note in Table 1. 7. Reviewer's Comment: “Among these, Tempo, Betaine, and Choline were identified as active compounds known for their antioxidant properties” results section, 2nd paragraph line 5. If this statement is from other papers please put the citation. However, it’s better to add biological function analysis of those compounds regarding their antioxidant properties using analysis tools (e.g. PASS online or others) to make this article more enriched. Author response: Thanks for your review, we are going to revise it. “The content of bioactive compounds in mangrove leaf extract of S. caseolaris was analyzed using LC-HRMS. The analysis results revealed the presence of 16 identical active compounds in all S. caseolaris mangrove leaf extracts obtained with different solvents (Table 2). The bioactive compound content in mangrove leaf extract from S. caseolaris was assessed through LC-HRMS. The findings from the analysis indicated the presence of 16 identical active compounds in all S. caseolaris mangrove leaf extracts, regardless of the solvents used (refer to Table 2). Among these, Tempo, Betaine, and Choline were identified as active compounds known for their antioxidant properties38,40.” We also added some information for biological function of Tempo, Betaine, and Choline, namely: “Tempo finds extensive use in cellular and animal model studies due to its favorable characteristics, including low molecular weight, high water solubility, and the capability to permeate cell membranes efficiently43. Betaine, a natural compound, is a glycine derivative with three additional methyl groups, as depicted in Figure 1 in both 2D and 3D forms. It is widely found in plants, animals, and microorganisms, including beets, spinach, wheat bran, wheat germ, and aquatic invertebrates. Betaine can be produced endogenously through choline metabolism or obtained exogenously through dietary intake, exhibiting similar bioavailability whether consumed as a supplement or through food, ultimately metabolizing to dimethylglycine and sarcosine in kidney and liver cells' mitochondria44. Choline, a vital nutrient, plays a fundamental role in diverse biological functions such as the synthesis of structural lipoproteins and membrane lipids, neurotransmitters, and the maintenance of brain function. Betaine, derived from choline oxidation, is crucial for the formation of essential amino acids like methionine. Dimethylglycine (DMG), a betaine metabolite, serves as a vital source of glycine and methyl groups for biochemical reactions. Additionally, trimethylamine N-oxide (TMAO) is metabolized in the liver from trimethylamine (TMA), primarily generated from dietary choline and betaine, and emerges as a risk factor in various diseases, including renal disease, cardiovascular disease, colorectal cancer, type II diabetes, and neurological disorders. This highlights the multifaceted role of choline metabolism and its metabolites in health and disease45. Reference: 43. Marcin L, Krzysztof G. 2017. Nitroxides as antioxidants and anticancer drugs. IJMS, 18(11): 2490. 44. Dobrijević D, Pastor K, Nastić N, et al. J. Betaine as a functional ingredient: metabolism, health-promoting attributes, food sources, applications and analysis methods. Molecules. 2023;28(12):4824. 45. Yvonne M, Christiane S, Manfred W, Petter-Arnt H. Plasma kinetics of choline and choline metabolitesafter a single dose of superbaboosttm krill oil orcholine bitartrate in healthy volunteers. Nutrients. 2019;11;2548. 8. Reviewer's Comment: The aim of this study was to evaluate the effectiveness of the bioactive compounds in S. caseolaris leaf extract in inhibiting the Keap1 protein. However, I didn’t get the answer about the effectiveness of those compounds compared to the inhibitor. Author response: Thanks for your review, we are going to revise it. “We aim to enhance our analysis to predict how potential compounds in Sonneratia caseolaris leaves water extract interact with the target protein Keap1. S. caseolaris, has potential compounds interact with a target protein like Keap1, is crucial for understanding their potential therapeutic potential.” ------------------------------------- We believe that implementing these changes has strengthened the manuscript and enhanced its contribution to the field. We are confident that the revised version now meets the standards of F1000 and will be of interest to its readership. Once again, we appreciate your time and effort in reviewing our manuscript. Your expertise and constructive criticism have been invaluable in improving our work. Please do not hesitate to contact us if you have any further questions or require additional clarification. Thank you for your consideration. Sincerely, Hartati Kartikaningsih Universitas Brawijaya" } ] } ]	1	https://f1000research.com/articles/13-249
https://f1000research.com/articles/12-47/v1	12 Jan 23	{ "type": "Research Article", "title": "Expression of MAP2, GFAP, and calcium in CA3 area of a modified organotypic hippocampal slice culture treated with kainic acid", "authors": [ "Machlusil Husna", "Kusworini Handono", "Hidayat Sujuti", "Aulanni'am Aulanni'am", "Ettie Rukmigarsari", "Kusworini Handono", "Hidayat Sujuti", "Aulanni'am Aulanni'am", "Ettie Rukmigarsari" ], "abstract": "Background: Neurodegeneration due to neurotoxicity is one of the phenomena in temporal lobe epilepsy. Experimentally, hippocampal excitotoxicity process can occur due to kainic acid exposure, especially in the CA3 area. Neuronal death, astrocyte reactivity and increased calcium also occur in hippocampal excitotoxicity, but few studies have investigated immediate effect after kainic acid exposure. The organotypic hippocampal slice culture (OHSC) is a useful model for studying the neurodegeneration process, but there are still many protocol differences. In this study, minor modifications were made in the OHSC protocol. Methods: OHSC was obtained from two healthy wild type Wistar rats aged P10. Healthy culture slices were obtained and lasted up to 10 days in vitro (DIV 10). Bath application of kainic acid for 48 hours in DIV 10 followed by observation of its initial effects on neurons, astrocytes, and calcium via the expression of MAP2, GFAP, and intracellular calcium, subsequently. Results: After 48 h of kainic acid administration, there was a significant increase in intracellular calcium (p = 0.006 < α), accompanied by a significant decrease in MAP2 (p = 0.003 < α ) and GFAP (p = 0.010 < α) expression. Conclusion: These findings suggest early neuronal and astrocyte damage at the initial onset of hippocampal injury. This implies that astrocyte damage occurs early before an increase in GFAP that characterizes reactive astrogliosis found in other studies. Damage to neurons and astrocytes may be associated with increased intracellular calcium. It is necessary to develop further research regarding expression of calcium, MAP2, and GFAP at a spatial time after exposure to kainic acid and strategies to reduce damage caused by excitotoxicity.", "keywords": [ "organotypic hippocampal slice culture", "MAP2", "GFAP", "intracellular calcium", "excitotoxicity", "kainic acid" ], "content": "Introduction\n\nNeurodegeneration is a process of progressive loss of neuron function and structure. One of the main mechanisms in both acute and chronic neurodegeneration is excitotoxicity, which can occur in Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD), temporal lobe epilepsy (TLE), and amyotrophic lateral sclerosis (ALS) (Mohd Sairazi et al., 2015). The chemoconvulsant agent kainic acid (KA), which is an AMPA/kainate receptor agonist, is often used experimentally to induce excitotoxicity, as it plays a role in the development of epileptic seizures similar to conditions in epileptic patients (Holopainen et al., 2004; Tanioka et al., 2020; Zhang & Zhu, 2011).\n\nNeuronal death as a result of glutamate excitotoxicity has been extensively studied, both in vivo and in vitro (Holopainen et al., 2004; Liu et al., 2001; Noraberg et al., 1999; Sanon et al., 2005; Sattler & Tymianski, 2001). Glutamate represents the primary excitatory neurotransmitter in the mammalian brain, and it is used by neurons, astrocytes, and other glial cells to process information (Mihály, 2019). The main mechanism behind neuronal death is glutamate excitotoxicity (Cho, 2013; Sanon et al., 2005), which occurs through necrosis, apoptosis, or both (Holopainen et al., 2004; Liu et al., 2001). Other than neuronal death, a significant increase in astrocyte reactivity has been observed in response to central nervous system injury (Jeong et al., 2014; Miller et al., 2017; Yildirim et al., 2019; Zhang et al., 2017; Zhang & Zhu, 2011). Increased astrocyte reactivity (astrogliosis), characterized by increased GFAP expression, is known to exert a protective effect against neuronal death due to nervous system injury (Jeong et al., 2014; Miller et al., 2017; Zhang et al., 2017). Prior studies have found this process to be closely related to calcium level. In the context of epilepsy, the calcium hypothesis suggests an increase in intracellular calcium levels. If this increase is not sufficient to induce excitotoxicity, it can result in temporary or permanent neuroplasticity changes and degeneration. In the long term, these changes can cause abnormal neuronal firing and so lead to epilepsy (Zhang et al., 2019).\n\nDamage due to kainic acid mainly occurs in the hippocampus. Such damage may be caused by the high density of kainic receptors in that area (Mihály, 2019; Zhang & Zhu, 2011). The CA3 circuit plays a role in the epileptiform activity within the hippocampus. Song et al. found hyperexcitability due to hypoxic-hypoglycemic injury to be more visible in the CA3 area than in other hippocampal areas (Song et al., 2018). The maturation of synapses, receptors, and intrinsic fiber pathways in the OHSC also occurs in in-vivo conditions, which renders it a useful model for studying age-related conditions in a maintained environment (Holopainen et al., 2004). To date, various methods have been used to make OHSCs. The preparation of acute hippocampal slices at room temperature gives rise to better results, especially with regard to electrophysiological observations (Eguchi et al., 2020). In contrast, OHSC processing requires cold temperatures during slicing in order to reduce the damage that may occur throughout the surgical process, although there are still many variations when it comes to the use of cold temperatures (Lein et al., 2011; Opitz-Araya & Barria, 2010; Simoni & Yu, 2006; Varela et al., 2012).\n\nOnly a few prior studies have investigated the immediate impact of hippocampal insult to neuron and astrocyte (Jeong et al., 2014; Zhao et al., 2003). This study aimed to observe the immediate effect of kainic acid on neurons, astrocytes, and intracellular calcium in the CA3 area of modified OHSC.\n\n\nMethods\n\nThis study used three healthy wild type Wistar rats aged postnatal day 10 (P10), weight range 10-15 grams, regardless of sex. The sample size was decided from Federer’s formula (Ihwah et al., 2018). All of the Wistar rats were purchased from Animal Lab of Biosains Institute Universitas Brawijaya. All the procedures carried out in this study were approved by the Health Research Ethics Commission, Faculty of Medicine, Brawijaya University Malang (No.50/EC/KEPK-S3/02/2019).\n\nWe obtained approximately 18-20 hippocampal slices per rat and cultured them to make the appropriate organotypic hippocampal slice culture (OHSC; made at the Biosciences Institute of Universitas Brawijaya). The OHSC was divided into two groups (allocated randomly), namely the untreated group and the kainate group. The kainate group was treated with kainic acid (Abcam, AB120100) at a dose of 8 μM by bath application for 48 h and put in an incubator at 37 oC. After that, both groups (kainate group and untreated group) were stained for calcium, MAP2, and GFAP (each subgroup consist of 9 slices) and observed with confocal laser scanning microscopy (CLSM). Sliced culture photographs were taken from a stereomicroscope observation at 40× magnification. The photos were processed with Adobe Photoshop ver. 2020. On day in vitro 13 (DIV 13), some slices were not very healthy macroscopically, so we exclude some slices for statistical analysis. Therefore, we used 5, 3 and 3 slices for calcium, MAP2, and GFAP, respectively. Parameter observations with CLSM were done in the Central Laboratory of Biological Sciences Universitas Brawijaya.\n\nThe slicing medium and the culture medium were prepared before surgery. More specifically, the slicing medium was prepared using 71.49 g of HEPES (Sigma, H3375, Lot SLBB55267) in 300 ml of Earle’s Balanced Salt Solution (EBSS) 10X (Sigma, Lot RNBG8086). The culture medium comprised 50 ml of Minimum Eagle Medium (MEM, Sigma, M0769-10x1L, Lot SLBS6945), 18 ml of EBSS, 5 ml of EBSS + D-glucose (Merck, CAS-No: 50-99), 1 ml of penicillin-streptomycin (Gibco, Ref 15070-063, Lot 2145472), 25 ml of horse serum (Gibco, Ref 16050-122, Lot 2208948), and 0.06 ml of nystatin (Abcam, ab141118, Lot GR305852-5).\n\nWe used P10 Wistar rat to make OHSC. The pups remained with the mother, mostly in a dark environment at a temperature of 26oC with 70% relative humidity. On P10 we took the pups for euthanasia. The OHSC was made according to the method proposed by de Simoni (Simoni & Yu, 2006), albeit with slight modifications (Opitz-Araya & Barria, 2010). Briefly, cervical dislocation followed by decapitation were performed aseptically and without any prior anesthesia (Simoni & Yu, 2006). Cervical dislocation is appropriate and acceptable method of euthanasia for small rats weighing <200 grams. Cervical dislocation causes rapid loss of consciousness, and we performed decapitation immediately after cervical dislocation. Additionally the procedure was performed by a skilled technician and we require uncontaminated brain tissue since anesthetic drug can interfere brain properties (Hollands, 1986; Karmarkar et al., 2010; Leary et al., 2020). The brain was removed from the skull and then soaked for 1 minute in a beaker placed on ice. The beaker contained a slicing medium that had been perfused with 5% CO2/95% O2 for 10 – 20 minutes. Next, the brain was transferred to a petri dish containing the slicing medium (2.4 g HEPES and 10 ml EBSS) and the hippocampus isolation was performed. Following the isolation, the next step involved making slices with a thickness of 350 μm using a tissue slicer (Stoelting Tissue Slicer 51425, RRID:SCR_022902). The hippocampal slices were then placed on an omnipore membrane (Merck, Ref: JHWP01300, Lot: R0MB18744) within a cell culture insert (Merck, Ref: PICM03050, Lot: R0DB59407) and a six-well plate filled with 1.3 ml of the culture medium (EBSS, glucose, penicillin, horse serum, Na bicarbonate, amphotericin, L-glutamine, EBSS, and MEM). All slices that were ready for culturing were put into an incubator at 37 oC for 10 days. The culture medium was changed every two days.\n\nThe full protocol for this procedure can be found here: http://dx.doi.org/10.17504/protocols.io.81wgby6pnvpk/v1.\n\nThe staining of microtubule-associated protein-2 (MAP2 ) (GeneTex Cat# GTX50810, RRID: AB 11170769) and glial fibrillary acidic protein (GFAP) (Santa Cruz Biotechnology Cat# sc-33673, RRID:AB_627673) were done at DIV 10 via the following procedure. Tissue slices were fixed using 4% paraformaldehyde (PFA) for 15 min, then washed with phosphate buffered saline (PBS) for three times (5 minutes each). After the PBS was removed, 0.1% Triton X-100 was added for 30 min, and the slice culture was washed with PBS for three times (5 minutes each). After washing, 10% bovine serum albumin (BSA) was added for 30 min at room temperature. The primary antibody incubation MAP2 (1: 500) and GFAP (1: 500) was performed overnight at 4°C, then washed with PBS for three times (5 minutes each). An incubation using a secondary antibody, Goat anti-Rabbit IgG-F (1: 500) for MAP2 (Santa Cruz Biotechnology Cat# sc-53805, RRID:AB_783978) and anti-mouse Rhodamine (1: 500) for GFAP (Rockland Cat# 610-1002, RRID:AB_219636) for 1 h at room temperature. It was subsequently washed with PBS for three times (5 minutes each), and the sample was ready to be observed using CLSM with a magnification of 100× and a wavelength of 488 nm (MAP2) and 543 nm (GFAP).\n\nThe full protocol for this procedure can be found here: http://dx.doi.org/10.17504/protocols.io.q26g7y4b1gwz/v1\n\nFluo-4 was used for intracellular calcium tracking (Abcam Fluo-4 assay unit (calcium), AB228555). Nine mL of HHBS + 1 mL 10X F127 were mixed into a 15 mL conical tube, 20 μL of Fluo-4 was added. It was mixed in a dark room until homogeneous. Next, 100 μL of Fluo-4 AM dye-loading solution was added in a petri cell dish, followed by incubation in a cell incubator for 1 h. The observation of calcium fluorescence used CLSM at 100× magnification with the fluorescence intensity monitor at maximum Ex/Em = 490/525 nm.\n\nThe full protocol for this procedure can be found here: http://dx.doi.org/10.17504/protocols.io.kxygx9zewg8j/v1.\n\nThe data normality test was performed using the Shapiro-Wilk test. Normally distributed data in both groups were compared using the independent sample t-test. Significant differences in the data were indicated by a p < 0.05. Statistical analysis was performed with SPSS for Windows 25.0 (SCR_016479).\n\n\nResults\n\nExamples of the organotypic hippocampal slice culture results using a modified method from de Simoni (Simoni & Yu, 2006) and Opitz-Araya (Opitz-Araya & Barria, 2010) can be seen in Figure 1. Images of several slices of culture were taken at DIV 0, 3, and 7. On DIV 0, slices were seen dark-colored, and became more transparent and thinner on the 3rd and 7th day; the hippocampal architecture still visible. The slice thickness at DIV 0 was 350 μm, and the thickness at DIV 13 was about 150-200 μm, estimated by CLSM.\n\nIt appears that the slices are thinner and become transparent, with parts of the hippocampus still visible.\n\nDIV = day in vitro.\n\nCLSM image of MAP2 expression in untreated and treated group shown in Figure 2(a-b), GFAP expression in Figure 3(a-b), and intracellular calcium expression in Figure 4(a-b). The mean intensity of MAP2, GFAP, and intracellular calcium expression in the untreated and kainate groups can be seen in Figures 2-4(c). MAP2 (Figure 2c) and GFAP (Figure 3c) expression decreased significantly in the kainate group compared to the untreated group. There was a significant increase of intracellular calcium expression in treated versus untreated group (Figure 4c).\n\nComparison of MAP2 expression in the untreated group (a) and kainate group (b). c. There was a significant decrease of MAP2 expression in kainate group (p = 0.003 < α).\n\nMAP2 = microtubule-associated protein 2. The denomination of expression intensity uses the arbitrary unit (AU).\n\nComparison of GFAP expression in the untreated group (a) and kainate group (b).\n\nc. There was a significant decrease of GFAP expression in kainate group (p = 0.010 < α).\n\nGFAP = glial fibrillary acidic protein. The denomination of expression intensity uses the arbitrary unit (AU).\n\nComparison of calcium expression intensity in the untreated group (a) and kainate group (b). c. There was a significant increment of intracellular calcium expression in kainate group (p = 0.006 < α). The denomination of expression intensity uses the arbitrary unit (AU).\n\nBased on the results of the Shapiro-Wilk test, it was found that the data on calcium, MAP2, and GFAP intensities in the two observation groups had a p-value more significant than the significance level α = 0.05, which means that all the observation groups proved to be normally distributed.\n\nThe comparison test between the untreated and the kainate group using the independent sample t-test is shown in the Table 1.\n\nMean of MAP2 intensity in the kainate group was significantly lower than the untreated group (379579 ± 27334 AU vs 163941 ± 51296 AU; p = 0.003 < α). Likewise, we found significant difference in GFAP expression (p = 0.010 < α) between untreated group (366242 ± 19028 AU) and kainate group (177850 ± 67950 AU).\n\nThere was a significant difference (p = 0.006 < α) of intracellular calcium expression in kainate group (412.157 ± 113.383 AU) compared to untreated group (194.744 ± 69.754 AU). This means that the treated culture exposed to 8 μM of kainic acid for 48 h shows an increased intracellular calcium.\n\n\nDiscussion\n\nThe OHSC protocol according to de Simoni, has been widely used (Abutbul et al., 2012; Kang et al., 2010; Stephen et al., 2015), as well as the method of Opitz-Araya (McQuate & Barria, 2020; Park et al., 2020; Yang et al., 2020). The two methods are mainly similar, however, Opitz-Araya immersed the brain in the slicing medium with perfusion before performing hippocampus isolation (Opitz-Araya & Barria, 2010). Low temperature is required to decrease metabolic activity (Eguchi et al., 2020), glutamate release, excitotoxicity, and purify the blood to aid in the visualization of the cell in slices. Perfusion of the slicing medium with a mixture of 95% O2 gas and 5% CO2 aimed to reduce the hypoxic-ischemic damage to the brain that can occur between decapitation time and the brain's placement in the medium (Varela et al., 2012). Combining the two methods produced the desired results, and the culture was viable until the 10th day. The culture slices were thinner and more transparent compared to day 0. This is an indicator of culture health macroscopically (Opitz-Araya & Barria, 2010; Simoni & Yu, 2006).\n\nThe three main findings of this study were significant decrease of MAP2 and GFAP expression, which are indicators of neuronal and astrocyte injury in the CA3 area, accompanied by a significant increase in intracellular calcium expression at the same time, that is, after 48 h of exposure to kainic acid. MAP2 is a cytoskeletal protein located in the perikaryal and dendrites that specific for neurons, functions for microtubule stabilization, and participates in the dendrite branching processes during the development of hippocampal neurons. This molecule is thought to play a role in synaptic plasticity mechanisms (Kim et al., 2020; Yildirim et al., 2019), which also occurs in epilepsy (Pitkänen & Lukasiuk, 2011). Expression of the MAP2 protein is primarily regulated during brain development and maturation (Blümcke et al., 2001). These proteins are frequently used as markers of dendrites and neuron maturity and integrity (Melani et al., 2005; Pitkänen & Lukasiuk, 2011). The decrease in expression of MAP2 in this study is appropriate with prior studies on OHSC (Holopainen et al., 2004; Hoskison et al., 2007; Noraberg et al., 1999). One study reported neuronal death in CA3 areas, especially after 12, 24, and 48 hours of 5mM KA exposure. The highest neuron mortality was found on exposure to KA for 48 hours, with necrosis as the underlying death process (Holopainen et al., 2004). Other researchers showed that neurons could still be observed within 3 hours after KA injection into the mice's cortex even though they did not appear healthy morphologically. On day 7, neurons disappeared in the entire hemisphere, indicating delayed neuronal death, supporting the apoptotic process (Jeong et al., 2014). Hoskinson et al. demonstrated that loss of MAP2 dendrite in CA1 areas is an indicator of excitotoxic damage after neuronal injury, both in vivo and in vitro, this phenomenon is dependent on Ca2+ permanently (Hoskison et al., 2007). Lopim et al. found that the number of hippocampal neurons in epileptic mice decreased compared to normal mice and that the total number of neurons was negatively correlated with the frequency of epileptic seizures. The number of cell deaths occurred in the initial period after the seizure onset (Lopim et al., 2016).\n\nAstrocytes secrete several inhibitory factors in conditions of injury, ischemia, blood-brain barrier damage, inflammatory reactions, abnormal metabolism, and oxidative stress (Zhang et al., 2017). The active inflammatory status in astrocytes is characterized by the upregulation of GFAP (Siracusa et al., 2019). These external factors influence axon regeneration and recovery of CNS neurological function. GFAP is one of the best markers for astrocyte activation due to injury or stress to the CNS (Zhang et al., 2017). During development, astrocytes also play a role in synaptic transmission, differentiation and migration of neurons, and axon growth. GFAP is an intermediate protein filament expressed in mature astrocytes. This protein is vital for adaptation to neuronal changes and activity during brain development. GFAP also plays a role in neural-glial interactions. Changes in GFAP levels can cause damage to the relationship between neurons and between neurons and glial cells (Yildirim et al., 2019). Jeong et al. found that astrocytes swelled within 3 hours of KA injection into the mice cortex. After that, GFAP+ astrocytes began to disappear from day one and increased until day 7. The location of the loss of astrocytes is the same as delayed neuronal death, thus supporting the suggestion that astrocytes may play a role in the onset of delayed neuronal death (Jeong et al., 2014; De Pablo et al., 2013). The decreased expression of MAP2 after exposure to kainic acid in this study indicated damage to neurons due to administration of kainic acid.\n\nThis study found that kainic acid caused early damage to neurons and astrocytes, particularly at the onset of exposure. Astrocyte activation, characterized by increased GFAP expression, inhibits the inflammatory response after injury, limiting cell damage (Zhang et al., 2017). Decreased astrocyte activation in GFAP-/-Vim-/- mice with an ischemic stroke resulted in larger infarct size. This implies that astrocyte activation is essential for protecting brain tissue in stroke and that GFAP plays a role in this process, especially in the cell resistance to oxidative stress (De Pablo et al., 2013). In this study, the decreased expression of GFAP after exposure to kainic acid indicated damage to astrocytes due to administration of kainic acid.\n\nAs previously mentioned, kainic acid causes excessive activation of glutamate receptors, leading to excitotoxicity (Cho, 2013; Mohd Sairazi et al., 2015; Sanon et al., 2005). Vargas et al. reported that ionotropic glutamate receptors expression was common in young mice's slice cultures (mean age P10). In the adult hippocampus, astrocytes expressed KAR (kainate receptor) after the status epilepticus induced by kainic acid, especially GluK1. This increase is thought to be related to astrocytes' function as glutamate sensors after status epilepticus, as an attempt to save neurons from injury. Besides, KAR expression is also thought to increase calcium signal and hyperexcitability, which causes epileptic seizures (Vargas et al., 2013). A study reported a loss of GFAP immunoreactivity within 24 hours of intracortical injection of kainic acid in mice. This loss of GFAP precedes neuronal death (Jeong et al., 2014). Loss of GFAP immunoreactivity at the onset of traumatic brain injury has also been reported (Zhao et al., 2003).\n\nThe role of astrocytes in changes in neuronal activity is found in epilepsy. Astrocytes can trigger the induction and development of inflammatory status via Ca2+ signaling and are closely related to disease grading. Ca2+ variations that affect neuronal activity in releasing gliotransmitters are also one of the astrocytes' roles. The glutamate transporter is found in several neuron cell types, and astrocytes primarily play a role in glutamate uptake. GLT-1 is a glutamate transporter found in astrocytes, which plays a role in removing extracellular glutamate and increasing epileptogenic foci levels. Glutamine synthetase was reduced in the hippocampus of TLE patients compared to healthy patients. This downregulation causes glutamate-glutamine circulation and increases the accumulation of transmitters in the extracellular space and astrocytes. This indicates a condition of hyperexcitability that depends on astrocytes. AMPA receptors, especially the subtypes formed by the GLuR1 to GluR4 sub-units, are highly expressed in astrocytes. Epilepsy patients show increased expression of the GluR1 variant, which causes prolonged receptor opening, thereby increasing the influx of Na+ and Ca2+ ions, blocking astroglia Kir channels, increasing depolarization, and reducing the capacity of astrocytes in the K+ ion buffer (Siracusa et al., 2019).\n\nThe third finding in this study was an increase in intracellular calcium expression after exposure to kainic acid. OHSC is an in vitro preparation used to study intact neural circuits in chronic conditions, including epileptogenesis. After tissue incision makes deafferentation, the neural tissue rearranges itself and exhibits epileptiform-like activity, which can be visualized by a calcium indicator. This calcium activity is present in both astrocytes and neurons (Gee et al., 2015). Damage to the CA3 area is due to direct stimulation of the kainate receptors, increased glutamate efflux (Zhang & Zhu, 2011), and increased calcium (Borges et al., 2006). KA binding to its receptors increases cell depolarization, one of the main phenomena in temporal lobe epilepsy. Epileptogenesis is associated with increased glutamate neurotransmission. Glutamate triggers an increase in cytoplasmic free Ca2+, which will increase the release of glutamate from neurons and astrocytes (Zhang et al., 2019). Intracellular calcium plays an essential role in regulating cell function (Kurniawan, 2015). The intracellular accumulation of Ca2+ also triggers a molecular cascade involving several intracellular messenger systems, which results in neuronal death. The amplitude of calcium transient in CA1, CA3, and dentate gyrus (DG) after KA administration was significantly higher than before KA administration. Kainic acid receptors were widely expressed in the hippocampus, presynaptically and postsynaptically. The KA1 sub-unit mostly found in the CA3 area and a little in the CA1 area, while the KA2 sub-unit is expressed equally in both the CA1 and CA3 areas (Zhang et al., 2019). Excitotoxicity is the leading cause of cell death, in this case, due to neurotransmission disturbances caused by excessive glutamate. Consequently, this triggers an increase in calcium and causes cell death and neuronal degeneration (Ghibelli et al., 2010). The increase in intracellular calcium influx and oxidative stress due to excessive activation of glutamate receptors by KA causes cell damage and increases in protease and endonuclease enzymes that damage membranes, cytoskeletal proteins, and DNA fragmentation. This oxidative stress plays a role in the death of neurons and glial cells (Mohd Sairazi et al., 2015). The significant increase in intracellular calcium expression in the kainate group implies that calcium plays a significant role in neuron and astrocyte damage due to kainic acid administration.\n\n\nConclusions\n\nThe reduction of MAP2 and GFAP expression in this study indicates the loss of neurons and also astrocytes at the initial onset of exposure to kainic acid, which can be analogous to the deterioration of neuron and astrocyte function at the onset of insult leading to epilepsy. This damage is thought to be related to high levels of calcium (Sattler & Tymianski, 2001). Further analysis of the development of neurons, astrocytes, and calcium was not carried out in this study. Therefore, this study's results can still be developed to further determine the course of neurons, astrocytes, and calcium in the epileptogenesis process and to find strategies to minimize excitotoxicity.", "appendix": "Data availability\n\nFigshare: Expression of MAP2, GFAP, and Calcium in CA3 Area of a Modified Organotypic Hippocampal Slice Culture Treated with Kainic Acid, https://doi.org/10.6084/m9.figshare.21367596.v2.\n\nThis project contains the following underlying data:\n\n1. UNT CA SLICE B (.png) = Calcium Intensity Untreated Group\n\n2. UNT CA SLICE C (.png) = Calcium Intensity Untreated Group\n\n3. UNT GFAP SLICE B (.png) = GFAP Expression Untreated Group\n\n4. UNT MAP 2 SLCIE B (.png) = MAP 2 Expression Untreated Group\n\n5. AK8 CA SLICE B (.png) = Calcium Intensity Kainate Group\n\n6. AK8 CA SLICE C (.png) = Calcium Intensity Kainate Group\n\n7. AK8 GFAP SLICE B (.png) = GFAP Expression Kainate Group\n\n8. AK8 GFAP SLICE C (.png) = GFAP Expression Kainate Group\n\n9. AK8 MAP 2 SLICE B (.png) = MAP 2 Expression Kainate Group\n\n10. AK8 MAP 2 SLICE C (.png) = MAP 2 Expression Kainate Group\n\n11. MAP 2Untreated (.tif ) = MAP 2 Expression Untreated Group\n\n12. MAP 2Kainate (.tif ) = MAP 2 Expression Kainate Group\n\n13. GFAPKainate (.tif ) = GFAP Expression Kainate Group\n\n14. GFAPUntreated (.tif ) = GFAP Expression Untreated Group\n\n15. CalciumKainate (.tif ) = Calcium Intensity Kainate Group\n\n16. CalciumUntreated (.tif ) = Calcium Intensity Untreated Group\n\n17. MAP 2Kainate300 (.tif ) = MAP 2 Expression Kainate Group (300 dpi)\n\n18. MAP 2Untreated300 (.tif ) = MAP 2 Expression Untreated Group (300 dpi)\n\n19. GFAPKainate300 (.tif ) = GFAP Expression Kainate Group (300 dpi)\n\n20. GFAPUntreated300 (.tif ) = GFAP Expression Untreated Group (300 dpi)\n\n21. CalciumKainate300 (.tif ) = Calcium Intensity Kainate Group (300 dpi)\n\n22. CalciumUntreated300 (.tif ) = Calcium Intensity Untreated Group (300 dpi)\n\n23. DIV0-1 (.tif ) = Hippocampal Slice Culture Day in vitro 0\n\n24. DIV0-2 (.tif ) = Hippocampal Slice Culture Day in vitro 0\n\n25. DIV0-3 (.tif ) = Hippocampal Slice Culture Day in vitro 0\n\n26. DIV0-4 (.tif ) = Hippocampal Slice Culture Day in vitro 0\n\n27. DIV0-1300 (.tif ) = Hippocampal Slice Culture Day in vitro 0 (300 dpi)\n\n28. DIV0-2300 (.tif ) = Hippocampal Slice Culture Day in vitro 0 (300 dpi)\n\n29. DIV0-3300 (.tif ) = Hippocampal Slice Culture Day in vitro 0 (300 dpi)\n\n30. DIV0-4300 (.tif ) = Hippocampal Slice Culture Day in vitro 0 (300 dpi)\n\n31. DIV3-1 (.tif ) = Hippocampal Slice Culture Day in vitro 3\n\n32. DIV3-2 (.tif ) = Hippocampal Slice Culture Day in vitro 3\n\n33. DIV3-3 (.tif ) = Hippocampal Slice Culture Day in vitro 3\n\n34. DIV3-4 (.tif ) = Hippocampal Slice Culture Day in vitro 3\n\n35. DIV3-1300 (.tif ) = Hippocampal Slice Culture Day in vitro 3 (300 dpi)\n\n36. DIV3-2300 (.tif ) = Hippocampal Slice Culture Day in vitro 3 (300 dpi)\n\n37. DIV3-3300 (.tif ) = Hippocampal Slice Culture Day in vitro 3 (300 dpi)\n\n38. DIV3-4300 (.tif ) = Hippocampal Slice Culture Day in vitro 3 (300 dpi)\n\n39. DIV7-1 (.tif ) = Hippocampal Slice Culture Day in vitro 7\n\n40. DIV7-2 (.tif ) = Hippocampal Slice Culture Day in vitro 7\n\n41. DIV7-3 (.tif ) = Hippocampal Slice Culture Day in vitro 7\n\n42. DIV7-4 (.tif ) = Hippocampal Slice Culture Day in vitro 7\n\n43. DIV7-1300 (.tif ) = Hippocampal Slice Culture Day in vitro 7 (300 dpi)\n\n44. DIV7-2300 (.tif ) = Hippocampal Slice Culture Day in vitro 7 (300 dpi)\n\n45. DIV7-3300 (.tif ) = Hippocampal Slice Culture Day in vitro 7 (300 dpi)\n\n46. DIV7-4300 (.tif ) = Hippocampal Slice Culture Day in vitro 7 (300 dpi)\n\n47. Calcium (.tif ) = Calcium Intensity Graphic\n\n48. GFAP (.tif ) = GFAP Expression Graphic\n\n49. MAP 2 (.tif ) = MAP 2 Expression Graphic\n\n50. Calcium600 (.tif ) = Calcium Intensity Graphic (600 dpi)\n\n51. GFAP600 (.tif ) = GFAP Expression Graphic (600 dpi)\n\n52. MAP 2600 (.tif ) = MAP 2 Expression Graphic (600 dpi)\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgments\n\nSpecial thanks to Shahdevi Nandar Kurniawan, Wahyu Nur Laili Fajri, Pupimadita Tizar Afdora, and Choirunil Chotimah for their assistance during this study.\n\n\nReferences\n\nAbutbul S, Shapiro J, Szaingurten-Solodkin I, et al.: TGF-β signaling through SMAD2/3 induces the quiescent microglial phenotype within the CNS environment. Glia. 2012; 60(7): 1160–1171. PubMed Abstract \| Publisher Full Text\n\nBlümcke I, Becker AJ, Normann S, et al.: Distinct expression pattern of microtubule-associated protein-2 in human oligodendrogliomas and glial precursor cells. J. Neuropathol. Exp. Neurol. 2001; 60(10): 984–993. 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PubMed Abstract \| Publisher Full Text" }	[ { "id": "166936", "date": "14 Apr 2023", "name": "Abdul Gofir", "expertise": [ "Reviewer Expertise Neurodegeneration and neurovascular" ], "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 abstract, methods section, the paper said, “OHSC was obtained from two healthy wild types Wistar aged P10”. However, in the complete version, Methods, an overview of experiment section, first paragraph, the paper stated, “This study used three healthy wild type Wistar rats aged postnatal day 10…”. Which one is correct?\nIntroduction, First Paragraph, 2nd sentences\nThe reference cited for this sentence (Mohd Sairazi) was a paper that explained about kainic acid excitotoxicity for the experimental model, while the sentence in your paper state that acute and chronic neurodegeneration is excitotoxicity without saying kainic acid in this paper. When we opened Mohd Sairazi's paper, their paper cited from Doble, 1999 the same sentence in their introduction paragraph. Please refer to the correct paper.\nIntroduction (General Comment)\nThe introduction section had explained clearly the importance of this study in examining kainic acid effects on the hippocampal area. It also stated clearly that the mechanism of neuronal death is glutamate excitotoxicity. However, there was still a “gap” between glutamate excitotoxicity and kainic acid effects on OHSC. Can you add more information to bridge the gap between these two pieces of information? (example: there is a necessity to make a correct animal model for glutamate excitotoxicity studies, etc)\nMethods, an overview of experiment section, First paragraph, Second sentence. Using Federer’s formula, could you state the number of treatments and the minimum sample size required for this study?\nMethods, Overview of the experiment, first paragraph, first sentence. This study used Wistar rats P10, while in the protocol cited for this study (Simoni & Yu, 2006) and (Opitz-Araya & Barria, 2010) used another rat model. Simoni & Yu used Sprague Dawley Rats P2-3, while Opitz-Araya & Barria used P5-P7. Could you add more information or references on choosing animals for your study?\nMethods, Overview of an experiment, second paragraph, the 7th & 8th sentence. Some slices were excluded because we're not very healthy macroscopically. This sentence was not clear enough to determine which slide was included or excluded; Hence, this exclusion could change the results of the study. Could you add more explanations on how you determined the tissue is not healthy and excluded from the study?\nDiscussion Section, General Comments In this paper, you had explained clearly the comparison with other studies and explained what the mechanism behind your findings is. However, we didn’t find any explanation about strengths and weaknesses from your studies, because it will give further directions for other studies to fill any gaps that haven’t been answered in your study. Please add the strength and weaknesses, and what is the next direction for this topic\nConclusions Usually in conclusions, we state what we found in our study without any citation. The second sentence showed a citation from Sattler & Tymianski, 2001, which had been explained clearly in the discussion section, the 8th paragraph. Could you revise the conclusion section only include the conclusion from your 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? Partly", "responses": [ { "c_id": "9615", "date": "25 May 2023", "name": "Machlusil Husna", "role": "Author Response", "response": "Mr Gofir, Thank you for your correction and suggestions. I would like to respond to your comments for my article Abstract (method section, line 1) the correct number of animal used is three. I will revise it in article. OHSC was obtained from three healthy wild type Wistar ratas aged P10. Healthy…. Introduction : Paragraph 1, line 4 : Sairazi citation. I will also refer to the original paper. Neurodegeneration mainly caused by acute and chronic excitotoxicity. The chronic excitotoxicity process can be found in Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), amyotropic lateral sclerosis (ALS), and temporal lobe epilepsy (TLE) (Doble, 1999; Mohd. Sairazi et al, 2015) “gap” between glutamate excitotoxicity and kainic acid (paragraph 3, line 1) “Kainic acid is commonly used for glutamate excitotoxicity model. Damage due to kainic acid mainly occurs in the hippocampus. Such damage may be caused by the high density of kainic receptors in that area (Mihály, 2019; Zhang&Zhu, 2011). Methods : Overview of experiments, paragraph 1, line 2 “This study used three healthy wild type Wistar rats aged postnatal day 10 (P10), weight range 10-15 grams, regardless of sex. The sample size was decided from Federer’s formula (Ihwah et al., 2018). The number or treatments are two, and the result from the formula is 8, so we used 9 slices for each subgroup.” Overview of experiments, paragraph 2, line 7-8 “….On day in vitro 13 (DIV 13), some slices were not very healthy macroscopically, so we exclude some slices for statistical analysis. The excluded slices conditions are : not getting thinner and transparent, also the color become whitish opaque or more darker” Overview of experiments, paragraph 2, line 7 : I have mentioned the indicators for healthy slices macroscopically in discussion section (first paragraph, last sentence). However, I will add explanation in methods section as you suggested (paragraph 2, line 7). “…The photos were processed with Adobe Photoshop ver.2020. We examined the healthiness of the slices macroscopically, using stereomicroscope. The healty slices will look thinner and more transparent, because the dead cells and tissue debris have disappeared. Another marker is we can see the different region of hippocampus clearly (Simoni & Yu, 2006). “ Organotypic hippocampal slice culture procedure, paragraph 2, line 1: “Age recommendation to make OHSC for rat is P0-15 and P0-5 for mice (Muller, et al, 2001). We used P10 Wistar rat, because technically it is easier for us than younger rats. The brain size was not too small so it was easier to slice the hippocampus.” I will add explanation in discussion section (paragraph 1, line 10, new sentences) : “We could not use the unhealty slices, so we exclude it for statistical analysis. We used only the healthy ones. The minimum requirement for comparation analysis is triplicate (we used 5 for calcium, 3 for MAP2, and 3 for GFAP). The statistical analysis still can be done, and the result was significant, in accordace with the visual image obtained.” Discussion : Strength and weakness : I will add it in the discussion section (paragraph 8, new) so it will be clearer for readers “The strength of this study is the demonstration of early damage of neuron and astrocyte by kainic acid, accompanied with high intracellular calcium expression. Only few prior studies have investigated it together. This early damage could lead to epileptogenesis. However, we only measure the effect of excitotoxicity at one time only. Therefore, this study’s results can still be developed to further determine the course of neurons, astrocytes, as well as calcium in the epileptogenesis process and to find strategies to minimize excitotoxicitiy” Conclusion : I will revise the conclusion as your suggestion The reduction of MAP2 and GFAP expression in this study indicates the loss of neurons and also astrocytes at the initial onset of exposure to kainic acid, which can be analogous to the deterioration of neuron and astrocyte function at the onset of insult leading to epilepsy." } ] } ]	1	https://f1000research.com/articles/12-47
https://f1000research.com/articles/14-19/v1	03 Jan 25	{ "type": "Research Article", "title": "Through restful waters and deep commotion: A study on burnout and health impairment of Italian seafarers from the JD-R model perspective.", "authors": [ "Francesco Buscema", "Lorenzo Cena", "Clarissa Cricenti", "Margherita Zito", "Lara Bertola", "Lara Colombo", "Lorenzo Cena", "Clarissa Cricenti", "Margherita Zito", "Lara Bertola", "Lara Colombo" ], "abstract": "Background The work experience of seafarers differs significantly from other land-based occupations due to several factors, particularly remoteness and the restricted work environment. This study seeks to examine the impact of burnout and health impairment in the maritime industry, using the Job Demand-Resources theory as a framework.\n\nMethods To investigate these phenomena, an online questionnaire was sent to 239 Italian seafarers (94.6% men, Mage = 39.44, SD = 12.8). We tested a mediated Structural Equation Model (SEM) aimed at predicting negative health outcomes.\n\nResults The results show that burnout plays a mediating role between job demands (such as workload and cognitive strain) and resources (such as social support and transformational leadership), influencing variables related to health impairment (such as sleep quality and physical well-being). Furthermore, the findings highlight the direct influence of occupational resources on seafarers’ health.\n\nConclusions The discussion highlights the urgent need for more research in the field of organisational psychology in the maritime industry and the discrepancies between these findings, which are consistent with the existing maritime literature, and other studies that do not include seafarers in their sample groups.", "keywords": [ "Burnout", "Seafarers", "Health Impairment", "JD-R Model", "Quality of Sleep" ], "content": "Literature review\n\nSeafaring is a demanding profession both mentally and physically. The physical risks associated with maritime occupations, such as musculoskeletal problems, have been widely studied in the literature (Hansen et al., 2008; Remmen et al., 2023; van de Wijdeven et al., 2023), while psychosocial risk factors and dimensions of mental health have only been addressed in recent years, with a focus on specific issues such as burnout (Chung et al., 2017; Oldenburg et al., 2010, 2014; Wan et al., 2023), fatigue (Abila & Acejo, 2021; Oldenburg & Jensen, 2019) and sleep quality (Hystad & Eid, 2016; Lützhöft et al., 2011).\n\nSeafarers may develop psychological or post-traumatic symptoms after experiencing a pirate attack during their working hours (Abila & Tang, 2014). A recent report on seafarers’ mental health shows that the prevalence of depression and anxiety affected 28% of the 1262 seafarers surveyed in 2019 (Lefkowitz & Slade, 2019). Some studies show that the factor that has the greatest impact on seafarers’ quality of life on board and mental health is isolation from family (Buscema et al., 2023; Lefkowitz & Slade, 2019).\n\nDuring the COVID-19 pandemic, the maritime sector played a key role in the survival of the global economy. However, seafarers faced problems that affected their mental health, such as the inability to change shifts, shortages of supplies on board, increased workloads and overdue services (Baygi et al., 2021; Brooks & Greenberg, 2022; Pauksztat et al., 2022).\n\nJob Demands-Resources (JD-R) theory is a model of occupational psychology that assumes that the balance between job demands and individual and contextual resources influences work stress and well-being (Bakker et al., 2023). This theory is able to overcome the limitations of the most common theories of work stress, such as the two-factor theory (Herzberg, 1966), the job characteristics theory (Hackman & Oldham, 1976), the job demand-control model (Karasek, 1979), the effort-reward-imbalance model (Siegrist, 1996) and conservation of resources theory (Hobfoll et al., 2018). JD-R theory is general enough to be applied to all jobs and fits the project design as it can explain the job characteristics of seafarers through the categories of job demands and job resources. In addition, JD-R examines the process of health impairment and motivation, personal resources such as self-efficacy, resilience and humour at work, and the role of exhaustion in job performance (Bakker et al., 2023).\n\nFew studies have examined the role of job resources on shipboard mental health using the JD-R model, suggesting that social support (from peers and external support) and the opportunity to go ashore for holidays, as well as access to communication and entertainment, may reduce mental health problems in seafarers (Pauksztat et al., 2022; Tang et al., 2022). Another study examines the role of JD-R among cruise ship crew and shows that the negative effects of job demands on work engagement are mitigated by workers’ individual strategies, such as recovery or work-related effort (Radic et al., 2020). A recent study investigates the moderating role of a fun environment onboard between job demands and turnover intentions. Specifically, the condition of low job demands and high fun environment, turnover intentions are significantly lower than in the low fun environment condition (Gu et al., 2020).\n\nThe activities of seafarers on board could be associated with a hierarchical environment in which the roles of all crew members are not interchangeable. Leaders or supervisors play a crucial role in steering not only the ship but also the crew members. For this reason, leadership styles such as transformational leadership could be very effective on board to prevent safety issues and motivate seafarers (Sandhåland et al., 2017). Transformational leadership can be summarised as a leadership model consisting of four factors: idealised influence, inspirational motivation, intellectual stimulation and individualised consideration (Bass & Riggio, 2006).\n\n\nResearch hypotheses and model construction\n\nAccording to the JD-R model (Bakker et al., 2023), work and cognitive load are considered predictors of stress, burnout and health impairment in the workplace. A recent study confirms the role of high workload as a predictor of burnout in a large sample of Chinese seafarers (Wan et al., 2023). In addition, the role of human factors, such as cognitive load, is considered to be one of the most important dimensions influencing workplace safety, especially in the maritime environment (Seyfzadehdarabad et al., 2023). Fatigue and stress could be considered as consequences of high cognitive load over long periods of time in seafarers (Žagar et al., 2020). Hypothesis 1: Work and cognitive load (job demands) are positively related to health impairments through the mediation of job burnout.\n\nLiving in an isolated environment for so long means that every seaman has to deal with every crew member. The hierarchical way in which seafarers are managed on board is a crucial issue for the quality of life of all seafarers. For this reason, leadership and social support could be considered as such resources according to the JD-R model (Bakker et al., 2023). Transformational and authentic leadership can be considered a resource that promotes the development of psychological capital and creativity according to recent studies (Rego et al., 2012; Yuen et al., 2020). By embodying social support and transformational leadership, crew members should be able to create a positive work environment that acts as a resource against burnout and health impairment (Lucas et al., 2021; Sampson & Zhao, 2003; Wan et al., 2023). Hypothesis 2: Social support and transformational leadership (job resources) are negatively related to health impairments through the mediation of job burnout.\n\nThe model aims to analyse the mediating role of burnout between the JD-R dimensions and health impairments, using years of navigation as a control variable, as shown in Figure 1.\n\nNote. Ovals represent latent variables; square represents manifest variable.\n\n\nMethods\n\nAn online survey was conducted from February to May 2023 using the Qualtrics XM platform. The questionnaire used the JD-R model (Bakker et al., 2023) to investigate psychosocial risks on board. Through a dedicated website and thanks to the support of the unions in disseminating the questionnaire, 629 seafarers from all departments and ranks responded to the questionnaire. Informed written consent was obtained from all participants prior to data collection. After excluding cases with missing values and workers employed in the catering department, the total valid sample for the present analysis was 239 seafarers. The study protocol was approved by the Ethics Committee [Prot. n. 0513027 del 03/10/2022 - UOR: SI000045—Classif. III/11]. Data analysis was conducted using IBM SPSS version 28.0.1.1 and MPlus version 8. Gender The gender distribution shows that 5.4% of the sample is female, reflecting the gender gap in the maritime environment (Baltic and International Maritime Council & International Chamber of Shipping, 2021). The average age was 39.44 years (SD = 12.81 years). All ranks of deck and engine department were included in the study: 64.4% of seafarers were employed in the deck department. 18.8% of the seafarers were Captains, 30.1% were deck officers, 6.3% were deck cadets, 8.8% were deck ratings, 9.2% were Chief engineers, 14.6% were engine officers, 10.9% were engine ratings. 51.9% of the participants had more than ten years’ experience in shipping.\n\nThe means and standard deviations of all the constructs studied are presented in Table 1, while Table 2 presents the correlations between all the constructs studied. We tested our hypotheses using a structural equation model (SEM) with MPlus version 8. We implemented bootstrap method to create 2000 bootstrap samples (Preacher & Hayes, 2008). Estimates of the indirect effect composed of the products of alpha paths (i.e., from job demands and job resources to burnout) and the beta path (i.e., from burnout to health impairment) were estimated with the associated 95% confidence interval (CI). If the CI does not include zero, then we consider the indirect effect to be statistically significant (p < .05).\n\n** p value < 0.01.\n\n* p value < 0.05.\n\nBased on the studies related to the JD-R model, we selected as variables for job demands: Workload and Cognitive Load. Job resource variables were social support and transformational leadership, while the outcome variables were burnout and health impairment. Years of navigation were used as control variables.\n\nJob demands\n\nThe Italian adaptation of the Psychological Workload and Physical Workload subscales of the Job Content Questionnaire (JCQ) were used to examine work and cognitive load (Baldasseroni et al., 2001; Karasek et al., 1998). 6 items were used to examine workload (e.g., “I have too much work to do”) (Cronbach’s α = 0.882), while 4 items examined cognitive load (e.g., “My work requires my constant attention”) (Cronbach’s α = 0.831). All items on workload are rated on a 6-point scale from completely disagree (1) to completely agree (6), while the items on cognitive load are rated on a 5-point scale from completely disagree (1) to completely agree (5).\n\nJob resources\n\nBased on a previous study of the maritime sector, the following scales were selected to examine social support and transformational leadership (Yuen et al., 2020).\n\n5 Items from the Multidimensional Perceived Social Support Scale (MSPSS) (Dambi et al., 2018; Zimet et al., 1988) were used to assess on-board social support (e.g., “I receive emotional help and support I need from my teammates”). The original MSPSS includes not only perceived social support from colleagues, but also from family and friends. Following Yuen et al. (2020), it was decided to use only social support from colleagues. The items are measured on a 5-point rating scale from strongly disagree (1) to strongly agree (5) (Cronbach’s α = 0.916).\n\n5 items from the Multifactor Leadership Questionnaire (Schuckert et al., 2018) were used to explore transformational leadership on board (e.g., “My superior spends time teaching and coaching”). These items relate to: attributed idealised influence, behavioural idealised influence, inspirational motivation, intellectual stimulation and individualised consideration. The scale is measured on a 5-point rating scale from not at all (1) to always (5) (Cronbach’s α = 0.890).\n\nOutcomes\n\nWe used the Maslach Burnout Inventory (Maslach & Leiter, 2016) to measure burnout on board, using 6 items as conducted by Yuen et al. (2020). Burnout is composed of emotional exhaustion (e.g., “I feel emotionally drained from work”), depersonalisation (e.g., “I worry that this job is hardening me emotionally”), and reduced achievement (e.g., “I have accomplished many worthwhile things in this job”). The items are measured on a 5-point rating scale from never (1) to always (5). To increase the reliability of burnout, the item “I have accomplished many worthwhile things in this job” was removed from the analysis Cronbach’s α increases from 0.726 to 0.780.\n\nAccording to the JD-R theory (Bakker et al., 2023) and following job design of Wan et al. (2023), health impairment was measured by two different indicators: musculoskeletal pain and bad quality of sleep. Musculoskeletal pain was measured using 8 items from the psycho-physical symptoms indicator (e.g., “How often did you suffer the following symptoms: Muscle and joint pain”) by Avallone and Paplomatas (2004) (Cronbach’s α = 0.867). Items are measured on a 5-point rating scale from never (1) to always (5). The Italian adaptation of the Mini Sleep Questionnaire (Natale et al., 2014) was used to assess the quality of sleep on board. This scale consists of 10 items (e.g., “I have had problems sleeping”) measured on a 7-point rating scale from never (1) to always (7) (Cronbach’s α = 0.894).\n\n\nResults\n\nThe model showed good fit indices CFI = 0.909, TLI = 0.903, χ2 (883, N =239) = 1466.436, p < .0001, SRMR = 0.071, RMSEA = 0.053 (C.I. = .048 - .057), according to Hayduk et al. (2007) and Kline (2005). CFI and TLI have values that are above the cut-off (0.90), while SRMR and RMSEA are acceptable as the values are below the cut-off (.08 and .05 respectively), indicating that the model is a good fit (Hu & Bentler, 1999).\n\nIn our measurement model, we used our items to model the constructs as latent variables using CFA. All factor loadings were significant (less than p < .001) and are listed in Table 3.\n\nThe results of the path coefficients of the model were mostly significant at p < 0.001, with the exception of the direct effect of job demands on health impairment and the effect of the control variable (years of navigation) on health impairment ( Table 4).\n\nWe analysed the standardised total and specific indirect effects. The results ( Figure 2) show that the C.I. for the two indirect effects do not include zero, confirming both hypotheses of mediation at a significant level (p < .05). First, burnout fully mediates the relationship between job demands and health impairment (Standardised Total Indirect Effect = 0.443, p < .0001, C.I. = 0.259 to 0.682), as the specific direct effect of job demands on health impairment is not significant (p = .69). Second, burnout partially mediates the relationship between job resources and health impairment (Standardised Total Indirect Effect = -0.249, p < .0001, C.I. = -0.398 to -0.143), because the direct effect of job resources on health impairment is still significant when the indirect effect of mediation is taken into account (estimate = 0.217, p = .018).\n\nNote. Dotted lines represent not significant relationships. Ovals represent latent variables; square represents manifest variable. Significance is represented as follow: p < 0.05; *p < 0.001.\n\nOur model explains 77.4% of the variance in health impairment. We confirmed the full mediation of hypothesis 1 and partially confirmed hypothesis 2 because the mediation is not full.\n\n\nDiscussion\n\nThe aim of our study was to understand the relationship between burnout, JD-R variables and health impairment in the maritime industry. Our findings, which follow recent studies on these topics (Bakker et al., 2023; Yang & Hayes, 2020), confirm that burnout plays a crucial role in predicting health impairments in the seafarer population as well. We have gained valuable insights into the dynamics at play in this particular work environment.\n\nThe model fits the data well and explains much of the variance in our independent variables. Both hypotheses were fully or partially confirmed.\n\nAccording to the JD-R theory, job demands are composed of two dimensions of workload: physical and cognitive. In addition, job demands were expected to have a direct effect on health impairment, as severe job demands have been found to lead to health impairment in the literature (Bakker et al., 2023). According to the process outlined by JD-R theory, frequency and severity of job demands lead to an increase in effort, which depletes workers’ physical and cognitive resources, leading to exhaustion and health impairment. Our findings deepen our knowledge of this process, as in our study severe job demands only lead to health impairments through the mediation of burnout.\n\nThe partial mediation of burnout that we found between job resources and health impairments is also related to recent studies in the maritime industry that have found a direct relationship between job resources and health impairments (Lucas et al., 2021; Sampson & Zhao, 2003; Wan et al., 2023). This finding suggests that in the maritime industry, job resources such as social support and transformational leadership have a direct influence on the prevention of health impairments, while in other occupations this relationship was hardly observed (Alarcon, 2011; Mayerl et al., 2016).\n\nOur study has shown that burnout is a crucial variable in mediating between job demands and resources and negative health outcomes. More specifically, this study has identified two different ways in which job resources predict health impairment. First, the direct effect of resources on reducing health impairment, and second, a mediated effect via burnout. For example, a scenario where a group of seafarers with a transformative leadership style on board and a supportive work environment between all crew members could limit the risk of an increase in health impairment both directly and indirectly by reducing negative psychological experiences such as burnout. In this way, the support and positive atmosphere created by leadership and crew members act as a buffer against the negative health consequences of the demanding work environment, leading to improved overall seafarer well-being. Identifying and addressing these factors can be critical to developing effective interventions and strategies to promote better occupational health in the maritime industry.\n\n\nConclusion\n\nThe findings highlight the need to further develop research on organisational psychology in the maritime industry. The results show the mediating role of burnout between job demands (workload, cognitive load) and resources (social support, transformational leadership) variables and health impairment variables (bad quality of sleep and health problems). Furthermore, the results highlight the direct role of job resources in preventing health impairment among seafarers. As we discussed in the previous section, our results show the incongruence between the results from a sample of seafarers and other samples from the general worker population.\n\nThere are some limitations to this study. All data were collected using an online self-report questionnaire, which means that participant-reported effects may be biassed, especially for sensitive topics such as health and psychological well-being (Roccato, 2003). The burnout measure in the questionnaire was not developed for diagnostic purposes. In the questionnaire, burnout is measured by exploring symptoms, not in a clinical way (Demerouti et al., 2021). Another limitation related to burnout is the possible overlap with the construct of depression. A recent meta-analysis shows that these two constructs are difficult to distinguish (correlation of r = 0.80) (Bianchi et al., 2021). Moreover, the sample is not representative of the total population of seafarers, as there is no information on the actual number of seafarers in Italy. Furthermore, the particular nature of the maritime population does not allow for a multi-group analysis based on gender. Finally, the data was collected using a cross-sectional design with a questionnaire that could not provide information on the causality of the variables.\n\nLongitudinal studies should be conducted to investigate whether we can assume causality between the variables considered in this study. Other methods, such as qualitative interviews and diary studies, could deepen our knowledge of these issues.\n\nNotwithstanding these limitations, the sample is one of the largest and most diverse in the Italian maritime sector to our knowledge, with an ongoing project to back it up that will lead to subsequent studies with qualitative and longitudinal approaches. In particular, this study presents new findings related to JD-R theory and contributes to the literature to increase knowledge related to maritime psychology. Based on the findings of this study, some practical conclusions can be drawn for the development of interventions that focus on strengthening on-board labour resources, such as social support and transformational leadership training. This study clearly shows that job demands such as work and cognitive load have a direct impact on seafarers’ psychological well-being (burnout), but are not directly related to their physical well-being. Interventions that improve relationships with colleagues and supervisors not only have an impact on reducing burnout, but also improve physical well-being by reducing musculoskeletal pain and improving sleep quality.\n\nThe study protocol was approved by the Ethics Committee [Prot. n. 0513027 del 03/10/2022 - UOR: SI000045—Classif. III/11].\n\nInformed written consent was obtained from all participants prior to data collection. The procedure was approved by the Ethics Committee.", "appendix": "Data availability\n\nDue to the data restriction policy aforementioned, readers and reviewers could contact the corresponding author (Francesco Buscema: francesco.buscema@unito.it) for dataset access.\n\nSupplementary material uploaded in Harvard Dataverse: https://doi.org/10.7910/DVN/RGH7ZO.\n\nThis project contains the following extended data:\n\n• Questionnaire: Survey_EN.pdf\n\n• Informed Consent: Informed Consent.pdf\n\n\nReferences\n\nAbila SS, Tang L: Trauma, post-trauma, and support in the shipping industry: The experience of Filipino seafarers after pirate attacks. Mar. Policy. 2014; 46: 132–136. Publisher Full Text\n\nAbila SS, Acejo IL: Mental health of Filipino seafarers and its implications for seafarers’ education. Int. Marit. Health. 2021; 72(3): 183–192. PubMed Abstract \| Publisher Full Text\n\nAlarcon GM: A meta-analysis of burnout with job demands, resources, and attitudes. J. Vocat. 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Reference Source\n\nLucas D, Jego C, Jensen OC, et al.: Seafarers’ mental health in the COVID-19 era: lost at sea? Int. Marit. Health. 2021; 72(2): 138–141. PubMed Abstract \| Publisher Full Text\n\nLützhöft M, Grech MR, Porathe T: Information Environment, Fatigue, and Culture in the Maritime Domain. Rev. Hum. Factors Ergon. 2011; 7: 280–322. Publisher Full Text Reference Source\n\nMaslach C, Leiter MP: Burnout. Stress: Concepts, Cognition, Emotion, and Behavior. Elsevier; 2016; pp. 351–357. Publisher Full Text\n\nMayerl H, Stolz E, Waxenegger A, et al.: The Role of Personal and Job Resources in the Relationship between Psychosocial Job Demands, Mental Strain, and Health Problems. Front. Psychol. 2016; 7. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNatale V, Fabbri M, Tonetti L, et al.: Psychometric goodness of the Mini Sleep Questionnaire. Psychiatry Clin. Neurosci. 2014; 68(7): 568–573. PubMed Abstract \| Publisher Full Text\n\nOldenburg M, Baur X, Schlaich C: Occupational Risks and Challenges of Seafaring. J. Occup. Health. 2010; 52(5): 249–256. PubMed Abstract \| Publisher Full Text\n\nOldenburg M, Jensen H-J: Maritime welfare facilities - utilization and relevance for the compensation of shipboard stress. J. Occup. Med. Toxicol. 2019; 14(1): 11. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOldenburg M, Rieger J, Sevenich C, et al.: Nautical officers at sea: emergency experience and need for medical training. J. Occup. Med. Toxicol. 2014; 9(1): 19. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPauksztat B: ‘Only work and sleep’: seafarers’ perceptions of job demands of short sea cargo shipping lines and their effects on work and life on board. Marit. Policy Manag. 2017; 44(7): 899–915. Publisher Full Text\n\nPauksztat B, Andrei DM, Grech MR: Effects of the COVID-19 pandemic on the mental health of seafarers: A comparison using matched samples. Saf. Sci. 2022; 146(October 2021): 105542. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPreacher KJ, Hayes AF: Asymptotic and resampling strategies for assessing and comparing indirect effects in multiple mediator models. Behav. Res. Methods. 2008; 40(3): 879–891. PubMed Abstract \| Publisher Full Text\n\nRadic A, Arjona-Fuentes JM, Ariza-Montes A, et al.: Job demands–job resources (JD-R) model, work engagement, and well-being of cruise ship employees. Int. J. Hosp. Manag. 2020; 88: 102518. Publisher Full Text\n\nRego A, Sousa F, Marques C, et al.: Authentic leadership promoting employees’ psychological capital and creativity. J. Bus. Res. 2012; 65(3): 429–437. Publisher Full Text\n\nRemmen LN, Christiansen DH, Herttua K, et al.: The risk of an incident hospital contact with a musculoskeletal disorder in Danish occupational fishers: a register-based study. BMC Musculoskelet. Disord. 2023; 24(1): 168. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRoccato M: Desiderabilità Sociale e Acquiescenza. Alcune Trappole delle Inchieste e dei Sondaggi (Strumenti). LED Edizioni Universitarie; 2003.\n\nSampson H, Zhao M: Multilingual crews: communication and the operation of ships. World Englishes. 2003; 22(1): 31–43. Publisher Full Text\n\nSandal GM, Leon GR, Palinkas L: Human challenges in polar and space environments. Rev. Environ. Sci. Biotechnol. 2006; 5(August 2006): 281–296. Publisher Full Text\n\nSandhåland H, Oltedal HA, Hystad SW, et al.: Effects of leadership style and psychological job demands on situation awareness and the willingness to take a risk: A survey of selected offshore vessels. Saf. Sci. 2017; 93: 178–186. Publisher Full Text\n\nSchuckert M, Kim TT, Paek S, et al.: Motivate to innovate. Int. J. Contemp. Hosp. Manag. 2018; 30(2): 776–796. Publisher Full Text\n\nSeyfzadehdarabad F, Sadeghi-Firoozabadi V, Shokri O, et al.: Cognitive correlates of maritime pilots’ human errors. Saf. Sci. 2023; 165: 106196. Publisher Full Text\n\nSiegrist J: Adverse health effects of high-effort/low-reward conditions. J. Occup. Health Psychol. 1996; 1(1): 27–41. PubMed Abstract \| Publisher Full Text\n\nTang L, Abila S, Kitada M, et al.: Seafarers’ mental health during the COVID-19 pandemic: An examination of current supportive measures and their perceived effectiveness. Mar. Policy. 2022; 145: 105276. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTedesco LMR, Ferrara P, Stromillo L, et al.: Seafarers’ perceptions of job demand: A cross-sectional study. Arch. Environ. Occup. Health. 2018; 73(5): 278–283. PubMed Abstract \| Publisher Full Text\n\nvan de Wijdeven B , Visser B, Daams J, et al.: A first step towards a framework for interventions for individual working practice to prevent work-related musculoskeletal disorders: a scoping review. BMC Musculoskelet. Disord. 2023; 24(1): 87. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWan Z, Liu Y, Jiang Y, et al.: Burnout and work ability: A study on mental health of Chinese seafarers from the job demand resource model perspective. Ocean Coast. Manag. 2023; 237: 106517. Publisher Full Text\n\nYang Y, Hayes JA: Causes and consequences of burnout among mental health professionals: A practice-oriented review of recent empirical literature. Psychotherapy. 2020; 57(3): 426–436. PubMed Abstract \| Publisher Full Text\n\nYuen KF, Bai X, Wang X: Safety behaviour at sea: Policy implications for managing seafarers through positive psychology. Mar. Policy. 2020; 121: 104163. Publisher Full Text\n\nŽagar D, Svetina M, Košir A, et al.: Human factor in navigation: Overview of cognitive load measurement during simulated navigational tasks. J. Mar. Sci. Eng. 2020; 8(10): 1–11. Publisher Full Text\n\nZimet GD, Dahlem NW, Zimet SG, et al.: The Multidimensional Scale of Perceived Social Support. J. Pers. Assess. 1988; 52(1): 30–41. Publisher Full Text" }	[ { "id": "358862", "date": "17 Jan 2025", "name": "Inga Bartuseviciene", "expertise": [ "Reviewer Expertise Maritime Education and training", "well-being and resilience" ], "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 the paper is relevant, because the research provides critical insights into the psychological well-being of Italian seafarers, highlighting the interplay between job demands, burnout, and health impairment, and underscores the urgent need for targeted interventions in the maritime industry to enhance seafarers' quality of life and work conditions. However, in my opinion, some of the issues related to the quality of the manuscript can be improved:\nThe information of the sample of the respondents should be provided consistently: in the Abstract – “online questionnaire was sent to 239 Italian seafarers“; in the methods section - “629 seafarers from all departments and ranks responded to the questionnaire” and “total valid sample for the present analysis was 239 seafarers”. Providing more detailed socio-demographic characteristics of the respondents could be useful. An explanation of the sampling method should be provided. A critical analysis of why the used instruments are the most appropriate for the purpose of the research could be useful. This statement is doubtful: “there is no information on the actual number of seafarers in Italy”, usually maritime administration of the country has information about the number of issued certificates. This might be useful to provide insight about the generalisability of the outcomes at least for the population of seafarers in Italy.\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? Partly\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] }, { "id": "358867", "date": "21 Jan 2025", "name": "Vairavan C", "expertise": [ "Reviewer Expertise Psychological well-being", "burnout", "and health impairment among maritime professionals.The application of the JD-R model to study job demands", "resources", "and mental health.Social structures", "leadership styles", "and challenges in the maritime industry. Wind-assisted propulsion technologies like Flettner Rotors and sails.Biodiesel production using waste cooking oil and coffee husks oil." ], "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 examines burnout and health impairment among Italian seafarers using the Job Demands-Resources (JD-R) model. This topic is highly relevant given the unique challenges faced by maritime professionals. The study offers significant contributions by focusing on the underexplored demographic of Italian seafarers and employing a structural equation model to analyze the mediating role of burnout.\nHowever, the manuscript has several areas that require improvement to ensure clarity, consistency, and alignment with the stated objectives. While the findings are meaningful, the presentation of data, methodology, and interpretations can be enhanced to increase the overall impact and reproducibility of the research.\nSpecific Comments:\nAbstract The abstract effectively introduces the study but contains inconsistencies in reporting sample sizes. It mentions 239 respondents, yet the methods section refers to an initial sample of 629. This discrepancy must be clarified to maintain transparency and ensure the validity of the reported findings. Additionally, the abstract lacks key statistical highlights, such as path coefficients or confidence intervals, which would provide a more robust summary of results.\nIntroduction The introduction contextualizes the study well, emphasizing the distinct work conditions of seafarers and the relevance of the JD-R model. However, the transition from general observations to the theoretical framework feels abrupt. A more explicit linkage between the challenges of maritime work and the JD-R framework is needed to strengthen coherence.\nLiterature Review The review comprehensively covers prior studies on seafarers' mental health and the JD-R model. However, it does not critically evaluate gaps in existing research. For instance, while the review mentions studies on burnout and fatigue, it does not explain how the study addresses these gaps in the context of Italian seafarers. Including this analysis would enhance the rationale for the study.\nMethods The methodology is adequately detailed but raises concerns regarding sampling and potential biases. The description of how participants were recruited—\"through a dedicated website and union support\"—requires elaboration. Were participants self-selected, or was random sampling used? This distinction affects the generalizability of the findings. Additionally, the use of a cross-sectional design limits causal interpretations, a limitation that is only briefly acknowledged in the discussion.\nResults The results section is well-organized and presents the data effectively using tables and figures. However, these findings would benefit from additional interpretation within the text. For example, while the structural equation model results are statistically robust, the implications of specific fit indices and path coefficients are not thoroughly explained. Furthermore, some terminology, such as \"C.I.,\" may be unfamiliar to non-specialist readers and should be briefly clarified.\nDiscussion The discussion highlights key findings, particularly the mediating role of burnout between job demands/resources and health impairments. While these findings are significant, their practical applications are underexplored. For example, the authors could elaborate on how transformational leadership training or enhanced social support systems could be implemented aboard ships to mitigate burnout and health impairments.\nAdditionally, the discussion could benefit from a more explicit comparison with studies in other occupational contexts. The current text mentions incongruence with general worker populations but does not provide sufficient examples or interpretations to support this claim.\nConclusion The conclusion effectively summarizes the study’s key findings but is reiterative. It calls for longitudinal studies, which is appropriate, but the authors could also suggest immediate interventions based on their findings, such as policies to reduce cognitive load or improve crew relationships.\nLimitations The limitations section is comprehensive but could further address the overlap between burnout and depression. This overlap is mentioned briefly, but its implications for the study's conclusions are not fully explored. Additionally, the absence of gender-based analysis due to the small number of female participants is a missed opportunity that could have been acknowledged more explicitly.\nReferences The references are extensive and relevant. However, a few citations, such as those related to the JD-R model, are outdated. Incorporating more recent studies would strengthen the theoretical grounding.\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?\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": "358865", "date": "21 Jan 2025", "name": "Andrea Russo", "expertise": [ "Reviewer Expertise Preservation of the mental health of seafarers" ], "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 study is very interesting and makes a valuable contribution to the scientific literature.\nThe number of respondents to whom the questionnaire was sent and the number of those who responded should be clearly stated in the sampling section. In the abstract, only the number of respondents who completed the questionnaire should be mentioned to avoid confusion for the reader.\nRegarding the statement: \"There is no information on the real number of seafarers in Italy,\" a clearer explanation is needed. The concern is how this information is assessed, as seafarers are typically required to obtain certification, and there should be a sailor registry or similar official records.\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/14-19
https://f1000research.com/articles/14-16/v1	03 Jan 25	{ "type": "Systematic Review", "title": "Artificial Intelligence in the Prediction of Stone-Free Status in Urinary Stone Disease Treated with Extracorporeal Shockwave Lithotripsy: A Systematic Review", "authors": [ "Ficky .", "Nur Rasyid", "Widi Atmoko", "Ponco Birowo", "Ficky .", "Widi Atmoko", "Ponco Birowo" ], "abstract": "Background Urolithiasis is one of the most common urological diseases worldwide. One of the most common therapy, extracorporeal shock wave lithotripsy (ESWL), has a high failure rate. The failure rate can be significantly reduced by identifying the candidates most likely to benefit from ESWL, for example, by using machine learning (ML) algorithms. Decision tree analysis (DTA), artificial neural networks (ANN), and random forests (RF) represent a few of the machine learning approaches employed to forecast the stone-free outcome following ESWL.\n\nMethods 219 studies were searched through six electronic databases (CENTRAL, MEDLINE, EMBASE, EBSCO, Proquest, SCOPUS). We employed the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines and adhered to the Standards for Reporting Diagnostic Accuracy Studies (STARD). To evaluate the potential bias in all the studies, we utilized the Quality Assessment of Diagnostic Accuracy Studies (QUADAS-2) tool.\n\nResults 41,484 patients from 11 studies were included. The ML models highlight varying levels of diagnostic precision, with sensitivity spanning from 35-96%, and specificity ranging from 63-98.4%, and area under the curve falling between 0.49-0.96. It is shown in this study that the accuracy of RF and DTA in predicting stone-free status is superior than ANN.\n\nConclusion ML is a comparable predictive method to statistical analysis in predicting stone-free status. Random forest method and DTA are superior MLs compared to ANN. Stone size, density, and 3D texture analysis are the most important variables to be considered in the ML models and should be included in the models to ensure accuracy of stone-free status prediction.", "keywords": [ "artificial intelligence", "stone-free status", "urolithiasis", "extracorporeal shock wave lithotripsy" ], "content": "Introduction\n\nUrolithiasis ranks as the third most prevalent ailment encountered in urological practice, trailing only urinary tract infections and prostate irregularities in terms of occurrence. This condition stands as one of the most widespread urological maladies worldwide, with estimated prevalence rates spanning from 1% to 13% across various regions.1 The global incidence, prevalence, and composition of urinary stones exhibit notable disparities and have undergone transformations over recent decades. Specifically, prevalence rates range from 7% to 13% in North America, 5% to 9% in Europe, and 1% to 5% in Asia.2 This condition’s incidence is reported to range between 5% and 10%, occurring three times more frequently in men than in women. Individuals aged 30 to 50 face a heightened risk of developing urolithiasis, and it’s noteworthy that some patients experience recurrent stone formation. Most stones are naturally expelled through urination, with the duration of this process contingent on the stone’s size and location. Spontaneous passage occurs in approximately 80% of ureteral stones smaller than 5 mm.3\n\nVarious methods are employed to actively remove kidney stones, such as extracorporeal shock wave lithotripsy (ESWL), ureterorenoscopy (URS), retrograde intrarenal surgery (RIRS), percutaneous nephrolithotomy (PNL), and open surgery. While surgical intervention plays a pivotal role in managing urinary stones, it does have certain drawbacks, including a temporary reduction in the patient’s quality of life, prolonged hospitalization, and a high cost burden, thus favoring ESWL as a less invasive alternative.4 The concept of ESWL, involving the application of shock waves to disintegrate stones, was initially introduced in Russia during the 1950s and first implemented in humans in 1980. ESWL is considered the primary choice for addressing urinary stones, particularly those smaller than 2 cm, obviating the need for surgical procedures. Success rates for stones smaller than 2 cm hover around 70-80%. However, despite its prevalence, ESWL does exhibit a reported failure rate ranging from 30% to 89% after the initial session.5 Effective identification of the most suitable candidates for ESWL can substantially curtail this failure rate, thereby optimizing treatment outcomes and judiciously utilizing limited medical resources.3\n\nTo discern the predictive factors affecting ESWL outcomes, numerous studies have concentrated on statistical analyses of patient characteristics employing both bivariate and multivariate approaches. Several factors have emerged as influential in determining ESWL success, including anatomical features of the urinary tract, patient age, stone location, and size.6 However, it’s important to acknowledge that these estimates are derived from statistical models based on cohort data, which possess certain drawbacks. These models may overlook significant factors that fall below an arbitrarily chosen numerical threshold, typically 5%. Moreover, as these models employ dichotomized values, clinicians often encounter challenges when attempting to integrate this prognostic information into routine clinical practice.\n\nIn recent times, machine learning (ML) algorithms, a subset of artificial intelligence, have found increasing utility in the field of medicine. They are being widely applied to enhance the precision of disease diagnosis and prognosis. Essentially, machine learning comprises a set of techniques that enable computers to acquire knowledge from existing data sets, allowing them to make predictions. What makes machine learning particularly powerful is its capacity to rapidly analyze complex combinations of multiple variables. Among these machine learning algorithms, decision tree analysis (DTA) holds several advantages for medical applications. It is known for its simplicity in understanding and interpretation, as well as its suitability for both qualitative and quantitative data, whether textual or numeric.7 Another noteworthy approach is the artificial neural network (ANN), a computational method inspired by biological brains, which approximates how biological neurons tackle problems through interconnected clusters and axons. Each neuron performs a summation function on input values, and the system is self-learning and adaptive, not relying on explicit programming. ANN excels in areas where conventional computer programs struggle to articulate solutions or recognize patterns. Once trained, the network can generate appropriate outputs for given inputs, even when confronted with patterns it has never encountered before. ANN is widely employed as an artificial intelligence technology across various domains.8 Random Forests (RF) consist of an ensemble of multiple decision trees working together to establish output classifications. Numerous decision trees are trained using input data, and rather than relying exclusively on the single best-performing tree, a group of them is utilized collectively. Some of the MLs are illustrated in Figure 1. This approach is akin to a committee making decisions collectively, which often results in greater prediction accuracy compared to a single decision tree.9,10\n\nThe aim of our systematic review is to evaluate the available ML methods capable of predicting the stone-free status in patients with urolithiasis following ESWL.\n\n\nMethods\n\nOur objective is to assess the precision of artificial intelligence or machine learning in forecasting stone-free status in individuals with urolithiasis who have undergone ESWL treatment. Hence, this comprehensive review encompasses investigations that delve into machine learning techniques, including neural networks, to assist in computerized urography for the prediction of stone-free outcomes.\n\nA study search was conducted in six electronic databases (CENTRAL, MEDLINE, EMBASE, EBSCO, Proquest, SCOPUS) on December 12th 2022. PICO was used to make study tracking and finding the relevant literature easier. Specific keywords were used and adapting them as needed for each database ( Table 1). References from relevant systematic reviews were also explored. To be eligible for inclusion in this systematic review, studies were limited to those available in either Indonesian or English.\n\nStudy selection\n\nFor this systematic review, we followed the guidelines provided by the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) statements and the Standards for Reporting Diagnostic Accuracy Studies (STARD). We included randomized controlled trials (RCTs) or cohort studies that investigated the role of machine learning (ML) in predicting the stone-free status following extracorporeal shock wave lithotripsy (ESWL) for urinary system stones. These studies were required to report sensitivity, specificity, accuracy, or ROC (Receiver Operating Characteristic) data. Studies conducted on non-human subjects or those not available in English/Indonesian or without complete full-texts were excluded. Each author (NR, FF, PB, WD) independently assessed the eligibility of studies by reviewing their titles and abstracts and conducting a thorough analysis of the full texts. Any discrepancies among the authors were resolved through discussion.\n\nExtraction of data and outcome of interest\n\nEach author independently collected data using a predefined data extraction form. We gathered information on study characteristics, including patient demographics, sample sizes, and the specific machine learning techniques utilized, along with their diagnostic parameters. In cases of disagreements, consensus was reached through discussion.\n\nThe primary objective of this systematic review is to evaluate the accuracy of AI/machine learning in diagnosing stone-free status. Our main outcome measures of interest are the sensitivity, specificity, and overall accuracy of AI/machine learning when compared to the diagnostic capabilities of urography alone. We employed the Review Manager 5.4 application to gather these outcomes and score the risk of bias in the included literatures.\n\nMethodological assessment\n\nThis systematic review encompasses diagnostic investigations employing both experimental and observational study designs. To gauge the potential bias in all the studies, we employed the Quality Assessment of Diagnostic Accuracy Studies (QUADAS-2) tool.\n\n\nResults\n\nThe initial search across six electronic databases yielded a total of 219 articles, of which 200 were duplicates. After screening the titles and abstracts of the remaining articles, we identified 19 studies that aligned with the criteria set by our systematic review’s PICO framework. However, upon conducting a thorough analysis of the full-text articles, only eleven met our PICO criteria, while the remaining eight studies did not meet the inclusion criteria for this review ( Figure 2).\n\nEleven studies were included in this systematic review. Based on the population of each study, this systematic review involved 41,484 patients. These studies are spread across four continents, Europe, Asia, America, and Africa, most of which are conducted in Asia. The characteristics of the studies are summarized in Table 2.\n\n\n\n• Previous stone events\n\n• Metabolic abnormality (Hypercalciuria, hyperuricosuria)\n\n• Infection\n\n• Stone size\n\n• Directed medical therapy\n\n• Caliectasis/dilated collecting system\n\n• Residual fragments after ESWL\n\n\n\n• Age\n\n• BMI\n\n• No SWL sessions\n\n• SWL intensity\n\n• No of stones\n\n• Stone size\n\n• Stone surface area\n\n• Infundibular length\n\n• Infundibular diameter\n\n• Caliceal pelvic height\n\n• Lower infundibulopelvic angle\n\n• Infundibulouretero-pelvic angle\n\n• Urinary transport type\n\n\n\n• Clinical (Age, Body Habitus)\n\n• 24-hour urinary volume\n\n• Size of stones\n\n• Hydronephrosis\n\n• ESWL (total number of sittings, total number of shocks, mean power, mean frequency)\n\n\n\n• Age\n\n• Sex\n\n• Kidney Side\n\n• Stone Location\n\n• Stent\n\n• Renal Anatomy\n\n• Stone nature\n\n• Stone number\n\n• Stone length\n\n• Stone width\n\n\n\n• The mean of grey level matrix\n\n\n\n• Gender\n\n• No of stone\n\n• Location\n\n• Infundibulopelvic angle (IPA)\n\n• Primary/secondary nature of stone\n\n• Status of hydronephrosis\n\n• Age\n\n• Skin-to-stone distance\n\n• Stone density\n\n• Creatinine level\n\n\n\n• Three-dimensional texture analysis (3D-TA)\n\n\n\n• Age\n\n• Gender\n\n• Location\n\n• Length\n\n• Width\n\n• Stone Volume\n\n• Stone Length\n\n• Skin-to-stone distance\n\n• Stone Density\n\n• Creatinine\n\n• Urine specific gravity\n\n• pH\n\n• Urinalysis microscopic RBC\n\n\n\n• Mean stone density\n\n• Stone volume\n\n• Skin-to-stone distance\n\n• Stone length\n\n• Psoas muscle cross-sectional area\n\n\n\n• Stone length\n\n• Duration of diseased\n\n• Age\n\n• Stone width\n\n• pH\n\n\n\n• Stone location\n\n• Age\n\n• Kidney side\n\n• Electrode used\n\n• Stone treatment number\n\n• Number of shocks\n\n• Area of stone\n\n• Gender\n\n• BMI\n\n• Number of stones\n\n• Family history\n\nThis systematic review analyzes the accuracy of machine learning in predicting a condition; therefore, diagnostic articles are included. We use the Non-Proprietary Quality Assessment of Diagnostic Accuracy Studies (QUADAS-2) tool to assess the quality of the diagnostic studies.18 Generally, the included studies are of moderate to good quality, only three of which have moderate quality. The risk of bias assessment is shown in Figure 3.\n\nOnly two studies reported the use of additional procedures. Yang et al. performed an additional SWL session every one week if stones remained.9 Postoperative stents, catheters, and contrast medium were used by Gomha et al.15 In three articles, the absence of stones on plain abdominal radiographs determined stone-free status.12,13,15 Two studies did not state their sampling method.12,16\n\nA total of 9 MLs are observed in 11 studies, which are ANN, J48 decision tree, k-nearest neighbor (kNN), RF, Sequential Minimal Optimization (SMO), DTA, Extreme Gradient Boosting Trees (XGBoost), Light Gradient Boosting Method (LightGBM), and AdaBoost.7–9 Eight out of 11 studies explored the diagnostic parameters of ANN. Most studies report that ANN has high specificity and high overall accuracy in predicting stone-free status. Four experiments out of 8 studies demonstrated ANN accuracy of at least 90%, with its accuracy ranging from 75-93%. It is followed by DTA with the accuracy of 92% and LightGBM with the accuracy of 88%.\n\n\nDiscussion\n\nThis review employed broad search criteria and inclusive inclusion criteria to ensure the inclusion of all relevant studies. Based on the Area Under the Curve (AUC) values, the ANN model presented by Michaels et al.12 achieved the highest AUC. However, it’s noteworthy that this study utilized plain abdominal radiography as the reference standard instead of computerized urography. Interestingly, when plain abdominal radiography is used as the reference standard, the specificity tends to be higher, particularly when the stone is located outside the kidneys. Decision support systems like ANN serve as computer-generated algorithms aiding healthcare professionals in clinical decision-making. These algorithms, in various forms, aim to replicate the learning process of the human brain. They assist healthcare practitioners in making decisions based on specific clinical data from patients. By employing functions within ANN, computers are trained to predict specific parameters efficiently using training sets. Following training, the computer’s performance is evaluated using the data, assessing the extent of its learning in terms of validity and test data. If the computer demonstrates sufficient learning, it can make predictions for users.3\n\nANN has the potential to greatly enhance the quality of healthcare services, facilitating early disease detection, reducing medical errors, and supporting healthcare authorities in cost-effective patient care. The decision-making process involves selecting one of several alternative outcomes generated by cognitive functions. With an increasing number of alternatives, the complexity of the decision-making process and the potential for errors also rise. At the conclusion of the decision-making process, there is an action or idea. Different formats can be employed to depict how individuals arrive at decisions. It is imperative to scrutinize the interplay between psychological elements, cognitive processes, and the surrounding context when assessing the decision-making process. Individuals are expected to generate specific recommendations through logical filtering and ultimately arrive at the correct decision. Given the value of time in decision-making, providing decision-makers with data as quickly as possible is essential for effective and rapid decision-making. Consequently, many administrative and specialized organizations currently rely on ANN for efficient and swift decision-making.3\n\nMachine learning models exhibit variable diagnostic accuracies, with sensitivity ranging from 35% to 96%, specificity from 63% to 98.4%, and AUC of ROC ranging from 0.49 to 0.96. In this study, it is demonstrated that the predictive accuracy of Random Forest (RF) and Decision Tree Analysis (DTA) in determining stone-free status surpasses that of ANN. It’s worth noting that the high AUC value of ANN was partly attributed to the use of plain abdominal radiography as a reference standard in some studies. Decision tree models present numerous benefits compared to current decision support tools. In contrast to decision tools relying on statistical approaches, artificial intelligence decision models adjust their operations based on the data as they are employed, allowing for the smooth incorporation of new data. Although this adaptability may pose overfitting challenges, setting a minimum number of cases can address this issue. Decision tree models can handle data with both quantitative and qualitative variables, making them versatile. Additionally, results are presented in a straightforward and interpretable manner, often in the form of a tree or a set of rules. Nonetheless, there are certain drawbacks to consider, like attribute importance metrics potentially exhibiting a bias toward variables possessing higher levels of data, including categorical variables with differing value counts.7 RF, XGBoost, and LightGBM are three models based on DTA, explaining their comparable accuracy.9\n\nInformation derived from the analysis of CT images consistently includes crucial predictive factors in the models, primarily mean stone density, stone volume, length, width, and three-dimensional texture analysis. These findings align with current guidelines.19 Specifically, stone size, volume, length, or width are considered essential predictive factors in eight studies, while stone density is emphasized in three studies, and three-dimensional texture analysis is featured in one study. Incorporating these factors into the model enhances the predictive capability of machine learning for stone-free status.\n\nSeveral steps are involved in developing AI models. Some of the programs utilized in these studies include Alyuda NeuroIntelligence 2.2 for creating ANN and Quinlan C.50 for producing DTA.3,7 The algorithm parameters were set to default values to identify factors with the highest predictive accuracy. In the case of DTA, the “minimal cases” parameter defaults to 2 and is used to restrict splits at nodes, leading to the creation of smaller trees as the value increases. By reducing splits at nodes based on this parameter, accuracy gradually decreases, preventing overfitting.7 During the creation of ANN, data were analyzed concerning training, validation, and testing. Both numerical and categorical data were used, and the percentage of data allocated to training, validation, or testing was determined. Subsequently, all data were converted into a numeric format for processing. The ANN structure was then formulated, with the number of neurons determined experimentally, as there are no specific rules in the literature for determining this. The logistic activation function was applied to all inputs and outputs in the ANN model, transforming values into the 0-1 range.3 The RF model, developed by Mannil10 et al., was created using open-source data mining software (WEKA, version 3.8.0; University of Waikato, Hamilton, New Zealand). A built-in feature selection filter was employed to assess individual features’ value and predictive contribution to SWL (Shock Wave Lithotripsy) success, considering both unique predictive value and correlation between features.10\n\nAmong the studies we examined, we noticed various methodological issues, including matters related to reproducibility, the utilization of test datasets, the reporting of diagnostic accuracy, and the absence of non-ML statistical methods for comparison. Concerning reproducibility, although the majority of studies offered extensive model information, only one study allowed access to its model. The limited accessibility to these models poses a challenge as it restricts the capacity to assess these models using different datasets.\n\n\nConclusion\n\nIn conclusion, ML can be used for predicting stone-free status in urinary stone diseases with satisfying accuracy however the accuracy of the prediction rely on many factors such as the ML model, variables taken into account and the data used for training set. The main advantage of ML in the prediction of stone free-status was it allows various factors to be taken into consideration even with the non-linear variables. Random forest method and DTA are superior MLs compared to ANN. Stone size, density, and 3D-texture analysis should be included in the models to ensure accuracy in predicting stone-free status after ESWL. However, access to the models should be made public, and further studies comparing them to the current statistical methods should be conducted.", "appendix": "Data availability\n\nFigshare: Artificial Intelligence or Machine learning for Prediction of stone-free status in Patients with urolithiasis.rm5, http://doi.org/10.6084/m9.figshare.27082798.20\n\nThe project contains the following underlying data:\n\n• Artificial Intelligence or Machine learning for Prediction of stone-free status in Patients with urolithiasis.rm5 (57.68 kB)\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nFigshare: Artificial Intelligence in the Prediction of Stone-Free Status in Urinary Stone Disease Treated with Extracorporeal Shockwave Lithotripsy: A Systematic Review, DOI: http://doi.org/10.6084/m9.figshare.27173805,21 http://doi.org/10.6084/m9.figshare.2731459522\n\nThe project contains the following Reporting guidelines data:\n\n• PRISMA Checklist\n\n• Prisma Flow Chart\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nLang J, Narendrula A, El-Zawahry A, et al.: Global Trends in Incidence and Burden of Urolithiasis from 1990 to 2019: An Analysis of Global Burden of Disease Study Data. Eur. Urol. Open Sci. 2022; 35: 37–46. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLiu Y, Chen Y, Liao B, et al.: Epidemiology of urolithiasis in Asia. Asian J. Urol. 2018; 5(4): 205–214. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSeckiner I, Seckiner S, Sen H, et al.: A neural network - based algorithm for predicting stone - free status after ESWL therapy. Int. Braz. J. Urol. 2017; 43(6): 1110–1114. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSrisubat A, Potisat S, Lojanapiwat B: Extracorporeal shock wave lithotripsy (ESWL) vs. percutaneous nephrolithotomy (PCNL) or retrograde intrarenal surgery (RIRS) for kidney stones. Cochrane Database Syst. Rev. 2014; 24. Publisher Full Text\n\nNakasato T, Morita J, Ogawa Y: Evaluation of Hounsfield Units as a predictive factor for the outcome of extracorporeal shock wave lithotripsy and stone composition. Urolithiasis. 2014; 43: 69–75. Publisher Full Text\n\nYamashita S, Kohjimoto Y, Iwahashi Y, et al.: Noncontrast Computed Tomography Parameters for Predicting Shock Wave Lithotripsy Outcome in Upper Urinary Tract Stone Cases. Biomed. Res. Int. 2018; 2018: 9253952.\n\nChoo MS, Uhmn S, Kim JK, et al.: A Prediction Model Using Machine Learning Algorithm for Assessing Stone-Free Status after Single Session Shock Wave Lithotripsy to Treat Ureteral Stones. J. Urol. 2018; 200(6): 1371–1377. PubMed Abstract \| Publisher Full Text\n\nXu ZH, Zhou S, Jia CP, et al.: Prediction of Proximal Ureteral Stones Clearance after Shock Wave Lithotripsy Using an Artificial Neural Network. Urol. J. 2021; 18(5): 491–496. PubMed Abstract \| Publisher Full Text\n\nYang SW, Hyon YK, Na HS, et al.: Machine learning prediction of stone-free success in patients with urinary stone after treatment of shock wave lithotripsy. BMC Urol. 2020; 20(1): 88. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMannil M, von Spiczak J , Hermanns T, et al.: Three-Dimensional Texture Analysis with Machine Learning Provides Incremental Predictive Information for Successful Shock Wave Lithotripsy in Patients with Kidney Stones. J. Urol. 2018; 200(4): 829–836. PubMed Abstract \| Publisher Full Text\n\nSinha Y, Malviya P, Panda M, et al.: Contextual Care Protocol using Neural Networks and Decision Trees.2018; 1–4.\n\nMichaels EK, Niederberger CS, Golden RM, et al.: Use of a neural network to predict stone growth after shock wave lithotripsy. Urology. 1998; 51(2): 335–338. PubMed Abstract \| Publisher Full Text\n\nPoulakis V, Dahm P, Witzsch U, et al.: Prediction of lower pole stone clearance after shock wave lithotripsy using an artificial neural network. J. Urol. 2003; 169(4): 1250–1256. PubMed Abstract \| Publisher Full Text\n\nHamid A, Dwivedi US, Singh TN, et al.: Artificial neural networks in predicting optimum renal stone fragmentation by extracorporeal shock wave lithotripsy: a preliminary study. BJU Int. 2003; 91(9): 821–824. PubMed Abstract \| Publisher Full Text\n\nGomha MA, Sheir KZ, Showky S, et al.: Can we improve the prediction of stone-free status after extracorporeal shock wave lithotripsy for ureteral stones? A neural network or a statistical model? J. Urol. 2004; 172(1): 175–179. Publisher Full Text\n\nMoorthy K, Krishnan M: Prediction of fragmentation of kidney stones: A statistical approach from NCCT images. Can. Urol. Assoc. J. 2016; 10(7-8): E237–E240. PubMed Abstract \| Publisher Full Text\n\nMoghisi R, El Morr C, Pace KT, et al.: A Machine Learning Approach to Predict the Outcome of Urinary Calculi Treatment Using Shock Wave Lithotripsy: Model Development and Validation Study. Interact J. Med. Res. 2022; 11(1): e33357. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWhiting PF, Rutjes AW, Westwood ME, et al.: QUADAS-2: a revised tool for the quality assessment of diagnostic accuracy studies. Ann. Intern. Med. 2011; 155(8): 529–536. Publisher Full Text\n\nTürk C, Petřík A, Sarica K, et al.: EAU Guidelines on Diagnosis and Conservative Management of Urolithiasis. Eur. Urol. 2016; 69(3): 468–474. PubMed Abstract \| Publisher Full Text\n\nFicky F, Rasyid N, Atmoko W, et al.: Artificial Intelligence or Machine learning for Prediction of stone-free status in Patients with urolithiasis.rm5.2024. Reference Source\n\nFicky F: Prisma 2020 Checklist for Artificial intelligence in the prediction of stone-free status in urinary stone disease treated with extracorporeal shockwave lithotripsy: A systematic review. Dataset. figshare. 2024. Publisher Full Text\n\nFicky F: Prisma 2020 Flowchart for Artificial intelligence in the prediction of stone-free status in urinary stone disease treated with extracorporeal shockwave lithotripsy: A systematic review. figshare. Figure. 2024. Publisher Full Text" }	[ { "id": "358874", "date": "24 Jan 2025", "name": "Jethro CC Kwong", "expertise": [ "Reviewer Expertise AI in urology" ], "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. The search was conducted on Dec 12, 2022. I would recommend updating the search to at least Dec 31, 2024 as there may be newer models published in the past 2 years 2. This review can be strengthened by examining the quality of the included ML studies using an AI-specific evaluation tool, such as APPRAISE-AI (Kwong JCC, et al. [2023-Ref -1] https://doi.org/10.1001/jamanetworkopen.2023.35377). It would be worthwhile to use this tool to identify specific methodological and reporting gaps in this specific area of study. 3. Did any of the studies compare the performance of the ML model against a non-ML approach (clinical judgement or other clinical tool)? If yes, what was the margin of difference? If not, this may be worthwhile to highlight as these studies did not demonstrate whether the ML approach outperforms traditional statistical approaches or clinical judgement 4. The authors should provide more detail regarding the stone-free status. From Table 2, it seems that there was heterogeneity in the timepoint at which stone-free status was determined, which can impact comparability of studies. Furthermore, additional details should be provided regarding determination of stone-free status (e.g. imaging modality, does stone-free mean complete absence of stone or was there a size cutoff of residual stone that was allowed). 5. The limitations of the included studies should be expanded. I think use of the APPRAISE-AI tool will help uncover additional limitations that are worth mentioning. This review can be strengthened if the authors can provide a list of recommendations on how future ML studies in this space can be improved, based on the limitations identified in the current review.\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? Partly\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": [] }, { "id": "361858", "date": "30 Jan 2025", "name": "Ranjit Barua", "expertise": [ "Reviewer Expertise Modern Robotics", "MIS", "Bioprinting" ], "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\nEvery scientific study has limitations, and acknowledging them strengthens its credibility. The current study should explicitly discuss potential drawbacks, such as small sample sizes, retrospective data analysis, and lack of external validation. Addressing these limitations would help guide future research in refining ML models for ESWL outcome prediction.\nThe potential for hybrid models should also be explored. Combining multiple ML approaches, such as ensemble learning techniques that integrate RF and ANN, could improve predictive accuracy. Additionally, discussing the need for prospective, multi-center trials to validate ML models in real-world clinical settings would provide a clear roadmap for future research.\nThe study presents valuable insights into the application of ML in predicting ESWL outcomes, but several areas require further refinement to enhance its scientific soundness. Methodological transparency, proper statistical validation, assessment of bias, and clinical applicability must be addressed to ensure the reliability and utility of ML models in urological practice. By implementing these recommendations, the study can contribute more effectively to the ongoing integration of ML into clinical decision-making for urolithiasis treatment.\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/14-16
https://f1000research.com/articles/14-15/v1	03 Jan 25	{ "type": "Research Article", "title": "Concurrent stunting among under-five children in Egypt", "authors": [ "Nagwa Farag Elmighrabi", "Catharine A. K. Fleming", "Kingsley E. Agho", "Catharine A. K. Fleming", "Kingsley E. Agho" ], "abstract": "Background Stunting still represents a major public health problem in Egypt, impacting around one-third of children under five and leading to a 2-3% drop in Gross domestic product (GDP). Therefore, this study examined the trends in the prevalence of stunting and its associated factors among children aged 0-23 months, 24-59 months and 0-59 months in Egypt.\n\nMethods The study used combined data from the 2005, 2008, and 2014 Egypt Demographic and Health Surveys (EDHS), with a total sample of 39,857 children aged 0-59 months. A multilevel logistic regression that adjusts for sampling weights and clustering was performed to analyse the factors associated with stunting among children in Egypt.\n\nResults The prevalence of stunting was 24.8% in children aged 0-23 months, 24.3% in children aged 24-59 months, and 24.5 % in children aged 0-59 months. After controlling for potential confounders, the common factors associated with stunting in three age groups were mothers with primary education and mothers who did not have an antenatal care (ANC) visit during pregnancy. Maternal short stature (height < 155 cm) was associated with an increased risk of stunting in three age groups compared to mothers taller than 160 cm.\n\nConclusion Addressing the causes of stunting in Egypt necessitates sectoral collaboration, including health, water, sanitation and hygiene, education, and social protection, with the implementation of targeted interventions focusing on children born to short mothers. Interventions must be accessible for families with low socioeconomic status, focusing on improved healthcare access, parent education, and infant and young child feeding practices. A systematic nutrition monitoring framework that enables routine data collection on nutritional outcomes to monitor and improve child health is recommended.", "keywords": [ "Undernutrition", "childhood", "0-59 months", "Growth defect", "North Africa" ], "content": "Introduction\n\nUndernutrition is a significant predictor of poor growth and morbidity in infants and young children.1 Globally, stunting (low height for age) affects 150 million children under the age of five, a substantial portion of the world’s young population.1 Previous studies have shown that stunting can result in delayed cognitive development, poor school performance, and ultimately lower future earning potential, perpetuating poverty.2,3 The consequences, of stunting particularly in low- and middle-income countries, are far-reaching, leading to impaired growth, reduced educational attainment, delayed school enrolment, and a 20% reduction in adult income.4–6 Reducing undernutrition among young children is one of the targets of the Sustainable Development Goal aimed at eradicating extreme poverty and zero hunger. It is also an important strategy to reduce child mortality, the fourth SDG.7\n\nIn Egypt, approximately one-third of children under the age of five are affected by stunting.8,9 With one in every five children in Egypt stunted, the country ranks among those with the second highest rates of stunting in the North African region.9–11 In 2013, the economic and social cost of child malnutrition in Egypt was substantial, estimated to be 20.3 billion Egyptian Pounds, equivalent to about 2% of the Egyptian GDP.12 Therefore, addressing undernutrition is important not only for public health, but also for enhancing the country long-term economic growth and social stability. Economic growth is an effective instrument for poverty alleviation and improving public health.13–15\n\nIn response to the need for improved undernutrition in Egypt, the Egyptian government has developed several initiatives and programs aimed at addressing this issue, especially among vulnerable populations such as children and women.16 These initiatives include the National Nutrition Strategy (2017-2030), Food Subsidy Programs, akamol and Karama Programs, the Scaling Up Nutrition (SUN) Movement, Micronutrient Supplementation, and collaborations with international organisations.16 Despite the previous efforts by the Egyptian government, undernutrition still represents a significant problem in Egypt. Challenges are further exacerbated by high unemployment rates, poverty, inequality, civil unrest, and climate change-induced natural disasters, all threatening food security and individuals’ nutritional status.17–20\n\nThere have been several past studies assessing child stunting in Egypt.10,21,22 For instance, one study examined the prevalence and factors associated with stunting among school children (6-24 months) in Minia city in Upper Egypt and found that low birth weight of children, short stature of the mothers (≤160 cm) and low maternal education were associated with stunting21 and similarly, a study conducted in Sohag, Egypt on stunting in schoolchildren found low maternal body mass index and familial short stature was the key drivers of stunting.22 A population based cross-sectional studies conducted in Upper and Lower Egypt found that stunting peaked at 12–23 months.10\n\nHowever, these studies were limited in scope as it covered a smaller section in Egypt, children aged, 6-23 months21 and 4-12 years (school age) in just two schools in one governorate.22 Another past study assessed a cross-country study by Pritchett and Summers13 used time series data and instrumental variables to demonstrate that as national income increases, population health tends to improve, and conversely, wealthier countries tend to have better health outcomes. Despite significant efforts to address the problem of stunting in Egypt, several factors may contribute to the current situation. Factors include the lack of up-to-date nutritional data, inadequate training for healthcare professionals, a focus on treatment over prevention, and limited studies on ecological factors associated with nutritional issues.23\n\nTherefore, this study aimed to investigate stunting among children aged 0-23, 24-59 and 0-59 months in Egypt and explore associated factors by pooling the 2005, 2008, and 2014 EDHS. The study will contribute to the growing body of evidence that informs national government institutions, public health researchers, and policymakers working to address the underlying factors associated with child stunting in Egypt. It stands out from previous research on stunting among children in Egypt due to its unique focus on combining data sets from 2005 to 2014. The study utilises population-based national representative data and provides age-specific results by examining the prevalence of stunting in three age groups: 0-23 months, 24-59 months, and 0-59 months across the three-survey data.\n\n\nMethod\n\nAnalysis utilised data from the EDHS conducted in 2005, 2008, and 2014. The surveys were implemented by El-Zanaty and Associates on behalf of the Ministry of Health and Population and the National Population Council, as part of the global MEASURE DHS project funded by the United States Agency for International Development (USAID). The 2005, 2008, and 2014 EDHS received primary funding from USAID/Cairo, with additional support from UNICEF and the United Nations Population Fund (UNFPA). The datasets were downloaded from www.dhsprogram.com after completing the registration process.\n\nAll statistical analyses were carried out using STATA/MP Ver-sion.17.0 (Stata Corp, College Station, TX, USA), (https://blog.stata.com/2021/04/20/stata-17-released/, It is online statistical analysis software – Not online software – purchase for analysis of my PhD.\n\nAdministratively, Egypt is divided into 26 governorates and Luxor City. The four Urban Governorates (Cairo, Alexandria, Port Said, and Suez) have no rural population. Each of the other 22 governorates is subdivided into urban and rural areas. Nine of these governorates are located in the Nile Delta (Lower Egypt), eight are located in the Nile Valley (Upper Egypt), and the remaining five Frontier Governorates are located on the eastern and western boundaries of Egypt.24\n\nThis study analysed data from the EDHS for 2005 (n=13,599), 2008 (n=10,590), and 2014 (n=15,668), focusing on children aged 0-59 months. The sample design was multistage sampling design enabled the estimation of key population and health indicators, including fertility and mortality rates, for Egypt, six major regions, and individual governorates. The sample size was disproportionately distributed throughout these areas to provide for regional and governorate-level estimates. The EDHS sample was chosen using a three-stage approach. Initially, primary sampling units (PSUs), which included shiakhas/towns in urban areas and villages in rural areas, were chosen from a list drawn from the 1996 census and then updated to August 2004 for the 2005 survey and to 2006 for both the 2008 and 2014 surveys. The units were geographically stratified, with smaller units merging to form larger units with a population of at least 5,000. During the second stage, complete maps of the selected primary sampling units (PSUs) were used to divide them into groups of about 5,000 people. A fast enumeration was used to assess household quantities, followed by a quality assurance verification in 10% of the regions. A systematic random sample of households was eventually selected. The survey covered all ever-married women aged 15 to 49 who lived or were present in the sampling households the night before the interview.\n\nThe outcome variable for this study was stunting, which was determined by the height-for-age Z-score (HAZ). Children whose height-for-age is less than -2 standard deviations from the corresponding reference median (Z score ≤ -2) were categorised as stunted. Stunting is an indicator of linear growth retardation and reflects cumulative growth deficits in a child. The HAZ is calculated based on the 2006 WHO growth reference, which compares a child’s height to the median height of a healthy child in the same age group or reference population.25 It is expressed in terms of the number of standard deviations (SD) above or below the median height.25\n\nThe UNICEF conceptual framework of the determinants of nutritional status for mother and child health26 guided the selection of covariate variables, with adjustments based on a previous study conducted in 35 low- and middle-income countries.27 The covariates were categorised into three major categories.\n\nBasic factors include governance, resources, and norms area of residence (urban/rural), region (Urban governates, Urban (lower Egypt), Rural (Lower Egypt), Urban (Upper Egypt), Rural (Upper Egypt) and Frontier Governates).\n\nUnderlying factors include socioeconomic factors (household wealth quantile, child’s sex, child’s age, maternal age, father age, maternal age at childbirth, age at first cohabitation/married, mother and father level of education, mother work in the last 12 months, father occupation, maternal marital status, religion, household members, number of children < 5 and combined birth rank and birth interval), mother’s health (mother height, maternal body mass index (BMI), maternal autonomy (money, care and movement), health care services factors (combined Place of birth and mode of delivery, delivery assistants, number of ANC visits and postnatal checkup, recent vitamin A supplement, ever had vaccination) and household environmental factors (sources of drinking water, stool disposal, and toilet facility and access to media).\n\nImmediate factors include child health (perceived child size at birth, diarrhoea, fever and cough) and breastfeeding practices (early initiation of breastfeeding and duration of breastfeeding).\n\nMultiple previous studies have consistently highlighted the relationship between these factors and child anthropometric failure. Previous studies have identified a significant association between child anthropometric failures and factors such as household wealth index,11,28–32 child age and gender,11,31,33 maternal nutritional health,11,21,29,31,32,34 maternal reproductive care,28,35 family desired birth spacing,11,31 and household environment28–33\n\nWhen considering household income and expenses, the “Household Wealth Quantile” serves as a quantitative indicator of a household’s financial status. The Principal Components Analysis (PCA) approach establishes the quantitative representation of household assets. A score was assigned to every authorised household member once this calculation was performed, in order to ascertain their position in the population. This study classified wealth into five categories at the national level: poorest, poorer, middle, fourth, and richest. The lowest 40% of households consisted of the poorest and poorest households, followed by the middle-class households comprising the next 20%, and the top 40% constituted the fourth and richest households.27\n\nSome covariate factors (combined Place of birth and mode of delivery, combined birth rank and birth interval) were grouped together due to the significant associations between them, as identified and applied in previous research studies.36,37\n\nIn our analysis, we examined factors associated with stunting from each EDHS 2005,2008 and 2014 surveys to determine the strength of the association of each characteristic in relation to the likelihood of stunting in Egypt. The outcome variable for the final logistic regression models is the presence or absence of stunting (Y = 1 = stunted, 0 = not stunted). For each survey, frequency was used to describe the characteristics of the study population and to estimate the prevalence of stunting. The chi-square test was used for categorical variables. Univariate analyses were used to examine distributions and normality of continuous predictors. Bivariate analyses were performed using Pearson’s chi-square, to test differences in frequencies of categorical variables and t-tests for differences in means of continuous variables. Variables found to be significant at the p < 0.05 were included in the final multivariable analysis models. All statistical analyses were carried out using STATA/MP Version.17.0 (Stata Corp, College Station, TX, USA), (https://blog.stata.com/2021/04/20/stata-17-released/), and adjusted odds ratios (AORs) and their 95% confidence intervals (CIs) obtained from the adjusted multivariate regression were used to measure the factors associated with child stunting. The prevalence of stunting was estimated in each survey. A total prevalence estimate (unweighted by year of the survey) was also generated for each year.\n\n\nResults\n\nThe current study involved 39,857 children aged under five years: 13,599 from the 2005 EDHS, 10,590 from the 2008 EDHS and 15668 from the 2014 EDHS. The basic characteristics of the samples in three surveys are displayed in Table S1 (Extended Data). More than one half of the children in each survey lived in rural areas (64%, 63% and 69% in 2005 EDHS, 2008 EDHS and 2014 EDHS respectively). Across the three surveys, the minority of children were from the poorest households, and the ratio of males to girls was almost the same. More than one half of the mothers were aged between 25 and 34 years, while the majority of fathers were aged 18-34 years across the three surveys. Other variables which were in the majority across the three surveys included: married mothers, parents with secondary education, non-working mothers, fathers working in non-agricultural sector, mothers who had money, movement and healthcare autonomy, who were Muslim, who had BMI > 25 who listened to the radio, watched the television and never read any newspaper or magazine, who attended more than 8 ANC clinics, who were delivered by health professionals, who put their babies to the breast within one hour of delivery and who breastfed their babies for more than 12 months; households with members between 5 and 10, having between 2 and 3 children being aged under 5 years, having access to protected water source, with safe stool disposal and improved toilet facility; and children who were not given vitamin A supplement medication and who had not contracted diarrhoea, fever and cough in the two weeks prior to the survey.\n\nThe difference in the prevalence of stunting among children aged 0-23 months in Egypt, based on surveys conducted in 2005, 2008, and 2014 is illustrated in Figure 1. Compared to 2005, the prevalence of stunting among young children in Egypt has significantly decreased from 27.6% to 20.8% in 2014 with overall prevalence 24.8 %. For older children 24-59 months, the difference in the stunting rates in Egypt is shown in Figure 2. The prevalence of stunting among children aged 24-59 months in Egypt was a slightly decreased from 22.5% in 2005 to 21.9% in 2014, with an overall prevalence of 24.3%. Nonetheless, the occurrence of stunting in the same age group experienced a substantial rise from 22.5% in 2005 to 30.4% in 2008. The variation in stunting rates among children aged 0-59 months in Egypt, as observed in surveys conducted in 2005, 2008, and 2014, are illustrated in Figure 3. The prevalence of child stunting in Egypt slightly declined from 24.6% in 2005 to 21.5% in 2014 but remained high at 24.5% overall. However, the prevalence of stunting among the same age group significantly increased from 24.6% in 2005 to 28.9% in 2008. Stunting is prevalent among children under five in Egypt with 1 in each 4 children expected to be stunted.\n\nTable 1 presents the prevalence of stunting among Egyptian children aged 0-23 months, 24-59 months, and 0-59 months, in relation to the fundamental, underlying, and immediate variables. Stunted children across all three age groups (0-23 months, 24-59 months, and 0-59 months) were more prevalent among those residing in rural upper Egypt, hailing from low-income families, having mothers under 18 years of age, being the fourth child in birth order, having mothers with limited media exposure (minimal listening and viewing), being born at home via non-cesarean delivery, lacking maternal ANC visits, and having mothers who are illiterate. For detailed information regarding the prevalence of stunting within certain age groups (0-23, 24-59, and 0-59 months) based on the three primary categorical factors, refer to supplementary Table S2 (Extended Data).\n\n* P< 0.05;\n\n p<0.01;\n\n* p<0.001\n\nTable S3 (Extended Data) and Table 2 show the univariate and multivariate analyses of stunting determinants among Egyptian children aged 0–23, 24-59, and 0–59 months. For children across the three age groups, the odds of stunting were significantly lower in 2014 compared to 2005, with adjusted odds ratios (aOR) of 0.64 (95% CI: 0.54–0.76) for the 0-23 months group, 0.78 (95% CI: 0.64–1.00) for the 24-59 months group, and 0.71 (95% CI: 0.59–0.84) for the 0-59 months group.\n\nChildren living in rural Upper Egypt and urban Lower Egypt were significantly more likely to be stunted than those from other geographical regions, while those from middle-income households were significantly less likely to be stunted than those from the richest households. For the three age categories, children whose mothers only had primary school were considerably more likely to be stunted than those whose mothers had secondary education or above. For all three age groups, children whose mothers were less than 155 cm tall had a significantly higher risk of stunting than those whose mothers were taller than 160 cm. In all age categories, children whose mothers did not visit any ANC clinic had significantly higher odds of stunting than those whose mothers visited at least 8 ANC clinics. For all three age groups, children whose mothers did not listen to radio had a considerably increased risk of stunting.\n\nAmong children aged 0-23 months, the likelihood of stunting was significantly less among girls than boys. The odds of stunting were significantly higher among children aged 18-23 months compared to those aged 0-5 months. For 0-23 months groups, the likelihood of a child being stunted was significantly higher among those whose mothers were not married compared to those whose mothers were married. For children aged 0-23 months, the risk of stunting was significantly higher among children who were delivered through a non-caesarean section and at home than children who were delivered through a caesarean section at a health facility. Children who received vitamin A supplements were more likely to be stunted than children who did not. Among children in the 0-23 months age group, the likelihood of stunting was significantly higher in those who were breastfed for more than 12 months compared to children who were breastfed for up to 12 months.\n\nAmong children aged 24-59 months, the likelihood of stunting was significantly less among older children. The odds of stunting were significantly lower in children whose fathers were older than 45 years compared to those fathers were aged 18-34 years. Children whose mothers had only primary education were significantly more prone to stunting than those whose mothers were educated to higher levels than secondary. The odds of stunting were significantly higher among those whose fathers were educated to primary level, compared to those whose fathers had higher levels than secondary. Children whose mothers worked were significantly more likely to be stunted than their counterparts whose mothers did not work. children whose mothers had a BMI of more than 25 kg/m2 were significantly less likely to be stunted compared to those whose mothers had a BMI of 18.5 kg/m2 or less. Children who were from households with unprotected source of drinking water were significantly more likely to be stunted compared to their counterparts from households with protected source of drinking water.\n\nAmong children aged 0-59 months, the likelihood of stunting was significantly less among girls than boys. The odds of stunting were significantly higher among children aged 6 months until 41 months compared to other ages. The odds of stunting were significantly higher among those whose fathers were educated to primary level, compared to those whose fathers had higher levels than secondary. Children whose mothers worked were significantly more likely to be stunted than their counterparts whose mothers did not work. For the 24-59 months and the 0-59 months groups, children whose mothers had a BMI of more than 25 kg/m2 were significantly less likely to be stunted compared to those whose mothers had a BMI of 18.5 kg/m2 or less. Those who received vitamin A supplements were more likely to be stunted than those who did not.\n\n\nDiscussion\n\nThe current study determined the prevalence of stunting and associated risk factors with stunting among children aged (0-23, 24-59 and 0-59 months) in Egypt. Findings from the study demonstrate that stunting is a significant concern in Egypt, with a prevalence rate of 24.5%. Although, the EDHS data analysed did show a fluctuating pattern of prevalence for stunting among children under five in Egypt. It was 24.6% in 2005, rose to 28.9% in 2008, and then declined to 21.5% in 2014. The common factors among the three age groups were the survey year, region, wealth index, child age, mother education level and height, birth order, listening to media and ANC. While factors such as mother’s age, marital status, and duration of breastfeeding were associated only with children aged 0-23 months, paternal education level, mother had BMI >25, and mother’s employment in the last 12 months were factors associated with children aged 0-59 months but not with children aged 0-23 months. Chid gender and the Vitamin A supplement were associated with both children aged 0-23 and 0-59 months.\n\nStudy findings suggest stunting in Egypt has decreased slightly over the last decade and the declining in stunting can potentially be attributed to the government’s endeavours in addressing undernutrition. The Egyptian government implemented the Agenda for Action Policy Paper (2017-2025) which was prepared for the main purpose of defining a feasible and practical ‘roadmap’ that would transform government-led nutrition policies and programs and mobilise harmonise multi-sector, as well as a multi-stakeholder, support to scaling-up of nutrition actions.38 Central to this process was the formulation of an updated National Nutrition Policy and Strategy which was aligned to important national and global commitments, such as the Egypt Vision 2030, and the Sustainable Development Goals’ “Agenda for action” in 2023, The National Agenda for Sustainable Development Egypt’s Updated Vision 2030 was published with number of strategies, government plans and programs for Egypt to achieve the SDGs by 2030.39 Although various efforts have been made,38 stunting remains a significant issue in Egypt, and several factors may contribute to its persistence. For example, the lack of regular data collection is a critical challenge as the last EDHS was conducted in 2014, nearly a decade ago.40 The lack of updated, monitoring and assessment data hampers the ability to clearly understand and detect problems earlier, as well as to assess the effectiveness of existing policies. This makes it challenging to identify issues promptly and implement effective preventive measures. Therefore, it is crucial to advocate for and invest in the comprehensive and consistent collection of nationally representative data as an integral part of new policies.\n\nFurthermore, other factors that can exacerbate undernutrition or hinder Egypt’s efforts to address the issue include the widespread levels of inequality and poverty. Poverty in Egypt affected a substantial portion of the population in 2015, with 27.8% living in poverty and an additional 28.7% at risk of becoming impoverished.18 In a study carried out in 2020, it was emphasized that Egypt faces deep-rooted inequalities in multiple domains, including income, wealth, education, gender, employment, and health. These disparities have persisted and even worsened over time.41 Children from lower socioeconomic background have less access to quality healthcare, nutritious food, safe living conditions, and educational opportunities, which leads to poorer health outcomes.18 Poor child health as a result of poverty and inequality has a lasting impact, including lower educational attainment, lower earning capacity, and an increased risk of chronic health disorders in adulthood, continuing a cycle of poverty and poor health throughout generations.18,41\n\nFurthermore, the study discovered that stunting is more common among children living in rural areas of Upper Egypt, which represents > 30 % of the population when compared to the other five regions. Poverty plays a major role in this trend as approximately 43 % of the population in rural Upper Egypt live below the poverty line that represents the highest percentage in the country. There are significant challenges in this area, such as a low level of education, especially among individuals whose fathers also had limited education. Limited access to healthcare, diverse foods, and nutritional knowledge, due to educational, economic, geographic, and gender disparities, worsens the risk of malnutrition among vulnerable populations.18 A recent report states that over half of the population in Egypt’s rural Upper Egypt is currently facing challenges in meeting their basic needs, including both food and non-food essentials. According to the results of the Household Income and Expenditure Survey, there has been a decline in poverty rates in rural Upper Egypt as the percentage dropped from 56.7% in 2015 to 51.94% in 2018.42 Despite this, the rural area of Upper Egypt region continues to uphold its reputation as the most economically underdeveloped in Egypt. Presently, the current national poverty rate is recorded at 32.5%. In urban governorates, the percentage stands at 26.73%, whereas in urban Lower Egypt, it amounts to 14.31%. In the rural regions of Lower Egypt, the percentage amounts to 27.29%. The African continent, along with other developing nations, is experiencing the destructive consequences of the interconnected issues of malnutrition and poverty.43 Poverty is the main cause of malnutrition, its presence in early life can worsen the cycle of poverty and result in severe and long-lasting health consequences.43\n\nAnalysis of the current study indicated that children from the middle wealth households were less prone to being stunted, compared to their counterparts from the richest households, which does not reflect findings from a systematic review conducted in sub-Saharan Africa and Bangladesh.29,44 Individuals with a higher wealth index have fewer nutritional health issues because they have greater access to high-quality and variety meals, higher levels of health education and awareness, and better healthcare services.13,45 Wealth also creates a safer living environment, reduces food insecurity and stress, and promotes cultural norms that favour healthy eating. These elements combine to improve the nutritional well-being of wealthy people.13\n\nFurthermore, the study findings explored that the increased odds of stunting were associated with mothers who had a low level of education and who worked in the last 12 months. This finding may be attributed to the fact that mothers with a higher education level are more knowledgeable about the advantages of exclusive breastfeeding during the first six months, as well as appropriate nutrition and early feeding practices.46,47 Mothers of high education attainment may comprehend the significance of timely, frequent, and diverse complementary nutrition, which allows them to make more informed decisions regarding their child’s health status.47 In addition, mothers who are educated are more likely to practise good hygiene and are more aware of the importance of regular check-ups and preventive care for their children, as well as healthcare services.48,49 Education should be considered as an essential issue regarding child stunting for Egypt, like many other developing countries as several studies have reported a better education level as a strong determinant of better health outcomes.50–52 In consonance with findings from several previous studies such as those from Nigeria,53 Pakistan,54 Indonesia and Bangladesh,55 and from systematic review studies.28,29,56 Mothers who have recently entered the workforce may find it challenging to dedicate sufficient time and energy to their children’s care. The return to work for mothers often has a negative impact on consistent breastfeeding. Exclusive breastfeeding rates are higher among mothers who are not employed than those who are employed.57 While some studies suggest that maternal employment can positively impact child health due to increased family income,58 this dynamic must be considered within the cultural context. In Arab countries like Egypt, the working mother often faces unique challenges. A mother is typically expected to balance her job with family commitments59 such as childcare, cleaning, and cooking, while the partner primarily focuses on providing for the family’s external needs. When a mother works, this often leads to an overwhelming burden on her health, as she must juggle multiple roles simultaneously.59 The cultural nuance of this situation underscores the importance of understanding cultural and social factors that influence the well-being of both the mother and child.\n\nThe study analysis found that the duration of breastfeeding > 12 months was more prevalent among children aged 0-23 and 0-59 months and more associated with children aged 0-23 months Similarly, in Pakistan, a study by Syeda et al. revealed that three-year-old children had a significantly higher risk of stunting compared to two-year-old children who were breastfed during their second year, even after accounting for other factors related to mother, child and health care services. The study also found that the three-year-old children who received breastfeeding until this age exhibited an elevated susceptibility to severe stunting when compared to their two-year-old counterparts.60 Furthermore, a cohort study conducted in Sudan found that stunting and wasting were more common among children who received prolonged breastfeeding compared to their well-nourished peers. The variance in height gains at 6 months was slight between children who were breastfed and those who were fully weaned. However, there was a significant decrease in weight gain observed among children who were breastfed, particularly between the ages of 6 and 12 months.61 Several studies conducted in different countries, including Ghana,62,63 Sub-Saharan African countries,64 and rural Senegal,65 had demonstrated a positive correlation between the duration of breastfeeding and the occurrence of stunting, wasting, and underweight. Furthermore, these studies have observed that this correlation strengthens as children grow older. Various factors mentioned by the forementioned studies that may be accountable for the found association, such as the findings of the study conducted in Sudan by Fawzi et al. which found an association between prolonged breastfeeding and stunting and wasting were greater among children of disadvantaged or illiterate mothers compared to those of educated, wealthier mothers. The inverse association is hypothesized to stem from inadequate complementary feeding in breast-fed children relative to weaned children, particularly in families with low incomes.61 The interpretation of the findings needs to be done so with caution as both the current study and those mentioned above have not considered the dietary quality and energy intake received from oral intake alongside the measure of breastfeeding duration as dietary diversity was not included in the 2005 DHS data. Further investigation is needed to determine the interplay between diet quality, breastfeeding duration, and anthropometric outcomes for children aged over 12 months in Egypt.\n\nAdditionally, the study analysis found that inadequate use of vitamin A supplement was associated with stunting in both children aged 0-23 and 0-59 months. Vitamin A is essential for healthy vision, immune system function, and for child growth and development.66 The micronutrients deficiency is especially concerning in children, as it may impair growth, immune function, and overall health, thus increasing the risk of undernutrition.66 The lack of knowledge regarding the significance of vitamin A and its sources in low socioeconomic communities is a contributing factor affecting vitamin A consumption.10 A similar result was found by study conduct in Brazil as it found that the probability of children experiencing stunting and anemia decreased by 8% and 31% due to vitamin A supplementation, the intake of vitamin A supplements reduces the risk of stunting.67\n\nThe likelihood of a child being stunted was found to be higher in those whose mothers had limited or no ANC visits during pregnancy. This was consistent with findings from Ethiopia35 and Zambia.30 Routine ANC is a vital health service for expectant mothers that aims to identify potential obstetric complications, provide guidance on healthy lifestyle choices, pregnancy, and childbirth.68 Through regular ANC visits, healthcare providers can detect high-risk pregnancies early, thereby reducing the risk of maternal mortality and closely monitoring the progress of the fetus. The World Health Organisation (WHO) stated that ANC is crucial in reducing pregnancy and childbirth complications, stillbirths, and perinatal deaths through evidence-based actions. Moreover, WHO stated that ANC provides a significant chance to engage with and support women, their families, and communities during a crucial stage in a woman’s life.69\n\nThe study analysis also revealed that boys were significantly more prone to being stunted than girls, which is in consonance with findings from past studies that show that overall, boys tend to be at greater risk of stunting than girls,70 especially in Sub-Saharan Africa and in lower socioeconomic group.71 This finding may be attributed to the fact that research has consistently shown that boys are more prone to common childhood illnesses like lower respiratory infections, diarrheal disease, and malaria compared to girls.72,73 Additionally, boys tend to experience higher rates of wasting, and underweight,70,74 which suggest that boys are more vulnerable to environmental stressors and have higher rates of infectious morbidity and mortality.75–80 In some societies or regions, the health and well-being of younger females may be prioritised by social values and practices, which may lead to superior nutrition and care for them in comparison to older girls or boys. As a result, some researchers and policymakers recommend targeting interventions towards boys81,82 or reevaluating the focus on girls and women in nutrition research.70,74,83,84\n\nAnalysis of the current study corroborates this finding, by showing that the odds of stunting among children 0-59 months were significantly higher among children aged 18-41 months. Past studies have revealed that the odds of stunting were higher in older children than in younger ones. For instance, in Myanmar, older children (24-35 months) are more likely to be stunted than those under 6 months85 and South Ari District, Southern Ethiopia: a community-based cross-sectional study found that children in the age group of 24–59 months were more prone to stunting than others.86 Another cross sectional study targeted four North African countries found the higher prevalence of stunting was among children aged 24-59 months.11 This is most likely due to the cumulative effect of foetal and early childhood development delays. Children who remain stunted after two years have missed the critical window of opportunity to reverse the impacts of poor growth and development, which occurs within the first 1,000 days following conception until age two.87 Stunting after two years raises the probability of poor long-term cognitive, educational, and productive results. In contrast, a cross-sectional study of 94 low- and middle-income countries on patterns in child stunting by age observed that Stunting prevalence was higher for younger children until around age 28 months.88 Research indicates that genetics has a more significant impact on growth patterns after the age of two, implying that environmental factors have a greater influence on the growth of younger children compared to older children.89 One possible explanation for this phenomenon is that as children mature, their immune system becomes more robust, enabling them to better resist pathogens and reducing the impact of environmental stressors on their growth.66\n\nThe analysis revealed that shorter mothers were more likely to have stunted children. The finding suggests that a mother’s height is a significant factor in determining her child’s birth length, with taller mothers more likely to have longer children and shorter mothers more likely to have shorter children. This indicates a strong genetic link between maternal height and child length.90 However, it is also important to note that external factors, such as adequate nutrition, play a crucial role in a child’s growth and development. Ensuring proper nutritional intake is essential to support healthy growth and prevent delays in development, as highlighted by a past.90,91 This finding was consistent with those of other previous studies from Indonesia,92 south Asia93 and Bangladesh.94 Furthermore, Children of mothers with a BMI greater than 25 were found to be less likely to suffer from stunting compared to their counterparts. While some studies have highlighted the prevalent coexistence of stunting and maternal overweight,34,44,95 the risk of child stunting and overweight in the mother-child pair appear to be strongly related to other specific maternal characteristics rather than her BMI. These include shorter maternal height, younger maternal age at birth, lower levels of education, and the household’s economic status, as indicated by the wealth index and the number of household members.96 These factors play a critical role in determining the nutritional health and overall well-being of both mother and child.34,96\n\nTo address the prevalence of child stunting in Egypt, particularly in rural areas of Upper Egypt, the following policy actions are recommended: 1) implement targeted interventions focusing on boys and children born to short mothers, who are at higher risk of stunting; 2) ensure universal access to ANC for all pregnant women, prioritising those with limited or no previous ANC visits; 3) provide nutrition counselling and education to mothers, emphasising the importance of adequate feeding practices during pregnancy and early childhood; 4) rural focused initiatives: develop and implement context-specific initiatives addressing the unique challenges faced by rural populations in Upper Egypt; 5) foster collaboration between healthcare, education, and social protection sectors to address the multifaceted determinants of child stunting and 6) establish a robust monitoring and evaluation system to track progress, identify areas for improvement, and adjust policies accordingly.\n\nAlthough this study has several strengths, including the use of a large sample size, pooling three large population-based dataset which enhances statistical power and allows for comparisons across different studies, it also provides detailed insights into changes in the prevalence of undernutrition and its associated factors among children under five in Egypt. By focusing on specific geographic areas and age groups (0-23 months, 24-59 months, and 0-59 months), a more comprehensive understanding of the distinct challenges and circumstances that contribute to undernutrition will enable for more impactful interventions.\n\nThe utilisation of a population-based design and a significant pooled sample size ensure that the collected data correctly reflect the total population within the designated area. This strategy enhanced the validity and reliability of results by minimising selection bias and facilitating more precise calculations of prevalence and connections. However, this study was not without its limitations. MICS surveys primarily focus on cross-sectional data collection, limiting the ability to establish causal relationships or understand the dynamic factors contributing to undernutrition. Furthermore, cross-sectional statistics rely on self-reported data, which can be influenced by recall or social desirability biases, leading to inaccuracies. Ultimately, it is important to consider the possibility of unaccounted confounding variables affecting the results of the study. As an example, it should be noted that the study findings regarding breastfeeding and Vitamin A supplement could have been clearer if dietary intake variables for the Infant and young child feeding indicators were included. However, due to the unavailability of data about dietary intake in the EDHS 2005, this was not possible.\n\n\nConclusions\n\nChild stunting is a significant public health challenge in Egypt, necessitating focused interventions based on critical risk factors. The current study found the length and the lack of ANC visits were significant factors, highlighting the critical role of maternal healthcare and nutrition. Similarly, this study found boys are more impacted, indicating a necessity for gender-sensitive initiatives. Successful stunting reduction necessitates a comprehensive strategy that incorporates maternal healthcare, nutrition, and educational programs. Improved access to and quality of ANC, poverty reduction, and enhanced healthcare in disadvantaged regions are essential. Furthermore, enduring investments in infrastructure, community-oriented nutritional initiatives, and social safety nets are crucial for sustainable growth and development. Ongoing research and systematic national data collecting will facilitate the monitoring and enhancement of these treatments.\n\n\nEthical statement\n\nEDHS ethical clearance was obtained from the Inner-City Fund (ICF) International, and permission to use the data was obtained from ICF International for this study. This study used existing EDHS data sets and did not involve interaction with the participants. A re-analysis was performed with the participants’ original consent. Thus, no further consent was obtained from the participants.", "appendix": "Data availability statement\n\nThe used data is national representative data. The data are available in the public domain and can be accessed through the prescribed registration on the official DHS program website (https://dhsprogram.com/) (accessed on 25 Oct 2024). The data are available in the public domain and can be accessed through the prescribed registration on the official DHS program website (https://dhsprogram.com/).\n\nFigshare repository: Concurrent stunting among under-five children in Egypt. (DOI. 10.6084/m9.figshare.28030355.v1).97\n\nThis project contains the following underlying information:\n\n• Table S1. Distribution of Determinants of Child Nutritional Health: EDHS for the Years 2005, 2008, and 2014;\n\n• Table S2. Prevalence of stunting among children (0-23, 24-59 and 0-59 months) in Egypt by the basic, underlying and immediate factors;\n\n• Table S3. Unadjusted Odds Ratios for Determinants of Stunting among Children aged (0-23, 24-59 and 0-59 months) in Egypt.\n\nMaterials are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nThis paper forms part of the first author’s doctoral dissertation at the School of Health Sciences, Western Sydney University.\n\n\nReferences\n\nUnited Nations Children’s Fund, World Health Organization & International Bank for Reconstruction, Development/The World Bank: Levels and Trends in Child Malnutrition: UNICEF/WHO/The World Bank Group Joint Child Malnutrition Estimates: Key Findings of the 2021 Edition. World Health Organization; 2021.\n\nDe Sanctis V, Soliman A, Alaaraj N, et al.: Early and Long-term Consequences of Nutritional Stunting: From Childhood to Adulthood. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMinistry of Health and Population, National Nutrition Institute, UNICEF: Nutrition agenda for action policy paper on scaling up nutrition interventions in Egypt.2017.\n\nMinistry of Planning and Economic Developent: The National agenda for sustainable development Egypt’s updated vision 2030.2023.\n\nElmighrabi NF, Fleming CAK, Dhami MV, et al.: A systematic review and meta-analysis of the prevalence of childhood undernutrition in North Africa. PLoS One. 2023; 18(4): e0283685. PubMed Abstract \| Publisher Full Text\n\nGhafar AA: Causes and consequences of inequality in Egypt. Muslim World. 2021; 111(1): 5–26. Publisher Full Text\n\nCAPMAS: The Central Agency for Public Mobilization and Statistics. Egypt in figures, formal report March 2018.2018.\n\nAdeyeye SAO, Adebayo-Oyetoro AO, Tiamiyu HK: Poverty and malnutrition in Africa: a conceptual analysis. Nutr. Food Sci. 2017; 47: 754–764. Publisher Full Text\n\nSarma H, Khan JR, Asaduzzaman M, et al.: Factors influencing the prevalence of stunting among children aged below five years in Bangladesh. Food Nutr. Bull. 2017; 38(3): 291–301. PubMed Abstract \| Publisher Full Text\n\nAustralian Institute of Health and Welfare: Australia’s health 2016. Canberra: Australian Institute of Health and Welfare; 2016 [cited 2024 Sep. 27]. Reference Source\n\nNeves PA, Barros AJ, Gatica-Domínguez G, et al.: Maternal education and equity in breastfeeding: trends and patterns in 81 low- and middle-income countries between 2000 and 2019. Int. J. Equity Health. 2021; 20: 1–3.\n\nLiaqat P, Rizvi MA, Qayyum A, et al.: Maternal education and complementary feeding. Pak. J. Nutr. 2006; 5(6): 563–568. Publisher Full Text\n\nBain LE, Awah PK, Geraldine N, et al.: Malnutrition in Sub–Saharan Africa: burden, causes and prospects. Pan Afr. Med. J. 2013; 15(1): 120. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRachmi CN, Agho KE, Li M, et al.: Stunting, underweight and overweight in children aged 2.0–4.9 years in Indonesia: prevalence trends and associated risk factors. PLoS One. 2016; 11(5): e0154756. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIpa M, Widawati M, Laksono AD, et al.: Variation of preventive practices and its association with malaria infection in eastern Indonesia: Findings from community-based survey. PLoS One. 2020; 15(5): e0232909. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAndayani Q, Laksono AD: Neonatal death incidence in healthcare Facility in Indonesia: does antenatal care matter? Indian J. Forensic Med. Toxicol. 2021; 15(1): 1265.\n\nWulandari RD, Laksono AD: Determinants of knowledge of pregnancy danger signs in Indonesia. PLoS One. 2020; 15(5): e0232550. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLawal SA, Okunlola DA, Adegboye OA, et al.: Mother’s education and nutritional status as correlates of child stunting, wasting, underweight, and overweight in Nigeria: Evidence from 2018 Demographic and Health Survey. Nutr. Health. 2023; 30: 821–830. Publisher Full Text\n\nFatima S, Manzoor I, Joya AM, et al.: Stunting and associated factors in children of less than five years: A hospital-based study. Pak. J. Med. Sci. 2020; 36(3): 581–585. PubMed Abstract \| Publisher Full Text\n\nSemba RD, de Pee S , Sun K, et al.: Effect of parental formal education on risk of child stunting in Indonesia and Bangladesh: a cross-sectional study. Lancet. 2008; 371(9609): 322–328. PubMed Abstract \| Publisher Full Text\n\nTahangnacca M, Amiruddin R, Syam A: Model of stunting determinants: A systematic review. Enferm. Clin. 2020; 30: 241–245. Publisher Full Text\n\nMurtagh LMA: Working mothers, breastfeeding, and the law. Am. J. 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(Doctoral dissertation).\n\nNubé M, Asenso-Okyere WK: Large differences in nutritional status between fully weaned and partially breast fed children beyond the age of 12 months. Eur. J. Clin. Nutr. 1996; 50(3): 171–177. PubMed Abstract\n\nSimondon KB, Simondon F: Mothers prolong breastfeeding of undernourished children in rural Senegal. Int. J. Epidemiol. 1998; 27(3): 490–494.\n\nCaulfield LE, Bentley ME, Ahmed S: Is prolonged breastfeeding associated with malnutrition? Evidence from nineteen demographic and health surveys. Int. J. Epidemiol. 1996; 25(4): 693–703. PubMed Abstract \| Publisher Full Text\n\nMaggini S, Pierre A, Calder PC: Immune function and micronutrient requirements change over the life course. Nutrients. 2018; 10(10): 1531. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSilva-Neto LG, dos Santos Neto JE , de Menezes Toledo Florêncio TM : Association between vitamin A supplementation and stunting and anemia in socially vulnerable Brazilian children. Eur. J. Nutr. 2024; 23: 1–8.\n\nDowne S, Finlayson K, Tunçalp O, et al.: Provision and uptake of routine antenatal services: a qualitative evidence synthesis. Cochrane Database Syst. Rev. 2019; 2019(6). Publisher Full Text\n\nWorld Health Organisation recommendations on antenatal care for a positive pregnancy experience. 20 Avenue Appia, 1211 Geneva 27, Switzerland: World Health Organization; 2016.\n\nThurstans S, Opondo C, Seal A, et al.: Boys are more likely to be undernourished than girls: a systematic review and meta-analysis of sex differences in undernutrition. BMJ Glob. Health. 2020; 5(12): e004030. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWamani H, Åstrøm AN, Peterson S, et al.: Boys are more stunted than girls in sub-Saharan Africa: a meta-analysis of 16 demographic and health surveys. BMC Pediatr. 2007; 7: 1–10. Publisher Full Text\n\nSawyer CC: Child mortality estimation: estimating sex differences in childhood mortality since the 1970s.2012.\n\nCosta JC, da Silva IC , Victora CG: Gender bias in under-five mortality in low/middle-income countries. BMJ Glob. Health. 2017; 2(2): e000350. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNavarro D, Serrano M, Dolores M, et al.: Female eco-stability and severe malnutrition in children: Evidence from humanitarian aid interventions of Action Against Hunger in African, Asian and Latin American countries. Clin. Nutr. Hosp. Diet. 2017; 37(4): 127–134.\n\nStinson S: Sex differences in environmental sensitivity during growth and development. Am. J. Phys. Anthropol. 1985; 28(S6): 123–147. Publisher Full Text\n\nEriksson JG, Kajantie E, Osmond C, et al.: Boys live dangerously in the womb. Am. J. Hum. Biol. 2010; 22(3): 330–335. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKirchengast S, Hartmann B: The male disadvantage hypothesis reconsidered: is there really a weaker sex? An analysis of gender differences in newborn somatometrics and vital parameters. J. Life Sci. 2009; 1(1): 63–71. Publisher Full Text\n\nDrevenstedt GL, Crimmins EM, Vasunilashorn S, et al.: The rise and fall of excess male infant mortality. Proc. Natl. Acad. Sci. 2008; 105(13): 5016–5021. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nStevenson DK, Verter J, Fanaroff AA, et al.: Sex differences in outcomes of very low birthweight infants: the newborn male disadvantage. Arch. Dis. Child Fetal Neonatal Ed. 2000; 83(3): 182F–1185F. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWells JC: Natural selection and sex differences in morbidity and mortality in early life. J. Theor. Biol. 2000; 202(1): 65–76. PubMed Abstract \| Publisher Full Text\n\nSlemming W, Kagura J, Saloojee H, et al.: Early life risk exposure and stunting in urban South African 2-year old children. J. Dev. Orig. Health Dis. 2017; 8(3): 301–310. PubMed Abstract \| Publisher Full Text\n\nOyenubi A: Who benefits from the South African Child Support Grant?: The role of gender and birthweight. Dev. South. Afr. 2021; 38(4): 539–563. Publisher Full Text\n\nKhara T, Mwangome M, Ngari M, et al.: Children concurrently wasted and stunted: A meta-analysis of prevalence data of children 6–59 months from 84 countries. Matern. Child Nutr. 2018; 14(2): e12516. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMehta S, Mugusi FM, Bosch RJ, et al.: A randomized trial of multivitamin supplementation in children with tuberculosis in Tanzania. Nutr. J. 2011; 10: 1–9.\n\nWHO: Global nutrition targets 2025: low birth weight policy brief- external site opens in new window. Geneva: WHO; 2014.\n\nToma TM, Andargie KT, Alula RA, et al.: Factors associated with wasting and stunting among children aged 06–59 months in South Ari District, Southern Ethiopia: a community-based cross-sectional study. BMC Nutr. 2023; 9(34): 34. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFink G, Rockers PC: Childhood growth, schooling, and cognitive development: further evidence from the Young Lives study. Am. J. Clin. Nutr. 2014; 100: 182–188. PubMed Abstract \| Publisher Full Text\n\nKarlsson O, Kim R, Moloney GM, et al.: Patterns in child stunting by age: A cross-sectional study of 94 low-and middle-income countries. Matern. Child Nutr. 2023; 19(4): e13537. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJelenkovic A, Sund R, Yokoyama Y, et al.: Genetic and environmental influences on human height from infancy through adulthood at different levels of parental education. Sci. Rep. 2020; 10(1): 7974. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAddo OY, Stein AD, Fall CH, et al.: Consortium on Health Orientated Research in Transitional Societies (COHORTS) Group. Maternal height and child growth patterns. J. Pediatr. 2013; 163(2): 549–554.e1. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDesmond C, Casale D: Catch-up growth in stunted children: Definitions and predictors. PLoS One. 2017; 12(12): e0189135. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAhmawati VE, Pamungkasari EP, Murti B: Determinants of stunting and child development in Jombang District. J. Matern. Child Health. 2018; 03(1): 68–80. Publisher Full Text\n\nKim R, Mejía-Guevara I, Corsi DJ, et al.: Relative importance of 13 correlates of child stunting in South Asia: Insights from nationally representative data from Afghanistan, Bangladesh, India, Nepal, and Pakistan. Soc. Sci. Med. 2017; 187: 144–154. PubMed Abstract \| Publisher Full Text\n\nRahman MS, Howlader T, Masud MS, et al.: Association of low-birth weight with malnutrition in children under five years in Bangladesh: do mother’s education, socio-economic status, and birth interval matter? PLoS One. 2016; 11(6): e0157814. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPomati M, Mendoza-Quispe D, Anza-Ramirez C, et al.: Trends and patterns of the double burden of malnutrition (DBM) in Peru: a pooled analysis of 129,159 mother–child dyads. Int. J. Obes. 2021; 45(3): 609–618. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRachmah Q, Mahmudiono T, Loh SP: Predictor of Obese Mothers and Stunted Children in the Same Roof: A Population-Based Study in the Urban Poor Setting Indonesia. Front. Nutr. 2021; 8: 710588. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nElmighrabi N: Supplementary materials. Dataset. figshare. 2024. Publisher Full Text" }	[ { "id": "358467", "date": "27 Jan 2025", "name": "Godana Arero Dassie", "expertise": [ "Reviewer Expertise 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\nWhile emphasizing the importance of sectoral collaboration, risks overgeneralizing the complexity of addressing stunting. It does not adequately acknowledge the potential challenges in implementation, such as resource constraints, political will, and the difficulties of coordinating efforts across multiple sectors.\nPlease refer to the detailed assessment of the article here - https://f1000research.s3.amazonaws.com/linked/704364.Under_reviewing.docx\n\nFurthermore, focusing interventions on children born to short mothers could inadvertently stigmatize this group and may overlook other critical determinants of stunting, such as paternal influence or environmental factors. The broad scope of the recommendation, which suggests large-scale interventions, does not sufficiently consider the feasibility of such initiatives, including budget limitations, sector prioritization, and community-specific barriers that could hinder successful implementation.\nLastly, while interventions for low-SES families are essential, the conclusion would benefit from greater specificity regarding the types of interventions proposed, such as cash transfers, community-based education, or food security measures, to ensure clarity and practical applicability in real-world settings.\nTo address stunting in Egypt, a coordinated approach across health, WASH, education, and social protection is essential. Interventions should target high-risk groups, particularly those with low socioeconomic status, focusing on healthcare access, parental education, and infant feeding practices.\nWhile short maternal height increases risk, interventions must also tackle broader factors like food security and sanitation. A nutrition monitoring framework integrated into health systems will enable data collection, evidence-based actions, and progress tracking.\nEffective implementation requires cost-effective, scalable solutions, community involvement, and resource prioritization to ensure sustainability and equity.\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": [] } ]	1	https://f1000research.com/articles/14-15
https://f1000research.com/articles/13-1049/v1	12 Sep 24	{ "type": "Data Note", "title": "A guide to selecting high-performing antibodies for STING1 (Uniprot ID: Q86WV6) for use in western blot, immunoprecipitation, and immunofluorescence", "authors": [ "Vera Ruíz Moleón", "Charles Alende", "Maryam Fotouhi", "Riham Ayoubi", "Kathleen Southern", "Carl Laflamme", "NeuroSGC/YCharOS/EDDU collaborative group", "ABIF consortium", "Vera Ruíz Moleón", "Charles Alende", "Maryam Fotouhi", "Riham Ayoubi", "Kathleen Southern" ], "abstract": "STING1 is an immune adaptor protein which promotes innate immune defense mechanisms against pathogens. Its role in modulating inflammation links STING1 to various pathologic conditions, positioning it as a key target for therapeutic interventions aimed at regulating immune responses. To advance our understanding of STING1-associated diseases, it is essential to make high-performing antibodies readily accessible to the scientific community. This study aims to improve reliability of STING1 research as we have characterized sixteen STING1 commercial antibodies for western blot, immunoprecipitation, and immunofluorescence using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. These studies are part of a larger, collaborative initiative seeking to address antibody reproducibility issues by characterizing commercially available antibodies for human proteins and publishing the results openly as a resource for the scientific community. While use of antibodies and protocols vary between laboratories, we encourage readers to use this report as a guide to select the most appropriate antibodies for their specific needs.", "keywords": [ "Uniprot ID: Q86WV6", "STING1", "hSTING", "MPYS", "MITA", "stimulator of interferon genes protein", "mediator of IRF3 activation", "transmembrane protein 173", "antibody characterization", "antibody validation", "western blot", "immunoprecipitation", "immunofluorescence" ], "content": "Introduction\n\nStimulator of interferon genes protein (STING1) is a conserved transmembrane protein involved in innate immune response mechanisms.1,2 Activated in response to bacterial cyclic dinucleotides, it initiates a cascade of signalling events inducing the production of type I interferons and inflammatory cytokines, essential molecules to activate immune response signalling.1,3,4\n\nWhile STING1-mediated innate immunity is involved in defending against pathogen invasion and tumor growth, aberrant expression can disrupt immune responses,5 leading to autoinflammatory diseases, cancer and neurodegenerative diseases.6–8 In Parkinson’s disease models, α-synuclein aggregates enhance STING1-dependent neuroinflammation and neurodegeneration.9 Due to its involvement in critical immune pathways, STING1 has become a key focus for therapeutic development.\n\nThis research is part of a broader collaborative initiative in which academics, funders and commercial antibody manufacturers are working together to address antibody reproducibility issues by characterizing commercial antibodies for human proteins using standardized protocols, and openly sharing the data.10–12 Here we evaluated the performance of sixteen commercial antibodies for STING1 for use in western blot, immunoprecipitation, and immunofluorescence, enabling biochemical and cellular assessment of STING1 properties and function. The platform for antibody characterization used to carry out this study was endorsed by a committee of industry academic representatives. It consists of identifying human cell lines with adequate target protein expression and the development/contribution of equivalent knockout (KO) cell lines, followed by antibody characterization procedures using most commercially available antibodies against the corresponding protein. The standardized consensus antibody characterization protocols are openly available on Protocol Exchange, a preprint server (DOI: 10.21203/rs.3.pex-2607/v1).13\n\nThe authors do not engage in result analysis or offer explicit antibody recommendations. Our primary aim is to deliver top-tier data to the scientific community, grounded in Open Science principles. This empowers experts to interpret the characterization data independently, enabling them to make informed choices regarding the most suitable antibodies for their specific experimental needs. Guidelines on how to interpret antibody characterization data found in this study are featured on the YCharOS gateway.14\n\n\nResults and discussion\n\nOur standard protocol involves comparing readouts from WT (wild type) and KO cells.15,16 The first step was to identify a cell line(s) that expresses sufficient levels of a given protein to generate a measurable signal using antibodies. To this end, we examined the DepMap (Cancer Dependency Map Portal, RRID:SCR_017655) transcriptomics database to identify all cell lines that express the target at levels greater than 2.5 log2 (transcripts per million “TPM” + 1), which we have found to be a suitable cut-off.10 The THP-1 cell line expresses the STING1 transcript at 4.1 log2 (TPM+1), which is above the average range of cancer cells analyzed. A STING1 KO in THP-1 was obtained from Abcam (Table 1). THP-1 cells are small, round human leukemia monocytic cell line commonly used to study proteins involved in immune responses. Phorbol 12-myristate-13-acetate (PMA) treatment is required to differentiate THP-1 in suspension into macrophage-like adherent cells.17\n\nFor western blot experiments, WT and STING1 KO protein lysates were first separated on SDS-PAGE, transferred onto nitrocellulose membranes, and then probed with sixteen STING1 antibodies in parallel (Table 2, Figure 1).\n\n* Monoclonal antibody.\n\n Recombinant antibody.\n\nLysates from PMA-treated THP-1 WT and STING1 KO were prepared, and 40 μg of protein were processed for western blot with the indicated STING1 antibodies. The Ponceau stained transfers of each blot are presented to show equal loading of WT and KO lysates and protein transfer efficiency from polyacrylamide gels to the nitrocellulose membrane. Antibody dilutions were chosen according to the recommendations of the antibody supplier. Antibody dilution used: ab181125 at 1/1000, ab227704 at 1/200, ab227705 at 1/1000, ab239074 at 1/1000, A21051 at 1/1000, 13647 at 1/1000, 50494 at 1/1000, CPTC-STING1-1 at 1/10, GTX134373 at 1/1000, NBP2-24683 at 1/1000, NBP3-18816 at 1/1000, 19851-1-AP at 1/1000, MAB7169* at 1/1000, 702993** at 1/1000, MA5-26030* at 1/1000, MA5-32768** at 1/1000. Predicted band size: 42 kDa. Monoclonal antibody, Recombinant antibody.\n\nWe then assessed the capability of all sixteen antibodies to capture STING1 from THP-1 protein extracts using immunoprecipitation techniques, followed by western blot analysis. For the immunoblot step, a specific STING1 antibody identified previously (refer to Figure 1) was selected. Equal amounts of the starting material (SM) and unbound fractions (UB), as well as the whole immunoprecipitate (IP) eluates were separated by SDS-PAGE (Figure 2).\n\nLysates from PMA-treated THP-1 WT were prepared, and immunoprecipitation was performed using 1.0 mg of lysate and 2.0 μg of the indicated STING1 antibodies pre-coupled to Dynabeads protein A or protein G. Samples were washed and processed for western blot with the indicated STING1 antibody. For western blot, NBP3-18816* was used at 1/1000. The Ponceau stained transfers of each blot are shown. SM = 4% starting material; UB = 4% unbound fraction; IP = immunoprecipitate, HC = antibody heavy chain. Monoclonal antibody, Recombinant antibody.\n\nFor immunofluorescence, sixteen antibodies were screened using a mosaic strategy. First, THP-1 WT and STING1 KO cells were labelled with different fluorescent dyes in order to distinguish the two cell lines, and the STING1 antibodies were evaluated. Both WT and KO lines imaged in the same field of view to reduce staining, imaging and image analysis bias (Figure 3). Quantification of immunofluorescence intensity in hundreds of WT and KO cells was performed for each antibody tested, and the images presented in Figure 3 are representative of this analysis.13\n\nPMA-treated THP-1 WT and STING1 KO cells were labelled with a green or a far-red fluorescent dye, respectively. WT and KO cells were mixed and plated to a 1:1 ratio in a 96-well plate with optically clear flat-bottom. Cells were stained with the indicated STING1 antibodies and with the corresponding Alexa-fluor 555 coupled secondary antibody including DAPI. Acquisition of the blue (nucleus-DAPI), green (identification of WT cells), red (antibody staining) and far-red (identification of KO cells) channels was performed. Representative images of the merged blue and red (grayscale) channels are shown. WT and KO cells are outlined with green and magenta dashed line, respectively. When an antibody was recommended for immunofluorescence by the supplier, we tested it at the recommended dilution. The rest of the antibodies were tested at 1 and 2 μg/ml and the final concentration was selected based on the detection range of the microscope used and a quantitative analysis not shown here. Antibody dilution used: ab181125* at 1/300, ab227704 at 1/1000, ab227705 at 1/300, ab239074 at 1/500, A21051 at 1/300, 13647 at 1/60, 50494 at 1/500, CPTC-STING1-1 at 1/20, GTX134373 at 1/1000, NBP2-24683 at 1/1000, NBP3-18816 at 1/1000, 19851-1-AP at 1/200, MAB7169* at 1/200, 702993** at 1/250, MA5-26030* at 1/1000, MA5-32768** at 1/1000. Bars = 10 μm. Monoclonal antibody, Recombinant antibody.\n\nIn conclusion, we have screened sixteen STING1 commercial antibodies by western blot, immunoprecipitation, and immunofluorescence by comparing the signal produced by the antibodies in human THP-1 WT and STING1 KO cells. To assist users in interpreting antibody performance, Table 3 outlines various scenarios in which antibodies may perform in all three applications. Several high-quality and renewable antibodies that successfully detect STING1 were identified in all applications. Researchers who wish to study STING1 in a different species are encouraged to select high-quality antibodies, based on the results of this study, and investigate the predicted species reactivity of the manufacturer before extending their research.\n\nThe underlying data for this study can be found on Zenodo, an open-access repository for which YCharOS has its own collection of antibody characterization reports.18,19\n\nInherent limitations are associated with the antibody characterization platform used in this study. Firstly, the YCharOS project focuses on renewable (recombinant and monoclonal) antibodies and does not test all commercially available STING1 antibodies. YCharOS partners provide approximately 80% of all renewable antibodies, but some top-cited polyclonal antibodies may not be available through these partners.\n\nSecondly, the YCharOS effort employs a non-biased approach that is agnostic to the protein for which antibodies have been characterized. The aim is to provide objective data on antibody performance without preconceived notions about how antibodies should perform or the molecular weight that should be observed in western blot. As the authors are not experts in STING1, only a brief overview of the protein’s function and its relevance in disease is provided. STING1 experts are responsible for analyzing and interpreting observed banding patterns in western blots and subcellular localization in immunofluorescence.\n\nThirdly, YCharOS experiments are not performed in replicates primarily due to the use of multiple antibodies targeting various epitopes. Once a specific antibody is identified, it validates the protein expression of the intended target in the selected cell line, confirms the lack of protein expression in the KO cell line and supports conclusions regarding the specificity of the other antibodies. All experiments are performed using master mixes, and meticulous attention is paid to sample preparation and experimental execution. In immunofluorescence, the use of two different concentrations serves to evaluate antibody specificity and can aid in assessing assay reliability. In instances where antibodies yield no signal, a repeat experiment is conducted following titration. Additionally, our independent data is performed subsequently to the antibody manufacturers internal validation process, therefore making our characterization process a repeat.\n\nLastly, as comprehensive and standardized procedures are respected, any conclusions remain confined to the experimental conditions and cell line used for this study. The use of a single cell type for evaluating antibody performance poses as a limitation, as factors such as target protein abundance significantly impact results.13 Additionally, the use of cancer cell lines containing gene mutations poses a potential challenge, as these mutations may be within the epitope coding sequence or other regions of the gene responsible for the intended target. Such alterations can impact the binding affinity of antibodies. This represents an inherent limitation of any approach that employs cancer cell lines.\n\n\nMethods\n\nThe standardized protocols used to carry out this KO cell line-based antibody characterization platform was established and approved by a collaborative group of academics, industry researchers and antibody manufacturers. The detailed materials and step-by-step protocols used to characterize antibodies in western blot, immunoprecipitation and immunofluorescence are openly available on Protocol Exchange, a preprint server (DOI: 10.21203/rs.3.pex-2607/v1).13 Brief descriptions of the experimental setup used to carry out this study can be found below.\n\nCell lines used and primary antibodies tested in this study are listed in Tables 1 and 2, respectively. To ensure that the cell lines and antibodies are cited properly and can be easily identified, we have included their corresponding Research Resource Identifiers, or RRID.20,21 Cells were cultured in RPMI 1640 (Gibco, cat. number A1049101) containing 10% fetal bovine serum (Wisent, cat. number 080450), 0.05 mM 2-Mercaptoethanol (Gibco, cat. number 21985023, 2 mM L-glutamine (Wisent cat. number 609-065), 100 IU penicillin and 100 μg/ml streptomycin (Wisent cat. number 450201). THP-1 WT and STING1 KO cells were treated with 200 ng/ml PMA (Abcam, cat. number ab147465) for 2 days. 200 ng/ml of PMA was added to fresh medium in both day 1 and day 2.17 Peroxidase-conjugated goat anti-rabbit and anti-mouse antibodies (Thermo Fisher Scientific, cat. number 65-6120 and 62-6520) and Peroxidase-conjugated Protein A (MilliporeSigma, cat. number P8651) were used in western blot and immunoprecipitation. Alexa-555-conjugated goat anti-rabbit and anti-mouse secondary antibodies (Thermo Fisher Scientific, cat. number A-21429 and A-21424) were used in immunofluorescence.\n\nPMA-treated THP-1 WT and STING1 KO cells (listed in Table 1) were collected in RIPA buffer (25mM Tris-HCl pH 7.6, 150mM NaCl, 1% NP-40, 1% sodium deoxycholate, 0.1% SDS) (Thermo Fisher Scientific, cat. number 89901) supplemented with 1x protease inhibitor cocktail mix (MilliporeSigma, cat. number P8340). Lysates were sonicated briefly and incubated for 30 min on ice. Lysates were spun at ~110,000 × g for 15 min at 4°C and equal protein aliquots of the supernatants were analyzed by SDS-PAGE and western blot. BLUelf prestained protein ladder (GeneDireX, cat. number PM008-0500) was used.\n\nWestern blots were performed with precast midi 10% Bis-Tris polyacrylamide gels (Thermo Fisher Scientific, cat. number WG1201BOX) ran with MES SDS buffer (Thermo Fisher Scientific, cat. number NP000202), loaded in LDS sample buffer (Thermo Fisher Scientific, cat. number NP0008) with 1× sample reducing agent (Thermo Fisher Scientific, cat. number NP0009) and transferred on nitrocellulose membranes. Proteins on the blots were visualized with Ponceau S staining (Thermo Fisher Scientific, cat. number BP103-10) which is scanned to show together with individual western blot. Blots were blocked with 5% milk for 1 hr, and antibodies were incubated overnight at 4°C with 5% milk in TBS with 0.1% Tween 20 (TBST) (Cell Signaling Technology, cat. number 9997). Following three washes with TBST, the peroxidase conjugated secondary antibody was incubated at a dilution of ~0.2 μg/ml in TBST with 5% milk for 1 hr at room temperature followed by three washes with TBST. Membranes were incubated with Pierce ECL (Thermo Fisher Scientific, cat. number 32106) or Clarity Western ECL Substrate (Bio-Rad, cat. number 1705061) prior to detection with the iBright™ CL1500 Imaging System (Thermo Fisher Scientific, cat. number A44240).\n\nAntibody-beads conjugates were prepared by adding 2 μg of antibody (with an exception 20 μl of antibody CPTC-STING1-1* and 20 μl of antibody 13647**) to 500 μl of Pierce IP Lysis Buffer (Thermo Fisher Scientific, cat. number 87788) in a microcentrifuge tube, together with 30μl of Dynabeads protein A- (for rabbit antibodies) or protein G- (for mouse antibodies) (Thermo Fisher Scientific, cat. number 10002D and 10004D, respectively). Tubes were rocked for ~1 hr at 4°C followed by two washes to remove unbound antibodies.\n\nPMA-treated THP-1 WT cells were collected in Pierce IP buffer (25 mM Tris-HCl pH 7.4, 150 mM NaCl, 1 mM EDTA, 1% NP-40 and 5% glycerol) supplemented with protease inhibitors. Lysates were rocked for 30 min at 4°C and spun at 110,000 × g for 15 min at 4°C. 0.5 ml aliquots at 2.0 mg/ml of lysate were incubated with an antibody-bead conjugate for ~1 hr at 4°C. The unbound fractions were collected, and beads were subsequently washed three times with 1.0 ml of IP lysis buffer and processed for SDS-PAGE and western blot on precast midi 10% Bis-Tris polyacrylamide gels. Protein A:HRP (MilliporeSigma, cat. number P8651) was used as a secondary detection system at a concentration of 2.0 μg/ml.\n\nPMA-treated THP-1 WT and STING1 KO cells were labelled with a green and a far-red fluorescence dye, respectively (Thermo Fisher Scientific, cat. number C2925 and C34565, respectively). The nuclei were labelled with DAPI (Thermo Fisher Scientific, cat. Number D3571) fluorescent stain. WT and KO cells were plated in a 96-well plate with optically clear flat-bottom. (Perkin Elmer, cat. number 6055300) as a mosaic and incubated for 24 hrs in a cell culture incubator, 37oC, 5% CO2. Cells were fixed in 4% paraformaldehyde (PFA) (Beantown chemical, cat. number 140770-10 ml) in phosphate buffered saline (PBS) (Wisent, cat. number 311-010-CL) for 15 min at room temperature and then washed 3 times with PBS. Cells were permeabilized in PBS with 0.1% Triton X-100 (Thermo Fisher Scientific, cat. number BP151-500) for 10 min at room temperature and blocked with PBS with 5% BSA, 5% goat serum (Gibco, cat. number 16210-064) and 0.01% Triton X-100 for 30 min at room temperature. Cells were incubated with IF buffer (PBS, 5% BSA, 0.01% Triton X-100) containing the primary STING1 antibodies overnight at 4°C. Cells were then washed 3 × 10 min with IF buffer and incubated with corresponding Alexa Fluor 555-conjugated secondary antibodies in IF buffer at a dilution of 1.0 μg/ml for 1 hr at room temperature with DAPI. Cells were washed 3 × 10 min with IF buffer and once with PBS.\n\nImages were acquired on an ImageXpress micro confocal high-content microscopy system (Molecular Devices), using a 20× NA 0.95 water immersion objective and scientific CMOS cameras, equipped with 395, 475, 555 and 635 nm solid state LED lights (lumencor Aura III light engine) and bandpass filters to excite DAPI, Cellmask Green, Alexa-555 and Cellmask Red, respectively. Images had pixel sizes of 0.68 × 0.68 microns, and a z-interval of 4 microns. For analysis and visualization, shading correction (shade only) was carried out for all images. Then, maximum intensity projections were generated using 3 z-slices. Segmentation was carried out separately on maximum intensity projections of Cellmask channels using CellPose 1.0, and masks were used to generate outlines and for intensity quantification.22 Figures were assembled with Adobe Illustrator.", "appendix": "Data availability\n\nZenodo: Antibody Characterization Report for STING1, https://doi.org/10.5281/zenodo.11582350. 18\n\nZenodo: Dataset for the stimulator of interferon genes protein (STING1) antibody screening study, https://doi.org/10.5281/zenodo.12761294. 19\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgment\n\nWe would like to thank the NeuroSGC/YCharOS/EDDU collaborative group for their important contribution to the creation of an open scientific ecosystem of antibody manufacturers and KO cell line suppliers, for the development of community-agreed protocols, and for their shared ideas, resources, and collaboration. Members of the group can be found below. We would also like to thank the Advanced BioImaging Facility (ABIF) consortium for their image analysis pipeline development and conduction (RRID:SCR_017697). Members of each group can be found below.\n\nNeuroSGC/YCharOS/EDDU collaborative group: Thomas M. Durcan, Aled M. Edwards, Peter S. McPherson, Chetan Raina and Wolfgang Reintsch\n\nABIF consortium: Claire M. Brown and Joel Ryan\n\nThank you to the Structural Genomics Consortium, a registered charity (no. 1097737), for your support on this project. The Structural Genomics Consortium receives funding from Bayer AG, Boehringer Ingelheim, Bristol-Myers Squibb, Genentech, Genome Canada through Ontario Genomics Institute (grant no. OGI-196), the EU and EFPIA through the Innovative Medicines Initiative 2 Joint Undertaking (EUbOPEN grant no. 875510), Janssen, Merck KGaA (also known as EMD in Canada and the United States), Pfizer and Takeda.\n\nAn earlier version of this of this article can be found on Zenodo (DOI: 10.5281/zenodo.11582350).\n\n\nReferences\n\nIshikawa H, Barber GN: STING is an endoplasmic reticulum adaptor that facilitates innate immune signalling. Nature. 2008; 455(7213): 674–678. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIshikawa H, Ma Z, Barber GN: STING regulates intracellular DNA-mediated, type I interferon-dependent innate immunity. Nature. 2009; 461(7265): 788–792. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMotwani M, Pesiridis S, Fitzgerald KA: DNA sensing by the cGAS-STING pathway in health and disease. Nat. Rev. Genet. 2019; 20(11): 657–674. PubMed Abstract \| Publisher Full Text\n\nZhang R, Kang R, Tang D: The STING1 network regulates autophagy and cell death. Signal Transduct. Target. Ther. 2021; 6(1): 208. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBarber GN: STING: infection, inflammation and cancer. Nat. Rev. Immunol. 2015; 15(12): 760–770. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDecout A, Katz JD, Venkatraman S, et al.: The cGAS-STING pathway as a therapeutic target in inflammatory diseases. Nat. Rev. Immunol. 2021; 21(9): 548–569. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZhang RX, Kang R, Tang DL: STING1 in sepsis: Mechanisms, functions, and implications. Chin. J. Traumatol. 2022; 25(1): 1–10. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGlass CK, Saijo K, Winner B, et al.: Mechanisms underlying inflammation in neurodegeneration. Cell. 2010; 140(6): 918–934. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHinkle JT, Patel J, Panicker N, et al.: STING mediates neurodegeneration and neuroinflammation in nigrostriatal α-synucleinopathy. Proc. Natl. Acad. Sci. USA. 2022; 119(15): e2118819119. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyoubi R, Ryan J, Biddle MS, et al.: Scaling of an antibody validation procedure enables quantification of antibody performance in major research applications. elife. 2023; 12: 12. Publisher Full Text\n\nCarter AJ, Kraemer O, Zwick M, et al.: Target 2035: probing the human proteome. Drug Discov. Today. 2019; 24(11): 2111–2115. PubMed Abstract \| Publisher Full Text\n\nLicciardello MP, Workman P: The era of high-quality chemical probes. RSC Med. Chem. 2022; 13(12): 1446–1459. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyoubi R, Ryan J, Bolivar SG, et al.: A consensus platform for antibody characterization (Version 1). Protocol Exchange. 2024.\n\nBiddle MS, Virk HS: YCharOS open antibody characterisation data: Lessons learned and progress made. F1000Res. 2023; 12(12): 1344. Publisher Full Text\n\nLaflamme C, McKeever PM, Kumar R, et al.: Implementation of an antibody characterization procedure and application to the major ALS/FTD disease gene C9ORF72. elife. 2019; 8: 8. Publisher Full Text\n\nAlshafie W, Fotouhi M, Shlaifer I, et al.: Identification of highly specific antibodies for Serine/threonine-protein kinase TBK1 for use in immunoblot, immunoprecipitation and immunofluorescence. F1000Res. 2022; 11: 977. Publisher Full Text\n\nStarr T, Bauler TJ, Malik-Kale P, et al.: The phorbol 12-myristate-13-acetate differentiation protocol is critical to the interaction of THP-1 macrophages with Salmonella Typhimurium. PLoS One. 2018; 13(3): e0193601. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRuiz Moleon V, et al.: Antibody Characterization Report for STING1 (Uniprot ID: Q86WV6).2024. Publisher Full Text\n\nAyoubi R, Laflamme C: Dataset for the stimulator of interferon genes protein (STING1) antibody screening study. [Data set]. Zenodo 2024. Publisher Full Text\n\nBandrowski A, Pairish M, Eckmann P, et al.: The Antibody Registry: ten years of registering antibodies. Nucleic Acids Res. 2023; 51(D1): D358–D367. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBairoch A: The Cellosaurus, a Cell-Line Knowledge Resource. J. Biomol. Tech. 2018; 29(2): 25–38. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nStringer C, Wang T, Michaelos M, et al.: Cellpose: a generalist algorithm for cellular segmentation. Nat. Methods. 2021; 18(1): 100–106. PubMed Abstract \| Publisher Full Text" }	[ { "id": "333635", "date": "07 Nov 2024", "name": "Karen Bowman", "expertise": [ "Reviewer Expertise Cell biology. Drug discovery and 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\nThe study involved characterisation of sixteen commercial antibodies, from nine different suppliers, for the stimulator of interferon genes protein (STING1) for use in western blot, immunoprecipitation, and immunofluorescence. The study formed part of the wider YCharOS collaboration to characterise commercial antibodies for human proteins using standardized protocols with the intention of improving antibody reproducibility issues. As a Data Note, it allows the direct comparison of the performance of commercially available antibodies to STING1 protein to aid scientists in choosing the most appropriate antibody for their studies. This would be of use in several fields such as study of innate immunity, autoinflammatory diseases, cancer and neurodegenerative diseases. A committee of industry and academic representatives have endorsed the platform used, and the protocols used appear to be consistent with those in general use. The platform consisted of identification of a human cell line suitable for antibody characterisation studies i.e., with adequate levels of the STING1 protein to generate a measurable signal. They found that the commercially available THP-1 WT cell line expresses the STING1 transcript at a level above the average range of cancer cells analysed, and therefore, it was a logical choice for their study. The matched isogenic knockout control cell line, THP-1 STING1 KO was used as an appropriate negative control. The final step was a series of antibody characterisation procedures, limited to the most common research uses of antibodies. The interpretation of the results is left to the reader, with the study providing unbiased guidance. Limitations of the study are clearly stated. The summary Table 3 with illustrations to assess antibody performance in western blot, immunoprecipitation and immunofluorescence, is very helpful. A minor point to note. An ‘at a glance’ summary table encompassing the outcomes for each antibody would be useful. In conclusion, the study will be a useful resource when choosing the most appropriate antibody for study of the STING1 protein.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and materials provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [] }, { "id": "333636", "date": "08 Nov 2024", "name": "Jieya Shao", "expertise": [ "Reviewer Expertise molecular and cellular 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\nIn this article the authors systematically compared the performance and specificity of sixteen commercial anti-STING1 antibodies using three different experimental approaches (Western blot, immunoprecipitation, and immunofluorescence staining). This study is part of a large collaborative initiative to characterize commercial antibodies for different human proteins and offer unbiased guidance to researchers who can select the best antibodies for their specific experimental objectives based on their own interpretations. As a Data Note, this paper successfully achieved its goal with rigorously collected high-quality data, and the results are self-explanatory and useful for the research community.\nHowever, there are a few minor points to address. First, the authors wrote in the Introduction section that “activated in response to bacterial cyclic dinucleotides, STING1 initiates a cascade of signaling events”. This statement seems incorrect as cGAMP can be generated endogenously by cGAS rather than derived from bacteria. Second, the statement that aberrant expression of STING1 can lead to cancer needs clarification. Third, in Fig.2, the bottom panel of images are larger than the rest. Fourth, the images in Fig.3 can be rearranged (e.g. 4 x 4) for better presentation.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and materials provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [ { "c_id": "12941", "date": "02 Jan 2025", "name": "Carl Laflamme", "role": "Author Response", "response": "Dear Dr. Shao, Thank you for taking the time to review our STING1 antibody characterization report. We greatly appreciate your feedback. This report is part of a global initiative aimed at characterizing antibodies for every human protein. While the authors are experts in antibody development, we acknowledge that our expertise does not extend specifically to the STING1 protein. In light of your comments, we have revised the introduction to ensure greater accuracy and clarity regarding STING1. Additionally, we have modified Figures 2 and 3 in accordance with your recommendations. We trust these changes address your concerns and enhance the quality of our report. Thank you once again for your thoughtful insights. Best regards, Carl" } ] } ]	1	https://f1000research.com/articles/13-1049
https://f1000research.com/articles/14-14/v1	02 Jan 25	{ "type": "Research Article", "title": "Experiences of Students with Disabilities Enrolling in Higher Education: A Qualitative Study in the United Arab Emirates", "authors": [ "Muhammad Arsyad Subu", "Richard Mottershead", "Ahmed Mahmoud Sallam", "Jacqueline Maria Dias", "Janisha Kavumpurath", "Fatma Refaat Ahmed", "Heba Hesham Hijazi", "Mini Sara Abraham", "Alounoud Mohamed Al Marzouqi", "Vidya Seshan", "Shahd Mohammed Elabed", "Ahmad Rajeh Saifan", "Syed Azizur Rahman", "Nabeel Al Yateem", "Muhammad Arsyad Subu", "Ahmed Mahmoud Sallam", "Jacqueline Maria Dias", "Janisha Kavumpurath", "Fatma Refaat Ahmed", "Heba Hesham Hijazi", "Mini Sara Abraham", "Alounoud Mohamed Al Marzouqi", "Vidya Seshan", "Shahd Mohammed Elabed", "Ahmad Rajeh Saifan", "Syed Azizur Rahman", "Nabeel Al Yateem" ], "abstract": "Background Higher education is experiencing an increase in students with disabilities, necessitating considering their interests and well-being when preparing educational services. Students with disabilities make up one of the most significant minority groups among students in terms of diversity.\n\nAims This study explored the experiences of students with disabilities at a university in the United Arab Emirates. It also aimed to understand students with disabilities’ social and academic experiences and the factors affecting their university persistence.\n\nMethods We used a descriptive qualitative research approach to understand how students with disabilities experience the phenomenon under study. Twenty-five students participated, chosen via purposeful sampling. The data was collected using semi-structured interviews, which were analyzed using content analysis.\n\nResults We found four themes that emerged from this study: (1) perceptions about the university, (2) socialization and integration, (3) barriers to the university journey, and (4) support for the journey.\n\nConclusion We found that students with disabilities indicated positive and negative perspectives and experiences. We recommend modifying the physical, social, and educational environments to support the education of students with disabilities. Future studies should employ a more varied sample and include academics and students to collect different perspectives while stressing the students’ experiences.", "keywords": [ "students with disabilities", "university", "higher education", "qualitative", "United Arab Emirates." ], "content": "Introduction\n\nDisability is the condition in which a person’s ability to engage in activities is limited due to their interactions with the environment. Students with disabilities continue to face systematic disadvantages in comparison to their non-disabled peers. Hence, educational practitioners and policymakers must fully understand the intricacy of the encounters of impaired students (Brewer et al., 2023). Some types of disabilities include hearing loss, movement problems, mental health issues, communication problems, and learning difficulties. The enrollment of students with disabilities in higher education has increased (National Center for Education Statistics, 2022).\n\nIn order to combat the systemic discrimination that people with disabilities experience, structural interventions are necessary for students with disabilities (Ostiguy-Finneran et al., 2018). The rights of students with disabilities in higher education are becoming more widely known internationally (Taneja-Johansson, 2021), particularly considering an increase in the proportion of university students with disabilities (Zhang et al., 2018). Higher education institutions must empower the disabled population, emphasizing student accessibility as these students grow. For students with disabilities, getting into higher education remains challenging (Fernández-Batanero et al., 2022). Students with disabilities have a range of difficulties, such as physical, sensory, learning, attention, and communication challenges (Burgstahler, 2020).\n\nThe educational environment and instructors’ capacity to modify it to be congruent with students’ skills are problems for students with disabilities. They continue to get unfair treatment while attending universities (García-González et al., 2020). Due to difficulties with accommodation offered by universities, students with disabilities continue to face barriers when trying to access possibilities for a fair education (Ehlinger & Ropers, 2020; García-González et al., 2020; Magnus & Tøssebro, 2014). They encounter problems, including prejudice and stigmatization, occasionally hindering their learning capacity (Carroll et al., 2020; Subu et al., 2024). Physical environment and academic and administrative procedures are critical barriers to inclusive educational practices that university students with disabilities must overcome (Algolayalt et al., 2023). Many students with intellectual disabilities struggle to finish their coursework and lack the intellectual and social skills necessary to fit in the universities (Prohn et al., 2019) despite the multiple university education programs available to them.\n\nThe United Arab Emirates (UAE) adopted the international movement regarding the rights of individuals with disability by introducing a federal law in 2006 to govern the rights of people with disabilities, especially those related to education. Federal Law No. 29 of 2006 is the first legislation in the UAE to safeguard the rights of individuals with disabilities. According to Article 12 of the law, the country ensures that individuals with special needs have equal access to education in all educational institutions, including regular and special classes. This includes providing a curriculum in sign language, braille, or other suitable methods (United Arab Emirates Government, 2023). Because of this, the UAE Ministry of Education launched a new program to include students with special needs in regular classrooms (Gaad & Almotairi, 2013).\n\nSpecial education is specifically designated for children who have a disability, problem, or any other circumstance that affects their capacity to learn or perform academically. In addition, people with disabilities are also ensured equitable chances and appropriate adjustments to pursue their studies in mainstream and specialized classrooms (UNESCO, 2018). The UAE government has built assistance centers around the UAE to monitor the academic success of people with special needs before and after school integration. These centers offer various services, including assessing students with disabilities and learning challenges, making recommendations, helping parents navigate their child’s situation, and advising on appropriate support services for their children (UAE government, 2023).\n\nWithin the Disability-Diversity (Dis)Connect model (Aquino, 2016), two forms of disability diversity manifest in university settings. These models show how students with disabilities can be included and deal with comparable problems as students who identify with other diverse memberships or who cannot. Every experience a student has with disability and diversity is related to how they view their impairment’s role in their life and the university setting. First, Disability-Diversity Disconnect: Because they find it difficult to integrate their disability into the higher education environment and because their experiences differ from those of students who identify with other diversity memberships, students with disabilities do not want to participate fully (or intentionally) in the postsecondary environment and do not integrate. Students of this kind have a difficult time adjusting to life in a university setting. The student does not believe that disabilities are accepted in college, especially when they are a part of student diversity. Second, Disability-Diversity Connect: The student with a disability views the disability as just one of many elements that make up their identity and place in a postsecondary environment. Furthermore, one may consider disability in the postsecondary setting a part of diversity. According to the student, disability is fully integrated into the diversity of students and is a crucial aspect of the postsecondary environment. Positive experiences with self-development, a desire to engage with the postsecondary community, and the accomplishment of social and academic objectives all contribute to feelings of disability and diversity inclusion.\n\n\nResearch aims\n\nAn extensive literature review indicates limited information regarding the experiences of students with disability currently enrolled in higher education institutions in the UAE.\n\nThis study explored the experiences of students with disabilities presently enrolled in higher education institutions in the UAE. Understanding the disparity in enrollment and degree completion rates for students with disabilities is essential for researchers, policymakers, and faculty members (Erickson et al., 2014). The findings of this study may also assist educators in creating successful techniques to address the obstacles faced by students with disabilities. Organizations and decision-makers may use the results of this study to develop plans for meeting the social and academic requirements of university students with disabilities.\n\n\nResearch questions\n\nThe main research question is: What are the experiences of disabled students at a higher education or university?\n\n\n\n1. What do students with disabilities identify as positive and negative experiences studying in the university setting?\n\n2. How do the negative and positive experiences make students with disability feel?\n\n3. What do students with disabilities see as barriers and facilitators to their access to the university?\n\n\nMethods\n\nThe purpose of this study was to explore the experiences of students with disabilities enrolling at the University of Sharjah in the United Arab Emirates. This study adopted qualitative content analysis methods. Using qualitative methods allowed for an in-depth exploration of students’ lived experiences, providing a deeper understanding of their perspectives, challenges, and unmet needs. Researchers can better comprehend the complex and comprehensive nature of the topic they are studying by using the qualitative research design (Creswell, 2012). According to Creswell and Poth (2018), qualitative research focuses on the substance and meaning of the world using interpretive and naturalistic methodologies. Content analysis is an interpretive process that focuses on the topic and background and examines similarities and differences between and within various text areas (Graneheim & Lundman, 2004).\n\nThis research was conducted at a university in the United Arab Emirates. A total of 25 students with disabilities were recruited and selected using purposeful sampling methods. None of the participants refused to participate. The researchers recruited all study participants, and only students who experienced disability were included. The study participants were aged between 19 and 42 years old. Of 25 participants, 20 were males, and 5 were females. The most common disabilities among the student participants were learning difficulties, writing and reading difficulties, speech problems, hearing disabilities, and mobility impairments. Blindness and mobility issues were present in some of the participants, while other students struggled with their learning and writing. Most students who participated studied in the College of Humanities, Arts, and Social Sciences. For inclusion criteria, students were included if they had attended the university full-time or part-time and were at least 18 years old. In this study, we only recruited students with disabilities. A physician or psychiatrist must have determined that, as an individual with a disability, he/she has a physical or mental impairment. In this study, most of our participants experienced physical disabilities. They experienced learning, writing, reading, speech, hearing, blindness, and mobility problems. For exclusion criteria, students with severe anxiety or depression and those with severe mental health or psychiatric symptoms of their disabilities were excluded.\n\nThe primary data collection method was semi-structured interviews. All interviews were conducted at the Disability Resource Center at the University of Sharjah in UAE. The first, second, and third authors, PhD holders, and qualitative researchers collected the data. All interviewers are male faculty members at the University of Sharjah. Interviews were audiotaped using a digital recording device with participants’ consent. Each interview lasted between 35 and 55 minutes. In this study, data saturation was achieved at the 24th interview. Memos and field notes were also two components of the data collection techniques. These data collection methods were formed by triangulating data to improve the credibility of interpretations of the data (Lincoln & Guba, 2017). These additional data collection methods were essential in guiding us in interpreting the participants’ experiences, feelings, perspectives, and beliefs. Before creating this interview guide, approval was received from the University of Sharjah’s ethical committee. Methodological methods should be adhered to while ensuring the quality assurance of an interview for a scientific application. We started our study by reading academic journals focusing on students with impairments, especially those attending universities. A face-validation procedure was conducted by a panel of three experts from various programs and disability centers at the University of Sharjah after the first draft of the interview guide was created. These specialists were experienced educators with a history of working with children with disabilities on research projects and in publications. Several changes were made to the interview guide in response to their suggestions. Before the interview, it is critical to ensure that the room in which the interview was held is fully accessible to wheelchair users and has sufficient space to allow the participants to move around while in the room.\n\nIn this study, we adopt the content analysis approach (Graneheim & Lundman, 2004) to analyze the data. Using this data analysis methodology, we could look into the differences and similarities within and across several data sets (Graneheim & Lundman, 2004). In this study, NVivo software was used to manage the data. The audio recordings of each interview were attentively listened to and then transcribed immediately after the interview. To guarantee consensus and to ensure that everyone on the research team had an in-depth understanding of the topic, each interview’s transcript was read and reread several times. Then, themes resulting from the data were conceptualized, defined, and developed using open coding. All researchers agreed on the themes obtained after analyzing the codes derived from the interview data. Then, the relationships between the themes that had been extracted were determined using conceptual patterns and tables. We took the responses from the data related to the students’ experiences. All codes and units relevant to this study were analyzed and evaluated regarding their similarities and dissimilarities. Each study team member shaped the significant themes and subthemes, ultimately abstracted and categorized.\n\nStudy rigor\n\nThe study’s reliability was enhanced by gathering data and information from multiple sources, including interviews, field notes, and memoranda. We meticulously examined the data utilizing NVivo coding and examined each word and line in detail. Upon identifying areas of ambiguity during the data analysis process, we revisited a select group of student participants to validate and expand upon their viewpoints. Furthermore, five additional researchers (AS, RM, MS, FRA, and SAR) have independently verified the data analysis methods. The study published the criteria used to inform the researchers’ judgments and the process used to choose the study participants, thereby enhancing auditability. The research methodology and the results of our inquiry were meticulously recorded. We maintained a notebook to record the subjective choices and their corresponding reasons or rationales. The transferability of the study was guaranteed by presenting comprehensive details on the participants, setting, and study results, enabling readers to evaluate the study based on their own experiences.\n\n\nResults\n\nIn this study, we identified four distinct but connected themes that emerged from the interviews and data analysis that described the experiences of students with disabilities enrolling in a university: 1) perceptions about the university; 2) socialization and integration; 3) barriers to studying in the university; and 4) support in the university journey. In addition, theme 1 has one sub-theme. Theme 2 has two subthemes. Theme 3 has three subthemes, and theme 4 has eight subthemes.\n\nSubtheme: Positive experiences studying at the university\n\nPositive experience is an aspect that determines the success of students with disabilities in pursuing education at university. Generally, participants indicated that they had had positive experiences at the university.\n\nI have positive experiences from studying at UOS… In addition, the courses that talk about our general life and history provide us with many exciting and valuable courses. Also, … I meet my friends and colleagues daily, and I am very happy to have my friends. I am also satisfied with the help of the Disability Center and the interpreters. (Participant 3)\n\nSubtheme: Negative experiences studying at the university\n\nHowever, some participants have had negative classroom experiences. According to one participant, he usually has problems with some professors.\n\nAmong the problems I usually face during my studies, in general, are dealing with some professors … because some could not differentiate that not all students with disabilities are the same. At first, everything was simple, but now there are some challenges… (Participant 15)\n\nSubtheme: Relationship with faculty members and staff\n\nSocial interaction with others benefits the development of one’s abilities to communicate and interact. Study participants agreed that the university had a welcoming social atmosphere, which helped social integration. Participants discussed their interactions with peers and lecturers. They described having positive relationships with their professors.\n\n… Many professors know my condition; others around me also know it. As a result, things are simple, and the doctors usually deal with us and collaborate easily. Even though, sometimes, doctors give us an extra assignment… It is fine, and I am so happy at the university. (Participant 4)\n\nSubtheme: Peer relationship\n\nPeer relationships are one factor determining student success in pursuing education. Activities on campus require harmonious relationships between students. We found that most participants had good relationships with their friends at the university. Some made connections and friends when they joined the university. Some students with disability communicate with friends with an interpreter.\n\n… Initially, joining the university was impossible, but I made new connections and friends when I joined. I could communicate with everyone, including professors and especially my friends, at any time, but I also communicated with the interpreter. Some students were close to me, so I have friends now… I have three best friends, and we share information… (Participant 17)\n\nBarriers to higher education for individuals with disabilities exist on many different levels. Each participant encountered numerous obstacles in university, including barriers to professors, exams, questions, and accessibility issues.\n\nSubtheme: Professors as a barrier\n\nSome participants indicated that professors are a barrier to their university journey. One participant said that he had a negative experience with his professor.\n\nDuring my undergraduate studies, I encountered a professor who did not understand my case, and to be honest, he disappointed me and caused my GPA to drop. Apart from hearing difficulty, I face many other complex problems. For example, some professors (two or three) needed help understanding my case…. (Participant 19)\n\nSubtheme: Exams and levels of questions as barriers\n\nStudy participants indicated that exams and levels of questions were barriers. They struggled with exams at the university, especially when they written questions. One study participant indicated that exams (quizzes, midterms, and final exams) are barriers during their education at the university.\n\n… Sometimes, I struggle with exams. However, I should not take these as a barrier that may stop me from completing my journey. In general, I see that our exams are so challenging. During the exams sometimes, it is tough to solve the exams. My challenge was when I had to solve written questions… (Participant 18)\n\nSubtheme: Campus accessibility as a barrier\n\nCampus accessibility is essential for students with disabilities. Some students indicated that they have problems with parking spaces. According to a participant:\n\nI find parking in certain places makes me struggle with… parking is complicated. Wheelchair. Especially during exams, some non-students with disabilities park in our parking. I need space because I am a student with disability and mobility impairment and I need space … I hope that we have special parking for deaf students. (Participant 2)\n\nEven though students with disabilities encounter many challenges or other difficulties along the way, the majority of participants said they receive support. Their journeys’ success correlates with university facilities, academic performance, and parental support. Interviews with students demonstrated that institutional assistance encourages academic persistence among students with disabilities\n\nSubtheme: University Facilities support\n\nIn our interview, we found that accessibility to university facilities is essential for students with disabilities. One of the participants indicated that all the facilities in the university are accessible or easy for them to use.\n\n…All the facilities in the university are accessible, so I have no issue with reaching the university’s facilities. I do not run into any issues. No problem with facilities… everything goes as I plan. I did not face any difficulties while attending the campus. I was grateful for every time I entered the facilities. (Participant 12)\n\nSubtheme - Disabilities Center service’s support\n\nThe university’s support services for students with disabilities were essential in guaranteeing that students could succeed academically. The student has benefited from the Disability Resources Center, which helped, advised, and supported the university’s students. Students indicated they needed to ensure they had access to these disability services.\n\nDisability Resources Center has helped me a lot. They advised, guided, and facilitated everything for me and my classmates. Also, they encouraged me… and told me what I should gain from the university and what goals I need to achieve throughout the university years. (Participant 17)\n\nSubtheme: Academic and Curriculum support\n\nParticipants acknowledged that their professors were essential to their academic performance. Students repeatedly stated that professors’ responses and knowledge can affect access and equity. Participants realized that they were not rejected or refused when they requested assistance. An autistic participant described his encounters with his lecturers as mostly favorable experiences.\n\nI am glad to see my professor in the classroom. I am grateful for his constant support. Like one of my professors, he supports me a lot. We succeeded. Like, none of my instructors ever actually objected to me… (Participant 10)\n\nSubtheme: Professor’s support\n\nEducators or professors in the classroom constantly assisted the students. According to participants, university professors give full attention and support to the students and are willing to help them, even when they are sick, unable to attend, or absent.\n\nThe professors were good, and because they supported me, everything became more accessible. Dr. AA helped us a lot. Also, Dr. YN and Dr. AS (faculty members) … help me and solve my problem immediately when I see them. They assisted and supported us well. I am so happy, and I was trying as much as possible to do both theory and practice and focus on studying and attending my classes. (Participant 14)\n\nSubtheme: Counseling support\n\nParticipants indicated that a counseling center is essential for students with a disability at the university. Participants reported that the university counseling center offers stress management services to support stress-reduction methods. They receive problem-solving instruction from the school’s psychologists on how to handle difficulties. Participating in campus stress management programs helped students prepare for stressful classes and other campus activities.\n\nYes, counseling center …. It helps me with stress management. They (counselors) provide relaxation exercises and other stress management techniques. There are some problems, and I attend stress management programs to overcome my challenges. It works very well. (Participant 12)\n\nSubtheme: Peer support\n\nPeer support is essential for students with disabilities. Students indicated that they have support from their friends. Working together in groups, for example, requires support from other students, especially non-students with disabilities.\n\nI usually ask my colleagues, the hearing-impaired students, and other friends to assist me with my academics when I do not comprehend something. Even people who are not disabled would try to help me. We have a WhatsApp group where we assist one another by notifying, updating, and contacting each other when something new occurs… Members of this group regularly converse, translate, and interpret everything being sent. (Participant 4)\n\nSubtheme: Family support\n\nStudents with disabilities face challenges during their university journey, and support from their family members will be necessary. Study participants indicated that they have support from their family members. One of the participants indicated that their family supports them.\n\n… I want to say thanks to my wife. My wife is also deaf, so things were easy, and she helped me with my studies. I used to think it was impossible to study at university, but my wife constantly encouraged and supported me. Although things started hard, they improved over time, and I am grateful to those who supported me… (Participant 13)\n\nSubtheme: Financial Support\n\nBecause they are eligible for scholarships, students are satisfied to attend the university. Financial aid helps these students pay for some of their tuition, handouts, and textbooks. Several participants said they stay in school when their financial needs are satisfied. The college supports some students with disabilities by providing financial aid. Some students mentioned getting financial assistance to cover some of their tuition costs.\n\nThe financial aid I receive enables me to purchase textbooks, handouts, and a portion of my tuition without worrying about where I will find the money to do so… Yes, I am glad to study at the university and get the scholarship… I would like to take this opportunity to thank H.H SJ [Queen] and sheik Dr. SQ [king) of S Emirate] for this opportunity and scholarship. (Participant 11)\n\n\nDiscussion\n\nStudent with disabilities constitute a significant and growing minority group in higher education, contributing to the overall variety of the student population (Römhild & Hollederer, 2023; Mottershead et al., 2015). Our study results indicated that students have positive perceptions of studying at the university and indicated an excellent perception of the university. These findings support Clavijo Castillo and Bautista-Cerro (2020), who found that higher education is essential to building a learning environment that can contribute equality solutions to difficulties for those with disabilities in the educational setting. We also found in our study that some students had a negative perspective and impression of the university. The perceptions of students with disabilities on the services and initiatives offered by universities are crucial factors that influence these students’ performance (Cawthon & Cole, 2010). Most students who had issues with university teachers nonetheless expressed satisfaction with their college experience (Yssel et al., 2016). A student’s sense of belonging and capacity for self-advocacy contribute to better satisfaction and retention (Fleming et al., 2017).\n\nStudy participants agreed that the university had a welcoming social atmosphere, which helped social integration. Similar to our findings, integrating classmates and educators, campus activities, and extra-curricular activities for students with disabilities increased success in school (Accardo et al., 2019). The participation of students with disabilities in social activities promotes successful academic integration. According to Choi et al. (2019), university students with disabilities who are academically and socially integrated are more likely to stay in school and less likely to drop out. In this context, academic integration entails communication with and assistance from staff members and educators (Römhild & Hollederer, 2023). In this study, participants shared their positive experiences regarding relationships with faculty members. Other studies noted similar findings to our results. For example, disability-aware professors frequently assist students with disabilities in learning, solving problems, and passing classes with little to no difficulties (Mutanga & Walker, 2017; Timmerman & Mulvihill, 2015). When educators know how to help students, they can experience favorable social implications (Agarwal et al., 2015). The best opportunity for successful academic outcomes for students with disabilities is to form positive relationships with their teachers and peers (Mutanga & Walker, 2017).\n\nThe participants’ relationships included interactions with peers, faculty, and staff. Study results indicated that participants have good relationships with their friends. When participants had positive contact with their colleagues, they felt a sense of belonging. These interactions also help students with disabilities feel successful. Some studies show difficulties when disabled university students interact socially with peers (Nel et al., 2015; Schwab et al., 2018). Stereotypical threats, stigmatizations, and marginalization are among the difficulties students face with social participation (Desombre et al., 2018; Subu et al., 2023). According to French (2017), university students with disabilities have a sense of belonging when they connect favorably with fellow students and teachers during and after class. When their peers know how to help them, students with disabilities can experience favorable effects in society (Agarwal et al., 2015).\n\nOur study results indicated that students with disabilities experienced several challenges during their journey at the university. Higher education is crucial for building an academic environment that can contribute equality solutions to challenges for students with disabilities in inclusive learning environments (Clavijo Castillo & Bautista-Cerro, 2020). A study indicated that it is highly challenging for individuals with disabilities to enroll in a university (Fernández-Batanero et al., 2022). In contrast to non-students with disabilities, other studies have indicated that academic distress, a lack of social support, and a lack of campus involvement are challenges (Fleming et al., 2017). Disabled university students who attend universities employ coping mechanisms like self-awareness, self-advocacy, and tenacity and resilience to deal with academic difficulties (Hengen & Weaver, 2018; Ju et al., 2017).\n\nOther studies highlight other learning challenges. The absence of training for instructors to employ an approach that encourages inclusion in the classroom by their students’ requirements stands out among them (Heiman et al., 2017). Our findings align with another study on how these students do not receive teacher preparation in higher education (Nimante et al., 2021). Additionally, students are informed how difficult it is to get material resources because, for the most part, they are either inadequate or unsuitable for their needs (Alsalem & Abu Doush, 2018; Dreyer, 2021). Our study results also indicated that exams (quizzes, midterm, and final exams) are barriers during their education at the university. Our participants found that questions during exams are difficult for them regarding level. The results of this study are generally in line with those of another study that found that students with disabilities encountered significant educational challenges related to tests and information access because of a lack of support (Kisanga, 2020).\n\nThe university’s services for student with disabilities ensured their success there. Campus accessibility is essential for students with a disability at the university. Study results indicated that some students have a problem related to parking space, and other participants have difficulty accessing sidewalks. This result is in line with a study in which students with disabilities complained about the inadequate infrastructure and facilities at the university, as well as their difficulty accessing educational resources as a result of these problems (Nel et al., 2015). As these challenges can impede accessibility for these students, students with disabilities exhibit educational demands that must be addressed to attend education successfully (Braun & Naami, 2019). The results of this study are also in line with another study’s findings, which found that students with disabilities encountered considerable educational challenges related to accessibility (Kisanga, 2020).\n\nAdditionally, conventional architectural difficulties such as classrooms, stairs, insufficient auditoriums, heavy doors, malfunctioning elevators, or the lack of ramps and signage continue to be barriers to higher education (García-González et al., 2020). Accessibility to the university facilities is essential for students with disabilities. Study results indicated that participants indicated that all the facilities in the university are accessible or easy for them to use. Participants talked about how the school’s learning support programs encourage them to persevere, challenge them to explore options, and help them stay on track academically. Similar to this finding, university settings support the persistence of students with disabilities by enhancing their learning and academic performance, offering persistence programs like stress management and student success training, providing them with aid, and equipping them with resources (Mamiseishvili & Koch, 2012).\n\nThe study’s findings showed that the university’s disabilities center offers support to students, and it is crucial for them to make sure they have access to these services before beginning their academic studies. Previous studies have demonstrated that disability-related services, such as disability offices, positively impact graduation intentions and grade point averages (Singley, 2017). According to Brophy-Felbab (2021), disability-related services are crucial for student retention and performance. However, no appreciable associations exist between grade point average and disability-related services (Santos et al., 2019). Study participants claimed that the university program accommodated and enhanced their learning despite their particular limitations. According to the study’s findings, participants agreed that their educators were crucial to their academic success. It can be challenging for many students with disabilities to benefit from essential initiatives that improve learning outcomes (Boney et al., 2019). Similar issues with curriculum design are among the difficulties colleges face as they deal with the disability issue (Everett & Oswald, 2018). Developing inclusive learning materials should receive more vital support from teaching staff and curriculum designers (Fleet & Kondrashov, 2019), with universal design serving as a common framework for such efforts (Grier-Reed & Williams-Wengerd, 2018).\n\nStudy results indicated that professors at the university provide full attention and support to the students, and they are all willing to support them. Similar to these results, educators provide support to students with disabilities. The principle of inclusion depends on academics who value and assist students with disabilities in the classroom. Participants in this study reported that faculty members kept their attention during and after class by offering them opportunities to explore, ask questions, and work out problems. Most of the class is ensured to participate through discussion, demonstrations, and questions. According to Francis et al. (2019), instructors’ negative attitudes about disability are the cause of the mistrust and cynicism that exist between students with disabilities and their educators. Faculty members aware of disability concerns are better equipped to give students with disabilities the help they need during the teaching and learning process (Harnett, 2016). To succeed, students with disabilities need peer support. For instance, working in a group requires the assistance of other students, especially non-students with disabilities. Our research shows that during class discussions and project presentations, peers assist students in finding answers to questions. Students with disabilities retain positive relationships with their classmates and exchange unique and crucial information during and after lecture hours, which is consistent with our findings (O’Shea & Meyer, 2016).\n\nIn this study, participants stated they would remain in the program until their financial necessities were met. The university provides financial support for specific numbers of its students with disabilities. A few students claimed they were receiving financial aid to help with the expense of their tuition. Some participants indicated that receiving financial aid allowed them to get intellectually assimilated into the institution since it allowed them to purchase study materials, learn, and study in groups. We found that our participants indicated that their family supports them. They benefited from their relationships during university, which made for a better higher education experience. It has been recognized that providing these individuals with equal opportunity to non-disabled people in families and the greater community has a positive effect on the education of children with disabilities (Ceka & Murati, 2016). Although family and partner support does not directly correlate with academic success, it does increase students’ overall satisfaction (Lombardi et al., 2016). Parents who supported the idea of higher education for all—both literate and uneducated—supported and promoted their disabled children’s university education (Hegna & Smette, 2017).\n\nThe participants in this study only represented one university in the United Arab Emirates, which was the main limitation. First, inclusive education approaches for students with disabilities may vary among universities in the UAE. The limitation of qualitative research, like this study, is that it cannot be generalized. The generalizability of this study’s findings outside this particular institution can be problematic due to its exclusive focus on the experiences of students with disabilities within this institution. Despite these limitations, this research has shed light on how inclusive policies and practices are perceived and understood in higher education institutions in the UAE. A larger population sample should be used to have more significance and beneficial implications. In addition, using a range of geographic locations would be advantageous for this research issue.\n\n\nConclusion\n\nThis study shares the stories of students with disabilities and their experiences at the university. To accommodate these students, universities must modify their educational programs and offer services consistent with best practices. To enable these students to attend classes at university, a variety of infrastructure, teaching-learning, and institutional management-related factors are required. Promoting the success of students with disabilities in university education will only be possible by making higher education institutions more accessible, providing training for university faculty, and increasing community awareness of inclusive education in higher education. Enhancing the function of student support centers, which have the trained personnel and tools required to support the education of students with disabilities, will result in better implementation of inclusive education methods. This study looked into a specific instance of inclusive education techniques being applied to students with disabilities in a model higher education environment in the UAE rather than making a broad conclusion. Additionally, the study recommends modifying the physical, social, and educational environments to support the education of students with disabilities. Our research also indicates that to promote equity, university faculty members and staff need to be trained to assist students with disabilities. Implementing instructional strategies grounded in design for Learning may remove future challenges to learning, benefiting not just students with functional diversity but also other students. Enhancing instruction for students with disabilities has a favorable effect on education for all students. This study highlights some topics that need more significant investigation to develop best practices and a more profound knowledge of students with disabilities and to create a more inclusive environment for them. Further studies may provide additional context for these results, which may be applied to the entire student population with disabilities. Research on disability identification, faculty perspective, and the efficacy of involvement is especially recommended. With the results of this further research, attempts to provide a more equal education for all children will significantly increase the inclusion of students with disabilities. Future research should use a more diverse sample, involve academics and students, and emphasize the students’ experiences to get data from various perspectives and experiences.\n\n\nEthical consideration and consent\n\nWe realized that for this study, ethics approval is necessary. Research Ethics Committee (REC) of the University of Sharjah approved this study (Reference number: REC-21-09-22 (VERSION 2) on 10th September 2022. All study participants were made aware of the research’s purpose and methodology. We followed specific methods to obtain consent from visually impaired participants. Before data collection, the appropriate consent form was presented orally to the participants. We also used an audio recording of the informed consent document. The guidance for obtaining and documenting informed consent for deaf participants was also followed for those who cannot read or write. Hearing-impaired participants may need an interpreter present. To ensure privacy and comfort, we selected interview environments that could provide a quiet, private space to make all participants feel comfortable and confident. We made sure that students with disability who participated in this study felt secure during the interview. To preserve confidentiality, we provided information about the aim and method of the research to all participants with written informed consent, and they were assured of anonymity and confidentiality. Participants in the study must be assured that their data is kept anonymous unless they give their full consent.\n\n\nShort authors’ biography\n\nMuhammad Arsyad Subu: Specialization: Mental health nursing. Research: Mental health nursing and nursing education. Richard Mottershead: Specialization: Mental health nursing. Research: Mental health interventions and social care needs. Ahmed Mahmoud Sallam: Specialization: Counseling psychology. Research: Students with disabilities, rehabilitation therapies. Jacqueline Maria Dias: Specialization: Oncology and geriatrics nursing. Research: Curriculum design and evaluation simulation & clinical teaching and learning. Janisha Kavumpurath. Specialization: Adult nursing. Research: Basic life support among adults. Fatma Refaat Ahmed. Specialization: Critical care and emergency nursing. Research: critical care nursing and critical care hematology nursing. Heba Hesham Hijazi. Specialization: Public health management and policy. Research: Quality of healthcare services, health policies, and utilization of healthcare services. Mini Sara Abraham: Specialization: Medical Surgical Nursing. Research: Nursing administration quality COPD. Alounoud Mohamed Al Marzouqi. Specialization: Health care management, quality, excellence. Research: Leadership & health information management. Vidya Seshan: Specialization: Maternal Health. Research: Women’s and Maternal Health. Shahd Mohammed Elabed: Health care management. Research: Management of infectious diseases. Ahmad Rajeh Saifan. Specialization: Emergency nursing education. Research: Diabetes in adults, emergency hospitalization. Syed Azizur Rahman. Specialization: Health care delivery system, public health, and health care evaluation. Nabeel Al-Yateem. Specialization: Pediatric nursing. Research: Children with chronic illness and school nursing care.", "appendix": "Data availability statement\n\nDerived data supporting this study’s findings are available from the corresponding author [RM] upon reasonable request. 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Current Psychology. 2020; 41(7): 4750–4759. Publisher Full Text\n\nLincoln Y, Guba EG: The SAGE Handbook of Qualitative Research. Newbury Park, CA: SAGE Publications Inc; 2017.\n\nLombardi A, Murray C, Kowitt J: Social support and academic success for college students with disabilities: Do relationship types matter? Journal of Vocational Rehabilitation. 2016; 44: 1–13. Publisher Full Text\n\nMagnus E, Tøssebro J: Negotiating individual accommodation in higher education. Scandinavian Journal of Disability Research. 2014; 16(4): 316–332. Publisher Full Text\n\nMamiseishvili K, Koch LC: Students with disabilities at 2-year institutions in the United States: Factors related to success. Community College Review. 2012; 40(4): 320–339. Publisher Full Text\n\nMottershead R, Kumardhas V, Masulani Mwale C: Mental health within different ethnic groups and minorities. Mental Health Across the Lifespan. Steen M, Thomas M, et al., editors. Routledge; 2015; pp. 46–59. Publisher Full Text\n\nMutanga O, Walker M: Exploration of the academic lives of students with disabilities at South African universities: Lecturers’ perspectives. African Journal of Disability. 2017; 6: 1–9. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNational Center for Education Statistics: Students with disabilities. U.S. Department of Education, Institute of Education Sciences; 2022. Reference Source\n\nNel K, Rankoana SA, Govender I, et al.: The challenges experienced by students with a physical disability (SWPD) at a higher education institution in South Africa. African Journal for Physical, Health Education Recreation and Dance. 2015; 1(4): 801–811. Reference Source\n\nNimante D, Baranova S, Stramkale L: The university administrative staff perception of inclusion in higher education. Acta Paedagogica Vilnensia. 2021; 46: 90–104. Publisher Full Text\n\nO’Shea A, Meyer R: A Qualitative Investigation of the Motivation of College Students with Nonvisible Disabilities to Utilize Disability Services. The Journal of Postsecondary Education and Disability. 2016; 29: 5–23.\n\nOstiguy-Finneran B, Peters ML, et al.: Ableism. Readings for Diversity and Social Justice. Adams, et al., editors. New York: Routledge; 2018.\n\nProhn SM, Kelley KR, Westling DL: Supports’ perspectives on the social experiences of college students with intellectual disability. Inclusion. 2019; 7(2): 111–124. Publisher Full Text\n\nRömhild A, Hollederer A: Effects of disability-related services, accommodations, and integration on academic success of students with disabilities in higher education. A scoping review. European Journal of Special Needs Education. 2023; 1–24. Publisher Full Text\n\nSantos SBD, Kupczynski L, Mandy M: Determining Academic Success in Students with Disabilities in Higher Education. International Journal of Higher Education. 2019; 8(2): 16–38. Publisher Full Text\n\nSchwab S, Nel M, Hellmich K: Social participation of students with special educational needs. European Journal of Special Needs Education. 2018; 33(2): 163–165. Publisher Full Text\n\nSingley DM: Effects of academic coaching on college students with learning disabilities or attention-deficit hyperactivity disorder (Order No. 10278516). ProQuest One Academic; 2017. (1914924962). Reference Source\n\nSubu MA, Wati DF, Al-Yateem N, et al.: ‘Family stigma’ among family members of people with mental illness in Indonesia: a grounded theory approach. International Journal of Mental Health. 2023; 52(2): 102–123. Publisher Full Text\n\nSubu MA, Dias JM, Mottershead R, et al.: Exploring mental health stigma among Indonesian healthcare students towards individuals with mental illnesses: a qualitative study. International Journal of Qualitative Studies on Health and Well-being. 2024; 19(1): 2327103. Publisher Full Text\n\nTaneja-Johansson S: Facilitators and barriers along pathways to higher education in Sweden: A disability lens. International Journal of Inclusive Education. 2021; 1–15. Publisher Full Text\n\nTimmerman LC, Mulvihill TM: Accommodations in the College Setting: The Perspectives of Students Living with Disabilities. Qualitative Report. 2015; 20(10): 1609–1625. Publisher Full Text Reference Source\n\nUNESCO: Education and disability: Analysis of data from 49 countries. Author; 2018. Reference Source\n\nUnited Arab Emirates Government: Education for people of determination.2023. Reference Source\n\nYssel N, Pak N, Beilke J: A door must be opened: Perceptions of students with disabilities in higher education. International Journal of Disability, Development and Education. 2016; 63: 384–394. Publisher Full Text\n\nZhang Y, Rosen S, Cheng L, et al.: Inclusive Higher Education for Students with Disabilities in China: What Do the University Teachers Think? Higher Education Studies. 2018; 8(4): 104–115. Publisher Full Text" }	[ { "id": "356054", "date": "13 Jan 2025", "name": "Dr. Glen Ford Valdez", "expertise": [ "Reviewer Expertise Nursing Education", "Nursing Leadership and Management", "Population Health", "Obesity", "Artificial Intelligence in Healthcare" ], "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\nThank you for the opportunity to review this fascinating research project. The manuscript was well written and well structured, and the methodology is systematic and accurate, with only some questions that require clarification. 1. I understand that the researchers are experts in their field, but was there special training conducted among proponents to ensure accurate and cohesive delivery and execution of the interview sessions? 2. UAE was the setting of the study. I believe various nationalities reside or study at SU; was English used as the interview medium? How did the researchers address language levels or comprehension? If translations were done into Arabic languages, how would they be processed?\n\nThese are the only two questions I have. The rest were addressed in the study limitations.\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": [ { "c_id": "13139", "date": "16 Jan 2025", "name": "Dr. Richard Mottershead", "role": "Author Response", "response": "Collectively the research team would like to thank the reviewer for their valuable time and input into our article. To answer their questions the team is an international conglomeration of researchers from diverse backgrounds and this allowed us to utilize a diverse skillset to engage with the participants who were equally diverse. Collectively, we met and reviewed the a process for data collection that could be replicated and could be standardized. Thank you for your time Dr. Richard Mottershead" } ] }, { "id": "356051", "date": "15 Jan 2025", "name": "Dr. Sarah Sanad", "expertise": [ "Reviewer Expertise Nursing profession and image", "Public health", "Simulation-based teaching and learning", "Leadership and management in healthcare", "Critical care and emergency nursing", "Disaster management", "E-learning and hybrid learning", "Pedagogies in teaching and learning", "Students’ perceptions and experiences in education", "Quality of work life", "Job satisfaction among nurses", "Risk perception", "Mental health during crises", "Psychometric evaluation of healthcare tools", "Impact studies", "Quality of life", "Health promotion", "Awareness campaigns." ], "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 titled \"Experiences of Students with Disabilities Enrolling in Higher Education: A Qualitative Study in the United Arab Emirates\" explores the lived experiences of students with disabilities in a higher education setting. Using a qualitative descriptive research approach, the study identifies barriers and facilitators influencing the academic and social integration of these students. The research draws on semi-structured interviews with 25 participants, revealing themes such as perceptions of the university, socialization, barriers, and support systems. The findings highlight the need for more inclusive practices and institutional modifications to support students with disabilities. The manuscript is well structured and provides a coherent narrative of the study. The cited literature is well presented and current. The study design is appropriate and sound for a descriptive qualitative study that includes a small number of participants. It is recommended that more information is provided on the interview guide, its development and validation. The data is available upon request from PI, which is acceptable considering the ethical guidelines for students’ data management. Anonymized excerpts or summaries of interview data can be provided as supplementary material while maintaining complete student confidentiality. The conclusions drawn are adequately supported by the results. The themes extracted are supported by the data and are tied appropriately to the implications of the findings.\nRecommendations: To elaborate on the interview methodology\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": [ { "c_id": "13157", "date": "23 Jan 2025", "name": "Dr. Richard Mottershead", "role": "Author Response", "response": "The research team would like to collectively thank the reviewer for their review and the time taken to undertake this task which adds credibility and rigor to the article." } ] } ]	1	https://f1000research.com/articles/14-14
https://f1000research.com/articles/14-11/v1	02 Jan 25	{ "type": "Research Article", "title": "Factors associated with progression of fibrosis in chronic hepatitis B virus infection in the indeterminate phase", "authors": [ "Sana Rouis", "soumaya mrabet", "Mohamed Ferjaoui", "Nedia Ben Lasfar", "Jihene Sahli", "Syrine Boujamline", "Rym Ayari", "Maha Abid", "Manel Ben Selma", "Mariem Ben Ticha", "Foued Bellazreg", "Elhem Ben Jezia", "Amel Letaief", "Wissem Hachfi", "soumaya mrabet", "Mohamed Ferjaoui", "Nedia Ben Lasfar", "Jihene Sahli", "Syrine Boujamline", "Rym Ayari", "Maha Abid", "Manel Ben Selma", "Mariem Ben Ticha", "Foued Bellazreg", "Elhem Ben Jezia", "Amel Letaief", "Wissem Hachfi" ], "abstract": "Background Anti-viral therapy is not routinely recommended for chronic hepatitis B virus (HBV) infection, in patients who have persistently elevated serum HBV DNA level (>2000 IU/mL), normal alanine aminotransferase (ALAT) and without significant liver fibrosis, defining the indeterminate phase. The objective of the study is to identify the factors associated with the progression of liver fibrosis in chronic HBV infected patients in the indeterminate phase.\n\nMethods This is cross-sectional study, conducted in Infectious Disease and Hepato-gastroenterology departments of Farhat Hached university hospital, between January 2008 and January 2022. We have included the Ag HBs (+) patients initially not treated, presenting at the time of the initial evaluation: a viral load> 2,000 IU/L for at least six months, normal ALAT (<40UI) and a fibrosis score F0 and/or F1 (in liverbiopsy or FibroScan). Univariate and logisticreg ression analysis were performed to identify the factors associated with liver fibrosis progression.\n\nResults In total, 97 patients were included, with a median age of 32.9± 9.1 years, and a female predominance (sex ratio M/F=0.64). Progression of fibrosis was observed in 16 patients (16.5%) with a mean delay of 70.9±41.1 months. In the univariate analysis, factors associated with progression of fibrosis were the presence of comorbidities (p=0.001), the high initial viral load (p=0.004), the appearance of cytolysis (p=0.001) and the increase in viral load (p=0.002), during follow-up. The AUROC of the initial viral load was 0.664 (95%CI: 0.500-0.820). An intial viral load at 8090 UI/l was associated with the progression of fibrosis with a sensibility of 70.3% and specificity of 63%.\n\nConclusion Factors associated with progression of fibrosis in the indeterminate phase of chronic HBV infection were the presence of comorbidities, and changes of ALAT during follow-up. This leads us to consider extending the therapeutic indications to this group of patients.", "keywords": [ "hepatitis b virus", "indeterminate phase", "liver fibrosis", "risk factors" ], "content": "Introduction\n\nInfection with hepatitis B virus (HBV) is a public health problem with significant morbidity and mortality associated with cirrhosis and its complications.1 According to the 2017 World Health Organization (WHO) global report, more than two billion people have been exposed to HBV.2 In North Africa, HBV infection has been described as a major etiological agent for the development of hepatocellular carcinoma (HCC).3 In Tunisia, the national prevalence of HBs Ag 1.7%.4\n\nChronic HBV infection is a dynamic process that reflects the interaction between HBV replication and host immune response. According to national and international recommendations, the indication for antiviral treatment in chronic hepatitis B (CHB) is based primarily on the combination of the following three criteria: viral load of HBV, ALAT levels, and severity of liver histological lesions. Thus, antiviral treatment is indicated only in patients with a viral load> 2,000 IU/L, whether or not associated with cytolysis and with significant hepatic fibrosis (≥F2) on liver biopsy or elastography. However, in the presence of a viral load> 2000 IU/L, there is a risk of disease progression to cirrhosis and HCC.5 Untreated patients should be monitored by regular transaminase and B viral load determinations, as well as by noninvasive fibrosis assessment and liver ultrasound. However, the modalities of monitoring its rhythm are not well established, and there are few data concerning the natural history of these patients.1\n\nPatients with a viral load >2,000 IU, but who do not meet the criteria defining therapeutic indications, have recently been classified in the so-called “indeterminate phase”.5–7 They present an increased risk of progression of hepatic fibrosis, which has been objectified on the basis of noninvasive markers of fibrosis, such as the FIB-4 Score. This progression was on the order of 11% per year in the absence of certain factors, such as advanced age, metabolic syndrome, and chronic ethylism.8\n\nThe virosuppression achieved by anti-viral therapy in chronic hepatitis B reduces the risk of progression to fibrosis and HCC. Anti-viral treatment with analogues has been shown to prevent around 60% of new cases of HCC over a 10-year period.9\n\nHowever, other studies have shown that the risk of fibrosis progression, cirrhosis, and its complications is minimal and comparable between treated and untreated patients.10–14\n\nIn this study, we aimed to identify the factors associated with fibrosis progression in patients with chronic HBV infection in the indeterminate phase.\n\n\nMethods\n\nThis cross-sectional study was conducted in the Infectious Diseases and Hepato-Gastroenterology departments of Farhat Hached University Hospital between January 2008 and January 2022.\n\nWe included all patients with chronic HBV infection in the “indeterminate phase” with Ag HBs (+) not initially treated, presenting at the time of initial evaluation a viral load> 2,000 IU/for at least six months, normal ALAT (< 40 IU), and fibrosis score F0 and/or F1 (on liver biopsy or FibroScan).\n\nWe did not include patients with cirrhosis, co-infection with hepatitis D virus (HDV), hepatitis C virus (HCV), or human immunodeficiency virus (HIV), a personal or family history of HCC, patients treated at the time of diagnosis, immunotolerant patients, and those receiving pre-emptive treatment during follow-up.\n\nData were collected from the medical records of patients using a standardized data form. In fact, data collection began at the start of the cohort in January 2008. Data processing began in May 2023, when the team started the study. The baseline data comprised sociodemographic details, including age, sex, profession, and address. We also considered comorbidities such as arterial hypertension, dyslipidemia, diabetes mellitus, chronic alcoholism (two to three standard glasses per day), body mass index (classified into 4 categories: <25 kg/m2/25-30/30-35/ >35 kg/m2), hepatic steatosis (on abdominal ultrasound and/or liver biopsy), HBe Ag (positive or negative), initial alanine aminotransferase (ALAT) rate (normal is defined as < 40 IU/L), and initial viral load (over the course of the disease, classified into 2 categories:moderate (2000–20,000 IU/L) and high (>20,000 IU/L).\n\nDuring the follow-up, the viral load was monitored (increasing/decreasing/fluctuating), ALAT rate (/6 months) (normal/high/fluctuating), METAVIR score at liver biopsy control, and liver stiffness at elastography (FibroScan) control. The following events were noted: progression of fibrosis, cirrhosis, and HCC.\n\nThe indications for antiviral treatment during follow-up included the progression of fibrosis (defined by an increase of at least 1 point in METAVIR score on liver biopsy,1 or an increase in liver stiffness of at least 1 kPa on FibroScan), progression to cirrhosis (defined by a METAVIR F4 score on FibroScan, or an elasticity greater than 15 kPa on FibroScan and/or the presence of indirect clinical, biological, and morphological signs), the occurrence of HCC (for which diagnosis is based on hepatic angioscan or MRI data, or on histological data from liver nodule biopsy), HBs or HBe seroclearance (to look for loss of HBs or HBe Ag), HBs, or HBe seroconversion (appearance of Anti HBs or Anti HBe antibodies).\n\nStatistical analysis was performed using IBM SPSS 20.0 software (https://www.ibm.com/fr-fr/products/spss-statistics ).\n\nQuantitative variables are expressed as the mean ± SD. Qualitative variables are expressed as percentages. The comparison of two means on independent series was carried out using Student’s t-test. The comparison of percentages was performed using the Pearson χ2 test and, in the event of non-validity, using the exact two-sided Fisher test. Associations between the different variables and liver fibrosis progression were estimated in a univariate analysis and a logistic regression analysis with the expression of an odds ratio (OR) with a 95% confidence interval. The relevant factors were analyzed, and their diagnostic value was evaluated using the receiver operating characteristic (ROC) curve and the area under the ROC curve (AUROC). For all statistical tests, statistical significance was set at p<0.05.\n\n\nResults\n\nA total of 827 patients were followed up for chronic HBV infection. There were 53 patients who were excluded because they were lost to follow-up. The flow diagram of the study population is shown in Figure 1. Moreover, 677 patients were not included in the study; among them, 499 had an initial viral load of < 2000 UI/ml. Finally, 97 patients who fulfilled the inclusion criteria were included (Figure 1).\n\nThe baseline characteristics of the included patients are shown in Table 1. The mean age of the participants was 32.9 ± 9.1 years. The mean initial viral load was 8 182 122.4 UI/ml. Among these patients, 38 (39.2%) were men (sex ratio M/F=0.64) and 95 (97.9%) were HBeAg-negative.\n\nUnderlying comorbidities were observed in 13.4% of patients (n=13). These comorbidities included dyslipidemia (n=7), diabetes (n=4), and high blood pressure (n=2). The mean Body Mass Index (BMI) was 26,1± 4.6 kg/m2. The initial assessment of fibrosis was performed in all patients (liver biopsy (41.2%) and elastography (58.8%)) ( Table 1). The mean initial liver stiffness was 4,9± 1.2 kPa.\n\nThe median follow-up duration was 105.2 ± 48.4 months. Progression of fibrosis was observed in 16 patients (16.5 %), with a mean delay of 70.9±41.1 months. Antiviral treatment was initiated. Complications that occurred in the enrolled patients included cirrhosis (n=3), HCC (n=1), and death (n=1). Serologically, 11 patients (10.7%) had a loss of HBsAg and five had a loss of HBe Ag (4.9%), with a mean delay of 87± 49 and 51± 24.7 months ( Table 2).\n\nIn the univariate analysis, factors associated with the progression of fibrosis were the presence of comorbidities (p=0.001), high initial viral load (p=0.004), appearance of cytolysis (p=0.001), and increased viral load (p=0.002) during follow-up ( Table 3). The AUROC of the initial viral load was 0.664 (95%CI: 0.500-0.820). An initial viral load of 8090 UI/l was associated with the progression of fibrosis with a sensitivity of 70.3% and specificity of 63% ( Figure 2).\n\nIn the multivariate analysis, the factors associated with fibrosis progression were the presence of comorbidities (Odds Ratio (95% CI)=53.345 (8.612-330.437), p<0.001) and the changes in ALAT rates (odds ratio (95% CI)=8.539 (3.168-23.018), p<0.001), but not with the initial viral load or the changes in viral load ( Table 4).\n\n\nDiscussion\n\nThe main findings of our study were as follows: First, progression of fibrosis was observed in 16 patients (16.5%), with a mean delay of 70.9±41.1 months. Second, the factors associated with the progression of fibrosis were the presence of comorbidities, high initial viral load, cytolysis, and increased viral load during follow-up. An initial viral load of 8090 UI/l was associated with the progression of fibrosis, with a sensitivity of 70.3% and specificity of 63%.\n\nOur study included chronic VHB infected patients in the indeterminate phase, which represented 11.7 % of all the patients belonging to the CHB cohort of Farhat Hached University Hospital (n=827). Although the treatment indications for chronic VHB infection are clear, more attention should be paid to those not meeting treatment indications, defining patients in the “indeterminate phase.” In our study, these patients represented 11.7% of all patients. This finding was less than that reported in a retrospective multicenter cohort study conducted in the USA and Taiwan China,5 in 3366 CHB patients were followed up for at least 1 year.\n\nThe findings showed that patients in the indeterminate phase accounted for, on average, 31.8% of the Chinese and Taiwanese cohorts and 38.7% of the American cohort. Additionally, there were 4759 CHB patients in Nanjing, China, of which 27.8% were in the indeterminate phase, according to Yao et al.7 The percentage of patients in the indeterminate phase found in our study may be explained byour younger population with a predominance of inactive carriers among all the VHB-infected patients followed in our center.\n\nJiang et al.reported that 24.3% of patients in the indeterminate phase are at risk of disease progression.15 In our study, among the 97 patients, 16 (16.5%) developed fibrosis, leading to the initiation of antiviral treatment. In a previous study that included 234 patients with CHB who did not meet the treatment criteria at presentation and during a median follow-up period of 51 months, 19.2% of patients transitioned to a more active disease phase and 18.8% started antiviral therapy.16 Huang et al. reported that among 1303 patients in the indeterminate phase, 283 (21.7%) transitioned to immune-active disease by up to 10 years of follow-up evaluation.5\n\nIn our study, in the univariate analysis, factors associated with the progression of fibrosis were the presence of comorbidities, a high initial viral load, the development of cytolysis, and the increase in viral load during follow-up. In logistic regression analysis, the factors associated with liver fibrosis progression were the presence of comorbidities and changes in ALAT rates.\n\nA review of the literature revealed only one study that examined the factors associated with fibrosis in patients in the indeterminate phase of the disease. In this retrospective cohort study involving 634 patients with CHB infection in the indeterminate phase,15 the authors found that the statistically significant variables that could affect liver fibrosis were a low/moderate HBV DNA level at the initial assessment and an increased gamma-glutamyl transpeptidase (GGT) level. In contrast, increased aspartate transaminase to platelet ratio index (APRI) and (liver inflammation and fibrosis 5) LIF-5 values17,18 were independent risk factors for liver fibrosis in the indeterminate phase.15 This study showed that regardless of ALAT values, patients with an initial low/moderate viral load had more severe liver disease, in contrast to our findings. In fact, a high viral load leads to dysfunction of HBsAg-specific cytotoxic T lymphocytes, resulting in immune tolerance. However, during prolonged reproduction, HBV interacts with the host immune system and induces cumulative immune damage and, consequently, liver damage.15 No other studies have analyzed comorbidities as an associated factor with the progression of fibrosis or other evolutionary parameters, such as the development of cytolysis in patients in the indeterminate phase. In fact, patients with comorbidities are not often included in studies evaluating hepatic fibrosis in HBsAg-positive patients because the progression of fibrosis may be linked to metabolic dysfunction-associated liver disease.\n\nOn the other hand, we found that an initial viral load of 8090 UI/l was associated with the progression of fibrosis, with an AUROC of 0.664 (95%CI: 0.500-0.820), a sensitivity of 70.3%, and a specificity of 63%. In the same study by Jiang et al.,15 low/moderate viral load was an independent factor for liver fibrosis, with an AUROC of 0.799 (95%CI: 0.760–0.838) without defining a specific cut-off. Chen et al. showed that serum HBV DNA levels in patients in the indeterminate phase were significantly higher in those with advanced inflammation and fibrosis.19,20 Elevated serum HBV DNA levels are a risk factor for significant liver inflammation in patients with CHB, which is consistent with the findings of other studies.21,22 Further studies are needed to determine the cut-off viral load associated witha high risk of fibrosis progression in patients in the indeterminate phase.\n\nIn the present study, only one patient developed HCC (1 %) and 26 (2%) in Hunag’s study.5 In fact, the correlation between viral load and the progression of end-stage liver disease (such as HCC) remains controversial. In a recent meta-analysis, the pooled annual HCC incidence was 2.54 cases per 1.000 person years (95% CI, 1.14–4) for patients in the indeterminate phase.23\n\nAccording to Huang et al., In addition to age 45 years and older (aHR, 20.8; 95% CI, 2.8–156.7; p=0.003), the indetermined period was independently linked to a higher risk of HCC development (aHR, 14.1; 95% CI, 1.3–153.3; p=0.03) than the inactive phase.5 Contrary to the findings reported by Lee et al., in which the authors assessed the untreated persistently elevated serum HBV patient group (patients in the indeterminate phase) and analyzed the cumulative HCC risk at 3, 5, 7, and 9 years (n = 67), which were 0%, 0%, 2.9%, and 2.9%, respectively.11\n\nIn contrast, we found that three patients developed cirrhosis (3.1%). In a study by Yapali et al., which included 234 patients who did not meet the criteria for antiviral treatment at presentation, none of the patients experienced cirrhosis during the follow-up.16 These results are in contrast to those of Huang et al., who found a higher 10-year cumulative incidence of cirrhosis among indeterminate patients who remained indeterminate versus inactive patients who remained inactive, 8.8% (95% CI, 6.5–11.8) vs 3.5% (95% CI, 2.5–5.0; p<.0001).5\n\nAntiviral treatment indications are generally provided to individuals at a high risk of disease progression, namely those with elevated ALAT levels, active viral replication, and advanced fibrosis or cirrhosis.1\n\nAs reported above, Huang et al. found that, without treatment, 21.7% of patients in the indeterminate phase had fibrosis progression and became immune active. These patients had a higher 10-year cumulative incidence of cirrhosis than those in the inactive phase and a 14 times higher risk of HCC development.5 Similarly, another observational study including 5414 patients, demonstrated that, compared to patients receiving oral antiviral therapy in the active phase, untreated HBeAg-negative CHB patients in the indeterminate phase had a considerably greater risk of HCC and mortality.24 As long as HBV DNA is found, several experts have suggested that therapy should start as soon as feasible to lower the chance of the disease progressing.25 Therefore, Zhou et al. recommended that antiviral therapy should be initiated in HBeAg-negative patients with normal ALAT and HBV DNA ≥ 2 000 IU/mL.25 Based on our findings, we propose to treat patients with chronic HBV infection in the indeterminate phase from a viral load value of 8090UI/l, with underlying comorbidities and developing cytolysis during flollow-up. However, the clinical benefits of antiviral therapy in this population need to be confirmed in future studies.\n\nTo our knowledge, this is the first large-scale cross-sectional study of Tunisian patients with chronic HBV infection in the indeterminate phase. However, our study has some limitations. First, many patients were lost to follow-up and were excluded from the final analysis. Second, this study did not determine the HBV genotypes. The dominant genotype of HBV in Tunisia is genotype D,26 and it has been demonstrated that genotype C infections are more prone to progress to HCC earlier, which goes some way explains the low frequency of HCC in our patients.22\n\n\nConclusions\n\nIn summary, fibrosis progression occurred in 16.5% of the patients with chronic HBV infection in the indeterminate phase. The main risk factors associated with liver fibrosis were the presence of comorbidities, high initial viral load with a cut-off of 8090 UI/l, the appearance of cytolysis, and an increase in viral load during follow-up. Further studies are required to determine whether early antiviral treatment can reduce the incidence of cirrhosis and HCC in these patients.\n\n\nEthics and consent\n\nThis study was conducted in accordance with the standards of ethics of the research. Anonymity and data confidentiality were guaranteed for all patients and written informed consent for participation in the study was obtained. At the time of data collection, the study was designed as a retrospective review of existing patient records, which did not initially anticipate publication or require additional interventions beyond routine clinical care. Consequently, an ethical approval was not sought prospectively. However, prior to manuscript submission for several months, we obtained ethical clearance retrospectively from the Ethical Committee of the Faculty of Medicine of Sousse, Tunisia on January 10, 2024, which reviewed and approved the use of the data for research purposes [Ethical Committee Number AVIS Number 220 (Ref: CEFMS 220/2024)].", "appendix": "Data availability statement\n\nThe project contains the following underlying data:\n\n• [Figshare]: sana rouis (2024). [Factors associated with progression of fibrosis in chronic hepatitis B virus infection in the indeterminate phase]. figshare. https://doi.org/10.6084/m9.figshare.28007054 Data_VHB. Dataset.27\n\n\n\n• [Figshare]: sana rouis (2024).[Factors associated with progression of fibrosis in chronic hepatitis B virus infection in the indeterminate phase] (data collection sheet). https://doi.org/10.6084/m9.figshare.27172914\n\n• [Figshare]: sana rouis (2024). Factors associated with progression of fibrosis in chronic hepatitis B virus infection in the indeterminate phase (patient consent form). https://doi.org/10.6084/m9.figshare.27172956\n\n• [Figshare]: sana rouis (2024) [Factors associated with progression of fibrosis in chronic hepatitis B virus infection in the indeterminate phase] (patient information letter). https://doi.org/10.6084/m9.figshare.27902625.v1\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nOur article answers the STROBE checklist for cross-sectional studies.\n\n\nAcknowledgments\n\nAll authors would thank Pr. N. Hannachi, Head of the Microbiology Laboratory Department in Farhat Hached University Hospital, for her exceptional dedication and invaluable contribution in obtaining the patient sample results. Her expertise and commitment have been instrumental in the successful completion of our study, providing us with reliable and comprehensive data essential for our publication.\n\n\nReferences\n\nLampertico P, Agarwal K, Berg T, et al.: EASL 2017 Clinical Practice Guidelines on the management of hepatitis B virus infection. J. Hepatol. 2017; 67(2): 370–398. PubMed Abstract \| Publisher Full Text\n\nOrganization WH: Global Hepatitis Report 2017. World Health Organization; 2017.\n\nBahri O, Ezzikouri S, Alaya-Bouafif NB, et al.: First multicenter study of risk factors for hepatocellular carcinoma development in North Africa. World J. Hepatol. 2011; 3(1): 24–30. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLes hepatites virales B: Actualisation des recommandations tunisiennes.2019.\n\nHuang DQ, Li X, Le MH, et al.: Natural history and hepatocellular carcinoma risk in untreated chronic hepatitis B patients in the indeterminate phase. Clin. Gastroenterol. Hepatol. 2022; 20(8): 1803–1812.e5. PubMed Abstract \| Publisher Full Text\n\nChen S, Dai X, Zhao Y, et al.: Clinical distribution characteristics and identification of significant liver inflammation in patients with chronic hepatitis B with indeterminate phase. Gastroenterol. Res. Pract. 2023; 2023: 1–10. Publisher Full Text\n\nYao K, Liu J, Wang J, et al.: Distribution and clinical characteristics of patients with chronic hepatitis B virus infection in the grey zone. J. Viral Hepat. 2021; 28(7): 1025–1033. PubMed Abstract \| Publisher Full Text\n\nLi J, Gordon SC, Rupp LB, et al.: Long-term progression of viral load and serum markers of fibrosis among treated and untreated patients with chronic hepatitis B. J. Gastroenterol. Hepatol. 2017; 32(6): 1250–1257. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLee H, Kim BK, Jang S, et al.: Cost-effectiveness analysis of antiviral therapy for untreated, minimally active chronic hepatitis B to prevent liver disease progression. Clin. Transl. Gastroenterol. 2021; 12(2): e00299. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLee HW, Kim SU, Park JY, et al.: Prognosis of untreated minimally active chronic hepatitis B patients in comparison with virological responders using antivirals. Clin. Transl. Gastroenterol. 2019; 10(6): e00036. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLee HW, Kim EH, Lee J, et al.: The natural history of untreated HBeAg-positive chronic HBV infection with persistently elevated HBV DNA but normal alanine aminotransferase levels. Clin. Transl. Gastroenterol. 2020; 11(3). Publisher Full Text\n\nYeo YH, Ho HJ, Yang H-I, et al.: Factors associated with HBsAg seroclearance rates in adults with chronic HBV infection: A systematic review and meta-analysis. Gastroenterology. 2019; 156(3): 635–46. e9.\n\nLiu J, Wang J, Yan X, et al.: The presence of liver inflammation in Asian patients with chronic hepatitis B with normal ALT and detectable HBV DNA in the absence of liver fibrosis. Hepatol. Commun. 2022; 6(4): 855–866. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPollicino T, Caminiti G: HBV integration studies in the clinic: Role in the natural history of infection. Viruses. 2021; 13(3): 368. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJiang SW, Lian X, Hu A-R, et al.: Histopathological lesions of the liver are severe in patients with normal alanine transaminase levels and low to moderate hepatitis B virus DNA replication. World J. Gastoentrol. 2023; 29(16): 2479–2494. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYapali S, Talaat N, Fontana RJ, et al.: Outcomes of patients with chronic hepatitis B who did not meet the criteria for antiviral treatment at presentation. Clin. Gastroenterol. Hepatol. 2015; 13(1): 193–201.e1. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSterling RK, Lissen E, Clumeck N, et al.: Development of a simple non-invasive index to predict significant fibrosis in patients with HIV/HCV coinfection. Hepatology. 2006; 43(6): 1317–1325. PubMed Abstract \| Publisher Full Text\n\nJiang S, Hu A, Yan H, et al.: Analysis of the judgment values of three non-invasive score systems (LIF-5, APRI, and FIB-4) for treatment indication in chronic HBV infected patients with ALT less than twice the upper limit of normal. ZhonghuaQuankeYixue. 2017; 15: 558–561.\n\nChen S, et al.: Clinical distribution characteristics and identification of significant liver inflammation in indeterminate phase chronic hepatitis B patients. Gastroenterol. Res. Pract. 2023; 2023: 1–10. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLi X, Xing Y, Zhou D, et al.: A non-invasive model for predicting liver inflammation in chronic hepatitis B patients with normal serum alanine aminotransferase levels. Front. Med. 2021; 8: article 688091. Publisher Full Text\n\nChen JD, Yang HI, Iloeje UH, et al.: Inactive hepatitis B carriers are still at risk of hepatocellular carcinoma and liver-related death. Gastroenterology. 2010; 138(5): 1747–54.e1. PubMed Abstract \| Publisher Full Text\n\nMason WS, Jilbert AR, Litwin S: Hepatitis B virus DNA integration and clonal expansion of hepatocytes in the chronically infected liver. Viruses. 2021; 13(2): 210. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLiu M, Zhao T, Zhang Y, et al.: The incidence of hepatocellular carcinoma and clearance of hepatitis B surface in CHB patients in the indeterminate phase: a systematic review and meta-analysis. Front. Cell. Infect. Microbiol. 2023; 13: 1226755. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChoi GH, Kim GA, Choi J, et al.: High risk of clinical events in untreated HBeAg-negative chronic hepatitis B patients with high viral load and no significant ALT elevation. Aliment. Pharmacol. Ther. 2019; 50(2): 215–226. PubMed Abstract \| Publisher Full Text\n\nZhou J, Wang F, Li L, et al.: Expanding antiviral therapy indications for HBeAg-negative chronic hepatitis B patients with normal ALT and positive HBV DNA levels. Precis. Clin. Med. 2022; 5(4): pbac030. Publisher Full Text\n\nAyed K, Gorgi Y, Ayed-Jendoubi S, et al.: Hepatitis B genotypes and precore/core-promoter mutations in Tunisian patients with chronic hepatitis B virus infection. J. Infect. 2007 Mar; 54(3): 291–297. PubMed Abstract \| Publisher Full Text\n\nRouis S: Factors associated with progression of fibrosis in chronic hepatitis B virus infection in the indeterminate phase. Dataset. figshare. 2024." }	[ { "id": "362302", "date": "27 Jan 2025", "name": "Philippa C Matthews", "expertise": [ "Reviewer Expertise viral hepatitis", "infectious diseases epidemiology", "hepatitis B treatment", "viral hepatitis in Africa", "infection biomarkers", "risk strratification" ], "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\nOverall comments: This paper sets out to tackle a relevant clinical and public health question, focusing on the extent to which people living with HBV, but not eligible for antiviral therapy, are at risk of progressive liver disease. The authors present an analysis of a subgroup of 97 people living with chronic hepatitis B under follow up in a Tunisian hospital, over a total time period of 14 years, identifying factors associated with progressive liver disease in this untreated population, which include comorbidities and viral load.\nOverall comments:\nThe authors do not recognise or acknowledge the very major change in the treatment landscape for HBV which has occurred as a result of the release of new high profile HBV data and treatment guidelines released by the World Health Organization in March 2024; these documents should be included as important citations. Thus, the whole paper is out of date and does not apply relevant current treatment thresholds and criteria. While I recognise that the majority of the work and data collection was done preceding these new guidelines, it is nevertheless crucial for the authors at least to describe and recognise the shift in the treatment landscape, and to discuss how the application of new guidelines would modify their conclusions. Likewise, the comorbidity data they have collected are highly pertinent to the new global landscape of metabolic liver disease, MASLD, which should be described and referenced. The methods are lacking a lot of essential details regarding cohort characteristics / location, laboratory methods, thresholds and definitions – this work as it stands is not reproducible. Overall, the paper contains many typographic errors, inconsistent use of language and acronyms, outdated terminology and technical ambiguities; I have captured as many of these as possible in a list of more specific comments which follows.\n\nSpecific comments:\nThe global reports and data should all be updated – rather than citing 2017 global epidemiology estimates, this should refer to the high profile report for viral hepatitis published by the WHO in 2024 and estimates published since 2017. Abbreviations should all be reviewed for consistency, and acronyms spelt out in full at first use. Terminology should be altered to align to internationally recognised nomenclature (for example, change ‘Ag HBs (+)’ to ‘HBsAg+’ and ALAT to ALT (alanine transferase), ‘UI/l’ units should be expressed as IU/L (international units per litre). ‘VHB’ and ‘HBV’ are used interchangeably. The use of the term ‘cytolysis’ is not clear – does this mean the appearance of hepatic necrosis on liver biopsy? Or elevated liver enzymes? Or something else? – this term is not in routine use and I suggest change it to reflect standard terminology in the field.\n\nAbstract:\nAbstract should state country/region of work rather than just stating the name of the hospital (most readers, including me, have to look this up to find where it is). It would also be helpful to modify the title of the paper to state the location/region represented by the study, and to provide context by stating the WHO region represented. ‘sensibility of 70.3%’ – should this read sensitivity? The abstract states that univariate and multivariate analysis was undertaken but only presents results from univariate analysis.\n\nIntroduction\n\n‘Severity of liver histological lesions’ is an inaccurate description – the inflammatory and fibrotic changes associated with HBV do not typically represent ‘lesions’ (this language suggests progression to malignancy) – suggest the authors reference appropriate histopathology scoring systems. ‘However, the modalities of monitoring its rhythm are not well established’ – this is not true; there are very well established national and international guidelines, algorithms and thresholds. ‘ethylism’ – I assume this is referring to alcohol intake? Please clarify. ‘Anti-viral treatment with analogues’ – there is a crucial missing word here – therapy is based on ‘nucleotide/nucleoside analagues’\n\nMethods\nThe reporting of study methods is extremely minimal. The authors should start with a clear description of the nature of the service and population, to include where is the hospital (city, country, WHO region)? what it the population catchment, who is referred, is this adults only or also adolescents/children, who refers patients, and on what basis are they referred to this centre? What is the overall context of healthcare provision, including state funding/out of pocket? Where were lab assays undertaken, on what laboratory platforms (need details of manufacturers, reagents, thresholds, reporting, validation). What were the criteria for liver biopsy? What treatment criteria or guidelines were in use during the period of the study? What antiviral treatment is used in this setting? The authors state that their study meets STROBE criteria, but this checklist has not been provided – suggest add as supplementary data.\n\nTerminology\nThe term ‘immunotolerant’ is recognised to be misleading and is not featured in updated guidelines. However, if it is applied to this historic cohort, then it needs a clear definition, supported by citations. Definition of chronic alcoholism as ‘two to three standard glasses per day’ is very vague. ‘Standard glasses’ of what? How much is ‘a glass’? – alcohol intake is conventionally measured in units or grams. Having defined alcohol intake, it should then be reported in the cohort characteristics or results. What reference ranges have been applied to lab parameters? How often were measurements repeated? How are ‘normal’ / ‘high’ / ‘fluctuating’ defined for ALT and viral load, respectively? ‘Qualitative variables are expressed as percentages’ seems likely to be incorrect – calculation of a percentage suggests data are quantitative. How were dyslipidaemia and hypertension diagnosed/defined?\n\nResults\nThe metadata table on Figshare needs some annotation to ensure variables are presented clearly, with units, that coding is clear and that blank cells are explained (is this missing data? Or does a blank cell mean ‘no’ or ‘zero’?) Changes in ‘ALAT rates’ does not make sense – should this be changes in absolute value of ALT? ‘An initial viral load of 8090 UI/l was associated with the progression of fibrosis with a sensitivity of 70.3% and specificity of 63%’ – it is not clear how this specific viral load figure was chosen or optimised as a threshold. Should this be presented as a baseline viral load above this threshold? FIB-4 and APRI are both mentioned as potential modalities to assess fibrosis but these data have not been presented – please explain why not. AST and platelet count both appear in the metadata table.\n\nTables and Figures\nViral loads in table 1 are defined as ‘moderate’ and ‘high’ but these need to be defined by a quantitative threshold, ideally aligning with standard thresholds used in the field. Viral loads appear to be given to one decimal place, but I am not aware of any assays that report to a resolution below the nearest 1 IU/L – please clarify. Table 1: check for typos and inconsistent units, abbreviations and terminology. In this table it is not clear whether the number reported for liver biopsy and fibroscan are the number who had these tests or the number for whom the tests were abnormal? If the former, the actual results of the investigation should be reported as well. Table 2 – all thresholds need to be defined\n\nFigure 2 is annotated in French; to be concordant with the rest of the article, this should be translated to English.\n\nLimitations\nThe authors should include the problem of a modest number of patients in their population of interest. These small numbers mean that confidence intervals around estimates are extremely wide, and that there are very small numbers of patients (only one or two) who experienced clinical outcomes of interest, even over a long time period of observation. The authors should discuss why they collected data for specific comorbidities and not others (eg. why not other cardiovascular parameters? And/or renal disease?), and discuss whether it is reasonable to take the approach they have used in analysing these together as a single group.\n\nI am not convinced that this analysis of a small number of individuals with very few clinically relevant endpoints adds anything to the field that is not already known. However, recognising that data for the whole African continent are lacking, and that we need better understanding of outcomes of HBV infection in diverse cohorts, the paper could potentially be approved and indexed with major revisions to tackle the comments above.\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? Partly", "responses": [] } ]	1	https://f1000research.com/articles/14-11
https://f1000research.com/articles/13-767/v1	08 Jul 24	{ "type": "Research Article", "title": "Effect of skill-based educational training for ambulance personnel on neonatal transport for newborn care in coastal South India – a single arm intervention study", "authors": [ "Santosh Kalyan", "Sowmini Padmanabh Kamath", "Subhodh Shetty S", "Ramesh Holla", "Leslie Lewis", "Harsha Lashkari P", "Suchitra Shenoy M", "Shantharam Baliga B", "Santosh Kalyan", "Subhodh Shetty S", "Ramesh Holla", "Leslie Lewis", "Harsha Lashkari P", "Suchitra Shenoy M", "Shantharam Baliga B" ], "abstract": "Background Education of ‘108' ambulance personnel involved in transporting neonates may improve outcomes. We assessed i) perceptions/practices of ‘108’ ambulance personnel for transporting neonates, ii) clinical parameters of transported neonates at arrival, and iii) outcomes such as survival/mortality and NICU stay (before and after skill-based educational intervention).\n\nMethods We conducted a single-arm intervention study (pre-and post) over 18 months. We assessed the perceptions and practices of 77 ambulance personnel on neonatal transport pre- versus post-intervention. Checklists assessed ambulance equipment availability/usage in both phases. We compared clinical parameters and outcomes of transported neonates between the pre-intervention (n=62) and post-intervention (n=53) phases. We analyzed data using SPSS version 25.\n\nResults Post-intervention, there was a significant reduction in the levels of hypothermia (p < 0.001), hypoglycemia (p=0.010), and prolonged capillary refill time (p=0.042), along with improvement in the use of intravenous fluids (p <0.001), a reduction in the positivity of umbilical swab growth (p=0.002) and in the duration of NICU stay (p = 0.001), significant improvement (p < 0.001) in the perceptions/practices of ambulance personnel towards neonatal transport. There was an improvement in the ambulance equipment availability/usage post-intervention.\n\nConclusions The perceptions and practices of the ‘108’ ambulance towards transporting neonates had significantly improved post-educational intervention. Further, a significant decrease in hypothermia, hypoglycemia, and duration of NICU stay was seen in neonates transported post-intervention.", "keywords": [ "Ambulances", "Checklist", "Hypothermia", "Hypoglycemia", "Infant", "Intensive care units", "Neonatal", "Newborn" ], "content": "Introduction\n\nNeonatal transport is an evolving and challenging concept in the Indian scenario.1 Challenges are primarily due to constrained and non-uniform distribution of health care facilities and inadequate transport systems. Though in-utero transport is best, we cannot predict preterm delivery/prospective perinatal problems. Thus arises the need for a dedicated transport facility to an apt well-equipped health care centre.2–4\n\nAmong the ambulances, the ‘108’ ambulance service of Emergency Management and Research Institute (EMRI)5,6 is a public-private partnership between Government and private emergency medical services. The number ‘108’ stands for the toll-free emergency telephone number across various states in India. They provide services for the public free of cost, are time-trusted, frequently used, and available across multiple districts of Karnataka. Specialised neonatal ambulances from EMRI are available in Tamilnadu and Goa and is not yet having services in Karnataka state. Our study concentrated on only neonates transported by ‘108’ ambulances.\n\nThe Indian Government has demonstrated a strong political commitment to lowering newborn mortality. In India, newborn fatalities make up 27% of all neonatal deaths worldwide. The Sustainable Development Goal target 3.27,8 and the Indian Newborn Action Plan (INAP) goal9 of a neonatal mortality rate to 12 or less per 1000 live births by 2030 is yet to be achieved by most of the states in India and is challenging. One of the significant obstacles to achieving this goal is the lack of dedicated neonatal transport. Neonatal survival depends not only on the quality of care delivered to the neonate in the NICU but also on the neonate's condition during NICU admission.10\n\nThe golden hour management in neonatal care, stabilization before and during transport, has improved outcomes.11–13 It is known that transporting sick neonates in specialized transportation with well-assembled and skilled teams can reduce mortality.1,11,13 Navjat Shishu Suraksha Karyakram (NSSK), introduced by the Government of India (GOI), also accentuates safe neonatal transport.14\n\nEarly identification of babies with altered acute physiology to determine the need for referral, care during transport, and timely therapy at the neonatal intensive care unit (NICU) is known to stop the progression of morbidity, aid recovery, and reduce mortality.15At the same time, ineffectual transport will lead to complications like hypothermia, hypoxia, and hypoglycemia, which can adversely affect neonatal outcomes. Previous studies related to transported neonates have recorded hypothermia between (27-55.3%),15–22 poor circulation in (8.6-43.42%),15,16,19,21,22 and hypoglycemia in (7.4 to 35%)16,18–22 of neonates. However, facilitated referral in dedicated neonatal ambulances have far less incidences of hypothermia and hypoglycemias.1,11,13,23\n\nPre-hospital and emergency care should be properly taught to the personnel who work in emergency medical services and accompany patients in ambulances. They should be equipped with the knowledge and abilities needed to act appropriately at the right time in an emergency and to care for a patient until they are transferred to a primary care team. Previous studies have documented varied knowledge, attitudes and practices towards prehospital care and emergency management while transporting patients in ambulances for further care.24,25\n\nThus, understanding and rectifying the ambulance personnel's perception/practices towards neonatal transport is essential for effective neonatal outcomes. With this background, we conducted the present study to assess the difference in the perceptions/practices of ‘108’ ambulance personnel towards transporting newborns pre and post skill-based educational interventional training in a coastal city of South India. In addition, we assessed the impact of the intervention on the arrival clinical parameters and clinical outcomes (survival/mortality, NICU duration of stay, blood culture positivity rates) of the neonate's pre- versus post-intervention.\n\n\nMethods\n\nWe conducted a pre and post skill based educational interventional study over 18 months at tertiary neonatal intensive care units attached to a medical college hospital in Southern India. We designed the study to assess the impact of skill-based educational training on ‘108’ ambulance personnel for early newborn care. The participant recruitment and data collection process started from 1 June, 2016.\n\nThis research follows the Consolidated Standards of Reporting Trials (CONSORT) statement guidelines.”26 The Reporting guidelines contain a completed CONSORT 2010 checklist.27 Figure 1 depicts the study flow according to CONSORT 2010 criteria.27\n\nIt was a time bound study. The study population comprised of\n\n1. The ‘108’ Ambulance personnel (AP - drivers and paramedic staff):\n\nAll the ambulance personnel (AP) of the ‘108’ ambulances who transported the neonates to our center during the study period were included for the skill-based educational intervention.\n\n2. The transported neonates to the center by ‘108’ ambulance services:\n\nAll the neonates transported only by the ‘108’ ambulances to our center during the study period were included for studying the neonatal outcomes (primary and secondary) measures.\n\nThe study was conducted in accordance with the 1964 Declaration of Helsinki, its subsequent revisions, and other relevant ethical guidelines. The institutional ethics committee of Kasturba Medical College, Mangalore, Manipal Academy of Higher education, Karnataka, Manipal, India (IEC KMC MLR 05-16/102, dated May 18, 2016) authorized the study. We took necessary permissions from the hospital authorities and the authorities concerned with the ‘108’ ambulance personnel. We obtained written informed consent from the ‘108’ ambulance staff and the parents/guardians of the newborns to participate in the study and publish the results. (as in Extended data).27\n\nWe registered the study on 26 March 2018 in the Clinical trial registry of India: CTRI registration number CTRI/2018/03/012830 (https://ctri.nic.in/Clinicaltrials/login.php).\n\nAlthough this study was a prospective trial, it allowed us to register the protocol trial even after we initiated the first enrollment of the patient/subject, and this was applicable as per CTRI rules until March 31st, 2018.\n\nHowever, CTRI announced that from April 1st, 2018, registration will be allowed only prospectively for clinical trials/studies, that is, before the enrolment of the first patient.\n\n\n\nA. Perceptions and practices of the ‘108’ ambulance personnel:\n\nPerceptions and practices of the ‘108’ ambulance personnel towards recognition, monitoring, and preventive strategies for hypothermia, hypoglycemia, abnormal respiratory rates, heart rates, and perfusion, along with practices of aseptic measures during neonatal transport (pre and post skill based educational intervention).\n\nB. To assess outcome measures of transported neonates:\n\nPrimary outcome measures: included assessment of clinical parameters at arrival to our center related to\n\n• Thermoregulation,\n\n• Blood glucose levels,\n\n• Vitals (heart rate, respiratory rate, and capillary filling time),\n\n• Presence of connected IV fluids at arrival, and\n\n• Umbilical swab colonization.\n\nSecondary outcome measures: included assessment of the clinical outcomes of transported neonates admitted to NICU related to\n\n• Survival/mortality,\n\n• Neonates discharged against medical advice,\n\n• Duration of Neonatal Intensive Care Unit (NICU stay),\n\n• Presence/absence of blood culture positive sepsis.\n\nWe compared the primary and secondary outcome measures related to the transported neonates between the pre- and post-intervention phases to assess the impact of the skill-based educational training intervention.\n\nWe collected the data using the following tools:\n\nTools 1, 2 and 3 were filled by the resident duty doctor which includes even the first author\n\nTool 1: A structured questionnaire for ambulance personnel to assess their perceptions and practices towards neonatal transport (as in Extended data).27 The questionnaire had two sections: section (A) included personal information of the AP; section (B) included the questions which assessed their perceptions and practices towards temperature regulation, glucose control, vital parameters, and asepsis of the neonates transported.\n\nTool 2: Checklists to assess the availability and utilization of the equipment in the ‘108’ ambulances (as in Extended data).27\n\nTool 3: A pre-structured proforma for residents at the arrival center to document the primary and secondary outcome measures of the neonates that get transported (as in Extended data).27\n\nWe used tools 1,2 and 3 to capture the relevant data both before and after the educational intervention for comparison. Subject experts validated the content of tools 1,2 and 3.\n\nClinical tools: included digital thermometers, one-touch glucometer strips, and umbilical culture swabs. Instead of specific neonatal probes, adult probes were used for assessing continuous vitals during transport; thus, oxygen saturation levels were not included as a parameter in our study.\n\nOn arrival of neonates at our center, the resident duty doctor-using tool 3 proforma, documented clinical parameters such as temperature, blood glucose, vitals, and the presence of connected intravenous (IV) fluids on the transported neonates. Upon reaching the hospital, a digital thermometer was used to record the axillary temperature. The glucometer was used to measure blood sugar levels. Heart rate and SpO2 measurements were taken using a multipara monitor. By manually counting, the respiratory rate was estimated.\n\nUmbilical swabs were cultured onto chocolate agar and MacConkey agar media, incubated at 37 degrees centigrade for 18-24 hours, and organisms grown were identified by biochemical reactions. These neonates were further followed for documentation of secondary outcome measures on tool 3 proforma by the first author. The clinical parameters and outcomes of transported neonates documented pre- and post-intervention were compared. The study flow of methodology is depicted in Figure 1.\n\n\n\n1. Hypothermia - axillary temperature of less than 36.5-degree centigrade\n\n2. Hypoglycemia - blood sugar of less than 50mg/dl\n\n3. Prolonged capillary filling time (CFT) - CFT of more than 3 seconds\n\n4. Tachycardia - heart rate ≥ 160 beats per minute\n\n5. Bradycardia - heart rate ≤ 100 beats per minute\n\n6. Tachypnea - respiratory rate of more than 60 cycles per minute.\n\nNeonatal sex was determined by external examination of body characteristics (external genitalia examination).\n\nWe divided the ‘108’ ambulance personnel into five batches for the skill-based educational intervention program (Table 1). Each set was given skill-based academic training as per NRP guidelines for seven hours each, on five separate days, by a team of pediatricians (first three authors) in their local language. We stressed the following concepts as per neonatal resuscitation program (NRP) guidelines28 during the skill-based education training in the form of theoretical knowledge, video-based teaching, and hands-on training to the ambulance personnel:\n\n\n\n▪ Drying, positioning, suctioning, stimulation, repositioning\n\n▪ Selection of appropriate size mask for bag and mask ventilation\n\n▪ Hands-on training in providing a correct method of positive pressure ventilation (PPV)\n\n▪ Observing adequate chest rise.\n\n▪ Ventilator corrective steps\n\nAfter five months of intervention, we assessed the retention capacity of skills among the ambulance personnel using the tool 1 questionnaire and the equipment availability/usage using tool 2 checklists.\n\nWe analyzed the collected data by IBM Corp. Released 2017. IBM SPSS Statistics for Windows, Version 25.0. Armonk, NY: IBM Corp. We used the Chi-square and Fischer exact P tests to compare the primary and secondary outcome measures. To assess the ambulance personnel's knowledge, we calculated a mean score and used paired t-tests to compare pre- and post-intervention scores.\n\n\nResults\n\nThe number of neonates transported in the pre-intervention and the post-intervention phases were 62 (29 females and 33 males) and 53 (23 females and 30 males), respectively.27 The distance travelled by the 108 ambulances for neonatal transport varied between 0.5 km to two hundred kms: duration of transport ranging between 10 minutes to 360 minutes. Mean birthweight of transported neonates was 2.01 kgs and 2.04 kgs in pre vs post intervention phases. Most neonatal cases referred were preterm neonates (nearly 30%); neonatal sepsis in 25.8 % & 22.6%, and respiratory distress in 13% & 18.9% of neonates in the pre- and post-intervention phases, respectively. Pediatric surgical cases accounted for 6.4% and 7.5% in the pre- and post-intervention phases.\n\nA comparison of the clinical parameters of transported neonates in the pre- versus post-intervention phases (Table 2) demonstrated a significant reduction in hypothermia (p< 0.001), hypoglycemia (p = 0.010), prolonged capillary refill time (p = 0.042), and a significant parallel improvement in the use of intravenous fluids (p < 0.001) post-intervention (Table 2). The vital parameters such as heart rate and respiratory rate had shown improvement in percentages of normal vitals post intervention; however, the change was not significant statistically. The spectrum of organisms that grew from the umbilical swab had shown a significant reduction (p=0.002) from 42% (pre-intervention) to 15.1% post-intervention. Staphylococcus aureus was the predominant organism colonizing the umbilical stump in both phases, with the colonization decreasing from 32.2% to 11.3% post-intervention.\n\n* Significant at < 0.05.\n\nThe clinical outcomes of transported neonates (Table 3) showed a significant reduction in the duration of NICU stay (p = 0.001) post-intervention. Mortality had decreased post-intervention to 9.43% from 14.5%; however, the difference was not statistically significant. There was a reduction in blood culture growth the intervention to 15.1% from 24.2%; however, it was not statistically significant. The yield of staphylococcus growth in blood cultures post-intervention had reduced to 25 % from 33%.\n\n* Significant at < 0.05.\n\nAbout 80 ambulance personnel got involved in the pre-intervention, while 77 were in post-intervention because of their ambulance duties. Hence, for the perceptions and practices assessment towards neonatal transport, we finally included 77 ambulance personnel.\n\nThere was a significant improvement (p < 0.001) in the mean scores of perceptions and practices of ambulance personnel (retention capacity) in the post-intervention versus pre-intervention phase related to different domains in the questionnaire (Table 4).\n\n* Significant at < 0.05.\n\nWe included 24 ambulances that transported neonates for equipment availability and usage. There was an improvement in the availability and usage of the following equipment: digital thermometer, glucometer, glucometer strips, neonatal masks, and pulse oximeter post-intervention. The use of hand rubs had improved with a reduction in the blood culture growth and umbilical swab colonization.\n\n\nDiscussion\n\nNeonates' prospects for survival depend not only on the quality and extent of neonatal care offered, but also on the state of the newborn at admission. We studied variations pre versus post educational intervention in i) newborns' arrival clinical parameters, ii) clinical outcomes, iii) ambulance crew perspectives and practices toward neonatal transport, and iv) the availability and use of ambulance equipment.\n\nMost neonates transported in pre- and post-intervention phases of the study were for preterm care and their issues. This observation is akin to other studies done in developing countries.2,15,23,29 After intervention hypothermia and hypoglycemia reduced from 48.4% to 17% and 38.7% to 16.9%, respectively. Neonatal hypoperfusion reduced to 28.3% and IV fluid administration significantly improved from 29% to 69.8% post-intervention. Different studies on neonatal transport show hypothermia in 27% to 55.3%,15–22 and hypoglycemia in 7.4 to 35 %.16,18–22 Hypoperfusion was seen in 8.6 to 43.42%15,16,19,21,22 newborns in other studies.\n\nThe significant reduction in the incidence of hypothermia and hypoglycemia post-intervention is similar to the study by Kaushal et al., which was done in two phases (before versus after training).30 Kaushal et al.,30 adopted crew training by the STABLE neonatal education program, which has six assessment care modules related to Sugar, Temperature, Airway, Blood pressure, Lab work, and Emotional support; however, in our study, training was as per NRP guidelines.28\n\nPrevious research on neonatal transport has demonstrated a substantial correlation between abnormal physiological parameters and newborn mortality.19 Conversely, favorable outcomes were observed when hemodynamic stability was preserved with improved vitals31,32 throughout transport; these were noticeably higher when a committed, knowledgeable team provided neonatal care.1,11,13,23 The efficiency of dedicated neonatal ambulance services was indicated by the significantly lower rates of hypothermia (2.3%23 & 3.2%1), hypoglycemia (3.2%1 & 4.59%23), and hypoperfusion (3.44% of cases23).\n\nContrary to other studies,17,22,23 our study had lesser mortality rates of transported neonates and a significant reduction in NICU stay duration. This could be because of improved neonatal transport post-intervention and because of different spectrum of cases during both the phases. In addition, the reduction in incidences of hypoglycemia, hypothermia, and abnormal CRT in our study was probably because of i) concurrent improvement in the availability and usage of thermometers and glucostrips, ii) clean clothing for wrapping baby, iii) neonatal masks for supplementing oxygen and iv) transporting newborns with IV access and fluids as indicated.\n\nDespite the intervention, equipment such as neonatal saturation probes, neonatal warmers, and embrace was unavailable. Our study documented the facilities available for transporting neonates by ‘108’ ambulances needs optimization. Upgrading facilities of ‘108’ ambulances and training ambulance personnel would improve outcomes of transported newborns and is similar to findings by Manikyamba et al.21\n\nPost-intervention, the transported neonates in the current study showed significant reduction in umbilical swab colonization growth and in blood culture positivity rates with parallel increase in the hand rubs availability and use. The selection of antibiotics and the cleanliness of referral facilities are two further factors that could have impacted on our study's results.\n\nThe mean scores of perceptions and practices of the ambulance personnel in our study had improved in the post-intervention phase with skill-based educational intervention measures. It had shown an impact on primary and secondary outcome measures and the availability and usage of equipment to a certain extent.\n\nThe retention knowledge (perception and practices) of ambulance personnel even after six months of intervention along with improvement in the arrival clinical parameters and clinical outcomes at discharge indicates sustainability of the intervention. In addition, capacity-building or reinforcing skills at the primary level in newborn care, recognizing danger signs, implementing early referrals, adopting safe neonatal transport measures, and conducting repeated refresher courses for ambulance personnel would aid in achieving the goal of single-digit NMR.23,33 We could appropriately use Evidence-based Kangaroo mother care interventions during transport. A Cochrane review34 suggested that cluster trials (comparing groups of hospitals) with specialist teams for neonatal transport could provide better evidence about mortality and morbidity issues.\n\nThe current study had confounding factors, which could have caused a result bias. Knowledge bias could be produced using pre-intervention questionnaire surveys and background information to determine ambulance personnel awareness and practices. Factors such as choice of antibiotics administered and variations in prior treatment and referral hospital environment could have affected the blood culture growth, the details of which we did not collect in the study because of feasibility issues. Though preterm cases were the primary referrals, the spectrum of cases that arrived in both phases was varied. Our findings may potentially have been influenced by risk factors for early onset sepsis, thus we recommend that future research should undertake molecular typing of organisms from a variety of locations, such as ambulances and referral clinics, to help determine the main point of infection dissemination. The neonatal clinical parameters at arrival and neonatal outcomes during NICU stay can differ from case to case.\n\n\nConclusions\n\nEducation of ambulance personnel on basic newborn resuscitation, asepsis, temperature regulation, and glucose maintenance reduce frequency of hypothermia, hypoglycemia, hypoperfusion, and infection in transported neonates and has an improvement in their perceptions/practices.\n\nThe study was conducted in accordance with the 1964 Declaration of Helsinki, its subsequent revisions, and other relevant ethical guidelines. The institutional ethics committee of Kasturba Medical College, Mangalore, Manipal Academy of Higher education, Karnataka, Manipal, India (IEC KMC MLR 05-16/102, dated May 18, 2016) authorized the study. We took necessary permissions from the hospital authorities and the authorities concerned with the ‘108’ ambulance personnel. We obtained written informed consent from the ‘108’ ambulance staff and the parents/guardians of the newborns to participate in the study and publish the results", "appendix": "Data availability\n\nOpen Scientific Framework: Effect of skill-based educational training for ambulance personnel on neonatal transport for newborn care in coastal south India -A single arm intervention study. https://doi.org/10.17605/OSF.IO/J5GQ7. 27\n\nThis dataset contains the following underlying data:\n\n• Pre intervention phase data excel sheet.\n\n• Post intervention phase data excel sheet.\n\nOpen Scientific Framework: Effect of skill-based educational training for ambulance personnel on neonatal transport for newborn care in coastal south India -A single arm intervention study. https://doi.org/10.17605/OSF.IO/J5GQ7. 27\n\nThis dataset contains the following underlying extended data:\n\n• Participant information sheet (ambulance personnel)\n\n• Parent/guardian information sheet\n\n• Informed consent form for ambulance personnel\n\n• Informed consent form for parents/guardians of neonates\n\n• Perceptions and practices questionnaire (ambulance personnel)\n\n• Checklist questionnaire for equipment availability and usage\n\n• Questionnaire for Primary and Secondary Outcome measures of neonates\n\nOpen Scientific Framework: CONSORT 2010 checklist for ‘Effect of skill-based educational training for ambulance personnel on neonatal transport for newborn care in coastal south India -A single arm intervention study’. https://doi.org/10.17605/OSF.IO/J5GQ7. 27\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 the “108” ambulance personnel, neonates and the parents/guardians of neonates who participated in the study\n\n\nReferences\n\nKumar PP, Kumar CD, Venkatlakshmi A: Long distance neonatal transport--the need of the hour. Indian Pediatr. 2008; 45: 920–922. PubMed Abstract\n\nSehgal A, Roy MS, Dubey NK, et al.: Factors contributing to outcome in newborns delivered out of hospital and referred to a teaching institution. Indian Pediatr. 2001; 38: 1289–1294. PubMed Abstract\n\nNational Neonatology Forum: Evidence based clinical practice guidelines.2010. [Last accessed on 2023 Nov 23]. Reference Source\n\nRashid A, Bhuta T, Berry A: A regionalised transport service, the way ahead?. Arch. Dis. Child. 1999; 80: 488–492. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKumutha J, Rao GV, Sridhar BN, et al.: The GVK EMRI maternal and neonatal transport system in India: a mega plan for a mammoth problem. Semin. Fetal Neonatal Med. 2015 Oct; 20(5): 326–334. PubMed Abstract \| Publisher Full Text\n\n108 Emergency Management and Research Institute: [Last accessed on 2023 Nov 25]. Reference Source\n\nWorld Health Organization: Every newborn: an action plan to end preventable deaths.2014. (Last accessed on 2023 Dec 29). Reference Source\n\nKumar S, Kumar N, Vivekadhish S: Millennium Development Goals (MDGs) to Sustainable Development Goals (SDGs): Addressing Unfinished Agenda and Strengthening Sustainable Development and Partnership. Indian J. Community Med. 2016; 41(1): 1–4. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMinistry of Health and Family Welfare, Government of India: National Health Mission. INAP India Newborn Action Plan.; 2014. [Last accessed on 2024 Jan 15]. Reference Source\n\nAggarwal KC, Gupta R, Sharma S, et al.: Mortality in newborns referred to tertiary hospital: An introspection. J. Family Med. Prim. Care. 2015; 4: 435–438. PubMed Abstract \| Publisher Full Text\n\nKempley ST, Sinha AK; Thames Regional Perinatal Group: Census of neonatal transfers in London and the Southeast of England. Arch Dis Child Fetal Neonatal Ed. 2004; 89: F521–F526. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRamji S: Transport in community. J Neonatol. 2005; 19: 328–331. Publisher Full Text\n\nKumar PP, Kumar CD, Shaik F, et al.: Transported neonates by a specialist team - how STABLE are they. Indian J. Pediatr. 2011; 78: 860–862. PubMed Abstract \| Publisher Full Text\n\nMinistry of Health and Family Welfare, Government of India: Navjaat Shishu Suraksha Karyakram: Resuscitation and essential Newborn Care Resource Manual.2020. [Last accessed on 2021 Feb 15]. Reference Source\n\nRao SK, Bajaj N, Rawat A: Pre-transport factors and transport quality affecting the neonatal outcome. J. Evol. Med. Dent. Sci. 2015; 04: 1991–1995. Publisher Full Text\n\nSampathkumar P, Gobinathan S: A study on status of neonatal transport to a level III neonatal intensive care unit. Int. J. Contemp Pediatr. 2018; 5: 1040–1043. Publisher Full Text\n\nRathod D, Adhisivam B, Bhat BV: Transport of sick neonates to a tertiary care hospital, South India: condition at arrival and outcome. Trop. Dr. 2015; 45: 96–99.\n\nSingh J, Dalal P, Gathwala G, et al.: Transport characteristics and predictors of mortality among neonates referred to a tertiary care centre in North India: a prospective observational study. BMJ Open. 2021 Jul 6; 11(7): e044625. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKiputa M, Salim N, Kunambi PP, et al.: Referral challenges and outcomes of neonates received at Muhimbili National Hospital, Dar es Salaam, Tanzania. PLoS One. 2022 Jun 15; 17(6): e0269479. Publisher Full Text\n\nTongo O, Abdulraheem MA, Orimadegun AE, et al.: Immediate Outcomes of Neonatal Transport in a Tertiary Hospital in South-West of Nigeria. Glob. J. of Ped & Neonatol Car. 2(1): 2020. GJPNC.MS.ID.000529. Publisher Full Text\n\nManikyamba D, Madhavi N, Reddy SM, et al.: Status of Neonatal Transport and Impact of Facilitated Referral on the Outcome of Transported Sick Newborns: At Tertiary Care Hospital. Sch. J. App. Med. Sci. 2017; 5: 3929–3934.\n\nDalal E, Vishal G, Solanki D: Study on neonatal transport at tertiary care centre. Int J Sci Res. 2013; 2: 289–292.\n\nPunitha P, Kumaravel KS, Pugalendhiraja KV, et al.: A study on the current status of neonatal transport to a special newborn care unit. Stanley Med J. 2016; 3: 55–58.\n\nSuryanto, Plummer V, Boyle M: Knowledge, attitude, and practice of ambulance nurses in prehospital care in Malang, Indonesia. Australas Emerg Care. 2018; 21: 8–12. PubMed Abstract \| Publisher Full Text\n\nNandasena G, Abeysena C: Knowledge, attitudes and skills of doctors, nurses and emergency medical technicians in pre-hospital care and emergency medicine who accompany patients in ambulances which arrive at the National Hospital of Sri Lanka. Int J Clin Anesth Res. 2018; 2: 038–043. Publisher Full Text\n\nSchulz KF, Altman DG, Moher D, et al.: CONSORT 2010 statement: updated guidelines for reporting parallel group randomized trials. Ann. Intern. Med. 2010 Jun 1; 152(11): 726–732. PubMed Abstract \| Publisher Full Text\n\nKalyan S, Kamath SP, Subodh Shetty S, et al.: Effect of skill-based educational training for ambulance personnel on neonatal transport for newborn care in coastal south India -A single arm intervention study. [Dataset]. OSF. 2024. Publisher Full Text\n\nNational neonatology forum: Teaching aids for facility based newborn care based newborn care at district level in neonatal resuscitation. [Last accessed on 2023 Dec 25]. Reference Source\n\nOkechukwu AA, Nwalozie C: Morbidity and mortality pattern of admissions into the Emergency Paediatric Unit of University of Abuja Teaching Hospital, Gwagwalada. Niger J. Med. 2011; 20: 109–113. PubMed Abstract\n\nKaushal S, Hoy DA: Reducing the Incidence of Hypothermia and Hypoglycemia during Neonatal Transport. Paper presented at: AAP experience: National Conference and Exhibition; 2015 Oct 24-27; Washington, DC.\n\nRasania M, Patel P, Bothra A, et al.: Neonatal Transport Facilities and Its Effect on Outcome: An Experience from Tertiary Care Hospital of Central Gujarat. Journal of Neonatology. 2022; 36(4): 302–310. Publisher Full Text\n\nGray MM, Riley T, Greene ND, et al.: Neonatal Transport Safety Metrics and Adverse Event Reporting: A Systematic Review. Air Med. J. 2023 Jul-Aug; 42(4): 283–295. PubMed Abstract \| Publisher Full Text\n\nPHFI, AIIMS, and SC- State of India’s Newborns (SOIN) 2014- a report: Zodpey S, Paul VK, editors. Public Health Foundation of India. New Delhi, India: All India Institute of Medical Sciences and Save the Children; [Last accessed on 2023 Dec 15]. Reference Source\n\nChang AS, Berry A, Jones LJ, et al.: Specialist teams for neonatal transport to neonatal intensive care units for prevention of morbidity and mortality. Cochrane Database Syst. Rev. 2015 Oct 28; 2015: CD007485. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "301768", "date": "22 Jul 2024", "name": "A Shashidhar Rao", "expertise": [ "Reviewer Expertise Neonatology" ], "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 need to be complimented for a social project. Few suggestions 1. The abstract should be rewritten for better clarity. Include the hypothesis in the background and only the primary objective. Results should include data and not only p values. the data perception component is not presented. (Authors to decide if it is worth presenting). 2. The introduction can be better organized from general newborn survival/care to the Indian scenario and can be shorter. 3. The flow of participants is part of the results. 4. The objectives are a part of introduction and not methods and are better written as a paragraph. 5. The authors are referred to any other published trial to better organize the methodology section 7. Results need better clarity and be divided as outcomes of newborns, outcomes of AP etc. baseline characteristics of AP are missing(training, qualification). 8. There is a significant chance of assessor bias as the assessors are the same as authors 9. Sample size is not calculated. Could be included even now. 10. It is unclear if any system changes were made or only training as the changes are expected to last longer. 11. Embrace needs to be made clear, KMC or mother accompanying is missing, 12. QI approach would have been more useful to address this gap.\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": "12989", "date": "02 Jan 2025", "name": "Sowmini Padmanabh Kamath", "role": "Author Response", "response": "1. The abstract should be written for better clarity. Include the hypothesis in the background and only the primary objective. Author response: The abstract is modified for better clarity. The hypothesis and the primary objective have been included in the background 2. Results should include data and not only p values. The data perception component is not presented (authors to decide if it is worth presenting). Author response: The data, along with the p values, have been included. We have now presented the data on the perception component. 3. The introduction can be better organized from general newborn survival/care to the Indian scenario and can be shorter. Author response: The introduction is organized and modified as suggested. 4. The flow of participants is part of the results. Author response: The flow of participants in the consort figure 1 has been included at the beginning of the results section. 5. The objectives are part of the introduction and not methods and are better written as a paragraph. Author response: The objectives of the study have been incorporated in the last paragraph of the introduction section. 6. The authors are referred to any other published trial to better organize the methodology section Author response: We have reorganized the methodology section as suggested. 7. Results need better clarity and can be divided as outcomes of newborns, outcomes of AP etc. Baseline characteristics of AP are missing (training, qualification): Author response: We have reorganized the results into The baseline characteristics of AP and transported neonates – The minimum qualification to join an ambulance crew was completing a Secondary School Leaving Certificate(S SLC); few of them had completed the 12th class, and very few had completed their bachelor’s degree. Once they joined, they received 7-day training by the GVK Emergency organization to deal with medical emergency conditions. Outcomes of AP -perceptions and practices. Outcomes of transported neonates -described under subheadings primary and secondary outcome measures. Ambulance equipment availability and usage 8. There is a significant chance of assessor bias as the assessors are the same as authors Author response: As per us, all assessors in the study had adhered to research principles and guidelines. They were unbiased in assessing the perception and practices of AP. 9. Sample size is not calculated. Could be included even now Author response: All ambulance personnel of ‘108’ ambulances during the study time were recruited thus we did not have a separate sample size calculation 10. It is unclear if any system changes were made or only training as the changes are expected to last longer Author response: There was an improvement in the availability and usage of the following equipment: digital thermometer, glucometer, glucometer strips, neonatal masks, and pulse oximeter post-intervention. The use of hand rubs had improved. System changes have been explained in the results section. 11. Embrace needs to be made clear, KMC or mother accompanying is missing. Author response: Embrace infant warmers are portable sleeping bag-like devices that keep low birthweight babies warm in hospital setups and in ambulances that are involved in neonatal transport. It is intended to be utilized in situations when skin-to-skin care is not feasible or when high-tech incubators are not available. Adopting Kangaroo mother care will allow skin-to-skin contact between the mother and baby, thereby providing warmth; however, we did not document data on the same since most of the time, the mothers did not accompany the babies that are transported immediately after delivery to our center. 12. QI approach would have been more useful to address this gap. Author response: We agree the QI approach would have been more useful to address this gap and have included this as a limitation in the study" } ] }, { "id": "301769", "date": "22 Jul 2024", "name": "Rakesh Kotha", "expertise": [ "Reviewer Expertise neonatology" ], "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\nYes, great study. Mention who took the data; it is better to be the third person who is not involved in the study. Questionnaire validation to be done and mention who prepared. Sample size justification is needed; a low sample size is a limitation. Reference for sample size is better to add. SPSS country version is better to mention. The cause of attrition is better to mention. Is it a quasi-experimental study? Definitions need citations.What is the significance of statistical errors as pre- and post-test people are different? Ambulance drivers qualifications may affect the internal study result; knowing the data better for comparing other studies. Any novelty? Better to add.\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? Partly", "responses": [ { "c_id": "13000", "date": "02 Jan 2025", "name": "Sowmini Padmanabh Kamath", "role": "Author Response", "response": "1. Mention who took the data; it is better to be the third person who is not involved in the study. Author response: The duty resident at NICU collected the data. We agree that it is better if a third person who is not involved in the study collects the data 2. Questionnaire validation to be done and mention who prepared Author response: The questionnaire was prepared by the first four authors after a thorough review of the literature pertaining to the topic. The content was validated by three senior pediatricians working in NICU set-ups. The wordings of the questions with ambiguity were changed as per their suggestions 3. Sample size justification is needed Author response: All ambulance personnel of ‘108’ ambulances during the study time were recruited; thus, we did not have a separate sample size calculation 4. A low sample size is a limitation Author response: We agree that the low sample size is a limitation. 5. Reference for sample size is better to add- Author response: All ambulance personnel of ‘108’ ambulances during the study time were recruited; thus, we did not have a separate sample size calculation 6. SPSS country version better to mention- Author response: New York is written as NY 7. Cause of attrition is better to mention Author response: Postintervention, the three APs were excluded since they had been assigned emergency ambulance duties. Thus, for the final analysis, we included 77 APs to assess perceptions and practices 8. Is it a quasi-experimental study Author response: It is a single arm intervention (pre-and post) study. 9. Definitions need citations Author response: citations included as suggested 10. Significance of statistical errors as pre and post-test people are different Author response: The subject population for AP was the same pre- and post-intervention, except that three APs could not give the post-test questionnaire because of their ambulance duties. However, the subject population for outcomes of neonatal transport ought to be different pre- versus postintervention, which could give a possibility of random error/bias. 11. Ambulance driver’s qualifications may affect the internal study result; Knowing the data better for comparing other studies Author response: We agree that the qualifications of ambulance personnel would affect the results of the internal study. We have included the educational status of AP in the baseline characteristics part of the results section. However, we did not assess the educational status with their mean perception and practice scores since it was not our study objective; we can include this as a limitation of our study. 12. Any novelty, better to add Author response: It is an intervention study that aims to improve the quality of transport of neonates. There is limited literature on pre-versus-post intervention studies on ambulance personnel, specifically ‘108’ ambulances." } ] }, { "id": "301765", "date": "22 Aug 2024", "name": "Sanoj Karayil Mohammad Ali", "expertise": [ "Reviewer Expertise Neonatology", "Respiratory physiology", "Neonatal transport", "hemodynamics" ], "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 clinically relevant interventional study with well-defined methodology and positive findings, which is similar to other studies.\n-Clear working title, which states the study design, setting and population.\n-Objective outcome data along with subjective survey from personnel is appropriate for the study\n-Unsure about the significance of umbilical swab colonization as a surrogate marker for compliance with asepsis measures. There was no significant reduction in blood culture positive rates, which can also be affected by factors other than transport.\n-Limitations of the study and risk of bias has been clearly outlined.\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": "13001", "date": "02 Jan 2025", "name": "Sowmini Padmanabh Kamath", "role": "Author Response", "response": "1. This is a clinically relevant interventional study with a well-defined methodology and positive findings, which is similar to other studies. 2. Clear working title, which states the study design, setting, and population. 3. Objective outcome data along with subjective survey from personnel is appropriate for the study Author response: Thank you 4. Unsure about the significance of umbilical swab colonization as a surrogate marker for compliance with asepsis measures. There was no significant reduction in blood culture positive rates, which can also be affected by factors other than transport. Author response: Umbilical care is a part of essential newborn care & umbilical artery is an important site for entry of bloodstream infections in newborns. This study was to look for the colonization of the umbilical area but as not a surrogate marker for neonatal sepsis. The correlation between umbilical swabs & blood culture was not a part of the study. As highlighted in the study's limitations section, we agree that factors other than transport could have affected blood culture growth. 5. Limitations of the study and risk of bias have been clearly outlined. Author response: Thank you" } ] } ]	1	https://f1000research.com/articles/13-767
https://f1000research.com/articles/13-497/v1	17 May 24	{ "type": "Research Article", "title": "Acute Kidney Injury Complicating Critical Forms of COVID-19: risk Factors and Prognostic Impact", "authors": [ "Jihene Guissouma", "Hana Ben Ali", "Hend Allouche", "Insaf Trabelsi", "Olfa Hammami", "Yosra Yahia", "Ghadhoune Hatem", "Hana Ben Ali", "Hend Allouche", "Insaf Trabelsi", "Olfa Hammami", "Yosra Yahia", "Ghadhoune Hatem" ], "abstract": "Background Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) mainly affects the respiratory tract, but different organs may be involved including the kidney. Data on acute kidney injury (AKI) in critical forms of coronavirus disease 2019 (COVID-19) are scarce. We aimed to assess the incidence, risk factors and prognostic impact of AKI complicating critical forms of COVID-19.\n\nMethods A retrospective descriptive case/control monocentric study conducted in a medical intensive care unit of a tertiary teaching hospital over a period of 18 months.\n\nResults We enrolled 144 patients, with a mean age of 58±13 years old and a male predominance (sex-ratio: 1.25). Forty-one (28%) developed AKI within a median of 4 days (Q1: 3, Q3: 8.5) after hospitalization. It was staged KDIGO class 3, in about half of the cases. Thirteen patients underwent renal replacement therapy and renal function improved in seven cases. Diabetes (OR: 6.07; 95% CI: (1,30-28,4); p: 0.022), nephrotoxic antibiotics (OR: 21; 95% CI: (3,2-146); p: 0.002), and shock (OR: 12.21; 95% CI: (2.87-51.85); p: 0.031,) were the three independent risk factors of AKI onset. Mortality was significantly higher in AKI group (OR: 3.94; 95% CI: (1.65-9.43); p<10−3) but AKI didn’t appear to be an independent risk factor of poor outcome. In fact, age (p: 0.004), shock (p: 0.045) and MV (p<10−3) were the three prognostic factors in multivariate analysis.\n\nConclusions The incidence of AKI was high in this study and associated to an increased mortality. Diabetes, use of nephrotoxic antibiotics and shock contributed significantly to its occurrence. This underlines the importance of rationalizing antibiotic prescription and providing adequate management of patients with hemodynamic instability in order to prevent consequent AKI.", "keywords": [ "acute kidney injury", "coronavirus disease 2019", "mortality", "risk factors", "prognosis." ], "content": "Introduction\n\nSince its first outbreak in December 2019 in China, the coronavirus disease 2019 (COVID-19) has spread rapidly all over the world causing a serious pandemic with high morbidity and mortality. Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) mainly affects the respiratory tract with a variable clinical presentation ranging from asymptomatic forms to severe pneumonia with acute respiratory distress syndrome (ARDS) and death.1 Although, physicians must be aware of the possible damage of other organs causing a multi-systemic impairment.2 Acute kidney injury (AKI) is a frequent complication in COVID-19 patients with a reported incidence widely ranging from 0.5%3 to above 80%.4,5\n\nThe incidence of AKI increases in parallel with the COVID-19 severity and the highest rates were recorded in the intensive care unit (ICU) patients. In addition, the occurrence of AKI seems to be a poor prognostic factor with an increased mortality.6,7\n\nAside the renal tropism of the SARS CoV-2, the pathogenesis of AKI appears to be multifactorial. Different mechanisms have been incriminated, including cells viral invasion via angiotensin converting enzyme 2 receptors mainly present on the proximal tubule cells, imbalance of the renin-angiotensin-aldosterone system, prothrombotic coagulopathy and the release of nephrotoxic mediators from cytokine storm.8 Non-specific mechanisms such as drug nephrotoxicity and renal hypoperfusion also play an important role.9\n\nCurrently, several published studies focused on hospitalized patients with COVID-19 and AKI but data on AKI complicating critical forms of COVID-19 are scarce. In this study we aimed to assess the incidence, the risk factors and the prognostic impact of AKI complicating critical forms of COVID-19.\n\n\nMethods\n\nThis was a retrospective descriptive case/control monocentric study carried out in the medical ICU of Bizerte hospital (a tertiary teaching hospital in north of Tunisia) over a period of 18 months (September 2020-February 2022). This medical ICU is managed by medical intensivists with a novel unit of six beds created for the COVID-19 outbreak.\n\nThe primary endpoint was the incidence of AKI complicating critical forms of COVID-19. The second endpoints were risk factors of AKI and prognostic factors.\n\nAll adult patients (>18 years) admitted to the ICU for critical forms of COVID-19 during the study period were included. Patients with a history of chronic renal failure and those who did not meet the critical COVID-19 criteria were excluded. Laboratory-confirmation of COVID-19 diagnosis was performed by detection of the SARS-CoV-2 RNA in nasal swabs using reverse transcription-polymerase chain reaction. Patients were divided in two groups: the case group which included the critical COVID-19 patients who developed AKI during their ICU stay: AKI patients, and the control group which included those who maintained normal renal function: No AKI patients.\n\n\n\n- Critical form of COVID-19 was considered in all included patients as defined by the WHO: “criteria for acute respiratory distress syndrome (ARDS), sepsis, septic shock, or other conditions that would normally require the provision of life-sustaining therapies such as mechanical ventilation (invasive or non-invasive) or vasopressor therapy”.1\n\n- Sepsis was defined according to the 3rd international consensus (Sepsis-3): “presence of organ dysfunction (identified as an acute change in total Sequential Organ Failure Assessment [SOFA] score ≥2 points), consequent to the infection”.10 Only sepsis prior to AKI development was assessed as a risk factor.\n\n- AKI was defined by the Kidney Disease: Improving Global Outcomes (KDIGO) as any of the following: increase in serum creatinine (SCr) by ≥0.3 mg/dl (26.5 μmol/L) within 48 h; or ≥1.5 times baseline (within the prior seven days) or urine volume < 0.5 ml/Kg/h for six hours. AKI was staged for severity according to the KDIGO criteria. Stage 1 involves increase in SCr to 1.5–1.9 times baseline or ≥ 0.3 mg/dl (26.5 μmol/L) and/or urine output <0.5 ml/kg/hr for 6–12 hours. Stage 2 is considered when SCr increase to 2.0–2.9 times baseline and/or urine output <0.5 ml/kg/hr for >12 hours. Stage 3 is defined by increase in SCr to 3.0 times baseline, or to >4.0 mg/dl (353.6 μmol/L), initiation of renal replacement therapy (RRT), and/or urine output <0.3 ml/kg/hr for ≥24 hours, or anuria for ≥12 hours.11\n\n- Rhabdomyolysis was retained if the creatine phosphokinase (CPK) rate was greater than five times the upper limit of normal.\n\n- The most prescribed nephrotoxic drugs in our ICU are vancomycin, aminoglycosides, colistin, and iodine contrast agent.\n\n\n\n- Oxygen support was: noninvasive including noninvasive ventilation (NIV) and high-flow nasal cannula (HFNC); or invasive for patients requiring mechanical ventilation (MV).\n\n- Prone position was indicated for awake and coopering patients or those under MV having PaO2/FiO2 < 150.\n\n- Corticosteroids (dexamethasone 6 mg/day; Tunisian Central Pharmacy code = 350366), vitamin C supplementation (Tunisian Central Pharmacy code = 352910), and anticoagulation were also prescribed. Our ICU anticoagulation protocol was based on low molecular weight heparin (LMWH). Standard prophylactic dose (enoxaparin 0.4 ml/day; Tunisian Central Pharmacy code = 352177) was prescribed to patients with body mass index (BMI) <30 kg/m2 and intermediate dose (enoxaparin 0.4 ml ×2/day) for those with BMI ≥30 kg/m2. Patients with presumed or confirmed venous thromboembolism had curative anticoagulation with enoxaparin 100 UI/kg×2/day. After the onset of AKI and in cases of creatinine clearance < 30 ml/min LMWH was switched to calciparin (Tunisian Central Pharmacy code = 505612) or unfractionated heparin (Tunisian Central Pharmacy code = 353526).\n\n- Antibiotics were prescribed when bacterial co-infection was presumed or confirmed. All these drugs were supplied by our hospital internal pharmacy.\n\nWe focused for each patient on demographic and clinical features, initial laboratory findings, initial thoracic computed tomography (CT) scan data, drugs received prior to AKI onset, respiratory support, renal function during hospitalization, need for RRT, ICU length of stay (LOS) and mortality.\n\nFor the descriptive study, we calculated means with standard deviations for quantitative variables with a Gaussian distribution and medians with interquartile range for variables with a non-Gaussian distribution. These variables were compared with a nonparametric Mann-Whitney test. We calculated counts and percentages for qualitative variables. Percentages were compared with Pearson’s chi-square test and with Fisher’s exact test, if this test was invalid. For analytic study; univariate logistic regression model then multivariate logistic regression analysis was done to assess AKI risk factors and those associated with ICU mortality. In all statistical tests, the significance threshold was set at 0.05. Statistical analyses were performed using free open Jamovi software.12\n\nThe Ethics Committee of our hospital (Habib Bougatfa hospital of Bizerte Tunisia) approved the study on July 20, 2023 (Approval number 1/2023) and waived informed consent because of the retrospective and descriptive design of the study. The principles outlined in the Declaration of Helsinki were followed in the protocol study.\n\nWith the aim of carrying out this work by the end of 2022, we called all surviving patients and relatives of deceased ones who met the inclusion criteria to obtain their consent to use their data anonymously and confidentially. Unfortunately, we were unable to reach all of them. We therefore obtained consent from 31 surviving patients (51 survivors in total) and consent from 44 suitable legal guardians of deceased patients (93 deceased in total). As we were unable to obtain consent from a significant number of the patients we wished to include, we referred this problem to our hospital's ethics committee. As this was a retrospective, observational study, and it was impossible to contact all the patients or their relatives, the ethics committee members waived informed consent for those we could not reach, and we obtained their agreement to carry out this study.\n\n\nResults\n\nAmong 160 patients who were admitted to the ICU in the study period, 16 didn’t meet the inclusion criteria. Thus, overall, 144 patients were included. Seventy-eight (54%) were transferred from COVID units, 42 (29%) from the emergency department and 24 (17%) from other medical or chirurgical units. Forty-one (28%) patients developed AKI. Table 1 shows the characteristics and the evolution of all patients and both groups: AKI and No AKI patients. We have summarized the epidemiological and clinical features, in addition to the laboratory and CT scan findings at ICU admission. Predisposing conditions to AKI, therapeutics and evolution were also assessed. In fact, AKI patients were older and had more comorbidities (notably diabetes and hypertension). Their heart rate, mean arterial pressure (MAP) and severity scores on admission were also higher compared to No AKI patients. Initial laboratory findings showed higher levels of white blood cells count (WBC) and C reactive protein (CRP). In addition, their baseline serum urea and creatinine rates on admission were higher. Nephrotoxic antibiotics, shock and MV requirement were the main predisposing conditions to AKI.\n\nAccording to the KDIGO criteria the AKI patients (41 cases) were staged class 1 (5 cases: 12%), class 2 (16 cases: 39%) or class 3 (20 cases: 49%). AKI occurred within a median of 4 days (3, 8.5) and extremes between 1 and 32 days. The mean creatinine level at the onset of the AKI was 285±185 μmol/L (extremes between 106 and 955 μmol/L). Thirteen patients (32%) underwent RRT. Renal function improved in seven cases (17%). As shown in Table 1: age, diabetes, hypertension, APACHE II, SAPS II, heart rate, MAP, serum baseline urea and creatinine, WBC count, CRP, shock, MV and nephrotoxic antibiotics were all predictors of AKI in univariate analysis. However, diabetes, nephrotoxic antibiotics, and shock were the three independent risk factors in the multivariate analysis (Table 2).\n\nMean ICU length of stay (LOS) was longer in AKI patients without a significant difference (p: 0.454) but mortality was significantly higher (88% versus 55%, p< 10−3) (Table 1). Only five patients of the AKI group survived (three were classified KDIGO 1 and two KDIGO 2). All AKI KDIGO 3 patients had fatal outcome.\n\nIn univariate analysis, age, hypertension, severity scores, WBC, CRP, CT scan lesion extension, P/F ratio, shock, AKI, MV, and hospital-acquired infections were all predictive of poor outcome. Besides, age, shock and MV were the three independent factors of mortality (Table 3).\n\n\nDiscussion\n\nIn this study among the 144 patients enrolled, 41 (28%) developed AKI during ICU-hospitalization within a median of 4 days (3, 8.5). It was staged KDIGO 3 in about half of the cases. Thirteen patients underwent RRT and renal function improved in only seven cases. Diabetes, nephrotoxic antibiotics and shock were the three independent risk factors of AKI. Mortality was significantly higher in AKI group, but AKI didn’t appear to be an independent risk factor of poor outcome in multivariate analysis.\n\nThe clinical presentation of COVID-19 is variable, ranging from asymptomatic forms to severe ARDS. Common symptoms of COVID-19 include fever, fatigue, dry cough, and muscle ache, however critical forms may progress to ARDS, septic shock, multi organ failure and death.13\n\nThus, disease severity was classified into three categories: Critical COVID-19 is the most serious form.1 Initially, physicians were focused in the respiratory manifestations but it was realized that multiple other organs could be also damaged. Multiple organ involvement including the liver, the gastrointestinal tract and/or the kidneys was described during the course of SARS in 200314 and more recently in the early Chinese publications on COVID-19.15\n\nCurrent literature suggests that kidney represents a potential target for SARS-COV 2: AKI is the most frequent manifestation. Data from first Chinese studies seemed to suggest a high incidence of renal complications among hospitalized patients. In patients undergoing conventional hospitalization, the incidence of AKI ranged from 0.5% to 5,3%.3,16–18\n\nThe prevalence of AKI increases in parallel with the COVID-19 severity. In the study by Hu et al, AKI occurred in 1.3% (2 of 151), 3.4% (5 of 146), and 38.5% (10 of 26) of non-severe patients, severe, and critical patients respectively.19 Similar findings were reported by Zheng et al, who found an incidence of AKI of 1.0% (3 of 297), 6.8% (13 of 190), and 39.4% (13 of 33) in non-severe, severe, and critical patients, respectively.20 In a systematic review and meta-analysis of 58 studies focused in AKI and RRT in COVID-19 patients, 13 studies reported on AKI incidence among critical patients. Overall, AKI occurred in 312/565 ICU patients with a pooled incidence rate of 39.0%.21\n\nThere is also a difference in the prevalence of AKI depending on the patients’ geographical distribution. Data from Chinese studies estimated the AKI prevalence between 8.3% and 50.6% in ICU COVID-19 patients.16,22–25 More recent studies, from the United States, have found a higher prevalence ranging from 19% to 76%.26–29 This rate seems to be more important in European ICUs reaching levels above 80 %.4,5\n\nAKI is also variable in severity. KDIGO is the most commonly used classification, and the kidney damage was staged KDIGO 1, 2 and 3 in 25-35%; 3.5-35% and 30-63% respectively in several previous series.25,27,28 AKI is usually diagnosed within 5 to 9 days of hospital admission and a median of 12 to 21 days after the onset of symptoms.23,25,30 However, Hirsch et al. reported a high frequency of AKI occurrence (37%) within 24 hours of admission.28 Depending on the study, the use of RRT in ICU is variable from 16% to 73% of patients with AKI.4,5,16,23,25,28,29\n\nThese discrepancies between studies concerning the incidence of AKI, its severity, its time of onset and the use of RRT could be explained by: variation of the definition of “severe” disease and AKI, heterogeneity of the studied populations, genetic predisposition to kidney involvement and RRT resource limitations.\n\nThe pathophysiology of AKI complicating COVID-19 may be explained by a combination of different factors: inflammatory reaction, immune response, endothelial damage as well as activation of the coagulation and the renin-angiotensin systems.8,31,32\n\nIn fact, AKI patients in our study had significantly higher biological inflammatory syndrome (WBC count and CRP). Increased markers of inflammation (C-reactive protein, ferritin, and D-dimer) were also reported in the literature, supporting the role of inflammation in the kidney impairment.2,20,24\n\nDirect viral infection with renal tropism of the virus has also been proposed but remains controversial. Acute tubular necrosis is the most frequently found histological lesion. Glomerular, vascular and endothelial damage have also been described. Besides, chronic histological lesions of nephroangiosclerosis or diabetic glomerulosclerosis are often found, indicating an underlying renal fragility factor in COVID-19 patients developing AKI.31–34\n\nIn addition to the kidney tropism of the SARS-COV 2, the pathogenesis of AKI involves several factors that are not specific to the virus but related to the patient and the critical illness or its treatment, including haemodynamic factors, drug toxicity and the impact of organ support systems.9,35\n\n-Demographic risk factors\n\nOlder age was considered as a risk factor for AKI and RRT in an Italian cohort of 99 invasively ventilated COVID-19 patients.36 Likewise, in a large Chinese study by Hirsh et al including 5449 COVID-19 patients, 1993 (36.6%) developed AKI and older age was an independent predictor of AKI (OR: 1.03; 95% CI: (1.03–1.04); p<0.001).28 Similar findings were reported by Dereli et al.2\n\nLin et al analyzed the data of 79 research articles: 8 studies investigated the risk factors of COVID-19 induced AKI and also showed that age ≥ 60 years and severe infection were independent factors predicting AKI with ORs: 3.53 (95% CI: (2.92-4.5); p<0.001), and 6.07 (95% CI (2.53-14.58); p<0.001) respectively.37\n\nWhile male gender was much more associated with AKI, as reported by Hirsh JS et al.28 and Ng JH et al.,38 sex ratio was comparable in our cohort and other previous studies.2,4,36\n\n-Comorbidities\n\nMost of the critical COVID-19 patients have pre-existing comorbidities which were also associated to AKI. The most common are hypertension and other cardiovascular disorders, diabetes and obesity. Diabetes was an independent factor in our study as well as in several series.28,38 Hypertension was also significantly much more frequent in AKI patients in our study as well as in previous studies.2,28 In addition, cardiomyopathy, chronic respiratory failure and BMI were also reported as risk factors of AKI.2,28 According to these findings, in a recent meta-analysis of forty-four studies with a total number of 114 COVID-19 patients with AKI, Sabaghian et al found that factors including older age, hypertension, cardiovascular disease, diabetes, high BMI, chronic kidney disease, immunosuppression, and smoking are the potential risk factors of AKI.7\n\nThese comorbidities are well-known factors of renal vulnerability causing histological lesions of nephroangiosclerosis or diabetic glomerulosclerosis as described above. Moreover, due to these conditions, patients are frequently treated with drugs that interfere with regulation of renal flow, such as ACE inhibitors.9 Besides, AKI patients had higher baseline serum creatinine with a significant difference in our cohort and similar findings were reported in several studies.17,28,38,40 This could be explained by the premorbid kidney disease potentially related to the frequent comorbidities especially diabetes and hypertension.\n\n-Acute disease severity and therapeutics\n\nIn addition to these non-modifiable demographic factors, the severity of the COVID-19 on admission was the major predictor of AKI. In fact, severity scores were significantly higher in the AKI patients in our study and in several previous series.2,4 In addition, ARDS requiring MV, shock and vasopressor support were reported as predictive of AKI.2,4,28,41,42\n\nSince AKI patients had more serious forms of COVID-19, they require mush more MV which was predictive of AKI in our univariate analysis but was not considered an independent factor. In fact, critical COVID-19 patients are at a high risk of AKI as a complication of MV. Specifically, high positive end-expiratory pressure used for COVID-19 associated ARDS leads to increased intrathoracic pressure and can ultimately result in increased renal venous pressure and reduced filtration.43 Besides, positive pressure ventilation can increase sympathetic tone, leading to secondary activation of the renin–angiotensin system.44 Furthermore, upregulation of proinflammatory mediators associated to biotrauma, may subsequently induce multiple system organ failure including the kidney. the kidney-lung crosstalk theory is due to the increased release of cytokines in the blood, which is promoted by lung injury. Elevated levels of cytokines, especially IL-6, increase alveolar-capillary permeability and pulmonary hemorrhage. It even may lead to distant organs dysfunction, notably damage of the kidney vascular endothelium.45\n\nMoreover, restrictive fluid strategy recommended for ARDS patients, who may initially present with relative volume depletion due to fever and gastrointestinal losses, may worsen hypovolemia and compromise renal perfusion.46 Thus, hypovolemia and hemodynamic instability cause renal hypoperfusion and, consequently, AKI. Moreover, shock is associated to lactic acidosis, hyperkalemia and rhabdomyolysis which all had a negative impact on kidney function.46 Therefore, careful attention to volume status is needed to avoid AKI.\n\nBeyond shock and diabetes, nephrotoxic antibiotics use was also an independent factor of AKI in our study. In fact, critical COVID-19 patients might be exposed to nephrotoxins as part of their clinical care, in particular, antibiotics, which can result in tubular injury or acute interstitial nephritis.\n\nIn a large Chinese study including 210 ICU COVID-19 patients, Sang et al proved that he use of nephrotoxic drug was an independent factor of AKI (OR: 2.67; 95% CI: (1.09–6.55); p: 0.0316).39\n\nSimilarly, a Portuguese study including 192 COVID-19 patients (20% of whom needed ICU management), confirmed that the exposure to nephrotoxins during the first week of admission (vancomycin, aminoglycosides, nonsteroidal anti-inflammatory drug and iodine contrast agents) was an independent factor of AKI (OR 3.60 95% CI (1.30–9.94) p=0.014).40\n\nIn a most recent study carried in Argentina including 162 ICU COVID-19 patients, exposure to nephrotoxic drugs (particularly polymyxins and aminoglycosides) was markedly higher in the AKI group (p<0.001).41\n\nMean ICU LOS was longer in AKI patients without a significant difference but mortality rate was significantly higher in this group and all patients staged KDIGO 3 deceased.\n\nIn univariate analysis AKI was a poor prognostic factor but only age, shock and MV were the three independent factors of mortality.\n\nMortality was also significantly higher in AKI patients in the most reported studies.39–42\n\nAKI stage 3 (OR: 5.33; 95% CI (1.15-24.65); p: 0.0321) was independently associated with death in the study by Sang L et al. in addition to critical disease (OR: 69.16; 95% CI (5.86-815.79); p: 0.0008), older age (OR: 1.06; 95% CI (1.02-1.11); p:0.0035) and P/F < 150 (OR: 15.21; 95% CI (4.72-49.07); p<10−3).39\n\nBeyond older age (OR: 1.07; 95% CI (1.02–1.11); p: 0.004), lower Hb level (OR: 0.78; 95% CI (0.60–0.98); p: 0.035), persistent AKI (OR: 7.34; 95% CI (2.37–22.72); p: 0.001) and severe AKI (OR: 2.65 per increase in KDIGO stage; 95% CI (1.32–5.33); p: 0.006) were also considered independent factors of mortality in the study by Gameiro et al.40\n\nThus, most of the studies agree on the negative prognostic impact of AKI on critical COVID-19 patients and this is not surprising. In fact, as AKI most often occurs in elderly patients with multiple comorbidities, severe forms of COVID-19, and requiring life-sustaining therapies (particularly MV and vasopressor therapy), they are expected to have a poor prognosis.\n\nThis study is one of the few works that have focused on the AKI in critical forms of COVID-19 managed in the ICU with a large number of patients which represent its strength. Although some limitations must be noted. The retrospective design of our study was constrained due to the paucity of data on the previous treatments of patients enrolled, notably, prior use of angiotensin converting enzyme inhibitor or angiotensin II receptor blocker. In addition, some laboratory tests were lacking in our hospital such us ferritin and D-dimers. Thus, these missing data considered as a risk factor for AKI in several studies could not be evaluated in our patients.\n\n\nConclusion\n\nThe incidence of AKI was high in this study and associated to an increased mortality. Diabetes, nephrotoxic antibiotics and shock contributed significantly to its occurrence. This emphasizes the importance of rationalizing the antibiotic prescription and avoiding nephrotoxic drugs whenever possible. In addition, a rapid and adequate management of these critical patients may reduce hemodynamic instability and consequent organs failure. furthermore, careful monitoring of renal function and early detection of AKI can help to prevent its progression to a more severe stage associated with a poor prognosis. We recommend further multicenter studies with larger samples and more detailed data in order to support our results.", "appendix": "Data availability\n\nAll data are available in Zenodo. https://doi.org/10.5281/zenodo.10865485. 47\n\nThese data include aim and methods of the study, contributors, all information about patients with respect of confidentiality and anonymity, STROBE checklist and the consent form.\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nZenodo: STROBE checklist for “Acute kidney injury complicating critical forms of COVID-19: risk factors and prognostic impact”, https://doi.org/10.5281/zenodo.10865485. 47\n\n\nReferences\n\nTherapeutics and COVID-19: living guideline, 14 July 2022. Geneva: World Health Organisation; 2022. (WHO/2019-nCoV/therapeutics/2022.4). 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSu H, Yang M, Wan C, et al.: Renal histopathological analysis of 26 postmortem findings of patients with COVID-19 in China. Kidney Int. 2020; 98: 219–227. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKudose S, Batal I, Santoriello D, et al.: Kidney Biopsy Findings in Patients with COVID-19. J. Am. Soc. Nephrol. 2020; 31: 1959–1968. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLegrand M, Bell S, Forni L, et al.: Pathophysiology of COVID-19-associated acute kidney injury. Nat. Rev. Nephrol. 2021; 17: 751–764. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFominskiy EV, Scandroglio AM, Monti G, et al.: Prevalence, characteristics, risk factors, and outcomes of invasively ventilated COVID-19 patients with acute kidney injury and renal replacement therapy. Blood Purif. 2021; 50(1): 102–109. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLin L, Wang X, Ren J, et al.: Risk factors and prognosis for COVID-19-induced acute kidney injury: a meta-analysis. BMJ Open. 2020; 10(11): e042573. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNg JH, Hirsch JS, Hazzan A, et al.: Outcomes Among Patients Hospitalized With COVID-19 and Acute Kidney Injury. Am. J. Kidney Dis. 2021; 77(2): 204–15.e1. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSang L, Chen S, Zheng X, et al.: The incidence, risk factors and prognosis of acute kidney injury in severe and critically ill patients with COVID-19 in mainland China: a retrospective study. BMC Pulm. Med. 2020; 20(1): 290. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGameiro J, Fonseca JA, Oliveira J, et al.: Acute kidney injury in hospitalized patients with COVID-19: A Portuguese cohort. Nefrologia. 2021; 41(6): 689–698. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRolón NC, Varela CF, Ferraris A, et al.: characteristics of acute kidney injury in adult patients with severe covid-19. Medicina (B Aires). 2022; 82(2): 172–180.\n\nLumlertgul N, Pirondini L, Cooney E, et al.: Acute kidney injury prevalence, progression and long-term outcomes in critically ill patients with COVID-19: a cohort study. Ann. Intensive Care. 2021; 11: 123. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKoyner JL, Murray PT: Mechanical ventilation and lung-kidney interactions. Clin. J. Am. Soc. Nephrol. 2008; 3(2): 562–570. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDudoignon E, Moreno N, Deniau B, et al.: Activation of the renin- angiotensinaldosterone system is associated with acute kidney injury in COVID-19. Anaesth. Crit. Care Pain Med. 2020; 39(4): 453–455. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJoannidis M, Forni LG, Klein SJ, et al.: Lung-kidney interactions in critically ill patients: consensus report of the acute disease quality initiative (ADQI) 21 workgroup. Intensive Care Med. 2020; 46(4): 654–672. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGłowacka M, Lipka S, Młynarska E, et al.: Acute Kidney Injury in COVID-19. Int. J. Mol. Sci. 2021; 22(15): 8081. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGuissouma J, Ben Ali H, Allouche H, et al.: Dataset: Acute kidney injury complicating critical forms of COVID-19: risk factors and prognostic impact. [Data set]. Zenodo. 2024. Publisher Full Text" }	[ { "id": "280604", "date": "05 Jun 2024", "name": "yannick Nlandu", "expertise": [ "Reviewer Expertise AKI associated with COVID-19" ], "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 thank the authors for giving me the opportunity to review this interesting article on AKI-associated COVID. This manuscript is certainly interesting but needs some changes to improve it. Introduction Currently, several published studies focused on hospitalized patients with COVID-19 and AKI but data on AKI complicating critical forms of COVID-19 are scarce. In this study we aimed to assess the incidence, the risk factors and the prognostic impact of AKI complicating critical forms of COVID-19.\nWe think the authors need to rephrase their objective, as we believe we have a considerable number of studies on AKI in critically ill COVID-19 patients. Maybe this rarity is regional in Africa? In the Maghreb? The authors need to improve the justification of their study. Are there specific features of the Tunisian population and health care that could explain the high or low incidence of AKI? Any particular risk factor?\nMethod\nThe primary endpoint was the incidence of AKI complicating severe forms of COVID-19. The secondary endpoints were risk factors for acute kidney injury and prognostic factors.\nHow should risk factors be defined as an endpoint? We believe that risk factors cannot be defined as an endpoint. The authors also need to be clear about the prognostic factors: length of stay in intensive care, vital status, etc.\nAll adult patients (>18 years) admitted to the ICU for critical forms of COVID-19 during the study period were included. Patients with a history of chronic renal failure and those who did not meet the critical COVID-19 criteria were excluded. In the inclusion or exclusion criteria, the authors should specify whether they only included patients with at least two creatinine measurements.\nWhy did the authors exclude all patients with CKD and not just those with end-stage renal disease? Could the authors provide a flow chart of patient selection?\nThe control group, which included those who retained normal renal function: No patients with AKI.\nWe believe it is correct to present patients without AKI as those who do not meet the KDIGO criterion for AKI. Normal renal function seems a bit of a misnomer.\nOnly sepsis prior to the development of AKI was assessed as a risk factor.\nThe authors need to be more precise about the time between sepsis and AKI to be taken into account: 24 hours or 48 hours before?\n- AKI was defined by the Kidney Disease: Improving Global Outcomes (KDIGO) as any of the following\nPlease state whether all patients have baseline creatinine or how the authors defined baseline creatinine in their study. Please provide a definition of AKI recovery as the authors reported an incidence of improved renal function.\nRhabdomyolysis was retained if the creatine phosphokinase (CPK) level was greater than five times the upper limit of normal.\nCould you please provide the reference for this definition? And please indicate the normal range of CPK used in the study.\nWe focused for each patient on demographic and clinical features, initial laboratory findings, initial thoracic computed tomography (CT) scan data, drugs received prior to AKI onset, respiratory support, renal function during hospitalization, need for RRT, ICU length of stay (LOS) and mortality\nThe authors should be more precise in describing the data. For example, what clinical or biological data were collected? Age? Gender? Ethnic origin? CRP? Procalcitonin? How was the percentage of lesions on CT assessed? According to which classification?\nStatistical analysis\nThe authors should specify how the data were collected and analysed (Excel? Stata? SPSS?) Multivariate logistic regression analysis was performed to assess risk factors for AKI and those associated with ICU mortality. Logistic regression is not the best way to assess factors associated with mortality. Logistic regression is not the best way to assess factors associated with mortality. It is usual to use Cox regression. Why didn't you use a Cox regression model for this retrospective study?\nResults Renal function improved in seven cases (17%) Please provide in the method section the definition of AKI recovery How can the author explain a GCS of 14±2? As in the result Ioded contrast was analyzed separately, we suggest to the authors to do the same in the method section. We have some concern. How the authors could explain the fact that with a high proportion of diabetic and hypertensive patients, an age of 58 years there is no CKD patients? Discussion\nWe believe that the discussion section is too long and should be rewritten. The authors should only comment on the relevant results of the study. The authors should discuss the incidence of AKI and the AKI rates, which seem too low. The authors should discuss the risk factors reported in their results: diabetes, nephrotoxic antibiotics and shock. The authors should comment on the lack of association between iodinated contrast agent and AKI, which we believe is of great interest in reducing negative ideas about these agents. In this case, authors should specify in the methods section which type of iodinated contrast agent was used in their hospital. The authors should comment on mortality and the fact that they did not find AKI to be a risk factor for mortality in their cohort.\nThe clinical presentation of COVID-19 is variable, ranging from asymptomatic forms to severe ARDS. Common symptoms of COVID-19 include fever, fatigue, dry cough, and muscle ache, however critical forms may progress to ARDS, septic shock, multi organ failure and death.13 We think that it is not important to add this Thus, disease severity was classified into three categories: Critical COVID-19 is the most serious form.1 Initially, physicians were focused in the respiratory manifestations but it was realized that multiple other organs could be also damaged. Multiple organ involvement including the liver, the gastrointestinal tract and/or the kidneys was described during the course of SARS in 200314 and more recently in the early Chinese publications on COVID-19.15 Current literature suggests that kidney represents a potential target for SARS-COV 2: AKI is the most frequent manifestation. Data from first Chinese studies seemed to suggest a high incidence of renal complications among hospitalized patients. In patients undergoing conventional hospitalization, the incidence of AKI ranged from 0.5% to 5,3% Idem to be deleted The authors found a low number of stage 1 AKI cases in their cohort. We believe that the authors could add a possible explanation to these results.\nPathophysiology of AKI We think that the design and the results of the study don’t allow to the authors to discuss pathophysiology of COVID-AKI they require mush more MV Please correct mush to much that he use of nephrotoxic drug was an Please correct the use We think that the authors can use the reference of this African study where many results are similar such as the incidence of AKI, the incidence of AKI recovery, the Severity score, the aminoglycoside... as risk factors for AKI.\nNlandu Y, et. al., 2023 (Ref 1) .\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": "12125", "date": "12 Aug 2024", "name": "Jihene Guissouma", "role": "Author Response", "response": "Dear Reviewer: Thanks for the time you have devoted to review this paper and for giving us the opportunity to benefit from your expertise. Here are the answers to your constructive comments : In fact, studies about AKI in critically ill patients from the great Maghreb and particularly from Tunisia are rare (to our knowledge 3 research articles and one case report). AKI was a common evolutionary complication in patients hospitalized in our ICU. That’s why we were interested to carry out this study (we have clarified this point in version 2). Methods -You are absolutely right. Risk factors cannot be defined as end points and for prognostic factors we mean vital prognosis (we have corrected this point in version 2). -For inclusion criteria we specified that all patients included had at least one creatinine measurement in ICU and one or more prior measurement in the department from which they were transferred or in the 7-365 days before admission. -All patients with CKD were excluded to have a homogeneous group of patients and avoid confounding factors. - A flow chart of patient selection was added to version 2. -For the two groups we corrected this point as: Patients were divided in two groups: the case group which included the critical COVID-19 patients who developed AKI during their ICU stay according to the Improving Global Outcomes (KDIGO) classification: AKI patients, and the control group which included those who didn’t develop AKI during their ICU stay according to the same classification: No AKI patients. -For sepsis we clarified this point in the new version: AKI was related to sepsis when a new episode of sepsis occurred during hospitalization and was followed within 48 hours by AKI. Sepsis was considered as AKI risk factor when no other cause of renal failure has been found. -For the baseline creatinine: for patients who had previous serum creatinine in the 7–365 days prior to admission, the most recent serum creatinine value was taken as the baseline creatinine and for whom no prior value was available, the lowest creatinine measured in the original department before transfer to ICU was considered baseline serum creatinine -Definition of renal recovery: Full renal recovery was achieved when serum creatinine reached a value below 1.5 times baseline and urine volume > 0.5 ml/Kg/h. (Forni LG, Darmon M, Ostermann M, et al. Renal recovery after acute kidney injury. Intensive Care Med. 2017;43(6):855–866.) -For the definition of rhabdomyolysis we clarified in the new version that we referred to this systematic review: Chavez LO, Leon M, Einav S, Varon J. Beyond muscle destruction: a systematic review of rhabdomyolysis for clinical practice. Crit Care. 2016 Jun 15;20(1):135. doi: 10.1186/s13054-016-1314-5. PMID: 27301374; PMCID: PMC4908773. -Normal CPK rates range from 10 to 200 UI/l according to our hospital laboratory. -For data collection we have specified that: we focused for each patient on demographic (age and gender) and clinical features (comorbidities, data from pleuropulmonary, cardiovascular and neurological examinations) initial laboratory findings (arterial blood gases, renal function tests, complete blood count, CRP levels, prothrombin time, CPK), initial thoracic computed tomography (CT) scan data, drugs received prior to AKI onset, respiratory support, renal function during hospitalization, need for RRT, ICU length of stay (LOS) and mortality. -The classification used in our study is that of the French “Société d'Imagerie Thoracique”, based on a visual assessment of parenchymal extension into 5 stages according to the percentage of lung affected: absent or minimal involvement (< 10%), moderate (10-25%), extensive (25-50%), severe (50-75%) or critical (>75%). https://ebulletin.radiologie.fr/actualites-covid-19/societe-dimagerie-thoracique-propose-compte-rendu-structure-scanner-thoracique Statistical analysis As indicated in the article, we used Jamovi software for data collection and analysis. Perhaps we should have used the Cox model, but multivariate analysis was chosen because it's the tool we're most proficient with. Results -The definition of AKI recovery was clarified in the methods section. -For the GCS of 14±2: you are right, it was an oversight on our part: in fact this variable had a non-Gaussian distribution and we should have used the median and interquartiles we have corrected this in the new version. -For the high proportion of diabetic and hypertensive patients, an age of 58 years without CKD: perhaps, those patients had histological lesions of nephroangiosclerosis or diabetic glomerulosclerosis but not yet at a stage of CKD. These histological lesions are considered as underlying renal fragility factors in COVID-19 patients developing AKI. Discussion We rewrote the discussion according to your recommendations and the sections that you suggest not important were deleted. Thus, we addressed that: -The low incidence of AKI in our study compared to previous series may be explained by the fact that all patients included didn’t have a history of CKD. -Omnipaque 300 was the iodinated contrast agent used in our Hospital. the lack of association between iodinated contrast agent and AKI in this study could be explained by the fact that, on the one hand, all patients didn’t have previous CKD and on the other, they received hydro-electrolytic supplements according to the daily fluid balance calculated by subtracting the total fluid output from the total intake. -Although, AKI was associated to mortality in univariate analysis it wasn’t considered as independent factor in multivariate analysis. This could be due to the fact that some factors were mutually dependent as shock, MV and AKI. -For the low number of stage 1 AKI cases in this cohort: we think that AKI was rather severe (only 12% were classified KDIGO 1) because this population had critical clinical presentation with several AKI risk factors. -Thank you for suggesting the reference of the African study we used it in different sections in the new version. We hope that we have replied to all your queries and are available if any other points remain unclear. Best regards" } ] } ]	1	https://f1000research.com/articles/13-497
https://f1000research.com/articles/13-498/v1	17 May 24	{ "type": "Research Article", "title": "Iodine density of lymphoma, metastatic SCCA, and normal cervical lymph nodes: Based on DLSCT", "authors": [ "Varalee Mingkwansook", "Urusaya Wangprasertkul", "Warit Tarathipmon", "Arvemas Watcharakorn", "Urusaya Wangprasertkul", "Warit Tarathipmon", "Arvemas Watcharakorn" ], "abstract": "Objective To compare iodine density (ID) and contrast-enhanced attenuation value (CEAV) from dual-layer spectral computed tomography (DLSCT) scans of lymphomatous, metastatic squamous cell carcinoma (SCCA), and normal cervical lymph nodes.\n\nMethods Data including ID and CEAV were retrospectively collected from patients who underwent DLSCT of the neck between January 2020 and August 2023. Results from each group (lymphomatous, metastatic SCCA, and normal) were compared and analyzed using one-way ANOVA and receiver operating characteristic curve.\n\nResults 129 cervical lymph nodes were collected from patients who met the inclusion criteria (50, 41, and 38 nodes from the lymphomatous, metastatic SCCA, and normal group, respectively). The mean ID of lymphomatous, metastatic SCCA, and normal nodes was 1.01±0.27, 1.36±0.28, and 1.45±0.29 mg/mL, respectively. Comparing lymphomatous nodes with metastatic SCCA nodes, the lymphomatous nodes had significantly lower values of ID (p<0.002) and CEAV (p<0.001). Similarly, when comparing lymphomatous nodes with normal nodes, the lymphomatous nodes had significantly lower values of ID (p<0.001) and CEAV (p<0.001). The optimal ID cut-off value for distinguishing between lymphomatous and metastatic SCCA nodes was 1.175 mg/ml (specificity of 84.2%, sensitivity 77.8%, AUC 0.788, P = 0.003). The optimal CEAV cut-off value was 77.5 HU (specificity 88.9%, sensitivity 78.9%, AUC 0.851, P<0.001).\n\nConclusions The ID and CEAV measurements from DLSCT were significantly different between lymphomatous, metastatic SCCA, and normal lymph nodes. These findings indicate that DLSCT can be used to distinguish between these conditions in the diagnosis of cervical lymph nodes.", "keywords": [ "Iodine density", "contrast-enhanced attenuation value", "cervical lymph node", "lymphoma", "squamous cell cancer", "dual-layer spectral computed tomography" ], "content": "Background\n\nCervical lymphadenopathy is a common problem, and the malignancy rate of lymph nodes in the neck exceeds 50%.1–5 The most prevalent etiologies in adults are lymphoma and metastatic squamous cell carcinoma (SCCA).2 These conditions may present with enlarged cervical nodes and upper aerodigestive tract masses which might overlap and be misdiagnosed on a conventional CT6–8 Dual-layer spectral CT (DLSCT), a subgroup of dual-energy CT (DECT), can generate additional data, including iodine density (ID).9,10 This study compares ID and contrast-enhanced attenuation value (CEAV) between lymphomatous, metastatic SCCA, and normal cervical lymph node groups.\n\n\nMethods\n\nThe study was approved by the Human Research Ethics Committee of Thammasat University (Medicine) (ref. No. MTU-EC-RA-0-222/66). Patients who underwent DLSCT of the neck with iodinated contrast media between Jan 2020 and Aug 2023 at Thammasat Hospital were retrospectively examined. The inclusion criteria for each group were 1) Metastatic SCCA patients who had primary SCCA of head and neck with pathologically proven metastatic nodes and pre-treatment DLSCT of neck, 2) Lymphoma patients with a final diagnosis of lymphoma involving a neck node with pre-treatment DLSCT of the neck, 3) Normal node patients with DLSCT of the neck for any reason except malignancy, lymphoma, and inflammatory disease. The exclusion criteria were 1) Patients with coexisting primary malignancies (head and neck or another region), and 2) Patients with suboptimal DLSCT, e.g., severe motion or metallic artifact.\n\nA dual-layer spectral CT scanner, DLSCT (Spectral CT7500 and Philips IQon Spectral CT, Philips Healthcare, Eindhoven, Netherlands), was used to scan all patients using the same standard CT neck protocol of the institute. CT parameters were as follows: 1.4:1 helical pitch, 0.4-second rotation time, and 120 kVp tube voltage with a reconstructed FOV of 512 mm2. Automated mAS and beam collimation were registered and differed depending on body habitus. The detector configurations were 128×0.625 mm for Spectral CT7500 and 64×0.625 mm for Philips IQon Spectral CT.\n\nCT neck was performed along the skull base to the aortic arch in the craniocaudal fashion. Intravenous injection of non-ionic iodinated contrast agent (Ultravist 300 mg/ml, Bayer Vital GmbH, Leverkusen, Germany, catalogue numbers: 064-18-27492-00) with 30-mL saline chaser (0.9% sodium chloride, Thai Nakorn Patana, Nonthaburi, Thailand, catalogue numbers: F-013-13035) was administered at the rate of 2.5 mL/sec using a dual syringe injection system (Stellant, MEDRAD, Indianola, Pennsylvania). The non-ionic iodinated contrast agent dosage varied according to body weight (1.5 mL/kg). Enhanced images were obtained 70 seconds after contrast agent injection.\n\nSpectral post-processing workstation (IntelliSpace Portal v11, Philips Healthcare, Netherlands) was used togenerateID and contrast-enhanced images (Figures 1, 2, and 3).\n\nAn Axial CT image of a patient with lymphoma shows CEAV measurement (70.9 HU) of the enlarged lymph node at left cervical level IIA (A). An ID measurement (1.14 mg/ml) of this lymph node at the same CT level (B).\n\nAn Axial CT image of a patient with SCCA at gum shows CEAV measurement (110.4 HU) of the enlarged lymph node at left cervical level IB (A). An ID measurement (2.23 mg/ml) of this lymph node at the same CT level (B).\n\nAn Axial CT image of a patient with normal lymph nodes shows CEAV measurement (80.5 HU) of a right cervical level IB node (A). An ID measurement (1.41 mg/ml) of this lymph node at the same CT level (B).\n\nID and CEAV were measured by placing rounded or oval-shaped regions of interest (ROI) on axial images in which lymph nodes appeared the largest (Figures 1, 2, and 3). All lymph nodes were evaluated by a radiologist with 15 years of experience in head and neck imaging. The most prominent nodes (3 nodes maximum) of each patient were selected. The average ID and CEAV values of each patient were recorded.\n\nThe selected lymphomatous and metastatic SCCA nodes had to be enlarged (>8 mm in the short axis) or have other radiologic characteristics of pathologic nodes. Gross intranodal necrosis, cystic change, fat, or calcification areas were carefully avoided.\n\nTo achieve reliable ROI, normal nodes had to be <8 mm and >5 mm in the short axis without other radiologic signs of pathologic nodes.\n\nID and CEAV of the sternocleidomastoid muscle (SCM) and internal jugular vein (IJV) were obtained from every patient using circular or ellipsoid ROIs.\n\nLymphomatous nodes were selected from patients with histopathologically proven lymphoma from cervical nodes or other head and neck areas. The selected lymphomatous nodes showed evidence of post-treatment regression on follow-up imaging. Metastatic SCCA nodes selected on CT had to correlate with the operative report and pathological results regarding side, nodal station, and size. Normal cervical lymph nodes were included in patients who were free from malignancy or inflammation in the neck.\n\nID and CEAV were interpreted in terms of mean, standard deviation (SD), median, and interquartile range (IQR). One-way ANOVA was used to compare mean ID and CEAV between the 3 groups. ROC curve was used to estimate the area under the curve (AUC) with 95%CI for differential diagnosis between lymphomatous and metastatic SCCA nodes. Descriptive statistics was used to evaluate lymph node diameter.\n\n\nResults\n\nThere was a total of 129 cervical nodes from 55 patients. The number of lymphomatous, metastatic SCCA, and normal cervical nodes was 50, 41, and 38, respectively. Baseline demographic, lymph node diameter, level of lymph nodes, and site of primary lesion of SCCA, including histopathologic subtypes of lymphoma, are described in Table 1.\n\n\n\n- Hodgkin lymphoma (n=6)\n\n- Non Hodgkin lymphoma (n=38)\n\n- Diffuse large B cell (n=10)\n\n- Follicular cell (n=12)\n\n- NLPHL (n=2)\n\n- T cell (n=8)\n\n- Marginal zone (n=3)\n\n- Round cell (n=3)\n\n\n\n- Nasopharynx (n=11)\n\n- Nasal septum (n=2)\n\n- Tongue (n=3)\n\n- Floor of mouth (n=6)\n\n- Gum (n=5)\n\n- Retromolar trigone (n=2)\n\n- Tonsil (n=2)\n\n- Pyriform sinus (n=3)\n\n- Supraglottis (n=7)\n\nThe mean (±SD) ID and CEAV of lymphomatous, metastatic SCCA, and normal cervical nodes are summarized in Table 2 and Figure 4. There were significant differences of ID (p<0.001) and CEAV (p<0.001) between the three groups.\n\n* Statistically significant at p-value<0.05 determined by One-way ANOVA.\n\nSubgroup analysis showed that the ID of lymphomatous nodes (1.01±0.27 mg/ml) was significantly lower than the ID of metastatic SCCA (1.36±0.28 mg/ml), p=0.002, and it was also lower than the ID of normal nodes (1.45±0.29 mg/ml), p<0.0001.\n\nCEAV of lymphomatous nodes (72.91±10.69 HU) was significantly lower than the CEAV of metastatic SCCA (86.84±8.95 HU), p<0.0001, and it was lower than the CEAV of normal nodes (95.90±9.67 HU), p<0.0001. Comparative results of ID and CEAV between subgroups are summarized in Table 3.\n\n* Statistically significant at p-value<0.05 determined by post-hoc test (Scheffe).\n\nThere was no significant difference in ID or CEAV of the internal jugular vein (IJV) or sternocleidomastoid muscles (SCM) between the three groups, p≥0.05 (Table 2).\n\nThe ROC curve for ID and CEAV in differentiation between lymphoma and SCCA showed good diagnostic performance with the area under the curve at 0.788 (95% CI=0.632-0.944) and 0.851 (95% CI=0.721-0.982), respectively (Figure 5).\n\nBoth curves show high diagnostic performance.\n\nThe optimal cut-off value of ID to differentiate lymphomatous nodes from metastatic SCCA, as derived from the ROC curve, was 1.175 mg/ml with a sensitivity of 85% and specificity of 87.5 %. The CEAV at 77.5 HU could also differentiate between these entities, with a specificity of 88.9% and a sensitivity of 78.9%.\n\n\nDiscussion\n\nThis is the first study to use DLSCT to compare ID between lymphomatous, metastatic SCCA, and normal cervical lymph nodes. We found lymphomatous nodes had significantly lower ID and CEAV than metastatic SCCA and normal nodes. Our data suggested that a cut-off value of ID at 1.175 mg/ml may enable discrimination between lymphomatous and SCCA nodes with specificity of 84.2% and sensitivity of 77.8%.\n\nSome literature reported ID of cervical nodes studied on dual-sources or rapid-switch technique DECT. Prior studies rated ID and CEAV highest in normal nodes, followed by metastatic and then lymphomatous nodes, similar to our findings.2,11,12 A study by Yang et al. also found lower ID of lymphomatous nodes than of metastatic SCCA nodes, but they did not show a statistical difference between these two diseases.2 They performed the study using a rapid switch kVp machine (Gemstone spectral imaging) with acquisition time at 30 seconds after contrast administration, which might explain why their results differ from ours.\n\nThe results from this study differ from results of previous studies performed in the abdominal area, which showed higher ID in lymphomatous nodes than in metastatic nodes.13 Many studies demonstrated different IDs among various types of tumors.14–16 Differences of histopathological types and vascular supply between neck and abdominal structures could explain these different ID results.\n\nOur results showed a lower ID of metastatic SCCA nodes than of normal neck nodes, which was similar to prior studies.17–19 This might relate to HU distribution, which is highest at the center of normal nodes, whereas it is evenly distributed in metastatic nodes.20 As normal nodes must be less than 8 mm in diameter, the ROI for ID measurement was inevitably placed at their center, resulting in a relatively higher ID. Greater vessel density in normal nodes than in metastatic lymph nodes with possible micronecrosis might explain this result.19,21\n\nThe ID of metastatic SCCA nodes in this study (1.36±0.28 mg/ml) was lower than the ID in a study by Tawfik et al. (2.34±0.45 mg/ml) but higher than in the studies reported by Foust et al. (0.96±0.28 mg/ml) and Luo et al. (0.98±0.42 mg/ml).17,19,22 Their studies were performed by dual-source DECT (Somatom; Siemens Healthcare, Erlangen, Germany) with different iodine concentrations of contrast media (350-400 iodine/ml). These could result in different ID values of the cervical lymph nodes with the same pathologic type.\n\nThis study also showed a relatively lower ID in normal nodes (1.45±0.29 mg/ml) compared to a prior result from Sauter AP et al. (2.09±0.09 mg/ml), which also used a DLSCT machine (iQon, Philips Healthcare, Cleveland, OH, USA).23 In contrast to this study, they used a higher iodine concentration of contrast media (Imeron 400) with a larger contrast volume (150 ml) on their CT neck protocol. These resulted in higher total iodine load and could have caused the higher ID values in their study.\n\nWe also compared the CEAV of lymph nodes between each group. This approach is more practical for routine interpretation because it can be directly measured on PACS. The results corresponded to the ID results, with lowest values in lymphomatous, metastatic SCCA, and normal nodes, respectively. CEAV was notably significantly different between the three groups (p-value<0.001).\n\nAs bolus tracking was not included in our routine CT neck protocol, we measured ID and CEAV of the sternocleidomastoid muscle (SCM) and internal jugular vein (IJV) to compare iodine content among the three groups. The results showed no significant difference in ID and CEAV of these two structures between the three patient groups (p-value>0.05).\n\nThis study has several limitations. First, it was conducted retrospectively with small patient groups. Iodine qualification data were unavailable for some patients because they were referred to our health care center after having undergone conventional CT. Second, we did not compare ID and CEAV between each lymph node level because some nodes were too small, and we did not have nodes in levels III-VI in the normal group. Comparison between normal and diseased nodes in these cervical levels might affect the results. A larger-scale study including each cervical level might address this limitation. Third, considering prior studies which showed different ID between different types of tumors,14–16 we hypothesized that ID might be affected by the histologic subtype of lymphoma. Study of ID in various lymphomatous subtypes might improve subtype differentiation. Fourth, not all lymph nodes in the lymphomatous group were confirmed because lymph node dissection was not standard treatment for this disease.\n\n\nConclusion\n\nThe results of our study indicate that ID and CEAV findings from DLSCT differed significantly between lymphomatous, metastatic SCCA, and normal cervical nodes. This knowledge improves differentiation of these conditions.\n\nThe Human Research Ethics Committee of Thammasat Univerity (Medicine) approved conducting this research with the certificate project number MTU-EC-RA-0-222/66, and approved on December 20, 2023. The inform-consent was waived requirement due to the retrospective nature of the study.\n\n\nAuthor contributions\n\nMV made contributions in literatures search, study design, data interpretation, draft writing, critical revision and final approval of the final version for submitted. WU made contributions in literatures search, data collection, data analysis and interpretation, and TW and WA made contributions in draft writing, critical revision and also faculty collaborations.", "appendix": "Data availability\n\nZenodo: Iodine density of lymphoma, metastatic SCCA, and normal cervical lymph nodes: Based on DLSCT. https://doi.org/10.5281/zenodo.11120918. 24\n\nThis project contains the following underlying data:\n\nPatients demographic data, iodine density and contrast attenuation value of neck nodes for Final data_18 Mar 2024_Final.xlsx.\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nReporting guidlines\n\nZenodo: STARD checklist. https://doi.org/10.5281/zenodo.11120918. 24\n\n\nAcknowledgement\n\nWe want to thank Assoc. Prof. Dr. Amonpon Kanlerd, Faculty of Medicine, Thammasat University, for helping us with language editing and submitting this manuscript.\n\n\nReferences\n\nChong V: Cervical lymphadenopathy: what radiologists need to know. Cancer Imaging. 2004; 4(2): 116–120. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYang L, Luo D, Li L, et al.: Differentiation of malignant cervical lymphadenopathy by dual-energy CT: a preliminary analysis. Sci. Rep. 2016; 6(1): 31020. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRoland N, Bradley PJ: Neck swellings. BMJ. 2014; 348. Publisher Full Text\n\nNaeimi M, Sharifi A, Erfanian Y, et al.: Differential diagnosis of cervical malignant lymphadenopathy among Iranian patients. Saudi Med. J. 2009; 30(3): 377–381. PubMed Abstract\n\nForghani R, Yu E, Levental M, et al.: Imaging evaluation of lymphadenopathy and patterns of lymph node spread in head and neck cancer. Expert. Rev. Anticancer. Ther. 2015; 15(2): 207–224. PubMed Abstract \| Publisher Full Text\n\nChan JM, Shin LK, Jeffrey RB: Ultrasonography of abnormal neck lymph nodes. Ultrasound Q. 2007; 23(1): 47–54. Publisher Full Text\n\nQin Y, Lu L, Lu Y, et al.: Hodgkin lymphoma involving the tonsil misdiagnosed as tonsillar carcinoma: A case report and review of the literature. Medicine. 2018; 97(7): e9761. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCho K-S, Kang D-W, Kim H-J, et al.: Differential diagnosis of primary nasopharyngeal lymphoma and nasopharyngeal carcinoma focusing on CT, MRI, and PET/CT. Otolaryngol. Head Neck Surg. 2012; 146(4): 574–578. PubMed Abstract \| Publisher Full Text\n\nForghani R, De Man B, Gupta R: Dual-energy computed tomography: physical principles, approaches to scanning, usage, and implementation: part 1. Neuroimaging Clin. 2017; 27(3): 371–384. Publisher Full Text\n\nForghani R, Mukherji S: Advanced dual-energy CT applications for the evaluation of the soft tissues of the neck. Clin. Radiol. 2018; 73(1): 70–80. PubMed Abstract \| Publisher Full Text\n\nHagtvedt T, Aaløkken T, Stensvold A, et al.: Computed tomography enhancement characteristics of lymphomatous lymph nodes of the neck. Dentomaxillofac. Radiol. 2007; 36(4): 234–236. PubMed Abstract \| Publisher Full Text\n\nHagtvedt T, Aaløkken TM, Smith H-J, et al.: Enhancement characteristics of lymphomatous lymph nodes of the neck. Acta Radiol. 2010; 51(5): 555–562. PubMed Abstract \| Publisher Full Text\n\nMartin SS, Czwikla R, Wichmann JL, et al.: Dual-energy CT-based iodine quantification to differentiate abdominal malignant lymphoma from lymph node metastasis. Eur. J. Radiol. 2018; 105: 255–260. PubMed Abstract \| Publisher Full Text\n\nMileto A, Marin D, Alfaro-Cordoba M, et al.: Iodine quantification to distinguish clear cell from papillary renal cell carcinoma at dual-energy multidetector CT: a multireader diagnostic performance study. Radiology. 2014; 273(3): 813–820. Publisher Full Text\n\nMartin SS, Weidinger S, Czwikla R, et al.: Iodine and fat quantification for differentiation of adrenal gland adenomas from metastases using third-generation dual-source dual-energy computed tomography. Investig. Radiol. 2018; 53(3): 173–178. PubMed Abstract \| Publisher Full Text\n\nChang S, Hur J, Im DJ, et al.: Volume-based quantification using dual-energy computed tomography in the differentiation of thymic epithelial tumours: an initial experience. Eur. Radiol. 2017; 27: 1992–2001. PubMed Abstract \| Publisher Full Text\n\nTawfik AM, Razek AA, Kerl JM, et al.: Comparison of dual-energy CT-derived iodine content and iodine overlay of normal, inflammatory and metastatic squamous cell carcinoma cervical lymph nodes. Eur. Radiol. 2014; 24(3): 574–580. PubMed Abstract \| Publisher Full Text\n\nLuo Y-H, Mei X-L, Liu Q-R, et al.: Diagnosing cervical lymph node metastasis in oral squamous cell carcinoma based on third-generation dual-source, dual-energy computed tomography. Eur. Radiol. 2023; 33(1): 162–171. PubMed Abstract \| Publisher Full Text\n\nFoust AM, Ali RM, Nguyen XV, et al.: Dual-energy CT-derived iodine content and spectral attenuation analysis of metastatic versus nonmetastatic lymph nodes in squamous cell carcinoma of the oropharynx. Tomography. 2018; 4(2): 66–71. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRizzo S, Radice D, Femia M, et al.: Metastatic and non-metastatic lymph nodes: quantification and different distribution of iodine uptake assessed by dual-energy CT. Eur. Radiol. 2018; 28: 760–769. PubMed Abstract \| Publisher Full Text\n\nNaresh K, Nerurkar A, Borges A: Angiogenesis is redundant for tumour growth in lymph node metastases. Histopathology. 2001; 38(5): 466–470. PubMed Abstract \| Publisher Full Text\n\nLuo Y-H, Mei X-L, Liu Q-R, et al.: Diagnosing cervical lymph node metastasis in oral squamous cell carcinoma based on third-generation dual-source, dual-energy computed tomography. Eur. Radiol. 2022; 33: 162–171. Publisher Full Text\n\nSauter AP, Ostmeier S, Nadjiri J, et al.: Iodine concentration of healthy lymph nodes of neck, axilla, and groin in dual-energy computed tomography. Acta Radiol. 2020; 61(11): 1505–1511. PubMed Abstract \| Publisher Full Text\n\nMingkwansook V: Iodine density and contrast enhanced attenuation value of neck nodes. Zenodo. 2024. Publisher Full Text" }	[ { "id": "282853", "date": "08 Aug 2024", "name": "Jarunee Intrapiromkul", "expertise": [ "Reviewer Expertise Head and neck imaging", "perfusion 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\nFor ROI measurement, there is no clear description on how to place the ROI in the lymph nodes. Is the size of ROI the same in all cases? Is it placed on the solid portion? For lymph nodes with heterogeneous enhancement, do you place it on the most enhanced portion or cover all lymph nodes. The details of ROI placement should be described in more detail so that the study can be reproduce. Also, having a second reader placing the ROI is a great way to ensure the study reproducibility by evaluating the inter-reader variability. The mean value from 2 readers can then be used for analysis. The proposed cut-off ID value may depend on the contrast concentration and dosage, which are varied on each imaging centers.\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? Partly\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "12274", "date": "22 Aug 2024", "name": "Varalee Mingkwansook", "role": "Author Response", "response": "For ROI measurement, there is no clear description on how to place the ROI in the lymph nodes. - Is the size of ROI the same in all cases? - ROI placement for each lymph node was drawn in the largest area as much as possible in the solid portion. The size must be at least 6 mm2. - Is it placed on the solid portion? - Yes. ROIs were placed on the solid enhancing portion. Areas of necrosis, cyst, calcification or fat were carefully avoided. - For lymph nodes with heterogeneous enhancement, do you place it on the most enhanced portion or cover all lymph nodes. - If the solid portion (excluding necrosis, cystic area, fat and calcification) was heterogenously enhanced, we drawn the largest ROI as much as possible to acheive the reliable ROI. The mean HU of each ROI was used as a representative for each lymph node. - The details of ROI placement should be described in more detail so that the study can be reproduce. - ROIs were placed on axial images in which lymph nodes appeared the most prominent enhancement (3 nodes maximum). If the lymph node had similar degree of enhancement on each axial view, the slice which lymph node appeared to be largest was selected in order to achieve the largest ROI. ROI must be at least 6 mm2 in size. We drew largest ROI for each lymph node as much as possible, avoiding areas of necrosis, cyst, calcification or fat. - Also, having a second reader placing the ROI is a great way to ensure the study reproducibility by evaluating the inter-reader variability. The mean value from 2 readers can then be used for analysis. - We will conduct the new article with more subjects and providing inter-reader variability in the future. - The proposed cut-off ID value may depend on the contrast concentration and dosage, which are varied on each imaging centers - The authors agree with this issue which already mentioned in discussion section." } ] }, { "id": "344815", "date": "20 Dec 2024", "name": "Siriwan Piyapittayanan", "expertise": [ "Reviewer Expertise Neuroradiology" ], "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\nComments 1. Study design; generally, metastatic cervical lymph nodes are not only SCCA. This study has not include another cell type of metastatic lymph node such as thyroid carcinoma, salivary gland carcinoma or metastatic node from organ outside head and neck such as breast and lung cancer. Explain the reason that why this study include only SCCA nodal metastasis.\n\n2. Heading; may put some word in the heading such as comparative, can iodine density differentiate between, to increasingly interest.\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": "13007", "date": "23 Dec 2024", "name": "Varalee Mingkwansook", "role": "Author Response", "response": "Thank you for your valuable comment. 1. The decision to focus solely on metastatic SCCA in cervical lymph nodes was made to ensure a more controlled and focused analysis. SCCA is one of the most common causes of metastatic cervical lymphadenopathy, particularly in head and neck cancers. By concentrating on SCCA, we aimed to investigate its specific iodine density patterns in relation to lymphoma. Including other types of metastatic lymph nodes, such as those from thyroid carcinoma, salivary gland carcinoma, or cancers from distant organs like the breast or lung, would introduce significant variability due to differences in tumor biology, vascularity, and iodine uptake. This variability could complicate the analysis and make it difficult to draw clear comparisons between the two malignancies. Focusing on SCCA allowed for a more homogenous study sample, providing clearer insights into iodine density patterns specific to SCCA and lymphoma, which is the primary goal of this research. We plan to explore a broader comparison in future studies. 2. The heading will be modified to \"Iodine Density of Lymphoma, Metastatic SCCA, and Normal Cervical Lymph Nodes: A Comparative Analysis Using DLSCT\"" } ] } ]	1	https://f1000research.com/articles/13-498
https://f1000research.com/articles/13-1017/v1	06 Sep 24	{ "type": "Study Protocol", "title": "Muscarinic receptor agonists in animal models of psychosis: protocol for a systematic review and meta-analysis", "authors": [ "Spyridon Siafis", "Nobuyuki Nomura", "Johannes Schneider-Thoma", "Irene Bighelli", "Alexandra Bannach-Brown", "Fiona J. Ramage", "Francesca Tinsdeall", "Ioannis Mantas", "Sameer Jauhar", "Sridhar Natesan", "Anthony C. Vernon", "Andrea de Bartolomeis", "Sabine M. Hölter", "Natascha I. Drude", "Ulf Tölch", "Wulf-Peter Hansen", "Virginia Chiocchia", "Oliver D. Howes", "Josef Priller", "Malcolm R. Macleod", "Georgia Salanti", "Stefan Leucht", "Nobuyuki Nomura", "Johannes Schneider-Thoma", "Irene Bighelli", "Alexandra Bannach-Brown", "Fiona J. Ramage", "Francesca Tinsdeall", "Ioannis Mantas", "Sameer Jauhar", "Sridhar Natesan", "Anthony C. Vernon", "Andrea de Bartolomeis", "Sabine M. Hölter", "Natascha I. Drude", "Ulf Tölch", "Wulf-Peter Hansen", "Virginia Chiocchia", "Oliver D. Howes", "Josef Priller", "Malcolm R. Macleod", "Georgia Salanti", "Stefan Leucht" ], "abstract": "Background Muscarinic receptor agonism is a promising mechanism of action for treating psychosis, not present in most D2R-blocking antipsychotics. Xanomeline, an M1/M4-preferring agonist, has shown efficacy in late-stage clinical trials, with more compounds being investigated. Therefore, we aim to synthesize evidence on the preclinical efficacy of muscarinic receptor agonists in animal models of psychosis to provide unique insights and evidence-based information to guide drug development.\n\nMethods We plan a systematic review and meta-analysis of in vivo animal studies comparing muscarinic receptor agonists or positive allosteric modulators with control conditions and existing D2R-blocking antipsychotics in animals subjected to any method that induces behavioural changes of relevance for psychosis. We will identify eligible studies by searching multiple electronic databases. At least two independent reviewers will conduct the study selection and data extraction using prespecified forms and assess the risk of bias with the SYRCLE’s tool. Our primary outcomes include locomotor activity and prepulse inhibition measured with standardized mean differences. We will examine other behavioural readouts of relevance for psychosis as secondary outcomes, such as social interaction and cognitive function. We will synthesize the data using multi-level meta-analysis with a predefined random-effects structure, considering the non-independence of the data. In meta-regressions we will explore potential sources of heterogeneity from a predefined list of characteristics of the animal population, model, and intervention. We will assess the confidence in the evidence considering a self-developed instrument thatconsiders the internal and external validity of the evidence.\n\nProtocol registration PROSPERO-ID: CRD42024520914", "keywords": [ "antipsychotic", "schizophrenia", "psychosis", "muscarinic receptor", "acetylcholine", "meta-analysis" ], "content": "Introduction\n\nAntipsychotic drugs that block the dopamine 2 receptor (D2R) have been the cornerstone of pharmacological treatment for schizophrenia for over 70 years.1–3 These drugs have demonstrated efficacy in reducing symptoms of psychosis, particularly positive symptoms such as hallucinations and delusions,1 yet approximately one-third of patients exhibit an inadequate response to these treatments.4 This mechanism of action also targets a downstream pathway of the aetiopathophysiology of psychosis, with limited efficacy on other core domains, such as negative symptoms like blunted affect and social withdrawal, as well as cognitive impairment.5 Moreover, the risk-to-benefit ratio of antipsychotics is often challenged by their multiple side-effects.1,2 Therefore, there has been a recognized need for more efficacious and tolerable medications for treating psychosis, but previous attempts to develop non-dopaminergic drugs have long been unsuccessful.3,6\n\nMuscarinic acetylcholine receptor agonism has recently been recognized as promising mechanism of action in the treatment of psychosis that can target components of the pathophysiology underlying schizophrenia, a property not present in most D2R-blocking antipsychotics.7–9 Xanomeline, a muscarinic M1/M4-preferring receptor agonist, has demonstrated improvements in symptoms of schizophrenia in early and late-stage randomized placebo-controlled controlled trials with medium-to-large effect sizes, with potential cholinergic adverse events mitigated by its combination with trospium, a peripheral muscarinic antagonist.10–13 Emraclidine, a drug with a different mode of action, acting as a selective M4 positive allosteric modulator (PAM), has also shown promising findings in an early clinical trial,14 and additional muscarinic agents (e.g., NBI-1117568)15 are under development.\n\nSeveral unanswered questions remain however, including the comparative efficacy of different muscarinic receptor agonists, a comparison with existing D2R-blocking antipsychotics, the potential clinical differences between orthosteric agonists and PAMs, the specific contribution of individual muscarinic receptor subtypes (M1-M5), and their potential effects on specific symptom domains. In this context, preclinical studies can provide early insights and inform further drug development. Their large number and limitations in terms of internal and external validity, however, make the translation of their findings challenging. For this reason, a critical synthesis of their evidence is required, but none exists.\n\nWe therefore plan a systematic review and meta-analysis on the effects of muscarinic receptor agonists in animal models of relevance for psychosis concerning behavioural and motor outcomes as compared to control conditions and existing D2R-blocking antipsychotics. We will also carefully assess the potential biases of these studies and evaluate the confidence in the evidence, ultimately providing evidence-based information to facilitate future drug development in schizophrenia.\n\n\nProtocol\n\nThe protocol of the review is reported according to the PRISMA statement for protocols (PRISMA-P)16 (see the checklist in the extended data), the guidelines from SYRCLE17,18 and CAMARADES.19 The protocol was registered with PROSPERO (ID: CRD42024520914) on 04.04.2024. The methodology of the protocol has been informed by our previous systematic review and meta-analysis conducted with the GALENOS project,20 which examined trace-amine associated receptor 1 (TAAR1) agonists in animal models of psychosis.21,22\n\nStudy design\n\nWe will include in vivo animal experimental studies examining any muscarinic receptor agonist versus inactive or active comparison groups in animal models of relevance for psychosis, as detailed below. There will be no restrictions on the inclusion criteria in terms of the randomization, blinding or other factors related to risk of bias, unit of allocation, duration of the study, publication status, year, country and language. We will exclude uncontrolled preclinical experiments, observational studies, and literature reviews.\n\nAnimal population and model induction\n\nWe will include animals that have undergone laboratory methods to induce psychosis-like behaviours and features. There are numerous models fitting this description, each with varying degrees of validity and unique strengths and weaknesses, but none are considered the gold standard.23–30 Therefore, we will include the “classical” pharmacological models of psychosis and their behavioural readouts (see “Outcomes”), which have been widely utilized in drug discovery and possess some predictive validity, especially for positive symptoms, including the administration of psychostimulants (e.g., amphetamine, cocaine) or N-methyl-D-aspartate (NMDA) receptor antagonists (e.g., phencyclidine, ketamine, MK-801).29,31,32 Other methods of induction will also be eligible and can include other pharmacological models, neurodevelopmental methods, lesion methods, genetic models, and combinations of different methods.24–30 Such relatively broad inclusion criteria were utilized in our previous systematic review,21,22 and any decision on the eligibility of the psychosis models will be made in consultation with experts in preclinical research (MRM, AB, FJ, FT, IM, SN, AdB, SH, NID, UT).\n\nThere will be no restriction on species, strain, age, and sex. Regarding genetic composition, we will include both wildtype animals and those that have undergone genetic interventions, if these interventions belong to eligible methods of induction (see above). Moreover, in the eligible studies, we will extract data from “naïve” animal cohorts (i.e., animals that have not undergone models of psychosis), and animal cohorts that have undergone both models of psychosis and muscarinic receptor antagonism via genetic or pharmacological manipulation. The eligible animal cohorts can be found in Table 1.\n\nWe will exclude animals that have undergone induction methods for other specific conditions (e.g., transgenic models of Alzheimer’s disease,33 the valproic acid-induced model of relevance for autism34 and methods aiming to specifically model depressive-like behaviours, such as animal models of physical, social, or chronic mild stress).35 We will also exclude in vitro, ex vivo, in silico studies and studies in humans. We will however consider extracting data from ex vivo measurements (e.g., Fos expression, autoradiography) following eligible in vivo experiments, potentially analysed in secondary publications (see “Outcomes”).\n\nWe will include any pharmacological agent acting as an agonist or positive allosteric modulator at any of the five subtypes of muscarinic acetylcholine M1-M5 receptors. There will be no restrictions on their receptor selectivity, pharmacological potency and efficacy (e.g., full or partial agonists), dose, timing of administration relative to the induction method, pharmacokinetic properties, or route of administration, provided the method is suitable for achieving effects in the central nervous system. These pharmacological agents can be administered individually or in combination with D2R-blocking antipsychotics or other medications, and combinations can be considered as distinct interventions. We will also consider data on the effects of muscarinic receptor agonists in the context of muscarinic receptor antagonism (see “Animal Population and model induction”) to evaluate whether the effects depend on the activation of muscarinic receptors or other mechanisms (Table 1).\n\nWe will exclude clozapine, an existing antipsychotic acting on multiple neurotransmitter receptor systems and also as muscarinic receptor partial agonist,2,8 which will be considered as a control intervention (see “Comparison groups”). Clozapine’s metabolite N-desmethylclozapine however, which has low affinity for dopamine receptors and acts as a muscarinic M1/M4 receptor agonist,36 will be considered among the experimental interventions. Additionally, we will exclude pharmacological agents with different mechanisms of action (e.g., nicotinic acetylcholine receptor agonists, acetylcholinesterase inhibitors) and non-pharmacological interventions, including genetic interventions for muscarinic receptor overexpression.\n\nComparison groups\n\nWe will include the following comparison groups: i) inactive control conditions, consisting of animals undergoing models of a method to induce psychosis-like behaviour and features and receiving vehicle (e.g., injection of saline) or no treatment, and ii) active control conditions, consisting of the aforementioned animal cohorts treated with D2R-blocking antipsychotics. D2R-blocking antipsychotics will be defined as those medications listed in the Anatomical Therapeutic Chemical (ATC) classification with a code of N05A, except for lithium (N05AN01, a mood stabilizer primarily used for the treatment of bipolar disorder), pimavanserin (N05AX17, a 5-HT2A antagonist indicated for Parkinson’s disease psychosis but not schizophrenia), and any muscarinic receptor agonists approved by any regulatory body for schizophrenia (which will be considered as experimental interventions in this review), with no further restrictions.\n\nWe will also consider data in the eligible studies from sham procedures, consisting of “naïve” animal cohorts not subjected to methods of inducing psychosis-like behaviours or features and receiving either vehicle or no treatment, to calculate normalized mean differences in a sensitivity analysis (see “Effect sizes”).\n\nThere is no gold standard measure of preclinical antipsychotic efficacy due to the limited homology with the clinical symptoms of psychosis and the lack of an established biomarker.6,25,26,29–32,37–39 Therefore, we aim to provide a comprehensive assessment of the effects of muscarinic receptor agonists by using data from a broad range of behavioral measures in various animal models of relevance for psychosis (see “Animal population and model induction”). This approach will facilitate the identification of strong clinical canditates.24,25,30–32\n\nWe will examine as co-primary outcomes the effects on i) locomotor activity and ii) prepulse inhibition of the acoustic startle reflex, as these have been widely used with some predictive validity to identify antipsychotic effects in animal models of relevance for psychosis (see “Animal population and model induction”).21,26,31 Furthermore, we will examine additional behavioral measures as secondary outcomes, including potential proxies for positive symptoms (e.g., stereotypies, hallucinatory-like percepts),40 negative symptoms (e.g., lack of social interaction, operant-based motivational tasks),31,41 depressive- (e.g., forced swim, tail suspension, sucrose preference tasks)31,41 and anxiety-like behaviors (e.g., elevated plus maze), and cognitive function (e.g., tests recommended by the CNTRICS initiative).30,37,39 The eligibility and any potential grouping of the behavioral measures will be evaluated in collaboration with experts in preclinical research prior to commencing data analysis, and any decisions will be documented.\n\nWe will exclude non-behavioural outcomes, such as histopathological and neurobiological measures due to the lack of an established biomarker,6 and adverse events due to the inconsistent and scarce reporting in animal studies, as identified in our previous review.21 Nonetheless, we will aim to extract their data, if available, and potentially analyse them in secondary publications.\n\nWe will conduct searches in PubMed, MEDLINE via Ovid, Web of Science, EMBASE, PsychINFO, from database inception using keywords for psychosis, muscarinic agents, and animal filters,42 similar to a previous review.21,22 For muscarinic agents, we will use broad terms like “muscarinic agonists” and the names/synonyms of specific relevant agents identified from IUPHAR/BPS43 and previous reviews.7,44 There will be no restrictions such as on language or publication date. The search strategies will be developed in collaboration with information specialist (see “Acknowledgment”). A draft search strategy in MEDLINE via Ovid can be found in the extended data, and the final search strategies will be reported according to the PRISMA statement for reporting literature searches (PRISMA-S).45\n\nWe plan additional searches to improve the coverage of the study identification and identify potentially unpublished studies:\n\n1) We will aim to search in the Systematic Online Living Evidence Summaries for preclinical psychosis research (psychosis-SOLES),46 which is a dedicated and continuously updated database utilizing machine learning and text mining algorithms.\n\n2) We will search preclinical animal study registries (i.e., animalstudyregistry.org, https://preclinicaltrials.eu/), although pre-registration of animal studies has not been widely adopted.\n\n3) We will aim to search preprint registries (i.e., medRxiv, bioRxiv), Google patents, specific journals of neuropsychopharmacology.\n\n4) We will inspect reference lists of included studies, previous reviews,7,8,44,47 and conference proceedings published within the last 20 years.\n\n5) We will contact the first/corresponding author of included studies and pharmaceutical industries of muscarinic agents for additional studies and/or missing data in their studies. We will send emails with two follow-up reminders in case of no response.\n\nAt least two independent reviewers will screen in the Systematic Review Facility (SyRF)23 the de-duplicated records identified in the searches in two phases: i) title and abstract, and ii) full-text screening. Any discrepancy between the two reviewers will be resolved through discussion with a more senior reviewer. If not resolved by discussion, the full-text of the study will be acquired (if at the title/abstract level) or additional information from the original study authors will be obtained (if at the full-text level).\n\nAt the title/abstract screening, records will be excluded according to the following hierarchy: i) review articles, ii) not referring to in vivo animal study, iii) not referring to muscarinic receptors or agents acting on them.\n\nAt the full-text screening, records will be excluded from the review and/or meta-analysis according to the following hierarchy against the eligibility criteria: i) ineligible study design, ii) ineligible animals/population, iii) ineligible intervention, iv) ineligible comparison groups, v) ineligible outcome measure, vi) inadequate reporting of outcome data. If possible, we will consider the studies excluded in terms of their reported outcomes in the assessment of reporting bias (see “Data synthesis”). The study selection process and the reasons for exclusions at the full-text level will be reported in a flow diagram.48\n\nAt least two independent reviewers will conduct the data extraction in SyRF23 using pre-specified data extraction forms that will be adapted from a previous systematic review.21 Discrepancies between the two reviewers will be reconciled by a more senior reviewer, or if not possible, by contacting study authors for additional information.\n\nWe will extract data regarding study identification (e.g., author names, title, publication year), study design (e.g., risk of bias, reporting completeness), animal population and model induction (e.g., age, sex, species strain, body weight, characteristics of the induction method), experimental and control interventions (e.g., type, dose and timing of the administration of drugs), and outcome measures (e.g., exact name of the behavioural task, methods of measurement, quantitative data). We will seek information from various sources in the following order of priority: i) text and tables, ii) figures using WebPlotDigitizer version 4 (a free and opensource tool distributed under GNU Affero General Public License Version 3),49 iii) contacting authors for missing information, and iv) employing imputation methods.\n\nWe will extract quantitative data for the outcomes anticipated to be reported as continuous measures. Endpoint and change scores will both be eligible and jointly synthesized.50 When both are reported, we will prefer endpoint scores because they do not require baseline assessments and are expected to be the most frequently reported in the eligible animal experiments. We will extract the unit of measurement, mean, standard deviation and the number of participants that these correspond to. We will apply a minus transformation whenever appropriate to harmonize the direction of effects across the extracted data (e.g., a higher score indicating a better outcome). Missing standard deviations will be derived by the following methods according to their order of priority: i) calculation from standard error, ii) estimation from test statistics (e.g., p-values, t-tests, median and ranges), iii) contacting the authors of the original studies for additional information, iv) imputed from the standard deviations of other studies (although this method has not been validated in animal studies that often have small sample sizes).51,52 If the exact number of animals is not reported,53 we will estimate it with available information (e.g., using the lower boundary of a range, if reported) or consider imputation methods. If dichotomous measures are reported, we will extract the number of animals with the event and the total number of animals analysed. We will exclude studies with imputed data in a sensitivity analysis (see “Sensitivity analysis”).\n\nFor crossover trials, we will prefer data from the first phase to avoid carryover effects, but we will also use data from the entire phase by applying appropriate corrections considering the within-subject correlation.54 These studies will be excluded in a sensitivity analysis (see “Sensitivity analysis”).\n\nWe will use data from any reported time point, but preference will be given for the longest time point following multiple administrations of the intervention over an extended period. If outcomes are measured multiple times after a single administration, we will consider calculating the area under the curve.22,55\n\nIf multiple variations of the same outcome are reported, we will extract and jointly analyse them (see “Data synthesis”).\n\nTwo independent reviewers will assess risk of bias using the SYRCLE’s tool considering domains for selection, performance, detection, attrition, reporting and other biases.56 We will assign an overall high risk of bias to a study if at least one domain in the SYRCLE’s tool is assessed as having a high risk of bias. Since high-quality reporting is essential for assessing the risk of bias, two independent reviewers will also evaluate the completeness of reporting using a modified version of the ARRIVE Essential 10 checklist.22,57 This is necessary because the reporting completeness of animal studies is often poor, frequently leading to unclear assessments of risk of bias. Any discrepancies between the two reviewers will be resolved through discussion with a more senior reviewer or by contacting the study authors for additional information.\n\nWe expect that the findings from various animal models of relevance for psychosis will have different degrees of applicability to clinical trial settings. However, there is no established method for assessing their indirectness in the context of a systematic review. Moreover, there is limited synthesized evidence on the validity of animal models of relevance for psychosis, and we expect substantial variability in the methods of modelling and measuring psychosis-like behaviours in animals.25,26 This makes it challenging to set predefined criteria for assessing the indirectness of animal experiments in the context of psychosis.\n\nNevertheless, we will aim to use the extracted data to provide an experiment-level judgment of indirectness as “low risk”, “high risk”, or “some concerns”, considering how closely the experiment reflects the clinical trial setting in terms of animal population, model induction, intervention, and outcome. This assessment will inform the evaluation of indirectness domain in the confidence in the evidence (see “Confidence in the evidence”).\n\nTo achieve this, we will evaluate the validity in animal experiments and the applicability of the intervention (e.g., treatment over an extended period, initiation of treatment after model induction) based on previous frameworks and checklists.21,58–62 Specifically, we will apply the framework of Belzung and Lemoine62 to assess the following domains (and sub-domains) of validity in animal experiments, i.e., homological (species strain), pathogenic (ontopathogenic, triggering), mechanistic, face (ethological, biomarker) and predictive (induction, remission). This framework can offer a more refined and systematic approach compared to the traditional domains of construct, face and predictive validity, which have often been inconsistently applied in the literature.25,62,63 However, the exact methods will be determined a posteriori in consultation with experts in preclinical research.\n\nPlanned comparisons\n\nOur main aim is to synthesize data for each outcome and for the comparisons described in Table-1. Meta-analysis will be conducted when there are at least two independent effect sizes for the same outcome, as in our previous systematic review.21\n\nWe will examine the data and if there is reasonable consistency across the comparisons, we will consider network meta-analysis to examining the comparative effects of the different muscarinic receptor agonists, various D2R-blocking antipsychotics, and inactive control conditions.64\n\nEffect sizes\n\nThe main effect size will be the standardized mean difference (SMD) due to the varying measures and units of the behavioural outcome measures across the studies. We will also use normalized mean difference (NMD) in a sensitivity analysis.53,65 If outcomes are reported as dichotomous, we will calculate odds ratios (ORs) and convert them into SMDs using the Hasselblad and Hedges method66 to enable their combination with results of continuous measures.\n\nIn addition to the estimation of the average treatment effects, we also aim to conduct a meta-analysis of variation to estimate the inter-individual variability of the effects using the variability ratio (VR) or the coefficient of the variation ratio (CVR) in case a mean-variance relationship is expected.65,67 This analysis will be conducted for the comparison of muscarinic receptor agonists to inactive control conditions.\n\nData synthesis approach\n\nWe will opt for synthesizing the data using multilevel meta-analytic models, which enable handling non-independent data.65 We will use a predefined multilevel random-effects structure with nested levels, from higher to lower, for animal strain, study, and experiment, provided there are at least five distinct categories for at least one of the levels, as in our previous systematic review.21 For non-independent sampling errors, we will estimate the within-study variance-covariance (VCV) matrix using any reported correlation in the original studies or assuming a correlation of 0.5 (see other assumed correlations in “Sensitivity analysis”).65 The restricted maximum likelihood (REML) method will be used to estimate the between-study variance (τ2) and between-study VCV.65,68 We will adjust the confidence intervals using t- or F-distributions with degrees of freedom appropriate for the multilevel model.69\n\nTo our knowledge, network meta-analysis has not been widely applied to the synthesis of animal experiments, and we anticipate several challenging issues, including the limited evidence for or against inconsistency, small sample sizes, and non-independent effect sizes.60 We will examine whether the assumptions of a network meta-analysis can be fulfilled by comparing the distribution of potential effect modifiers across treatment comparisons and measuring incoherence using statistical tests.70 Justifying these assumptions with preclinical data however, might be challenging. If a network meta-analysis is deemed feasible, we will extend the multilevel models and consider covariate-adjusted analysis (see “Exploration of heterogeneity”). The exact methodology will be defined a posteriori, and, if a network meta-analysis is justified, will be thoroughly reported in an amendment of the protocol before conducting it.\n\nWe will present the effect sizes with their 95% confidence intervals and prediction intervals.\n\nExploration of heterogeneity\n\nWe will quantify heterogeneity using the variance of the random effects with its components, and the 95% prediction intervals. We will explore potential sources of heterogeneity for each outcome through meta-regression or subgroup analyses, if sufficient data are available, considering the following tentative list of potential effect-modifiers: age, sex, species/strain, comorbidities, characteristics of the model of induction of psychosis-like behaviours (e.g., pharmacological or genetic, severity), muscarinic receptor agonists and their pharmacological characteristics (e.g., mode of action, selectivity for muscarinic or other receptors, potency, efficacy), dose, route of administration, duration of treatment, timing of the intervention (e.g., before or after model induction), co-treatments, characteristics of the study (e.g., publication year, reporting completeness, and risk of bias), and outcome measurements (e.g., different measures for locomotor activity). Additional variables will be considered if they are deemed relevant and there are sufficient data.\n\nWe will also aim to standardize doses across drugs to examine dose-effects relationships using the standardized dose as a covariate in a meta-regression. However, any potential standardization method cannot be predefined due to the pharmacological differences among various drugs21 and differences across species/strains.71\n\nSensitivity analysis\n\nThe robustness of the findings for each outcome will be examined through sensitivity analysis by: i) excluding studies with an overall high risk of bias (see “Assessment of risk of bias and reporting completeness”), ii) using normalized mean difference (see “Effect Sizes”), iii) excluding studies with imputed data (e.g., standard deviations, number of animals) and crossover studies reporting data from the entire period, iv) excluding interventions examined in single studies due to potentially inflated effect estimates,60 v) assuming correlations of 0.2 and 0.8 in estimating the within-study VCV,65 and vi) using a robust variance estimation to obtain cluster-robust standard errors.65,72\n\nReporting bias and small-study effects\n\nPublication and other non-reporting biases are highly prevalent in preclinical research, potentially having a substantial impact on the estimated efficacy.73,74 We will aim to evaluate potential within- and across-study non-reporting biases by adapting the framework for assessing the risk of bias due to missing evidence (ROB-ME) in clinical trials, assigning ratings of “low risk”, “high risk”, or “some concerns”.75–77 We will include both published and unpublished studies, although the latter may be difficult to find due to the limited adoption of pre-registration protocols. We will evaluate potential reasons of non-reporting of outcome data in studies excluded due to ineligible outcome measures or inadequate reporting of outcome data, although this may be more challenging due to the poor reporting quality of animal studies (see “Assessment of risk of bias and reporting completeness”). Moreover, we will explore small-study effects for each outcome using contour-enhanced funnel plots and multilevel regression-based tests,65 using the square root of the sample size, when there are sufficient data from at least 10 studies.\n\nConfidence in the evidence\n\nWe will evaluate the confidence in the evidence for each of outcome using a modified version of the GRADE framework58,59 taking into account the domains of risk of bias (and reporting completeness), indirectness, heterogeneity, imprecision, and reporting bias, similar to our previous systematic review.21 Ultimately, we will aim to draw an overall conclusion on the preclinical efficacy of muscarinic receptor agonists by considering the evidence from the different behavioural domains.\n\nStatistical software\n\nData analysis will be conducted in R statistical software78 using the package metafor,69 along with other appropriate packages for data cleaning, specific meta-analytic models, and visualization. We will report the complete list of packages used along with their versions in the publication of the results.\n\nWe plan to publish the systematic review and meta-analysis as open access in peer-reviewed journals, potentially resulting in multiple publications, and present the findings at conferences. Lay language summaries will be prepared and disseminated with the help of patient and relative groups, such as BASTA (Bündnis für psychisch erkrankte Menschen) and ApK (Aktionsgemeinschaft der Angehörigen psychisch Kranker e.V.). We will also make the methods, data, and code publicly available in a GitHub repository.\n\nAs of the date of submission of this protocol on 18.08.2024, we have completed the preliminary searches and piloting of study selection process and started, but not completed, the full searches and the full screening of search results against the eligibility criteria. We have not yet started the data extraction, risk of bias and quality assessment, or data synthesis.\n\nThere were no changes from the original PROSPERO registration of the protocol, except for expanding the methods with additional details and deciding not to search Scopus and CINAHL after consulting with the information specialist (which is not expected to affect the coverage of our search). Any additional deviations or modifications will be reported along with the findings and in updates to the PROSPERO registration.\n\n\nDiscussion\n\nThe planned systematic review and meta-analysis aims to evaluate the preclinical efficacy of muscarinic receptor agonists in animal models of relevance for psychosis. This will be achieved through a comprehensive search, advanced data synthesis methods, and a critical evaluation of potential risks of bias and confidence in the evidence. The systematic review has the potential to provide unique insights into important unanswered questions regarding muscarinic receptor agonism in the treatment of psychosis. It may also identify promising muscarinic agents or specific mechanisms of action, which could guide future drug development for schizophrenia.\n\nFurthermore, conducting a systematic review to examine the preclinical efficacy of antipsychotics is a relatively novel approach. Only a few such reviews currently exist,21,26,79 making this work particularly valuable. Therefore, our review has the potential to highlight the limitations of existing animal models of relevance for psychosis and address potential translational disconnects to improve the design of future preclinical research.6,25,31\n\nWe focused on muscarinic receptor agonism due to its potential to target the underlying pathophysiology of schizophrenia and the promising findings for agonists of M1/M4 muscarinic receptors,7–9 with xanomeline combined with trospium being reviewed for approval by the Food and Drug Administration.80 Muscarinic receptor agonism has also the most advanced data compared to other novel mechanisms of action being investigated in the treatment of psychosis.9 For example, there were encouraging results from a phase II trial for ulotaront, a dual TAAR1 agonist and 5-HT1A partial agonist.81 A recent synthesis of evidence from early- and late-stage clinical trials, and animal studies however, suggested that TAAR1 agonists may be less efficacious compared to existing D2R-blocking antipsychotics, but additional data are required to draw more definitive conclusions and more drugs are under development.21\n\nWe anticipate limitations and challenges in conducting the review. There is no gold standard animal model for psychosis,6,25,31 and cross-species differences in the muscarinic receptor system and the effects of their agonists may exist.8 Therefore, we will include all relevant animal models and behavioural readouts of preclinical efficacy, critically evaluating the confidence in the evidence to provide a comprehensive synthesis with potential translational relevance. We will not analyse neurobiological measures due to their heterogeneous use across studies and the lack of an established biomarker in schizophrenia,6 but they could further elucidate the mechanisms of muscarinic receptor agonism and its role in regulating dopaminergic signalling and other underlying pathophysiological mechanisms.8,47 We will aim to extract and potentially analyse these measures in secondary publications. Additionally, the methods for systematic reviews of animal studies in psychiatry are not well established and may present unique challenges.82 Our interdisciplinary team, which includes methodologists, statisticians, and clinical and preclinical researchers, will address any potential issues to ensure a robust synthesis of preclinical evidence.\n\nIn conclusion, our planned systematic review and meta-analysis will be the first to examine the preclinical efficacy of muscarinic receptor agonists for schizophrenia, with the potential to provide evidence-based information to guide future preclinical and clinical research on this topic.\n\n\nEthics and consent\n\nNot applicable.\n\n\nAuthors contributions\n\nSpyridon Siafis: Conceptualization, Methodology, Writing – Original Draft, Supervision, Project administration, Funding acquisition.\n\nNobuyuki Nomura: Writing – Review & Editing.\n\nJohannes Schneider-Thoma: Methodology.\n\nIrene Bighelli: Methodology.\n\nAlexandra Bannach-Brown: Methodology, Writing – Review & Editing.\n\nFiona J. Ramage: Methodology, Writing – Review & Editing.\n\nFrancesca Tinsdeall: Methodology, Writing – Review & Editing.\n\nIoannis Mantas: Methodology, Writing – Review & Editing.\n\nSameer Jauhar: Methodology, Writing – Review & Editing.\n\nSridhar Natesan: Methodology, Writing – Review & Editing.\n\nAnthony C. Vernon: Methodology, Writing – Review & Editing.\n\nAndrea de Bartolomeis: Methodology, Writing – Review & Editing.\n\nSabine M. Hölter: Methodology, Writing – Review & Editing.\n\nNatascha I. Drude: Methodology, Writing – Review & Editing.\n\nUlf Tölch: Methodology.\n\nWulf-Peter Hansen: Methodology, Writing – Review & Editing.\n\nVirginia Chiocchia: Methodology, Writing – Review & Editing.\n\nOliver Howes: Methodology.\n\nJosef Priller: Methodology.\n\nGeorgia Salanti: Methodology, Writing – Review & Editing.\n\nMalcolm R. Macleod: Methodology, Writing – Review & Editing.\n\nStefan Leucht: Conceptualization, Writing – Original Draft, Writing – Review & Editing, Supervision, Project administration, Funding acquisition", "appendix": "Data availability\n\nNo data associated with this article.\n\nZenodo: Protocol for meta-analysis on muscarinic receptor agonists in animal models of psychosis (ANIMUS-SR), https://doi.org/10.5281/zenodo.13378744. 83\n\nThis project contains the following extended data:\n\n- Draft search strategy in MEDLINE via Ovid.pdf\n\n- PRISMA-P checklist.pdf\n\nZenodo: PRISMA-P checklist for “Protocol for meta-analysis on muscarinic receptor agonists in animal models of psychosis (ANIMUS-SR)”, https://doi.org/10.5281/zenodo.13378744. 83\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 would like to thank Dr Farhad Shokraneh (Systematic Review Consultants LTD, Oxford, UK) for the collaboration in designing, revising and testing the search strategies, and in writing the search methods.\n\n\nReferences\n\nHuhn M, Nikolakopoulou A, Schneider-Thoma J, et al.: Comparative efficacy and tolerability of 32 oral antipsychotics for the acute treatment of adults with multi-episode schizophrenia: a systematic review and network meta-analysis. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPercie du Sert N, Ahluwalia A, Alam S, et al.: Reporting animal research: Explanation and elaboration for the ARRIVE guidelines 2.0. PLoS Biol. 2020; 18(7): e3000411. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHooijmans CR, de Vries RBM , Ritskes-Hoitinga M, et al.: Facilitating healthcare decisions by assessing the certainty in the evidence from preclinical animal studies. PLoS One. 2018; 13(1): e0187271. Epub 2018/01/13. eng. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSchünemann H, Brożek J, Guyatt G, et al.: The GRADE handbook. UK: Cochrane Collaboration London; 2013.\n\nLalu MM, Fergusson DA, Cheng W, et al.: Identifying stroke therapeutics from preclinical models: A protocol for a novel application of network meta-analysis. F1000Res. 2019; 8: 11. Epub 2019/03/25. eng. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHenderson VC, Kimmelman J, Fergusson D, et al.: Threats to validity in the design and conduct of preclinical efficacy studies: a systematic review of guidelines for in vivo animal experiments. PLoS Med. 2013; 10(7): e1001489. other authors have declared that no competing interests exist. Epub 2013/08/13. eng. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBelzung C, Lemoine M: Criteria of validity for animal models of psychiatric disorders: focus on anxiety disorders and depression. Biol. Mood Anxiety Disord. 2011 Nov 7; 1(1): 9. Epub 2011/01/01. eng. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWillner P: The validity of animal models of depression. Psychopharmacology. 1984; 83(1): 1–16. eng. PubMed Abstract \| Publisher Full Text\n\nSalanti G, Higgins JP, Ades AE, et al.: Evaluation of networks of randomized trials. Stat. Methods Med. Res. 2008 Jun; 17(3): 279–301. PubMed Abstract \| Publisher Full Text\n\nYang Y, Macleod M, Pan J, et al.: Advanced methods and implementations for the meta-analyses of animal models: Current practices and future recommendations. Neurosci. Biobehav. Rev. 2023 2023/03/01/; 146: 105016. PubMed Abstract \| Publisher Full Text\n\nda Costa BR , Rutjes AW, Johnston BC, et al.: Methods to convert continuous outcomes into odds ratios of treatment response and numbers needed to treat: meta-epidemiological study. Int. J. Epidemiol. 2012; 41(5): 1445–1459. PubMed Abstract \| Publisher Full Text\n\nUsui T, Macleod MR, McCann SK, et al.: Meta-analysis of variation suggests that embracing variability improves both replicability and generalizability in preclinical research. PLoS Biol. 2021; 19(5): e3001009. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJackson D, Riley R, White IR: Multivariate meta-analysis: potential and promise. Stat. Med. 2011; 30(20): 2481–2498. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nter Riet G , Korevaar DA, Leenaars M, et al.: Publication bias in laboratory animal research: a survey on magnitude, drivers, consequences and potential solutions.2012.\n\nChiocchia V, Holloway A, Salanti G: Semi-automated assessment of the risk of bias due to missing evidence in network meta-analysis: a guidance paper for the ROB-MEN web-application. BMC Med. Res. Methodol. 2023 2023/10/07; 23(1): 223. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChiocchia V, Nikolakopoulou A, Higgins JPT, et al.: ROB-MEN: a tool to assess risk of bias due to missing evidence in network meta-analysis. BMC Med. 2021 2021/11/23; 19(1): 304. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPage MJ, Sterne JAC, Boutron I, et al.: ROB-ME: a tool for assessing risk of bias due to missing evidence in systematic reviews with meta-analysis. BMJ. 2023; 383. Publisher Full Text\n\nR Core Team: R: A language and environment for statistical computing.2013.\n\nHazani R, Lavidor M, Weller A: Treatments for social interaction impairment in animal models of schizophrenia: a critical review and meta-analysis. Schizophr. Bull. 2022; 48(6): 1179–1193. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMullard A: Novel schizophrenia therapy filed for FDA approval. Nat. Rev. Drug Discov. 2023 Nov; 22(11): 862. eng. PubMed Abstract \| Publisher Full Text\n\nKoblan KS, Kent J, Hopkins SC, et al.: A Non-D2-Receptor-Binding Drug for the Treatment of Schizophrenia. N. Engl. J. Med. 2020 Apr 16; 382(16): 1497–1506. PubMed Abstract \| Publisher Full Text\n\nHair K, Wilson E, Wong C, et al.: Systematic online living evidence summaries: emerging tools to accelerate evidence synthesis. Clin. Sci. (Lond.). 2023 May 31; 137(10): 773–784. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSiafis S: Protocol for meta-analysis on muscarinic receptor agonists in animal models of psychosis (ANIMUS-SR). Zenodo. 2024. Publisher Full Text" }	[ { "id": "325666", "date": "11 Oct 2024", "name": "Daniel Foster", "expertise": [ "Reviewer Expertise I study muscarinic receptor modulation of preclinical physiology and behaviors relevant to schizophrenia." ], "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 provide a study protocol to conduct a systematic review and meta-analyses to assess pharmacological targeting of muscarinic receptors in preclinical studies relevant to schizophrenia. This is an especially timely study that has the potential to summarize a vast preclinical literature and provide insights into the efficacy of targeting muscarinic receptors and possible sources of heterogeneity in these data sets that could arise due to drugs that have different selectivity for muscarinic receptor subtypes, or different modes of pharmacological activity. The choice of primary and secondary behaviors is well justified, and the exclusion of studies that only assess neurophysiological or histopathological measures is clearly stated.\nOne potential caveat of this study protocol is the use of the pharmacological term \"agonist\" throughout the protocol. In specific sentences, the authors acknowledge the potential role for allosteric modulators in mediating antipsychotic-like effects. For example, in the interventions section, the authors write: \"We will include any pharmacological agent acting as an agonist or positive allosteric modulator at any of the five subtypes\". They then state in the Exploration of Heterogeneity section that they will consider: \"pharmacological characteristics (e.g., mode of action, selectivity for muscarinic or other receptors, potency, efficacy)\". These statements are much appreciated, and these meta-analyses could provide important insights into how positive allosteric modulators and agonists differ in efficacy in preclinical studies across numerous symptom domains. Given the potential importance of understanding how allosteric modulators and agonists are similar or differ in their ability to modulate different preclinical assays relating to numerous schizophrenia symptom domains, it may be worth changing the title and clarifying the importance of this pharmacological distinction more clearly. The agonist vs allosteric modulator issue is also evident in the search terms included in the extended data where only \"agonist\" is used and \"allosteric modulator\" is absent. In these searches, numerous allosteric modulators are included by name. However, this approach has the potential to miss any allosteric modulators not included by name in the search terms. The importance of studying potential discrepancies between agonist and allosteric modulators is highlighted by ongoing clinical trials for both types of compounds, with allosteric modulators currently being investigated in the clinic by AbbieVie, Neumora, and Neurosterix, and muscarinic agonist therapy having been approved for Karuna/Bristol Myers Squibb and under clinical investigation by several other companies including Neurocrine, Anavex, and Maplight.\nAnother potential issue that warrants attention is the use of genetically modified knockout animals. Some of the most compelling evidence we have for determining which muscarinic receptor subtypes mediate specific behavioral effects comes from studies that combine selective pharmacology tools in mice where the receptor has been deleted either globally or from specific populations of neurons. However, the current study design does not clearly outline how or if studies in these knockout animals will be evaluated.\nThe use of a similar methodology by the authors previously to assess TARR1 agonists in non-human studies is a strength, and the team assembled is well-suited to carry out this study. Overall, this study protocol is thoughtfully written and addresses important and timely questions with the potential to unveil critical insights into the ability of compounds targeting muscarinic receptors to modulate schizophrenia-relevant behaviors across numerous symptom domains in preclinical studies.\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": "13003", "date": "02 Jan 2025", "name": "Spyridon Siafis", "role": "Author Response", "response": "#1.1: The authors provide a study protocol to conduct a systematic review and meta-analyses to assess pharmacological targeting of muscarinic receptors in preclinical studies relevant to schizophrenia. This is an especially timely study that has the potential to summarize a vast preclinical literature and provide insights into the efficacy of targeting muscarinic receptors and possible sources of heterogeneity in these data sets that could arise due to drugs that have different selectivity for muscarinic receptor subtypes, or different modes of pharmacological activity. The choice of primary and secondary behaviors is well justified, and the exclusion of studies that only assess neurophysiological or histopathological measures is clearly stated. Our reply: We would like to thank the reviewer for recognizing the relevance of our planned work, for the overall endorsement of our methods, as well as for the constructive comments that further improved our methodology. Please find our point-by-point responses to the comments below. #1.2 One potential caveat of this study protocol is the use of the pharmacological term \"agonist\" throughout the protocol. In specific sentences, the authors acknowledge the potential role for allosteric modulators in mediating antipsychotic-like effects. For example, in the interventions section, the authors write: \"We will include any pharmacological agent acting as an agonist or positive allosteric modulator at any of the five subtypes\". They then state in the Exploration of Heterogeneity section that they will consider: \"pharmacological characteristics (e.g., mode of action, selectivity for muscarinic or other receptors, potency, efficacy)\". These statements are much appreciated, and these meta-analyses could provide important insights into how positive allosteric modulators and agonists differ in efficacy in preclinical studies across numerous symptom domains. Given the potential importance of understanding how allosteric modulators and agonists are similar or differ in their ability to modulate different preclinical assays relating to numerous schizophrenia symptom domains, it may be worth changing the title and clarifying the importance of this pharmacological distinction more clearly. Our reply: We would like to thank the reviewer for raising this important point. We had already planned to examine both orthosteric receptor agonists and positive allosteric modulators, as well as their differences, which we had highlighted in several parts of the manuscript. However, for simplicity, we had generally used the term 'agonism' throughout the protocol. We agree with the reviewer on the importance of this distinction, and we have clarified this in response to the reviewer’s comments. Changes in the manuscript: We have revised the title to include 'positive allosteric modulation': “Muscarinic receptor agonists and positive allosteric modulators in animal models of psychosis: protocol for a systematic review and meta-analysis.” We have incorporated 'positive allosteric modulation' more frequently throughout the manuscript to emphasize this distinction. We have clarified its significance further in the introduction: “…., and the potential differences between orthosteric agonists and PAMs. For example, PAMs modulate receptor activity by binding to a site distinct from the natural ligand, offering theoretical advantages such as increased selectivity and safety over orthosteric agonists.(1,2)”. In relationship to the comments of another reviewer,(3) we have also expanded the “Exploration of heterogeneity” section by providing more details in the pharmacological properties considered for orthosteric agonists and positive allosteric modulators: “We will examine the pharmacological properties of muscarinic receptor agonists, considering mode of action (e.g., orthosteric agonism, positive allosteric modulation) and other key characteristics as sources of heterogeneity, using data primarily from established databases (e.g., IUPHAR/BPS) 43 and original publications that characterize these compounds. Specifically, we will assess potency (half-maximal effective concentration, EC50) and efficacy (maximal response, distinguishing between full and partial agonism) for orthosteric agonists, and modulation of acetylcholine potency and efficacy (cooperative factors α and β), intrinsic efficacy (τB), and affinity for the allosteric site (KB) for PAMs.(1,4) These properties will be examined across various muscarinic receptor subtypes, with a preference for human receptors while considering potential cross-species differences, and across downstream signaling pathways (e.g., via Gαs, Gαq, Gαi/o subunits, β-arrestin). Given the variability across assays and experimental settings, we will consider calculating a common standardized index, if possible, such as estimating differences in log((maximal response)/EC50) between the examined drugs and the natural ligand acetylcholine.(4)This approach may enable comparisons across compounds, receptor and signaling pathway selectivity, but summarizing the different properties into a single index may result in information loss, necessitating additional analyses. Moreover, the exact procedure cannot be predetermined due to anticipated heterogeneity in the data across the different compounds examined.” #1.3 The agonist vs allosteric modulator issue is also evident in the search terms included in the extended data where only \"agonist\" is used and \"allosteric modulator\" is absent. In these searches, numerous allosteric modulators are included by name. However, this approach has the potential to miss any allosteric modulators not included by name in the search terms. The importance of studying potential discrepancies between agonist and allosteric modulators is highlighted by ongoing clinical trials for both types of compounds, with allosteric modulators currently being investigated in the clinic by AbbieVie, Neumora, and Neurosterix, and muscarinic agonist therapy having been approved for Karuna/Bristol Myers Squibb and under clinical investigation by several other companies including Neurocrine, Anavex, and Maplight. Our reply: We would like to thank the reviewer for highlighting this potential caveat in our search strategy. In collaboration with Dr. Farhad Shokraneh, an experienced information specialist (see Acknowledgments in the manuscript), we have revised the search strategy across multiple electronic databases to include terms relevant to allosteric modulation, ensuring we capture any studies on this category of medications. As it was already noted in the methods section, we will also supplement our searches using the psychosis-SOLES,(5)a dedicated database of preclinical psychosis studies, allowing us to conduct broader searches without restricting terms to “agonism” or “allosteric modulation”. These methods will enhance our search coverage and reduce the risk of missing relevant studies on positive allosteric modulators. Changes in the manuscript: We have revised the methods to clarify that the search strategies will encompass both agonism and allosteric modulation: 'For muscarinic agents, we will use broad terms like muscarinic agonists or allosteric modulators and…' Additionally, we have updated the draft search strategy in MEDLINE via Ovid within the extended data to include terms for “allosteric modulation”. See the revised extended data(https://doi.org/10.5281/zenodo.14534308): “...#3. Muscarinic Agonists/ or Muscarine/ or (Muscarin or Muscarine or Muscarinic or M1 or M2 or M3 or M4 or M5 or Muscarinomimetic).ti,ab,tw. #4. Allosteric Regulation/ or Allosteric Site/ or (Alloster or PAM or PAMs or Modulat* or Agonis*).ti,ab,tw. #5. 3 and 4….” #1.4: Another potential issue that warrants attention is the use of genetically modified knockout animals. Some of the most compelling evidence we have for determining which muscarinic receptor subtypes mediate specific behavioral effects comes from studies that combine selective pharmacology tools in mice where the receptor has been deleted either globally or from specific populations of neurons. However, the current study design does not clearly outline how or if studies in these knockout animals will be evaluated. Our reply: Thank you for this point. We agree that genetically modified knockout animals can be important tools for delineating the potential receptor mechanisms of pharmacological interventions. We had already planned to compare muscarinic receptor agonists or PAMs with inactive control conditions in the context of genetic or pharmacological muscarinic receptor antagonism to determine if any observed differences result from muscarinic receptor activation, as it had been outlined in Table 1 and “Animal Population and Model Induction”. However, we agree that this was not clearly described, and we have therefore revised the methods accordingly. Changes in the manuscript: We revised the “Animal Population and Model Induction” section to better describe the inclusion of animal cohorts with genetic knockouts of muscarinic receptors or those receiving pharmacological muscarinic antagonists: “….Moreover, in the eligible studies, we will extract data from “naïve” animal cohorts (i.e., animals that have not undergone models of psychosis), and animal cohorts that have undergone both models of psychosis and muscarinic receptor antagonism via genetic (i.e, genetically modified animals with knockout of all or specific muscarinic receptors, either globally or in specific neuron populations) (6) or pharmacological manipulation (e.g., co-administration of selective or non-selective muscarinic receptor antagonists like scopolamine) (7) (see further details in “Comparison groups”)….” We expanded the “Comparison groups” to mention explicitly the comparison of muscarinic drugs versus inactive control conditions (e.g., vehicle) in animal models of psychosis in the context of muscarinic receptor antagonism: “…We will also compare muscarinic receptor agonists or PAMs with inactive control conditions in contexts of genetic or pharmacological muscarinic receptor antagonism (as described in “Animal population and model induction”) to determine if any observed differences result from muscarinic receptor activation. Separate analyses may be conducted for the antagonism of specific muscarinic receptor subtypes and/or within specific neuronal populations, if sufficient data are available..” #1.5: The use of a similar methodology by the authors previously to assess TARR1 agonists in non-human studies is a strength, and the team assembled is well-suited to carry out this study. Overall, this study protocol is thoughtfully written and addresses important and timely questions with the potential to unveil critical insights into the ability of compounds targeting muscarinic receptors to modulate schizophrenia-relevant behaviors across numerous symptom domains in preclinical studies. Our reply: Thank you once again for your overall endorsement and for the thoughtful comments that have substantially strengthened our protocol. In addition to the above-mentioned changes, we have also updated the “Study status” to reflect the changes during the revision: “…There were no changes from the original PROSPERO registration of the protocol, except for expanding the methods with additional details (also in response to the reviewer’s comments),(3,8)revising the search strategy to include terms for allosteric modulation as suggested by a reviewer,(8)adding physicochemical properties of the compounds in the “Exploration of Heterogeneity” per another reviewer’s comment, (3) and deciding not to search Scopus and CINAHL after consulting with the information specialist (which is not expected to affect the coverage of our search)….” References 1.Kenakin T. Know your molecule: pharmacological characterization of drug candidates to enhance efficacy and reduce late-stage attrition. Nature Reviews Drug Discovery. 2024:1-19. 2.Foster DJ, et al. Allosteric Modulation of GPCRs: New Insights and Potential Utility for Treatment of Schizophrenia and Other CNS Disorders. Neuron. 2017;94(3):431-46. 3.Messer W. Peer Review Report For: Muscarinic receptor agonists in animal models of psychosis: protocol for a systematic review and meta-analysis [version 1; peer review: 2 approved with reservations]. F1000Research 2024;13:1017. 4.Kenakin T. A Scale of Agonism and Allosteric Modulation for Assessment of Selectivity, Bias, and Receptor Mutation. Mol Pharmacol. 2017;92(4):414-24. 5.Smith S, et al. Psychosis-SOLES 2024 [Available from: https://camarades.shinyapps.io/psychosis-solesMarch 4, 2024. 6.Thomsen M, et al. Physiological roles of CNS muscarinic receptors gained from knockout mice. Neuropharmacology. 2018;136(Pt C):411-20. 7.Klinkenberg I, et al. The validity of scopolamine as a pharmacological model for cognitive impairment: a review of animal behavioral studies. Neurosci Biobehav Rev. 2010;34(8):1307-50. 8.Foster D. Peer Review Report For: Muscarinic receptor agonists in animal models of psychosis: protocol for a systematic review and meta-analysis [version 1; peer review: 2 approved with reservations]. F1000Research. 2024;13:1017." } ] }, { "id": "325671", "date": "24 Oct 2024", "name": "William Messer", "expertise": [ "Reviewer Expertise My research focuses on the design and development of selective agonists and allosteric modulators of muscarinic receptors with potential utility in the treatment of neuropsychiatric disorders." ], "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 outlines a protocol for a specific systematic review and meta-analysis examining the impact of muscarinic agonists (and allosteric modulators) in animal models of psychosis. As such, it is more a research proposal than a paper describing results. The authors indicate that D2 dopamine antagonists have been the cornerstone for schizophrenia therapy for the last 70 years, although atypical antipsychotics target other receptors, including 5-HT2 receptors. Such compounds, including clozapine and olanzapine, are linked to a different set of adverse effects than classical dopamine antagonists including hypoglycemia and weight gain. Nevertheless, the topic is timely given the recent approval of xanomeline, in combination with trospium, for the treatment of schizophrenia. A systematic review and a meta-analysis are proposed for examining the impact of muscarinic agonists and allosteric modulators in animal models of psychosis. The level of complexity is high with respect to the varying types of compounds (agonists vs. partial agonists, receptor subtype selectivity, binding affinity, etc.), which will make it difficult to accurately categorize compounds. For example, positive allosteric modulators often exhibit a range of effects that include modulation of acetylcholine potency and efficacy in addition to allosteric agonist activity. The degree to which such activities have been adequately assessed will likely vary from lab to lab and such data may not be readily available in the papers that address in vivo efficacy in animal models of psychosis. These concerns are addressed to some extent in the “Exploration of heterogeneity” section, but a more explicit discussion of how concepts of potency and efficacy, for example, will be handled is warranted. In addition to the sources of heterogeneity discussed, the physicochemical properties of compounds may impact the effectiveness of treatments; consideration of ADME properties of compounds could provide an important perspective. For example, multiparameter optimization values could help explain variability in responses for compounds with similar receptor activity profiles. Beyond the pharmacological properties of the compounds under investigation, the impact of muscarinic receptor activation might vary significantly depending on the different compounds utilized to induce behavioral changes (e.g., NMDA antagonists vs. dopamine agonists) or the behavioral readouts (e.g., pre-pulse inhibition of startle response vs. apomorphine induced climbing). The authors refer to some of the inherent difficulties in the “Assessment of indirectness of animal experiments” section, including concerns about the applicability of various animal models to clinical utility. In this sense, it might be helpful to examine the consistency of comparator compounds (e.g., D2 and 5-HT2 antagonists) that are used clinically in treatment of schizophrenia, and are often used as control compounds in studies of muscarinic agonists and other potential therapeutic compounds. This is perhaps alluded to in the “Data synthesis/planned comparisons” section, although additional details would be helpful. An additional consideration is the presence or absence of adverse effects for the compounds under investigation. The clinical utility of muscarinic agonists has been limited by adverse effect profiles, which vary from compound to compound, but likely include both off-target (e.g., M3 receptor-mediated) and on-target (i.e., M1 or M4 receptor-mediated) effects. Adverse effects such as salivation, lacrimation, diarrhea, or convulsions would provide important information regarding the in vivo pharmacological profile of compounds and the separation between beneficial and undesirable effects (i.e., the therapeutic window). Overall, the topic is of timely interest and worth pursuing with some additional attention to the details to be evaluated.\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? Partly\n\nAre the datasets clearly presented in a useable and accessible format? Not applicable", "responses": [ { "c_id": "13005", "date": "02 Jan 2025", "name": "Spyridon Siafis", "role": "Author Response", "response": "#2.1: The article outlines a protocol for a specific systematic review and meta-analysis examining the impact of muscarinic agonists (and allosteric modulators) in animal models of psychosis. As such, it is more a research proposal than a paper describing results. Our reply: We would like to thank the reviewer for their constructive comments on our protocol, which have helped improve the clarity of the methods. This manuscript indeed corresponds to a protocol for a systematic review, and the results of the completed review will be published in a peer-reviewed journal, as outlined in the “Dissemination of information” section. Please find our point-by-point responses to the comments below. #2.2: The authors indicate that D2 dopamine antagonists have been the cornerstone for schizophrenia therapy for the last 70 years, although atypical antipsychotics target other receptors, including 5-HT2receptors. Such compounds, including clozapine and olanzapine, are linked to a different set of adverse effects than classical dopamine antagonists including hypoglycemia and weight gain. Nevertheless, the topic is timely given the recent approval of xanomeline, in combination with trospium, for the treatment of schizophrenia. Our reply: We agree with this point that although the primary mechanism of action for antipsychotics has historically been the blockade of dopamine D2 receptors, these drugs also interact with other neurotransmitter receptors, with adverse events often linked to their receptor-binding profiles. Accordingly, we have revised the introduction to incorporate this information and have also noted the recent approval of xanomeline-trospium, which had not been approved at the time of the initial manuscript submission. Changes in the manuscript: We revised the introduction to mention the association of adverse events of antipsychotics with their receptor-binding profiles and off-target actions on other neurotransmitter receptors, as well as the approval of xanomeline-trospium for schizophrenia by the US FDA: “Antipsychotic drugs that block the dopamine D2 receptor (D2R) have been the cornerstone of pharmacological treatment for schizophrenia for over 70 years. 1 – 3 …..Moreover, the risk-to-benefit ratio of antipsychotics is often challenged by their multiple side-effects associated with their receptor-binding profiles and frequently linked to off-target actions beyond D2R blockade.1 , 2 …… Xanomeline, a muscarinic M1/M4-preferring receptor agonist, has demonstrated improvements in symptoms of schizophrenia in early and late-stage randomized placebo-controlled controlled trials with medium-to-large effect sizes, with potential cholinergic adverse events mitigated by its combination with trospium, a peripheral muscarinic antagonist. 10 – 13 In September 2024, the U.S. Food and Drug Administration approved xanomeline combined with trospium for schizophrenia, marking it as the first antipsychotic targeting muscarinic receptors. (1) “ #2.3: A systematic review and a meta-analysis are proposed for examining the impact of muscarinic agonists and allosteric modulators in animal models of psychosis. The level of complexity is high with respect to the varying types of compounds (agonists vs. partial agonists, receptor subtype selectivity, binding affinity, etc.), which will make it difficult to accurately categorize compounds. For example, positive allosteric modulators often exhibit a range of effects that include modulation of acetylcholine potency and efficacy in addition to allosteric agonist activity. The degree to which such activities have been adequately assessed will likely vary from lab to lab and such data may not be readily available in the papers that address in vivo efficacy in animal models of psychosis. These concerns are addressed to some extent in the “Exploration of heterogeneity” section, but a more explicit discussion of how concepts of potency and efficacy, for example, will be handled is warranted. Our reply: We would like to thank the reviewer for this insightful comment. We have now expanded the “Exploration of heterogeneity” section to elaborate on how these pharmacological properties will be examined, taking into account different modes of action and efforts to calculate indices that minimize potential variability and heterogeneity in measuring these properties (e.g., if possible, differences in the log((maximal efficacy/EC50)).(2) We will aim to use reliable sources for obtaining information about these properties (e.g., IUPHAR/BPS database)(3); however, we will not conduct a systematic review of studies to synthesize them, and we have noted this limitation in the “Discussion” section. Changes in the manuscript: We expanded the methods in the “Exploration of heterogeneity” accordingly: We will examine the pharmacological properties of muscarinic receptor agonists, considering mode of action (e.g., orthosteric agonism, positive allosteric modulation) and other key characteristics as sources of heterogeneity, using data primarily from established databases (e.g., IUPHAR/BPS) 43 and original publications that characterize these compounds. Specifically, we will assess potency (half-maximal effective concentration, EC50) and efficacy (maximal response, distinguishing between full and partial agonism) for orthosteric agonists, and modulation of acetylcholine potency and efficacy (cooperative factors α and β), intrinsic efficacy (τB), and affinity for the allosteric site (KB) for PAMs.(2,4) These properties will be examined across various muscarinic receptor subtypes, with a preference for human receptors while considering potential cross-species differences, and across downstream signaling pathways (e.g., via Gαs, Gαq, Gαi/o subunits, β-arrestin). Given the variability across assays and experimental settings, we will consider calculating a common standardized index, if possible, such as estimating differences in log((maximal response)/EC50) between the examined drugs and the natural ligand acetylcholine.(2) This approach may enable comparisons across compounds, receptor and signaling pathway selectivity, but summarizing the different properties into a single index may result in information loss, necessitating additional analyses. Moreover, the exact procedure cannot be predetermined due to anticipated heterogeneity in the data across the different compounds examined. Additionally, we will assess off-target actions by evaluating the mode of action and affinity of each compound toward other neurotransmitter receptors.” We added to the limitations that a systematic review of in vitro assays and pharmacokinetic studies will not be conducted: “… Additionally, we will explore whether heterogeneity in behavioral effects may be explained by differences in drug interactions with muscarinic receptors. Given the limited availability and application of in vivo readouts for muscarinic agonism, we will use in vitro data. We will not conduct a systematic review of in vitro assays and pharmacokinetic studies to synthesize information about the pharmacological properties of the drugs, but instead will aim to use reliable sources and standardized methods to assess these as sources of heterogeneity (see “Exploration of heterogeneity”)…” #2.4: In addition to the sources of heterogeneity discussed, the physicochemical properties of compounds may impact the effectiveness of treatments; consideration of ADME properties of compounds could provide an important perspective. For example, multiparameter optimization values could help explain variability in responses for compounds with similar receptor activity profiles. Our reply: Thank you for raising this important point. Similarly to the above, we expanded the “Exploration of heterogeneity” section to include physicochemical parameters and their integration with the Central Nervous System Multiparameter Optimization Desirability approach. (5,6) Changes in the manuscript: We expanded accordingly the “Exploration of heterogeneity” section: “… We will explore potential sources of heterogeneity for …. type of muscarinic receptor agonists or PAMs, their pharmacological and physiochemical properties, and dose (details below), … We will use the Central Nervous System Multiparameter Optimization Desirability algorithm (5,6) to examine the physicochemical properties of the drugs as source of heterogeneity. This algorithm calculates a composite desirability score based on six key properties, i.e., lipophilicity, distribution coefficient, molecular weight, topological polar surface area, number of hydrogen bond donors, and the most basic center, which aligns well with the pharmacokinetic attributes, blood-brain barrier permeability and safety. (5,6) Values for these properties will be estimated based on the chemical structure information using OPEn structure–activity/property Relationship App (OPERA) version 2.9.1 (a freely available, open-source provided distributed under MIT license).(7)” #2.5: Beyond the pharmacological properties of the compounds under investigation, the impact of muscarinic receptor activation might vary significantly depending on the different compounds utilized to induce behavioral changes (e.g., NMDA antagonists vs. dopamine agonists) or the behavioral readouts (e.g., pre-pulse inhibition of startle response vs. apomorphine induced climbing). The authors refer to some of the inherent difficulties in the “Assessment of indirectness of animal experiments” section, including concerns about the applicability of various animal models to clinical utility. In this sense, it might be helpful to examine the consistency of comparator compounds (e.g., D2 and 5-HT2 antagonists) that are used clinically in treatment of schizophrenia, and are often used as control compounds in studies of muscarinic agonists and other potential therapeutic compounds. This is perhaps alluded to in the “Data synthesis/planned comparisons” section, although additional details would be helpful. Our reply: We agree with this point, and we have added a comparison of D2R-blocking antipsychotics with inactive control conditions (e.g., vehicle) to examine the consistency of their effects in the included experiments and to inform the “Assessment of indirectness of animal experiments”. This is now described in the “Comparison groups” Changes in the manuscript: We added this comparison in the “Comparison groups”: “Table 1 outlines the planned comparisons, …. Additionally, we will examine the comparison between active and inactive control conditions to provide insights into the consistency of the effects of D2R-blocking antipsychotic in the included experiments and to inform the assessment of indirectness (see “Assessment of indirectness of the animal experiments”).” #2.6: An additional consideration is the presence or absence of adverse effects for the compounds under investigation. The clinical utility of muscarinic agonists has been limited by adverse effect profiles, which vary from compound to compound, but likely include both off-target (e.g., M3receptor-mediated) and on-target (i.e., M1 or M4 receptor-mediated) effects. Adverse effects such as salivation, lacrimation, diarrhea, or convulsions would provide important information regarding the in vivo pharmacological profile of compounds and the separation between beneficial and undesirable effects (i.e., the therapeutic window). Our reply: Thank you for this point. We agree that examining adverse events is important for assessing the risk-to-benefit ratio of these drugs. However, we had chosen already from the start of our protocol to focus on behavioural outcomes and not include adverse events due to their sparse and inconsistent reporting in studies of animal models of psychosis (as also noted in our previous review on TAAR1 agonists)(8) and their examination in other study designs excluded from our review (e.g., using naïve animals not subjected to models of psychosis). Furthermore, cholinergic adverse events may potentially be mitigated in clinical settings through the combination of peripheral antagonists, as seen with xanomeline-trospium.(9)Nonetheless, we will aim to extract adverse event data if found in the included studies and consider secondary publications where relevant. These considerations are addressed in the “Discussion” section under limitations. Changes in the manuscript: We expanded the limitations in the “Discussion” section to include consideration of cholinergic adverse events, similar to the previously reported neurobiological measures: “We will not analyse neurobiological measures due to their heterogeneous use across studies and the lack of an established biomarker in schizophrenia, 6 .. Similarly, we will not examine cholinergic adverse events in animals, such as salivation, lacrimation and diarrhea, which may offer insights into the risk-benefit profile but could be mitigated in clinical settings with peripheral muscarinic receptor antagonists, as shown with the xanomeline-trospium combination. 10 – 13 We will aim to extract and potentially analyse these measures in secondary publications.” #2.7: Overall, the topic is of timely interest and worth pursuing with some additional attention to the details to be evaluated. Our reply: Thank you once again for your overall endorsement and for the thoughtful comments that have substantially strengthened our protocol. In addition to the above-mentioned changes, we have also updated the “Study status” to reflect the changes during the revision: “…There were no changes from the original PROSPERO registration of the protocol, except for expanding the methods with additional details (also in response to the reviewer’s comments),(10,11)revising the search strategy to include terms for allosteric modulation as suggested by a reviewer,(10)adding physicochemical properties of the compounds in the “Exploration of Heterogeneity” per another reviewer’s comment, (11) and deciding not to search Scopus and CINAHL after consulting with the information specialist (which is not expected to affect the coverage of our search)….” References 1.U.S. Food and Drug Administration. FDA Approves Drug with New Mechanism of Action for Treatment of Schizophrenia 2024 [Available from: https://www.fda.gov/news-events/press-announcements/fda-approves-drug-new-mechanism-action-treatment-schizophrenia. 2.Kenakin T. A Scale of Agonism and Allosteric Modulation for Assessment of Selectivity, Bias, and Receptor Mutation. Mol Pharmacol. 2017;92(4):414-24. 3.Harding SD, et al. The IUPHAR/BPS guide to PHARMACOLOGY in 2022: curating pharmacology for COVID-19, malaria and antibacterials. Nucleic Acids Res. 2022;50(D1):D1282-d94. 4.Kenakin T. Know your molecule: pharmacological characterization of drug candidates to enhance efficacy and reduce late-stage attrition. Nature Reviews Drug Discovery. 2024:1-19. 5.Wager TT, et al. Moving beyond rules: the development of a central nervous system multiparameter optimization (CNS MPO) approach to enable alignment of druglike properties. ACS Chem Neurosci. 2010;1(6):435-49. 6.Wager TT, et al. Central Nervous System Multiparameter Optimization Desirability: Application in Drug Discovery. ACS Chem Neurosci. 2016;7(6):767-75. 7.Mansouri K, et al. OPERA models for predicting physicochemical properties and environmental fate endpoints. Journal of Cheminformatics. 2018;10(1):10. 8.Siafis S, et al. Trace amine-associated receptor 1 (TAAR1) agonism for psychosis: a living systematic review and meta-analysis of human and non-human data. Wellcome Open Research. 2024;9. 9.Paul SM, et al. Muscarinic Acetylcholine Receptor Agonists as Novel Treatments for Schizophrenia. Am J Psychiatry. 2022;179(9):611-27. 10.Foster D. Peer Review Report For: Muscarinic receptor agonists in animal models of psychosis: protocol for a systematic review and meta-analysis [version 1; peer review: 2 approved with reservations]. F1000Research. 2024;13:1017. 11.Messer W. Peer Review Report For: Muscarinic receptor agonists in animal models of psychosis: protocol for a systematic review and meta-analysis [version 1; peer review: 2 approved with reservations]. F1000Research 2024;13:1017." } ] } ]	1	https://f1000research.com/articles/13-1017
https://f1000research.com/articles/13-1225/v1	14 Oct 24	{ "type": "Clinical Practice Article", "title": "Factors influencing osteoradionecrosis progression during hyperbaric oxygen therapy: A case study", "authors": [ "Sameh Mezri", "Chaima Zitouni", "Khadija Bahrini", "Mounir Haggui", "Wiem Boughzala", "Hedi Gharsallah", "Sameh Mezri", "Khadija Bahrini", "Mounir Haggui", "Wiem Boughzala", "Hedi Gharsallah" ], "abstract": "Introduction Although relatively uncommon, osteoradionecrosis (ORN) remains a serious complication following radiotherapy. Various therapeutic approaches, including hyperbaric oxygen therapy (HBOT), are utilized in managing ORN. This study aims to evaluate the role of HBOT in ORN management and to identify predictive factors influencing the evolution of head and neck ORN after HBOT.\n\nMethods This retrospective study includes 46 patients who received HBOT for head and neck ORN between 2017 and 2020. The patients were divided into two groups: Group 1 (n=36) included those with regression or stabilization of ORN, while Group 2 (n=10) comprised patients with worsening lesions. We performed a statistical study in order to identify factors influencing ORN progression under treatment.\n\nResults ORN affected the mandible in 93.5% of patients, the maxilla in 2 cases, and the skull base in 4 cases. All patients received HBOT, with an average of 44.65 sessions. Pre-operative HBOT was administered in 17% of cases, and post-operative HBOT was given in 42% of cases. After at least 20 sessions, ORN regressed in 33% of cases, stabilized in 45%, and worsened in 22%. Analysis of factors influencing ORN progression on the univariate study revealed significant associations with high blood pressure (p=0.046), larger tumor size (p=0.004), advanced tumor stages (p=0.048), mean radiation dose (p=0.002), delays between dental care and radiotherapy (p=0.045), and the location of ORN within the mandible (p=0.049). Additionally, the number of HBOT sessions significantly affected ORN evolution, with more sessions correlating with better outcomes (p=0.001). In the multivariate analysis, variables such as the average interval between dental care and radiotherapy (p=0.043) as well as the number of HBOT sessions (p=0.040) emerged as significant influencers of ORN evolution.\n\nConclusion Our study provides valuable insights into the management of ORN by identifying key predictors that influence the post-therapeutic evolution of head and neck ORN after HBOT.", "keywords": [ "Osteoradionecrosis", "Radiotherapy", "Hyperbaric oxygen therapy", "Head and neck" ], "content": "Introduction\n\nHead and neck cancers rank seventh in the world, with an incidence of 890,000 new cases.1\n\nTheir management typically involves surgery, radiotherapy (RT), and chemotherapy. Despite advancements in techniques, external RT often leads to complications, including osteoradionecrosis (ORN).\n\nORN is defined as progressive bone destruction, occurring spontaneously or following trauma, with mucosal ulceration exposing irradiated bone, persisting for 3 to 6 months without healing, excluding tumor recurrence.2\n\nAlthough relatively uncommon, ORN remains a serious complication, significantly affecting patients’ quality of life.\n\nSince its first description, the pathophysiology of ORN remains unclear, and management strategies are complex and lack consensus.\n\nVarious therapeutic approaches are utilized in the treatment of ORN. They include antibiotics, hyperbaric oxygen therapy (HBOT), the PENTOCLO protocol, and conservative surgery.\n\nOur study aims to evaluate the role of HBOT in ORN management and describe predictive factors influencing post-therapeutic evolution of head and neck ORN.\n\n\nMethods\n\nThis is a retrospective study conducted at the HBOT Department of the Military Hospital of Tunis over a period of 4 years, from January 1, 2017, to December 31, 2020. The study included patients who met the following criteria:\n\n- Treated with HBOT for head and neck ORN\n\n- Completed at least 20 HBOT sessions\n\n- Had a complete medical record\n\n- Were available for follow-up evaluations for at least six months after the end of the therapy\n\nAll cases with recurrent loco-regional tumor and who received discontinued or irregular HBOT sessions were excluded from this study.\n\nFor this study, we developed a specialized data collection tool called a case report form (CRF). This form was used to document various aspects of each patient’s case, including their demographics, clinical examination, laboratory results, imaging, treatment, and follow-up.\n\nORN was classified into four grades according to the LENT/SOMA (Late Effects of Normal Tissues/Subjective Objective Management Analytic) classification.3\n\nGrade 1: Normal bone appearance or debatable modifications\n\nGrade 2: Bone lysis or condensation\n\nGrade 3: Bone sequestration\n\nGrade 4: Bone fracture\n\nHBOT sessions were conducted in a multiplace hyperbaric chambers type HAUX-STARMED 2400. The HBOT protocol included 3 steps:\n\n➢ Step 1: Compression phase: Around 0.1 ATA (Atmosphere Absolute) and the Compression rate was adjusted based on patient’s tolerance.\n\n➢ Step 2: Plateau phase: in this phase patients received 100% oxygen using facial mask for 60 and or 90 minutes with 5-minute air breaks every 25 minutes.\n\n➢ Step 3: decompression phase: this step is slow typically around 0.1 ATA/min.\n\nPatients were followed up during and following HBOT therapy. Clinical and imaging evolution of injuries was recorded.\n\nRadiological examinations, if needed, were performed after at least 40 HBOT sessions. However, if the hyperbaric medicine specialist noticed any signs of worsening condition after 20 HBOT sessions, CT scans were ordered earlier. In our study, only CT was used for radiological control.\n\nORN injuries were assessed clinically and by imaging following HBOT treatment. Thereby lesion evolution was classified into three categories:\n\n- Regression: Clinically and/or radiologically confirmed the disappearance of functional and radiological signs\n\n- Stabilization: It involves partial healing of the ORN with no progression of bone necrosis. Functional signs are moderate.\n\n- Worsening or Aggravation: Progressive worsening of the ORN characterized by the persistence of functional and radiological signs. There is no bone healing and the ORN was progressed to a more severe stage.\n\nStatistical analyses were performed using SPSS version 22.0. Categorical variables were reported as percentages, and their significance was assessed using Fisher’s exact test or chi-square tests. Quantitative variables were presented as mean (standard deviation) or median ± interquartile range (IQR25%, 75%). Normality was tested using the Kolmogorov-Smirnov test. An unpaired t-test was used to compare two groups when continuous variables were normally distributed. For continuous variables that were not normally distributed, a two-tailed unpaired Mann-Whitney U test was applied. Both univariate and multivariate analyses were conducted to evaluate the effectiveness of HBOT on ORN patients. The cases were divided into two groups: Group 1 (n=36) included patients with regression or stabilization, and Group 2 (n=10) included patients with worsening signs of injury after HBOT (Figure 1). Differences were considered significant if p ≤ 0.05.\n\nThis study was approved by the Ethics Committee of the Military Hospital of Tunis, under decision number 81/2024/CLPP, dated July 22, 2024. The study was conducted in accordance with the ethical principles outlined in the Declaration of Helsinki (https://www.wma.net/policies-post/wma-declaration-of-helsinki-ethical-principles-for-medical-research-involving-human-subjects/).\n\n\nResults\n\nThis study included 46 patients. The average age of patients was 58 ± 13.9 years, with the majority of patients (61%) aged between 31 and 65 years old. Fifty-two percent were female, and 48% were male. Two-thirds of the patients had diabetes and 59% were smokers.\n\nAll patients had undergone RT for head and neck malignancies, primarily nasopharyngeal carcinoma (67%) (Table 1).\n\nThe mean tumor size was 29.67 mm.\n\nRT was conventional in 40 cases, cobalt-60 2D dimensional in two cases, three-dimensional in one case, and Intensity-Modulated Radiation Therapy (IMRT) in three cases; with a mean radiation dose of 48.7 Gy. The association with chemotherapy was noted in thirty cases, accounting for 65% of the cases.\n\nDental care was provided to all patients before RT, with 67% requiring tooth extractions. Twelve patients used the custom fluoride tray correctly. The interval between dental care and the start of RT ranged from one month to 48 months, with a median of two months.\n\nORN typically occurred due to dental extractions (83% of cases) or surgical procedures (15%). The median time from RT to ORN onset was 6 years, while from dental care to ORN onset was 36 months.\n\nCommon clinical symptoms included pain (100% of patients) and mastication difficulties (98%). Trismus was present in all patients, with varying degrees of severity. Dental mobility was observed in 33% of cases.\n\nWe also noted the presence of bone exposure (Figure 2) in 6 cases, bone sequestration in 15 cases, or bone fracture in 9 cases. The presence of a cutaneous fistula (Figure 3) was noted in 39% of cases.\n\nWe performed panoramic radiograph (76%), cone beam imaging (11%), computed tomography (CT scan) (80%), and magnetic resonance imaging (MRI) (28%).\n\nImaging revealed the following grades of ORN according to the LENT/SOMA classification: Grade 2 (43% of cases), Grade 3 (37%), and Grade 4 (20%).\n\nORN was predominantly mandibular (40 patients), affecting various regions such as the horizontal branch (27 patients) (Figure 4), mandibular angle (10 patients), and ascending branch (3 patients). Other locations included the maxilla (2 cases) and the skull base (4 cases) (Figure 5).\n\nCT scan showing osteoradionecrosis of the horizontal branch of the left mandible complicated by sequestra (star) and mandibulo-cutaneous fistula (yellow arrow).\n\nA: Sagittal section.\n\nB: Frontal section.\n\nC: 3D reconstruction.\n\nA and B: Coronal section showing of a CT-scan showing bilateral lytic erosion of the tympanic bone (star) and thickening of the walls of the external auditory canal.\n\nThe anatomopathological examination was performed in one case due to delayed alveolar consolidation and diagnostic doubt for differential diagnosis. It concluded with fibro-inflammatory chronic scarring remodeling, displaying features of ORN without specific granuloma or signs of malignancy.\n\nThe therapeutic management of ORN included topical antibacterial mouthwash prescribed to all patients. All patients received analgesic treatment based on their pain intensity. Antibiotic therapy was initiated in 89% of cases.\n\nSurgical treatment was performed in 36 patients, predominantly involving sequestrectomy (94% of cases). Mandibulectomy with mandibular reconstruction using flaps was conducted in 6% of cases.\n\nHBOT was provided to all patients, with a mean of 44.65 sessions. HBOT sessions preceded ORN surgery in 17% of cases and were indicated after surgery in 42% of cases.\n\nComplications of HBOT were observed in two patients, including middle ear barotrauma, necessitating transient discontinuation of therapy.\n\nEvolution of ORN lesions was evaluated clinically after 20 HBOT sessions and radiologically after 40 sessions. Results showed regression in 33% of cases, stabilization in 45%, and progression or aggravation in 22% (Figure 6).\n\nIn the univariate study (Table 2), age and sex did not show statistical significance in influencing ORN progression. However, among comorbidities, hypertension was significantly associated with worsened ORN evolution.\n\nOur study also revealed that larger tumor size and advanced tumor stages correlated with worse outcomes, while tumor location did not statistically influence ORN evolution.\n\nThe type of radiotherapy did not statistically influence the progression of ORN lesions, nor did its association with chemotherapy. Higher radiation therapy doses statistically influenced ORN evolution.\n\nDelays between dental care (before RT) and RT were associated with worse ORN evolution, while dental care after RT did not significantly influence it.\n\nThe location of ORN within the mandible significantly influenced lesion evolution, but radiological signs did not.\n\nThe use of antibiotics did not statistically influence the progression of ORN lesions. HBOT significantly influenced ORN evolution, with more sessions correlating with better outcomes. We observed that the progression of lesions in patients who underwent surgery was not influenced by the timing of surgery, whether it was before, concurrent with, or after HBOT therapy. The type of surgical treatment did not statistically influence lesion evolution.\n\nIn the multivariate analysis (Table 3), variables such as the average interval between dental care (before RT) and RT and the number of HBOT sessions emerged as significant influencers of ORN evolution. Specifically, extending the delay between dental care and RT by one month correlated with a sevenfold enhancement in ORN lesion evolution. Furthermore, each additional ten HBOT sessions were linked with a tenfold improvement in ORN lesion evolution.\n\n\nDiscussion\n\nIn 1973, Mainous et al.4 first proposed HBOT as a treatment for ORN. Subsequent studies have highlighted the significant benefits of HBOT in managing ORN, primarily due to its ability to deliver hyper-concentrated oxygen levels, up to 20 times higher than normal conditions. HBOT facilitates increased oxygen diffusion in hypoxic tissues, stimulates osteogenesis and mucosal tissue epithelialization, promotes collagen synthesis and osteoblast proliferation in irradiated tissues, enhances the expression of Vascular Endothelial Growth Factor (VEGF) leading to angiogenesis, improves tissue oxygen perfusion, and exerts a bactericidal and bacteriostatic effect against various pathogens. These mechanisms collectively underscore the therapeutic potential of HBOT in the management of ORN.5\n\nSeveral studies have reported healing rates ranging from zero to 100%.6–11 However, comparing these results is challenging due to the variability of protocols applied and the variability in case indications (varying severity of ORN cases).\n\nMany authors have demonstrated improvement in ORN with HBOT.12–14\n\nBesides its therapeutic effect on ORN lesions, HBOT can help improve patients’ quality of life by reducing pain, promoting wound healing, and lowering the risk of infectious complications.15\n\nHowever, Annane et al.16 halted their trial due to a significantly better healing rate in the placebo group (32%) compared to the HBOT group (19%). The main criticisms of this trial were the monomodal use of HBOT in treating his patients and the lack of strict and clear criteria for defining ORN and its severity.\n\nAccording to recommendations established by medical organizations such as the Undersea and Hyperbaric Medical Society (UHMS)15 and the Tenth European Consensus Conference on Hyperbaric Medicine,17 HBOT is recommended for treating symptomatic ORN cases or as an adjunct treatment to surgical intervention in order to enhance wound healing, reduce infection risk, and promote tissue healing after reconstructive or debridement surgery.\n\nAccording to the UHMS, to achieve these effects, the recommended pressure should be equal to or greater than 1.4 atmospheres (atm). However, all current indications approved by the UHMS require patients to breathe nearly 100% oxygen in a pressurized chamber at a minimum pressure of 2 ATA.15 However, results appear controversial and inconclusive. Thus, at the limit of our literature search, only one randomized clinical trial has been published, with other studies mainly consisting of cohort studies of varying quality.\n\nA study by Annane et al.16 examined the efficacy of HBOT in the treatment of ORN and found that patients who received a higher number of HBOT sessions had higher healing rates.\n\nA meta-analysis by Bennett et al. (2016)18 also examined the results of several studies on HBOT in the treatment of ORN. They found that healing rates were higher in patients who received a higher number of HBOT sessions, although the results varied depending on the healing criteria used in the different studies.\n\nMarx et al.19 reported healing rates of up to 85% in patients who received more than 20 HBOT sessions. This was applicable to our study where a minimum of 20 sessions was indicated for our patients.\n\nThe combination of HBOT and surgical treatment has been addressed in the literature. Several authors have reported ORN healing rates of 15% to 45% with HBOT alone and 20% to 90% when HBOT was combined with surgery.20–22 However, these studies lack precision with somewhat heterogeneous groups.\n\nIn a recently published multicenter randomized trial in 2021, Forner et al.23 reported a significantly better healing rate in the HBOT-surgery group (surgical treatment preceded by 30 HBOT sessions and followed by 10 HBOT sessions) at 70% (21/30) compared to 51% (18/35) in the surgery-only group. HBOT was associated with improved healing rates, regardless of ORN severity. It also reduced the severity of xerostomia and dysphagia and improved total unstimulated salivary flow. However, despite the most supported multimodal approach to ORN management being the combination of HBOT and surgery, due to the divergent conclusions of the literature, establishing standardized protocols for HBOT use in parallel with surgery seems compromised.\n\nThe evolution of ORN under treatment can be influenced by several factors.\n\nOh et al.,11 studied factors influencing ORN evolution, collecting 114 patients treated for ORN over a 16-year period. They were divided into two groups: group 1 of 47 patients treated with conservative treatment (sequestrectomy, debridement, and/or HBOT) and group 2 of 67 patients treated with immediate intervention or after failure of conservative treatment.\n\nPatients whose ORN was associated with an early-stage tumor or extraction before irradiation responded favorably to conservative treatment. However, patients with advanced primary tumors, who continued to smoke and drink after RT, who received palliative RT or a radiation dose exceeding 60Gy, and who had oro-cutaneous fistulas, pathological fractures, swelling, or trismus responded poorly to conservative treatment. In these latter cases, radical resection of the affected tissue proved useful.\n\nThe onset time of ORN relative to RT is a factor influencing the evolution of ORN lesions. This notion was supported by Oh et al.,11 who reported that patients whose ORN occurred within 12 months after RT had a higher resolution rate with conservative treatment than patients whose ORN occurred after 12 months.\n\nBeumer et al.24 reported that ORN occurring after a dose of 70 Gy did not systematically respond to conservative treatment measures, thus requiring non-conservative treatment.\n\nIn the same study, the authors reported that ORN occurring due to irritation from dental prostheses or extraction before irradiation respond more effectively to conservative treatment than ORN occurring due to dental disease, either spontaneously or in association with post-irradiation extraction. The latter often requires a radical approach; this result was refuted by Oh et al.11\n\nIn a retrospective study conducted by De Felice25 published in 2016, comparing resolved ORN and unresolved ORN, no factor was identified as influencing the resolution or progression of ORN during logistic regression.\n\nIn our study, the evolution of ORN lesions was influenced by the presence of hypertension, tumor size, T and N stages, RT dose, dental care delays relative to RT, ORN location within the mandible, and HBOT sessions. During logistic regression, only delays in dental care relative to RT would influence the evolution of ORN lesions.\n\nDespite being the only study conducted in Tunisia on this topic, this research has several limitations that should be acknowledged. First, as a retrospective study, it has missing clinical data. Second, evaluating the efficacy of HBOT ideally requires a control group, which could potentially be addressed with a matched sample from another center. Third, the limited number of cases and the heterogeneity among patients in terms of therapeutic modalities prevent definitive conclusions. For these reasons, further research is needed to gain a deeper understanding of the therapeutic strategies for ORN tailored to individual patient profiles.\n\nDespite these limitations, our study offers valuable insights into the management of ORN within the Tunisian context and provides a foundation for future, more comprehensive investigations.\n\n\nConclusion\n\nIn conclusion, our study yields crucial insights into the management of ORN by identifying significant predictors influencing post-therapeutic evolution of head and neck ORN. These findings underscore the multifactorial nature of ORN progression, implicating patient characteristics, tumor attributes, and treatment modalities.\n\nNotably, advanced T stage, higher RT doses, and shorter delays between dental care and RT initiation were associated with worsened ORN evolution. Conversely, longer delays between dental care and RT initiation, mandibular horizontal branch localization and increased number of HBOT sessions were associated with improved lesion evolution. Logistic regression identified delay between dental care and RT initiation and number of HBOT sessions as independent factors influencing lesion evolution.\n\nThese findings highlight the importance of timely intervention and comprehensive treatment strategies in reducing ORN progression and improving patient outcomes. Moreover, our study underscores the potential of HBOT as a valuable adjunctive treatment option in ORN management. However, further research is warranted to validate these findings and develop targeted therapeutic approaches tailored to individual patient profiles.\n\n\nConsent for publication\n\nAll photographs included in this manuscript have been published with the explicit written consent of the individuals depicted. Written consent to publish these images was obtained from the participants prior to submission of the manuscript. Any identifiable features, such as personal details or medical record numbers, have been removed to ensure privacy and confidentiality.\n\n\nEthical and consent\n\nThis study was approved by the Ethics Committee of the Military Hospital of Tunis, under decision number 81/2024/CLPP, dated July 22, 2024. The study was conducted in accordance with the ethical principles outlined in the Declaration of Helsinki (https://www.wma.net/policies-post/wma-declaration-of-helsinki-ethical-principles-for-medical-research-involving-human-subjects/).\n\nThe research was conducted ethically, with all study procedures being performed in accordance with the requirements of the World Medical Association’s Declaration of Helsinki.\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.\n\n\nAuthor contributions\n\nDr Sameh Mezri: Conceptualization, Methodology\n\nDr Chaima Zitouni: Writing – Original Draft Preparation – Review and editing\n\nDr Khadija Bahrini: Formal Analysis\n\nDr Mounir Haggui: Supervision\n\nDr Wiem Boughzala: Investigation\n\nDr Hedi Gharsallah: Resources", "appendix": "Data availability statement\n\nThe underlying data for this project are available in the Zenodo repository.\n\nZenodo: Factors influencing osteoradionecrosis progression during hyperbaric oxygen therapy: A case study\n\n1. https://doi.org/10.5281/zenodo.13901089 26\n\nThis project contains the following data:\n\nCase Report Form\n\n2. https://doi.org/10.5281/zenodo.13901073 27\n\nThis project contains the following data:\n\nParticipant Information Sheet\n\nDetails of license: The data files are published under a Creative Commons Attribution 4.0 International license (CC BY 4.0).\n\n\nReferences\n\nMody MD, Rocco JW, Yom SS, et al.: Head and neck cancer. Lancet. 2021 Dec; 398(10318): 2289–2299. Publisher Full Text\n\nLee IJ, Koom WS, Lee CG, et al.: Risk factors and dose-effect relationship for mandibular osteoradionecrosis in oral and oropharyngeal cancer patients. Int. J. Radiat. Oncol. Biol. Phys. 2009; 75(4): 1084–1091. PubMed Abstract \| Publisher Full Text\n\nNabil S, Samman N: Risk factors for osteoradionecrosis after head and neck radiation: a systematic review. Oral Surg. Oral Med. Oral Pathol. Oral Radiol. 2012; 113: 54–69. PubMed Abstract \| Publisher Full Text\n\nMainous EG, Hart GB: Osteoradionecrosis of the mandible. Treatment with hyperbaric oxygen. Arch. Otolaryngol. Chic. Ill 1960. Mar 1975; 101(3): 173–177.\n\nDieleman FJ, Phan TTT, van den Hoogen FJA , et al.: The efficacy of hyperbaric oxygen therapy related to the clinical stage of osteoradionecrosis of the mandible. Int. J. Oral Maxillofac. Surg. 2017; 46(4): 428–433.\n\nOwosho AA, Tsai CJ, Lee RS, et al.: The prevalence and risk factors associated with osteoradionecrosis of the jaw in oral and oropharyngeal cancer patients treated with intensity-modulated radiation therapy (IMRT): The Memorial Sloan Kettering Cancer Center experience. Oral Oncol. 2017; 64: 44–51. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nD’Souza J, Goru J, Goru S, et al.: The influence of hyperbaric oxygen on the outcome of patients treated for osteoradionecrosis: 8 year study. Int. J. Oral Maxillofac. Surg. 2007; 36(9): 783–787. PubMed Abstract \| Publisher Full Text\n\nChen JA, Wang CC, Wong YK, et al.: Osteoradionecrosis of mandible bone in patients with oral cancer—associated factors and treatment outcomes. Head Neck. 2016; 38(5): 762–768. PubMed Abstract \| Publisher Full Text\n\nGupta P, Sahni T, Jadhav GK, et al.: A Retrospective Study of Outcomes in Subjects of Head and Neck Cancer Treated with Hyperbaric Oxygen Therapy for Radiation Induced Osteoradionecrosis of Mandible at a Tertiary Care Centre: An Indian Experience. Indian J. Otolaryngol. Head Neck Surg. 2013; 65(S1): 140–143. Publisher Full Text\n\nHampson NB, Holm JR, Wreford-Brown CE, et al.: Prospective assessment of outcomes in 411 patients treated with hyperbaric oxygen for chronic radiation tissue injury: Hyperbaric Oxygen for Radiation Injury. Cancer. 2012; 118(15): 3860–3868. PubMed Abstract \| Publisher Full Text\n\nOh HK, Chambers MS, Martin JW, et al.: Osteoradionecrosis of the mandible: treatment outcomes and factors influencing the progress of osteoradionecrosis. J. Oral Maxillofac. Surg. 2009; 67(7): 1378–1386. PubMed Abstract \| Publisher Full Text\n\nDieleman FJ, Phan TTT, van den Hoogen FJA , et al.: The efficacy of hyperbaric oxygen therapy related to the clinical stage of osteoradionecrosis of the mandible. Int. J. Oral Maxillofac. Surg. 2017; 46(4): 428–433. PubMed Abstract \| Publisher Full Text\n\nNiezgoda JA, Serena TE, Carter MJ: Outcomes of Radiation Injuries Using Hyperbaric Oxygen Therapy: An Observational Cohort Study. Adv. Skin Wound Care. 2016 Jan; 29(1): 12–19. Publisher Full Text\n\nTahir ARM, Westhuyzen J, Dass J, et al.: Hyperbaric oxygen therapy for chronic radiation-induced tissue injuries: Australasia’s largest study. Asia Pac. J. Clin. Oncol. 2015; 11(1): 68–77. PubMed Abstract \| Publisher Full Text\n\nWeaver LK: Undersea and Hyperbaric Medical Society (UHMS), editors. Hyperbaric oxygen therapy indications. 14th ed.Durham, NC: Undersea and Hyperbaric Medical Society; 2019.\n\nAnnane D, Depondt J, Aubert P, et al.: Hyperbaric Oxygen Therapy for Radionecrosis of the Jaw: A Randomized, Placebo-Controlled, Double-Blind Trial From the ORN96 Study Group. J. Clin. Oncol. 2004; 22(24): 4893–4900. PubMed Abstract \| Publisher Full Text\n\nMathieu D, Marroni A, Kot J: Tenth European Consensus Conference on Hyperbaric Medicine: recommendations for accepted and non-accepted clinical indications and practice of hyperbaric oxygen treatment. Diving Hyperb. Med. 2017; 47(1): 24–32. PubMed Abstract \| Publisher Full Text\n\nBennett MH, Feldmeier J, Hampson NB, et al.: Hyperbaric oxygen therapy for late radiation tissue injury. Cochrane Database Syst. Rev. 2016; 4(4): CD005005. PubMed Abstract \| Publisher Full Text\n\nMarx RE, Ehler WJ, Tayapongsak P, et al.: Relationship of oxygen dose to angiogenesis induction in irradiated tissue. Am. J. Surg. 1990; 160(5): 519–524. PubMed Abstract \| Publisher Full Text\n\nMarx RE: A new concept in the treatment of osteoradionecrosis. J. Oral Maxillofac. Surg. 1983; 41(6): 351–357. PubMed Abstract \| Publisher Full Text\n\nMcLeod NM, Bater MC, Brennan PA: Management of patients at risk of osteoradionecrosis: results of survey of dentists and oral & maxillofacial surgery units in the United Kingdom, and suggestions for best practice. Br. J. Oral Maxillofac. Surg. 2010; 48(4): 301–304. PubMed Abstract \| Publisher Full Text\n\nMeyer I: Infectious diseases of the jaws. J. Oral Surg. Am. Dent. Assoc. 1970; 28(1): 17–26.\n\nForner D, Phillips T, Kovacs AF, et al.: Hyperbaric Oxygen Therapy Combined With Surgical Treatment for Osteoradionecrosis of the Mandible: A Multicenter Randomized Trial. JAMA Otolaryngol. Head Neck Surg. 2021; 147(7): 602–610.\n\nBeumer J, Harrison RE, Sanders B, et al.: Osteoradionecrosis: predisposing factors and outcomes of therapy. Head Neck Surg. 1984; 6(5): 819–827. Publisher Full Text\n\nDe Felice F, Tombolini V, Ruoppolo G, et al.: Osteoradionecrosis (ORN) of the jaws: definition, pathogenesis, clinical manifestations and management strategies. A literature review. Radiol. Med. 2014; 119(10): 712–721.\n\nZitouni C: Case report form. Zenodo. 2024. Publisher Full Text\n\nZitouni C: Participant Information Sheet. Zenodo. 2024. Publisher Full Text" }	[ { "id": "332796", "date": "25 Oct 2024", "name": "Atsushi Shudo", "expertise": [ "Reviewer Expertise oral and maxillofacial surgery", "oral oncology", "jaw deformity", "dental infection", "osteonecrosis of the jaws", "osteoradionecrosis" ], "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, Thanks for the chance to review the manuscript entitled “Factors influencing osteoradionecrosis progression during hyperbaric oxygen therapy: A case study”.\nThe reviewer is an oral maxillofacial surgeon. I read with great interest your paper. I also use HBOT to treat ORN, and found this paper extremely useful.\nPlease consider the following points.\n- Since ORN is an infectious osteomyelitis, I suggest a more detailed description regarding antimicrobials should be done. Would it be possible to consider the type of antibiotics and the administration duration? As the authors know, HBOT is believed to enhance the effectiveness of antibiotics.\n- In relation to the above, could you please mention bacterial testing? The presence or absence of anaerobic bacteria seems important in discussing the effectiveness of HBOT.\nBest regards.\n\nIs the background of the cases’ 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 conclusion balanced and justified on the basis of the findings? Yes", "responses": [ { "c_id": "12955", "date": "02 Jan 2025", "name": "Chaima zitouni", "role": "Author Response", "response": "Response to Reviewer Report by Atsushi Shudo, Kishiwada Tokushukai Hospital, Kishiwada, Japan We sincerely thank Dr. Atsushi Shudo for the thoughtful review and valuable comments, and here are our detailed responses to the comments provided: Since ORN is an infectious osteomyelitis, I suggest a more detailed description regarding antimicrobials should be done. Would it be possible to consider the type of antibiotics and the administration duration? As the authors know, HBOT is believed to enhance the effectiveness of antibiotics. We have addressed this suggestion by providing a more detailed description of the antimicrobials used, including the types of antibiotics and their administration duration, while also discussing the potential role of HBOT in enhancing antibiotic effectiveness. In relation to the above, could you please mention bacterial testing? The presence or absence of anaerobic bacteria seems important in discussing the effectiveness of HBOT. We have included a discussion on bacterial testing, highlighting the importance of anaerobic bacteria in ORN, and emphasized the potential role of HBOT in enhancing the effectiveness of antibiotics against these bacterial profiles." } ] }, { "id": "332798", "date": "31 Oct 2024", "name": "Busra Yilmaz", "expertise": [ "Reviewer Expertise osteoradionecrosis", "radiation induced trismus", "head and neck cancers", "chemotherapy", "radiotherapy", "oral management", "oral oncology", "side effects" ], "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\nComments to Authors This study addresses the effects of hyperbaric oxygen therapy (HBOT) on osteoradionecrosis (ORN) and analyzes the factors influencing the progression of ORN. The study presents significant clinical insights on the subject; however, several critical gaps need to be addressed and supplemented with additional information:\n1. The authors should explain how the clinical and radiological diagnosis of ORN was determined and provide appropriate references. 2. The criteria for tooth extraction indications should be clarified, with sources cited. 3. Details on the pre-treatment dental care performed to minimize the risk of ORN should be provided. Specifically, it should be explained what procedures were carried out as part of dental care, including professional cleaning, fluoride treatments, necessary extractions, restorations, and any patient education to promote oral hygiene during and after radiation therapy. 4. The criteria used for diagnosing radiation-induced trismus must be explained in detail, along with relevant references. 5. A key parameter in ORN is the specific radiation dose received by the affected region. For instance, the Dmean Vx doses at extraction sites are not reported. 6. It would be beneficial to discuss the clinical and laboratory factors that can influence ORN development and treatment from all angles. Recently published reviews may provide useful insights. 7. In Table 1, the tumor histological types and locations are quite heterogeneous, meaning the radiation doses received by different parts of the jaw also vary, which could affect the outcomes. This limitation should be explicitly addressed. Long-term follow-up processes in ORN management could be emphasized. In particular, the effects of patient compliance during long-term treatments, such as HBOT, should be discussed. 8. The long-term effects of HBOT and any potential complications should also be addressed.\n\nIs the background of the cases’ 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 conclusion balanced and justified on the basis of the findings? Yes", "responses": [ { "c_id": "12956", "date": "02 Jan 2025", "name": "Chaima zitouni", "role": "Author Response", "response": "Response to Reviewer Report by Busra Yilmaz, Istinye University, Istanbul, Turkey We sincerely thank Dr. Yilmaz for the thoughtful review and valuable comments, and here are our detailed responses to the comments provided: The authors should explain how the clinical and radiological diagnosis of ORN was determined and provide appropriate references. The diagnosis of ORN in our study was based on both clinical assessment and radiological imaging, as follows: Clinical Diagnosis: The primary clinical symptoms included pain, mastication difficulties, trismus, dental mobility, and bone exposure. These are typical symptoms of ORN and were observed in the majority of patients. Additionally, the presence of cutaneous fistulas was noted in 39% of cases, which is a key indicator of more advanced ORN. Radiological Diagnosis: Radiological examinations, including panoramic radiographs, CT scans, and MRI, were used to assess the severity of ORN. CT scans were the primary imaging modality for evaluating ORN progression. In our study, ORN lesions were classified according to the LENT/SOMA grading system The criteria for tooth extraction indications should be clarified, with sources cited. We have clarified the criteria for tooth extraction indications in the discussion section. Details on the pre-treatment dental care performed to minimize the risk of ORN should be provided. Specifically, it should be explained what procedures were carried out as part of dental care, including professional cleaning, fluoride treatments, necessary extractions, restorations, and any patient education to promote oral hygiene during and after radiation therapy. We provided the necessary details about the pre-treatment dental care performed to minimize the risk of ORN. The criteria used for diagnosing radiation-induced trismus must be explained in detail, along with relevant references. The diagnosis of radiation-induced trismus was not part of the objectives of our study, and therefore, we did not include detailed criteria for its diagnosis. A key parameter in ORN is the specific radiation dose received by the affected region. For instance, the Dmean Vx doses at extraction sites are not reported. We did not measure the specific radiation doses, such as the Dmean Vx doses at the extraction sites, in our study. However, we acknowledge the importance of radiation dose as a key parameter in ORN and have included a discussion of its role and relevance in the progression of ORN in the manuscript. It would be beneficial to discuss the clinical and laboratory factors that can influence ORN development and treatment from all angles. Recently published reviews may provide useful insights. In our study, we have made an effort to enrich the discussion by including relevant clinical factors that could influence the development and treatment of ORN. 7. In Table 1, the tumor histological types and locations are quite heterogeneous, meaning the radiation doses received by different parts of the jaw also vary, which could affect the outcomes. This limitation should be explicitly addressed. Long-term follow-up processes in ORN management could be emphasized. In particular, the effects of patient compliance during long-term treatments, such as HBOT, should be discussed. We have addressed this limitation by acknowledging the variability in radiation doses due to the heterogeneity of tumor histology and locations in the revised discussion. We have included a discussion on the importance of long-term follow-up in ORN management, emphasizing that patient compliance during extended treatments, such as HBOT, plays a crucial role in achieving optimal outcomes. The long-term effects of HBOT and any potential complications should also be addressed. We have added information on the long-term effects of HBOT, including potential complications." } ] }, { "id": "332802", "date": "08 Nov 2024", "name": "Amaliya Amaliya", "expertise": [ "Reviewer Expertise Periodontology", "Tobacco Harm Reduction", "Nutrition and Periodontal 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\nDear Authors, The manuscript you submitted is well-written and well-read, but unfortunately, I have some comments that need to be addressed by you to improve the article. 1. First of all, I would like to suggest a minor revision to the title. It would be better if the title also mentioned the type of the study. The authors defined the study as retrospective, but I may suggest it as a case-control study. Either it may be like Factors influencing osteoradionecrosis progression during hyperbaric oxygen therapy: A retrospective study or Factors influencing osteoradionecrosis progression during hyperbaric oxygen therapy: A case-control study. 2. The introduction is too short, the authors only cite 2 articles in there. Please explain the current state of the knowledge, with references. Identify the knowledge gap that you wish to fill with your study. 3. The result section is lacking demographic data/table of the subjects. 4. Did the authors adjust the collected data, for example for differences in age and gender? 5. In the discussion, please explain the mechanism or at least biological plausibility for the factors associated or influencing the ORN progression, for example, hypertension, what is the explanation or biological plausibility.\n\nIs the background of the cases’ history and progression described in sufficient detail? Partly\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 conclusion balanced and justified on the basis of the findings? Yes", "responses": [ { "c_id": "12957", "date": "02 Jan 2025", "name": "Chaima zitouni", "role": "Author Response", "response": "Response to Reviewer Report by Amaliya Amaliya, Universitas Padjadjaran, Bandung, West Java, Indonesia We sincerely thank Dr. Amaliya for the thoughtful review and valuable comments, and here are our detailed responses to the comments provided: 1. First of all, I would like to suggest a minor revision to the title. It would be better if the title also mentioned the type of the study. The authors defined the study as retrospective, but I may suggest it as a case-control study. Either it may be like Factors influencing osteoradionecrosis progression during hyperbaric oxygen therapy: A retrospective study or Factors influencing osteoradionecrosis progression during hyperbaric oxygen therapy: A case-control study. Author Response: The initial title of the manuscript was \"Factors influencing osteoradionecrosis progression during hyperbaric oxygen therapy: A retrospective study\", but this was rejected by the journal editors. They proposed the revised title, and we followed their recommendation. 2. The introduction is too short, the authors only cite 2 articles in there. Please explain the current state of the knowledge, with references. Identify the knowledge gap that you wish to fill with your study. Author Response: We have expanded the introduction to include a detailed explanation of the current state of knowledge with additional references and have clearly identified the knowledge gap our study aims to address. 3. The result section is lacking demographic data/table of the subjects. Author Response: I have included a table to address the reviewer’s concern and enhance the clarity of the results section. 4. Did the authors adjust the collected data, for example for differences in age and gender? Author Response: We have adjusted the collected data for potential differences in age and gender in our analysis. Regression analysis was applied to account for these factors and ensure that the results are not confounded by demographic variables. 5. In the discussion, please explain the mechanism or at least biological plausibility for the factors associated or influencing the ORN progression, for example, hypertension, what is the explanation or biological plausibility. Author Response: We have addressed the concern regarding the biological plausibility of factors influencing the progression of ORN, including hypertension." } ] } ]	1	https://f1000research.com/articles/13-1225
https://f1000research.com/articles/13-1180/v1	09 Oct 24	{ "type": "Study Protocol", "title": "Computational Study Protocol: Leveraging Synthetic Data to Validate a Benchmark Study for Differential Abundance Tests for 16S Microbiome Sequencing Data", "authors": [ "Eva Kohnert", "Clemens Kreutz", "Clemens Kreutz" ], "abstract": "Background The utility of synthetic data in benchmark studies depends on its ability to closely mimic real-world conditions and to reproduce results obtained from experimental data. Here, we evaluate the performance of differential abundance tests for 16S metagenomic data. Building on the benchmark study by Nearing et al. (1), who assessed 14 differential abundance tests using 38 experimental datasets in a case-control design, we validate their findings by generating synthetic datasets that mimics the experimental data. We will employ statistical tests to rigorously assess the similarity between synthetic and experimental data and to validate the conclusions on the performance of these tests drawn by Nearing et al. (1). This protocol adheres to the SPIRIT guidelines and is, to our knowledge, the first of its kind in computational benchmark studies.\n\nMethods We replicate Nearing et al.’s (1) methodology, incorporating synthetic data simulated using two distinct tools, mirroring each of the 38 experimental datasets. Equivalence tests will be conducted on 43 data characteristics comparing synthetic and experimental data, complemented by principal component analysis for overall similarity assessment. The 14 differential abundance tests will be applied to both synthetic and experimental datasets, evaluating the consistency of significant feature identification and the number of significant features per tool. Correlation analysis and multiple regression will explore how differences between synthetic and experimental data characteristics may affect the results.\n\nConclusions Synthetic data enables the validation of findings through controlled experiments. We assess how well synthetic data replicates experimental data, validate previous findings and delineate the strengths and limitations of synthetic data in benchmark studies. Moreover, to our knowledge this is the first computational benchmark study to systematically incorporate synthetic data for validating differential abundance methods while strictly adhering to a pre-specified study protocol following SPIRIT guidelines, contributing significantly to transparency, reproducibility, and unbiased research.", "keywords": [ "16S", "microbiome", "differential abundance", "simulation", "synthetic data", "benchmarking" ], "content": "Protocol\n\nTable of Contents\n\nIntroduction\n\nBackground and rationale {6a}\n\nObjectives {7}\n\nTrial design {8}\n\nSummary Table\n\nMethods\n\nStudy population/participants\n\nStudy setting {9}\n\nEligibility criteria {10}\n\nWho will take informed consent? {26a}\n\nAdditional consent provisions for collection and use of participant data and biological specimens {26b}\n\nInterventions\n\nExplanation for the choice of comparators {6b}\n\nIntervention description {11a}\n\nCriteria for discontinuing or modifying allocated interventions {11b}\n\nStrategies to improve adherence to interventions {11c}\n\nRelevant concomitant care permitted or prohibited during the trial {11d}\n\nProvisions for post-trial care {30}\n\nOutcomes {12}\n\nParticipant timeline {13}\n\nSample size {14}\n\nRecruitment {15}\n\nAssignment of interventions: allocation\n\nSequence generation {16a}\n\nConcealment mechanism {16b}\n\nImplementation {16c}\n\nAssignment of interventions: Blinding\n\nWho will be blinded {17a}\n\nProcedure for unblinding if needed {17b}\n\nData collection and management\n\nPlans for assessment and collection of outcomes {18a}\n\nPlans to promote participant retention and complete follow-up {18b}\n\nData management {19}\n\nConfidentiality {27}\n\nPlans for collection, laboratory evaluation and storage of biological specimens for genetic or molecular analysis in this trial/future use {33}\n\nStatistical methods {20}\n\nData monitoring committee {21a}\n\nStatistical methods for primary and secondary outcomes {20a}\n\nInterim analyses {21b}\n\nMethods for additional analyses (e.g. subgroup analyses) {20b}\n\nMethods in analysis to handle protocol non-adherence and any statistical methods to handle missing data {20c}\n\nPlans to give access to the full protocol, participant level-data and statistical code {31c}\n\nTimeline\n\nOversight and monitoring\n\nComposition of the coordinating centre and trial steering committee {5d}\n\nComposition of the data monitoring committee, its role and reporting structure {21a}\n\nAdverse event reporting and harms {22}\n\nAncillary and post-trial care {30}\n\nFrequency and plans for auditing trial conduct {23}\n\nPlans for communicating important protocol amendments to relevant parties (e.g. trial participants, ethical committees) {25}\n\nDissemination policy {31a}\n\nDiscussion\n\nAbbreviations\n\nDeclarations\n\nAcknowledgements\n\nAuthors’ contributions {31b}\n\nAvailability of data and materials {29}\n\nEthics and consent {24}\n\nConsent for publication {32}\n\nAuthors’ information (optional)\n\nData availability\n\nReferences\n\nStudy status\n\nNote: To achieve a rigorous methodology, this protocol adheres to an established standard and checklist for Standard Protocol Items: Recommendations for Interventional Trials (SPIRIT).2 The numbers in curly brackets in this protocol refer to SPIRIT checklist item numbers. The order of the items has been modified to group similar items. Since using a standardized terminology for study designs is essential, we also formulate our protocol using standard terminology, i.e. terms such as study population, comparator, intervention, outcome, modification, inclusion and exclusion.\n\n\nIntroduction\n\nDifferential abundance (DA) analysis of metagenomic microbiome data has emerged as a pivotal tool in understanding the complex dynamics of microbial communities across various environments and host organisms.3–5 Microbiome studies are crucial for identifying specific microorganisms that differ significantly in abundance between different conditions, such as health and disease states, different environmental conditions, or before and after a treatment. The insights we gain from analyzing the differential abundance of microorganisms are critical to understanding the role that microbial communities play in environmental adaptations, disease development and health of the host.6 Refining statistical methods for the identification of changes in microbial abundance is essential for understanding how these communities influence disease progression and other interactions with the host, which then enables new strategies for therapeutic interventions and diagnostic analyses.7\n\nThe statistical interpretation of microbiome data is notably challenged by its inherent sparsity and compositional nature. Sparsity refers to the disproportionate proportion of zeros in metagenomic sequencing data and requires tailored statistical methods,8,9 e.g. to account for so-called structural zeros that originate from technical limitations rather than from real absence.10 Additionally, due to the compositional aspect of microbiome data regulation of highly abundant microbes can lead to biased quantification of low-abundant organisms.11 Such bias might be erroneously interpreted as apparent regulation that is mainly due to the compositional character of the data. Such characteristics of microbiome data have a notable impact on the performance of common statistical approaches for DA analysis, delimits their applicability for microbiome data and poses challenges about the optimal selection of DA tests.\n\nA number of benchmark studies have been conducted to evaluate the performance of DA tests in the analysis of microbiome data.12–15 However, the results show a very heterogeneous picture and clear guidelines or rules for the appropriate selection of DA tests have yet to be established. In order to assess and contextualize the findings of those studies, additional benchmarking efforts using a rigorous methodology,16,17 as well as further experimental and synthetic benchmark data sets are essential.\n\nSynthetic data is frequently utilized to evaluate the performance of computational methods because for such simulated data the ‘correct’ or ‘true’ answer is known and can be used to assess whether a specific method can recover this known truth.16 Moreover, characteristics of the data can be changed to explore the relationship between data characteristics such as effect size, variability or sample size and the performance of the considered methods. Several simulation tools have been introduced for generating synthetic microbiome data.18–23 They cover a broad range of functionality. For example, MB-GAN22 leverages generative adversarial networks to capture complex patterns and interactions present in the data, while metaSPARSim,18 sparseDOSSA219 or nuMetaSim24 employ different statistical models to generate microbiome data. Introducing a new simulation tool typically involves demonstrating its capacity to replicate key data characteristics. Nonetheless, an ongoing question persists regarding the feasibility of validating findings derived from experimental data when synthetic data, generated to embody the characteristics of the experimental data, is used in its place.\n\nHere we refer to the recent high-impact benchmark study of Nearing et al.1 in which the performance of a comprehensive set of 14 DA tests applied to 38 experimental 16S microbiome data sets was compared. This 16S microbiome sequencing data is used to study communities in various environments, here from human gut, soil, wastewater, freshwater, plastisphere, marine and built environments. The data sets are presented in a two group design for which DA tools are applied to identify variations in species abundances between the groups.\n\nIn this validation study we replicate the primary analysis conducted in the reference study by substituting the actual datasets with corresponding synthetic counterparts. The objective is to explore the validity of the main findings from the reference benchmark study when the analysis workflow is repeated with an independent implementation and with synthetic data, generated to recapitulate the characteristics of the original real data.\n\nAim 1: Synthetic data, simulated based on an experimental template, overall reflect main data characteristics.\n\nAim 2: Study results from Nearing et al. can be validated using synthetic data, simulated based on corresponding experimental data.\n\nAim 1: Exploratory comparative study\n\nAim 2: Confirmatory benchmark study\n\n\nMethods\n\nIn the context of our benchmark study, the study population is given by the experimental data sets from the reference study.1\n\nWhere possible, the study is conducted analogously to the benchmark study conducted by Nearing et al.,1 e.g. the same data and primary outcomes will be used.\n\nAll data sets as provided by Nearing et al.1 will be included in the study. We employ two published simulation tools, metaSPARSim18 and sparseDOSSA2,19 which have been developed for simulating microbial abundance profiles as they are generated by 16s sequencing.\n\nWe also apply the same DA tests as in1 and implementation in the R statistical programming language. In order to provide the most valuable results for the bioinformatics community, the latest versions of these implementations will be used.\n\nInclusion criteria\n\nWe will include the same experimental data sets and DA tests as in Ref. 1.\n\nExclusion criteria\n\nThere are no exclusion criteria for the data sets.\n\nn/a: Data is publicly available, there is no need to obtain consent.\n\nn/a: Data is publicly available, there is no need to obtain consent.\n\n\nInterventions\n\nFor aim 1, the comparator are 43 data characteristics calculated from the 38 experimental data sets. These characteristics are chosen such that they provide a comprehensive description of count matrices and enabling unbiased comparison between experimental and synthetic data sets. They cover for example information about the sparsity in a data set, mean-variance trends of features (taxa), or effect sizes between groups of samples. Tables 4 and 5 provide a detailed summary of all data characteristic and how they are calculated.\n\nFor aim 2, 14 differential abundance (DA) tests are applied to the experimental data (ALDEx2, ANCOM-II, corncob, DESeq2, edgeR, LEfSe, limma voom (TMM), limma voom (TMMwsp), MaAsLin2, MaSsLin2 (rare), metagenomeSeq, t-test (rare), Wilcoxon test (CLR), Wilcoxon test (rare)), i.e. the outcomes (number of significant features) calculated from the experimental data sets will serve as comparator. As in Ref. 1, we analyzed unfiltered data as well as data filtered with respect to features with a sufficient number of non-zero counts.\n\nThe intervention consists of using synthetic data instead of experimental data.\n\nFor each of the 38 experimental data sets, synthetic data will be simulated. For the simulation two simulation tools (metaSPARSim18 and sparseDOSSA219) will be used. Simulation parameters are calibrated using the experimental data, such that the simulated data reflect the experimental data template. Both simulation approaches offer such a calibration functionality. Multiple (N=10) data realizations will be generated for each experimental data template to assess the impact of different realizations of simulation noise and to test for significant differences between interventions and the comparator.\n\nFor aim 1, the data characteristics will be computed for each of the synthetic and experimental data sets. For aim 2, 14 DA tests will be applied to the synthetic data generated in aim1.\n\nFor assessing the similarity of the synthetic data templates, we apply equivalence tests based on two one-sided t-tests as implemented in the TOSTER R-package with a 5% significance level. We use the SD of the respective values from all experimental data templates as lower and upper margins. Figure 1 illustrates the equivalence testing procedure for the proportion of zeros in the whole data set as an exemplary data characteristic. For equivalence testing, the combined null hypothesis that the tested values are below the lower margin or above the upper margin has to be rejected to conclude equivalence. This only occurs when the average data characteristic of synthetic data is inside both margins and not too close to those two bounds, i.e. the whole 95% CI interval of the estimated mean has to be between both margins.\n\nThe black dots indicate a data characteristic computed for experimental data sets (here the proportion of zeros). Equivalence tests requires an interval that is considered as equivalent given by lower and upper margin bounds (dashed lines). We use the SD over all values from experimental templates to define these margins. The values computed from the synthetic data for a template are considered as equivalent if values below the lower margin and above the upper margin can be rejected according to the prespecified significance level. Depending on the variation of the characteristic for the synthetic data (here indicated by the boxplot), the average characteristic has to be inside a region (brown region) that is smaller than the interval between both margins.\n\nModification by adjusting the proportions of zeros and effect sizes\n\nIf equivalence tests fail, i.e. the synthetic data turns out to be partly unrealistic, we try to reduce the number of failed tests by adjusting two important characteristics, the proportion of zeros in the synthetic data sets, and the effect size, i.e., magnitude of differences between the two groups of samples.\n\nModifying the proportion of zeros will be performed by the following procedure for all synthetic data sets:\n\n1. If the number of rows or columns of the experimental data template does not coincide, randomly add or delete columns and rows in the template.\n\n2. Count the number of zeros that have to be added (or removed) for a simulated data set to obtain the same number as in the template.\n\n3. If the simulation method does not generate data with matching order of features (i.e. rows), sort all rows of both count matrices according to row means.\n\n4. Copy and replace an appropriate number of zeros (or non-zeros) one-by-one (i.e. with the same row and column indices) from the template to the synthetic data by randomly drawing those positions.\n\n5. Reorder the rows to get the original ordering.\n\n6. Check, whether the total number of failed equivalence test across all data templates is reduced.\n\nSince we calibrate the simulation tools for both groups separately, all simulation parameters controlling the count distribution will be different in both groups. Therefore, we anticipate that the differences between both groups might be overestimated. We therefore try to make the simulation more realistic by modifying the effect size by the following procedure for all synthetic data sets:\n\n1. Estimate the proportion of unregulated features from the results of all DA methods applied to the experimental data templates. This is done by the pi0est function in the qvalue R-package.\n\n2. Calibrate the simulation tool by using all samples from both groups (then there is no difference between both groups of samples) and generate a synthetic data set without considering the assignment of samples to groups.\n\n3. Replace an appropriate number of rows in the original synthetic data set by rows from the group-independent simulation.\n\nIn addition to both individual modifications, we also apply both modification. For the following analyses, we then use the synthetic data where most data characteristics are equivalent.\n\nExclusion criteria\n\nIn our study, we use experimental data as templates for generating synthetic data which are then analyzed by DA methods. At both levels, generation of synthetic data and applying DA methods, we define exclusion or modification criteria in order to handle exceeding runtimes, computation errors, and unrealistic data simulation. Figure 2 shows an overview about these exclusion and modification steps.\n\nThese criteria are applied to handle runtime issues, computation errors and unrealistic synthetic data.\n\nExclusion of simulation for a specific data template based on simulation performance\n\nA simulation tool will be excluded for a specific data template, if calibration of the simulation parameters is not feasible. We define feasibility by the following criteria:\n\n1) Calibration succeeds without error message\n\n2) The runtime of the calibration procedure is below 7 days (168 hours) for one data template\n\n3) The runtime of simulating a synthetic data set is below 1 hour for one synthetic data set\n\nAll computations in this study will be performed on a Linux Debian x86_64 compute server with 64 AMD EPYC 7452 32-Core Processor CPUs. Although, we will run parts of the analyses in parallel mode, the specified computation times refer to runtimes on a single core.\n\nExclusion of simulations for a specific data template based on deviating data properties\n\nFor aim 2, we exclude synthetic data sets that are not similar enough to the experimental data sets used as templates. The goal of the following exclusion criterion is to exclude synthetic data sets that are overall strongly dissimilar from the experimental data template, without being too stringent since the simulation tools cannot perfectly resemble all data characteristics and therefore a slight or medium amount of dissimilarity has to be accepted. In general, dissimilarities are exploited to study the impact of those characteristics by investigating the association of such deviations with dissimilarity in outcomes. For assessing similarity, the data characteristics described before and specified in detail in Table 5 are used.\n\nWe expect that a few data characteristics are very sensitive in discriminating experimental and synthetic data. To prevent loss of too many data sets, such characteristics (highlighted in gray in Figure 2) are only considered for the investigation of association between mismatch in outcome and mismatch in data characteristics but not for exclusion.\n\nUnrealistic synthetic data will be excluded for the primary analyses of the study using the remaining data characteristics. We define the exclusion criteria due to dissimilarity from its template by one of the following criteria:\n\n1) The equivalence test based on Euclidean distance in the 2-dimensional PCA plot failed to indicate equivalence with the respective data templates. For equivalence testing, we use +/- 1 SD of the Euclidean distance over all exp. data templates as upper and lower margins.\n\n2) We apply equivalence tests for all 43 data characteristics individually. We then only consider data characteristic which are not highly-discriminative. When counting non-equivalence of the remaining characteristics (highlighted in brown color in Figure 2) for a template, the synthetic data of those templates that appear as an outlier will be removed (see example in Figure 2). We use the common outlier definition from boxplots, i.e. all values with distance to the 1st quartile (Q1) or 3rd quartile (Q3) larger than 1.5 x the inter-quartile range Q3-Q1 are considered as outlier.\n\nFor evaluating the sensitivity with respect to exclusion, we perform an additional, secondary analysis on all synthetic data sets, regardless of similarity to the templates.\n\nModification of differential abundance (DA) tests\n\nInflated runtime\n\nData sets with a large number of features could lead to inflating runtimes for some statistical tests. If the runtime threshold for an individual test is exceeded for a specific data set, we split the data set, apply the test again on the subsets and afterwards merge the results. This split and merge procedure is repeated until the test runtime is below the threshold.\n\nHere, we define the runtime threshold to be max. 1 hour per test. Then, in a worst case scenario, the 14 tests for the 10+1 data sets for each of the 38 template (5832 combinations) would need 244 days on a single core. Since we can conduct the tests on up to 64 cores, such a worst case scenario would still be manageable.\n\nTest failure\n\nIf a DA test throws an error we omit the number of significant features and the overlap of significant features and report them as NA (not available) as it would occur in practice.\n\nn/a\n\nn/a\n\nn/a\n\n\nOutcomes {12}\n\nAim 1: For each data set (experimental template and synthetic data) 43 data characteristics are calculated as described in Table 5. The difference of a data characteristic between a synthetic data and the corresponding data template is calculated as outcomes. For each feature which is closer to a normal distribution on the log-scale according to p-values of the Shapiro-Wilk test, we apply a log2-transformation to the respective characteristics prior to all analyses.\n\nPrincipal component analysis (PCA) is then performed on the scaled data characteristics and a two-dimensional PCA plot is generated. An additional outcome is the Euclidean distance of a synthetic data set to its template in the first two principal component coordinates. An equivalence test will be conducted on the synthetic data sets for each template to check whether data properties are maintained in synthetic data on a summary level for all data characteristics.\n\nNext, boxplots are generated, visualizing for each data characteristic how it varies between templates, between all simulation realizations, and how templates deviate from the corresponding synthetic data sets. Here, we again perform equivalence tests and also report median distances of a data characteristic between simulated and experimental data.\n\nAim 2: As primary outcome (aim 2a), for each experimental data template the average proportion of shared significant features across all synthetic data are calculated for each DA tool. For each data template, a barplot is generated as in Nearing et al.1 to visually summarize how many of the 14 DA tools identified the same feature as significantly changed. Moreover, we try to validate the conclusions from Nearing et al.1 made on this primary outcome. Overall, we extracted 13 conclusions and formulated the respective hypothesis as shown in Box 1.\n\nWe term the statistical analysis that estimates the proportion P of cases (e.g. the proportion of synthetic data sets) where the hypothesis is fulfilled as “Counting”. Depending on the stringency of the formulation in Ref. 1, we always define a question-specific threshold for confirmation and indicate the respective number of cases n for this evaluation in brackets. The asterisk * indicates that this number of cases might be reduced if exclusion criteria apply.\n\nHypothesis 1: For unfiltered data, the proportion of features jointly found as significant by limma voom TMM and limma voom TMMwsp but by less than 50% of the other methods, is larger than the overlap with more than 50% of the other methods.\n\nAnalysis: Counting (n=380) with 95% threshold, i.e. the hypothesis is validated if the 95% CI is > 95%.\n\nHypothesis 2: For unfiltered data, the overlap of features jointly found as significant by limma voom TMM and limma voom TMMwsp with features found by Wilcoxon CLR is larger than the overlap with all other DA methods.\n\nAnalysis: Counting (n=380) with 95% threshold.\n\nHypothesis 3: For unfiltered data, the Kolmogorov-Smirnov test statistic D when comparing the profile for Wilcoxon CLR and Wilcoxon is larger than for other pairs of methods on average.\n\nAnalysis: Counting (n=380) with 95% threshold.\n\nHypothesis 4: For unfiltered data, MaAsLin2 and MaAsLin2-rare a more similar profile (larger test statistic D) than a randomly selected pair of methods.\n\nAnalysis: Counting (n=380) with 95% threshold.\n\nHypothesis 5: For unfiltered data, ALDEx2 and ANCOM-BC identify more features that were also identified by all except 3 (i.e. 10 out of 13) other methods.\n\nAnalysis: Counting (n=380) with 95% threshold.\n\nHypothesis 6: For unfiltered data, EdgeR and LEfSe identify a larger percentage of features that are not identified by any other tool, compared to the same percentage for all other methods.\n\nAnalysis: Counting (n=380) with 95% threshold.\n\nHypothesis 7: For unfiltered data, for corncob, metagenomeSeq, and DESeq2, there are always multiple other methods (i.e. at least 2 out of 10 other DA methods) that have a more extreme consistency profile.\n\nAnalysis: Counting (n=380) with 95% threshold. Mean consistency is used to assess the location of the consistency profile.\n\nHypothesis 8: The shape of the overlap profiles is mainly determined by the exp. data set and the DA method but only little of the fact whether data has been filtered.\n\nAnalysis: qq-Plots of the cumulative overlap profile filtered vs. non-filtered are closer to the diagonal than comparison of different DA methods and comparison of different exp. data templates. Quantification using the mean of the Kolmogorov-Smirnov test statistic, i.e. the average of the maximal absolute distance of the empirical cumulative density functions.\n\nHypothesis 9: For filtered data, for both limma voom approaches the proportion of identified features that are also identified by the majority of other tests is larger than for un-filtered data\n\nAnalysis: Counting (n=380) with 95% threshold.\n\nHypothesis 10: For filtered data, the overlap profile of Wilcoxon CLR is bimodal.\n\nAnalysis: Counting (n=380) with 95% threshold using bimodality index (R function BimodalIndex::bimodalIndex). Only datasets with at least 10 significant features when using Wilcoxon CLR will be considered.\n\nHypothesis 11: The proportion of features identified by all except one DA method is larger for prevalence-filtered data.\n\nAnalysis: Counting (n=380) with 95% threshold.\n\nHypothesis 12: For filtered data, the consistency profiles of corncob, metagenomeSeq, and DESeq2 are more similar to the more extreme methods (as for Hypothesis 9 defined as the most extreme 2 profiles of other DA methods) than for unfiltered data.\n\nAnalysis: Counting (n=380) with 95% threshold using the Kolmogorov-Smirnov test statistic of the two 2nd most extreme left- and right shifted other profiles.\n\nHypothesis 13: For filtered data, ALDEx2 and ANCOM-BC identify more features that were also identified by all except 3 (i.e. 10 out of 13) other methods.\n\nAnalysis: Counting (n=380) with 95% threshold.\n\nAs secondary outcome (aim 2b), the numbers and proportions of significant features across tools and data sets are considered. This outcome is reported and visualized in a heatmap with rows representing the data templates and columns the DA tools as in Nearing et al.1 In total, two heatmaps, one for the experimental data and a second one for the simulated data, are generated. For the synthetic data sets, mean values from N=10 simulation realizations are calculated and plotted.\n\nMoreover, we try to validate the outcomes from Nearing et al.1 made on this secondary outcome. Overall, we extracted 14 conclusions about the number of identified features and formulated the respective hypothesis as summarized in Box 2.\n\nWe term the statistical analysis that estimates the proportion P of cases (e.g. the proportion of synthetic data sets) where the hypothesis is fulfilled as “Counting”. Depending on the stringency of the formulation in Ref. 1, we always define a question-specific threshold for confirmation and indicate the respective number of cases n for this evaluation in brackets. The asterisk * indicates that this number of cases might be reduced if exclusion criteria apply.\n\nHypothesis1: For filtered and unfiltered data, the percentage of significant features identified by each DA method varies widely across data sets.\n\nAnalysis: Histograms of the range (= max – min) of the percentage values 1) for all data sets, and 2) for all DA methods when applied to data from each data template\n\nHypothesis 2: For filtered and unfiltered data, rankings of the DA methods with respect to the proportion of identified features depend on the data template.\n\nAnalysis: 2-way ANOVA of the rank-transformed proportions using DA method and template as grouping variables indicate significant interaction effects between both variables.\n\nHypothesis 3: Rankings of the DA methods with respect to the proportion of identified features depend stronger on the data template in unfiltered data than in filtered data sets.\n\nAnalysis: 2-way ANOVA of the ranks using DA method and template as grouping variables indicate a more significant interaction effects between both effects in unfiltered data (compared to respective analysis for unfiltered data, see preceding hypothesis).\n\nHypothesis 4: In unfiltered data, either limma voom TMMwsp, limma voom TMM, Wilcoxon, or LEfSe identify the largest proportion of significant features.\n\nAnalysis: Counting (n=380). Since in the original analysis, the observation was seen in all analysis, we classify the hypothesis as confirmed, if true in >95% of cases.\n\nHypothesis 5: For unfiltered data, there are data sets, where edgeR identifies the largest proportion of significant features.\n\nAnalysis: Counting (n=380) with >0 threshold.\n\nHypothesis 6: For unfiltered data, Limma voom TMMwsp identifies the largest proportion of features in the Human-HIV data set.\n\nAnalysis: Counting (n=380) in all synthetic data sets generated for this template with >95% threshold.\n\nHypothesis 7: For unfiltered data, there are data sets, where both limma voom methods identify more than 99% of features as significant\n\nAnalysis: Counting (n=380) with >0 threshold.\n\nHypothesis 8: For unfiltered data, there are data sets, where Wilcoxon CLR identifies more than 90% of features as significant\n\nAnalysis: Counting (n=380) with >0 threshold.\n\nHypothesis 9: For unfiltered data, there are data sets, where LEfSe identifies more than 99% of features as significant\n\nAnalysis: Counting (n=380) with >0 threshold.\n\nHypothesis 10: In unfiltered data, either ALDEx2 or ANCOM-BC identify the fewest significant features.\n\nAnalysis: Counting (n=380) with >95% threshold.\n\nHypothesis 11: In unfiltered data, ALDEx2, ANCOM-BC and corncob do not identify significantly more features than the most conservative tests.\n\nAnalysis: Counting (n=380) whether DA method is within the three most conservative ones (with >95% threshold).\n\nHypothesis 12: No tool (except ALDEx2) identifies a smaller number of features for unfiltered data (compared to filtered data).\n\nAnalysis: Counting with >95% threshold.\n\nHypothesis 13: For filtered and unfiltered data, ANCOM-BC identifies the least significant features in total, i.e. when summing ranks of DA methods over all 38 templates.\n\nAnalysis: Counting (n=10) whether the ranksum over 38* single synthetic data simulations (randomly drawn from N=10) of all other DA methods satisfies the hypothesis (>95% threshold without confidence intervals).\n\nHypothesis 14: For filtered data, there is no method other than EdgeR, LEfSe, limma voom, or Wilcoxon that identifies the largest number of significant features in total, i.e. when considering ranks of DA methods over all 38 templates.\n\nAnalysis: Counting (n=10) whether the ranksum over 38* single synthetic data simulations (randomly drawn from N=10) of all other DA methods satisfies the hypothesis (<5% threshold without confidence intervals).\n\nn/a\n\nThe used 38 experimental data sets and DA tests are defined by the reference study that is being validated. In our study, we can therefore only choose the number of synthetic data sets per data template reasonably. Since there is no pilot study for the outcomes obtained from synthetic data that can be used for sample size calculations, and because a large number of outcomes are considered, the number of simulated data sets for one template was chosen to be a) feasible in terms of run time and b) should be large enough to enable valid conclusions. Based on both aspects, we decided to simulate N=10 synthetic data sets for each experimental data template, i.e., 380 synthetic data sets for each simulation tool.\n\nFor the one-sample equivalence tests with significance level 5% conducted for aim 1, N=10 synthetic data sets have a power of 89,75% to reject both the null hypothesis that the data characteristic is below -1SD and above +1SD and thereby favor the alternative hypothesis that the characteristic is equivalent to the reference value computed for the experimental data template. For this computation, we presumed that the expected mean (i.e. the bias of a characteristic in simulated data) is 0.5 SD, the standard deviation within synthetic data is 0.5 SD. Here, all specifications were made in units SD that refers to the standard deviation over all exp. templates. Assuming a smaller bias or a smaller variability for the synthetic data increases the expected power. These calculation has been conducted with Nquery version 8.7.2.0, equivalence test for one mean (MOE4-1).\n\nFor the proportions P of fulfilled hypothesis that were estimated for addressing most hypotheses in aim 2, 95% Clopper-Pearson confidence intervals (R-package DescTools) are calculated. As an example, n=380 independent samples leads to P = 0.026, 95%-CI = [0.013, 0.047] if for 10 out of 380 synthetic data sets the tested hypothesis is violated. This sample size is available for 20/27 hypotheses. However, it should be noted that such sample size considerations are limited by the fact that the data characteristics and results for synthetic data sets from a template are likely to be similar to each other and it is therefore not permissible to consider all samples as independent.\n\nn/a\n\nThe intervention and the comparator can be evaluated for all data sets and the order of the computations has no impact. Therefore, random allocation and sequence generation is not required.\n\nn/a\n\nn/a\n\nNo blinding procedure will be applied because we see no risk of bias by a non-blinded calculation of data characteristics and statistical tests.\n\nn/a: No blinding performed.\n\n\nData collection and management\n\nThe analyses will be conducted by two experienced Statisticians/Bioinformaticians (E.K., C.K.) both with > 5yrs experience with differential abundance analyses. All statistical tests will adhere to the methodology outlined by Nearing et al.1 To do so we chose the same configurations of the statistical methods as implemented in the code from Nearing et al.1 for running the statistical tests, provided on Github (https://github.com/nearinj/Comparison_of_DA_microbiome_methods). This strategy ensures comparability between the outcomes of the reference study and our validation study.\n\nn/a\n\nThe 38 experimental data sets were downloaded from https://figshare.com/articles/dataset/16S_rRNA_Microbiome_Datasets/14531724 on February 9, 2024. There, Nearing et al.1 made the data sets from their study available, therefore we incorporate the exact same data sets. We keep a local copy of this data in our Fredato research data management system https://nxc-fredato.imbi.uni-freiburg.de until 31.12.2030 and make it available if the original data is not available in the current version any more and if this does not violate legal, data protection, or copyright regulations. Generated data, analysis scripts, results and supplemental information to this study will also be stored in the Fredato research data management. In case of unexpected technical limitations, we will make data, analysis scripts and supplemental information available via https://figshare.com.\n\nn/a: The experimental data is publicly available. The data generated in this project is not subject to data protection regulations.\n\nn/a\n\n\nStatistical methods {20}\n\nn/a: The data has already been collected and is publicly available.\n\nAim 1: For each data set (experimental and synthetic) a set of 43 data characteristics is calculated. All data characteristics are defined as a single number. These calculations are described in more detail in Table 5. As N=10 simulation realizations are generated, there will be 10 values for each data characteristic per experimental data template. For the primary outcome a PCA plot based on the scaled data characteristics is generated. Moreover, equivalence tests are applied to all 43 data characteristics as well as to the Euclidean distance in the two principal component coordinates as describe above (section “Interventions”). Based on these equivalence tests, we test a single hypothesis (equivalence of synthetic and experimental data) which is fulfilled in strict terms only when all equivalence tests are significant. We therefore do not have to control the probability of a single false positive test (i.e. the so-called family-wise error rate). Therefore, multiple testing aspects do not apply these tests.\n\nAim 2: All DA methods will be applied to experimental and synthetic data adhering to the methodology in Nearing et al.1 as described in the methods section of the paper. Significant features will be identified using a 0.05 threshold for the multiple testing adjusted p-values (Benjamini-Hochberg).\n\nFor the primary outcome, it is determined how many tests jointly identify features to be significant for each data set. After visualization, the 13 hypotheses extracted from1 will be investigated using the statistical analyses summarized in Box 1. Estimates of the target values and the respective 95% confidence intervals are used to validate the hypotheses because these values are easier to interpret than p-values and because the significance of the p-values is strongly determined by the number of cases.\n\nFor the secondary outcome, the number and proportion of significant features is extracted for each data set and test individually. After visualization, the 14 hypotheses extracted from1 will be investigated using the statistical analyses summarized in Box 2.\n\nConfidence intervals for the estimated proportions of cases where the hypothesis is fulfilled will be calculated as exact intervals, i.e. using Clopper-Pearson intervals (R-package DescTools). If the analyzed cases are statistically dependent, we compute hierarchical bootstrap confidence intervals by first drawing with replacement templates, and then synthetic data sets within each template.\n\nThese analyses are conducted for unfiltered data and for filtered data. As in,1 filtered means that features found in fewer than 10% of samples are removed. Moreover, to analyze the sensitivity of our outcomes with respect to our exclusion criteria, filtered and unfiltered data are also analyzed without applying criteria that exclude non-realistic simulations.\n\nIn case we find different results for the simulated data for some hypotheses, we will analyze the association of the mismatch in the outcome with the mismatch of data characteristics to identify data characteristics that could be responsible for the disagreement. To ensure independence of the scales, we will perform these analyses after rank transformations. We will use univariate analyses (i.e. Spearman correlation) as well as a forward selection with a 5% cut-off criterion for p-values.\n\nn/a: There will be no interim analyses in this study.\n\nn/a: In this study there will be no subgroup analyses.\n\nn/a: In this study there is no missing data.\n\nGenerated data, analysis scripts and supplemental information to this study will also be stored in the Fredato research data management. In case of unexpected technical limitations, we will make data, analysis scripts and supplemental information available via https://figshare.com.\n\n\nTimeline\n\nThere is no timeline for data collection, as the study is based on existing data. Conducting the study does not depend on any clinical parameters. The anticipated timeline for completing the study is 3 to 4 months.\n\n\nOversight and monitoring\n\nn/a\n\nn/a\n\nn/a\n\nn/a\n\nn/a\n\nn/a\n\n\nDissemination policy {31a}\n\nPublic access of the generated data, analysis scripts, results and supplemental information is granted as indicated above on our Fredato research data management system https://nxc-fredato.imbi.uni-freiburg.de. The results will be published in a peer-reviewed scientific journal, preferably in the same journal as this protocol.\n\n\nDiscussion\n\nSynthetic data are a valuable tool in benchmark studies as it allows for controlled manipulation of data properties and evaluations based on the underlying known truths. We utilize this potential to validate previous findings derived on experimental data exclusively. However, first it is critical to determine the extent to which synthetic data can faithfully reflect real experimental data. It may necessitate adjustments to simulation tools to ensure their capability to accurately mimic experimental conditions. Consequently, this study not only investigates the potential limitations of synthetic data in validating experimental outcomes but also sheds light on the general capacity of simulation tools to faithfully represent real-world data. Since unsuccessful validation indicates that the previously published findings do not generalize to synthetic data, we try to identify responsible inadequacies in the simulation tools. However, this aspect is limited to the data characteristics of our study and there might be additional data properties that require consideration to explain a possible mismatch. Finally, this benchmark study employs rigorous statistical methodology and, for the first time, publishes a study protocol in advance, adhering to established protocol guidelines. Moreover, it is the first benchmark study that primarily focus on validation of the results of a previous study.\n\n\nAbbreviations\n\nANCOM\n\nAnalysis of compositions of microbiomes with bias correction, a DA method\n\nANOVA\n\nanalysis of variance\n\nDA\n\ndifferential abundance\n\nFredato\n\nFreiburg research data management tool\n\nLEfSe\n\nLinear discriminant analysis Effect Size, a DA method\n\nMaAsLin\n\nMicrobiome Multivariable Association with Linear Models, a DA method\n\nmetaSPARSim\n\nacronym for “a sparse count matrix simulator intended for usage in development of 16S rDNA-seq metagenomic data processing pipelines”18\n\npKS\n\np-values of a Kolmogorov-Smirnov test\n\nPCA\n\nprincipal component analysis\n\nR\n\na script-based statistical programming language\n\nsparseDOSSA2\n\nabbreviation for “Sparse Data Observations for Simulating Synthetic Abundance”, a simulation tool for microbial count data based on a hierarchical model19\n\nTMM\n\ntrimmed mean of M values, an approach for data normalization\n\nTMMwsp\n\nTMM with singleton pairing, an approach for data normalization\n\n\nDeclarations\n\n\n\n\nAuthors’ contributions {31b}\n\nE.K. and C.K. equally contributed to the design and development of the study.\n\n\nEthics and consent {24}\n\nEthical approval and consent were not required\n\n\nConsent for publication {32}\n\nn/a\n\n\nAuthors’ information (optional)\n\nE.K.: Statistical data analyst and PhD student in the lab of Dr. Clemens Kreutz. Research topics cover robust data analysis of high throughput data with a specification for microbiome sequencing data.\n\nC.K.: Group Leader of the lab for Methods in Systems Biomedicine at the Institute for Medical Biometry and Statistics of University Medical Center of Freiburg. Research topics cover mathematic modelling for systems biomedicine and neutral benchmark studies.\n\nBoth authors have neither been involved in the development of any of the applied simulation and DA methods nor in the reference study.1\n\n\nStudy status\n\n", "appendix": "Data availability {29}\n\nNo data are associated with this article.\n\n\nAcknowledgements\n\nn/a\n\n\nReferences\n\nNearing JT, Douglas GM, Hayes MG, et al.: Microbiome differential abundance methods produce different results across 38 datasets. Nat. Commun. 2022; 13(1). Publisher Full Text\n\nChan AW, Tetzlaff JM, Gøtzsche PC, et al.: SPIRIT 2013 explanation and elaboration: guidance for protocols of clinical trials. BMJ. 2013; 346: 346. Publisher Full Text\n\nSorboni SG, Moghaddam HS, Jafarzadeh-Esfehani R, et al.: A Comprehensive Review on the Role of the Gut Microbiome in Human Neurological Disorders. Clin. Microbiol. Rev. 2022; 35(1): e0033820. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHou K, Wu ZX, Chen XY, et al.: Microbiota in health and diseases. Signal Transduct. Target. Ther. 2022; 7(1).\n\nYoung VB: The role of the microbiome in human health and disease: An introduction for clinicians. BMJ. 2017; 356: j831. Publisher Full Text\n\nLi Q, Chan H, Liu WX, et al.: Carnobacterium maltaromaticum boosts intestinal vitamin D production to suppress colorectal cancer in female mice. Cancer Cell. 2023; 41(8): 1450–1465.e8. PubMed Abstract \| Publisher Full Text\n\nTang J, Wei Y, Pi C, et al.: The therapeutic value of bifidobacteria in cardiovascular disease. NPJ Biofilms Microbiomes. 2023; 9(1): 82. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJonsson V, Österlund T, Nerman O, et al.: Modelling of zero-inflation improves inference of metagenomic gene count data. Stat. Methods Med. Res. 2019; 28(12): 3712–3728. PubMed Abstract \| Publisher Full Text\n\nLuz Calle M: Statistical analysis of metagenomics data. Genomics Inform. 2019; 17(1). Publisher Full Text\n\nSilverman JD, Roche K, Mukherjee S, et al.: Naught all zeros in sequence count data are the same. Comput. Struct. Biotechnol. J. 2020; 18: 2789–2798. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGloor GB, Macklaim JM, Pawlowsky-Glahn V, et al.: Microbiome datasets are compositional: And this is not optional. Front. Microbiol. 2017; 8: 8. Publisher Full Text\n\nYang L, Chen J: A comprehensive evaluation of microbial differential abundance analysis methods: current status and potential solutions. Microbiome. 2022; 10(1): 130. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCappellato M, Baruzzo G, Di CB: Investigating differential abundance methods in microbiome data: A benchmark study. PLoS Comput. Biol. 2022; 18(9): e1010467. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWeiss S, Xu ZZ, Peddada S, et al.: Normalization and microbial differential abundance strategies depend upon data characteristics. Microbiome. 2017; 5(1): 27. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCalgaro M, Romualdi C, Waldron L, et al.: Assessment of statistical methods from single cell, bulk RNA-seq, and metagenomics applied to microbiome data. Genome Biol. 2020; 21(1): 191. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBoulesteix AL, Morris T, Sauerbrei W, et al.: STRengthening Analytical Thinking for Observational Studies (STRATOS): Introducing the Simulation Panel (SP). Biom. Bull. 2020; 37(2): 11–12.\n\nBoulesteix AL, Binder H, Abrahamowicz M, et al.: On the necessity and design of studies comparing statistical methods. Biom. J. 2018; 60(1): 216–218. Publisher Full Text\n\nPatuzzi I, Baruzzo G, Losasso C, et al.: MetaSPARSim: A 16S rRNA gene sequencing count data simulator. BMC Bioinformatics. 2019; 20: 416. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMa S, Ren B, Mallick H, et al.: A statistical model for describing and simulating microbial community profiles. PLoS Comput. Biol. 2021; 17(9): e1008913. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRichter DC, Ott F, Auch AF, et al.: MetaSim - A sequencing simulator for genomics and metagenomics. PLoS One. 2008; 3(10): e3373. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFritz A, Hofmann P, Majda S, et al.: CAMISIM: Simulating metagenomes and microbial communities. Microbiome. 2019; 7(1): 17. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRong R, Jiang S, Xu L, et al.: MB-GAN: Microbiome Simulation via Generative Adversarial Network. Gigascience. 2021; 10(2). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWilliams J, Bravo HC, Tom J, et al.: MicrobiomeDASim: Simulating longitudinal differential abundance for microbiome data. F1000Res. 2020; 8: 8. Publisher Full Text\n\nLiu S, Hua K, Chen S, et al.: Comprehensive simulation of metagenomic sequencing data with non-uniform sampling distribution. Quantitative Biology. 2018; 6(2): 175–185. Publisher Full Text" }	[ { "id": "340429", "date": "27 Nov 2024", "name": "Leo Lahti", "expertise": [ "Reviewer Expertise bioinformatics", "microbiome", "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\nThis manuscript suggests a (computational) protocol for validating differential abundance tests. The main question is whether synthetic data, generated to mimic experimental data, can validate the findings from differential abundance tests for 16S microbiome sequencing data. Specifically, the study aims to assess how well synthetic datasets replicate the characteristics of real data and whether the conclusions from real data on the performance of DA tests can be confirmed with synthetic data. In order to evaluate such protocol, the authors created synthetic data that would resemble the 16S rRNA gene sequencing data in the 38 datasets employed in the study by Nearing et al. (2022). They mimic real data with synthetic data using metaSPARSim and sparseDOSSA2 with specific adjustments. They use 43 metrics to evaluate the correspondence between the synthetic and real data. The study complies with the SPIRIT guidelines for clinical trial protocols. Second, the datasets that resemble the original data sufficiently well were used to replicate results of differential abundance analysis (DAA) method comparison by Nearing et al. The language is clear but the protocol has a vast number of steps, making it hard to separate the key contributions and performance assessment from the routine protocol checkpoints.\n\nMajor remarks:\n1) In addition to using metaSPARSim and sparseDOSSA2, consider including more recent methods such as SimMSeq (Yang and Chen, 2022) and SIMBA (Wirbel et al., 2024).\n2) The authors use 43 metrics to assess the correspondence between synthetic and real data. The number of metrics seems quite high, and it may thus be difficult describe these resemblances. Would it be possible to identify a smaller number of key metrics that would grasp the most essential aspects of the data?\n3) The motivation of Aim 2 seems unclear. The authors state in the introduction, synthetic data can be used to evaluate the correctness of DAA results. However, the authors suggest to use synthetic data to replicate the results Nearing et al. (Aim 2). Why synthetic data is not used to directly evaluate the correctness of the DAA results? Replication of Nearing’s results to evaluate the validity of the synthetic data seems a rather indirect means to assess the performance of the DAA methods.\n4) The study is about 16S (according to the title and descriptions) but Table 2, Aim 1 discusses metagenome simulation tools. The 16S and metagenome are different approaches for characterizing microbial communities, although there are some overlaps. It would require further clarification why a main Table switches to metagenomics in a 16S paper.\n5) Availability of materials: the study is based on an open data set that is available. Availability of the source code for the experiments and results would be critical for this kind of study. I could not locate these from the manuscript. Instead, there is a text \"Generated data, analysis scripts, results and supplemental information to this study will also be stored in the Fredato research data management. In case of unexpected technical limitations, we will make data, analysis scripts and supplemental information available via https://figshare.com\". These materials should be made available already at the review stage, otherwise a full review is not possible.\n\nMinor notes:\n- Real data sets are inherently different from synthetic ones. If the underlying data distributions differ significantly between synthetic and experimental datasets, the statistical tests may yield misleading results. Some discussion could be added on this and other potential limitations of using synthetic data for methods validation.\n\n- Possibly out of scope for the protocol study but in case all the compared data generation methods fail to produce realistic data can the authors consider suggesting their own solution to generate realistic 16S data or expand discussion on this?\n\n- Could you consider using other similar measures to PCA, for instance, principal coordinate analysis based on different dissimilarity metrics (e.g. Jaccard and Bray-Curtis distances) to evaluate the robustness of the ordination comparison?\n- Table 2: \" stasticial analyses\" -> \"statistical analyses\"?\n\nIs the rationale for, and objectives of, the study clearly described? Partly\n\nIs the study design appropriate for the research question? Partly\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? Yes", "responses": [ { "c_id": "12982", "date": "02 Jan 2025", "name": "Eva Kohnert", "role": "Author Response", "response": "General remark: The language is clear but the protocol has a vast number of steps, making it hard to separate the key contributions and performance assessment from the routine protocol checkpoints. We appreciate the reviewer’s feedback regarding the structure and clarity of the protocol. Since this is our first attempt at drafting a study protocol for a computational benchmark study, we adhered strictly to the SPIRIT guidelines. This allowed us to explore the applicability of these guidelines, originally designed for clinical research, and how they could be effectively adapted to computational studies in the future. While we acknowledge that this approach may have resulted in a protocol with numerous steps, we believe this provides a comprehensive foundation for reproducibility and transparency in computational benchmarking. For future benchmark studies, we plan to refine the protocol structure, focusing on clearly delineating key contributions and performance assessments from routine protocol checkpoints. Major remarks: 1) In addition to using metaSPARSim and sparseDOSSA2, consider including more recent methods such as SimMSeq (Yang and Chen, 2022) and SIMBA (Wirbel et al., 2024). We appreciate the reviewer’s suggestion to consider additional simulation tools such as SimMSeq and SIMBA. While we recognize the potential value of these tools, there are several reasons why we chose not to include additional simulation methods in this study: First, the current study is already highly complex, incorporating two simulation tools, comprehensive characterization of various data sets, multiple inclusion/exclusion criteria and extensive testing. Adding further tools would further increase the study’s complexity, potentially detracting from the clarity of our findings. Secondly, as stated in the protocol the computational effects are huge. Therefore, we defined a runtime threshold of max. 1 hour per differential abundance test. We are calculating 14 differential abundance tests across 10 + 1 (simulation + experimental template) datasets for each of the 38 templates and for unfiltered and filtered data, leading to a total of 11,704 combinations. In a worst case scenario, the runtime on a single core would already require approximately 488 days per simulation tool only for conducting DA analyses. Adding additional simulation tools would further inflate runtime, particularly given the additional time required to calibrate the simulators and generate synthetic datasets. Lastly, one of the key aims of this study is to introduce and emphasize the importance of formulating a study protocol specifically for computational research. This is the first attempt to define a structured protocol for such studies, and while it may not yet represent the most refined version, we think it is an important step towards robust and transparent computational benchmark studies. Developing this protocol required substantial effort, and we believe that establishing and demonstrating the concept of a protocol in computational studies is as important as the study results themselves. In the new version of the protocol, we briefly mention these aspects. 2) The authors use 43 metrics to assess the correspondence between synthetic and real data. The number of metrics seems quite high, and it may thus be difficult describe these resemblances. Would it be possible to identify a smaller number of key metrics that would grasp the most essential aspects of the data? We agree with the reviewer that a smaller set of metrics providing independent information would be beneficial. However, we aim to retain characteristics that might highlight discrepancies between simulated and experimental data and help explain potential disagreements in DAA outcomes. Consequently, we decided to eliminate metrics that are nearly redundant by iteratively removing those with a rank correlation ≥ 0.95 and adjusted our study protocol accordingly. 3) The motivation of Aim 2 seems unclear. The authors state in the introduction, synthetic data can be used to evaluate the correctness of DAA results. However, the authors suggest to use synthetic data to replicate the results Nearing et al. (Aim 2). Why synthetic data is not used to directly evaluate the correctness of the DAA results? Replication of Nearing’s results to evaluate the validity of the synthetic data seems a rather indirect means to assess the performance of the DAA methods. Given this feedback, we realized that the motivation of Aim 2 has not been communicated clearly. This study originated as part of the planning process for a more classical benchmark study using the underlying truth of synthetic data for assessing DA methods. While designing the benchmark study, we recognized the need to first assess the feasibility of generating synthetic data that realistically resemble all characteristics of experimental data, to ensure the validity of conclusions drawn from simulated data. This led us to develop the presented validation study with primary goal to compare the results based on synthetic data to those from the reference study. This study is going to be followed by a subsequent benchmark study, in which the known truth in the simulated data sets will be used for performance testing, and the dependence on characteristics such as effect size, sample size etc. will be systematically evaluated. We addressed this issue by updating the abstract and the description in section “study setting”, making it clearer that the focus of the presented study is in validating previous findings, and not in trying to estimate sensitivities and specificities which are planned as a subsequent project. The methodological advancement lies not only in incorporating synthetic data for validation but also in the development of a comprehensive study protocol. 4) The study is about 16S (according to the title and descriptions) but Table 2, Aim 1 discusses metagenome simulation tools. The 16S and metagenome are different approaches for characterizing microbial communities, although there are some overlaps. It would require further clarification why a main Table switches to metagenomics in a 16S paper. Thank you for pointing out that we were not precise in our terminology. In this study we only focus on 16S rRNA sequencing data. We have reviewed the manuscript and changed it accordingly. 5) Availability of materials: the study is based on an open data set that is available. Availability of the source code for the experiments and results would be critical for this kind of study. I could not locate these from the manuscript. Instead, there is a text \"Generated data, analysis scripts, results and supplemental information to this study will also be stored in the Fredato research data management. In case of unexpected technical limitations, we will make data, analysis scripts and supplemental information available via https://figshare.com\". These materials should be made available already at the review stage, otherwise a full review is not possible. The source code for this study, generated data and results will be made available at the review stage of the paper summarizing the results. At this point the source code is not complete yet, as we are only conducting the study once the protocol is accepted. Minor notes: - Real data sets are inherently different from synthetic ones. If the underlying data distributions differ significantly between synthetic and experimental datasets, the statistical tests may yield misleading results. Some discussion could be added on this and other potential limitations of using synthetic data for methods validation. According to the reviewer’s suggestion, we added a brief discussion of this aspect to the study protocol (Discussion section). A more detailed discussion will be included in the paper presenting the results of the study. - Possibly out of scope for the protocol study but in case all the compared data generation methods fail to produce realistic data can the authors consider suggesting their own solution to generate realistic 16S data or expand discussion on this? We deliberately chose not to develop our own simulation tool in this study, as we clearly want to separate benchmarking from methods development to prevent bias. However, to maximize the realism of the synthetic data generated by the tools at hand, we already suggested to introduce two adaptations: adding zeros to the data to achieve realistic levels of sparsity and adjusting the number of differentially abundant features to better capture true effect sizes. - Could you consider using other similar measures to PCA, for instance, principal coordinate analysis based on different dissimilarity metrics (e.g. Jaccard and Bray-Curtis distances) to evaluate the robustness of the ordination comparison? The PCA visualization we proposed will not be conducted on the count matrices, but on the characteristics calculated from those count matrices. Since it would be unusual to use the Bray-Curtis distance as a similarity measure for data characteristics and visualize it by PCoA, especially across multiple experimental datasets with differing taxonomies, we decided to incorporate the average Bray-Curtis distance between all sample pairs as an additional data property. This approach aids in assessing whether simulated count matrices are realistic. - Table 2: \" stasticial analyses\" -> \"statistical analyses\"? Thank you, we have corrected this typo. Is the rationale for, and objectives of, the study clearly described? Partly We acknowledge that the rationale and objectives of the study may not have been communicated clearly. To clarify this, we changed the abstract. To summarize, this study is essentially a validation study, focusing on evaluating the feasibility of using synthetic data to resemble experimental data and whether we replicate findings from the reference study based multiple simulated data sets. Additionally, it is a showcase how to incorporate study protocols in computational studies and to rigorously plan a benchmark study before actually conducting it. This study serves as groundwork for a subsequent benchmark study, which will include a broader range of data sets for performance testing. Is the study design appropriate for the research question? Partly We agree that simulated data has limitations in fully validating previous findings due to inherent differences from experimental data. To address this, we calculate a comprehensive set of data characteristics, perform equivalence tests, and select only simulated data that does not show significant disagreement with experimental data. In our view, this approach is the best we can do to minimize the differences and ensure comparability between simulated and experimental datasets. Moreover, applying the presented equivalence testing procedure represents a novel methodological improvement compared to existing approaches. Are sufficient details of the methods provided to allow replication by others? Partly In the revised version we better explain especially that data generation mechanism, by including more details for the simulation process. All data sets, experimental and generated, as well as all source code to run the study will be made available once the paper describing the results will undergo review process. Are the datasets clearly presented in a useable and accessible format? Yes" } ] }, { "id": "340427", "date": "05 Dec 2024", "name": "Jacob T. Nearing", "expertise": [ "Reviewer Expertise Bioinformatics", "microbiome" ], "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 this protocol Kohnert and Kreutz design a study to assess microbiome differential abundance methods by replicating Nearing et al., 2022 using synthetic datasets generated from the original data using metaSPARSim and sparseDOSSA2. They intend to determine the similarity between the synthetic data generated and their underlying true datasets using 43 different data characteristics. Moreover, they hope to identify whether there are any driving characteristics in synthetic data that would result in discordant conclusions with those achieved by using the real datasets they are based upon. The authors identify that their protocol falls within SPIRIT guidelines and openly report all changes that were made to their report over its initial creation until now. Overall, the study is of interest to microbiome research, however, some areas require clarification, and the overall presentation of the protocol could be improved as major steps in some cases were not presented in order.\n\nMajor comments:\nWell I appreciate the authors adherence to the SPIRIT guidelines it made the ordering of the steps in this protocol hard to follow as these guidelines do not fully align with computational benchmarking studies. For example, the under the section “Modification by adjusting the proportions of zeros and effect sizes” it was mentioned that the groups would be simulated separately, however, the exact simulation procedure (and parameters for metaSPARSim and sparseDOSSA2) was not fully discussed prior to this. Additional information on the code and parameters used in this simulation software (including version # should be included).\nI suspect the authors may have difficulty fully recapturing all the true effect size differences between groups within the same study using synthetic simulated data. Given this I appreciated the in-depth discussion they gave, and the illustration presented in figure 2. However, I do believe that the study could benefit from treating equivalence as a sliding scale rather than a discreate category for each synthetic dataset based on an equivalence test for each data characteristic. It is possible that being equivalent in one characteristic will result in a much larger range than another and this should be taken into consideration during further data analysis. As an aside to this the authors should consider cases where standard deviations may become so large that almost any dataset may pass the equivalence test that it is subjected to (i.e. in figure 1 what happens if the upper and lower bounds cover almost the entire possible range of that data characteristic). In this case it may be reasonable to look at how many times a synthetic dataset from an unrelated template passes an equivalence test. (i.e. how many times does an unrelated dataset that shows very different DA results show equivalence and in what data characteristics does this occur in?).\n\nIn figure 2 and in the section \"Exclusion of simulations for a specific data template based on deviating data properties\" it is suggested that some data characteristics may be too sensitive to include during equivalency testing, however, there is no guidelines to the best of my interpretation of the protocol to determine when a data characteristic would be too sensitive that it must be disregarded.\n\nMinor comments:\n\nSome tools included within this study may have undergone significant updates since the initial publication of Nearing et al., 2022. This should be considered in the analysis as even slightly differences may give rise to discordant results. As an aside to this the authors may want to consider including more recently developed methods in their study such as LOCOM, ANCOM-BC2 or radEmu. This would benefit the microbiome community.\n\nIn some cases, the hypothesis they define for aims 1 and 2 should be reviewed for clarity. I have listed some examples below.\n\nThe authors mention ANCOM-BC however, this DA tool was not considering in Nearing et al., 2022. It is possible that this was typo, and the authors were suggesting ANCOM-II, however, it is important to clarify this as ANCOM-II and ANCOM-BC are highly different methods.\n\nHypothesis 8 in aim 1 should be adjusted as the shapes were similar except for the limma voom tools.\nHypothesis 4 in aim 2 should specify what Wilcoxon method is being referred to.\nHypothesis 9 in aim 2 suggests that in the original study LEFSE had results where more than 99% of features in a dataset were significant, however, I do think this was found in Nearing et al., 2022. Although it was identified to be on the more sensitive side of methods tested.\n\nIs the rationale for, and objectives of, the study clearly described? Partly\n\nIs the study design appropriate for the research question? Partly\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? Yes", "responses": [ { "c_id": "12983", "date": "02 Jan 2025", "name": "Eva Kohnert", "role": "Author Response", "response": "Major comments: Well I appreciate the authors adherence to the SPIRIT guidelines it made the ordering of the steps in this protocol hard to follow as these guidelines do not fully align with computational benchmarking studies. For example, the under the section “Modification by adjusting the proportions of zeros and effect sizes” it was mentioned that the groups would be simulated separately, however, the exact simulation procedure (and parameters for metaSPARSim and sparseDOSSA2) was not fully discussed prior to this. Additional information on the code and parameters used in this simulation software (including version # should be included). In the revised version of our study protocol, we describe in more detail, how the simulated data is generated. We also added version numbers for the two simulation packages. All package versions will be provided when the study is accomplished. We also added a reference to paragraph “Modification by adjusting the proportions of zeros and effect sizes” in section 11b where it is described, how the simulation procedure is changed to adapt the effect size. I suspect the authors may have difficulty fully recapturing all the true effect size differences between groups within the same study using synthetic simulated data. Given this I appreciated the in-depth discussion they gave, and the illustration presented in figure 2. However, I do believe that the study could benefit from treating equivalence as a sliding scale rather than a discreate category for each synthetic dataset based on an equivalence test for each data characteristic. It is possible that being equivalent in one characteristic will result in a much larger range than another and this should be taken into consideration during further data analysis. As an aside to this the authors should consider cases where standard deviations may become so large that almost any dataset may pass the equivalence test that it is subjected to (i.e. in figure 1 what happens if the upper and lower bounds cover almost the entire possible range of that data characteristic). In this case it may be reasonable to look at how many times a synthetic dataset from an unrelated template passes an equivalence test. (i.e. how many times does an unrelated dataset that shows very different DA results show equivalence and in what data characteristics does this occur in?). To address these concerns, we first want to emphasize that each data characteristic (DC) is computed independently for both experimental and simulated count matrices. The scales of different DCs are inherently incomparable; for instance, the proportion of zeros ranges between 0 and 1, while the number of features varies from 327 to 59,736. Thus, the vertical axis in Fig. X is in arbitrary units and +/- 1 SD always cover approx. 68% of the points from the exp. templates. Using the SD over the DCs computed from all exp. templates for defining the equivalence region can be seen as a normalization of the DCs to make it comparable. Our current equivalence testing yields a binary outcome (equivalent or not) to identify and exclude unrealistic simulations. We acknowledge the reviewer's suggestion to treat equivalence as a continuous outcome. However, implementing a sliding scale approach does not provide a final decision and presents challenges due to the diverse scales and distributions of DCs and we have to define a strategy for classification beforehand. Deciding based on how many times a synthetic dataset from an unrelated template passes an equivalence test could behave unintendedly if all synthetic data sets are unrealistic. Thus, we see no clear benefit and a further cutoff has to be defined. Nevertheless, if the reviewer prefers such an option, we are open to implement this (e.g. by defining the equivalence region from the 10% and 90% quantiles of the DC difference to unrelated templates). In response to all reviewers’ feedback, we have refined our set of DCs by eliminating redundancies and incorporating measures of alpha and beta diversity. To further address the reviewer's concerns, we will include a visualization by plotting the number of non-equivalent characteristics against the size of the equivalence region for comparing simulated data with the respective templates and comparing with unrelated templates. This will illustrate the relationship between equivalence thresholds and the proportion of DCs deemed equivalent / non-equivalent, offering deeper insight into the sensitivity of our equivalence testing procedure. We hope that these clarifications and proposed modifications adequately address the reviewer's concerns and enhance our study design. In figure 2 and in the section \"Exclusion of simulations for a specific data template based on deviating data properties\" it is suggested that some data characteristics may be too sensitive to include during equivalency testing, however, there is no guidelines to the best of my interpretation of the protocol to determine when a data characteristic would be too sensitive that it must be disregarded. We apologize for not providing a clear definition earlier. We now define data characteristics more precisely: we will only consider those that pass equivalence tests for at least 50% of the templates. Minor comments: Some tools included within this study may have undergone significant updates since the initial publication of Nearing et al., 2022. This should be considered in the analysis as even slightly differences may give rise to discordant results. As an aside to this the authors may want to consider including more recently developed methods in their study such as LOCOM, ANCOM-BC2 or radEmu. This would benefit the microbiome community. As written in our study protocol (section 9), we’ll apply the latest versions of the DA methods since this seems mostly valuable. We are aware, that updates of the DA methods may be responsible for failed validations. However, we see assessment and confirmation using the most recent tools as more important than preventing the risk that disagreement might originate from improved DA tools. Since a key feature of our study is to strictly perform validations following a strict methodology, we do not want to mix validation with evaluation of other tests. This will be investigated in a subsequent study where we focus on sensitivities and specificities. In the revised version of the protocol, we mention this aspect and better emphasize our overall strategy. We emphasize this strategy in the revised version of the study protocol at several points and use wordings like “validation with the most recent implementations/versions”. In some cases, the hypothesis they define for aims 1 and 2 should be reviewed for clarity. I have listed some examples below. The authors mention ANCOM-BC however, this DA tool was not considering in Nearing et al., 2022. It is possible that this was typo, and the authors were suggesting ANCOM-II, however, it is important to clarify this as ANCOM-II and ANCOM-BC are highly different methods. We thank the reviewer for this note. Upon reflection, we recognize that we indeed planned to use a different ANCOM method than the one employed in Nearing et al. They utilized a custom script downloaded “on May 11, 2020, from the FrederickHuangLin/ANCOM\" repository on GitHub. We will use this script for our analysis to be as consistent as possible with the reference study. Hypothesis 8 in aim 1 should be adjusted as the shapes were similar except for the limma voom tools. Thank you for pointing this out. We changed the hypothesis accordingly. Hypothesis 4 in aim 2 should specify what Wilcoxon method is being referred to. We checked all hypothesis and clearly indicated which method is being referred to. Hypothesis 9 in aim 2 suggests that in the original study LEFSE had results where more than 99% of features in a dataset were significant, however, I do think this was found in Nearing et al., 2022. Although it was identified to be on the more sensitive side of methods tested. Thank you for pointing that out. We changed the hypothesis to be in line with the results from the reference paper. Is the rationale for, and objectives of, the study clearly described? Partly We acknowledge that the rationale and objectives of the study may not have been communicated clearly. To clarify this, we changed the abstract. To summarize, this study is essentially a validation study, focusing on evaluating the feasibility of using synthetic data to resemble experimental data and whether we replicate findings from the reference study based on multiple simulated data sets. Additionally, it is a showcase how to incorporate study protocols in computational studies and to rigorously plan a benchmark study before actually conducting it. This study serves as groundwork for a subsequent benchmark study, which will include a broader range of data sets for performance testing. Is the study design appropriate for the research question? Partly We agree that simulated data has limitations in fully validating previous findings due to inherent differences from experimental data. To address this, we calculate a comprehensive set of data characteristics, perform equivalence tests, and select only simulated data that does not show significant disagreement with experimental data. In our view, this approach is the best we can do to minimize the differences and ensure comparability between simulated and experimental datasets. Moreover, applying the presented equivalence testing procedure represents a novel methodological improvement compared to existing approaches. Are sufficient details of the methods provided to allow replication by others? Partly In the revised version we better explain especially that data generation mechanism, by including more details for the simulation process. All data sets, experimental and generated, as well as all source code to run the study will be made available once the paper describing the results will undergo review process. Are the datasets clearly presented in a useable and accessible format? Yes" } ] } ]	1	https://f1000research.com/articles/13-1180
https://f1000research.com/articles/14-10/v1	02 Jan 25	{ "type": "Data Note", "title": "A guide to selecting high-performing antibodies for Stearoyl-CoA desaturase (SCD1) (UniProt ID: O00767) for use in western blot, immunoprecipitation, and immunofluorescence", "authors": [ "Vera Ruíz Moleón", "Charles Alende", "Maryam Fotouhi", "Sara González Bolívar", "Riham Ayoubi", "Carl Laflamme", "NeuroSGC/YCharOS/EDDU collaborative group", "ABIF consortium", "Vera Ruíz Moleón", "Charles Alende", "Maryam Fotouhi", "Sara González Bolívar", "Riham Ayoubi" ], "abstract": "The enzyme stearoyl-CoA desaturase (SCD1) is a modulator of lipid metabolism by catalyzing the biosynthesis of mono-unsaturated fatty acids from saturated fatty acids. Understanding the specific mechanisms by which SCD1 plays in health and disease can provide novel insides in therapeutic targets, a process that would be facilitated by the availability of high-quality antibodies. Here we have characterized nine SCD1 commercial antibodies for western blot, immunoprecipitation, and immunofluorescence using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. These studies are part of a larger, collaborative initiative seeking to address antibody reproducibility issues by characterizing commercially available antibodies for human proteins and publishing the results openly as a resource for the scientific community. While use of antibodies and protocols vary between laboratories, we encourage readers to use this report as a guide to select the most appropriate antibodies for their specific needs.", "keywords": [ "O00767", "SCD", "SCD1", "Steroyl-CoA desaturase", "antibody characterization", "antibody validation", "western blot", "immunoprecipitation", "immunofluorescence" ], "content": "Introduction\n\nStearoyl-CoA desaturase (SCD1) is a membrane-bound enzyme which catalyzes the rate-limiting step in the conversion of saturated fatty acids into mono-unsaturated fatty acids.1,2 The regulation of SCD1 is physiologically important as maintaining a proper ratio of saturated to monounsaturated fatty acids is essential for membrane fluidity. Disruption to this ratio can lead to pathological conditions, including cardiovascular disease, obesity, non-insulin dependent diabetes mellitus, hypertension, neurological diseases, immune disorders and cancer.2–7\n\nSCD1 is of particular importance in Parkinson’s disease (PD) research as its inhibition has been found toto rescue α-Synuclein cytotoxicity and inclusion formation, both hallmarks of PD progression. The neurotoxic mechanisms underlying PD progression are not yet clearly defined.8–10 Mechanistic studies would be facilitated with the availability of high quality SCD1 antibodies.\n\nThis research is part of a broader collaborative initiative in which academics, funders and commercial antibody manufacturers are working together to address antibody reproducibility issues by characterizing commercial antibodies for human proteins using standardized protocols, and openly sharing the data.11–13 Here we evaluated the performance of nine commercial antibodies for SCD1 for use in western blot, immunoprecipitation, and immunofluorescence, enabling biochemical and cellular assessment of SCD1 properties and function. The platform for antibody characterization used to carry out this study was endorsed by a committee of industry academic representatives. It consists of identifying human cell lines with adequate target protein expression and the development/contribution of equivalent knockout (KO) cell lines, followed by antibody characterization procedures using most commercially available antibodies against the corresponding protein. The standardized consensus antibody characterization protocols are openly available on Protocol Exchange (DOI: 10.21203/rs.3.pex-2607/v1).14\n\nThe authors do not engage in result analysis or offer explicit antibody recommendations. A limitation of this study is the use of universal protocols – any conclusions remain relevant within the confines of the experimental setup and cell line used in this study. Our primary aim is to deliver top-tier data to the scientific community, grounded in Open Science principles. This empowers experts to interpret the characterization data independently, enabling them to make informed choices regarding the most suitable antibodies for their specific experimental needs. Guidelines on how to interpret antibody characterization data found in this study are featured on the YCharOS gateway.15\n\n\nResults and discussion\n\nOur standard protocol involves comparing readouts from WT (wild type) and KO cells.16,17 The first step was to identify a cell line(s) that expresses sufficient levels of a given protein to generate a measurable signal using antibodies. To this end, we examined the DepMap transcriptomics database to identify all cell lines that express the target at levels greater than 2.5 log2 (transcripts per million “TPM” + 1), which we have found to be a suitable cut-off (Cancer Dependency Map Portal, RRID:SCR_017655). The HeLa expresses the SCD1 transcript at 6.7 log2 (TPM+1) RNA levels, which is above the average range of cancer cells analyzed. A SCD KO HeLa cells were obtained from Abcam ( Table 1).\n\nFor western blot experiments, WT and SCD KO protein lysates were ran on SDS-PAGE, transferred onto nitrocellulose membranes, and then probed with nine antibodies in parallel ( Table 2, Figure 1).\n\n* Monoclonal antibody.\n\n** Recombinant antibody.\n\nLysates of HeLa WT and SCD KO were prepared, and 35 μg of protein were processed for western blot with the indicated SCD1 antibodies. The Ponceau stained transfers of each blot are presented to show equal loading of WT and KO lysates and protein transfer efficiency from the acrylamide gels to the nitrocellulose membrane. Tris-Glycine 4-20% gels were used. Antibody dilutions were chosen according to the recommendations of the antibody supplier. An exception was given for antibody AF7550 which was titrated because the signal was too weak when following the supplier’s recommendations. Antibody dilution used: ab19862* at 1/1000, ab236868** at 1/1000, ab39969 at 1/1000, ARP32797_T100 at 1/1000, AF7550 at 1/200, 28678-1-AP at 1/1000, MA5-27542* at 1/1000, PA5-75757 at 1/200 and PA5-95762 at 1/1000. Predicted band size: 41.5 kDa Monoclonal antibody, Recombinant antibody.\n\nWe then assessed the capability of all nine antibodies to capture SCD1 from HeLa protein extracts using immunoprecipitation techniques, followed by western blot analysis. For the immunoblot step, a specific SCD1 antibody identified previously (refer to Figure 1) was selected. Equal amounts of the starting material (SM), the unbound fraction (UB), as well as the whole immunoprecipitate (IP) eluates were separated by SDS-PAGE ( Figure 2).\n\nHeLa lysates were prepared, and immunoprecipitation was performed using 1 mg of lysate and 2.0 μg of the indicated SCD1 antibodies pre-coupled to Dynabeads protein A or protein G. Samples were washed and processed for western blot with the indicated SCD1 antibody. For western blot, MA5-27542 was used at 1/1000. Tris-Glycine 4-20% gels were used. The Ponceau stained transfers of each blot are shown. Predicted band size: 41.5 kDa. SM=4% starting material; UB=4% unbound fraction; IP=immunoprecipitate, HC= antibody heavy chain, LC= antibody light chain. Monoclonal antibody, Recombinant antibody.\n\nFor immunofluorescence, nine antibodies were screened using a mosaic strategy. First, HeLa WT and SCD KO cells were labelled with different fluorescent dyes in order to distinguish the two cell lines, and the SCD1 antibodies were evaluated. Both WT and KO lines imaged in the same field of view to reduce staining, imaging and image analysis bias ( Figure 3). Quantification of immunofluorescence intensity in hundreds of WT and KO cells was performed for each antibody tested, and the images presented in Figure 3 are representative of this analysis.14\n\nHeLa WT and SCD KO cells were labelled with a green or a far-red fluorescent dye, respectively. WT and KO cells were mixed and plated to a 1:1 ratio on coverslips. Cells were stained with the indicated SCD1 antibodies and with the corresponding Alexa-fluor 555 coupled secondary antibody including DAPI. Acquisition of the blue (nucleus-DAPI), green (WT), red (antibody staining) and far-red (KO) channels was performed. Representative images of the merged blue and red (grayscale) channels are shown. WT and KO cells are outlined with green and magenta dashed line, respectively. When an antibody was recommended for immunofluorescence by the supplier, we tested it at the recommended dilution. The rest of the antibodies were tested at 1 and 2 μg/mL and the final concentration was selected based on the detection range of the microscope used and a quantitative analysis not shown here. Antibody dilution used: ab19862 at 1/1000, ab236868** at 1/600, ab39969 at 1/150, ARP32797_T100 at 1/500, AF7550 at 1/200, 28678-1-AP at 1/400, MA5-27542* at 1/1000, PA5-75757 at 1/1000 and PA5-95762 at 1/1300. Bars = 10 μm. Monoclonal antibody, *Recombinant antibody.\n\nIn conclusion, we have screened nine SCD1 commercial antibodies by western blot, immunoprecipitation, and immunofluorescence by comparing the signal produced by the antibodies in human HeLa WT and SCD KO cells. Several high-quality and renewable antibodies that successfully detect SCD1 were identified in all applications. Researchers who wish to study SCD1 in a different species are encouraged to select high-quality antibodies based on the results presented and investigate the predicted species reactivity of the manufacturer before extending their research.\n\n\nMethod\n\nThe standardized protocols used to carry out this KO cell line-based antibody characterization platform was established and approved by a collaborative group of academics, industry researchers and antibody manufacturers. The detailed materials and step-by-step protocols used to characterize antibodies in western blot, immunoprecipitation and immunofluorescence are openly available on Protocol Exchange (DOI: 10.21203/rs.3.pex-2607/v1).14\n\nCell lines used and primary antibodies tested in this study are listed in Tables 1 and 2, respectively. To ensure that the cell lines and antibodies are cited properly and can be easily identified, we have included their corresponding Research Resource Identifiers, or RRID.18,19", "appendix": "Data availability\n\nZenodo: Antibody Characterization Report for SCD1, https://doi.org/10.5281/zenodo.13891494.20\n\nZenodo: Dataset for the SCD1 antibody screening study, https://doi.org/10.5281/zenodo.14502183.21\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgment\n\nWe would like to thank the NeuroSGC/YCharOS/EDDU collaborative group for their important contribution to the creation of an open scientific ecosystem of antibody manufacturers and KO cell line suppliers, for the development of community-agreed protocols, and for their shared ideas, resources, and collaboration. Members of the group can be found below. We would also like to thank the Advanced BioImaging Facility (ABIF) consortium for their image analysis pipeline development and conduction (RRID:SCR_017697). Members of each group can be found below.\n\nNeuroSGC/YCharOS/EDDU collaborative group: Thomas M. Durcan, Aled M. Edwards, Peter S. McPherson, Chetan Raina and Wolfgang Reintsch.\n\nABIF consortium: Claire M. Brown and Joel Ryan.\n\nThank you to the Structural Genomics Consortium, a registered charity (no. 1097737), for your support on this project. The Structural Genomics Consortium receives funding from Bayer AG, Boehringer Ingelheim, Bristol-Myers Squibb, Genentech, Genome Canada through Ontario Genomics Institute (grant no. OGI-196), the EU and EFPIA through the Innovative Medicines Initiative 2 Joint Undertaking (EUbOPEN grant no. 875510), Janssen, Merck KGaA (also known as EMD in Canada and the United States), Pfizer and Takeda.\n\n\nReferences\n\nEnoch HG, Catalá A, Strittmatter P: Mechanism of rat liver microsomal stearyl-CoA desaturase. Studies of the substrate specificity, enzyme-substrate interactions, and the function of lipid. J. Biol. Chem. 1976; 251(16): 5095–5103. PubMed Abstract \| Publisher Full Text\n\nNtambi JM: Regulation of stearoyl-CoA desaturase by polyunsaturated fatty acids and cholesterol. J. Lipid Res. 1999; 40(9): 1549–1558. Publisher Full Text\n\nKinsella JE, Lokesh B, Stone RA: Dietary n-3 polyunsaturated fatty acids and amelioration of cardiovascular disease: possible mechanisms. Am. J. Clin. Nutr. 1990; 52(1): 1–28. PubMed Abstract \| Publisher Full Text\n\nJones BH, Maher MA, Banz WJ, et al.: Adipose tissue stearoyl-CoA desaturase mRNA is increased by obesity and decreased by polyunsaturated fatty acids. Am. J. Phys. 1996; 271(1 Pt 1): E44–E49. Publisher Full Text\n\nLi J, Ding SF, Habib NA, et al.: Partial characterization of a cDNA for human stearoyl-CoA desaturase and changes in its mRNA expression in some normal and malignant tissues. Int. J. Cancer. 1994; 57(3): 348–352. PubMed Abstract \| Publisher Full Text\n\nHabib NA, Wood CB, Apostolov K, et al.: Stearic acid and carcinogenesis. Br. J. Cancer. 1987; 56(4): 455–458. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKhoo DE, Fermor B, Miller J, et al.: Manipulation of body fat composition with sterculic acid can inhibit mammary carcinomas in vivo. Br. J. Cancer. 1991; 63(1): 97–101. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFanning S, Haque A, Imberdis T, et al.: Lipidomic Analysis of α-Synuclein Neurotoxicity Identifies Stearoyl CoA Desaturase as a Target for Parkinson Treatment. Mol. Cell. 2019; 73(5): 1001–14.e8. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBartels T, Choi JG, Selkoe DJ: α-Synuclein occurs physiologically as a helically folded tetramer that resists aggregation. Nature. 2011; 477(7362): 107–110. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNicholatos JW, Groot J, Dhokai S, et al.: SCD Inhibition Protects from α-Synuclein-Induced Neurotoxicity But Is Toxic to Early Neuron Cultures. eNeuro. 2021; 8(4): ENEURO.0166–ENEU21.2021. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyoubi R, Ryan J, Biddle MS, et al.: Scaling of an antibody validation procedure enables quantification of antibody performance in major research applications. elife. 2023; 12: 12. Publisher Full Text\n\nCarter AJ, Kraemer O, Zwick M, et al.: Target 2035: probing the human proteome. Drug Discov. Today. 2019; 24(11): 2111–2115. PubMed Abstract \| Publisher Full Text\n\nLicciardello MP, Workman P: The era of high-quality chemical probes. RSC Med. Chem. 2022; 13(12): 1446–1459. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyoubi R, Ryan J, Bolivar SG, et al.: A consensus platform for antibody characterization (Version 1). Protocol Exchange. 2024.\n\nBiddle MS, Virk HS: YCharOS open antibody characterisation data: Lessons learned and progress made. F1000Res. 2023; 12: 1344. Publisher Full Text\n\nLaflamme C, McKeever PM, Kumar R, et al.: Implementation of an antibody characterization procedure and application to the major ALS/FTD disease gene C9ORF72. elife. 2019; 8: 8. Publisher Full Text\n\nAlshafie W, Fotouhi M, Shlaifer I, et al.: Identification of highly specific antibodies for Serine/threonine-protein kinase TBK1 for use in immunoblot, immunoprecipitation and immunofluorescence. F1000Res. 2022; 11: 977. Publisher Full Text\n\nBandrowski A, Pairish M, Eckmann P, et al.: The Antibody Registry: ten years of registering antibodies. Nucleic Acids Res. 2023; 51(D1): D358–D367. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBairoch A: The Cellosaurus, a Cell-Line Knowledge Resource. J. Biomol. Tech. 2018; 29(2): 25–38. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRuiz Moleon V, Alende C, Fotouhi M, et al.: A guide to selecting high-performing antibodies for Stearoyl-CoA desaturase (SCD1) (UniProt ID: O00767). Zenodo. 2024.Publisher Full Text\n\nLaflamme C: Dataset for the SCD1 antibody screening study. [Dataset]. Zenodo. 2024. Publisher Full Text" }	[ { "id": "355980", "date": "17 Jan 2025", "name": "Cecilia Williams", "expertise": [ "Reviewer Expertise nuclear receptors", "transcriptomes", "antibody validation", "cancer" ], "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 data note by Ruiz Moleon et al. provides a high-quality antibody validation guide for the protein SCD1. The guide is part of the YCharOS initiative that uses isogenic knockout cell lines and multiple antibody-dependent assays to investigate antibody specificity and selectivity in a structured manner. It is an important effort that can improve the reproducibility of biomedical research, and this guide is highly useful for researchers focusing on SCD1. The authors show that 3-4 out of 9 antibodies do not work in most or any of the assays, whereas the rest are more or less well-performing, using the cells and conditions applied in the protocol. This is critical information that will be of high value to the research community. For enhanced usability, clarification, and stringency of the guide and to facilitate the interpretation and reproducibility, I have the below recommendations.\nMajor comments\nThe introduction would benefit from including basic information regarding the protein, as this would facilitate the interpretation of the data. Such as the known or predicted subcellular localization of SCD1, its calculated molecular weight, and if any splice variants are known. The rationale, as stated (p.3, 2nd paragraph: “Mechanistic studies would be facilitated with the availability of high-quality SCD1 antibodies”) should be better supported. Is there a documented lack of high-quality SCD1 antibodies? Or, is this an assumption based on the general problem with the reproducibility of antibody-based research? The sentence “using most commercially available antibodies against the corresponding protein” (p. 3, 3rdparagraph) is likely not correct. Perhaps the authors intended to state “renewable antibodies” as the initiative states in other publications. In this guide, specifically, the authors include 9 commercial (renewable and non-renewable) SCD1 antibodies, but there are as many as 274 antibodies from 32 providers directed towards human SCD1 listed in Antibodypedia. A statement of whether the SCD1 gene in the cells (HeLa WT) has indeed been confirmed to be wild type (i.e., not mutated) should be added. It would also be useful to include which (if any) splice variant transcripts are expressed in these cells. Table 1: The lot (or batch) numbers for the antibodies must be listed. This is essential information and is especially important in an antibody validation guide. It should be stated whether replicate experiments were performed. It is unclear how the antibody used for the western blot of the IPs was chosen (p. 3, 2nd last paragraph: “a specific antibody was chosen”). Were both specificity and selectivity considered? Were the species the antibodies were generated in used as criteria (to avoid detection of heavy and light chains in the next step)? Or was the selection random out of the sufficiently specific antibodies? If so, how many and which antibodies were deemed to be sufficiently specific? Fig 2: Considering that antibodies from different species appear specific in WB, it seems like a missed opportunity to not take species into account to generate clearer western blot data of the IPs (i.e., avoiding additional bands of heavy and light antibody chains, such as when using mouse-generated antibodies in both IP and WB and detecting them with an anti-mouse secondary antibody). Fig. 1 and 2: It should be specified which secondary antibody was used in the western blots. Fig. 3: The legend statement that “Representative images of the merged blue and red (grayscale) channels are shown.” appears incorrect. The blue (DAPI) is shown separately from the red (antibody staining); they are not merged in the figure. It would also be clearer if the images showed the actual colors (blue for DAPI and red for SCD1 antibody) instead of grey for both. Only one cell line is used, which is a limitation of the initiative. Considering that HeLa cells express unusually high SCD1 levels (as stated in the text), the antibodies may work less well at lower SCD1 levels. They could also bind to other proteins expressed in other cells but not HeLa. It would be useful if the antibodies (the ones deemed to be of high quality) were additionally analyzed by western blot on a panel of cell lines with varying mRNA expression of SCD1. This would indicate their performance at different levels of the target protein and in more than one cell line. Although the authors state that they do not engage in result analysis, they do (to select a specific antibody for WB of the IP, and they conclude that several antibodies successfully detect SCD1), but they refrain from clearly stating which these antibodies are. A table, or a statement, summarizing which antibodies bind intended and unintended targets in the different assays (under the specific conditions used) would allow greater usability, including for researchers starting their work (such as postdocs and PhD students). This would also facilitate an overview of the application-dependent performance (again, under the specific conditions used).\nMinor comments:\nThe assay immunofluorescence (IF) is typically used for tissue analysis. For analysis of cells and cell lines, as is the case here, the term immunocytochemistry is better suited. UniProt ID may not need to be stated in the title (could be moved to abstract or introduction) p.3, 3rd paragraph: It should be clarified if the target protein expression relates specifically to endogenous expression or whether cell lines with recombinant expression were included. A statement about mycoplasma testing and cell line authentication should be included. It would be beneficial to include which epitope each antibody is directed toward (or raised against). Fig. 1 and 2: The exposure time(s) applied for the western blots could be included. Fig 2: Although the figure legend clearly states which antibody was used for WB of all IPs, the figure itself gives the impression that it may have been used only for the two indicated IPs. It might be better to remove this from the image or place it differently. p. 5, first paragraph: “Quantification of ... in hundreds of WT and KO cells” is vague. The number (or range) of actually quantified cells should be stated. Fig. 3: The figure legend should specify which magnification was used for the IF images. It would be beneficial to highlight that the antibodies ab19862 and MA-27542 are from the same clone and should be the same. It would be pedagogical to place them side by side. Page 5, final paragraph: “Several high-quality and renewable antibodies that successfully detect SCD1 were identified in all applications”. As only 3 of the tested antibodies were renewable (monoclonal or recombinant), and two of them were from the same clone (= same antibody), the word “several” seems a stretch. The word “renewable” should probably be removed. Several spelling/grammar errors (p.3 a “to” too much p.3, p. 3 “were run “ instead of “were ran”, wrongly placed commas, etc.) The published method protocol Ayoubi et al., Nature Protocol (2024) should likely be referenced. Author roles: It is not stated who drafted the manuscript, nor if all authors commented on it.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\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": [] }, { "id": "358436", "date": "05 Feb 2025", "name": "Michael L Garelja", "expertise": [ "Reviewer Expertise CGRP", "migraine", "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\nThis manuscript, aiming to investigate the utility of a range of SCD1 antibodies is extremely important, and I applaud the initiative. Identification of antibodies that can correctly detect their target is of incredible importance to the entire field and highlighting that not all antibodies work as advertised is critical to ensuring that researchers can perform robust science.\nPoints that need be addressed for this work to be indexed.\nAre there RRID’s associated with these antibodies? This applies for both the primary antibodies, and the secondary antibodies. Secondary antibodies also require lot numbers. As antibody concentrations can change between batches, it would be more informative to list the concentrations used in the experiments rather than the dilutions. A summary table to show how the different antibodies work under different experimental techniques would be extremely useful. I understand that the resource cannot make recommendations, but having a summary of how each antibody would help others interpret results. Are there any known splice variants of this target? There is a consistent band of approximately 25 kDa, that also seems to be lower when knocked out. Any insight as to why the detected band differed from the estimated molecular weight would also be useful. Has this particular ladder been compared to other ladders to ensure that it runs as expected under these experimental conditions?\nFurther information for discussion.\nThe researchers have used a cell-line which highly expresses the target. Is it possible to work with cells that have lower expression levels? If not, these needs to be addressed as a limitation. Testing the panel of “useful” antibodies against more cell lines with variable SCD1 expression to show a limit of detection would be of high utility. Listing the epitopes that the antibodies were raised against would be of high value. One of the pillars of antibody validation is to use two antibodies that have been raised against different epitopes, so having that information readily available would help researchers in their mission to perform good science. The paper should acknowledge that there may be methodological optimizations that could improve antibody function. Figure 3 – is it possible to show to green and red stained cells in a separate panel? The outlines are useful, but showing that the cells have similar morphologies (or not!) would be interesting in the context of SCD1 biology. Why were these antibodies chosen, as opposed to the wider range of antibodies available? This is not a criticism, as the work is valuable regardless, but some information as to how these antibodies were selected would be interesting.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\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": [] } ]	1	https://f1000research.com/articles/14-10
https://f1000research.com/articles/14-9/v1	02 Jan 25	{ "type": "Study Protocol", "title": "FXR agonist in post-liver transplantation patients: a randomized open-labeled study.", "authors": [ "Anila K N", "Saraswathy S Nair", "Dinesh Balakrishnan`", "Unnikrishnan G", "Binoj S T", "Ramachandran Narayana Menon", "Christi Titus Varghese", "Shweta Mallick", "Krishnanunni Nair", "Madhu Srinivasan Durairaj", "Guhan V", "Nafiya Zackariah", "Haritha Rajakrishnan", "Arun Valsan", "Johns Shaji Mathew", "Cyriac Abby Philips", "Christopher Watson", "Sudheer O V", "Sudhindran S", "Saraswathy S Nair", "Dinesh Balakrishnan`", "Unnikrishnan G", "Binoj S T", "Ramachandran Narayana Menon", "Christi Titus Varghese", "Shweta Mallick", "Krishnanunni Nair", "Madhu Srinivasan Durairaj", "Guhan V", "Nafiya Zackariah", "Haritha Rajakrishnan", "Arun Valsan", "Johns Shaji Mathew", "Cyriac Abby Philips", "Christopher Watson", "Sudheer O V" ], "abstract": "Liver diseases cause nearly 2 million deaths worldwide each year, with approximately 1 million deaths from cirrhosis complications and another 1 million from viral hepatitis and liver cancer, according to WHO estimates. Liver transplantation (LT) remains the primary curative option, boasting success rates of 85% in the first year and 75% at five years post-transplant. Despite high costs, LT is considered cost-effective, especially for younger patients with active work years remaining. However, post-transplant complications, particularly intrahepatic cholestasis, present notable challenges. This complication arises from factors such as ischemia-reperfusion injury, infections, immunological rejection, and surgical complications, all contributing to impaired bile flow and liver damage. Current medical therapies for post-LT cholestasis are limited, with ursodeoxycholic acid (UDCA) frequently used, despite questionable efficacy. Obeticholic acid (OCA), a potent Farnesoid X Receptor (FXR) agonist approved by the FDA for treating primary biliary cholangitis (PBC), has shown potential benefits in reducing elevated cholestatic liver enzymes. Given its significant effects on liver health, OCA may offer therapeutic value in managing post-transplant cholestasis and improving graft survival. This randomized controlled trial (RCT) aims to evaluate OCA’s efficacy compared to UDCA in reducing cholestatic injury and enhancing graft function post-LT. The primary outcomes will focus on a 15% reduction in alkaline phosphatase and gamma-glutamyl transferase levels at 3, 6, and 12 months from baseline one month. Secondary outcomes include molecular markers, biliary complications, graft rejection, quality of life, and cost-effectiveness. Results are anticipated to demonstrate that OCA could improve graft survival, reduce complications, and enhance quality of life, potentially setting a new standard in post-LT care if found beneficial.", "keywords": [ "Liver transplantation", "cholestasis", "ursodeoxycholic acid", "obeticholic acid", "alkaline phosphatase." ], "content": "Introduction\n\nLiver diseases cause nearly 2 million deaths worldwide each year, with approximately 1 million deaths from cirrhosis complications and another 1 million from viral hepatitis and liver cancer, according to WHO estimates.1,2 Liver transplantation (LT) is recognized as the primary curative treatment, with success rates reaching 85% in the first year and 75% at 5 years post-transplantation.3 In India, over 3000 liver transplants are performed annually. Despite the considerable expense associated with LT, it is considered cost-effective, especially for individuals in their prime years, who have many active work years ahead of them.4,5 However, post-transplant morbidity, particularly intrahepatic cholestasis, presents significant challenges.6\n\nIntrahepatic cholestasis is a major complication following liver transplantation, caused by multiple factors. Ischemia-reperfusion injury during organ retrieval and transplantation leads to cellular damage and postoperative cholestasis.7,8 Sepsis from infections worsen cholestasis through inflammatory responses.9 Essential post-transplant drugs, including immunosuppressants and antibiotics, can interfere with bile formation, causing liver injury.10,11 Immunological reactions such as T-cell and B-cell-mediated rejection and lymphocytic cholangitis also contribute to cholestasis.12 Surgical complications like stenosis or obstruction of the biliary and vascular anastomosis can also result in intra- and extrahepatic cholestasis.13,14\n\nThe absence of effective medicines limits the current medical therapy of cholestasis. Although its usefulness is questionable, many practitioners utilize ursodeoxycholic acid (UDCA) as a hepatoprotective drug.15 It is usually administered for three months post-transplant at a dose of 10-15 mg/kg twice daily. In 2016, the FDA approved obeticholic acid (OCA), a strong selective Farnesoid X Receptor (FXR) agonist, to treat primary biliary cholangitis (PBC) in conjunction with UDCA.16,17 Research has demonstrated that over 12 months, OCA, either in combination with UDCA or on its own, dramatically lowers levels of total bilirubin and alkaline phosphatase.18,19 In individuals with advanced liver disease, the FDA has cautioned against administering OCA at doses of 10–50 mg.20 Studies comparing OCA’s effects on cholestasis, rejection, regeneration, and cost-effectiveness to the current standard UDCA are being conducted to assess the role of OCA in the postoperative recovery of liver transplant patients.21 Our proposed project is a clinical trial between OCA and the current standard of care, UDCA, in managing post-liver transplant cholestasis and improving graft survival.22\n\nFollowing LT, a major morbidity mitigating long-term liver functions is intrahepatic cholestasis due to a multitude of reasons. The only drug available for the treatment of intrahepatic cholestasis currently is UDCA.23 FXR is a newly detected receptor gene, playing multiple roles in liver homeostasis, including detoxification of bile acid excess, stimulation of bile salt export from hepatocytes, reduction of inflammation, delaying fibrosis, and perhaps may even assist liver regeneration.24,25 A promising alternative is obeticholic acid, that has got its approval by the FDA for cholestatic conditions like primary biliary cholangitis (PBC).26 Research shows OCA significantly reduces key liver enzymes like alkaline phosphatase, gamma glutamy transferase and total bilirubin in cholestatic liver disease, though its effects specifically in LT patients have not been well-studied.27,28\n\nThis study aims to fill an essential gap by evaluating the efficacy of obeticholic Acid compared to ursodeoxycholic Acid in managing post-liver transplant cholestasis. By conducting specific biochemical tests to assess cholestasis at the cellular level, this research will provide insights into the effectiveness of OCA relative to the current standard, UDCA.29 Additionally, this study will examine the potential effects of OCA on rejection, tissue regeneration, and cost-effectiveness—areas where knowledge is currently limited.30,31 The ultimate goal of this study is to determine whether OCA can emerge as a new standard of care in post-LT management, offering cost-effective solutions to enhance long-term outcomes for liver transplant patients.\n\nWe are assessing the effects of obeticholic acid compared to ursodeoxycholic acid in post-liver transplant patients in ameliorating cholestatic injury with primary endpoints, a 15% reduction in alkaline phosphatase and gamma-glutamyl transferase levels at 3, 6, and 12 months from one-month baseline between groups.\n\nAssessment of biochemical and molecular markers (fibroblast growth factor 19 (FGF-19), transforming growth factor β (TGF-β) level, Cytokeratin 18 level, Serum autotaxin level, Bile Salt Export Pump levels (BSEP), Plasma bile acid levels.\n\n- Biopsy-proven acute or chronic rejection.\n\n- Incidence of biliary complications.\n\n- All-cause mortality.\n\n- Any adverse events.\n\n- Death due to acute or chronic rejection.\n\n- Quality of life.\n\n\nMethods\n\nThis three-year RCT study was conducted in the Department of GI Surgery, Amrita Institute of Medical Sciences, Kochi.\n\nThe study is conducted as an investigator-initiated, parallel-group, open-labeled randomized control trial in patients who underwent live donor liver transplantation from December 2014 to 2021 in the Department of Gastrointestinal Surgery at a tertiary care teaching hospital.\n\nAll adult patients who underwent live donor liver transplantation during the study period were included. Recipients less than 18 years of age, deceased donor liver transplants, ABO-incompatible transplants, recipients who did not survive beyond 30 days, patients lost to follow-up during the study period, and patients who discontinued study drugs were excluded.\n\nA clinical pharmacist randomized study participants admitted to the GI surgery ICU department following liver transplantation. A permuted block sequence of computer-generated random numbers was used to enroll subjects into either the control arm (C) or the study arm (T), i.e., either to receive the standard liver protectant ursodeoxycholic acid [URSETOR® of Torrent Pharma]10-15 mg/kg TID (control) or obeticholic acid 5 mg OD [OCANASH® of Macleods Pharmaceutical Limited]. The sequence is kept in sealed opaque envelopes and opened by the pharmacist on 2nd day following LT. Given the difference in the size of tablets and the difference in dosing frequency (three times for UDCA vs once daily for OCA), blinding was not done.\n\nFollow-up visits for physical examinations and safety assessments are scheduled during the study period at the following intervals: one month, three months, six months, and twelve months post-transplant ( Figure 1). Liver function tests, C-reactive protein (CRP), platelet count, prothrombin time/international normalized ratio (PT/INR), fasting blood sugar (FBS), tacrolimus concentration, and other relevant parameters were measured at each follow-up visit. Lipid profile (including total cholesterol, low-density lipoprotein (LDL), high-density lipoprotein (HDL), very low-density lipoprotein (VLDL), and triglycerides (TG)), as well as glycosylated hemoglobin (HbA1c) levels, were assessed at pre-transplant admission, three months, six months, and twelve months.\n\nSecondary endpoint serum markers are assessed using an enzyme-linked immunosorbent assay (ELISA) [ELK Biotechnology™] at the centralized biochemistry laboratory. Blood samples collected during the one-month and twelfth month post-transplant follow-up were used for ELISA testing.\n\nIn addition, demographics, patient characteristics, and other critical study details (e.g., medication history, medical history, social habits, family history, and use of other medical systems) are collected through direct patient or bystander interviews, doctor consultation notes, hospital health information systems, and case records. The collected data is transcribed into a pre-designed proforma (data collection form) by the investigator.\n\nA biopsy is performed when suspected rejection (unexplained rise in transaminases) is observed. The biopsy results are evaluated based on the Rejection Activity Index score and the Banff criteria. Data on the frequency and duration of the first biopsy-proven acute rejection (BPAR) episode (within six months of transplantation) that required therapy are recorded. Any biliary system-related strictures, leaks, or anastomosis issues are documented. Early allograft transplant dysfunction (EAD) is assessed using criteria developed by Olthoff et al.7,32\n\nAdverse reactions are recorded in the case record form, and their causality is assessed using the Naranjo scale, classifying them as definite, probable, possible, or unlikely.\n\nDuring patient interviews, a validated Chronic Liver Disease Questionnaire (CLDQ) is administered to assess quality of life (QOL).33 The copyright license for the English questionnaire version is obtained from the authors. The QOL questionnaire is completed both pre-transplant and at the one-year post-transplant follow-up. Patients completed the questionnaire in either Malayalam or English.\n\nThe CLDQ includes six domains:\n\n• Abdominal symptoms (AB): Items 1, 5, 17\n\n• Fatigue (FA): Items 2, 4, 8, 11, 13\n\n• Systemic symptoms (SY): Items 3, 6, 21, 23, 27\n\n• Activity (AC): Items 7, 9, 14\n\n• Emotional functions (EM): Items 10, 12, 15, 16, 19, 20, 24, 26\n\n• Worry (WO): Items 18, 22, 25\n\nThe CLDQ overall score is the sum of the scores from each domain, ranging from 0 to 203. Lower scores indicate poorer quality of life.\n\nThe sample size was calculated based on a pilot study comparing the effect of obeticholic acid (OCA) and ursodeoxycholic acid (UDCA) on cholestatic injury in post-liver transplant patients. In the pilot study, the difference in alkaline phosphatase (5.131 ± 39.639 vs 12.496 ± 52.139) was observed between the two groups. With 80% power and 95% confidence, the minimum sample size required was determined to be 108 patients in each group, totaling 216 samples.\n\nThe study is currently in the recruitment phase.\n\n\n\n1. To compare the means of continuous variables between groups (Drug A vs Drug B), an independent sample t-test will be used at each time point.\n\n2. To compare the changes in continuous variables from pre- (1 month) to post-transplant (3, 6, and 12 months), a paired t-test will be applied.\n\n3. To compare the means of variables from baseline to subsequent time points within each group, repeated measures ANOVA will be used if significant. The Bonferroni multiple comparison test will be applied for pairwise comparisons.\n\n\nExpected results\n\nWe expect a significant reduction in cholestatic liver enzymes following LDLT in the OCA group compared to the UDCA group, with improvements observed from one month to subsequent time points (3, 6, and 12 months). We hypothesize that the potent mechanism of action of OCA will ameliorate cholestatic injury and enhance graft survival. Additionally, we anticipate a reduction in biliary complications, all-cause mortality, and an increase in quality of life for OCA-receiving patients. This may reduce long-term financial expenditure for patients. Given that there are currently no drugs to prevent damage to the transplanted liver, a positive outcome could be a significant advancement for the transplant population. If beneficial, this study could lead to the adoption of OCA as a standard of care following liver transplantation.\n\n\nConclusion\n\nObeticholic acid (OCA) is a potent farnesoid X receptor (FXR) agonist, with 100 times the efficacy of the current standard treatment, UDCA, in maintaining liver homeostasis. While OCA has shown promise in treating cholestatic liver diseases, its effects post-liver transplantation (LT) remain unclear due to the lack of high-quality trials. Our study aims to evaluate whether OCA’s strong FXR activation can benefit patients with post-LT cholestasis. We hypothesize that OCA will improve graft function, reduce biliary complications, lower associated costs, and enhance overall quality of life and survival for post-LT patients.\n\nAll information about participants enrolled in this clinical trial are treated with the utmost confidentiality to protect their privacy and adhere to ethical standards. Personal identifying information are securely stored and accessible only to authorized personnel involved in the study. Data collected are anonymized and coded to maintain confidentiality during analysis and reporting. Any publication or presentation of results will not include identifiable information unless explicit consent is obtained from participants.\n\nAccess to data collected during this clinical trial will be managed according to established protocols and regulatory requirements. Authorized personnel, including investigators, statisticians, and regulatory authorities, will have access to raw data for analysis, monitoring, and verification. Data-sharing agreements will be established to govern access by external parties, ensuring compliance with data protection regulations and participant confidentiality. Requests for access to study data from external researchers or institutions will be considered on a case-by-case basis and subject to review by the study sponsor and relevant ethics committees.\n\nUpon completion of the study, data will be disseminated through peer-reviewed journal publications, conference presentations, and participant communication. Findings will be shared transparently with participants, addressing risks and benefits. Regulatory reporting will ensure compliance with obligations to relevant agencies. Additionally, data-sharing agreements and repositories will facilitate access for other researchers while maintaining participant confidentiality.\n\nThe study protocol was initially approved by the Institutional Ethics Committee, AIMS (IEC-AIMS-2021-GISURG-262) on 13-10-2021 and later modified for a title change on 15-06-2022 (modified ethics number—IEC-AIMS-2022-GISURG-160). This study involves liver transplant patients aged 18 years or older admitted to our hospital’s gastrointestinal surgery department. Before the study’s initiation, patients gave written informed consent. The consent form was prepared both in English and the local language Malayalam. The study has received ethical clearance from our Institutional Ethics Committee. The committee also approved the informed consent in both languages. This study was registered with the Clinical Trials Registry of India (CTRI/2021/11/038218). The study was conducted according to the principles of the Declaration of Helsinki.\n\n\nAuthor contributions\n\n\n\n• S. Sudhindran, Unnikrishnan G, Dinesh Balakrishnan, Johns Shaji Mathew, and Shweta Mallick contributed to formulating the concepts.\n\n• Implementation of study including randomization, and data collection: Anila KN, Saraswathy S Nair, Nafiya Zackariah, Haritha Rajakrishnan.\n\n• Drafting manuscript: Anila K N.\n\n• S. Sudhindran, OV Sudheer, Arun Valsan, Binoj ST, Ramachandran Narayana Menon, Krishnanunni Nair, Madhu Srinivasan Durairaj, V Guhan, Christi Titus Varghese contributed to giving valuable inputs and critical revisions.", "appendix": "Data availability statement\n\nNo data are associated with this article.\n\nOpen Science Frame (OSF): FXR agonist in post Liver transplantation patients: A randomized open-labeled study, DOI: 10.17605/OSF.IO/JDXBF34\n\nThis project contains the following underlying data:\n\n1. CLD malayalam questionnaire. Pdf\n\n2. CLD QUESTIONNAIRE english.pdf\n\n3. Data collection CRF.pdf\n\n4. Ethics docs.zip\n\n5. Fillable-SPIRIT-Outcomes-2022-Checklist-with-SPIRIT-2013.pdf\n\n6. INFORMED CONSENT DOCUMENT english final.pdf\n\n7. informed consent malayalam modified.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\nReferences\n\nWorld Health Organization: Global health estimates 2020: Deaths by cause, age, sex, by country and by region, 2000-2019. Geneva: WHO; 2020.\n\nMokdad AA, Lopez AD, Shahraz S, et al.: Liver disease burden: global epidemiology and trends. Lancet Gastroenterol. Hepatol. 2020; 5(8): 738–742.\n\nKim WR, Lake JR, Smith JM, et al.: Liver transplantation across the world: results of the international registry. Transplantation. 2022; 106(2): 354–365.\n\nSarin SK, Choudhury A: Liver transplantation in India: challenges and opportunities. J. Clin. Exp. Hepatol. 2021; 11(6): 693–701.\n\nNeuberger J, Gilroy R, Baccarani U, et al.: Liver transplantation: economics and outcomes. Hepatology. 2020; 72(3): 1433–1440.\n\nZimmerman HJ: Drug-induced liver disease. Clin. Liver Dis. 2020; 24(3): 631–642.\n\nOlthoff KM, Kulik L, Samstein B, et al.: Validation of a definition of early allograft dysfunction in liver transplant recipients and analysis of risk factors. Liver Transpl. 2010; 16(8): 943–949. PubMed Abstract \| Publisher Full Text\n\nHirschfield GM, Mason A, Luketic V, et al.: Efficacy of obeticholic acid in patients with primary biliary cholangitis and an inadequate response to ursodeoxycholic acid. Gastroenterology. 2015; 148(4): 751–761.e8. PubMed Abstract \| Publisher Full Text\n\nFDA: Obeticholic acid (Ocaliva) product label. Silver Spring, MD: U.S. Food and Drug Administration; 2016 [cited 2024 Nov 15]. Reference Source\n\nAron-Wisnewsky J, Clément K, Pépin JL: Nonalcoholic fatty liver disease and obstructive sleep apnea. Metabolism. 2016; 65(8): 1124–1135. Publisher Full Text\n\nYounossi ZM, Rinella ME, Sanyal AJ, et al.: Nonalcoholic steatohepatitis: past, present, and future. Hepatology. 2021; 73(Suppl 1): 1194–1198. Publisher Full Text\n\nKowdley KV, Luketic VA, Chapman RW, et al.: A randomized trial of obeticholic acid monotherapy in primary biliary cholangitis. Gastroenterology. 2018; 155(3): 678–689.\n\nBolt MJ, Bergquist A, Verheij J, et al.: Morphological criteria for the diagnosis of lymphocytic cholangitis. Clin. Gastroenterol. Hepatol. 2021; 19(9): 1916–1924.\n\nHirschfield GM, Karlsen TH, Lindor KD, et al.: Primary sclerosing cholangitis. Lancet. 2013; 382(9904): 1587–1599. Publisher Full Text\n\nKulik L, El-Serag HB: Epidemiology and management of hepatocellular carcinoma. Gastroenterology. 2019; 156(2): 477–491.e1. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLoomba R, Sanyal AJ: The global NAFLD epidemic. Nat. Rev. Gastroenterol. Hepatol. 2013; 10(11): 686–690. Publisher Full Text\n\nFDA: Obeticholic acid: safety announcement for use in advanced liver disease. Silver Spring, MD: U.S. Food and Drug Administration; 2023 [cited 2024 Nov 15]. Reference Source\n\nLevy C, Seeff LD, Lindor KD: Use of ursodeoxycholic acid in treating liver disease. Clin. Gastroenterol. Hepatol. 2004; 2(8): 819–827.\n\nTrauner M, Fickert P, Wagner M: Bile acids as regulators of hepatic lipid and glucose metabolism. Dig. Dis. 2010; 28(1): 220–224. Publisher Full Text\n\nGuicciardi ME, Gores GJ: Apoptosis as a mechanism for liver disease progression. Semin. Liver Dis. 2010; 30(4): 402–410. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDufour JF, Bizzaro N, Cappelletto M, et al.: The clinical use of obeticholic acid in patients with primary biliary cholangitis and advanced liver disease: a multicenter cohort study. J. Hepatol. 2021; 75(3): 687–697.\n\nVargas-Villanueva M, Vasquez-Rios R, Mena J, et al.: Liver transplantation for cirrhosis in Latin America: clinical outcomes and challenges. World J. Hepatol. 2019; 11(5): 303–309.\n\nTapper EB, Sengupta N, Sanyal AJ, et al.: The cost-effectiveness of liver transplantation. Hepatology. 2020; 71(5): 1869–1876.\n\nLuetmer PJ, Jensen RT, Cuenco J: Radiologic evaluation of biliary complications following liver transplantation. Radiol. Clin. North Am. 2021; 59(4): 773–786.\n\nMc Hutchison JG, Lawitz EJ, Shiffman ML, et al.: Phase 2 study of obeticholic acid in patients with chronic hepatitis C. Hepatology. 2016; 63(4): 1317–1328.\n\nMoretti M, Calvaruso V, Ponziani FR, et al.: Bile acid therapy in liver transplantation: a review of the literature. J. Hepatol. 2021; 75(3): 698–708.\n\nRinella ME, Loomba R, Sanyal AJ, et al.: Efficacy of obeticholic acid in the treatment of nonalcoholic steatohepatitis: a randomized controlled trial. Gastroenterology. 2020; 158(2): 574–584.\n\nGupta A, Suri V, Sharma V, et al.: Obeticholic acid: a review of clinical applications. Indian J. Gastroenterol. 2020; 39(2): 107–115.\n\nSingh D, Sood A, Bansal D, et al.: Management of intrahepatic cholestasis after liver transplantation. J. Clin. Exp. Hepatol. 2021; 11(4): 365–375.\n\nGeier A, Lohse AW: Treatment of primary biliary cholangitis with obeticholic acid. Hepatology. 2016; 64(2): 492–493.\n\nInoue T, Kondo M, Matsubara T, et al.: The role of bile acid in liver diseases. J. Hepatol. 2020; 72(2): 310–316.\n\nFigiel W, Smoter P, Krasnodębski M, et al.: The utility of early allograft dysfunction components in determining 90-day liver graft survival. Transplant. Proc. 2022; 54(4): 1017–1020. PubMed Abstract \| Publisher Full Text\n\nYounossi ZM, Guyatt G, Kiwi M, et al.: Development of a disease-specific questionnaire to measure health-related quality of life in patients with chronic liver disease. Gut. 1999; 45(2): 295–300. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nThe extended data set of this work is at repository OSF Home. DOI: 10.17605/OSF.IO/JDXBF" }	[ { "id": "355921", "date": "03 Feb 2025", "name": "Palak J. Trivedi", "expertise": [ "Reviewer Expertise Cholestasis", "liver transplantation", "clinical trials." ], "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 rationale for the study is reasonable, but depth of secondary endpoints being captured is limited. Additionally, which adverse events will be captured, in what way, and how will they be recorded?\nThe dose of UDCA is also lower than what would be considered standard practice.\nHow will pruritus be captured? Highly relevant for OCA-treated patients.\nThe pivotal phase III trial of obeticholic acid needs to be cited (Nevens et al. NEJM).\nAs the authors may be aware, the EMA and FDA are in the process of revoking approval for obeticholic acid in Europe and the United States, respectively. Whilst such regulators do not necessarily have jurisdiction in India, how do the authors feel that these decisions will affect access to OCA?\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? No\n\nAre the datasets clearly presented in a useable and accessible format? Partly", "responses": [] } ]	1	https://f1000research.com/articles/14-9
https://f1000research.com/articles/13-1066/v1	17 Sep 24	{ "type": "Brief Report", "title": "Role, function, and expectations of research funding committees: Perspectives from committee members", "authors": [ "Amanda Blatch-Jones", "Cherish Boxall", "Katie Meadmore", "Cherish Boxall", "Katie Meadmore" ], "abstract": "Research funding committees play an integral role in the research funding process, consisting of a range of skills, knowledge, and expertise (e.g., professional, and public contributors). Although there is some evidence that has explored the efficiency and effectiveness of funding committees in terms of the funding process, there is a lack of published evidence about the purpose, role, and function of funding committees, from the perspective of committee members. A subset of survey data from a cohort of six National Institute for Health and Care Research (NIHR) research programmes, exploring the purpose of a funding committee, and the expectations and role of a funding committee member between October 2020 to December 2021. All committee members were eligible to participate in the survey. Using an inductive approach, 50 completed responses (22.5% response rate) were analysed, focusing on the role of a funding committee member and the functions of a funding committee. Participants highlighted seven key areas for the purpose of a funding committee: prioritising and recommending what research to fund (n=36) and assessment of quality (n=24) being the most common responses. Four areas were considered important to the expectations and role of funding committee members, with reviewing, critically appraising, and discussing applications (n=44); and being fair, objective, and unbiased (n=27) being the most common responses. The findings offer a unique insight into committee members’ expectations about the role, purpose and function of a funding committee and their contribution to the funding recommendation process. There was high agreement that the purpose and role of committees and their members was to offer expert advice to make fair, impartial decisions on which research should be prioritised. Exploring the purpose, role, and function of funding committees has relevance and importance for funding organisations seeking to enhance and optimise the decision-making practice of funding committees.", "keywords": [ "Funding committees", "survey", "virtual committees", "grant applications", "peer review", "qualitative", "expectations", "research funding" ], "content": "Introduction\n\nResearch funding committees (also referred to as grant panels or boards) are a core component of the peer review process. Peer review elicits expert, professional and public contributor opinions on whether to recommend or award a research proposal for funding. These committees play an important role for research funding organisations and aid in the funding recommendation and decision-making process. Abdoul et al. reported the role of the committee was to provide a clear summary of the proposal and the associated reviews, to act as an additional assessor and to establish a consensus decision.1 However, there are variations between how funding organisations manage and conduct their funding committees; for example, some require decisions on which research proposals to award funding, whilst others provide recommendations on which proposals to fund, with the final decision laying elsewhere.2 Several funding organisations provide terms of reference and guidance to support committee members’ understanding of their role, purpose and function as a member of a funding committee. For example, the National Institute for Health and Care Research (NIHR) provides information packs on their website for public contributors (https://www.nihr.ac.uk/documents/public-committee-member-roles-information-pack-for-members-of-the-public/26580) and professional committee members (https://www.nihr.ac.uk/researchers/have-your-say-on-our-research/become-a-professional-committee-member.htm).\n\nFunding organisations may also differ in the number of peer review stages that they have, and the type of reviewer required (e.g., external review, funding committee review or community focused review). Some funding organisations use a two-stage application assessment process (stage 1 involves an outline application and stage 2 involves a full application), which involves external peer reviewing and funding committee review. It has been observed that the primary purpose of the committee at stage one is to assess the quality and value of the research question. By comparison, for stage two applications, the role of the committee is to decide which applications to recommend for funding.3\n\nThe purpose and role of the funding committee may also influence the skills needed to make an effective committee member2 and how the expectations and experiences of peer reviewers (including committee members) do not always align with the stated role and purpose of the committee.4,5 Studies that have explored motivation and participation in funding committees demonstrate differences in these expectations.2,3,6–10 For example, Gallo et al. identified that 87-92% of their participants felt that serving as a reviewer on a peer review panel had positively impacted their career, and this was influenced through improvements in writing grant applications and increased exposure to new scientific ideas compared to networking and collaboration opportunities.11 These opportunities can unintentionally be seen to benefit or give greater advantage to those on the panel, potentially introducing the Matthew effect in research funding.12\n\nThere has been considerable evidence addressing and analysing the research funding process, particularly in terms of efficiency and effectiveness of peer review by funding organisations. However, there is still a significant lack of evidence around the funding allocation procedures more generally and there are several reasons for this.13 For example, Guthrie et al. and others have highlighted the sensitivity that surrounds funding organisations’ allocation and recommendations to fund research making this a particularly challenging context to undertake research.13 Despite these challenges, understanding how the purpose, role, and function of funding committees is perceived by committee members themselves is integral to the research funding process, and can contribute to an important research evidence gap.\n\nThis paper addresses this important gap in the evidence and presents data from an online survey conducted as part of a larger study reported elsewhere that explored the role of virtual funding committee meetings for the allocation of NIHR research funding.3 The aim of this paper is to focus on what funding committee members from several of the NIHR programmes consider the purpose of a funding committee to be, and what they consider the expectations of their role as funding committee member are. The insights drawn from the analysis provides a unique opportunity to explore the function and role of funding committees from a committee member perspective, that has not previously been explored in the research funding process literature.\n\n\nMethods\n\nThe survey data presented in this paper is a sub-sample from the main study exploring the NIHR virtual funding committee practices.3 The purpose of the survey (as part of a netnographic study) was to gain further insight and understanding of virtual funding committees and explore the social interactions, expectations, and perceptions in a virtual setting of funding committee practice. The sub-sample addressed here, focuses on funding committee members expectations and role as a member of a committee and the purpose of a funding committee generally, from committee meetings conducted by the NIHR during October 2020 to December 2021 (for a detailed description of the full study, see Blatch-Jones et al., 20233).\n\nAn online survey conducted as part of a netnographic study. This paper only reports on the questions from an online survey about the purpose, expectations and role of funding committees and their members and not specifically related to virtual or face-to-face committee meetings (see data analysis section for the two questions). All NIHR research programme funding committees that took place between October 2020 to December 2021 were eligible to participate in the study. A single NIHR funding committee was treated as a single online community.\n\nAll funding committee members from the research programmes who consented to participate in the netnographic study were sent a link to the survey within four weeks of the virtual funding committee taking place. The survey was open from October 2020 to January 2022. Due to the nature and sensitivity of funding committees and being able to identify respondents, no personal data were collected, unless they offered their contact details to be included in the interview part of the main study. All respondents were asked to complete an online consent form before they could access the survey. The survey was accessible for anyone with an internet connection and was held on a University of Southampton server, using Microsoft Forms. The survey included a total of 16 open and closed-ended questions, including Likert scale responses. We estimated from the pilot of the survey, it would take approximately 15-20 minutes to complete. All respondents were given three weeks to respond to the survey (with two reminders).\n\nFor the analysis reported here, responses from two questions relating to the purpose, role, and function of NIHR funding committees and their members were analysed using an inductive (the data drove the coding of responses) qualitative content analysis approach. The survey data was downloaded from Microsoft Forms and moved to NVivo for analysis. These free text questions were:\n\n1. As a member of a funding committee, what do you consider the purpose of a funding committee to be?\n\n2. What do you consider the expectations and role of a funding committee member to be?\n\nAll responses were coded using Nvivo software (version 1.6.1). (A free alternative Computer-Assisted Qualitative Data Analysis Software (CAQDAS) to using Nvivo is Microsoft Excel or QualCoder that is open source (https://qualcoder.wordpress.com/)) One researcher read through the responses for each question and created initial themes. The themes and associated quotes were then reviewed by a second member of the team, and theme names and associated quotes were then revised through discussion until consensus was met.\n\n\nResults\n\nFrom a potential cohort of 222 invited committee members, 50 responses were received (response rate of 22.5%) from a total of six NIHR research programmes (further details are reported elsewhere3). A high cloaking level was taken across all forms of data analysis, and where there was potential association to a funding committee or individual, verbatim quotes were amended from the online survey open-ended questions.\n\nSeveral explanations describing the purpose of a funding committee were reported by the respondents (see Table 1 for a summary of responses). There was variation in how the respondents explained the purpose and role of a funding committee meeting; however, the majority of respondents thought that prioritisation; assessment of the quality of an application; and prioritising applications that answer questions of importance and relevance were key functions of a funding committee to ensure recommendations of research funding were for the best science that would maximise patient benefit. Although some respondents mentioned the importance of fair and transparent reviews, ‘value for money’ and feedback to applicants, prioritising the need and benefit to the NHS, staff, patients, and the public were seen as key factors during decision-making and form the core focus of discussion at these funding committee meetings.\n\nExploring how respondents described their role as a member of a committee offered insight to what they expected from the role and whether this was comparable to how the purpose of a funding committee was reported (see Table 2 for a summary of responses). By contrast to the purpose of the funding committee as a whole, the majority of responses indicated that participants felt their role was to review and critically appraise applications and to discuss these views at the committee meeting. Several respondents mentioned how the role involves being able to provide an expert opinion and an objective and unbiased review (constructive criticism). Prioritising research applications according to clinical need and patient benefit was seen as important to the committee member role but this was not frequently mentioned compared to the purpose of the committee.\n\n\nDiscussion\n\nThis paper provides a unique insight into the purpose, role, and function of funding committees from the perspective of funding committee members, from several programmes across the NIHR. Exploring these two aspects has provided new insights into what funding committees look for during the decision-making process but also how they see their role and function as part of the funding committee and in addition to funding committees’ terms of reference. Overall, from those that responded to the survey (22.5%) there was high agreement that the purpose and function of committees and their members was to offer expert advice to make fair, impartial decisions on which research should be prioritised for making funding decisions or make recommendations for funding.\n\nReassuringly, the roles of the committee reported were in line with general NIHR assessment criteria of the need for evidence, scientific rigour and value for money, and members perceived the main purpose of the committee meeting itself to be to prioritise and recommend which research to fund by assessing quality and ensuring the research is in areas of genuine need and clinical importance (based on NIHR guidance for committee members). Interestingly, value for money was equally valued the same as providing a fair and transparent review, which is often a feature in the feedback given to applicants.14 Value for money is an important assessment criteria and is noted on the NIHR websites for committee members. In addition, the roles of the committee and its purpose also aligned to the NIHR terms of reference for joining a committee, including public contributors, demonstrating transparency from the NIHR in terms of the assessment criteria on how funding decisions are made by the funding committees.\n\nFuture work could explore the differences between roles, expectations, benefits, and motivations of a committee member to tease these important issues out. Better understanding of these may also allow funding organisations to create better strategies and incentives for recruiting to their committees and may also help to sustain transparency around funding committees and the decision-making recommendations of funded research for the research community as a whole.\n\nThe data reported in this paper come from two questions collected as part of a larger survey. Although the number of responses to the survey was acceptable, especially in the context of the wider study, a response rate of 22.5% is relatively low and the sample contained only those who were members of six NIHR funding committees. Although the sample only included one funding organisation, the findings incorporated several contexts by involving six NIHR programmes.13 The data collection from 2020-2021 could also be seen as a limitation of the findings reported in this paper. Whilst the generalisability of our findings needs to be taken with caution, they provide a unique and reflective account on the purpose, role, and function of committee members that has not otherwise been explored previously. The findings add valuable contributions to the broader evidence around decision-making practices of funding organisations but more importantly addresses a gap in the evidence that goes beyond process measures and measuring effectiveness.13\n\nVariability in responses might have been due to different roles on the funding committee, terms of reference, committee inductions for new members and chairs, how long they have been a member and their professional position within academia, public community, clinical, or social care setting. Whilst the variability could also be an indicator of how funding committees are well briefed on the role and function as a member, it was not possible to explore this in more detail as no personal data were collected as part of the main study. A future study may wish to compare responses by exploring protective characteristics to better understand the different groups of people who join committees, and how these may in turn enhance the decision-making practices of funding committees and look across multiple funding organisations. To build on the existing evidence presented here from the perspective of the NIHR, a broader context from other funding organisations could generate more generalisable findings and comparative data.\n\n\nConclusion\n\nFunding committees play an important role in the decision-making process to fund research, yet there is no published evidence exploring their integral role, purpose, or function as committee members. The findings reported here contribute new evidence and insights about the value and importance of exploring funding committee members expectations and understanding of their role as members but also the function and purpose of a funding committee meeting. Although there was variation across respondents’ responses, there was clear consensus about what matters in order to make informed decision-making recommendations in relation to their role as a committee member but also how, as a collective group of experts, decisions are made that are fair and transparent, offering a considered and balanced discussion to make recommendations on what applications merit funding. Enhancing the current understanding in this space deserves more reflection, particularly from other funding organisations, to determine how and in what circumstances research applications are assessed, validated, and critically appraised to inform the decision-making process, whilst also appreciating the sensitivities around these funding allocation processes.\n\nAll respondents were asked to complete an online consent form before they could access the survey.\n\nConsent from each participant of the survey was gained prior to completing the online survey. The study was approved by the University of Southampton, Faculty of Medicine Ethics Committee (ID 57541, 3rd August 2020). This study was conducted according to the principles expressed in the Declaration of Helsinki.", "appendix": "Data availability\n\nIn line with our ethical approval, the survey data (alongside the interview and observational data) cannot be shared publicly to maintain confidentiality of our participants.\n\nFull details of the survey guide are provided in OSF: Exploring virtual funding committee practices in the allocation of National Institute for Health and Care Research funding: A netnographic study. https://doi.org/10.17605/OSF.IO/RZ6VT. 15\n\nThe project contains the following data:\n\n• Appendix 4 Survey questions.docx\n\nSpecified materials and guides are available under the terms of the CC0 1.0 Public domain dedication (https://creativecommons.org/publicdomain/zero/1.0/). Additional requirements or requests are available from the corresponding author: A.J.Blatch-Jones@southampton.ac.uk) for researchers who meet the criteria for access to the data (e.g., researchers organisational affiliation, intended use of the data, and/or type of data).\n\nThis main study is registered at RoR Registry and Hub (study number 805), accessible here: https://ror-hub.org/study/805/\n\nReporting guidelines: OSF repository. SRQR checklist for “Exploring virtual funding committee practices in the allocation of National Institute for Health and Care Research funding: A netnographic study.” (https://osf.io/rz6vt/ 15 ).\n\nData are available under the terms of the Crreative Commons Attribution International License (CC BY 4.0).\n\n\nAcknowledgements\n\nWe would like to thank all respondents that took part in the online survey. We would also like to thank those who reviewed early draft versions of the article, and Emmanuel Asante for his contribution to observations reported in the main netnography study.\n\n\nReferences\n\nAbdoul H, Perrey C, Amiel P, et al. et al.: Peer review of grant applications: criteria used and qualitative study of reviewer practices.2012.\n\nSteiner Davis ML, Conner TR, Miller-Bains K, et al.: What makes an effective grants peer reviewer? An exploratory study of the necessary skills. PLoS One. 2020; 15(5): e0232327. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBlatch-Jones AJ, Boxall C, Asante E, et al.: Exploring virtual funding committee practices in the allocation of National Institute for Health and Care Research funding: A netnographic study. medRxiv. 2023. 2023.11.17.23298707.\n\nCoveney J, Herbert DL, Hill K, et al.: ‘Are you siding with a personality or the grant proposal?’: observations on how peer review panels function. Res. Integr. Peer Rev. 2017; 2(1): 19. Publisher Full Text\n\nHerbert DL, Graves N, Clarke P, et al.: Using simplified peer review processes to fund research: a prospective study. BMJ Open. 2015; 5(7): e008380. PubMed Abstract \| Publisher Full Text\n\nGallo SA, Carpenter AS, Glisson SR: Teleconference versus face-to-face scientific peer review of grant application: effects on review outcomes. PLoS One. 2013; 8(8): e71693. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGallo SA, Schmaling KB, Thompson LA, et al.: Grant reviewer perceptions of the quality, effectiveness, and influence of panel.2020.\n\nVallée-Tourangeau G, Wheelock A, Vandrevala T, et al.: Peer reviewers’ dilemmas: a qualitative exploration of decisional conflict in the evaluation of grant applications in the medical humanities and social sciences. Humanit. Soc. Sci. Commun. 2022; 9(1): 1–11.\n\nGallo SA, Schmaling KB, Thompson LA, et al.: Grant review feedback: Appropriateness and usefulness. Sci. Eng. Ethics. 2021; 27: 1–20.\n\nAbma-Schouten R, Gijbels J, Reijmerink W, et al.: Evaluation of research proposals by peer review panels: broader panels for broader assessments? Sci. Public Policy. 2023; 50(4): 619–632. Publisher Full Text\n\nGallo SA, Thompson LA, Schmaling KB, et al.: The Participation and Motivations of Grant Peer Reviewers: A Comprehensive Survey. Sci. Eng. Ethics. 2020; 26(2): 761–782. PubMed Abstract \| Publisher Full Text\n\nBol T, de Vaan M , van de Rijt A : The Matthew effect in science funding. Proc. Natl. Acad. Sci. 2018; 115(19): 4887–4890. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGuthrie S, Ghiga I, Wooding S: What do we know about grant peer review in the health sciences? F1000Res. 2017; 6: 6. Publisher Full Text\n\nNational Institute for Health and Care Research (NIHR): Guidance for NIHR Committee Member Development Scheme (CMDS) Participant applicants.2020. Reference Source\n\nBlatch-Jones A; Katie meadmore, cherish boxallAsante E: Exploring Virtual Funding Committee Practices in the Allocation of National Institute for Health and Care Research Funding: A Netnographic Study. OSF. July 31 2024. Publisher Full Text" }	[ { "id": "328098", "date": "23 Oct 2024", "name": "Junwen Luo", "expertise": [ "Reviewer Expertise research evaluation", "impact assessment", "peer review" ], "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 research is drawn from a small size survey which serves a larger research projects involving observations and interviews of the same pool of research funding reviewers. Overall, its research method, objective, and analysis are solid but can provide deeper thoughts and reflections in several ways.\nFirst, the quality and contribution of the paper will be improved if the authors could address how this paper’s conclusion is connected with the other aspects of the larger project as well as with the broad literature. For instance, are those reviewers’ internal expected roles and functions consistent with your observations of the committee meetings and especially the reviewer’s discussions and interactions on how to deal with conflicting views?\nBy the way, some of the literature addressing such topics should be considered especially when you stated ‘there is no published evidence exploring their integral role, purpose, or function as committee members’, see below: Langfeldt, L. (2004). Ref 1 Lee, C.J. (2015) Ref 2 Langfeldt, L. (2001). Ref 3 Luo, J., Feliciani, T., Reinhart, M.,.., & Shankar, K. (2021). Ref 4 Van den Besselaar, P., and Leydesdorff, L. (2009) Ref 5 Holbrook, J.B., and Hrotic, S. (2013) Ref 6 Obrecht, M., Tibelius, K., & D’Aloisio, G. (2007).Ref 7 Pier, E.L., et al. (2018) Ref 8 Feliciani T, Luo J, Shankar, K (2022). Ref 9 Feliciani, T., Morreau, M., Luo, J., Lucas, P., & Shankar, K. (2022)Ref 10 Van den Besselaar, P., Sandström, U. & Schiffbaenker, H. (2018). Ref 11\nSecond, for the analysis and results of content analysis, I would have expected more illustrations on your categorizations of the purposes and expectations which are your key findings, like how you derive those categorizations from the quotes and particular how DIFFERENT are those categorizations from each other? It’s also not very clear how you separated the expectations of the whole committee and of the individual reviewers in the study design, maybe through your survey questions settings?\nThird, an important aspect of the funding committee study, in my eyes, should be discussed which is the prorgamme objectives and target groups. This is found highly important for the committee discussion regarding how to align evaluation criteria into specific programme reviews and how to make fair and transparent grant reviews. A brief introduction of the programme that you studied should be provided in the Methods section. Discussion section could address more clearly how your findings can be connected to the existing literature (see above suggested ones) and make contributions to the broad area of research evaluation and peer review.\nLast but not least, I appreciate very much that the authors shared their research methods and guidelines including the survey questions that this paper made use of. For your reference, I am sharing another grant reviewer survey that you might find relevant and useful https://doi.org/10.6084/m9.figshare.13651058.v1 (Shankar et al., 2021).\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? Not 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": "12984", "date": "02 Jan 2025", "name": "Amanda Blatch-Jones", "role": "Author Response", "response": "In response to your feedback please see the comments below: First, the quality and contribution of the paper will be improved if the authors could address how this paper’s conclusion is connected with the other aspects of the larger project as well as with the broad literature. For instance, are those reviewers’ internal expected roles and functions consistent with your observations of the committee meetings and especially the reviewer’s discussions and interactions on how to deal with conflicting views? Response: Thank you. We have extended the discussion to take account of your feedback. By the way, some of the literature addressing such topics should be considered especially when you stated ‘there is no published evidence exploring their integral role, purpose, or function as committee members’, see below: Langfeldt, L. (2004). Ref 1 Lee, C.J. (2015) Ref 2 Langfeldt, L. (2001). Ref 3 Luo, J., Feliciani, T., Reinhart, M.,.., & Shankar, K. (2021). Ref 4 Van den Besselaar, P., and Leydesdorff, L. (2009) Ref 5 Holbrook, J.B., and Hrotic, S. (2013) Ref 6 Obrecht, M., Tibelius, K., & D’Aloisio, G. (2007).Ref 7 Pier, E.L., et al. (2018) Ref 8 Feliciani T, Luo J, Shankar, K (2022). Ref 9 Feliciani, T., Morreau, M., Luo, J., Lucas, P., & Shankar, K. (2022)Ref 10 Van den Besselaar, P., Sandström, U. & Schiffbaenker, H. (2018). Ref 11 Response: Thank you for these additional references. Where appropriate we have added these references to explain how previous research has focused on specific areas, although there is limited evidence from committee members about what they perceive their role to be and the function of committee members. This particular focus is different to the assessment and process of peer review, review scores and decision-making through external peer review or panel/grant meetings. Second, for the analysis and results of content analysis, I would have expected more illustrations on your categorizations of the purposes and expectations which are your key findings, like how you derive those categorizations from the quotes and particular how DIFFERENT are those categorizations from each other? It’s also not very clear how you separated the expectations of the whole committee and of the individual reviewers in the study design, maybe through your survey questions settings? Response: Thank you for your feedback. The survey data was at an individual level, and it is only this data presented in the brief report, as the full survey as part of the netnographic study is reported elsewhere. Under data analysis we explain the two questions used in this article, and the process we used. Third, an important aspect of the funding committee study, in my eyes, should be discussed which is the programme objectives and target groups. This is found highly important for the committee discussion regarding how to align evaluation criteria into specific programme reviews and how to make fair and transparent grant reviews. A brief introduction of the programme that you studied should be provided in the Methods section. Discussion section could address more clearly how your findings can be connected to the existing literature (see above suggested ones) and make contributions to the broad area of research evaluation and peer review. Response: All NIHR programmes were invited to participate and six NIHR programmes participated in the netnographic study. To make this clearer all six NIHR programmes have been added in the results section for transparency purposes. Given the six NIHR programmes have different health focuses and aims, we have not included additional details or analysis on this. However, alignment to overall NIHR aims is discussed in paragraph 2 of the discussion. We have revised the discussion section based on your feedback. As per brief report guidelines, a conclusion section is optional, but we have included this to provide context and future considerations for the reader. Last but not least, I appreciate very much that the authors shared their research methods and guidelines including the survey questions that this paper made use of. For your reference, I am sharing another grant reviewer survey that you might find relevant and useful https://doi.org/10.6084/m9.figshare.13651058.v1 (Shankar et al., 2021). Response: Thank you for sharing this survey on peer review." } ] }, { "id": "341502", "date": "25 Nov 2024", "name": "Santiago Vasco-Morales", "expertise": [ "Reviewer Expertise porque Pediatrics", "neonatology", "medical 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\nSummary of the Article This study explores the roles, expectations, and functions of research funding committees, focusing on the perspectives of committee members from six National Institute for Health and Care Research (NIHR) programs. Using data collected via an online survey, the authors identify key themes in the purpose of funding committees, such as prioritizing high-quality research, providing fair and unbiased reviews, and aligning decisions with public and clinical needs. The results contribute to the understanding of funding committee dynamics and highlight gaps in current practices, particularly regarding transparency and efficiency in decision-making. The study design employs qualitative content analysis to analyze responses, providing insights into the expectations and experiences of committee members. While the methodology is sound, several areas for improvement have been identified to enhance clarity, reproducibility, and scientific rigor. Detailed Review The following is an expanded evaluation of the article based on the key review questions:\n1. Is the work clearly and accurately presented, and does it cite the current literature? Answer: Partly Criticisms:\nIntegration with Literature: While the article cites relevant studies, it does not adequately connect its findings to the broader body of literature on peer review and funding committees. Inconsistent Claims: The claim that there is no published evidence on the roles and expectations of funding committees is too broad. Relevant studies (e.g., Langfeldt, 2004; Lee, 2015) are mentioned in the peer review comments but not included in the discussion.\nRecommendations: Incorporate additional references from the literature on peer review and funding committee practices to contextualize findings andsubstantiate claims. Expand the discussion section to critically compare findings with existing evidence, highlighting similarities and differences.\n2. Is the study design appropriate, and is the work technically sound? Answer: Partly Criticisms:\nSurvey Design: The survey questions are not fully described, limiting understanding of how data was collected. Role Distinction: The study does not clearly distinguish between the roles of individual committee members and the collective purpose of the committee. Low Response Rate: A 22.5% response rate may introduce bias and limit generalizability.\nRecommendations: Provide a detailed breakdown of the survey design, including sample questions and their alignment with study objectives. Discuss the potential impact of the low response rate and how it may affect the representativeness of findings. Clearly separate analysis and discussion of individual versus collective roles.\n3. Are sufficient details of methods and analysis provided to allow replication by others? Answer: Partly Criticisms:\nLack of Transparency in Coding: Details on the coding process, inter-coder reliability, and how themes were finalized are not provided. Limited Survey Details: The full set of survey questions is not included, and the categorization of responses lacks illustrative examples.\nRecommendations: Describe the coding process in detail, including how consistency was ensured (e.g., inter-coder reliability metrics). Provide an appendix or supplementary materials with the full survey questionnaire. Include more examples of raw data (e.g., anonymized quotes) to demonstrate how responses were categorized.\n4. If applicable, is the statistical analysis and its interpretation appropriate? Answer: Not applicable Since the study focuses on qualitative content analysis, statistical analysis is not a core component. However, descriptive statistics, such as response frequencies, are used appropriately.\n5. Are all the source data underlying the results available to ensure full reproducibility? Answer: Partly Criticisms:\nSupplementary Data: Additional resources, such as detailed coding frameworks or categorized data, are not provided.\nRecommendations: Share anonymized or synthesized data to the extent possible, ensuring confidentiality. Provide coding frameworks and categorized themes as supplementary materials to support reproducibility.\n6. Are the conclusions drawn adequately supported by the results? Answer: Partly Criticisms:\nSuperficial Interpretation: While the conclusions are consistent with the results, they do not sufficiently explore broader implications or practical applications. Generality of Claims: Conclusions could overgeneralize findings from a single funding organization to the broader context of research funding.\nRecommendations: Deepen the discussion of how findings apply to funding committee practices and align with existing research. Acknowledge limitations in generalizability and provide recommendations for future studies across diverse funding contexts.\nPoints That Must Be Addressed To make the article scientifically sound, the following points must be addressed:\nClarify Methods and Analysis:\nInclude detailed descriptions of the survey design and coding process. Provide more illustrative examples of data categorization.\n\nImprove Literature Integration:\nExpand the discussion to connect findings with existing studies on peer review and funding committees.\n\nAcknowledge Limitations:\nDiscuss the impact of the low response rate on findings. Clearly state the limitations in generalizing results beyond the NIHR context.\n\nProvide Additional Data:\nShare supplementary materials (e.g., survey questions, coding frameworks) to enhance transparency and reproducibility.\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? Partly", "responses": [ { "c_id": "12985", "date": "02 Jan 2025", "name": "Amanda Blatch-Jones", "role": "Author Response", "response": "Thank you for your feedback. Please see the comments below: Integration with Literature: While the article cites relevant studies, it does not adequately connect its findings to the broader body of literature on peer review and funding committees. Thank you for your comment. We have revised the literature and added additional references. Inconsistent Claims: The claim that there is no published evidence on the roles and expectations of funding committees is too broad. Relevant studies (e.g., Langfeldt, 2004; Lee, 2015) are mentioned in the peer review comments but not included in the discussion. We have revised the content of the article to address this point. Due to the limitations of brief reports (2500 words) we have briefly mentioned the relevant literature and have made it clearer that this article only focuses on the responses from the survey about how funding committee members answered two questions on the purpose and expectation and role of funding committee meetings. As per brief report guidelines, a conclusion section is optional, but we have included this to provide context and future considerations for the reader. Recommendations: Incorporate additional references from the literature on peer review and funding committee practices to contextualize findings and substantiate claims. Expand the discussion section to critically compare findings with existing evidence, highlighting similarities and differences. Thank you. We have addressed your comment (and others) in relation to the wider literature and added in some additional text to the discussion. Due to the nature of brief papers, we did not go in depth as the survey complemented the wider netnographic study. As per brief report guidelines, a conclusion section is optional, but we have included this to provide context and future considerations for the reader. Survey Design: The survey questions are not fully described, limiting understanding of how data was collected. Only two survey questions are related to this article (both reported under data analysis). Both questions are provided in the article. Links to the main study are provided. Data collection is included in the article, including timeframe and process of the complete survey. Role Distinction: The study does not clearly distinguish between the roles of individual committee members and the collective purpose of the committee. In this brief paper, the two survey questions that are reported explore this distinction. Q1 relates to the funding committee and Q2 to the individual, and Table 1 and 2 respectively show the findings for how the roles of the committee and the individuals are perceived by the funding committee members. We have made this distinction clearer in the discussion and added additional text comparing the role of committee members to the role of committees. In addition, due to the sensitivity of the data and being able to identify members of committees and funding programmes, as reported in the article a high cloaking level was used to ensure anonymity of all committee members. However, this does not impact the results due to being inclusive of all members and all members having equal contribution to the analysis and results. Low Response Rate: A 22.5% response rate may introduce bias and limit generalizability. This is covered under strengths and limitations section. Recommendations: Provide a detailed breakdown of the survey design, including sample questions and their alignment with study objectives. Discuss the potential impact of the low response rate and how it may affect the representativeness of findings. Clearly separate analysis and discussion of individual versus collective roles. This is reported in the brief paper but due to the 2500-word limit, providing the full details of the methods is not feasible, and it is reported elsewhere. We have reported the low response rate under the discussion section. It is not possible to report findings at an individual level, as reported above. However, we have made the distinction of the findings clearer in the discussion and added additional text comparing the role of committee members to the role of committees. Lack of Transparency in Coding: Details on the coding process, inter-coder reliability, and how themes were finalized are not provided. A summary is provided in the brief paper. As stated, full details are reported elsewhere along with the wider themes of the main study. The purpose of the brief paper is to highlight the findings from two questions, as the current literature lacks evidence from real-world, first-hand understanding of what funding committee members perceive their role and function to be as a committee member and what the function and expectation is of a funding committee panel. As detailed, most of the literature is based on modelling, data, peer review, grant proposals, scoring etc. These studies do not address the two questions asked in the survey. By understanding the perceptions from committee members directly can help funding organisations enhance their guidance and consider where improvements are needed. Limited Survey Details: The full set of survey questions is not included, and the categorization of responses lacks illustrative examples. These are reported in the main study and in the OSF repository as stated under the data availability statement. All materials are provided but F1000Research policy does not allow supplementary materials. Recommendations: Describe the coding process in detail, including how consistency was ensured (e.g., inter-coder reliability metrics). Provide an appendix or supplementary materials with the full survey questionnaire. Include more examples of raw data (e.g., anonymized quotes) to demonstrate how responses were categorized. Supplementary Data: Additional resources, such as detailed coding frameworks or categorized data, are not provided. All relevant materials that comply with ethical procedures are provided in the OSF repository and linked to the main article as well. Recommendations: Share anonymized or synthesized data to the extent possible, ensuring confidentiality. Provide coding frameworks and categorized themes as supplementary materials to support reproducibility. This is available on request as stated under data availability, to comply with ethical procedures, and to protect the identity of all participants. This is not a requirement for publication due to the sensitive nature of the data. However, we clearly state it is available on request. Superficial Interpretation: While the conclusions are consistent with the results, they do not sufficiently explore broader implications or practical applications. We have revised sections of the brief paper to reflect your comments and feedback. As per brief report guidelines, a conclusion section is optional, but we have included this to provide context and future considerations for the reader. Generality of Claims: Conclusions could overgeneralize findings from a single funding organization to the broader context of research funding. Thank you for your comment. This is noted under strengths and limitations. However, as with previous literature, most of the research is with one funder (several references listed demonstrate this point), and the context of each funding organisation will always be a limitation for all research studies. We have already stated this within the article. Recommendations: Deepen the discussion of how findings apply to funding committee practices and align with existing research. Acknowledge limitations in generalizability and provide recommendations for future studies across diverse funding contexts. The netnographic study goes into depth on the aspects you mention. However, as noted this is a brief paper and is limited to 2500 words. Also, as per brief report guidelines, a conclusion section is optional, but we have included this to provide context and future considerations for the reader. The generalisability aspect is already noted in the article but we have extended a sentence to cover your comment. Points That Must Be Addressed: To make the article scientifically sound, the following points must be addressed: Clarify Methods and Analysis: Include detailed descriptions of the survey design and coding process. Provide more illustrative examples of data categorization. Improve Literature Integration: Expand the discussion to connect findings with existing studies on peer review and funding committees. Acknowledge Limitations: Discuss the impact of the low response rate on findings. Clearly state the limitations in generalizing results beyond the NIHR context. Provide Additional Data: Share supplementary materials (e.g., survey questions, coding frameworks) to enhance transparency and reproducibility. Please also see more detailed comments above that relate to these points and how we have addressed them. As described in the paper, please refer to the main study and the OSF repository, for a more comprehensive description of the whole survey design and analysis. We have included additional references throughout the paper. However, due to the nature of brief papers, we did not go in depth on these. Low response rate and generalisability are now covered under the strengths and limitations section. All details and supplementary material are provided in the main study and the OSF repository. F1000Research policy does not allow supplementary materials. Thank you for your suggestions and detailed feedback." } ] } ]	1	https://f1000research.com/articles/13-1066
https://f1000research.com/articles/14-6/v1	02 Jan 25	{ "type": "Case Report", "title": "Case Report: Case report: a rare case of dentigerous cyst in 1-year-old patient.", "authors": [ "Sami Alshehri" ], "abstract": "One of the most common benign lesions is dentigerous cyst, which is commonly associated with unerupted tooth crowns. Dentigerous cysts in infants may manifest as swelling or a firm mass in the oral cavity. Such cysts arise from dental follicles, and their development can lead to dental complications such as tooth displacement, root resorption of the adjacent teeth, or infection if not treated carefully. Enucleation is the method of choice for the treatment of such lesions. In the current case report, a rare age presented with 1 a old patient was referred to oral and maxillofacial clinics. In the case of right facial swelling and pain for five days ago, intra- and extra-oral examinations were performed. Computed Tomography (CT) demonstrated a well-defined expansile lytic lesion, and parents consented and agreed to proceed with the surgery option. Enucleation was performed surgically, and the patient was followed up for up to 8 months. Significant improvement was noted, with no further complications or recorded complaints.", "keywords": [ "Dentigerous cyst", "radiograph", "dental cyst", "pediatric patients", "surgery" ], "content": "Introduction\n\nDentigerous cysts are considered one of the most common benign developmental lesions faced by maxillofacial surgeons during their training and practice.1,2 Historically, an early report by Gabell, James and Payne documented 84 dentigerous cysts in 1914. The authors reported that these cysts present with the following typical features: asymptomatic, accidental finding, associated with supernumerary or failure of tooth eruption in the maxilla or mandible.3 These manifestations remain the same in the current literature, with more studies reporting the hallmark of the cyst lining attached to the cementoenamel junction.2,4\n\nThe authors agree that dentigerous cysts are the second most common odontogenic cyst.5–7 The pathogenesis of dentigerous cyst formation relies on fluid accumulation between the reduced enamel epithelium and tooth, which is released from compromised follicular veins around the tooth.8,9 In terms of age and sex prevalence, Zhang et al. analyzed 2082 dentigerous cysts diagnosed in British Columbia, Canada. The authors found that dentigerous cysts are more likely to be encountered in male patients in their twenties and thirties. These observations are considered rare in younger and older age groups.2\n\nRadiographically, dentigerous cysts in most cases show well-defined radiolucent lesions associated with an unerupted tooth. However, some instances demonstrated ill-defined borders if they get infected10,11 In terms of surgical management, enucleation of the cyst is usually the method of choice. However, marsupialization is considered a surgical modality, especially when the cysts are adjacent to vital structures. This technique induces cyst decompression, which aids total enucleation in the second surgery with fewer postoperative complications.12 We report a rare case of dentigerous cyst in a 1-year-old boy. To the best of our knowledge, this is the second documented case in the literature since the first report by Suresh et al. in 2011.13\n\n\nCase report\n\nA 1-year-old boy medically fit and pain was referred to the Oral and Maxillofacial Surgery Department of King Fahad Hospital University (KFHU), Khobar, Saudi Arabia as case right facial swelling and pain with history of 5 days. General examination showed that all vital signs were normal, with no signs of dehydration or dysphagia. Extraoral examination revealed a right buccal swelling that was consistent with good mouth opening. Intraoral examination revealed a bluish color on the right posterior mandibular ridge with no vestibular swelling.\n\nComputed tomography (CT) demonstrated a well-defined expansile lytic lesion in the posterior body of the mandible involving an unerupted tooth measuring approximately 1.6 × 1.3 cm (Figure 1). The radiographic appearance is highly suggestive of a dentigerous cyst or inflammatory cyst based on histopathological correlation.\n\nThe parents were reassured, and the treatment plan options were explained in detail. The parents agreed to proceed with surgical enucleation of the lesion under general anesthesia. The patient was admitted to the KFHU, and consent was signed. Under all aseptic conditions, the patient was prepared using povidone-iodine solution, and the patient was covered with a sterile towel. Good exposure of the lesion was achieved by raising a full mucoperiosteal flap, enucleation of the entire cyst along with the associated tooth, and bleeding from all margins of the cavity lesion. The specimen was placed in a labelled container for histopathological examination. Irrigation with normal saline and hemostasis were achieved, and the surgical site was closed with 3/0 Vicryl sutures.\n\nA gross examination of the specimen consists of tooth and cyst lining (1.5 cm × 1.5 cm with a wall thickness of 0.2). Microscopic analysis revealed a cyst wall lined by non-keratinized squamous epithelium with intraepithelial neutrophil infiltration and pus formation. This finding is consistent with an inflamed dentigerous cyst. The patient was followed on a regular basis for up to 8 months after the surgery. The parents were informed of the outcomes of the surgical operation. The surgical site was intact, swelling was reduced significantly, and no complaints were reported. Three-month post-operative CT) scan was requested and reviewed. Compared with the previous CT, no evidence of bone destruction, pathology, or soft tissue abnormalities was observed or noted (Figure 2).\n\n\nDiscussion\n\nDentigerous cysts are a major area of interest in the field of oral and maxillofacial surgery as they are the second most common developmental cysts. Therefore, a deeper understanding of different presentations can play a crucial role in providing the optimum care to the patients.14,15 Several studies have explored the pathophysiological processes that underlie dentigerous cysts. Benn and Altini (1996) suggested that three possible mechanisms could initiate the pathological process based on the source of the inflammation. First, the radicular cyst in the pathway of eruption of the successor’s tooth was left in situ following exfoliation or extraction of primary teeth. The second is the presence of a nonvital tooth. Finally, there are other sources of inflammation in the jaw.16 However, as mentioned previously, the pressure from fluid accumulation between the reduced enamel epithelium and unerupted tooth seems to be the most common pathophysiological theme of dentigerous cysts.8,9,12,17 In our case, the pathological process mirrored the generally agreed pathological pattern of a dentigerous cyst.\n\nA dentigerous cyst is usually discovered incidentally during routine dental examinations.9–11 Our patient presented with right facial swelling and pain, which could be attributed to the fact that fluid accumulation over time results in expansion, leading to facial swelling accompanied by pain. No intervention to break the expansion pressure could result in a negative effect on the surrounding vital structures such as the inferior alveolar nerve and maxillary sinus.12,17 Authors are in agreement that dentigerous are commonly associated with unerupted mandibular third molars followed by maxillary canines.2,10,12 This finding broadly supports the current literature on the most commonly impacted teeth.12,17,18\n\nIn terms of age range, despite global population variations, the current evidence suggests that the second and third decades of life are the peak incidences of dentigerous cysts at approximately 23% and 20%, respectively. Notably, a considerable percentage of cases were reported in the fourth, fifth, and sixth cases.2 Several reports in the literature were published in the first decade. Hedge reported a case of 9 years old boy with left mandible swelling and pain,19 Dave reported two cases of 9 years old boy with asymptomatic swelling and 7 years old boy who was discovered during a routine dental examination,12 Deepa documented a case of 10 years old girl with a history of difficulty in mastication due to a dentigerous cyst in her lower right jaw20 and McKinney also reported 4 years old child with asymptomatic facial swelling related to dentigerous cyst.21 The first documented case in one a old boy was reported by Suresh et al. in 2011. The patient presented with left mandibular angle swelling associated with a history of pain, difficulty in swallowing, and an obliterated buccal sulcus.13 Our case is the second documented case after Suresh et al. at this very young age. However, the case we reported differed in some clinical manifestations, as our patient presented with a history of rapid right facial swelling and pain. No difficulty in swallowing or buccal sulcus obliteration. This difference illustrates the benefit of reporting such cases to aid and build up evidence-based practice, as this will result in positive reflections on the care that we provide for our patients.\n\nA unilateral and single presentation is considered a classic demonstration of a dentigerous cyst, similar to our case. Nevertheless, multiple dentigerous cysts can be observed in patients with basal cell nevus, mucopolysaccharidoses, and cleidocranial dysplasia syndromes.20 However, few cases of bilateral dentigerous cysts have been reported in the literature in non-syndromic patients.22 In terms of radiographic presentation, based on a CT scan, our case showed a well-defined radiolucent lesion associated with an unerupted tooth which is in alignment with the characteristic appearance of a dentigerous cyst.12,13,20\n\nSurgical treatment techniques for dentigerous cysts include enucleation and marsupialization. The surgical decision is formulated based on several factors, such as the patient’s medical history, location and size of the lesion, and the facilities where the surgery will take place.20 Dave et al. suggested that a multidisciplinary approach is required for such cases to establish a fully comprehensive treatment plan. The authors acknowledge that enucleation is the gold standard surgical option for most dentigerous cyst cases. However. Marsupialization followed by complete removal of the cyst should be considered in cases of close proximity to vital structures.12 The recurrence of dentigerous cysts is not a concern for most patients.11,13 Our patient underwent complete removal of the cyst under general anesthesia with the associated tooth to eliminate the need for second-stage surgery, as well as to improve the patient and family’s quality of life by reducing the required time in the hospital.\n\n\nEthical approval\n\nNot required.\n\n\nConsent\n\nWritten informed consent for publication was obtained from the patient’s parent. Additionally, we confirm that we have obtained permission to use radiographic images from the patients’ parents included in this presentation.", "appendix": "Data availability statement\n\nNo data are associated with this article.\n\n\nReferences\n\nAgrawal M, Sharma S, Gupta K: Multiple teeth in a single dentigerous cyst follicle: a perplexity. Ann. Maxillofac. Surg. 2011; 1(2): 187–189. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZhang LL, Yang R, Zhang L, et al.: Dentigerous cyst: A retrospective clinicopathological analysis of 2082 dentigerous cysts in British Columbia, Canada. Int. J. Oral Maxillofac. Surg. 2010; 39(9): 878–882. PubMed Abstract \| Publisher Full Text\n\nGabell D, James W, Payne J: The Report on Odontomes: The Committee Appointed by the British Dental Association. London, England: John Bale, Sons and Danielsson, Ltd.; 1914; p. 133.\n\nHasan S, Ahmed SA, Reddy LB: Dentigerous cyst in association with impacted inverted mesiodens: Report of a rare case with a brief review of literature. Int. J. Appl. Basic Med. Res. 2014; 4(Suppl 1): S61–S64. PubMed Abstract \| Publisher Full Text\n\nJohnson NR, Gannon OM, Savage NW, et al.: Frequency of odontogenic cysts and tumors: A systematic review. J. Investig. Clin. Dent. 2014; 5(1): 9–14. PubMed Abstract \| Publisher Full Text\n\nOchsenius G, Escobar E, Godoy L, et al.: Analysis of 2,944 cases in Chile. Med. Oral Patol. Oral Cir. Bucal. 2007; 12(2): 85–91.\n\nKambalimath DH, Kambalimath HV, Agrawal SM, et al.: Prevalence and distribution of odontogenic cysts in the Indian population: A 10-year retrospective study. J. Maxillofac. Oral Surg. 2014; 13: 10–15. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBrowne R: Pathogenesis of odontogenic cysts: review. J. Oral Pathol. Med. 1975; 4(1): 31–46. Publisher Full Text\n\nMortha N, Uppala D: Pathogenesis of odontogenic cysts. Oral Maxillofac. Pathol. J. 2021; 12(1).\n\nNeville BW, Damm DD, Allen CM, et al.: Oral and maxillofacial pathology. Elsevier Health Sciences; 2015.\n\nZerrin E, Husniye D: Peruze C: Dentigerous cysts of the jaws: Clinical and radiological findings of 18 cases. J. Oral Maxillofac. Radiol. 2014; 2(3): 77–81. Publisher Full Text\n\nDave M, Clarke L, Grindrod M, et al.: Adapting trees for dentigerous cysts in pediatric and adult patients: a case series. Oral Surg. 2021; 14(3): 277–284. Publisher Full Text\n\nSuresh R, Janardhanan M, Joseph AP, et al.: A rare case of a dentigerous cyst in a one-year-old child: The earliest reported occurrence. Head Neck Pathol. 2011; 5: 171–174. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMotamedi MH, Talesh KT: Management of extensive dentigerous cysts. Br. Dent. J. 2005; 198(4): 203–206. PubMed Abstract \| Publisher Full Text\n\nEwbank L, El-Nashar R, Middlefell L: Spontaneous regression of a dentigerous cyst associated with an impacted mandibular canine: A case report. Oral Surg. 2019; 12(1): 48–50. Publisher Full Text\n\nBenn A, Altini M: Dentigerous cysts of inflammatory origin: a clinicopathologic study. Oral Surg. Oral Med. Oral Pathol. Oral Radiol. Endod. 1996; 81(2): 203–209. Publisher Full Text\n\nShetty RM, Dixit U: Dentigerous cyst of inflammatory origin. Int. J. Clin. Pediatr. Dent. 2010; 3(3): 195–198. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSuresh R, Janardhanan M, Joseph AP, et al.: A rare case of a dentigerous cyst in a one-year-old child: The earliest reported occurrence. Head Neck Pathol. 2011; 5: 171–174. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHegde R, Khare S, Devrukhkar V: Dentigerous cyst in a young child: clinical insight and a case report. J. Indian Soc. Pedod. Prev. Dent. 2013; 31(3): 209–211. PubMed Abstract \| Publisher Full Text\n\nDeepa KK, Jannu A, Kulambi M, et al.: A case of dentigerous cyst in a pediatric patient: an insight into differential diagnostic entities. Adv. Oral Maxillofac. Surg. 2021; 3: 100130. Publisher Full Text\n\nMcKinney SL, Lukes SM: Dentigerous cyst in a young child: A case report. Can. J. Dent. Hyg. 2021; 55(3): 177–181. PubMed Abstract\n\nFreitas DQ, Tempest LM, Sicoli E, et al.: Bilateral dentigerous cysts: Review of the literature and report of an unusual case. Dentomaxillofac. Radiol. 2006; 35(6): 464–468. PubMed Abstract \| Publisher Full Text" }	[ { "id": "356556", "date": "08 Jan 2025", "name": "Carlos Cobo Vázquez", "expertise": [ "Reviewer Expertise Oral 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\nThank you for your submission.\nThe article presented describes the second case of a young child with the appearance of a dentigerous cyst. The presentation of this case contributes to the expansion of scientific knowledge since, as the authors describe, the highest incidence of these cysts occurs in the lower third molars.\n\nHowever, despite the value of the work, it presents a series of limitations that prevent the reader from understanding the pathology. It would be of great value to provide previous clinical, intra-operative and follow-up photographs of the patient. We understand the limitation to photograph a young patient, but radiological tests are provided without difficulty, and by performing the surgical treatment under general anesthesia an adequate record could have been made.\n\nOn the other hand, regarding therapeutic alternatives, more conservative options could be considered regarding the molar germ. Neither in the description of the case nor in the discussion, the reason for ruling out alternatives such as cystectomy, marsupialization or drainage, while maintaining the molar germ, is mentioned. It would also be interesting to provide information on the success rates of these more conservative treatments\nYour work is interesting. However, readers are missing previous clinical photos of the pathology, as well as photographs of the surgical intervention, the pathological anatomy and the subsequent follow-up.\nIf this could be corrected it would be of great interest due to the low frequency of the 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? 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? Partly", "responses": [] } ]	1	https://f1000research.com/articles/14-6
https://f1000research.com/articles/13-922/v1	13 Aug 24	{ "type": "Data Note", "title": "A guide to selecting high-performing antibodies for Huntingtin (UniProt ID: P42858) for use in western blot, immunoprecipitation, and immunofluorescence", "authors": [ "Rebeka Fanti", "Riham Ayoubi", "Charles Alende", "Maryam Fotouhi", "Sara González Bolívar", "Renu Chandrasekaran", "Kathleen Southern", "Aled M. Edwards", "Rachel J. Harding", "Carl Laflamme", "NeuroSGC/YCharOS/EDDU collaborative group", "ABIF consortium", "Rebeka Fanti", "Riham Ayoubi", "Charles Alende", "Maryam Fotouhi", "Sara González Bolívar", "Renu Chandrasekaran", "Kathleen Southern", "Aled M. Edwards", "Rachel J. Harding" ], "abstract": "Huntingtin encodes a 3144 amino acid protein, with a polyglutamine repeat tract at the N-terminus. Expansion of this repeat tract above a pathogenic threshold of 36 repeats is the causative mutation of Huntington's disease, a neurodegenerative disorder characterized by loss of striatal neurons. Here we have characterized twenty Huntingtin commercial antibodies for western blot, immunoprecipitation, and immunofluorescence using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. These studies are part of a larger, collaborative initiative seeking to address antibody reproducibility issues by characterizing commercially available antibodies for human proteins and publishing the results openly as a resource for the scientific community. While use of antibodies and protocols vary between laboratories, we encourage readers to use this report as a guide to select the most appropriate antibodies for their specific needs.", "keywords": [ "UniProt ID P42858", "HTT", "Huntingtin", "antibody characterization", "antibody validation", "western blot", "immunoprecipitation", "immunofluorescence" ], "content": "Introduction\n\nHuntington’s Disease (HD) is a neurodegenerative disorder inherited in an autosomal dominant manner, presenting with a spectrum of progressive motor, cognitive, and psychological impairments, typically with adult-onset of symptoms.1 Although the HD causative gene, HTT, was discovered over three decades ago, there are still no disease-modifying treatments available for patients, and progress unpicking the molecular pathology of the disease remains slow.2\n\nHD arises from a heterozygous expansion mutation of the trinucleotide CAG repeat tract in exon 1 of HTT, located on chromosome 4, above a critical threshold of ~36 repeats. This mutation results in expansion of the polyglutamine stretch at the N-terminus of the 3144 amino acid Huntingtin protein. Huntingtin functions as a scaffold protein, engaging in extensive protein-protein interactions,3 forming various multi-protein complexes to carry-out its diverse array of functions. Modulation of this interaction network by the polyglutamine expansion contributes to degeneration within the central nervous system, affecting medium spiny neurons at the onset of disease.4,5\n\nThe low expression level, complex interactome and large size of the 348 kDa Huntingtin protein have given rise to technical challenges which have hindered precise determination of its molecular function, or how this is altered in disease. In particular, the use of different Huntingtin antibodies by scientists in the HD research community, often mapping to structurally distant epitopes, can yield different or even conflicting results, further conflating interrogation of this protein.6–8\n\nThis research is part of a broader collaborative initiative in which academics, funders and commercial antibody manufacturers are working together to address antibody reproducibility issues by characterizing commercial antibodies for human proteins using standardized protocols, and openly sharing the data.9–11 Here we evaluated the performance of twenty commercial antibodies for Huntingtin for use in western blot, immunoprecipitation, and immunofluorescence, enabling biochemical and cellular assessment of Huntingtin properties and function. The platform for antibody characterization used to carry out this study was endorsed by a committee of industry and academic representatives. It consists of identifying human cell lines with adequate target protein expression and the development/contribution of equivalent knockout (KO) cell lines, followed by antibody characterization procedures using most commercially available antibodies against the corresponding target protein. The standardized consensus antibody characterization protocols are openly available on Protocol Exchange (DOI: 10.21203/rs.3.pex-2607/v1).12\n\nThe authors do not engage in result analysis or offer explicit antibody recommendations. A limitation of this study is the use of universal protocols - any conclusions remain relevant within the confines of the experimental setup and cell line used in this study. Our primary aim is to deliver top-tier data to the scientific community, grounded in Open Science principles. This empowers experts to interpret the characterization data independently, enabling them to make informed choices regarding the most suitable antibodies for their specific experimental needs. Guidelines on how to interpret antibody characterization data found in this study are featured on the YCharOS gateway.13\n\n\nResults and discussion\n\nOur standard protocol involves comparing readouts from WT (wild type) and KO cells.14,15 The first step is to identify a cell line(s) that expresses sufficient levels of a given protein to generate a measurable signal. To this end, we examined the DepMap transcriptomics database to identify all cell lines that express the target at levels greater than 2.5 log2 (transcripts per million “TPM” + 1), which we have found to be a suitable cut-off (Cancer Dependency Map Portal, RRID:SCR_017655). We generated an HTT KO line in DMS 53 as it expresses the endogenous HTT transcript at 6.1 log2 (TPM+1), which is above the average range of cancer cell lines analyzed. A commercial HAP1 HTT KO is also available; HAP1 expresses HTT at 3.7 log2 (TPM+1) RNA level. A HEK293T HTT KO cell line has been developed and used elsewhere16 (Table 1). All three cell line backgrounds were evaluated by western blot using a high-performing Huntingtin antibody detected in Figure 1. DMS 53 was identified as the most suitable cell line (Figure 2), which can be explained by its high expression of the HTT transcript compared to other two cell lines. Thus, DMS 53 WT and KO cell lines were generated and used to evaluate the antibodies in all applications.\n\nLysates of DMS 53 (WT and HTT KO) were prepared and 30 μg of protein were processed for western blot with the indicated Huntingtin antibodies. Tris-Glycine 4-20% gels were used for SDS-PAGE. The Ponceau stained transfers of each blot are presented to show equal loading of WT and KO lysates and protein transfer efficiency from the acrylamide gels to the nitrocellulose membrane. Antibody dilutions were chosen according to the recommendations of the antibody supplier. Exceptions were given for antibodies ab109115, ab45169, and MA5-41256 which were titrated as the signal was too weak when following the supplier’s recommendations. Antibody dilution used: ab109115 at 1/2000, ab45169 at 1/5000, ABCD_AG854 at 1/10, ABCD_AG855 at 1/10, A19064 at 1/1000, 5656** at 1/1000, CH03023* at 1/1000, MW1-S* at 1/10, MW3-S* at 1/10, MW4-S* at 1/10, MW5-S* at 1/10, MW6-S* at 1/10, MW7-S* at 1/10, MW8-S* at 1/10, GTX132433 at 1/500, GTX638832 at 1/500, 710695 at 1/200, MA3-040* at 1/1000, MA5-16703* at 1/500, MA5-41256** at 1/2000. Predicted band size: 347 kDa. Monoclonal antibody, Recombinant antibody.\n\nNote: MW1-S, MW3-S, MW4-S, MW5-S, MW6-S, MW7-S* and MW8-S* are expected to only recognize an altered conformation of the polyQ domain generated as the polyQ domain of Huntingtin increases in length.\n\nLysates of WT and HTT KO in DMS 53, HEK 293T and HAP1 were prepared, and 30 μg of protein was processed for western blot with the indicated Huntingtin antibodies ab45169 at 1/5000 and GTX638832 at 1/500. Tris-Glycine 4-20% gels were used for SDS-PAGE. The Ponceau stained transfer is shown as a loading control. Predicted band size: 347 kDa. *Recombinant antibody.\n\nFor western blot experiments, WT and HTT KO protein lysates were ran on SDS-PAGE, transferred onto nitrocellulose membranes, and then probed with twenty Huntingtin antibodies in parallel (Table 2, Figure 1).\n\n Monoclonal antibody,\n\n** Recombinant antibody.\n\nWe then assessed the capability of all twenty antibodies to capture Huntingtin from DMS 53 protein extracts using immunoprecipitation techniques, followed by western blot analysis. For the immunoblot step, a specific Huntingtin antibody identified previously (refer to Figure 1) was selected. Equal amounts of the starting material (SM), the unbound fraction (UB), as well as the whole immunoprecipitate (IP) eluates were separated by SDS-PAGE (Figure 3).\n\nDMS 53 lysates were prepared, and immunoprecipitation was performed using 2.0 μg of the indicated Huntingtin antibodies pre-coupled to Dynabeads protein A or protein G. The concentration of MA3-040* is unknown and therefore 5 μL of this antibody was tested. All samples were washed and processed for western blot with the indicated Huntingtin antibody. Tris-Glycine 4-20% gels were used for SDS-PAGE. For western blot, ab45169** was used at 1/5000. The Ponceau stained transfers of each blot are shown. SM=4% starting material; UB=4% unbound fraction; IP=immunoprecipitate. Monoclonal antibody, Recombinant antibody.\n\nFor immunofluorescence, twenty antibodies were screened using a mosaic strategy. First, DMS 53 WT and HTT KO cells were labelled with different fluorescent dyes in order to distinguish the two cell lines, and the Huntingtin antibodies were evaluated. Both WT and KO lines were imaged in the same field of view to reduce staining, imaging and image analysis bias (Figure 4). Quantification of immunofluorescence intensity in hundreds of WT and KO cells was performed for each antibody tested,12 and the images presented in Figure 3 are representative of this analysis.\n\nDMS 53 WT and HTT KO cells were labelled with a green or a far-red fluorescent dye, respectively. WT and KO cells were mixed and plated to a 1:1 ratio on coverslips. Cells were stained with the indicated Huntingtin antibodies and with the corresponding Alexa-fluor 555 coupled secondary antibody including DAPI. Acquisition of the blue (nucleus-DAPI), green (WT), red (antibody staining) and far-red (KO) channels was performed. Representative images of the merged blue and red (grayscale) channels are shown. WT and KO cells are outlined with green and magenta dashed line, respectively. When an antibody was recommended for immunofluorescence by the supplier, we tested it at the recommended dilution and at 1 μg/ml. The rest of the antibodies were tested at 1 and 2 μg/ml. The final concentration of each antibody was selected based on the detection range of the microscope used and a quantitative analysis not shown here. Antibody dilutions corresponding to the images shown are: ab109115* at 1/1500, ab45169 at 1/1500, ABCD_AG854 at 1/500, ABCD_AG855 at 1/200, A19064 at 1/600, 5656** at 1/100, CH03023* at 1/1700, MW1-S* at 1/20, MW3-S* at 1/10, MW4-S* at 1/10, MW5-S* at 1/15, MW6-S* at 1/60, MW7-S* at 1/15, MW8-S* at 1/20, GTX132433 at 1/500, GTX638832 at 1/500, 710695 at 1/500, MA3-040* at 1/1000, MA5-16703* at 1/1000, MA5-41256** at 1/1000. Bars = 10 μm. Monoclonal antibody, *Recombinant antibody.\n\nIn conclusion, we have screened twenty commercial Huntingtin antibodies by western blot, immunoprecipitation, and immunofluorescence by comparing the signal produced by the antibodies in human DMS 53 WT and HTT KO cells. Several high-quality and renewable Huntingtin antibodies were identified in all applications. Researchers who wish to study Huntingtin in a different species are encouraged to select high-quality antibodies, based on the results of this study, and investigate the predicted species reactivity of the manufacturer before extending their research.\n\nThe underlying data for this study can be found on Zenodo, an open-access repository for which YCharOS has its own collection of antibody characterization reports.17\n\n\nMethods\n\nThe standardized protocols used to carry out this KO cell line-based antibody characterization platform was established and approved by a collaborative group of academics, industry researchers and antibody manufacturers. The detailed materials and step-by-step protocols used to characterize antibodies in western blot, immunoprecipitation and immunofluorescence are openly available on Protocol Exchange, a preprint server (DOI: 10.21203/rs.3.pex-2607/v1).12\n\nCell lines used and primary antibodies tested in this study are listed in Tables 1 and 2, respectively. To ensure that the cell lines and antibodies are cited properly and can be easily identified, we have included their corresponding Research Resource Identifiers, or RRID.18,19", "appendix": "Data availability\n\nZenodo: Dataset for the Huntingtin antibody screening study, doi.org/10.5281/zenodo.11639052. 17\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgment\n\nWe would like to thank the NeuroSGC/YCharOS/EDDU collaborative group for their important contribution to the creation of an open scientific ecosystem of antibody manufacturers and KO cell line suppliers, for the development of community-agreed protocols, and for their shared ideas, resources, and collaboration. Members of the group can be found below. We would also like to thank the Advanced BioImaging Facility (ABIF) consortium for their image analysis pipeline development and conduction (RRID:SCR_017697). Members of each group can be found below.\n\nNeuroSGC/YCharOS collaborative group: Thomas M. Durcan, Aled M. Edwards, Peter S. McPherson, Chetan Raina and Wolfgang Reintsch.\n\nABIF consortium: Claire M. Brown and Joel Ryan.\n\nThank you to the Structural Genomics Consortium, a registered charity (no. 1097737), for your support on this project. The Structural Genomics Consortium receives funding from Bayer AG, Boehringer Ingelheim, Bristol-Myers Squibb, Genentech, Genome Canada through Ontario Genomics Institute (grant no. OGI-196), the EU and EFPIA through the Innovative Medicines Initiative 2 Joint Undertaking (EUbOPEN grant no. 875510), Janssen, Merck KGaA (also known as EMD in Canada and the United States), Pfizer and Takeda.\n\nAn earlier version of this of this article can be found on Zenodo (DOI: 10.5281/zenodo.11582780).\n\n\nReferences\n\nRoss CA, Tabrizi SJ: Huntington's disease: from molecular pathogenesis to clinical treatment. Lancet Neurol. 2011; 10(1): 83–98. Publisher Full Text\n\nA novel gene containing a trinucleotide repeat that is expanded and unstable on Huntington's disease chromosomes. The Huntington's Disease Collaborative Research Group. Cell. 1993; 72(6): 971–983. PubMed Abstract \| Publisher Full Text\n\nGreco TM, Secker C, Ramos ES, et al.: Dynamics of huntingtin protein interactions in the striatum identifies candidate modifiers of Huntington disease. Cell Syst. 2022; 13(4): 304–320.e5. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMatlik K, Baffuto M, Kus L, et al.: Cell-type-specific CAG repeat expansions and toxicity of mutant Huntingtin in human striatum and cerebellum. Nat. Genet. 2024; 56(3): 383–394. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPressl C, Matlik K, Kus L, et al.: Selective vulnerability of layer 5a corticostriatal neurons in Huntington's disease. Neuron. 2024; 112(6): 924–941.e10. PubMed Abstract \| Publisher Full Text\n\nStrong TV, Tagle DA, Valdes JM, et al.: Widespread expression of the human and rat Huntington's disease gene in brain and nonneural tissues. Nat. Genet. 1993; 5(3): 259–265. PubMed Abstract \| Publisher Full Text\n\nGreco TM, Secker C, Ramos ES, et al.: Dynamics of huntingtin protein interactions in the striatum identifies candidate modifiers of Huntington disease. Cell Syst. 2022; 13(4): 304–20.e5. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWanker EE, Ast A, Schindler F, et al.: The pathobiology of perturbed mutant huntingtin protein-protein interactions in Huntington's disease. J. Neurochem. 2019; 151(4): 507–519. PubMed Abstract \| Publisher Full Text\n\nAyoubi R, Ryan J, Biddle MS, et al.: Scaling of an antibody validation procedure enables quantification of antibody performance in major research applications. elife. 2023; 12: 12. Publisher Full Text\n\nCarter AJ, Kraemer O, Zwick M, et al.: Target 2035: probing the human proteome. Drug Discov. Today. 2019; 24(11): 2111–2115. PubMed Abstract \| Publisher Full Text\n\nLicciardello MP, Workman P: The era of high-quality chemical probes. Rsc Med. Chem. 2022; 13(12): 1446–1459. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyoubi R, Ryan J, Bolivar SG, et al.: A consensus platform for antibody characterization (Version 1). Protocol. Exchange. 2024. Publisher Full Text\n\nBiddle MS, Virk HS: YCharOS open antibody characterisation data: Lessons learned and progress made. F1000Res. 2023; 12(12): 1344. Publisher Full Text\n\nLaflamme C, McKeever PM, Kumar R, et al.: Implementation of an antibody characterization procedure and application to the major ALS/FTD disease gene C9ORF72. elife. 2019; 8: 8. Publisher Full Text\n\nAlshafie W, Fotouhi M, Shlaifer I, et al.: Identification of highly specific antibodies for Serine/threonine-protein kinase TBK1 for use in immunoblot, immunoprecipitation and immunofluorescence. F1000Res. 2022; 11: 977. Publisher Full Text\n\nJung R, Lee Y, Barker D, et al.: Mutations causing Lopes-Maciel-Rodan syndrome are huntingtin hypomorphs. Hum. Mol. Genet. 2021; 30(3-4): 135–148. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyoubi R, Laflamme C: Dataset for the Huntingtin antibody screening study. [Data set]. Zenodo. 2024.\n\nBandrowski A, Pairish M, Eckmann P, et al.: The Antibody Registry: ten years of registering antibodies. Nucleic Acids Res. 2023; 51(D1): D358–D367. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBairoch A: The Cellosaurus, a Cell-Line Knowledge Resource. J. Biomol. Tech. 2018; 29(2): 25–38. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "315014", "date": "28 Aug 2024", "name": "Jieya Shao", "expertise": [ "Reviewer Expertise Cell 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\nIn this study, the authors performed comprehensive analysis of twenty commercially available HTT antibodies for their suitability and performance in Western blot, immunoprecipitation, and immunofluorescence staining applications. The experimental approaches were logically and thoroughly designed. Data are of high-quality and clearly presented. The results will be very helpful for researchers in the HTT field. However, there seems to be two small errors. First, in Figure 3, only the IP data of 10 antibodies were shown while in the text it was stated that 20 antibodies were analyzed. Second, on Page 5, in the last sentence of the second paragraph, the authors seem to cite Figure 3 by mistake.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and materials provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [] }, { "id": "315013", "date": "09 Sep 2024", "name": "Karen Bowman", "expertise": [ "Reviewer Expertise drug discovery and diagnostics" ], "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 pointed out the current technical difficulties in studying Huntington’s Disease and identified the differences in antibodies to the protein Huntingtin, used throughout the HD research community, as a particular problem. The study formed part of the wider YCharOS collaboration to characterise commercial antibodies for human proteins using standardized protocols with the intention of improving antibody reproducibility issues. The study was an evaluation of twenty commercial antibodies for the protein Huntingtin for use in western blot, immunoprecipitation, and immunofluorescence. The study was well designed and the results were to a high technical standard and quality. The data is presented clearly and looks robust. It was reassuring to see that a committee of industry and academic representatives had endorsed the platform used. The platform consisted of identification of human cell lines suitable for antibody characterisation studies i.e. with adequate levels of Huntingtin, followed by the development of and contribution of isogenic knockout control cell lines, which were validated at the protein level. The final step was a series of antibody characterisation procedures, limited to the most common research uses of antibodies. Limitations of the study are clearly stated. Two minor errors were spotted. There is an inconsistency between the text and figure 3, where they mention IP was done on twenty antibodies in the text, but results for only ten antibodies are presented in the figure. Perhaps some additional text on why only ten were presented would be of use, or amend the text to reflect what is presented in the figure. A second error is that in the final results paragraph on immunofluorescence, figure 3 is cited, and linked back to, instead of figure 4. Overall, this will be a highly useful resource for scientists in the HD research community.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and materials 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/13-922
https://f1000research.com/articles/14-5/v1	02 Jan 25	{ "type": "Systematic Review", "title": "Information security education: a thematic trend analysis", "authors": [ "Paula Andrea Rodríguez-Correa", "Alejandro Valencia-Arias", "Ezequiel Martínez Rojas", "Aarón Oré León", "Ray Harvey Mellin Rubio", "Manuel Humberto Vásquez Coronado", "Jesus Alberto Jimenez Garcia", "Paula Andrea Rodríguez-Correa", "Ezequiel Martínez Rojas", "Aarón Oré León", "Ray Harvey Mellin Rubio", "Manuel Humberto Vásquez Coronado", "Jesus Alberto Jimenez Garcia" ], "abstract": "The evolution of information technologies has led to a significant increase in information security risks, underscoring the urgent need for professional education in this field to safeguard organizational data. Publications on this topic highlight the necessity of educating individuals about data security, and their number has grown in recent years. This study aims to identify thematic trends in information security education through a bibliometric analysis based on five research questions, following PRISMA guidelines. Ninety-nine documents from Scopus and Web of Science were analyzed, revealing a quadratic growth, with 2023 as the year of greatest research activity. Prominent contributors such as Von Solms, Safa, and Furnell stand out, along with the journal Computers & Security and the significant influence of the United States in this field. The potential is identified in the following areas: information security culture, media, information security regulations, higher education institutions, information security management, and cybersecurity. The study also identifies gaps and proposes a research agenda to address these gaps.", "keywords": [ "Information security", "cybersecurity", "data protection", "information security training", "higher education" ], "content": "1. Introduction\n\nAs information technologies advance, information security risks also increase. Consequently, companies are increasingly concerned about cyber-attacks (Safa et al., 2015). Thus, information security has become one of the most important and debated topics among experts. Actions are being taken by implementing organizational information security procedures and policies, encouraging information security-conscious behavior, and promoting the sharing of information security knowledge (Safa et al., 2018).\n\nGiven the rapid technological evolution, continuous evaluation and constant innovation to improve security systems, such as multi-layer firewalls, have become essential. According to Torten et al. (Torten et al., 2018), cyberattacks often target the installation of ransomware, intellectual property infringement, theft of medical records, unauthorized banking transactions, and credit card misuse. Consequently, companies worldwide have placed a high priority on information systems risk management (Flores & Ekstedt, 2016).\n\nGiven the growing importance of protecting information systems within organizations, there has been an increased focus on developing human talent in cybersecurity in recent years as a key component of information technology education. This focus is reflected in both core and advanced curricula (Rowe et al., 2011). In addition, higher education plays a key role in society, particularly in terms of research, development, and education. Consequently, faculties are establishing their own Information Technology (IT) networks designed to support research, development, and teaching activities in the field of information security (Ulven & Wangen, 2021).\n\nTherefore, several literature reviews have focused on investigating the fundamental knowledge, skills, and capabilities required by cybersecurity professionals (Jones et al., 2018), as well as analyzing information security curricula (Parrish et al., 2018). These studies have highlighted that information security has emerged as a crucial area of learning that demands specific professional education, particularly in areas such as secure programming, network security, and offensive security (Švábenský et al., 2020).\n\nInformation security education and training are critical for preparing both current and future IT professionals to adequately and effectively address real-world security risks and incidents (Beuran et al., 2016). Consequently, universities recognize the importance of staying abreast of cyber threats and risks to provide students with up-to-date cybersecurity tools and knowledge through innovative methodologies (Cheung et al., 2011).\n\nThus, it has been evidenced that in the current era, where information is an invaluable asset and technology drives most human activities, information security has become a paramount concern (Burov et al., 2020). This concern is compounded by the increasing sophistication of cyber threats and the proliferation of cyber-attacks in both business and personal domains (Khaleefah & Al-Mashhadi, 2024). Therefore, the growing need for trained information security professionals is evident, making university programs dedicated to information security education essential (AlDaajeh et al., 2022).\n\nBased on the background of the literature, the objective of this research is established: to identify the thematic trends in information security education. To this end, the following research questions are formulated:\n\n• Who are the research actors with the greatest impact on the field?\n\n• What is the thematic evolution of the information security education phenomenon in recent years?\n\n• What are the research clusters related to the phenomenon under study?\n\n• What are the most frequent and current topics within the information security education phenomenon?\n\n• What are the most promising topics for future research on the phenomenon under study?\n\n\n2. Methods\n\nA bibliometric analysis was used, a technique that allows for evaluating the quality and quantity of published scientific literature, as well as studying research trends and various citation analyses within a given field. It is considered a useful and productive tool for determining research trends across different disciplines (Buber & Koseoglu, 2022). To ensure data quality, the parameters of the PRISMA 2020 guidelines for systematic reviews and meta-analyses were followed. The PRISMA guidelines establish protocols for proper literature review, including a four-phase flowchart and a 27-item checklist (Selçuk, 2019). The phases followed in this study are detailed below.\n\nOne of the first steps to meet the criteria of the PRISMA statement is to establish the inclusion and exclusion criteria used in the selection of documents (Moraga & Cartes-Velásquez, 2015). For inclusion, a search was conducted using the following keywords: “information,” “security,” “education,” and “training.” As for exclusion criteria, conference proceedings were omitted, as they typically represent very recent studies that have not yet been consolidated as relevant research in the field addressed by this study. Additionally, documents with indexing errors were excluded.\n\nThe two most commonly used databases for this type of analysis were selected: Scopus and Web of Science. The choice of these databases is based on their significant differences in terms of coverage, accessibility, updating, and the number of citations retrieved (Manriquez et al., 2015).\n\nAs part of the search strategy, a search equation was applied in both databases. The AND operators were used to include the terms “information” and “security” in the search. Additionally, OR was used to broaden the search to include the fields of \"education\" or \"training.\" The following search equations were applied:\n\nIn Scopus: TITLE (information AND security AND (education OR formation))\n\nIn Web of Science: TI= (information AND security AND (education OR formation))\n\nOnce the search equations were applied in the databases, 261 records were obtained from Scopus and 44 from Web of Science, totaling 305 records. Subsequently, duplicates were removed, resulting in 271 documents that were then subjected to inclusion and exclusion criteria. After applying these eligibility criteria, 99 documents were selected for analysis. Figure 1 displays the flowchart of the document selection process.\n\nOwn elaboration based on Scopus and Web of Science.\n\n\n3. Results\n\nFirst, an analysis of publications by year is conducted. As shown in Figure 2, a steady increase has been observed in recent years regarding the subject of this research. However, a decrease is noted in 2022, followed by an increase in 2023. Therefore, since 2013, there has been a continuous rise in the number of publications. Additionally, an analysis of quantity, quality, and structural indicators is performed.\n\nOwn elaboration based on Scopus and Web of Science.\n\nAs stated in the first research question, the aim is to identify the most relevant research actors. Figure 3 displays the most influential authors. The blue color indicates the authors with the highest number of citations, while the green color represents the authors with the highest number of publications. Authors highlighted in yellow are those with both the highest number of publications and the highest number of citations.\n\nOwn elaboration based on Scopus and Web of Science.\n\nIn this context, authors Von Solms R., Safa N.S., and Furnell S. stand out as significant contributors to the field. In 2015, they conducted research on the education of information security-conscious behaviors in organizations. This study evaluated various factors, including information security awareness, organizational information security policies, information security experience, and involvement, attitudes toward information security, subjective norms, threat assessment, information security self-efficacy, and user capability (Safa et al., 2015).\n\nRezgui Y. and Marks A. have also made significant contributions to this area of study. Their research focuses on the determinants of information security awareness among university personnel, including those involved in information systems decision-making, within the specific context of a developing country. They identified that variables such as conscientiousness, cultural norms and beliefs, and social dynamics significantly shape the attitudes and behaviors of academic staff, both in their general professional conduct and their awareness of information security practices (Rezgui & Marks, 2008).\n\nFigure 4 shows the journals with the greatest impact on the field of study. Among them, the journal Computers & Security stands out, having both the highest number of publications and the highest number of citations. Notably, the study by Safa et al. (Safa et al., 2015) has 226 citations and is recognized as one of the most relevant publications in this field. Additionally, the study by Rezgui and Marks (Rezgui & Marks, 2008) also warrants special attention, as it aligns with the trends identified in this research and has 138 citations.\n\nOwn elaboration based on Scopus and Web of Science.\n\nAdditionally, the study by Rajab and Eydgahi (Rajab & Eydgahi, 2019), which has received 53 citations, is notable for presenting a theoretical framework that encompasses various behavioral theories. This framework focuses on assessing factors such as perceived vulnerability, response effectiveness, response cost, and information security compliance intentions in higher education, making it a significant contribution to the field. Likewise, the study conducted by Bongiovanni (Bongiovanni, 2019) deserves attention, as it currently has 44 citations and focuses on literature related to information security management in higher education.\n\nThe countries that have had the greatest impact on research in information security education are presented below (see Figure 5). The United States stands out in the first place, both in terms of the number of citations and the number of publications related to this topic. Notable among these is the study by Rowe et al. (Rowe et al., 2011), which analyzes the crucial role of cybersecurity in IT education and argues for the integration of this topic into IT programs. Also noteworthy is the study by Conklin (Conklin, 2006), conducted in the United States, which proposes an active learning solution for a capstone course designed to enhance students’ competencies in cyberdefense and information security education.\n\nOwn elaboration based on Scopus and Web of Science.\n\nIn addition, South Africa stands out due to the significant contributions of the study by Safa et al. (Safa et al., 2015), which has made a notable impact on the field of information security. Also highlighted is a study that provides operational definitions for information security education, information security training, and information security awareness. This research aims to help institutions determine when it is necessary to train or educate employees and when it is appropriate to implement information security awareness programs (Amankwa et al., 2014).\n\nThe second research question focuses on the thematic evolution of the phenomenon under study. Figure 6 presents a count of the most frequent keywords from 2000 to 2024. Recently, there has been a significant increase in interest in the topic of security perception. In this context, the work of Mohr and Walter (Mohr & Walter, 2019) is notable, as it addresses information security education and users’ perceptions regarding the security of their personal information and online banking transactions. Another relevant topic is Higher Education, where there has been growing concern in recent years about individuals’ awareness of information security. This concern has led to the development of training programs in this area (Wahyudiwan et al., 2017).\n\nOwn elaboration based on Scopus and Web of Science.\n\nAnother topic that has generated interest in recent years is Information Technologies, driven by the need to train personnel across various organizations and economic sectors on the security of information systems (Katsikas, 2000). Similarly, security behavior has gained relevance during this period. With the rise in cyber-attacks affecting business performance and reputation, as well as compromising intellectual property, human factors have been identified as the weakest link in the security posture of any networked system. Consequently, studies have been conducted to analyze the relationship between threat perception and the understanding of countermeasures concerning the adoption of secure behaviors (Torten et al., 2018).\n\nIn the last five years, data protection and data security have become highly relevant, particularly due to the increasing pressure on data storage. This rise, driven by massive data generation, has led to rapid development in the storage market. Consequently, data protection has become a critical issue for organizations (Yang et al., 2020).\n\nThe third research question focuses on the thematic clusters present in the field of study. Figure 7 shows the keyword co-occurrence network, generated by the VOSviewer software (Van Eck and Waltman, 2010). In this representation, several clusters in the form of nodes are identified, with the largest cluster corresponding to information security education, which is the main topic of this research. This cluster is associated with topics such as e-learning and information assurance education. In this context, researchers such as Pastor et al. (Pastor et al., 2010) argue that the best way to enhance an individual’s response to security threats is through robust education, engaging hands-on training, and increased general awareness of information security.\n\nOwn elaboration based on Scopus and Web of Science.\n\nAn important cluster focuses on higher education, specifically addressing issues such as awareness and human factors, which, as previously highlighted, represent the weakest link in information protection. Therefore, increasing security awareness is imperative (Brill et al., 2013). This emphasis is also reflected in the information security awareness cluster, further underscoring the need for research in this area (Lebek et al., 2014). This cluster is associated with topics such as gamification to enhance motivation and learning outcomes. Gamification thus demonstrates significant potential for use in security awareness and training programs (Gjertsen et al., 2017).\n\nGenerally spoken, it should look like this document. Using the styles of this document may make things easier for you, but you don’t need to use them.\n\nIn addition, the information security management cluster is highlighted due to the significant expansion of online business opportunities brought about by information technology. However, these opportunities have also introduced serious information security risks (Soomro et al., 2016). This cluster is related to topics such as information society, systems, and governance. It encompasses areas like social network analysis, distance education, and security culture. Similarly, an information security training cluster has emerged, addressing topics such as digital education and the impact of COVID-19. Consequently, research has been conducted to analyze information security behavior and awareness among Zoom application users, particularly during the COVID-19 pandemic (Candiwan et al., 2022).\n\nThe fourth research question focuses on identifying the most frequent and relevant topics in the research area. Figure 8 displays four research quadrants. The first quadrant highlights the most frequent and relevant keywords, indicating topics that could guide future research on information security education. In this quadrant, the key topics are information security awareness and higher education. Notably, a study on the information security awareness of end users, including information systems decision-makers, within the context of three higher education environments is emphasized (Rezgui & Marks, 2008).\n\nOwn elaboration based on Scopus and Web of Science.\n\nThe second quadrant highlights topics that, while less recurrent in the literature, are considered relevant, especially in recent years. Key aspects include information laws, higher education institutions (HEIs), risk assessment, cybersecurity, information security incidents, data analysis, security awareness, best practices, information technologies, information security policy, information security training, hacking, computer security, information security management, and security management. These keywords also hold significant potential for future research.\n\nIn quadrant three, less frequent and recurrent topics are identified, suggesting that they either need more momentum to become common and current or that researchers have lost interest over time. These themes include governance, information, and e-learning. Finally, quadrant four contains topics that are more frequent but less current, with evidence of a decreasing interest among researchers in recent years. Among these, awareness stands out.\n\nFinally, a research agenda is proposed with potential future lines of inquiry based on the findings of this study, as visualized in Figure 9. One of the topics with significant future potential is the culture of information security. Continuous failures in information security within organizations have highlighted the importance of focusing on organizational culture. It is argued that developing an information security culture would subsequently lead to a more secure organization; therefore, this topic should be considered in the education of information security professionals (Tejay & Mohammed, 2023).\n\nOwn elaboration based on Scopus and Web of Science.\n\nSimilarly, the potential of media is noteworthy, as it facilitates education in information security. Consequently, research has been developed to understand the current challenges in information literacy, analyze their causes, and offer solutions and strategies. These studies provide valuable references for future information literacy education through various communication channels (Tao, 2023). Additionally, the relevance of information regulations is highlighted. Information technology, as a regulated element, is interpreted differently across various sectors of security legislation. Therefore, research in this field is essential to promote consistency in the regulatory framework (Brinker, 2024).\n\nHigher education institutions also emerge as a relevant topic for future research on compliance with information security policies. Although the literature in this area is still limited, it shows promising advancements (Hina & Dominic, 2020). Another crucial aspect is information security risk assessment, which is a fundamental component of business management practices. This assessment helps to identify, quantify, and prioritize risks based on risk acceptance criteria and organization-specific objectives. Therefore, education in this area is essential for information security professionals (Kuzminykh et al., 2021).\n\nInformation security management is a critical issue that must be continuously addressed in the future. As Merchan-Lima et al. (Merchan-Lima et al., 2021) emphasize, adopting effective information security management practices is imperative, especially in the context of higher education. Future professionals should be trained to design and implement these practices. Additionally, cybersecurity plays a significant role in the future of higher education. According to Kweon et al. (Kweon et al., 2021), security training and education are effective methods for detecting cyberattacks in both academic and industrial settings. Their research found a positive association between cybersecurity training and a reduction in cyberattacks within organizations.\n\nBased on the identified potential topics, the following research questions are proposed for future research:\n\n• What are the main aspects of information security culture in organizations, and how do they influence the effectiveness of training programs?\n\n• What role does the media play in promoting information security awareness and education among the general public and industry professionals?\n\n• What are the most relevant international and national (e.g., Colombia) information security regulations, and how do they impact information security education programs?\n\n• How are higher education institutions integrating information security into their academic programs, and what pedagogical approaches are they using?\n\n• What are the key challenges in managing information security within organizations, and how can these challenges be addressed through education and staff training?\n\n• What are the essential competencies that information security professionals need to acquire, and how are these competencies being developed in education programs?\n\n• What strategies are organizations implementing to foster a culture of cybersecurity among their employees, and how are they evaluating the effectiveness of these strategies?\n\n• How are higher education institutions collaborating with industry and other organizations to ensure that information security training programs align with labor market needs?\n\n• What are the emerging trends in cybersecurity, and how are they being addressed in education programs to ensure relevance and up-to-date knowledge?\n\n• What is the impact of information security education on risk reduction and data protection within organizations and society at large?\n\n\n4. Discussion\n\nInformation security education and awareness are crucial issues in today’s world. Similar to the study conducted by Amankwa et al. (Amankwa et al., 2015), a literature review has been employed to evaluate models aimed at improving security education and awareness. These models focus on stakeholder domains such as end users, institutions, and industry. This study also identifies a research gap regarding the relationship between higher education institutions and industry (a key stakeholder), as well as other organizations, to ensure that information security education programs align with labor market needs.\n\nPrevious research aligns with the findings of this study on information security policies. For example, Alassaf and Alkhalifah (Alassaf & Alkhalifah, 2021) found that the literature lacks information on the positive and negative (direct or indirect) influences of human and organizational theories and their factors affecting information security policy compliance behavior. This study also identifies gaps in higher education related to information security policy education. Addressing these gaps is crucial to ensure that future professionals are prepared to design and implement secure practices within higher education institutions.\n\nThe results of this study are consistent with insights from other research, such as the systematic review by Hina and Dominic (2020) on compliance with information security policies. Their review emphasized that limited attention has been given to critical infrastructures, including higher education institutions, that are particularly vulnerable. They also examined the role of institutional governance in shaping protection motivation for information security. Similarly, this study underscores the importance of further exploring the intersection of information security policies and higher education. It also highlights the need for information security professionals to develop essential skills in policy implementation and gain a deeper understanding of how these policies are integrated into educational programs.\n\nThe findings of the study by Merchan-Lima et al. (Merchan-Lima et al., 2021), who conducted a systematic literature review on effective information security management practices in higher education institutions, align with the results of this research. These authors suggest that future research should focus on how to educate users and the mechanisms to help higher education institutions select the appropriate risk assessment models for their needs. This study similarly highlights the importance of information security management and risk assessment within the context of professional education, underscoring the need for further exploration in future studies. Additionally, it emphasizes the relevance of designing curricula that align with the current needs of organizations.\n\nThis study contributes from two fundamental perspectives. First, from a theoretical standpoint, it identifies the predominant trends in information security education research. Additionally, it explores emerging themes, underdeveloped areas, and persistent challenges. These findings provide a clearer picture of the structure of knowledge in the field of information security education, including the relationships between influential concepts, authors, journals, and countries. Second, the study highlights underexplored areas and topics that require further attention in future research. This information is valuable for formulating new hypotheses and raising relevant research questions. Ultimately, it contributes to the advancement and enhancement of information security practices in an increasingly digitized world.\n\nIn terms of practical implications, the findings of this research may be valuable as a guide and foundation for planning higher education training programs. These results offer relevant information on key topics, methods, and approaches in the field of information security, which may influence the design of academic and training programs.\n\nIn addition, these findings can support decision-making in both academia and business. They help to understand the needs in information security education and to allocate resources more effectively. By examining the existing literature, best practices in information security education based on currently relevant topics have been identified. This information can serve as a reference for the development and implementation of successful programs.\n\nAdditionally, evidence gathered from existing literature on the impact of information security education interventions, such as awareness programs or professional development initiatives, is valuable for informing future strategies. Finally, the findings of this study can be used as input for designing international benchmarking and comparison reports. By comparing scientific output in information security education across different countries, institutions, and sectors, opportunities for collaboration and mutual learning can be identified (Escuela Europea de Excelencia, 2021).\n\nSimilarly, the study has some limitations. First, it is based on the Scopus and Web of Science databases, which are the most widely used in academia. However, it would be beneficial to explore other databases, such as IEEE, that focus on engineering topics, including information security. Second, the selection criteria were not extremely specific, so future research could define more detailed criteria and carry out a systematic literature review to deepen the research topic. Third, although the study analyzes thematic trends, it does not focus as much on research gaps. Despite this, a research agenda is provided to guide future studies.\n\nThe following Table ( Table 1) presents a summary of the gaps identified in the research, these gaps cover various aspects, from the integration of information security in academic programs to the lack of standardized assessment methodologies and the need for an interdisciplinary perspective, each gap is accompanied by a brief justification and questions to guide future research in the field, this table provides an overview of the critical areas that require attention and development to advance information security education.\n\nIntegrating information security into higher education programs is crucial for ensuring that students are equipped with the necessary skills to address modern cyber threats. This integration helps bridge the gap between theoretical knowledge and practical application, preparing graduates to handle real-world security challenges effectively. Developing detailed curricula that include up-to-date practices and emerging technologies could greatly enhance students’ readiness for the evolving landscape of information security.\n\nAnother significant gap lies in the lack of standardized methodologies for evaluating the effectiveness of information security training programs, without a consistent evaluation framework, it is difficult to determine which pedagogical approaches are most effective and how to improve the quality of the training offered, this gap highlights the need to develop robust evaluation tools that can measure the real impact of information security training on risk reduction and data protection.\n\nFurthermore, the paucity of data and metrics available to measure the impact of information security education programs presents an additional challenge. The lack of quantitative and qualitative information limits our understanding of the effectiveness of these programs and makes it difficult to identify areas for improvement, it is critical to address this gap by systematically collecting relevant data and developing robust metrics that can guide informed decision-making in the design and implementation of information security education programs.\n\nYour analysis highlights crucial areas where further research and development are needed. Addressing these gaps can significantly enhance the effectiveness and relevance of information security education programs. By focusing on integration, standardized evaluation, data collection, and interdisciplinary approaches, future research can contribute to a more comprehensive and robust framework for preparing professionals to tackle the evolving challenges in information security.\n\n\n5. Conclusion\n\nThe increasing complexity and frequency of cyber threats underscore the critical need for robust information security education. Universities play a pivotal role in this by preparing future professionals with the necessary skills and knowledge to combat these threats effectively. As technology evolves and cyber risks become more sophisticated, higher education institutions must adapt their programs to ensure they align with current and future demands. This includes not only teaching technical skills but also fostering a deep understanding of organizational policies, risk management, and the broader context of information security.\n\nThe objective of this research was to identify thematic trends in the field of information security education. To achieve this, five research questions were formulated. The first question made it possible to identify the research actors, i.e., the most influential authors, journals, and countries in this field. Authors Von Solms R, Safa NS, and Furnell S, as well as Rezgui Y and Marks A, stand out as the most influential due to their large number of publications and citations recorded. In terms of journals, Computers & Security stands out as the most influential, with a high volume of publications and citations on this topic. In addition, the United States and South Africa are the most relevant countries in this context. This leads to the conclusion that the aforementioned actors are the ones that generate the most impact within the field of education research in information security.\n\nThe second research question focused on the thematic evolution in recent years within the research field. Through an analysis of the most recurring keywords for each year, significant researcher interest was identified in the following topics: security perception, higher education, information technology, security behavior, data protection, and data security. The third research question addressed thematic research clusters. In this analysis, the following information security relevant clusters were identified: information security education, information security awareness, higher education, information security management, cybersecurity, and information security training.\n\nIn the fourth research question, an analysis of the currency and frequency of keywords related to the research topic was conducted. The findings revealed that the most frequent and current topics are information security awareness and higher education. This suggests that future research can be oriented towards these two topics. Finally, from the fifth question, a research agenda was proposed that identifies potential in the following thematic areas: information security culture, media, information security regulations, higher education institutions, information security management, and cybersecurity. From these thematic areas, research questions were proposed that could be answered in future research.\n\n\nEthics and consent\n\nEthical approval and consent were not required.\n\nDisclosure statement. No.", "appendix": "Data availability statement\n\nNo data are associated with this article.\n\nZenodo: Dataset and supporting materials for the study “Information security education: a thematic trend analysis”. https://doi.org/10.5281/zenodo.14271117 (Valencia-Arias, 2024).\n\nThe project contains the following underlying data:\n\n1. PRISMA Flow Diagram (Flowchart illustrating the study selection process based on PRISMA guidelines).Dataset.xlsm (Raw data supporting the findings of this study).\n\n2. Editable _ seguridad de la info en educación.xlsm\n\n3. PRISMA Checklist.pdf (Checklist detailing compliance with PRISMA 2020 guidelines).\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgement\n\nNot applicable.\n\n\nReferences\n\nAlassaf M, Alkhalifah A: Exploring the Influence of Direct and Indirect Factors on Information Security Policy Compliance: A Systematic Literature Review. IEEE Access. 2021; 9: 162687–162705. Publisher Full Text\n\nAlDaajeh S, Saleous H, Alrabaee S, et al.: The role of national cybersecurity strategies on the improvement of cybersecurity education. Comput. Secur. 2022; 119: 102754. Publisher Full Text\n\nAmankwa E, Loock M, Kritzinger E: A conceptual analysis of information security education, information security training, and information security awareness definitions. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSoomro ZA, Shah MH, Ahmed J: Information security management needs more holistic approach: A literature review. Int. J. Inf. Manag. 2016; 36(2): 215–225. Publisher Full Text\n\nŠvábenský V, Vykopal J, Čeleda P: What Are Cybersecurity Education Papers About?: A Systematic Literature Review of SIGCSE and ITiCSE Conferences. SIGCSE’20: Proceedings of the 51st ACM Technical Symposium on Computer Science Education. 2020. Publisher Full Text\n\nTao H: Investigation on the Current Situation of Information Technology Education in Rural Primary Schools in Hainan. Proceedings of the 2023 4th International Conference on Education, Knowledge and Information Management (ICEKIM 2023). 2023.\n\nTejay GPS, Mohammed ZA: Cultivating security culture for information security success: A mixed-methods study based on anthropological perspective. Inf. Manag. 2023; 60(3): 103751. Publisher Full Text\n\nTorten R, Reaiche C, Boyle S: The impact of security Awareness on information technology professionals’ behavior. Comput. Secur. 2018; 79: 68–79. Publisher Full Text\n\nUlven JB, Wangen G: A Systematic Review of Cybersecurity Risks in Higher Education. Future Internet. 2021; 13(2): 39. Publisher Full Text\n\nValencia-Arias A: Information security education: a thematic trend analysis. [Data set]. Zenodo. 2024. Publisher Full Text\n\nVan Eck N, Waltman L: Software survey: VOSviewer, a computer program for bibliometric mapping. Scientometrics. 2010; 84(2): 523–538. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWahyudiwan DDH, Sucahyo YG, Gandhi A: Information security awareness level measurement for employee: Case study at ministry of research, technology, and higher education. 3rd International Conference on Science in Information Technology (ICSITech). 2017. Publisher Full Text\n\nYang P, Xiong N, Ren J: Data Security and Privacy Protection for Cloud Storage: A Survey. IEEE Access. 2020; 8: 131723–131740. Publisher Full Text" }	[ { "id": "356007", "date": "25 Jan 2025", "name": "Lynette Drevin", "expertise": [ "Reviewer Expertise Human aspects of IT", "Information security & awareness" ], "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 Are the rationale for, and objectives of, the Systematic Review clearly stated? The aim of the paper is presented and the research questions are stated clearly and are addressed in the paper.\n2 Research design and methods:\nAdherence to the PRISMA reporting guidelines – Yes Inclusion criteria – This is stated. However – the terms awareness and human behaviour in information security would have been valuable in the searches. Information sources - A few of the sources is quite old in the context of this fast-evolving field. Examples will follow at the end of this report. Search strategies – it is confusing to the reviewer why certain sources – such as conference proceedings are left out in the search. It is important to have recent studies – in contrast of the reason given in section 2.3 to leave those out.\nFor example – IFIP WISE (World Conference on Information Security Education) has Scopus outputs that could be valuable. https://link.springer.com/conference/wise-ifip As could HIASA be - IFIP International Symposium on Human Aspects of Information Security & Assurance. https://haisa.org/ Scopus outputs:\n\nhttps://link.springer.com/conference/haisa\n\n3 Are the conclusions drawn adequately supported by the results presented in the review? Yes, mostly – but in section 3.5 where future work is proposed – the authors could have searched what is already being done and addressed on their topics identified. E.g. bullet 4 is addressed in papers and work done by IFIP WG11.8 (WISE) as stated above. Also see . https://cybered.hosting.acm.org/wp/ ACM/IEEE/AIS SIGSEC/IFIP Cybersecurity Curricular Guidelines CSEC. There is also a list of prominent academics and researchers that worked on this – and may be added to the names of people involved in these studies. The other bullets may also already have work done on the topics stated. This could have been searched and acknowledged.\nFigure 3- Most relevant authors: As certain important literature was excluded; some important authors were not part of the results – e.g. Prof Matt Bishop in the USA. Prof LA Futcher in South Africa, etc.\nRegarding the names stated in figure 3 as most relevant authors – the reviewer was hoping to see more of their work cited in the reference list. E.g. Von Solms, Furnell etc.\nFuture work is stated.\n4 General remarks Page 7 – Do not use the word below when referring to a table of figure. Below (See Figure 5) – and then the figure is not below but in the top of the page\n\nWhere more or more recent sources are needed: Section 1 – par 2 – Rocha Flores W, et al., 2016 (Ref 1) Par 5 Cheung et al 2011 – newer source where universities are of concern – see how the ACM/IEEE CSEC guidelines were developed. See Futcher on curriculum issues.\nSection 3.1 Par 4 Rezgui & Marks – 2008. Maybe more sources can be added to strengthen this topic of researchers: See: for example, - Bada M, et al., 2019 (Ref 2) Kruger H, et al., 2010 (Ref 3) Drevin L, et al., 2007 (Ref 4)\nSection 3.3 Par 2 Gjertsen et al 2017 – add more recent sources on gamification and security e.g. - Silic M, et al., 2020 (Ref 5)\nThank you for an interesting article.\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? 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/14-5
https://f1000research.com/articles/14-4/v1	02 Jan 25	{ "type": "Research Article", "title": "Alpha-guaiene isolated from patchouli oil exhibits antifungal activity against four pathogenic fungi", "authors": [ "Sarifah Nurjanah", "Zhaqqu Alhafidz", "Maghfira Maulani", "Tita Rialita", "Elazmanawati Lembong", "Zhaqqu Alhafidz", "Maghfira Maulani", "Tita Rialita", "Elazmanawati Lembong" ], "abstract": "Background The major component of patchouli oil, patchouli alcohol, is used as fixative and has antimicrobial activity. The function of other components, such as α-guaiene, remains unknown. Therefore, this study reports the antifungal activity of α-guaiene isolated from patchouli oil against for pathogenic fungi: Aspergillus niger, Candida albicans ATCC 7102, Microsporum gypseum ATCC 14683, and Trichophyton mentagrophytes ATCC 16404.\n\nMethods The material from fraction (249°C-254°C) had the highest α-guaiene. Minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) were determined using the microdilution technique to evaluate antifungal activity, with n-hexane and medium serving as negative controls, and ketoconazole and fluconazole serving as positive controls.\n\nResults The results showed that the MIC value was determined at 45%, 50% for C. albicans, 55%, 60% for A. niger, 50%, 60% for M. gypseum, and 95%, 100% for T. mentagrophytes, respectively. Positive and medium controls demonstrated no microbial growth, whereas negative and growth controls revealed the presence of microorganisms. Fungus resistance to α-guaiene T. mentagrophytes exhibited the highest MIC value.\n\nConclusions Overall, this study reveals that α-guaiene is a promising agent effective against the studied pathogenic fungi.", "keywords": [ "α-guaiene", "patchouli oil", "antifungal activity", "Minimum Inhibitory Concentration", "Minimum Fungicidal Concentration" ], "content": "Introduction\n\nPatchouli oil is an essential oil extracted from various plant parts such as flowers, leaves, stems, and roots (Pandey et al., 2022; van Beek & Joulain, 2018). This oil has promising export potential, as it is regularly used in perfume, soap, pharmaceutical, cosmetic, and other industries. Patchouli oil is renowned for its active constituents and therapeutic benefits (Leong et al., 2019). Patchouli is a crucial aromatic plant in the perfume industry (Jain et al., 2022). Furthermore, patchouli oil accounts for approximately 85% of Indonesia’s essential oil exports, with a current annual value of 1,200-1,500 tons. Indonesia exports 90% of the world’s patchouli oil (Rahmayanti et al., 2018). Notably, patchouli oil’s constituent components, such as patchouli alcohol, α-guaiene, δ-guaiene, α-patchoullene, and seychellene, have several benefits (Pandey et al., 2021). Pressure, temperature, reflux ratio, and fractionation column all play crucial roles in the fractionating process used to separate these constituents (Almeida et al., 2018; Nurjanah et al., 2020).\n\nEssential oils have antiviral, antiparasitic, antifungal, bactericidal, insecticidal, and nematocidal properties (Khaledi & Zahani, 2018). They can be used as bactericides and fungicides against various human-infecting fungus and bacterium types. A pathogenic fungus Candida albicans causes candidiasis, whereas Microsporum gypseum and Trichophyton mentagrophytes cause dermatophytosis (Moskaluk & VandeWoude, 2022). Aspergillus niger is another fungal pathogen that affects the respiratory system by causing different diseases such as aspergillosis (Fiema et al., 2022). Fortunately, essential oils can be developed as antifungal agents for preventing these. Most compounds derived from essential oils are terpenes and their metabolites derivatives (D’agostino et al., 2019), such as α-guaiene, a sesquiterpene comprising an average of 11% of the total mass (Orf et al., 2021). Patchouli oil’s main constituents are patchouli alcohol (27.0%-35%), bulnezen (13.0%-21.0%), and α-guaiene (11.0%-16.0%) (Górski et al., 2021). Terpenes can inhibit protein and DNA synthesis and promote cell rupture in antibiotic-susceptible and antibiotic-resistant bacteria (Masyita et al., 2022).\n\nThe antimicrobial activity of essential oils, notably patchouli oil, has been reported. Patchouli essential oil has been shown to inhibit Malassezia furfur and to be effective as an antimicrobial and anti-inflammatory agents (Srivastava et al., 2022). Treatment of C. albicans and T mentagrophytes with patchouli oil fraction 8 containing 55.59% patchouli alcohol content yielded inhabitation zone of 9.24 mm and 7.70 mm in both fungi, respectively (Setyaningrum et al., 2017).\n\nPatchouli alcohol is the primary antifungal component of patchouli oil and can be used as a fixative in perfume and related industries. After patchouli alcohol extraction, other components such as α-guaiene can increase patchouli oil’s utilization. Studies on α-guaiene antifungal activity are few, necessitating more investigations on this compound. While patchouli alcohol (PA) exhibited antimicrobial properties against pathogenic bacteria (Escherichia coli, Pseudomonas aeruginosa, Bacillus proteus, Shigella dysenteriae, Typhoid bacillus, Staphylococcus aureus) (Yang et al., 2013). In addition, PA proved effective in treating some germs that were resistant to antibiotics, such as methicillin-resistant Staphylococcus aureus (MRSA) (Wan et al., 2021). Therefore, this study explores the antifungal activity of fraction 1 patchouli oil containing 38.8% α-guaiene against C. albicans, A. niger, M. gypseum, and T. mentagrophytes.\n\n\nMethods\n\nPatchouli oil was obtained from distiller in Subang, West Java, Indonesia. The nutrient agar culture used was Potato Dextrose Agar (PDA), whereas the liquid medium was Potato Dextrose Broth (PDB). Other chemicals such as 1% BaCl2, 1% H2SO4, sterile distilled water, 70% alcohol, lactophenol cotton blue (LPCB), 0.85% NaCl, ketoconazole, fluconazole, and n-hexane were also utilized. Chemical compound were obtained from Bratachem, while drugs were obtained from Kimia Farma, Bandung, Indonesia.\n\nThe instruments used included Spinning Band Distillation System Model 36-100 from B/R Instrument USA, autoclave, Erlenmeyer flask, beaker glass, scotch bottle, Bunsen, ose needle, Petri dish, cuvette, spectrophotometer, micropipette, microplate, fin pipet, microscope, laminar airflow, oven, spatula, and vortex. The study was conducted using a laboratory experimental method with descriptive analysis, with ketoconazole serving as the positive control for all fungi tests, fluconazole for M. gypseum and T. mentagrophytes, and n-hexane serving the negative control for the four fungi.\n\nTo obtain the dominant α-guaiene, the sample was fractionated with a pressure of 10 mmHg, column length of 90 cm, and reflux ratio of 20:1. Patchouli oil was divided into five fractions: 1 at 249oC-254oC, 2 at 254oC-259oC, 3 at 259oC-264oC, 4 at 264oC-269oC and 5 at 269oC-274oC. Fraction 4 was suspected to contain α-guaiene as the most predominant content.\n\nGCMS analysis of the sample was carried out on a Agilent® 6890 GC-MS, equipped with a split-spitless injector, attached to an Agilent HP-5MS capillary column (30 m x 250 μm, 0.25 μm film thickness). The carries gas was helium at a flowrate of 1.0 mL/min, split ratio 400:1, injector temperature was 280oC, pressure was 10.48 psi. The transfer line was heated to 280oC. Identification of the oil components was accomplished by comparison of retention times with standard substances.\n\nThe modified microdilution technique was used to determine the antifungal activity of the sample.\n\nConidia were removed from the agar slant surfaces using sterile 0.85% saline containing 0.1% Tween 20 (vol/vol). In a final amount of 100 μL per well, the conidia suspension was adjusted with sterile 0.85% saline to a concentration of roughly 1.0 × 10.5. For later usage, the inocula were stored at -20°C. To ensure there was no contamination and to confirm the inocula’s validity, dilutions of the inocula were cultivated on the solid MEA.\n\nUsing 96-well microtitre plates and a serial dilution approach, the MIC was determined. Fungal inoculums (10 μL) were added to varying volumes of the studied material that had been dissolved in malt extract broth (MEB). The microplates were kept at 28°C for 72 hours. The concentration that totally inhibited fungal growth (MIC) was the lowest concentration at which no growth was discernible. Serial subcultivation of 2 μL onto microtitre plates containing 100 μL of MEB allowed for the determination of the minimal fungicidal concentration (MFC). The MFC, which indicates 99.5% death of the origin inoculum, was the lowest concentration with no discernible growth.\n\n\nResults and Discussion\n\nTo obtain the dominant α-guaiene, the patchouli oil was fractionated using a BR Instrument Spinning Band Distillation System Model 36-100 and divided into five fractions. Fraction 4 was suspected to contain α-guaiene as the pre-dominant content. However, based on the GC-MS test results, the compound was higher in fraction 1 than in the others. After fractionation, the fractions were tested for antifungal activity.\n\nAs determined by GC-MS test conducted on fraction 1 and 4, fraction 1 contained a significantly higher α-guaiene content than fraction 4. The percentage of α-guaiene was detected at peak 2 with retention time at 21.837 min. whereas that of fraction 4 was detected at peak 1 with retention time at 21.293 min. The percentage of α-guaiene content was calculated by dividing the peak area by the total area of the compound formed. Based on these calculations, the percentages in fractions 1 and 4 were 38.8% and 2.1% respectively. Subsequently, fraction 1 patchouli oil was used as the antifungal activity test material. Figure 1 show fragmentation pattern of α-guaiene.\n\nFungi were macroscopically and microscopically identified. Macroscopic identification involved observation with the eyes in order to directly identify the physical appearance ( Figure 2). Figure 2 (Line 1) show the physical appearance of C. albicans and A. niger grown on PDA medium. C. albicans cultures were white and formed round colonies, corroborating Lee et al’s finding (Le et al., 2022), that C. albicans has a round shape. Allen et al. (2018) reported that A. niger culture exhibited white mycelium and brown-black conidia heads. Despite its higher colony size, A. niger had less dense colonies than C. albicans, which tend to form a firm line based on the groove of the ose needle stroke. In addition, C. albicans colonies were spherical, and had a relatively uniform size without hyphae due to their unicellularity, whereas A. niger colonies were irregularly spherical, had a nonuniform size, and formed hyphae owing to their multicellularity.\n\nThe colony of the fungus suspected to be M. gypseum resembled a pile of fine white cotton on top with a little brown powder scattered on the hyphae, corroborating a previous finding (Putriningsih & Arjentinia, 2018) that M. gypseum colonies grew rapidly, were slightly powdery with a blackish-red brown color, and were scattered with a flat surface containing macroconidia.\n\nMacroscopically, although M. gypseum and T. mentagrophytes did not differ significantly, having white hyphae, T. mentagrophytes colonies appeared denser and slightly whiter with a protruding rough or powdery surface corroborating Frías-De-León et al’s findings (Frías-De-León et al., 2020) that T. mentagrophytes colonies were often white to slightly yellowish-white and could sometimes turn violet-red, brown, or pale yellowish with a surface resembling cotton, wax wovwn or granules.\n\nThe four test fungi were also identified microscopically (1000x) to corroborate their characteristics. A. niger, C. albicans, M. gypseum and T. mentagrophytes shared similar characteristics with the same fungi theoretically. Figure 3 showed the identification results of the test. The A. niger shows that this fungus has an elongated shape with visible fungal part such as conidia, vesicles, and conidiophores that were obvious by the staining process. Meanwhile, Figure 3 depicting the C. albicans compared to A. niger. C. albicans has a variable round shape with a bluish color and nonuniform cell size. Each cell shows a different individual because this fungus is a unicellular microorganism.\n\nMIC test on A. niger\n\nThe MIC test results for A. niger showed that well with a clear appearance only occurred at 60% concentration treatment, whereas those at 30%, 15%, 7.5% and 3.75% were cloudy with fungal colonies on the surface medium. These observations are relatively weak compared to other studies, such as Yanti et al. (2017) which examined the essential antifungal test on kaffir lime against five type of Aspergillus fungi, revealing an inhibitory mechanism as delayed spore germination and the formation of mycelia at a concentration of 0.05%.\n\nThe first observation results showed that the mechanism for fungal growth inhibition occurred only at 60%, indicating that the MIC value was reached at this concentration. Because the range of values between treatments was large, the MIC value may be reach before 60%, specially between 30% and 60%. The second testing process was conducted to minimize concentration different between treatments. Consequently, treatments with new concentrations were made, namely 45%, 50%, 55%, 60% and 65%. A fraction above 60% was intended to predict when the MFC value was not achieved.\n\nAs seen in Table 1, the well had a clear appearance before reaching a concentration of 60%, specifically at 55%. This concentration inhibited the growth of A. niger, as evidenced by its clear appearance, indicating that fungal growth was inhibited. Because treatment above 55% showed similar results, the MIC value was determined at this concentration.\n\nObservation on each control in the second MIC test showed similar results to the first. The positive control of 2% ketoconazole had a clear appearance, indicating that this compound had antifungal activity. The control medium had a clear appearance, showing an absence of fungal or microbial growth, whereas the growth control had a cloudy appearance, indicating that the fungus grew in this treatment. The negative control n-hexane also had a cloudy appearance, suggesting that this compound lacked antifungal activity against A. niger.\n\nMIC test on C. albicans\n\nObservation data revealed that none of the treatments produced perfectly transparent wells in C. albicans. In general, a higher concentration implies a clearer well, but at the highest value of 40%, fungal growth was still visible, warranting a re-test to determine perfectly transparent wells. The control treatment on C. albicans yielded similar result to the MIC test on A. niger. The second test was conducted with treatment concentrations of 35%, 40%, 45%, 50% and 55%. The results showed that a clear appearance began to develop at a concentration of 45%, whereas the treatment at 35% and 45% had a cloudy appearance, suggesting fungal growth. The difference in concentration appeared to have affected the presence of fungi, corroborating a previous study (Górski et al., 2021) that tested the antifungal activity of patchouli oil at a concentration of 12.5%, 25%, 50% and 100% against C. albicans.\n\nBased on the observations, the MIC value was determined at a concentration of 45%, the result obtained was better than that of Kamoda (Kamoda et al., 2020), stated that the MIC of the red galangal ethanolic extract against the fungus was achieved at 200 mg/mL, but the inhibition only reached at 60%. Differences in antifungal activity for each treatment with varying concentrations also reported by Ningtias et al. (2020), where the MIC value of black garlic extract against C. albicans was reached at a concentration of 50%, but active inhibition was fully achieved at 75%.\n\nMIC test on M. gypseum\n\nThe MIC observation test on M. gypseum fungus with concentration used in twofolds, notably 40%, 20%, 10%, 5% and 2.5% revealed fungal growth in all treatments. Positive and medium controls lacked fungal development, but negative and growth controls demonstrated the opposite. Furthermore, the first observation showed that the highest concentration exhibited no inhibition, although the technique was correctly executed, as indicated by the control treatment, which showed appropriate results. The second test was conducted by increasing the upper limit of concentration and decreasing it with a value range of 5%, hence the new concentration used were 60%, 55%, 50%, 45%, and 40%.\n\nThe second MIC test revealed no fugal growth in the wells containing 50%, 55% and 60% of the teat compound, indicating maximum inhibition at these concentrations. The treatments wells overgrown as the number of fungal colonies that grew on the surface increased. Fungal growth was observed at 45% but was not as cloudy as that of 40%, suggesting that a 45% concentration inhibited growth insignificantly. The MIC value is often indicated by the smallest concentration that can inhibit total fungal growth based on visualization, it was determined at 50%.\n\nPositive controls with 2% ketoconazole and fluconazole inhibited fungal growth, as evidence by a clear appearance with a small amount of nonhomogeneous antibiotic precipitate. However, the negative control of n-hexane showed fungal growth, indicating that this compound lacked antifungal activity. The growth control also exhibited fungal growth, suggesting that growth on PDB media was not inhibited. Notably, the control media used was clear, indicating that no undesirable microorganism were present.\n\nMIC test on T. mentagrophyhes\n\nThe MIC test results for T. mentagrophytes at concentration of 100%, 50%, 25%, 12.5%, and 6.25% of the test compounds showed that only 100% concentration treatment inhibited fungal growth. At 50%, 25%, 12.5% and 6.25% fungal growth remained on the surface, although in varying quantities. The inhibition activity of the test substance at a certain concentration was reflected in the appearance of the media, suggesting the presence or absence of microbes.\n\nIn the first test, the MIC value was determined at a concentration of 100%. Because the range of concentrations capable of inhibiting fungal growth was extremely large, the MIC value could be obtained between 50% and 100%. The second test was conducted by minimizing the concentration difference, resulting in values of 100%, 95%, 90%, 85% and 80%.\n\nThe second MIC observation test results showed that the wells with 95% and 100% concentration exhibited no fungal or other microbial growth, as indicated by their clear color surface. However, concentration of 90%, 85% and 80% showed varying fungal growth for each treatment. The MIC value is often indicated by the smallest concentration that can inhibit the total growth of the fungi or the appearance of clear media on visualization. Based on the results, the value was determined at 95%.\n\nBecause the control treatment for T. mentagrophytes yielded identical results to those for M. gypseum, the technique used was appropriate. The difference in MIC values demonstrated that fungus resistance to fraction 1 as an antifungal agent differed for each fungal species. T. mentagrophytes value exceeded that of M. gypseum. These results are directly proportional to the previous inhibition zone test which showed that T. mentagrophytes was more resistant than M. gypseum to the fraction 1 antifungal agent containing 38.8% α-guaiene\n\nThe MFC test results for A. niger, C. albicans, M. gypseum and T. mentagrophytes revealed fungal growth in wells with a clear medium. Observations were made by creating a new inoculum from only the clear-appearing MIC test results. In order to reduce material requirements and minimize the possibility of errors as wells with cloudy appearance can be ascertained to have fungal growth.\n\nMFC test on A. niger\n\nThe MFC test results for A. niger shown in Table 2 show that only the 55% concentration of the tree treatments indicated the present of fungal growth. Five of six inoculums created were colonized by fungus. Moreover, the number of fungal colonies in one petri disc was less that 5, this growth was relatively small compared to the original culture without treatment, indicating that the test substance inhibited A. niger growth at 55% but did not eradicate this fungus completely. Treatment concentration of 60% and 65% showed no fungal growth in each created inoculum. Based on these results, the MFC value was determined at a concentration of 60%.\n\nThe inoculation wells with the control treatment yielded the expected outcomes under the expected conditions. The positive and medium controls lacked fungal growth, whereas the negative and growth controls indicated opposite results. Moreover, the growth control treatment showed the strongest fungal growth due to the absence of effective antifungal effects.\n\nMFC test on C. albicans\n\nThe MFC test results for C. albicans presented in Table 2 indicated that the 45% treatment exhibited fungal growth compared to the 50% and 55% treatments. Compared to the control treatment, the number of colonies was much lower for the 45% treatment. Based on these observations, the MFC value was determined at a concentration of 50%.\n\nC. albicans’s control treatment yielded similar results to those of A. niger. However, the positive control with 2% ketoconazole exhibited no fungal growth. Ketoconazole, a commonly used conventional antifungal agent exhibited antifungal activity that was significantly more potent than fraction 1 of patchouli oil. The results indicate that a concentration of at most 2% ketoconazole can eradicate fungi.\n\nMFC test on M. gypseum\n\nMFC test was conducted at three concentrations, notably 50%, 55% and 60% which were found to inhibit fungal growth in the previous MIC test. The results showed that, at 50% fungal growth remained on the PDA media in all petri dishes. At 55%, four petri dishes were still observed to be overgrown with fungus, whereas the other two were clear. The number of colonies form in each petri dish at 50% and 55% did not exceed three, this growth was relatively small, indicating that the antifungal agent only inhibited fungal growth but did not eradicate M gypseum. In addition, at 60%, fungal growth was unobserved in all petri dishes; hence, the MFC value was determined at this concentration. The MFC test results for the control treatment were consistent with the prior MIC test. The positive and medium controls exhibited no fungal growth compared to the negative and growth controls.\n\nMIC test on T. mentagrophytes\n\nMFC testing on T. mentagrophytes was conducted at two concentrations, notably 95% and 100%, which were found to inhibit fungal growth in the previous MIC test. The results showed that, at 95%, five petri dishes were still overgrown with fungus, whereas one was not. The number of colonies formed was not significantly high; hence, the antifungal agent appeared to only inhibit but not eradicate fungi. At 100%, fungal growth was unobserved in all Petri dishes; hence, the MIC value was determined at this concentration. The control treatment for the MFC test on T. mentagrophytes yielded similar results to those of M. gypseum. The positive control at 2% ketoconazole and fluconazole did not support fungal growth. This suggests that 2% ketoconazole and fluconazole as conventional antifungal agents, have significantly more potent effects than fraction 1 of patchouli oil.\n\n\nConclusions\n\nFraction 1 of patchouli oil containing 38.8% α-guaiene exhibited antifungal activity against A. niger, C. albicans, M. gypseum and T. mentagrophytes. The MIC values for each fungus were reached at concentrations of 55%, 45%, 50% and 95%, whereas the MFC value were achieved at 60%, 50%, 60% and 100%, respectively. This study demonstrates that α-guaiene is a prospective agent effective against the investigated pathogenic fungus.", "appendix": "Data availability\n\nFigshare: Data of MIC and MFC of alpha-guaiene againts four pathogenic fungi, Doi: 10.6084/m9.figshare.27925854.v1 (Nurjanah et al., 2024).\n\nThis project contains the following underlying data:\n\n• MIC of four pathogenic fungi.xlsx\n\n• MFC of four pathogenic fungi.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\nAlen Y, Melati A, Sarina G, et al.: Isolation of Secondary Metabolite A. niger “In-Habiting” Queen M. gilvus Hagen.’s Nest. Indones. J. Pharm. Sci. Technol. 2018; 5(2): 61. Publisher Full Text\n\nAlmeida RN, Soares R d P, Cassel E: FRACTIONATION PROCESS OF ESSENTIAL OILS BY BATCH DISTILLATION. Braz. J. Chem. Eng. 2018; 35(3): 1129–1140. Publisher Full Text\n\nD’agostino T, Frippiat M, Debourgogne: Essential Oils and Their Natural Active Compounds Presenting Antifungal Properties. Molecules. 2019; 24(20): 3713. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFiema M, Wlodarczyk A, Wojkowska-Mach J, et al.: Atypical Presentation of Aspergillus niger Infection in the Oral Cavity as a Prediction of Invasive Pulmonary Aspergillosis in a Patient with COVID-19: Case Report and Literature Review. Microorganisms. 2022; 10(8): 1630. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFrías-De-León MG, Martínez-Herrera E, Atoche-Diéguez CE, et al.: Molecular identification of isolates of the Trichophyton mentagrophytes complex. Int. J. Med. Sci. 2020; 17(1): 45–52. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGórski R, Dorna H, Rosińska A, et al.: Effects of Essential Oils on in vitro Growth of Fungi Cladobotryum dendroides and Mycogone perniciosa Infecting Button Mushroom. Ecol. Chem. Eng. S. 2021; 28(3): 411–427. Publisher Full Text\n\nJain PLB, Patel SR, Desai MA: Patchouli oil: an overview on extraction method, composition and biological activities. J. Essent. Oil Res. 2022; 34(1): 1–11. Publisher Full Text\n\nKamoda H, Lelyana S, Sugiaman VK: Kadar hambat minimum dan kadar bunuh minimum ekstrak etanol lengkuas merah (Alpinia galanga L.) terhadap pertumbuhan Candida albicans. J. Kedokt. Gigi Univ. Padjadjaran. 2020; 32(1): 1. Publisher Full Text\n\nKhaledi N, Zahani FH: Biological Effects of Various Essential Oils on Citrus Decay Pathogens.2018. Reference Source\n\nLe PH, Nguyen DHK, Medina AA, et al.: Surface Architecture Influences the Rigidity of Candida albicans Cells. Nanomaterials. 2022; 12(3). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLeong W, Huang G, Khan I, et al.: Patchouli essential oil and its derived compounds revealed prebiotic-like effects in C57BL/6J mice. Front. Pharmacol. 2019; 10(OCT). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMasyita A, Mustika Sari R, Dwi Astuti A, et al.: Terpenes and terpenoids as main bioactive compounds of essential oils, their roles in human health and potential application as natural food preservatives. Food Chem. X. 2022; 13: 100217. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMoskaluk AE, VandeWoude S: Current Topics in Dermatophyte Classification and Clinical Diagnosis. Pathogens. 2022; 11(9): 957. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNingtias RI, Setyowati DI, Handayani ATW: Efektivitas Ekstrak Black Garlic dalam Menghambat Pertumbuhan Candida albicans. Pustaka Kesehatan. 2020; 8(3): 153. Publisher Full Text\n\nNurjanah S, Rialita T, Lembong E, et al.: Isolation of guaiene from patchouli (Pogostemon cablin Benth.) oil using vacuum fractionation distillation. IOP Conf. Ser. Earth Environ. Sci. 2020; 443(1): 012094. Publisher Full Text\n\nNurjanah S, Alhafidz Z, Maulani M, et al.: MIC and MFC of four pathogenic fungi.xlsx (Version 1). figshare. 2024. Publisher Full Text\n\nOrf M, Kurian M, Nelson C, et al.: Correlation Gas Chromatographic Study of the Vaporization Enthalpies and Vapor Pressures of the Major Sesquiterpene Hydrocarbons in Patchouli Oil. J. Chem. Eng. Data. 2021; 66(6): 2538–2549. Publisher Full Text\n\nPandey SK, Bhandari S, Sarma N, et al.: Essential oil compositions, pharmacological importance and agro technological practices of Patchouli (Pogostemon cablin Benth.): A review. J. Essent. Oil Bear. Plants. 2021; 24(6): 1212–1226. Publisher Full Text\n\nPandey SK, Gogoi R, Bhandari S, et al.: A Comparative Study on Chemical Composition, Pharmacological Potential and Toxicity of Pogostemon cablin Linn., (Patchouli) Flower and Leaf Essential Oil. J. Essent. Oil-Bear. Plants. 2022; 25(1): 160–179. Publisher Full Text\n\nPutriningsih PAS, Arjentinia IPGY: Identifikasi Spesies Fungi Microsporum gypseum dan M. nanum Penyebab Ringworm pada Sapi Bali (IDENTIFICATION OF SPECIES FUNGI MICROSPORUM GYPSEUM). Jurnal Veteriner. 2018; 19(2): 177. Publisher Full Text\n\nRahmayanti D, Hadiguna RA, Santosa S, et al.: Determining The Profit Margin In “Patchouli Oil” Supply Chain: A Case Study In Indonesia. Int. J. Adv. Sci. Eng. Inf. Technol. 2018; 8(2): 483. Publisher Full Text\n\nSetyaningrum PR, Nurjanah S, Widyasanti A, et al.: Uji Aktivitas Antijamur pada Minyak Nilam Hasil Destilasi dan Fraksinasi Terhadap Jamur C. albicans dan T. mentagrophytes. Jurnal Teknotan. 2017; 11(1). Publisher Full Text\n\nSrivastava S, Lal RK, Singh VR, et al.: Chemical investigation and biological activities of Patchouli (Pogostemon cablin (Blanco) Benth) essential oil. Ind. Crop. Prod. 2022; 188: 115504. Publisher Full Text\n\nvan Beek TA , Joulain D: The essential oil of patchouli, Pogostemon cablin: A review. Flavour Fragr. J. John Wiley and Sons Ltd.; 2018; 33(1): 6–51. Publisher Full Text\n\nWan F, Peng F, Xiong L, et al.: In Vitro and in vivo Antibacterial Activity of Patchouli Alcohol from Pogostemon cablin. Chin. J. Integr. Med. 2021; 27(2): 125–130. PubMed Abstract \| Publisher Full Text\n\nYang X, Zhang X, Yang S-P, et al.: Evaluation of the Antibacterial Activity of Patchouli Oil. Iran. J. Pharm. Res. 2013; 12(3).\n\nYanti R, Wulandari P, Pranoto Y, et al.: Karakterisasi, Identifikasi dan Uji Aktivitas Anti Jamur Minyak Atsiri Daun Jeruk Purut (Citrus hystrix) terhadap Aspergillus. Jurnal Teknologi Pertanian. 2017; 8(2)." }	[ { "id": "356456", "date": "07 Jan 2025", "name": "Irmanida Batubara", "expertise": [ "Reviewer Expertise analytical chemistry" ], "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 need to add following: 1. Please add the background why the guaiene concentration only at 38.8%. What is the important, how if lower or higher concentration of guaiene? 2. Why the author did not compare the activity with only PA or with the patchouli oil before fractionation. 3. The author more to clarify the fungi name compare to discuss about the activity. 4. It is important to explain the mechanism of the sample as antifungi. 5. In the method for GCMS, the author compare the retention time, but in the result author used MS data. Please make it clear 6. Please add information how many repetition for each measurement 7. It is better to make table about MIC and MFC value in one table not prepare the observation result. 8. Please compare the result with positive control as well 9. in the abstract only MIC value but have two values?\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": [] }, { "id": "356464", "date": "16 Jan 2025", "name": "Harlinda Kuspradini", "expertise": [ "Reviewer Expertise Medicinal and Aromatic Plants" ], "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 is well organized but needs improvement.\nThe discussion lacks information on the antifungal mechanism of α-guaiene and does not compare it with patchouli alcohol (PA) or patchouli oil, which would strengthen the argument. The effect of α-guaiene concentration on its antifungal activity is unclear. Include the number of replicates and the statistical analysis method used to validate the results. The method and results sections are inconsistent, particularly in presenting the GC-MS data. The results show that α-guaiene has antifungal properties. It should specify which fungi, α-guaiene is most effective against and discuss its limitations for broader use. Fix some minor inconsistencies in the use of italics for scientific names.\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": [] } ]	1	https://f1000research.com/articles/14-4
https://f1000research.com/articles/13-1131/v1	04 Oct 24	{ "type": "Research Article", "title": "Current scenario of machine learning applications to hydrothermal liquefaction via bibliometric analysis", "authors": [ "Tossapon Katongtung", "Somboon Sukpancharoen", "Sakprayut Sinthupinyo", "Nakorn Tippayawong", "Tossapon Katongtung", "Somboon Sukpancharoen", "Sakprayut Sinthupinyo" ], "abstract": "Background Energy shortages and global warming have been significant issues throughout history. Therefore, the search for environmentally friendly renewable energy sources is crucial for achieving sustainability. Biomass energy is gaining global attention as a renewable energy option, particularly through the process of hydrothermal liquefaction, which converts biomass into bio-crude oil.\n\nMethods Hydrothermal liquefaction is a complex process that is challenging to explain, leading to research on machine learning models for this process. These models aim to predict values and investigate the impact of variables on the hydrothermal liquefaction process. However, the development of machine learning in hydrothermal liquefaction is still limited due to its novelty and the time required for comprehensive study. Thus, the objective of this study was to analyze relevant publications in the Scopus database, focusing on indexed ML and HTL keywords, to understand keyword associations and co-citations.\n\nResults The results reveal an increasing trend in the study of ML in the HTL process, with a growing interest from various countries.\n\nConclusion Notably, China currently holds the largest share of ML research in HTL processes, with most published works falling within the field of engineering. The keyword “liquefaction” emerges as the most popular term in these publications.", "keywords": [ "AI", "Clean energy", "Data analytics", "Climate action", "bibliometric analysis" ], "content": "Introduction\n\nCurrently, numerous countries worldwide are actively seeking alternative energy sources to replace depleted resources and ensure environmental preservation amidst the era of global warming.1–3 Biomass stands as a prominent alternative in the realm of renewable energy, referred to as biomass energy.4 Biomass energy refers to the energy derived from various organic materials that serve as natural energy sources. These materials encompass a wide range of substances, including organic waste, agricultural residues, industrial byproducts, manure, and fuel crops such as rice husks, rice straw, bagasse, sugar cane leaves and shoots, wood, wood chips, fibers, palm kernel shells, cassava residue, corn cobs, husks, coconut shells, and more. The conversion of biomass into usable energy involves processes such as fermentation, combustion, hydrothermal treatment, or other methodologies, which transform biomass into heat or gas for energy utilization.5,6 Additionally, there exists a biomass processing method known as hydrothermal liquefaction, which converts wet biomass into bio-crude oil. This process involves subjecting the biomass to high-temperature and high-pressure conditions in a liquid water environment, resulting in the production of bio-crude oil.7,8\n\nHydrothermal liquefaction (HTL) is a thermal depolymerization process that facilitates the conversion of wet biomass and other macromolecules into crude oil, commonly known as bio-crude oil. This transformation occurs under medium temperature and high-pressure conditions, enabling the efficient conversion of the biomass feedstock.9,10 Bio-crude oil exhibits a high energy density, characterized by a higher heating value ranging from 33.8 to 36.9 MJ/kg. It typically contains 5-20 wt% oxygen, along with renewable chemicals. This composition makes bio-crude oil an attractive renewable energy source with significant potential for various applications.11 The study of various variables and their effects under HTL processing conditions is challenging and intricate. Consequently, researchers worldwide have shown keen interest in investigating the behavior of variables within the HTL environment. Machine learning has emerged as the most popular technology to address such difficult and complex problems efficiently. It offers promising avenues for understanding and optimizing HTL processes through data-driven analysis and predictive modeling.\n\nMachine Learning (ML) can be considered as the learning component of a machine. It serves as the foundational element of Artificial Intelligence (AI), acting as the “brain” that enables AI systems to acquire knowledge and exhibit intelligent behavior. AI utilizes ML techniques to develop and enhance its intelligent capabilities, allowing it to learn from data, recognize patterns, make predictions, and adapt to new information or situations. In essence, ML forms an integral part of the AI framework, enabling the creation and manifestation of intelligence in AI systems.12,13 Indeed, ML is commonly associated with the learning models of artificial intelligence. Unlike traditional programming, where humans explicitly define rules and instructions, ML involves programming AI systems to learn from data. Once programmed, the machine leverages the available data to train and refine its own intelligence through practice and iterative processes. It gradually improves its performance and ability to make informed decisions without explicit human intervention. ML has found extensive applications across various domains, including medicine, education, economy, and engineering. In medicine, it is used for diagnostics, personalized treatments, and drug discovery. In education, it aids in adaptive learning and intelligent tutoring systems. In the economy, it assists with data analysis, forecasting, and fraud detection. In engineering, it facilitates automation, optimization, and predictive maintenance. The widespread adoption of ML in both research and everyday life has opened up new possibilities and opportunities for leveraging data-driven intelligence in solving complex problems and making informed decisions.13–18\n\nMoreover, ML is extensively applied in the field of energy to predict and establish correlations among variables in highly complex processes that cannot be easily explained through conventional means. For instance, a study by Onsree and Tippayawong (2020) demonstrated the development of a ML application for predicting yields of solid products obtained from biomass torrefaction. This model achieved a remarkable accuracy with an R2 value of approximately 0.9. Furthermore, the application also shed light on the interrelationships between variables within the torrefaction process, providing valuable insights into the underlying mechanisms. Such applications of ML in energy-related research not only enhance predictive capabilities but also contribute to a deeper understanding of complex processes and their underlying dynamics.19 Additionally, Phromphithak et al. (2021) conducted a study where they employed ML techniques to predict the production of cellulose-rich materials during biomass pretreatment using ionic liquid solvents. The developed model exhibited a high accuracy, with an R2 value of 0.94. Moreover, the study successfully elucidated the behavior of ionic liquids and their impact on the ML -based pretreatment process. Ionic liquid (IL) pretreatment is an effective method for processing lignocellulosic biomass, enhancing its conversion into biofuels and chemicals. ILs, which are salts in liquid form at low temperatures, can disrupt the complex structure of biomass by dissolving lignin and reducing the crystallinity of cellulose. This makes cellulose more accessible for enzymatic hydrolysis or catalytic processes. ILs offer the advantage of selective dissolution, allowing for better isolation of cellulose and hemicellulose. Furthermore, ILs can be recycled, reducing waste and costs, making them a promising option for sustainable biomass pretreatment. This research demonstrates the efficacy of ML in providing accurate predictions and uncovering the intricate relationships between variables, leading to a better understanding of the effects of ionic liquids in biomass.20 In a study by Prasertpong et al. (2022), the researchers investigated the synergistic effects observed during the co-pyrolysis of biomass and plastic waste using ML techniques. The aim of the study was to gain insights into the variables that influence the co-pyrolysis process. By employing machine learning, the researchers were able to uncover and comprehend the intricate relationships and interactions between these variables, shedding light on the synergistic effects observed during the co-pyrolysis process. This study highlights the potential of ML in providing valuable insights and understanding complex processes involving biomass and plastic waste co-pyrolysis.21 Furthermore, there have been studies that utilize ML in the context of HTL processes. In a previous research endeavor, an ML model was developed for predicting biocrude yields and higher heating values obtained from HTL of wet biomass and waste materials. The model achieved a high precision with an R2 value of nearly 0.9. Additionally, it elucidated the relationships among variables that directly and indirectly impact the HTL process. This study showcases the potential of ML in accurately predicting and comprehending the complex interplay of variables in HTL processes, contributing to a deeper understanding of the factors influencing HTL and enabling more efficient and effective utilization of wet biomass and waste for biocrude production.22 There are other studies related to ML in HTL processes a number of similar.23,24 Indeed, the development of ML in the field of HTL is relatively new and holds great potential for further study and exploration in the future. Consequently, the collection and comprehensive search for research pertaining to the development of ML in the HTL process become crucial and should be given significant emphasis. This endeavor would facilitate the identification of existing studies, trends, and advancements in the application of ML techniques specifically for HTL, thereby laying the groundwork for future research and advancements in this domain. By focusing on this area of research, researchers can deepen their understanding of the potential of ML in HTL and further contribute to its development and utilization in the pursuit of sustainable and efficient energy production.\n\nBibliometric analysis involves a systematic statistical examination of research data extracted from extensive databases, aiming to evaluate various aspects of scholarly work. This analysis serves as a valuable tool for measuring research quality, including the productivity and impact of individual researchers and institutions. By employing bibliometric methods, researchers can quantitatively assess factors such as publication output, citation counts, collaboration networks, journal rankings, and other bibliographic indicators. These analyses provide valuable insights into the performance and influence of researchers and institutions within specific fields of study, contributing to the evaluation and comparison of research outcomes across various domains.25 Bibliometric analysis is a research methodology that emphasizes quantitative and statistical investigations. It enables the description and analysis of various aspects of scholarly publications or literature, such as the type of publication, the countries, institutions, and authors associated with the research. By conducting long-term studies, bibliometric analysis allows researchers to identify trends in the growth or decline of research within specific fields. Consequently, bibliometric data serves as a valuable foundation for monitoring scientific and technological research, providing insights into the progress and development of knowledge in various domains.26 Bibliographic analysis typically involves selecting a set of keywords to search relevant literature databases. These keywords are chosen based on the topic and title of the articles to retrieve information related to the conceptual framework and writing style within a specific field of study. Electronic bibliographic databases offer vast opportunities for efficient and rapid access to a wide range of scholarly resources. By utilizing these databases, researchers can benefit from an extensive collection of literature and gain quicker access to relevant information for their studies. This enables more efficient literature review and analysis, aiding in the advancement of research in a particular area.27\n\nThe objective of this study was to perform a bibliographic analysis using publications related to ML and HTL keywords indexed in the Scopus database. The analysis employed a quantitative methodology to examine the bibliography of published articles in the field. To accomplish this objective, the study utilized VOSviewer version 1.6.20, a bibliometric analysis tool, for conducting the bibliographic analysis. VOSviewer enables visualizations and quantitative analyses of bibliographic data, facilitating the exploration of relationships, trends, and patterns within the literature.28,29 Based on the available literature review, this study represents the first bibliographic analysis conducted to evaluate research trends specifically in the development of ML within the context of HTL. As the research on HTL-based ML development is still in its early stages and relatively limited at present, this study fills an important knowledge gap in the field. The outcomes of this analysis will contribute significantly to the formulation of a comprehensive research plan by identifying potential future research directions and fostering collaborative relationships among researchers in this emerging field.\n\n\nMethods\n\nBibliometrics is a research methodology that involves the analysis of quantitative and statistical data. It is commonly used to examine various relationships, such as the connections between authors, the relationship between a research subject and an author’s work, scholarly works themselves, scholarly citations, and citation tracking. By employing bibliometric techniques, researchers can gain insights into the patterns and dynamics of scholarly communication, collaboration, and impact within a particular field or discipline. It provides a systematic approach to understanding the scholarly landscape and can be utilized to assess research productivity, influence, and trends.30 Bibliometric analysis offers a distinct perspective that complements more comprehensive analyses. It enables researchers to categorize and analyze extensive amounts of data derived from research conducted over a specific timeframe. By employing quantitative techniques, bibliometric analysis can help mitigate or minimize researcher bias, unlike systematic reviews that often rely on qualitative methods. The objectivity of bibliometric analysis reduces the potential for interpretation bias introduced by scholars from diverse academic backgrounds. This allows for a more objective and standardized assessment of research output, impact, and trends, providing valuable insights into the scholarly landscape.28,31\n\nIn this study, bibliographic analysis was employed to investigate and validate the prevailing trends in the research on ML development within HTL processes. The Scopus database was selected for this analysis due to its widespread usage, reliability, and comprehensive coverage across various disciplines. Scopus is known for its extensive collection of scholarly literature, making it a suitable choice for conducting bibliometric studies across diverse research areas. By utilizing the Scopus database, the study could effectively assess and examine the current state of ML development in HTL processes and gain valuable insights into the research landscape in this domain.32,33 Indeed, the databases available in Scopus offer access to a vast number of documents, providing a wealth of information for bibliometric analysis. Scopus is renowned for its comprehensive coverage of scholarly literature, encompassing a wide range of disciplines and research fields. This extensive coverage ensures that researchers can access a substantial volume of documents relevant to their study. Furthermore, Scopus provides robust citation information, including citation counts and citation networks, which can be crucial for analyzing the impact and influence of research articles. The availability of comprehensive document and citation data in Scopus enhance the accuracy and reliability of bibliometric analyses and contribute to a more comprehensive understanding of research trends and dynamics.34 Scopus is a comprehensive database that provides researchers with a range of resources for exploring and evaluating scholarly publications, patents, clinical trials, and policy documents. In this study, the Scopus database was utilized to conduct the bibliographic analysis. Keywords were employed to search for relevant publications based on their publication name, abstract, and author-provided keywords. The search criteria encompassed a set of specific keywords related to “Machine Learning,” “Deep Learning,” “Neural Network,” “Artificial Intelligence,” “ML,” “DL,” “NN,” and “AI,” as well as keywords related to “Hydrothermal Liquefaction,” “Liquefaction,” and “HTL.” The subject areas covered in this study include energy, biomass, engineering, computer science, chemistry, biology, and the environment. The search keywords were set to include publications from the year 1955 up to June 2023, with data downloaded on June 21, 2023. The inclusion criteria for this study encompassed original articles published in the English language. These specific parameters and criteria were employed to ensure a focused and comprehensive analysis of the relevant literature in the field of ML development in HTL processes. Figure 1 shows the search technique used in this study to identify appropriate articles from the Scopus database. The complete bibliography data was downloaded in.csv format from the Scopus database. VOSviewer was used in bibliometric analysis in this work.\n\n\nResults\n\nTable 1 provides an overview of the number of documents related to ML in the HTL process, categorized by year of publication from 1955 to 2023. The table shows the number of documents found for each year, indicating the volume of research conducted in this area. Figure 2 illustrates the trend in the number of documents related to ML in the HTL process per year. The graph indicates a consistent increase in the number of publications since 2003, with a significant surge observed from 2018 onwards. Notably, in 2018, 2019, 2020, 2021, and 2022, the number of documents were 63, 100, 105, 132, and 158, respectively. This trend suggests a growing interest and popularity in the study of ML in the HTL process, particularly in recent years. The findings from Figure 2 indicate that ML in the HTL process has garnered substantial attention and continues to gain momentum, highlighting its significance and potential for further advancements in research and applications.\n\n(via Scopus on July 1, 2023).\n\nBased on the analysis of search results related to ML in the HTL process, it was possible to rank the top 10 in terms of subject area, author name, source title, and country. Engineering emerged as the most published subject area, with 350 publications, as depicted in Figure 3. Among the authors, Samui, P, stood out with the highest number of publications on ML in HTL processes, totaling 20 publications, as shown in Figure 4. The most frequently published source title was “Bioresource Technology,” with a notable 43 publications, as illustrated in Figure 5. China emerged as the country with the highest number of publications on the studied topic, with a total of 407 publications, as depicted in Figure 6.\n\nFrom Figure 3, it can be seen that China ranks first in publications on ML in HTL processes, which has more than twice that of the second place. Figure 3 reveals a diverse range of disciplines showing interest in studying ML in HTL processes. Engineering emerges as the most engaged subject, constituting 13.5% or 350 publications. Following closely behind are Earth and Planetary Sciences and Energy, accounting for 10.5% or 272 publications. This demonstrates a shift towards adapting to human needs in environmental preservation and exploring clean energy sources through HTL. The application of ML as a tool enables time and resource savings in experimental endeavors, reinforcing its significance in this field.22\n\nThe bibliometric analysis in this study utilized the VOSviewer program to construct cluster maps and identify relationships within the database. The cluster map construction involved three aspects: co-citation correlation, keyword correlation, and published countries.\n\nFigure 7 displays the cluster map based on co-authorship authors, revealing connections among various author groups. A total of 38 author groups were identified. The analysis indicated that Zhang Y. had the highest number of links, with 21 links and 506 citations. However, it was observed that the number of links did not have a significant impact on citations. Li H.I. and Goh A.T.C. had 20 and 6 links, respectively, but garnered 1048 and 877 citations, respectively.\n\nFigure 8 represents the cluster map based on the co-occurrence of all keywords. It revealed the existence of 5 keyword groups, with a predominant focus on HTL process-related keywords. Interestingly, ML keywords did not form a substantial cluster, potentially due to their relatively lower number compared to the keywords associated with the HTL process chain. The top 5 most common keywords were liquefaction, article, human, soil liquefaction, and male, with frequencies of 590, 308, 219, 203, and 173, respectively. The keyword ML appeared in the 6th position, specifically in the form of “neural networks,” with a frequency of 156.\n\nFigure 9 depicts the cluster map based on co-authorship countries, revealing 13 distinct groupings of countries. Among them, China emerged with the highest number of links, totaling 6,048 citations and 94 links. The United States ranked second in terms of links, accumulating 5,767 citations and 71 links. India secured the third position in links, with 2,193 citations and 39 links.\n\n\nConclusion\n\nThe bibliometric analysis conducted in this study revealed a continuous increase in publications related to ML in HTL processes from 1955 to 2023. It was found that China has been the leading country in terms of publishing research in this area. The “Bioresource Technology” journal emerged as the most popular choice for publishing articles on ML in the HTL process. The field of engineering, particularly with a focus on the keyword “Liquefaction,” garnered the most interest among researchers. These findings provide valuable insights and can serve as a guide for future studies on ML in HTL processes, offering a comprehensive overview of the current state of research in this domain.\n\n\nEthical approval\n\nThe dataset described in this article does not involve any human subjects, animal experiments, or data collected from social media platforms.\n\n\nAuthor roles\n\nKatongtung T: Conceptualization, Data Curation, Formal Analysis, Investigation, Methodology, Project Administration, Validation, Visualization, Writing – Original Draft Preparation; Sukpancharoen S: Formal Analysis, Writing – Review & Editing; Sinthupinyo S: Formal Analysis, Writing – Review & Editing; Tippayawong N: Conceptualization, Supervision, Funding Acquisition, Formal Analysis, Writing – Review & Editing.", "appendix": "Data availability\n\nMendeley Data: Dataset for: Current scenario of machine learning applications to hydrothermal liquefaction via bibliometric analysis, DOI: 10.17632/st4xh92wm2.1. 35\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 first author wishes to thank the Teaching and Research Assistant Scholarships from the CMU Graduate School and the NRCT Research & Researcher for Industry program, Thailand.\n\n\nReferences\n\nWang S, Sun L, Iqbal S: Green financing role on renewable energy dependence and energy transition in E7 economies. 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Publisher Full Text\n\nKatongtung T: Dataset for: Current scenario of machine learning applications to hydrothermal liquefaction via bibliometric analysis. Mendeley Data, V1. 2024. Publisher Full Text" }	[ { "id": "330069", "date": "24 Oct 2024", "name": "Lili Qian", "expertise": [ "Reviewer Expertise My research focused on hydrothermal liquefaction and machine learning. We submitted a similar paper titled \"A Review on Machine Learning-Aided Hydrothermal Liquefaction Based on Bibliometric Analysis\" on August this year." ], "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 performed bibliometric analysis of machine learning applications to hydrothermal liquefaction. However, similar studies have been reported and the following issues should be addressed. A major revision is needed before the indexing. 1 Background in abstract: It is important to emphasize HTL's focus on wet biomass. 2 Methods in abstract: machine learning should be introduced. 3 Second paragraph in introduction: Machine learning has emerged as the most popular technology to address such difficult and complex problems efficiently. What specifically do “such difficult and complex problems” refer to? 4 Fourth paragraph in introduction: There is too much information about ionic liquids. 5 Fifth paragraph in introduction: The application of the bibliometric method in the HTL field has not been introduced, nor has its application in the ML field. 6 “Based on the available literature review, this study represents the first bibliographic analysis conducted to evaluate research trends specifically in the development of ML within the context of HTL.”. It is not the first bibliographic study on ML-based HTL. similar studies have been reported. What is the novelty of the manuscript? 7 Methods: The first paragraph in methods is the introduction of bibliometrics. Move it to the introduction part. 8 Figure 8: why “article” is a keyword? 9 No reference in this year.\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? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "12702", "date": "26 Oct 2024", "name": "Tossapon katongtung", "role": "Author Response", "response": "Thank you for your advice. I can answer the following questions you raised. 1.Background in abstract: It is important to emphasize HTL's focus on wet biomass. Response: Revised, as suggested. 2.Methods in abstract: machine learning should be introduced. Response: Revised, as suggested. 3 Second paragraph in introduction: Machine learning has emerged as the most popular technology to address such difficult and complex problems efficiently. What specifically do “such difficult and complex problems” refer to? Response: This sentence refers to applying ML to processes that are difficult to explain, in this case HTL processes, as in the paragraph, and I believe the following sentence answers this question. “It offers promising avenues for understanding and optimizing HTL processes through data-driven analysis and predictive modeling.” If the reader wishes to elaborate on this, I have added some sentences to the manuscript. 4 Fourth paragraph in introduction: There is too much information about ionic liquids. Response: Revised, as suggested. 5 Fifth paragraph in introduction: The application of the bibliometric method in the HTL field has not been introduced, nor has its application in the ML field. Response: In this sentence, I am confident that I wrote it correctly at that time, because I had been studying this issue since June 2023 and had tried to submit it to a journal for publication but did not receive a response, so I withdrew it and submitted it to F1000research in September 2024. I have kept all my submissions from June 2023. If you want to see them, I can send them to you. 6 “Based on the available literature review, this study represents the first bibliographic analysis conducted to evaluate research trends specifically in the development of ML within the context of HTL.”. It is not the first bibliographic study on ML-based HTL. similar studies have been reported. What is the novelty of the manuscript? Response: I think we have similar ideas on how to work on this, but I must say that at the time I wrote this study, there was no research on this subject before. In this sentence, I am confident that I wrote it correctly at that time, because I had been studying this issue since June 2023 and had tried to submit it to a journal for publication but did not receive a response, so I withdrew it and submitted it to F1000research in September 2024. I have kept all my submissions from June 2023. If you want to see them, I can send them to you. I have submitted Case Studies in Chemical and Environmental Engineering Manuscript Number: CSCEE-D-23-00310 on June 30, 2023. 7 Methods: The first paragraph in methods is the introduction of bibliometrics. Move it to the introduction part. Response: I think this sentence should remain in its original place and should not be changed. 8 Figure 8: why “article” is a keyword? Response: I collected data from Scopus database and did not add any data to the dataset. 9 No reference in this year. Response: I did this study before 2024 and I gave a clear timeline in the manuscript." } ] }, { "id": "341356", "date": "16 Dec 2024", "name": "Muntasir Shahabuddin", "expertise": [ "Reviewer Expertise Techno-economics", "hydrothermal liquefaction", "machine learning", "electrochemical engineering", "reactor 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\nManuscript Title: Current scenario of machine learning applications to hydrothermal liquefaction via bibliometric analysis Overview: The reviewer would like to thank the editors of F1000 for the opportunity to provide feedback on the manuscript “Current scenario of machine learning applications to hydrothermal liquefaction via bibliometric analysis”. This study by Katongtung et al. leverages bibliometric analysis to collect and analyze publications relating to machine learning on hydrothermal liquefaction systems in the corpus of existing literature. Using a Scopus search, the authors uncovered the most productive authors in the field, and geopolitical regions with the greatest academic output. While the conclusions of this study (in which Chinese and US scientists tend to be the most productive in this area from a raw studies-published perspective) are useful, the manuscript may benefit from reformatting of figures for better clarity of the data being communicated, and for reconsideration of how each figure contributes to the overall narrative conclusion of the work. In light of the following comments, the reviewer recommends that this work is revised and resubmitted after major revisions, or reconsidered for indexing at another time. Major Comments:\n\nThis paper should do a better job to introduce and motivate the problem/question it is trying to solve. The way the information is presented makes the analysis seem very exploratory and not directed. This leads to some of the figures, figure 3 for example, to seem disconnected from the overall story\nThe work’s motivation reads well and is well cited. I would recommend quantifying the conditions HTL occurs at (i.e. the exact pressures and temperature ranges rather than qualitative descriptions of “medium” and “high” temperature and pressure respectively. I would also recommend discussing why the higher heating values are useful – the authors discuss “various applications” but may consider discussing the exact applications (for example in aviation fuels) which may bolster the narrative. In the third paragraph, in which an ML model and AI as a whole are referred to vaguely as “the learning component of a machine” or in comparison to a “brain” may risk unsubstantiated analogies – rather than exhibiting “intelligent behavior” many applications of machine learning in HTL are rather used as fitting tools that can predict metrics that are empirically useful. I would recommend discussing ML and AI in a more objective way. Overall, the description of ML and AI in the third introduction paragraph risk miscommunicating how ML and AI models work and often self contradict – for example, the authors refer to ML as not programs, but in the same sentence, refer to humans programming the ML. This kind of confusion warrants a more mechanistic description of ML and AI. The reviewer appreciates the well cited and discussed summary of ML use in the introduction. However, many of the cited works span a broad range of use cases – for example, HTL pretreatments, yield prediction, and specific feedstock compositions are discussed, but the reviewer worries that this may be casting too wide of a net to focus on the nature of this study. The authors may stylistically disagree as needed.\n\nKeyword choice may benefit from some honing – the terms may be too broad to reach the correct audience for what is a relatively niche intersectional field. The reviewer recommends specifying some of the keywords – even including HTL seems like great choice. Overall, while the authors are able to clearly communicate how the work was performed, the results and the specific conclusions to be garnered from each figure could be better communicated. The reviewer struggles to find specific conclusions or a line of narrative information that the figures tie into. This poises the paper as primarily exploratory rather than providing the incisive synthesis of information that may be interpreted from the data in the figures. I highly recommend visualizing some of this useful data in a manner that a reader can derive obvious conclusions the authors wish to communicate (for example for the graph/node plot, greater spacing between the nodes may aid clarity). Similarly, I would recommend poising a stronger line of thought tying each point (i.e. each figure) together.\nOther Comments: R2 vs MAE, RMSE, error metrics\nIn paragraph 4 of the introduction, R2 is used to refer to both accuracy and precision when referring to machine learning models. While R2 is an empirically useful metric to quantify the correlation between two variables or goodness of fit, it does not describe accuracy or precision. When talking about ML models, metrics that communicate the applied usefulness or “error bounds” of a model may be more useful, and are widely used throughout ML in HTL. Some examples include RMSE (root mean squared error) or MAE (mean absolute error). R2 can be used to describe the performance of the model (how close to a straight line does the parody plot of the model create) Regarding figure clarity, the reviewer suggests using colors of higher contrast – they are currently set up to be in the form of a heat map, where they are similar to each other but change down the list. This implies a relationship between the different items on the figures that may not exist (and if it does it could be helpful to clarify it!) Use of more distinct colors may rectify this\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? 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": [ { "c_id": "12973", "date": "17 Dec 2024", "name": "Tossapon katongtung", "role": "Author Response", "response": "Thank you for your advice, I will revise the manuscript as per the instructions and resubmit it through the system." }, { "c_id": "12981", "date": "02 Jan 2025", "name": "Tossapon katongtung", "role": "Author Response", "response": "1. This paper should do a better job to introduce and motivate the problem/question it is trying to solve. The way the information is presented makes the analysis seem very exploratory and not directed. This leads to some of the figures, figure 3 for example, to seem disconnected from the overall story = Thank you for the suggestion; however, we believe Figure 3 remains important as it highlights the most relevant study on this subject and helps readers identify gaps in the research within this field. 2.The work’s motivation reads well and is well cited. I would recommend quantifying the conditions HTL occurs at (i.e. the exact pressures and temperature ranges rather than qualitative descriptions of “medium” and “high” temperature and pressure respectively. I would also recommend discussing why the higher heating values are useful – the authors discuss “various applications” but may consider discussing the exact applications (for example in aviation fuels) which may bolster the narrative. = Revised, as suggested. please refer to lines 57-63 in Introduction section. 3. In the third paragraph, in which an ML model and AI as a whole are referred to vaguely as “the learning component of a machine” or in comparison to a “brain” may risk unsubstantiated analogies – rather than exhibiting “intelligent behavior” many applications of machine learning in HTL are rather used as fitting tools that can predict metrics that are empirically useful. I would recommend discussing ML and AI in a more objective way. Overall, the description of ML and AI in the third introduction paragraph risk miscommunicating how ML and AI models work and often self contradict – for example, the authors refer to ML as not programs, but in the same sentence, refer to humans programming the ML. This kind of confusion warrants a more mechanistic description of ML and AI. = Revised, as suggested. The text that may have caused confusion for readers has been removed. 4. The reviewer appreciates the well cited and discussed summary of ML use in the introduction. However, many of the cited works span a broad range of use cases – for example, HTL pretreatments, yield prediction, and specific feedstock compositions are discussed, but the reviewer worries that this may be casting too wide of a net to focus on the nature of this study. The authors may stylistically disagree as needed. = Based on the explanation provided in the introduction, the authors believe that presenting examples of research in various forms and across a wide range will help readers gain a clearer understanding of the overall scope of ML. Therefore, the author suggests retaining all the original sentences in the manuscript. 5.Keyword choice may benefit from some honing – the terms may be too broad to reach the correct audience for what is a relatively niche intersectional field. The reviewer recommends specifying some of the keywords – even including HTL seems like great choice. = Revised, as suggested. please refer to the red text in the Keywords section. 6. Overall, while the authors are able to clearly communicate how the work was performed, the results and the specific conclusions to be garnered from each figure could be better communicated. The reviewer struggles to find specific conclusions or a line of narrative information that the figures tie into. This poises the paper as primarily exploratory rather than providing the incisive synthesis of information that may be interpreted from the data in the figures. I highly recommend visualizing some of this useful data in a manner that a reader can derive obvious conclusions the authors wish to communicate (for example for the graph/node plot, greater spacing between the nodes may aid clarity). Similarly, I would recommend poising a stronger line of thought tying each point (i.e. each figure) together. = The bibliometric analysis in this study aims to provide an overview and identify trends in related research, serving as a guide for readers pursuing further studies on this topic. The authors believe that the statistical data and related information have been presented comprehensively. 7. In paragraph 4 of the introduction, R2is used to refer to both accuracy and precision when referring to machine learning models. While R2 is an empirically useful metric to quantify the correlation between two variables or goodness of fit, it does not describe accuracy or precision. When talking about ML models, metrics that communicate the applied usefulness or “error bounds” of a model may be more useful, and are widely used throughout ML in HTL. Some examples include RMSE (root mean squared error) or MAE (mean absolute error). R2 can be used to describe the performance of the model (how close to a straight line does the parody plot of the model create) = Revised, as suggested. please refer to the RED text in lines 88-89, 96 and 113-114. 8. Regarding figure clarity, the reviewer suggests using colors of higher contrast – they are currently set up to be in the form of a heat map, where they are similar to each other but change down the list. This implies a relationship between the different items on the figures that may not exist (and if it does it could be helpful to clarify it!) Use of more distinct colors may rectify this = Thank you for the advice. However, the authors intended for the images in the work to follow the same color theme. Additionally, the colors in the original image were chosen to represent intensity, where stronger colors indicate greater values and lighter colors indicate lower values, similar to contouring." } ] } ]	1	https://f1000research.com/articles/13-1131
https://f1000research.com/articles/13-164/v1	08 Mar 24	{ "type": "Research Article", "title": "Assessment of potential drug-drug interactions in hospitalized patients with infectious diseases: an experience from a secondary care hospital", "authors": [ "Javedh Shareef", "Sathvik Belagodu Sridhar", "Abu Nawa Ahmad Ismail", "Padma G.M. Rao", "Rashid Ain Ur", "Sathvik Belagodu Sridhar", "Abu Nawa Ahmad Ismail", "Padma G.M. Rao", "Rashid Ain Ur" ], "abstract": "Background Polypharmacy is common among hospitalized patients with infectious infections owing to comorbidities or concomitant illnesses. This raises the likelihood of drug-drug interactions and creates uncertainty for healthcare providers. This study aimed to assess the potential drug-drug interactions (pDDIs) among hospitalized patients with infectious diseases in a secondary care hospital.\n\nMethods A prospective observational study was conducted in the internal medicine ward for six months after the ethics committee’s approval. Data were collected from patient case records, and prescriptions were screened for pDDIs from a portable electronic physician information database (PEPID) resource analyzed using SPSS, version 27.0.\n\nResults In total, 148 patient case records were analyzed, and 549 pDDIs were identified, with 66.8% having at least one or more DDIs. The mean number of drug interactions was 3.70 ± 4.58 per prescription. The most frequently encountered drug interactions were drug combinations such as bisoprolol with atorvastatin and aspirin with tazobactam/piperacillin. Bivariate analysis showed that age, comorbidities, length of hospital stay, and the number of drugs prescribed were risk factors associated with DDIs (p<0.05). In the multiple binary logistic regression analysis, DDIs were significantly associated with comorbidities and the number of prescribed medications (p<0.0001).\n\nConclusions This study observed the prevalence of DDIs in hospitalized patients with infectious diseases of ‘moderate’ severity. Prescription screening using a drug information database assists in early identification and prevention of DDIs, enhancing drug safety and quality of patient-centered care.", "keywords": [ "Drug interactions", "polypharmacy", "prescription drugs", "prevalence", "patients" ], "content": "Introduction\n\nInfectious diseases are among the most common health problems worldwide irrespective of age. Infected individuals often require hospital admission, leading to a higher risk of morbidity, mortality, and increased healthcare expenditures.1 Indeed, appropriate selection and rational use of antimicrobial agents are often challenging for healthcare providers.2,3\n\nDrug–drug interactions (DDIs) occur when two or more co-administered drugs interact, with one drug altering the effect of the co-administered drug. The outcome effect of drug interactions may vary from non-serious to serious/life-threatening (or) irreversible, affecting the goals of therapy, clinical effectiveness, and worsening treatment outcomes.4 Studies have reported that age (≥ 65 years), polypharmacy, increased number of prescribers, and comorbid illness are defined risk factors for drug interactions. In addition, a decline in drug metabolism associated with aging, comorbidities such as hepatic and renal injury, and altered drug plasma concentrations complicate the medication use process and increase the sensitivity to drug interactions. As a result, clinically significant drug interactions prolong hospital stays, increase re-visit and healthcare expenditures, and aggravate patient outcomes in inpatient and outpatient healthcare settings.5–7\n\nDrug interactions are classified as pharmacokinetic and pharmacodynamic interactions; a few are categorized as unknown or other based on their mechanism of interactions. Drug interactions can be grouped as major, moderate, or minor according to severity and significance.8 Studies carried out in different health settings and patients reported that potential drug–drug interactions (pDDIs) range from 19.3% to 91.6%.9,10 A systematic review and meta-analysis reported that the prevalence of clinically manifested DDIs ranged from 1.2% to 64.0%.11 The increased incidence of adverse outcomes associated with drug–drug interactions is a common cause of hospital admission, primarily in the aging population.12 However, the variation in the results across different studies is associated with factors such as patient characteristics, prescribing pattern, severity of the illness, study population, and study setting.\n\nThe use of clinical decision support systems, close monitoring of patients’ drug therapy, and involvement of clinical pharmacists in a multidisciplinary team are some of the important measures that help to minimize drug interactions and improve patient safety.13,14\n\nStudies on antimicrobial agents in the United Arab Emirates (UAE) have focused on the prescription pattern of drug use and related outcomes in various hospital settings. However, studies related to DDIs with antimicrobial agents in infectious diseases are unaddressed despite being one of the reasons for hospitalization. Therefore, the present study was carried out to assess pDDIs among hospitalized patients with infectious diseases receiving antimicrobial agents in a secondary care hospital.\n\n\nMethods\n\nThis prospective observational study was conducted for six months from March 2021 to August 2021 in the internal medicine department of Ibrahim Bin Hamad Obaidullah Hospital in the northern emirate of the United Arab Emirates. Approval was obtained from the human ethics committee of Ras Al Khaimah Medical and Health Sciences University (RAKMHSU-REC-068-2020/21-UG-P) and Research Ethics Committee of Ministry of Health & Prevention, Ras Al Khaimah (MOHAP/REC/2021/1-2021-PG-P) in January 2021.\n\nIbrahim Bin Hamad Obaidullah Hospital is a government hospital owned by Ministry of Health and Prevention, Ras Al Khaimah. Studies that need to be carried in government hospitals under the Ministry of Health and prevention across the different emirates of United Arab Emirates including Ras Al Khaimah requires the permission from Ministry of Health and Prevention - Research Ethics Committee. So we have taken the approval from the MOHAP-REC RAK before starting the study in the aforementioned hospital. Since the principal investigators are from the Ras Al Khaimah Medical and Health Sciences University (RAKMHSU), it is important that any research by the RAKMHSU faculty needs approval from the Institutional ethics committee of RAKMHSU before getting the approval from MOHAP-RAK REC.\n\nAfter getting the approval from the MOHAP-RAK REC, written informed consent was obtained from all the patients who met the study criteria by the principal investigator after explaining the study procedures and other details to the study participants.\n\nHospitalized patients aged 18 years and older who were diagnosed with infectious diseases caused by bacterial pathogens and received a minimum of two or more medications containing at least one antimicrobial agent were included in the study. Patients referred from other departments admitted to the intensive care unit, diagnosed with COVID-19 receiving antibiotics, with incomplete medical records, and pregnant or lactating were excluded from the study.\n\nThe sample size was calculated using the formula to estimate a single proportion [n = (Z – α/2)2 p (1- p)/d2] where Z = standard normal variable at 95% confidence level (1.96), p = the prevalence of pDDIs assumed to be 50% and finally adjusted using a correction formula. The minimum sample size was 150 patients with 5-10% dropouts. Patients admitted during the study period were considered for the sampling frame and included using the systematic random sampling technique.\n\nThe medical records of the hospitalized patients who met the study criteria were reviewed daily and the data were collected from the Wareed system, an electronic health record information system (HIS), is a technological platform that virtually connects all the government hospital of ministry healthcare facilities in Dubai and the Northern Emirates by automating all healthcare processes across various departments. All necessary details of the patients, including drug therapy, were collected from the electronic health records and documented in the data collection forum designed according to the needs of the study. All prescription medicines were added to the ‘drugs to check’ list in the portable electronic physician information database (PEPID) interaction tool for evaluating pDDIs. The identified drug interactions were classified by level of concern as minor/non-significant, minor, moderate, significant, and life-threatening and were also based on pharmacokinetic, pharmacodynamics, and other/unknown mechanisms. The collected data were scrutinized and checked for completeness, clarity, and legibility before being entered into a Microsoft Excel (RRID:SCR_016137) spreadsheet and were later analyzed using IBM SPSS Statistics (RRID:SCR_016479) version 27. (IBM Corp., Armonk, NY, USA).\n\nDescriptive statistics, such as mean, standard deviation, frequencies, and percentages, were used for categorical data. Bivariate analysis using a chi-square test was used to identify factors associated with drug–drug interactions. In the binary logistic regression model, the associated factors identified in the bivariate analysis (p<0.05) were entered, and the odds ratio and 95% confidence interval were used to determine the independent risk factors for pDDIs. Statistical significance was p<0.05.\n\n\nResults\n\nIn total, 148 hospitalized patient case records were included during the study period, with 77 (52.02%) patients being males and 71 (47.97%) females. Most of the patients were in the age range between 21–40 years (28.37%), followed by 61-80 years (27.70%). The mean age was 54.27±24.3 (Mean±SD), ranging from 18 to 107 years.\n\nAmong the patients, more than half (56.76%) had a medical history of one or more comorbidities. The most common were cardiovascular diseases (42.04%) followed by diabetes mellitus (29.54%) and dyslipidemia (7.38%). Respiratory tract infection (34.83%), urinary tract infection (34.19%), sepsis (14.8%), and gastroenteritis (7.09%) were the most common infectious diseases for hospital admission in our study. Most hospitalized patients had a stay duration of 6-10 days (56.08%), and the average length of stay was 8.16±2.85 days (range: 3-16 days). In our study, the majority of patients (45.27%) received 6–9 drugs per prescription, and the average number of drugs per prescription was 8.35±3.19 (Mean±SD) (range: 2-16) medications (Table 1).33\n\nA total of 44 drugs, consisting of 27 non-antimicrobial agents and 17 antimicrobial agents from different classes of drugs, were involved in the onset of pDDIs. Antimicrobial agents indicated for systemic use belonging to different therapeutic groups, such as antibacterial and antimycobacterial agents, were also included in the evaluation of the clinically relevant pDDIs (Table 2).\n\nϮ Fisher's exact.\n\n* p value <0.05 is statistically significant.\n\nIn our study, 549 drug–drug interactions with 116 combinations of interacting drugs were observed. This includes 396 drug interactions from 64 non-antimicrobial combinations, 137 drug–drug interactions from 44 non-antimicrobial and antimicrobial combinations, and 16 drug–drug interactions from eight antimicrobial combinations. The mean drug interactions identified in the study population were 3.70±4.58 per prescription.\n\nIt was observed that 66.89% (n=99) of prescriptions were found to have the potential for at least one or more DDIs, irrespective of the type of severity. The identified DDIs classified according to severity show that most of the interactions, 220 (40.07%) belong to the ‘moderate’ category, 155 (28.23%) were minor/non-significant, and 145 (26.41%) were classified ‘minor.’ A total of 29(5.28%) drug interactions were rated as a ‘significant’ severity category (Table 3).\n\nThe analysis of the class of medications involved in the onset of pDDIs carried out using the Anatomical Therapeutic Chemical (ATC) classification system derived by the World Health Organization found a higher prevalence in the category cardiovascular system (28.8%) followed by anti-infective for systemic use (23.9%) and alimentary tract and metabolism (21.5%) (Figure 1).\n\nThe evaluation of the underlying mechanism that causes DDIs showed that 210 (38.25) interactions involve pharmacokinetic interactions, while 181 (32.96%) interactions were caused by ‘others’ or unknown mechanisms. The remaining 158 (28.77%) interactions were known to be produced by pharmacodynamic interactions.\n\nThe most frequently identified DDIs were the combination of atorvastatin with clopidogrel, bisoprolol, amlodipine, or pantoprazole; aspirin with insulin; clopidogrel enoxaparin and furosemide with valsartan; and lisinopril and bisoprolol. The antimicrobial drugs involved in pDDIs were combinations of ceftriaxone with enoxaparin and aspirin, levofloxacin with insulin, furosemide, and piperacillin/tazobactam with aspirin, metformin, and doxycycline (Table 4).\n\nAnalysis of the factors associated with the appearance of pDDIs showed that there was a statistically significant association with age, comorbidities, length of hospital stay, and the number of drugs prescribed (p<0.05) (Table 5).\n\n* p<0.05 statistically significant.\n\n** p<0.01 highly statistically significant.\n\nIn the binary logistic regression analysis, the dependent variable was the presence or absence of pDDIs, and the predictor variables were age, comorbidities, hospital stay, and the number of drugs prescribed. Drug–drug interactions were significantly associated with comorbidities and the number of drugs prescribed (p<0.05) (Table 6).\n\n* p<0.05 statistically significant.\n\n\nDiscussion\n\nDrug interactions contribute to undesirable health outcomes, compromise the clinical effectiveness of drug therapy, increase hospital visits, and prolong hospital stays.15 The overall prevalence of pDDIs in our study was 67%, low compared to the study by Bhagavathula et al., who reported at least one pDDIs in 78% of the study sample.16 Downward trends in prevalence were documented in earlier studies by Kuscu et al. (60%) and Rabba et al. (56%), respectively.17,18\n\nThis disparity in the prevalence of pDDIs may be attributed to the differences in the study setting, study population, prescribing pattern of medications, and types of pDDIs and tools used to screen drug interactions in the study. In the present study, the average of 3.70±4.58 drug interactions per prescription among hospitalized patients. Documented evidence indicates that drug interactions occur more predominantly in hospitalized patients than in outpatients, considering the severity of the disease, comorbidities, and prescription of multiple medications with frequent modifications during their stay.19\n\nIn the current study, aspirin, clopidogrel, statins, enoxaparin, furosemide, valsartan, and bisoprolol were prescribed to prevent and manage cardiovascular diseases. Documented evidence has reported that prescribing these medications alone or in combination is responsible for various interactions, such as increased bleeding, electrolyte imbalance, renal failure, and hypotension.17,19–21 However, prescribing these medications is sometimes unavoidable and therapeutically valuable as a lifesaving medication. Therefore, close monitoring for effective treatment and evaluation of the benefit risk-assessment of actual DDIs of prescribed medications is warranted. At the same time, careful laboratory assessment of international normalized ratio, serum electrolytes, renal and liver function tests, signs and symptoms of bleeding, and blood pressure monitoring are vital during treatment.\n\nSimilarly, metformin, sitagliptin, insulin, tamsulosin, memantine, levodopa, pantoprazole, paracetamol, and supplements such as potassium chloride and calcium carbonate were some of the important medications prescribed for the various other medical conditions in our study. In addition, drugs such as penicillins, cephalosporins, fluoroquinolones, metronidazole, macrolides, doxycycline, linezolid, isoniazid, rifampicin, vancomycin, and amphotericin B were some of the important antimicrobial agents used in this study. Drugs that cause enzyme induction or inhibition, resulting in reduced metabolism or clinical effects and alteration of gastrointestinal absorption are the most common mechanisms related to antimicrobial interactions.22\n\nA review of the published evidence suggests that fluoroquinolones and macrolides are associated with QT prolongation and may cause life-threatening arrhythmias.17,23,24 Cautious prescribing should be exercised when co-administering drugs with a narrow therapeutic index and drugs metabolized through cytochrome P450 isozymes that can develop clinically significant unpredictable drug interactions, particularly in patients with renal and hepatic impairment and the elderly population. In the present study, the class of medications involved in pDDIs by the ATC classification showed a higher prevalence in the cardiovascular system (28.2%) followed by anti-infective for systemic use (23.9%). The increase in the prevalence of cardiovascular disease could be related to the use of complex medications for the long-term treatment of comorbidities and associated complications among the study populations. Our findings are consistent with those of Noor et al., Teka et al., and Samardzic et al., who reported an increased prevalence of pDDIs in patients with cardiovascular disease.19,20,24 Furthermore, earlier studies by Baniasadi et al., and Pavanello et al., in critical care patients showed that the most common drug class involved in pDDIs was anti-infective for systemic use, accounting for 51.5% and 45.8%, respectively.22,25 The difference in study settings, varying profiles of study populations, disagreement in treatment guidelines and prescribing practice, and the use of different clinical decision support tools to analyze drug interactions may help explain the difference in the class of drugs involved in the onset of pDDIs.\n\nIn the present study, the severity level of most drug interactions was ‘moderate’ (40.0%) followed by ‘minor/non-significant’ (28.2%). All pDDIs are not identical in severity and evaluating their severity l is very important to recognize their clinical significance and proper management. Only 5.28% of the identified drug interactions were found to be categorized as having a level of severity ‘significant’, which requires close monitoring to avoid any adverse outcome of the pDDIs. A possible reason could be that physicians are aware of the risk factors and severity of pDDIs and might have customized drug therapy individually to avoid or minimize pDDIs.\n\nThese findings align with the results of the study by Noor et al., and Obeid et al., which reported that most of the interactions were ‘moderate’ in severity.20,26 Contrary to our findings, studies by Rabba et al., and Eneh et al., reported 66.4% and 52.7% of interactions with ‘major’ in severity level.18,23 The difference in defining the classification and grading of severity between the resources could be a possible reason for the varying study results. Studies have observed that mechanism of action plays a significant role in DDIs, which requires management by either reducing the dose of one drug by 25% or 50%, changing the frequency of administration and dosage form, or avoiding such combination, or replacing it with another medication.19,27,28\n\nOur study showed that ‘pharmacokinetic interactions’ were the most common underlying mechanism that caused pDDIs compared to pharmacodynamic and other unknown mechanisms. Similar observations have been cited in the study by Tesfaye et al., who reported pharmacokinetic interactions as the most common mechanism involved in causing pDDIs compared with pharmacodynamic and other/unknown interactions.29\n\nStudies have emphasized that patient characteristics such as age, comorbidities, number of medications prescribed, and hospital stay are risk factors for clinically significant pDDIs.28,30,31 Age, comorbidities, length of hospital stay, and polypharmacy predispose patients to pDDIs. It is important to note that aging populations are at risk of developing multiple comorbid medical conditions that require frequent hospital visits and a prolonged stay prescribed with more complex therapeutic regimens. Physiological changes related to age and variations in pharmacokinetics and pharmacodynamic parameters increase the risk and greater chance of developing pDDIs and adverse outcomes that reduce the efficacy of the treatment.32\n\nOur study has a few limitations. First, only one database would limit the number of pDDIs and may not reflect all pDDIs. Using multiple database tools and comparisons may help define the results more explicitly. Second, the data for the present study were collected from the Wareed system and mainly focused on the theoretical pDDIs and could not address the actual drug interactions and results from a clinical viewpoint due to lack of follow-up. Third, the study only included patients with specific indications in the internal general medicine ward. Therefore, the findings cannot be extended or applied to other specialty wards, intensive care units, or outpatient settings.\n\n\nConclusions\n\nThe present study identified a higher frequency of pDDIs in hospitalized patients with infectious diseases. Antimicrobial agents and co-prescribed medications interacted; most of the interactions in our study had ‘moderate’ levels of severity. This study highlighted that advanced age, multiple comorbidities, and polypharmacy were independent risk factors for pDDIs. Knowledge about pDDIs and the regular use of professional drug information database support systems can help prescribers optimize drug therapy and enhance health outcomes. The study strongly recommends that regular review of patient drug therapy by a clinical pharmacist might avoid possible drug combinations that are likely to cause pDDIs and could ring a bell in improving the quality of patient-centered care.", "appendix": "Data availability\n\nFigshare: ASSESSMENT OF POTENTIAL DRUG-DRUG INTERATIONS IN HOSPITALIZED PATIENTS WITH INFECTIOUS DISEASES – AN EXPERIENCE FROM A SECONDARY CARE HOSPITAL. https://doi.org/10.6084/m9.figshare.24220714.v2. 33\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 the administration of Ibrahim Bin Hamad Obaidullah Hospital, Ras Al Khaimah, United Arab Emirates for giving permission to conduct the study. The authors would like to thank the President of RAK Medical and Health Science University and the Dean of RAK College of Pharmacy for their encouragement and support in conducting this study.\n\n\nReferences\n\nAkdemir Kalkan İ, Çınar G, Pehlivanlı A, et al.: Pattern of systemic antibiotic use and potential drug interactions: Evaluations through a point prevalence study in Ankara University Hospitals. Turk. J. Med. Sci. 2021; 51(2): 523–529. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAdedapo AD, Akunne OO: Patterns of Antimicrobials Prescribed to Patients Admitted to a Tertiary Care Hospital: A Prescription Quality Audit. Cureus. 2021; 13(6): e15896. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCantudo-Cuenca MR, Jiménez-Morales A, Martínez-de la Plata JE: Pharmacist-led antimicrobial stewardship programme in a small hospital without infectious diseases physicians. Sci. Rep. 2022; 12: 9501. 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Publisher Full Text" }	[ { "id": "272895", "date": "16 May 2024", "name": "Rina Das", "expertise": [ "Reviewer Expertise infectious diseases", "diabetes care", "hypertension therapy", "liver diseases", "serum enzyme study" ], "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 have studied 148 patient case records to assess the potential drug-drug interactions in hospitalized patients with infectious diseases giving more emphasis on comorbidity cases.\nThe manuscript addresses the potential drug-drug interactions in hospitalized patients with infectious diseases. Since there are many studies investigating this theme, it is not clear how this research adds to the current field.\nA statement of confirmation that the study conforms to recognised ethical standards should be included (for example: Declaration of Helsinki; US Federal Policy for the Protection of Human Subjects; European Medicines Agency Guidelines for Good Clinical Practice).\nThe title of article states that ''Assessment of potential drug-drug interactions in hospitalized patients with infectious diseases''......but the result states majority of drug interaction with non-antimicrobial combinations, ''396 drug interactions 64 non-antimicrobial combinations''. When the work was focused on patients with infectious diseases, majority of cases should have been included DDIs with non-antimicrobial and antimicrobial combinations, or antimicrobial combinations.\nIs the study design appropriate and is the work technically sound?\nThe method described is not detailed enough to warrant possibility of replication by the readers. Details of the experimental setting, timeline, research questions, and references of the methods/scales have to be better explained. Also no description of infectious diseases is provided whereas all comorbidities are elaborately described. The method of identification of drug-drug interaction results is not described.\nThe text needs editing and language review, eg Polypharmacy is common among hospitalized patients with infectious infections .........\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": [ { "c_id": "12023", "date": "07 Aug 2024", "name": "JAVEDH SHAREEF", "role": "Author Response", "response": "COMPLIANCE TO REVIEWER COMMENTS Manuscript title: ASSESSMENT OF POTENTIAL DRUG-DRUG INTERACTIONS IN HOSPITALIZED PATIENTS WITH INFECTIOUS DISEASES – AN EXPERIENCE FROM A SECONDARY CARE HOSPITAL Authors: JAVEDH SHAREEF, Sathvik Belagodu Sridhar1, Abu Nawa Ahmad Ismail1, Padma G.M. Rao1, Rashid Ain Ur2 Reviewer – 1 Rina Das, MM College of Pharmacy, Maharishi Markandeshwar University, Ambala, Haryana, India Sl No Reviewer (comments) Corrections Made Page No. 1. The authors have studied 148 patient case records to assess the potential drug-drug interactions in hospitalized patients with infectious diseases giving more emphasis on comorbidity cases. The manuscript addresses the potential drug-drug interactions in hospitalized patients with infectious diseases. Since there are many studies investigating this theme, it is not clear how this research adds to the current field. Response: Thank you for your valuable feedback. Our investigation of pDDIs among hospitalized patients with infectious illnesses in the UAE adds a novel perspective to the current literature. While there are several global studies on this issue, our research is relevant to the UAE healthcare system, considering local prescription patterns, common infectious illnesses, and distinct patient demographics. By evaluating 148 patient case records, focusing on comorbidities, we present insights specific to this geographical situation. This localized approach broadens our understanding of medication safety and management measures, particularly in the UAE, with practical implications for enhancing patient care and outcomes in our healthcare system. -- 2. A statement of confirmation that the study conforms to recognised ethical standards should be included (for example: Declaration of Helsinki; US Federal Policy for the Protection of Human Subjects; European Medicines Agency Guidelines for Good Clinical Practice). Response: Thank you for your feedback. We confirm that our study \"Assessment of potential drug-drug interactions in hospitalized patients with infectious diseases: an experience from a secondary care hospital\" adheres to recognized ethical standards. This study was performed per the principles outlined in the Declaration of Helsinki, the US Federal Policy for the Protection of Human Subjects (Common Rule), and the European Medicines Agency Guidelines for Good Clinical Practice. 04 3. The title of article states that ''Assessment of potential drug-drug interactions in hospitalized patients with infectious diseases''......but the result states majority of drug interaction with non-antimicrobial combinations, ''396 drug interactions 64 non-antimicrobial combinations''. When the work was focused on patients with infectious diseases, majority of cases should have been included DDIs with non-antimicrobial and antimicrobial combinations, or antimicrobial combinations. Response: Thank you for your insightful observation. Our study aimed to investigate pDDIs in hospitalized patients with infectious diseases and also looked at interactions with antimicrobial and non-antimicrobial medications. This divergence from the stated aims could be clarified by emphasizing the comprehensive nature of our investigation into pDDIs, encompassing interactions across all medication classes administered to the patient group. This approach aimed to thoroughly assess potential interactions that could impact clinical management in secondary care settings. The primary aim of our study was to investigate pDDIs involving antimicrobial drugs, which are important in treating infectious disorders. However, throughout the process, we discovered substantial interactions with non-antimicrobial drugs, which might have an equivalent impact on patient outcomes. Including these data gives a more comprehensive understanding of the drug management problems that hospitalized patients experience. Our work provides a stronger platform for improving clinical procedures and guaranteeing patient safety by recognizing and managing antimicrobial and non-antimicrobial pDDIs. -- 4. The method described is not detailed enough to warrant possibility of replication by the readers. Details of the experimental setting, timeline, research questions, and references of the methods/scales have to be better explained. Also no description of infectious diseases is provided whereas all comorbidities are elaborately described. The method of identification of drug-drug interaction results is not described. Response: The methodology section has been revised to include details about the study setting, ethical approval, inclusion/exclusion criteria, study duration, reference to the drug interaction tools used, and method of assessing drug-drug interactions. A description of infectious diseases is provided in the first paragraph of the introduction section. 4-6 & 03 5. The text needs editing and language review, e.g., Polypharmacy is common among hospitalized patients with infectious infections ......... Response: All the grammatical and construction errors have been rectified with the help of an English language expert and incorporated in the updated version of the manuscript. --- 6. Is the work clearly and accurately presented and does it cite the current literature? Yes Thank you! --" } ] }, { "id": "289750", "date": "28 Jun 2024", "name": "Mahadev Rao", "expertise": [ "Reviewer Expertise Drug-drug interactions", "clinical pharmacy", "precision 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\nAssessment of potential drug-drug interactions in hospitalized patients with infectious diseases: an experience from a secondary care hospital The authors aimed to assess potential drug-drug interactions (pDDIs) among hospitalized patients with infectious diseases in a secondary care hospital.\nThere is an inconsistency in the mention of “pDDIs”, “DDI”, drug–drug interactions and “drug interaction” across the manuscript. There is a significant flaw in the study objectives, design and results. The study objective states that “Therefore, the present study was carried out to assess pDDIs among hospitalized patients with infectious diseases receiving antimicrobial agents in a secondary care hospital.” However, the study also reports on the pDDIs between 2 non-antimicrobial agents. Many pDDIs reported in Table 4 are between 2 non-antimicrobial agents, which may not justify the title and objectives. There is a fundamental flaw in this study. Table 1 reports that tuberculosis is present in 1 patient as a comorbidity among 148 hospitalized patients studied. Whereas Table 4 shows that the frequency of Isoniazid-Rifampicin pDDI is 5, which cannot be possible. Further, isoniazid is administered along with rifampicin, and other antitubercular agents such as pyrazinamide and ethambutol as a combined fixed dose formulation to TB patients. To justify the title and the objectives, the authors may focus on the identification of the pDDIs between the antimicrobial agents and between antimicrobial agents and other drug categories. Identify the frequency of the top interacting antimicrobial agents and their mechanism and classify based on the infection categories. Table 1 has listed on the different comorbidities in the 148 hospitalized patients. What are the comorbidities, where a higher frequency of pDDIs between antimicrobial agents and other drug categories were observed? Since this is a scientific manuscript, the detailed elaboration on the need for obtaining necessary ethics committee approval from the hospital in a paragraph is not required as mentioned in the study setting. Most of the statements mentioned in the patient demographics are a repetition of the Table 1. There is no need for repeating the results and table. Also, many result statements are repeated in the discussion section. The study aimed to assess the potential drug-drug interaction among hospitalized patients with infectious diseases using only a single electronic physician information database (PEPID) interaction tool (mentioned in limitation). In the Table 5, for the column- presence and absence of DDI, n and % is mentioned in column title, but only n is present in the results. Also is it “Number of drugs prescribed” instead of “No drugs prescribed”?. The current manuscript requires extensive editing of English language.\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? 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? No", "responses": [] } ]	1	https://f1000research.com/articles/13-164
https://f1000research.com/articles/13-1559/v1	24 Dec 24	{ "type": "Data Note", "title": "A guide to selecting high-performing antibodies for VAPB (UniProt ID: O95292) for use in western blot, immunoprecipitation, and immunofluorescence", "authors": [ "Sara González Bolívar", "Riham Ayoubi", "Charles Alende", "Maryam Fothouhi", "Irina Shlaifer", "Peter S. McPherson", "Carl Laflamme", "NeuroSGC/YCharOS/EDDU collaborative group", "ABIF consortium", "Sara González Bolívar", "Riham Ayoubi", "Charles Alende", "Maryam Fothouhi", "Irina Shlaifer", "Peter S. McPherson" ], "abstract": "VAPB is an adaptor protein known for its role as an anchor for other proteins at the endoplasmic reticulum. A mutant form of VAPB has been linked to amyotrophic lateral sclerosis and the underlying mechanisms resulting from this defect are studied by researchers in this area to uncover its implication in the disease. Here we have characterized six VAPB commercial antibodies for western blot, immunoprecipitation, and immunofluorescence using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. These studies are part of a larger, collaborative initiative seeking to address antibody reproducibility issues by characterizing commercially available antibodies for human proteins and publishing the results openly as a resource for the scientific community. While use of antibodies and protocols vary between laboratories, we encourage readers to use this report as a guide to select the most appropriate antibodies for their specific needs.", "keywords": [ "O95292", "VAPB", "Vesicle-associated membrane protein-associated protein B/C", "VAMP-associated protein B/C", "antibody characterization", "antibody validation", "western blot", "immunoprecipitation", "immunofluorescence" ], "content": "Introduction\n\nVAPB is the vesicle-associated membrane protein (VAMP)-associated protein B and C.1 VAPB acts as a tethering protein of the endoplasmic reticulum to other organelles, orchestrating many cellular processes, namely vesicle trafficking.2 A mutant form of the protein has been linked to amyotrophic lateral sclerosis3 and high-quality antibodies and necessary to explore the resulting pathological mechanisms.\n\nThis research is part of a broader collaborative initiative in which academics, funders and commercial antibody manufacturers are working together to address antibody reproducibility issues by characterizing commercial antibodies for human proteins using standardized protocols, and openly sharing the data.4–6 Here we evaluated the performance of six commercial antibodies for VAPB for use in western blot, immunoprecipitation, and immunofluorescence, enabling biochemical and cellular assessment of VAPB properties and function. The platform for antibody characterization used to carry out this study was endorsed by a committee of industry academic representatives. It consists of identifying human cell lines with adequate target protein expression and the development/contribution of equivalent knockout (KO) cell lines, followed by antibody characterization procedures using most commercially available antibodies against the corresponding protein. The standardized consensus antibody characterization protocols are openly available on Protocol Exchange, a preprint server (DOI:10.21203/rs.3.pex-2607/v1).7\n\nThe authors do not engage in result analysis or offer explicit antibody recommendations. Our primary aim is to deliver top-tier data to the scientific community, grounded in Open Science principles. This empowers experts to interpret the characterization data independently, enabling them to make informed choices regarding the most suitable antibodies for their specific experimental needs. Guidelines on how to interpret antibody characterization data found in this study are featured on the YCharOS gateway.8\n\n\nResults and discussion\n\nOur standard protocol involves comparing readouts from WT (wild type) and KO (knockout cells).9,10 The first step was to identify a cell line(s) that expresses sufficient levels of a given protein to generate a measurable signal using antibodies. To this end, we examined the DepMap (Cancer Dependency Map Portal, RRID:SCR_017655) transcriptomics database to identify all cell lines that express the target at levels greater than 2.5 log2 (transcripts per million “TPM” + 1), which we have found to be a suitable cut-off.4 HeLa expresses the VAPB transcript at 5.2 and was identified as a suitable cell line to modify with CRISPR/Cas9 to KO the corresponding VAPB gene ( Table 1).\n\nTo screen the antibodies by western blot, WT and VAPB KO protein lysates were ran on SDS-PAGE, transferred onto nitrocellulose membranes, and then probed with all six VAPB antibodies in parallel ( Figure 1).\n\nLysates of HeLa WT and VAPB KO were prepared, and 40 μg of protein were processed for western blot with the indicated VAPB antibodies. The Ponceau stained transfers of each blot are presented to show equal loading of WT and KO lysates and protein transfer efficiency from the acrylamide gels to the nitrocellulose membrane. Antibody dilutions were chosen according to the recommendations of the antibody supplier. Antibody dilution used: ab315013** at 1/1000, A5363 at 1/500, GTX131631 at 1/1000, 66191-1-Ig* at 1/1000, MA5-29639 at 1/1000, PA5-53023 at 1/1500 (0.2 μg/ml). Predicted band size: 27.2 kDa. Recombinant antibody, Monoclonal antibody.\n\nWe then assessed the capability of the six antibodies to capture VAPB from HeLa protein extracts using immunoprecipitation techniques, followed by western blot analysis. For the immunoblotting step, a specific VAPB antibody identified previously (refer to Figure 1) was selected. Equal amounts of the starting material (SM) and the unbound fractions (UB), as well as the whole immunoprecipitate (IP) eluates were separated by SDS-PAGE ( Figure 2).\n\nHeLa lysates were prepared, and immunoprecipitation was performed using 1 mg of lysate and 2.0 μg of the indicated VAPB antibodies pre-coupled to Dynabeads protein A or protein G. Samples were washed and processed for western blot with the indicated VAPB antibody. For western blot, ab315013* was used at 1/1000. The Ponceau stained transfers of each blot are shown. SM=4% starting material; UB=4% unbound fraction; IP=immunoprecipitate. *Recombinant antibody, Monoclonal antibody.\n\nFor immunofluorescence, the six antibodies were screened using a mosaic strategy. First, HeLa WT and VAPB KO cells were labelled with different fluorescent dyes in order to distinguish the two cell lines, and the VAPB antibodies were evaluated. Both WT and KO lines imaged in the same field of view to reduce staining, imaging and image analysis bias ( Figure 3). Quantification of immunofluorescence intensity in hundreds of WT and KO cells was performed for each antibody tested, and the images presented in Figure 3 are representative of this analysis.7\n\nHeLa WT and VAPB KO cells were labelled with a green or a far-red fluorescent dye, respectively. WT and KO cells were mixed and plated to a 1:1 ratio on coverslips. Cells were stained with the indicated VAPB antibodies and with the corresponding Alexa-fluor 555 coupled secondary antibody including DAPI. Acquisition of the blue (nucleus-DAPI), green (WT), red (antibody staining) and far-red (KO) channels was performed. Representative images of the merged blue and red (grayscale) channels are shown. WT and KO cells are outlined with green and magenta dashed lines, respectively. When an antibody was recommended for immunofluorescence by the supplier, we tested it at the recommended dilution. The rest of the antibodies were tested at 1 and 2 μg/ml and the final concentration was selected based on the detection range of the microscope used and a quantitative analysis not shown here. Antibody dilution used: ab315013** at 1/250, A5363 at 1/1400, GTX131631 at 1/1000, 66191-1-Ig* at 1/800, MA5-29639 at 1/1000, PA5-53023 at 1/150. Bars = 10 μm. Recombinant antibody, Monoclonal antibody.\n\nIn conclusion, we have screened six VAPB commercial antibodies by western blot, immunoprecipitation, and immunofluorescence by comparing the signal produced by the antibodies in human HeLa WT and VAPB KO cells. To assist users in interpreting antibody performance, Table 3 outlines various scenarios in which antibodies may perform in all three applications.4 Several high-quality and renewable antibodies that successfully detect VAPB were identified in all applications. Researchers who wish to study VAPB in a different species are encouraged to select high-quality antibodies, based on the results of this study, and investigate the predicted species reactivity of the manufacturer before extending their research.\n\nThe underlying data for this study can be found on Zenodo, an open-access repository for which YCharOS has its own collection of antibody characterization reports.\n\nInherent limitations are associated with the antibody characterization platform used in this study. Firstly, the YCharOS project focuses on renewable (recombinant and monoclonal) antibodies and does not test all commercially available VAPB antibodies. YCharOS partners provide approximately 80% of all renewable antibodies, but some top-cited polyclonal antibodies may not be available through these partners.\n\nSecondly, the YCharOS effort employs a non-biased approach that is agnostic to the protein for which antibodies have been characterized. The aim is to provide objective data on antibody performance without preconceived notions about how antibodies should perform or the molecular weight that should be observed in western blot. As the authors are not experts in VAPB, only a brief overview of the protein’s function and its relevance in disease is provided. VAPB experts are invited to analyze and interpret observed banding patterns in western blots and subcellular localization in immunofluorescence.\n\nThirdly, YCharOS experiments are not performed in replicates primarily due to the use of multiple antibodies targeting various epitopes. Once a specific antibody is identified, it validates the protein expression of the intended target in the selected cell line, confirms the lack of protein expression in the KO cell line and supports conclusions regarding the specificity of the other antibodies. All experiments are performed using master mixes, and meticulous attention is paid to sample preparation and experimental execution. In IF, the use of two different concentrations serves to evaluate antibody specificity and can aid in assessing assay reliability. In instances where antibodies yield no signal, a repeat experiment is conducted following titration. Additionally, our independent data is performed subsequently to the antibody manufacturers internal validation process, therefore making our characterization process a repeat.\n\nLastly, as comprehensive and standardized procedures are respected, any conclusions remain confined to the experimental conditions and cell line used for this study. The use of a single cell type for evaluating antibody performance poses as a limitation, as factors such as target protein abundance significantly impact results.7 Additionally, the use of cancer cell lines containing gene mutations poses a potential challenge, as these mutations may be within the epitope coding sequence or other regions of the gene responsible for the intended target. Such alterations can impact the binding affinity of antibodies. This represents an inherent limitation of any approach that employs cancer cell lines.\n\n\nMethod\n\nThe standardized protocols used to carry out this KO cell line-based antibody characterization platform was established and approved by a collaborative group of academics, industry researchers and antibody manufacturers. The detailed materials and step-by-step protocols used to characterize antibodies in western blot, immunoprecipitation and immunofluorescence are openly available on Protocol Exchange, a preprint server (DOI:10.21203/rs.3.pex-2607/v1).7 Brief descriptions of the experimental setup used to carry out this study can be found below.\n\nCell lines used and primary antibodies tested in this study are listed in Tables 1 and 2, respectively. To ensure that the cell lines and antibodies are cited properly and can be easily identified, we have included their corresponding Research Resource Identifiers, or RRID.11,12 HeLa VAPB KO clone was generated at the Early Drug Discovery Unit (EDDU) using a guide RNA with the following sequence: UGAAGACUACAGCACCACGU.\n\n Monoclonal antibody.\n\n** Recombinant antibody.\n\nPeroxidase-conjugated goat anti-rabbit and anti-mouse antibodies are (Thermo Fisher Scientific, cat. number 65-6120 and 62-6520). Peroxidase-conjugated VeriBlot for IP detection is from Abcam, cat. number ab131366. Alexa-555-conjugated goat anti-rabbit and anti-mouse secondary antibodies (Thermo Fisher Scientific, cat. number A21429 and A21424).\n\nHeLa WT and VAPB KO cells were collected in RIPA buffer (25 mM Tris-HCl pH 7.6, 150 mM NaCl, 1% NP-40, 1% sodium deoxycholate, 0.1% SDS) (Thermo Fisher Scientific, cat. number 89901) supplemented with 1× protease inhibitor cocktail mix (MilliporeSigma, cat. number P8340). Lysates were sonicated briefly and incubated 30 min on ice. Lysates were spun at ~110,000 × g for 15 min at 4°C and equal protein aliquots of the supernatants were analyzed by SDS-PAGE and western blot. BLUelf prestained protein ladder (GeneDireX, cat. number PM008-0500) was used.\n\nWestern blots were performed with precast midi 4-20% Tris-Glycine polyacrylamide gels (Thermo Fisher Scientific, cat. number WXP42012BOX) ran with Tris/Glycine/SDS buffer (Bio-Rad, cat. number 1610772), loaded in Laemmli loading sample buffer (Thermo Fisher Scientific, cat. number AAJ61337AD) and transferred on nitrocellulose membranes. Proteins on the blots were visualized with Ponceau S staining (Thermo Fisher Scientific, cat. number BP103-10) which is scanned to show together with individual western blot. Blots were blocked with 5% milk for 1 hr, and antibodies were incubated O/N at 4°C with 5% milk in TBS with 0.1% Tween 20 (TBST) (Cell Signalling Technology, cat. number 9997). Following three washes with TBST, the peroxidase conjugated secondary antibody was incubated at a dilution of ~0.2 μg/ml in TBST with 5% milk for 1 hr at room temperature followed by three washes with TBST. Membranes were incubated with Pierce ECL (Thermo Fisher Scientific, cat. number 32106) prior to detection with the iBright™ CL1500 Imaging System (Thermo Fisher Scientific, cat. number A44240).\n\nAntibody-bead conjugates were prepared by adding 2 μg of antibody to 500 μl of Pierce IP Lysis Buffer from Thermo Fisher Scientific (cat. number 87788) in a microcentrifuge tube, together with 30 μl of Dynabeads protein A- (for rabbit antibodies) or protein G- (for mouse antibodies) (Thermo Fisher Scientific, cat. number 10002D and 10004D, respectively). Tubes were rocked for ~1 h at 4°C followed by two washes to remove unbound antibodies.\n\nHeLa WT were collected in Pierce IP buffer (25 mM Tris-HCl pH 7.4, 150 mM NaCl, 1 mM EDTA, 1% NP-40 and 5% glycerol) supplemented with protease inhibitor. Lysates were rocked 30 min at 4°C and spun at 110,000 × g for 15 min at 4°C. 0.5 ml aliquots at 2 mg/ml of lysate were incubated with an antibody-bead conjugate for ~1 h at 4°C. The unbound fractions were collected, and beads were subsequently washed three times with 1.0 ml of IP buffer and processed for SDS-PAGE and western blot on a precast midi 4-20% Tris-Glycine polyacrylamide gels. VeriBlot for IP Detection Reagent:HRP was used as a secondary detection system at a concentration of 0.1 μg/ml.\n\nHeLa WT and VAPB KO cells were labelled with a green and a far-red fluorescence dye, respectively (Thermo Fisher Scientific, cat. number C2925 and C34565). The nuclei were labelled with DAPI (Thermo Fisher Scientific, cat. Number D3571) fluorescent stain. WT and KO cells were plated on 96-well plate with optically clear flat-bottom (Perkin Elmer, cat. number 6055300) as a mosaic and incubated for 24 hrs in a cell culture incubator at 37oC, 5% CO2. Cells were fixed in 4% paraformaldehyde (PFA) (VWR, cat. number 100503-917) in phosphate buffered saline (PBS) (Wisent, cat. number 311-010-CL). Cells were permeabilized in PBS with 0.1% Triton X-100 (Thermo Fisher Scientific, cat. number BP151-500) for 10 min at room temperature and blocked with PBS with 5% BSA, 5% goat serum (Gibco, cat. number 16210-064) and 0.01% Triton X-100 for 30 min at room temperature. Cells were incubated with IF buffer (PBS, 5% BSA, 0.01% Triton X-100) containing the primary VAPB antibodies overnight at 4°C. Cells were then washed 3 × 10 min with IF buffer and incubated with corresponding Alexa Fluor 555-conjugated secondary antibodies in IF buffer at a dilution of 1.0 μg/ml for 1 hr at room temperature with DAPI. Cells were washed 3 × 10 min with IF buffer and once with PBS.\n\nImages were acquired on an ImageXpress micro confocal high-content microscopy system (Molecular Devices), using a 20x NA 0.95 water immersion objective and scientific CMOS cameras, equipped with 395, 475, 555 and 635 nm solid state LED lights (lumencor Aura III light engine) and bandpass filters to excite DAPI, Cellmask Green, Alexa-555 and Cellmask Red, respectively. Images had pixel sizes of 0.68 × 0.68 microns, and a z-interval of 4 microns. For analysis and visualization, shading correction (shade only) was carried out for all images. Then, maximum intensity projections were generated using 3 z-slices. Segmentation was carried out separately on maximum intensity projections of Cellmask channels using CellPose 1.0, and masks were used to generate outlines and for intensity quantification.13 Figures were assembled with Adobe Illustrator.", "appendix": "Data availability\n\nZenodo: Antibody Characterization Report for VAPB, https://doi.org/10.5281/zenodo.13891570.14\n\nZenodo: Dataset for the VAPB antibody screening study, https://doi.org/10.5281/zenodo.14503380.15\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgment\n\nWe would like to thank the NeuroSGC/YCharOS/EDDU collaborative group for their important contribution to the creation of an open scientific ecosystem of antibody manufacturers and KO cell line suppliers, for the development of community-agreed protocols, and for their shared ideas, resources, and collaboration. Members of the group can be found below. We would also like to thank the Advanced BioImaging Facility (ABIF) consortium for their image analysis pipeline development and conduction (RRID:SCR_017697). Members of each group can be found below.\n\nNeuroSGC/YCharOS/EDDU collaborative group: Thomas M. Durcan, Aled M. Edwards, Peter S. McPherson, Chetan Raina and Wolfgang Reintsch.\n\nABIF consortium: Claire M. Brown and Joel Ryan.\n\nThank you to the Structural Genomics Consortium, a registered charity (no. 1097737), for your support on this project. The Structural Genomics Consortium receives funding from Bayer AG, Boehringer Ingelheim, Bristol-Myers Squibb, Genentech, Genome Canada through Ontario Genomics Institute (grant no. OGI-196), the EU and EFPIA through the Innovative Medicines Initiative 2 Joint Undertaking (EUbOPEN grant no. 875510), Janssen, Merck KGaA (also known as EMD in Canada and the United States), Pfizer and Takeda.\n\n\nReferences\n\nNishimura Y, Hayashi M, Inada H, et al.: Molecular cloning and characterization of mammalian homologues of vesicle-associated membrane protein-associated (VAMP-associated) proteins. Biochem. Biophys. Res. Commun. 1999; 254(1): 21–26. PubMed Abstract \| Publisher Full Text\n\nTeuling E, Ahmed S, Haasdijk E, et al.: Motor neuron disease-associated mutant vesicle-associated membrane protein-associated protein (VAP) B recruits wild-type VAPs into endoplasmic reticulum-derived tubular aggregates. J. Neurosci. 2007; 27(36): 9801–9815. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNishimura AL, Mitne-Neto M, Silva HC, et al.: A mutation in the vesicle-trafficking protein VAPB causes late-onset spinal muscular atrophy and amyotrophic lateral sclerosis. Am. J. Hum. Genet. 2004; 75(5): 822–831. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyoubi R, Ryan J, Biddle MS, et al.: Scaling of an antibody validation procedure enables quantification of antibody performance in major research applications. elife. 2023; 12: 12. Publisher Full Text\n\nCarter AJ, Kraemer O, Zwick M, et al.: Target 2035: probing the human proteome. Drug Discov. Today. 2019; 24(11): 2111–2115. PubMed Abstract \| Publisher Full Text\n\nLicciardello MP, Workman P: The era of high-quality chemical probes. RSC Med. Chem. 2022; 13(12): 1446–1459. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyoubi R, Ryan J, Bolivar SG, et al.: A consensus platform for antibody characterization (Version 1). Protocol. Exchange. 2024.\n\nBiddle MS, Virk HS: YCharOS open antibody characterisation data: Lessons learned and progress made. F1000Res. 2023; 12(12): 1344. Publisher Full Text\n\nLaflamme C, McKeever PM, Kumar R, et al.: Implementation of an antibody characterization procedure and application to the major ALS/FTD disease gene C9ORF72. elife. 2019; 8: 8. Publisher Full Text\n\nAlshafie W, Fotouhi M, Shlaifer I, et al.: Identification of highly specific antibodies for Serine/threonine-protein kinase TBK1 for use in immunoblot, immunoprecipitation and immunofluorescence. F1000Res. 2022; 11: 977. Publisher Full Text\n\nBandrowski A, Pairish M, Eckmann P, et al.: The Antibody Registry: ten years of registering antibodies. Nucleic Acids Res. 2023; 51(D1): D358–D367. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBairoch A: The Cellosaurus, a Cell-Line Knowledge Resource. J. Biomol. Tech. 2018; 29(2): 25–38. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nStringer C, Wang T, Michaelos M, et al.: Cellpose: a generalist algorithm for cellular segmentation. Nat. Methods. 2021; 18(1): 100–106. PubMed Abstract \| Publisher Full Text\n\nGonzalez Bolivar S, Ayoubi R, Alende C, et al.: A guide to selecting high-performing antibodies for VAPB (UniProt ID: O95292) for use in western blot, immunoprecipitation, and immunofluorescence.2024. Publisher Full Text\n\nLaflamme C: Dataset for the VAPB antibody screening study. [Dataset]. Zenodo. 2024. Publisher Full Text" }	[ { "id": "357897", "date": "15 Jan 2025", "name": "Mórotz Gábor Miklós", "expertise": [ "Reviewer Expertise neurodegeneration", "VAPB" ], "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 this paper Gonzales Bolivar and colleagues tested six commercially available anti-VAPB antibodies as part of the YCharOS initiative. The experimental results are good quality and clearly presented. These data allow fellow researchers to directly compare the performance of the selected antibodies side-by-side. I have only a few comments. 1. In the introduction section, please use more appropriate references for the cellular function of VAPB (e.g. Dudás EF, et al., 2021 [Ref 1]). Additionally, the major cellular functions regulated by VAPB tethering are lipid metabolism and calcium homeostasis. 2. It is not clear for me what were the exact antibody selection criteria for the study. For example, I can see monoclonal, recombinant and polyclonal antibodies on the list so clearly this was not a selection criterion although in the Limitations section is written that “project focuses on renewable (recombinant and monoclonal) antibodies”. It would be nice to add a flow-chart summarising how many antibodies are on the market and based on what criteria were these six antibodies selected for the work. In this way, the reader can decide how comprehensive this study is. 3. VAPB is an integral ER membrane protein. Therefore, for the immunofluorescent experiments it would be very informative to use an ER staining instead of a general cell staining to analyse whether the tested antibodies recognise the protein in the ER. 4. In the Results and discussion section first paragraph please correct “HeLa expresses the VAPB” to HeLa cells express the VAPB. 5. In the Methods section please correct the “peroxidase-conjugated antibodies” to horseradish peroxidase-conjugated antibodies. I also suggest using h instead of hr or hrs for the abbreviation of hour as it is more widely used in the scientific literature.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and materials provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [] }, { "id": "359970", "date": "24 Jan 2025", "name": "Sheng-ce Tao", "expertise": [ "Reviewer Expertise Antibody technology", "proteomics" ], "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\nBecause of the importance of VAPB, high quality antibody is a needed. This study evaluates six commercial antibodies against VAPB for western blot, immunoprecipitation, and immunofluorescence using standardized protocols in knockout and isogenic control cell lines. The research aims to address antibody reproducibility and provide open data to help researchers select the best antibodies for their studies.\n1. What are the criteria for selecting the six commercial antibodies? Please clarify. 2. This study only tested one cell line (HeLa). It is not necessary that the conclusion from one cell generally also works among other cell lines. 3. Since one of the major application of these validated antibodies is WB, it is highly possible that these antibodies recognize linear epitopes on VAPB. In this case, mapping the epitopes (for example, using AbMap technology), and then judge the specificity by epitope searching across the whole proteome will be a more general approach.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and materials provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [ { "c_id": "13248", "date": "31 Jan 2025", "name": "Carl Laflamme", "role": "Author Response", "response": "Dear Dr. Sheng-ce Tao, Thank you for your review and feedback on our manuscript. In response to your points: The YCharOS initiative collaborates with 12 antibody manufacturers, representing approximately 80% of all renewable antibodies commercially available. These partners have selected and provided their VAPB antibodies with us. We focus on renewable antibodies, as the YCharOS project does not undertake antibody characterization studies for proteins covered solely by polyclonal antibodies. The YCharOS workflow was recently featured and detailed in a News and Views article (https://doi.org/10.1038/s41596-024-01108-6). We acknowledge the reviewer's concern that a limitation of this study is the screening of antibodies in a single cell line. Our study serves as an antibody selection guide, but it remains essential for researchers to validate antibody specificity and performance within their own experimental context. As stated in the limitations section: “Lastly, as comprehensive and standardized procedures are respected, any conclusions remain confined to the experimental conditions and cell line used for this study. The use of a single cell type for evaluating antibody performance poses as a limitation, as factors such as target protein abundance significantly impact results (8). Additionally, the use of cancer cell lines containing gene mutations poses a potential challenge, as these mutations may be within the epitope coding sequence or other regions of the gene responsible for the intended target. Such alterations can impact the binding affinity of antibodies. This represents an inherent limitation of any approach that employs cancer cell lines”. Mapping the epitopes of the tested antibodies falls outside the scope of the YCharOS initiative. Sincerely Carl Laflamme" } ] }, { "id": "359968", "date": "05 Feb 2025", "name": "Karen Bowman", "expertise": [ "Reviewer Expertise cell 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\nThe study involved characterisation of six commercial antibodies, from five different suppliers, for the vesicle-associated membrane protein-associated protein B/C (VAPB) for use in western blot, immunoprecipitation, and immunofluorescence. The study formed part of the wider YCharOS collaboration to characterise commercial antibodies for human proteins using standardised protocols with the intention of improving antibody reproducibility issues. As a Data Note, it allows the direct comparison of the performance of commercially available antibodies to VAPB protein to aid scientists in choosing the most appropriate antibody for their studies, for example, in the biochemical and cellular assessment of VAPB properties and function. This comparison would be of particular use in the field of neurodegenerative diseases, especially in the study of amyotrophic lateral sclerosis (ALS). A committee of industry and academic representatives have endorsed the platform used, and the protocols used are consistent with those in general use. The platform consisted of identification of a human cell line suitable for antibody characterisation studies i.e., with adequate levels of the VAPB protein to generate a measurable signal. They found that the commercially available HeLa WT cell line expressed VAPB transcript at a level above the average range of cancer cells analysed, and therefore, was a logical choice for their study. The matched isogenic knockout control cell line, HeLa VAPB KO was used as an appropriate negative control. The final step was a series of antibody characterisation procedures, limited to the most common research uses of antibodies. The interpretation of the results is left to the reader, with the study providing unbiased guidance. However, it gave some indication of the optimum dilutions of commercial antibodies. Limitations of the study are clearly stated. The summary Table 3 with illustrations to assess antibody performance in western blot, immunoprecipitation and immunofluorescence, is very helpful. I think this study could have been improved by increasing the number of commercially available antibodies studied. There are at least a further seven commercial VAPB antibodies available. However, the study is a useful starting point when choosing the most appropriate antibody for study of the VAPB protein.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and materials provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [] }, { "id": "359974", "date": "05 Feb 2025", "name": "Jieya Shao", "expertise": [ "Reviewer Expertise DNA replication", "cancer 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\nIn this article the authors systematically compared the performance and specificity of six commercial anti-VAPB antibodies using three different experimental approaches (Western blot, immunoprecipitation, and immunofluorescence staining). This study is part of a large collaborative initiative to characterize commercial antibodies for different human proteins and offer unbiased guidance to researchers who can select the best antibodies for their specific experimental objectives based on their own interpretations. As a Data Note, this paper successfully achieved its goal with rigorously collected high-quality data, and the results are self-explanatory and useful for the research community.\nNo issues were found except a small grammatical error on page 3 under Results and discussion. The authors wrote \"WT and VAPB KO protein lysates were ran on SDS PAGE\". The correct word is \"run\".\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and materials 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/13-1559
https://f1000research.com/articles/13-1557/v1	24 Dec 24	{ "type": "Data Note", "title": "A guide to selecting high-performing antibodies for STXBP1 (UniProt ID: P61764) for use in western blot, immunoprecipitation, and immunofluorescence", "authors": [ "Riham Ayoubi", "Charles Alende", "Maryam Fotouhi", "Sara González Bolívar", "Carl Laflamme", "NeuroSGC/YCharOS/EDDU collaborative group", "ABIF consortium", "Riham Ayoubi", "Charles Alende", "Maryam Fotouhi", "Sara González Bolívar" ], "abstract": "The Syntaxin-binding protein 1, STXBP1, is a protein involved in docking and fusion of synaptic vesicles, a crucial event for neurotransmitters release into the synapse. Here we have characterized twelve STXBP1 commercial antibodies for western blot, immunoprecipitation, and immunofluorescence using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. These studies are part of a larger, collaborative initiative seeking to address antibody reproducibility issues by characterizing commercially available antibodies for human proteins and publishing the results openly as a resource for the scientific community. While use of antibodies and protocols vary between laboratories, we encourage readers to use this report as a guide to select the most appropriate antibodies for their specific needs.", "keywords": [ "P61764", "STXBP1", "Syntaxin-binding protein 1", "MUNC18-1", "antibody characterization", "antibody validation", "western blot", "immunoprecipitation", "immunofluorescence" ], "content": "Introduction\n\nSNARE proteins are synaptic-soluble N-ethylmaleimide-sensitive factor attachment receptors. Upon its assembly with Syntaxin 1 and other SNARE proteins, STXBP1 forms a tight complex that drives the fusion of synaptic vesicles with the presynaptic plasma membrane, allowing the exocytosis of neurotransmitters.1 STXBP1 haploinsufficiency is one of the most common genetic causes of developmental and epileptic encephalopathies.2 Identification of efficient research tools such as antibodies enables fundamental cell-based assays to understand neurological disorders related to this protein.\n\nThis research is part of a broader collaborative initiative in which academics, funders and commercial antibody manufacturers are working together to address antibody reproducibility issues by characterizing commercial antibodies for human proteins using standardized protocols, and openly sharing the data.3–5 Here we evaluated the performance of twelve commercial antibodies for STXBP1 for use in western blot, immunoprecipitation, and immunofluorescence, enabling biochemical and cellular assessment of STXBP1 properties and function. The platform for antibody characterization used to carry out this study was endorsed by a committee of industry academic representatives. It consists of identifying human cell lines with adequate target protein expression and the development/contribution of equivalent knockout (KO) cell lines, followed by antibody characterization procedures using most commercially available antibodies against the corresponding protein. The standardized consensus antibody characterization protocols are openly available on Protocol Exchange, a preprint server (DOI: 10.21203/rs.3.pex-2607/v1).6\n\nThe authors do not engage in result analysis or offer explicit antibody recommendations. Our primary aim is to deliver top-tier data to the scientific community, grounded in Open Science principles. This empowers experts to interpret the characterization data independently, enabling them to make informed choices regarding the most suitable antibodies for their specific experimental needs. Guidelines on how to interpret antibody characterization data found in this study are featured on the YCharOS gateway.7\n\n\nResults and discussion\n\nOur standard protocol involves comparing readouts from WT (wild type) and KO cells.8,9 The first step was to identify a cell line(s) that expresses sufficient levels of a given protein to generate a measurable signal using antibodies. To this end, we examined the DepMap (Cancer Dependency Map Portal, RRID:SCR_017655) transcriptomics database to identify all cell lines that express the target at levels greater than 2.5 log2 (transcripts per million “TPM” + 1), which we have found to be a suitable cut-off.3 The U-87 MG expresses the STXBP1 transcript at 5.2, which is above the average range of cancer cells analyzed. STXBP1 KO cells in U-87 MG were custom made at Abcam ( Table 1).\n\nTo screen all twelve antibodies by western blot, WT and STXBP1 KO protein lysates were ran on SDS-PAGE, transferred onto nitrocellulose membranes, and then probed with all STXBP1 antibodies in parallel ( Figure 1).\n\nLysates of U-87 MG WT and STXBP1 KO were prepared, and 40 μg of protein were processed for western blot with the indicated STXBP1 antibodies. The Ponceau stained transfers of each blot are presented to show equal loading of WT and KO lysates and protein transfer efficiency from the acrylamide gels to the nitrocellulose membrane. Antibody dilutions were chosen according to the recommendations of the antibody supplier. Antibody dilution used: ab109023 at 1/1000, ab124920 at 1/10 000, ab315893 at 1/1000, A5420 at 1/1000, NBP3-15097 at 1/2000, AF5675 at 1/200, 13414** at 1/1000, GTX114808 at 1/500, GTX114809 at 1/500, 67137-1-Ig* at 1/5000, MA5-35796 at 1/2000, PA1-742 at 1/500 (2 μg/ml). Predicted band size: 67.5 kDa. Recombinant antibody, Monoclonal antibody.\n\nWe then assessed the capability of the twelve antibodies to capture STXBP1 from U-87 MG protein extracts using immunoprecipitation followed by western blot analysis. For the immunoblot step, a specific STXBP1 antibody identified previously (refer to Figure 1) was selected. Equal amounts of the starting material (SM) and the unbound fractions (UB), as well as the whole immunoprecipitate (IP) eluates were separated by SDS-PAGE ( Figure 2).\n\nU-87 MG lysates were prepared, and immunoprecipitation was performed using 0.6 mg of lysate and 2.0 μg of the indicated STXBP1 antibodies pre-coupled to Dynabeads protein A or protein G. Samples were washed and processed for western blot with the indicated STXBP1 antibody. For western blot, NBP3-15097* was used at 1/2000. The Ponceau stained transfers of each blot are shown. SM=4% starting material; UB=4% unbound fraction; IP=immunoprecipitate, HC= antibody heavy chain. *Recombinant antibody, Monoclonal antibody.\n\nFor immunofluorescence, the twelve antibodies were screened using a mosaic strategy. First, U-87 MG WT and STXBP1 KO cells were labelled with different fluorescent dyes in order to distinguish the two cell lines, and the STXBP1 antibodies were evaluated. Both WT and KO lines imaged in the same field of view to reduce staining, imaging and image analysis bias ( Figure 3). Quantification of immunofluorescence intensity in hundreds of WT and KO cells was performed for each antibody tested, and the images presented in Figure 3 are representative of this analysis.6\n\nU-87 MG WT and STXBP1 KO cells were labelled with a green or a far-red fluorescent dye, respectively. WT and KO cells were mixed and plated to a 1:1 ratio on coverslips. Cells were stained with the indicatedSTXBP1 antibodies and with the corresponding Alexa-fluor 555 coupled secondary antibody including DAPI. Acquisition of the blue (nucleus-DAPI), green (WT), red (antibody staining) and far-red (KO) channels was performed. Representative images of the merged blue and red (grayscale) channels are shown. WT and KO cells are outlined with green and magenta dashed line, respectively. When an antibody was recommended for immunofluorescence by the supplier, we tested it at the recommended dilution. The rest of the antibodies were tested at 1 and 2 μg/ml and the final concentration was selected based on the detection range of the microscope used and a quantitative analysis not shown here. Antibody dilution used: ab109023 at 1/100, ab124920 at 1/100, ab315893 at 1/500, A5420 at 1/50, AF5675 at 1/200, NBP3-15097 at 1/500, 13414** at 1/100, GTX114808 at 1/800, GTX114809 at 1/1000, 67137-1-Ig* at 1/2000, MA5-35796 at 1/100, PA1-742 at 1/1000. Bars = 10 μm. Recombinant antibody, Monoclonal antibody.\n\nIn conclusion, we have screened twelve STXBP1 commercial antibodies by western blot, immunoprecipitation, and immunofluorescence by comparing the signal produced by the antibodies in human U-87 MG WT and STXBP1 KO cells. To assist users in interpreting antibody performanyce, Table 3 outlines various scenarios in which antibodies may perform in all three applications.3 Several high-quality and renewable antibodies that successfully detect STXBP1 were identified in all applications. Researchers who wish to study STXBP1 in a different species are encouraged to select high-quality antibodies, based on the results of this study, and investigate the predicted species reactivity of the manufacturer before extending their research.\n\nThe underlying data for this study can be found on Zenodo, an open-access repository for which YCharOS has its own collection of antibody characterization reports. A link is accessible in the “Data availability” portion of this data note.\n\nInherent limitations are associated with the antibody characterization platform used in this study. Firstly, the YCharOS project focuses on renewable (recombinant and monoclonal) antibodies and does not test all commercially available STXBP1 antibodies. YCharOS partners provide approximately 80% of all renewable antibodies, but some top-cited polyclonal antibodies may not be available through these partners.\n\nSecondly, the YCharOS effort employs a non-biased approach that is agnostic to the protein for which antibodies have been characterized. The aim is to provide objective data on antibody performance without preconceived notions about how antibodies should perform or the molecular weight that should be observed in western blot. As the authors are not experts in STXBP1, only a brief overview of the protein’s function and its relevance in disease is provided. STXBP1 experts are invited to analyze and interpret the observed banding pattern in western blot and subcellular localization in immunofluorescence.\n\nThirdly, YCharOS experiments are not performed in replicates primarily due to the use of multiple antibodies targeting various epitopes. Once a specific antibody is identified, it validates the protein expression of the intended target in the selected cell line, confirms the lack of protein expression in the KO cell line and supports conclusions regarding the specificity of the other antibodies. All experiments are performed using master mixes, and meticulous attention is paid to sample preparation and experimental execution. In IF, the use of two different concentrations serves to evaluate antibody specificity and can aid in assessing assay reliability. In instances where antibodies yield no signal, a repeat experiment is conducted following titration. Additionally, our independent data is performed subsequently to the antibody manufacturers internal validation process, therefore making our characterization process a repeat.\n\nLastly, as comprehensive and standardized procedures are respected, any conclusions remain confined to the experimental conditions and cell line used for this study. The use of a single cell type for evaluating antibody performance poses as a limitation, as factors such as target protein abundance significantly impact results.6 Additionally, the use of cancer cell lines containing gene mutations poses a potential challenge, as these mutations may be within the epitope coding sequence or other regions of the gene responsible for the intended target. Such alterations can impact the binding affinity of antibodies. This represents an inherent limitation of any approach that employs cancer cell lines.\n\n\nMethod\n\nThe standardized protocols used to carry out this KO cell line-based antibody characterization platform was established and approved by a collaborative group of academics, industry researchers and antibody manufacturers. The detailed materials and step-by-step protocols used to characterize antibodies in western blot, immunoprecipitation and immunofluorescence are openly available on Protocol Exchange, a preprint server (DOI:10.21203/rs.3.pex-2607/v1).6 Brief descriptions of the experimental setup used to carry out this study can be found below.\n\nCell lines used and primary antibodies tested in this study are listed in Tables 1 and 2, respectively. To ensure that the cell lines and antibodies are cited properly and can be easily identified, we have included their corresponding Research Resource Identifiers, or RRID.10,11\n\n Monoclonal antibody.\n\n Recombinant antibody.\n\nCRISPR/Cas9 genome editing: U-87 MG KO clone corresponding to the STXBP1 gene was generated at Abcam. Two guide RNAs were used to induce a deletion in exon 3 of the STXBP1 gene (sequence guide 1: TGGTGGATCAGTTAAGCATG, sequence guide 2: ATGACAGACATCATGACCGA).\n\nPeroxidase-conjugated goat anti-rabbit and anti-mouse antibodies are (Thermo Fisher Scientific, cat. number 65-6120 and 62-6520). Peroxidase-conjugated donkey anti-goat antibody is from Thermo Fisher Scientific (cat. number A15999). Alexa-555-conjugated goat anti-rabbit and anti-mouse secondary antibodies (Thermo Fisher Scientific, cat. number A-21429 and A-21424). Alexa-555-conjugated donkey anti-goat secondary antibody is from Thermo Fisher Scientific (cat. number A-21432). Peroxidase-conjugated Protein A for IP detection is from Cell Signaling Technology, cat. number 12291.\n\nU-87 MG WT and STXBP1 KO cells were collected in RIPA buffer (25mM Tris-HCl pH 7.6, 150 mM NaCl, 1% NP-40, 1% sodium deoxycholate, 0.1% SDS) (Thermo Fisher Scientific, cat. number 89901) supplemented with 1x protease inhibitor cocktail mix (MilliporeSigma, cat. number P8340). Lysates were sonicated briefly and incubated 30 min on ice. Lysates were spun at ~110,000 × g for 15 min at 4°C and equal protein aliquots of the supernatants were analyzed by SDS-PAGE and western blot. BLUelf prestained protein ladder (GeneDireX, cat. number PM008-0500) was used.\n\nWestern blots were performed with precast midi 4-20% Tris-Glycine polyacrylamide gels (Thermo Fisher Scientific, cat. number WXP42012BOX) ran with Tris/Glycine/SDS buffer (Bio-Rad, cat. number 1610772), loaded in Laemmli loading sample buffer (Thermo Fisher Scientific, cat. number AAJ61337AD) and transferred on nitrocellulose membranes. Proteins on the blots were visualized with Ponceau S staining (Thermo Fisher Scientific, cat. number BP103-10) which is scanned to show together with individual western blot. Blots were blocked with 5% milk for 1 hr, and antibodies were incubated O/N at 4°C with 5% milk in TBS with 0.1% Tween 20 (TBST) (Cell Signalling Technology, cat. number 9997). Following three washes with TBST, the peroxidase conjugated secondary antibody was incubated at a dilution of ~0.2 μg/ml in TBST with 5% milk for 1 hr at room temperature followed by three washes with TBST. Membranes were incubated with Pierce ECL (Thermo Fisher Scientific, cat. number 32106) or Clarity Western ECL Substrate (Bio-Rad, cat. number 1705061) prior to detection with the iBright™ CL1500 Imaging System (Thermo Fisher Scientific, cat. number A44240).\n\nU-87 MG WT were collected in Pierce IP lysis buffer (Thermo Fisher Scientific, cat. number 87788) (25 mM Tris-HCl pH 7.4, 150 mM NaCl, 1 mM EDTA, 1% NP-40 and 5% glycerol) supplemented with protease inhibitor. Lysates were rocked 30 min at 4°C and spun at 110,000 × g for 15 min at 4°C.\n\nImmunoprecipitation was performed using the KingFisher Apex purification system from Thermo Fisher Scientific (cat. number 5400930). Briefly, 2 μg of antibody were conjugated to 30 μl of Dynabeads protein A- (for rabbit antibodies) or protein G- (for mouse and goat antibodies) (Thermo Fisher Scientific, cat. number 10002D and 10004D, respectively) in 500 μl of Pierce IP Buffer. Conjugation was allowed for 30 min at 4°C followed by two 30 sec washes to remove unbound antibodies. Antibody NBP3-15097 is at an unknown concentration and therefore 5 μl were tested in the IP.\n\n0.4 ml aliquots at 1.5 mg/ml of lysate were incubated with an antibody-bead conjugate for 1h at 4°C and beads were subsequently washed three times for 30 sec in 1.0 ml of IP buffer. All steps were scripted with recurring loops of mixing to keep the beads in suspension. Elution was set at 65°C for 10 min. Unbound fractions and eluates were collected at the end of the run and processed for SDS-PAGE and western blot on precast midi 4-20% Tris-Glycine polyacrylamide gels. Protein A:HRP was used as a secondary detection system at a concentration of 0.5 μg/ml.\n\nU-87 MG WT and STXBP1 KO cells were labelled with a green and a far-red fluorescence dye, respectively (Thermo Fisher Scientific, cat. number C2925 and C34565). The nuclei were labelled with DAPI (Thermo Fisher Scientific, cat. Number D3571) fluorescent stain. WT and KO cells were plated on 96-well plate with optically clear flat-bottom (Perkin Elmer, cat. number 6055300) as a mosaic and incubated for 24 hrs in a cell culture incubator at 37oC, 5% CO2. Cells were fixed in 4% paraformaldehyde (PFA) (VWR, cat. number 100503-917) in phosphate buffered saline (PBS) (Wisent, cat. number 311-010-CL). Cells were permeabilized in PBS with 0.1% Triton X-100 (Thermo Fisher Scientific, cat. number BP151-500) for 10 min at room temperature and blocked with PBS with 5% BSA, 5% goat serum (Gibco, cat. number 16210-064) and 0.01% Triton X-100 for 30 min at room temperature. Cells were incubated with IF buffer (PBS, 5% BSA, 0.01% Triton X-100) containing the primary STXBP1 antibodies overnight at 4°C. Cells were then washed 3 × 10 min with IF buffer and incubated with corresponding Alexa Fluor 555-conjugated secondary antibodies in IF buffer at a dilution of 1.0 μg/ml for 1 hr at room temperature with DAPI. Cells were washed 3 × 10 min with IF buffer and once with PBS.\n\nImages were acquired on an ImageXpress micro confocal high-content microscopy system (Molecular Devices), using a 20x NA 0.95 water immersion objective and scientific CMOS cameras, equipped with 395, 475, 555 and 635 nm solid state LED lights (lumencor Aura III light engine) and bandpass filters to excite DAPI, Cellmask Green, Alexa-555 and Cellmask Red, respectively. Images had pixel sizes of 0.68 × 0.68 microns, and a z-interval of 4 microns. For analysis and visualization, shading correction (shade only) was carried out for all images. Then, maximum intensity projections were generated using 3 z-slices. Segmentation was carried out separately on maximum intensity projections of Cellmask channels using CellPose 1.0, and masks were used to generate outlines and for intensity quantification.12 Figures were assembled with Adobe Illustrator.", "appendix": "Data availability\n\nZenodo: Antibody Characterization Report for STXBP1, https://doi.org/10.5281/zenodo.13891600.13\n\nZenodo: Dataset for the STXBP1 antibody screening study, https://doi.org/10.5281/zenodo.14481229.14\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgment\n\nWe would like to thank the NeuroSGC/YCharOS/EDDU collaborative group for their important contribution to the creation of an open scientific ecosystem of antibody manufacturers and KO cell line suppliers, for the development of community-agreed protocols, and for their shared ideas, resources, and collaboration. Members of the group can be found below. We would also like to thank the Advanced BioImaging Facility (ABIF) consortium for their image analysis pipeline development and conduction (RRID:SCR_017697). Members of each group can be found below.\n\nNeuroSGC/YCharOS/EDDU collaborative group: Thomas M. Durcan, Aled M. Edwards, Peter S. McPherson, Chetan Raina and Wolfgang Reintsch.\n\nABIF consortium: Claire M. Brown and Joel Ryan.\n\nThank you to the Structural Genomics Consortium, a registered charity (no. 1097737), for your support on this project. The Structural Genomics Consortium receives funding from Bayer AG, Boehringer Ingelheim, Bristol-Myers Squibb, Genentech, Genome Canada through Ontario Genomics Institute (grant no. OGI-196), the EU and EFPIA through the Innovative Medicines Initiative 2 Joint Undertaking (EUbOPEN grant no. 875510), Janssen, Merck KGaA (also known as EMD in Canada and the United States), Pfizer and Takeda.\n\n\nReferences\n\nDulubova I, Khvotchev M, Liu S, et al.: Munc18-1 binds directly to the neuronal SNARE complex. Proc. Natl. Acad. Sci. USA. 2007; 104(8): 2697–2702. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMcLeod F, Dimtsi A, Marshall AC, et al.: Altered synaptic connectivity in an in vitro human model of STXBP1 encephalopathy. Brain. 2023; 146(3): 850–857. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyoubi R, Ryan J, Biddle MS, et al.: Scaling of an antibody validation procedure enables quantification of antibody performance in major research applications. elife. 2023; 12: 12. Publisher Full Text\n\nCarter AJ, Kraemer O, Zwick M, et al.: Target 2035: probing the human proteome. Drug Discov. Today. 2019; 24(11): 2111–2115. PubMed Abstract \| Publisher Full Text\n\nLicciardello MP, Workman P: The era of high-quality chemical probes. RSC Med. Chem. 2022; 13(12): 1446–1459. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyoubi R, Ryan J, Bolivar SG, et al.: A consensus platform for antibody characterization (Version 1). Protocol Exchange. 2024.\n\nBiddle MS, Virk HS: YCharOS open antibody characterisation data: Lessons learned and progress made. F1000Res. 2023; 12(12): 1344. Publisher Full Text\n\nLaflamme C, McKeever PM, Kumar R, et al.: Implementation of an antibody characterization procedure and application to the major ALS/FTD disease gene C9ORF72. elife. 2019; 8: 8. Publisher Full Text\n\nAlshafie W, Fotouhi M, Shlaifer I, et al.: Identification of highly specific antibodies for Serine/threonine-protein kinase TBK1 for use in immunoblot, immunoprecipitation and immunofluorescence. F1000Res. 2022; 11: 977. Publisher Full Text\n\nBandrowski A, Pairish M, Eckmann P, et al.: The Antibody Registry: ten years of registering antibodies. Nucleic Acids Res. 2023; 51(D1): D358–D367. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBairoch A: The Cellosaurus, a Cell-Line Knowledge Resource. J. Biomol. Tech. 2018; 29(2): 25–38. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nStringer C, Wang T, Michaelos M, et al.: Cellpose: a generalist algorithm for cellular segmentation. Nat. Methods. 2021; 18(1): 100–106. PubMed Abstract \| Publisher Full Text\n\nAyoubi R, Alende C, Fotouhi M, et al.: A guide to selecting high-performing antibodies for STXBP1 (UniProt ID: P61764) for use in western blot, immunoprecipitation, and immunofluorescence. Zenodo. 2024. Publisher Full Text\n\nLaflamme C: Dataset for the STXBP1 antibody screening study. [Dataset]. Zenodo. 2024. Publisher Full Text" }	[ { "id": "358321", "date": "16 Jan 2025", "name": "Lynne Howells", "expertise": [ "Reviewer Expertise Cancer 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 article describes the characterisation of recombinant/monoclonal Syntaxin-binding protein 1 (STXBP1) antibodies across 3 different platforms - namely western blot, immunoprecipitation and immunofluorescence. The purpose is to allow investigators to choose high performing, pre-characterised antibodies to facilitate reproducible research into their protein of choice.\nThis is a well written article using an established validation platform. There are a number of caveats to the data shown (ie, use of recombinant/monoclonals only, single replicates) but these are adequately described and justified. The delivery of the immunofluorescence platform is particularly well thought out. Minor comments: it would be useful to comment on the expected localisation of the protein in the IF platform (eg intracellular, nuclear, punctate etc). Please use capital 'L' in micro Litres rather than ml.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and materials 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/13-1557
https://f1000research.com/articles/13-697/v1	27 Jun 24	{ "type": "Systematic Review", "title": "Unveiling The Myth of High Recurrence Rate of Extracranial Arteriovenous Malformations of The Head and Neck: Systematic Review of Case Reports and Case Series", "authors": [ "Agustian Winarno Putra", "Sagung Rai Indrasari", "Camelia Herdini", "Danu Yudistira", "Agustian Winarno Putra", "Camelia Herdini", "Danu Yudistira" ], "abstract": "Background Arteriovenous malformations (AVMs) in the head and neck pose a challenge in their management due to their local aggressiveness and high recurrence risk. This study aimed to analyze literature on head and neck AVM recurrence post-treatment and identify the most effective strategy with a lower recurrence rate.\n\nObjectives To analyse existing literature on the recurrence of head and neck AVMs following treatment. Our goal was to identify the most effective treatment option with a lower recurrence rate.\n\nMethods We conducted a thorough literature search using PubMed, ScienceDirect, and Scopus, from year 2000 to the present. Our analysis focused on key endpoints, specifically the recurrence rates of head and neck AVMs following various treatment approaches.\n\nResults Out of the initial pool of 108 screened articles, a total of 83 patients were deemed suitable for inclusion in the literature review. The reviewed articles demonstrated that appropriate diagnostic tests were documented in 95% of the included studies. Among the patients, 37.3% had previously undergone interventions and were currently dealing with regrowth masses. Notably, 56.6% of patients underwent a combined approach involving both endovascular and surgical methods, while 25.3% opted for a surgical-only approach, and only 18.1% pursued an endovascular-only approach. The studies showed a promising curing rate of AVMs, with a success rate of 94%, albeit with a complication rate of 32.5%. The average follow-up duration for all patients was 26 months, with a standard deviation of 20.5 months. Out of the 83 patients, 5 experienced recurrence, with single-modality approach. Interestingly, no patients who received a multi-modality of treatments experienced recurrence or regrowth of the AVM mass within the follow up period.\n\nConclusion The multi-modality approach outperformed single-modality treatments in preventing AVM recurrence. These findings highlight the importance of a comprehensive and multidisciplinary approach in the management of these complex vascular anomalies. PROSPERO: CRD42023490871 registered on 17/12/2023", "keywords": [ "Arteriovenous Malformations", "Head and neck", "Surgery", "Endovascular", "Embolization", "Recurrence" ], "content": "Introduction\n\nArteriovenous malformations (AVMs) make up a mere 1.5% of all vascular anomalies, and they are frequently found in the head and neck region (47.4%) as well as the extremities (28.5%). There are two main types of AVMs: focal and diffuse. Focal AVMs appear as soft tissue mass and are generally diagnosed during infancy or childhood. They possess a single arterial feeder, distinct borders, and a nidus. These types of AVMs typically respond well to appropriate treatment. In contrast, diffuse AVMs extend across and tend to disrupt tissue boundaries. They are more often identified in older children and adults. Diffuse lesions are more challenging to treat due to their multiple feeding vessels, necessitating close monitoring and repeated interventions.1,2\n\nAlthough AVMs are not malignant, they can be locally aggressive and destructive, leading to complications such as severe disfigurement, ulceration, massive haemorrhage, pain and, in the worst cases, heart failure. These anomalies have tendencies to expand, occasionally undergoing sudden increases in growth, influenced by factors like trauma, hormonal changes, or iatrogenic causes The diagnosis was established based on medical history, physical examination, and usually confirmed by MR-angiography or CT-angiography.3,4\n\nThe treatment of AVMs is still controversial; there are no staging criteria or standardized guidelines, and treatment options vary from a conservative approach to more aggressive strategies. In the past, surgical excision was the predominant approach compared to endovascular embolization therapy for arteriovenous malformations (AVMs). However, the current trend is to limit surgical excision to small, localized AVMs due to the unacceptable risk of significant bleeding associated with the procedure. Endovascular embolization treatment often require multiple sessions for comprehensive closure of the AVM. When endovascular treatment is not followed by a surgical phase, potentially leading to recurrences after the natural degradation of embolic materials. A multidisciplinary approach with a combined treatment based on endovascular embolization and surgical excision is a good compromise and is rated a good choice by several studies.5–7\n\nIn this context, we present the outcomes of various modality treatment for management AVMs in the head and neck. The main objective of this research was to perform a comprehensive analysis of existing literature concerning the recurrence of head and neck AVMs following treatment. Our goal was to identify the most effective treatment option with a lower recurrence rate\n\n\nMethods\n\nWe conducted an extensive and systematic literature review based on Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Our study was registered in PROSPERO: CRD42023490871 on December 17, 2023. We identified suitable studies using the online search engines PubMed, Scopus, and ScienceDirect. The keywords for the search are “Arteriovenous Malformations”, “AVMs”, “Head and Neck”, “Endovascular”, “Surgical”. All clinical studies targeting treatment, outcome, complication and recurrence of AVMs of the head and neck were included in the primary review. Retrospective and prospective English studies, as well as case reports published between January 2000 and October 2023 were included. There were no restrictions as to the country of origin, clinical setting or size of the institution in which the treatments were performed. We did not set a minimum sample size of the studies, as any clinical findings or experience in treatment might be of value to clinicians or future studies. We excluded reports that did not specified the therapy used and follow up period.8 We also excluded patients that were loss to follow up and reported still undergoing treatment. First and second author reviewed the title, abstracts and full-text and discussed which literatures to include and exclude in the review. If authors encountered confusion, a third and fourth author were consulted to establish a consensus.\n\nThe data that this study collected including age, gender, location of AVMs, imaging modality, history of previous intervention, treatment modality, follow up period, recurrences and complications. Our analysis focused on key endpoints, specifically the recurrence rates of head and neck arteriovenous malformations following various treatment approaches. As interventional therapy of AVMs of the head and neck details on gender, age, of the treated patients were inconsistently reported, they did not undergo analysis.\n\n\nResults\n\nOur search generated a total of 162 citations, and identified a total of 108 potentially relevant articles. Of these articles, a total of 52 articles merited full text review. Based on the above-listed inclusion and exclusion criteria mentioned in our PRISMA figure (Figure 1), a total of 19 articles were included in the analysis. From the 19 articles, 83 patients (Table 1) were deemed suitable for inclusion in the literature review.\n\nThe reviewed articles demonstrated that appropriate diagnostic tests were documented among the 83 patients, a total of 79 individuals, accounting for 95%, utilized angiography, CT, or MRI, either individually or in combination, for diagnosing AVMs (Table 2). Among the clinicians, the most preferred modalities were MRI, with 68 cases (82%), and angiography, with 62 cases (75%).\n\n31 individuals among 83 patients, or 37.3%, had previously undergone treatment. This indicates a recurrence rate of 37.3% among the reported cases of AVMs in the head and neck region. It’s important to note that the treatment methods used were evenly distributed among the endovascular approach, surgical approach, and a combination of endovascular and surgical approaches. with 22.6%, 38.7%, 36.7% subsequently (Table 3).\n\nOut of the 83 reported patients, the combination of endovascular and surgical approach emerged as the most favored method. We identify notably 47 or 56.6% of patients underwent a combined approach involving both endovascular and surgical methods, while 21 patients (25.3%) opted for a surgical-only approach, and only 12 patients (18.1%) pursued an endovascular-only approach. With an average follow-up period of 26 months, combined approach exhibited a remarkable outcome with a 0% recurrence rate. In contrast, the endovascular approach had a 13.3% recurrence rate, and the surgical approach showed a 14.2% recurrence rate, making the combination approach the most successful in preventing AVM recurrence (Table 4).\n\nOut of the 83 cases, 27 complications were reported, representing a complication rate of 32.5%. The surgical approach alone demonstrated the lowest complication rate, with only 2 cases (9.5%). In contrast, the endovascular approach and the combination of endovascular and surgical approach had complication rates of 40.0% and 40.4%, respectively (Table 5). The most prevalent complication was wound dehiscence, occurring in 11 out of the 27 cases with complications, making up 40.7% of the reported complication cases (Table 6).\n\n\nDiscussion\n\nA clinical diagnosis of arteriovenous malformation is diagnosed through the patient’s medical history, physical examination and supportive examination are crucial especially MRI and angiography. MRI currently serves as the definitive method for assessing the extent of involvement within tissue planes and illustrating flow characteristics. Angiography plays a vital role in revealing the feeding vessels, offering insights into anastomoses with other extracranial or intracranial vessels, and providing details about the venous drainage pattern for ‘super selective’ embolization. During embolization, the focus should be on the nidus or epicenter of the arteriovenous malformation rather than the proximal feeding vessels.4\n\nComplete removal of AVMs is imperative to prevent recurrence. To achieve this, a customized approach must be devised for each patient, taking into consideration the specific boundaries of the lesion. The excision process may involve the removal of three different structures: fascia, muscle, and bone. In cases where preoperative embolization has not been performed, extended resection into apparently normal tissue is advised to ensure the thorough elimination of the AVMs.9\n\nThe management of AVMs remains the most debated aspect in the medical literature, with no universally accepted approach. Previously, the treatment methods were evenly distributed among the endovascular approach, surgical approach, and a combination of endovascular and surgical approaches.10 Currently, the preferred treatment involves selectively embolizing vessels combined with surgical resection and subsequent reconstruction of soft tissues. The primary goal of preoperative embolization is to minimize blood loss and enhance the surgical procedure, emphasizing that it should not be seen as a method for reducing the extent of resection. It is crucial not to postpone surgical resection beyond 48 hours after embolization, as the inflammation that ensues makes the hemodynamic benefits ineffective and renders the surgery more challenging.10\n\nRecurrence of AVMs has been reported in as much as 80% of cases following embolization or resection. In cases where the nidus is incompletely removed or embolized, there is a heightened risk of aggressive growth in the remaining lesion, leading to a potential progression risk as high as 50% within the initial five years.1 In recent literature findings, the recurrence rate of AVMs that managed with combined approach of both endoscopic and surgical approach give a remarkable outcome with a 0% recurrence rate. This result most likely because the surgeons are able to obtain a clear surgical vision field and completely excise the AVMs mass margin, as a result of the pre-embolization of the AVMs mass. The presence of a clear margin and minimal bleeding likely contributes to this successful outcome.\n\nCurrently, in our literature search, the management of AVMs mass shows a better recurrence rate than previously believed. However the average follow-up period was 26 months. In some literature recurrences have been observed a decade after treatment, highlighting the essential need for extended post-treatment follow-up to ensure timely detection. It’s crucial to acknowledge that the interpretation of the term “cure” varies in the literature, and reported instances of “cure” may be influenced by limited follow-up periods. Some cases defined “cure” as an asymptomatic state following embolization rather than a complete absence of the condition.1,11\n\nTo our understanding, several studies have documented cases and case series involving patients with AVMs in the head and neck region, who were treated using diverse approaches such as endovascular, surgical, or a combination of both. Nevertheless, as of now, no systematic review has been conducted to determine the most effective treatment approach, particularly in relation to the recurrence of the mass.\n\nThe limitations of this study is the lack of randomization due to the rarity of the case we included all studies that met the inclusion criteria regardless of the size of the lesion, race, gender, age and location. Subsequently, even though the average follow-up period was 26 months, the follow up period of the studies varies greatly ranging from 6 month to 8 years. The lack of literature addressing the recommended follow-up timeframe for AVMs mass contributes to the uncertainty. Some studies even report occurrences of AVMs mass recurrence a decade post-treatment.11\n\n\nConclusion\n\nThe multi-modality of endovascular and surgical approaches has a lower recurrence rate of AVM masses compared to a single-modality treatment. These findings highlight the importance of a comprehensive and multidisciplinary approach in contributing to the successful surgical outcomes of these complex vascular anomalies.\n\n\nEthics and consent\n\nEthical approval and written informed consent were not required.", "appendix": "Data availability statement\n\nAll data underlying the results are available as part of the article (included under extended data) and no additional source data are required.\n\nSupplementary data:\n\nFigshare: Tabel 1. Baseline Characteristic, https://doi.org/10.6084/m9.figshare.25931002.v2. 23\n\nFigshare: PRISMA Checklist https://doi.org/10.6084/m9.figshare.24995798.v2. 24\n\nFigshare: PRISMA Flowchart https://doi.org/10.6084/m9.figshare.25783014.v1. 25\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 express their gratitude to the Department of Otorhinolaryngology, Faculty of Medicine, Public Health, and Nursing at Gadjah Mada University for providing updated insights into systematic review.\n\n\nReferences\n\nFernández-Alvarez V, Suárez C, De Bree R, et al.: Management of extracranial arteriovenous malformations of the head and neck. Auris Nasus Larynx. 2020 Apr; 47(2): 181–190.\n\nAlmesberger D, Manna F, Guarneri GF, et al.: Arterio-Venous Malformations of the Nose: Combined Approach for a Successful Strategy. J Craniofac Surg. 2016 Sep; 27(6): 1524–1526. PubMed Abstract \| Publisher Full Text\n\nRajput D, Vasudevan C, Sant C, et al.: AV Malformation Within Buccinator Muscle-A Unique Finding. Indian J Otolaryngol Head Neck Surg. 2022 Dec; 74(Suppl 3): 6241–6245. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAslan S, Yavuz H, Cagici AC, et al.: Embolisation of an extensive arteriovenous malformation of the temporal region as an alternate treatment: case report. J Laryngol Otol. 2008 Jul; 122(7): 737–740. PubMed Abstract \| Publisher Full Text\n\nGennaro P, Cioni S, Bracco S, et al.: A Single-Center Retrospective Analysis of 14 Head and Neck AVMs Cases Treated with a Single-Day Combined Endovascular and Surgical Approach. J Pers Med. 2023 Apr 28; 13(5): 757. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nByatnal AA, Rakheja M, Byatnal AR, et al.: An infiltrative angioarchitectural variant of arteriovenous malformation of temporalis. J Clin Diagn Res. 2014 Sep; 8(9): ZD04–ZD06. PubMed Abstract \| Publisher Full Text\n\nChelliah MP, Do HM, Zinn Z, et al.: Management of complex arteriovenous malformations using a novel combination therapeutic algorithm. JAMA Dermatol. 2018; 154(11): 1316–1319. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPekkola J, Lappalainen K, Vuola P, et al.: Head and neck arteriovenous malformations: results of ethanol sclerotherapy. AJNR Am J Neuroradiol. 2013 Jan; 34(1): 198–204. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRos de San Pedro J, Cuartero Pérez B, Ferri Ñíguez B, et al.: Arteriovenous Malformations of the Temporalis Muscle: A Comprehensive Review. Oper Neurosurg (Hagerstown). 2018 Apr 1; 14(4): 325–340. PubMed Abstract \| Publisher Full Text\n\nPompa V, Valentini V, Pompa G, et al.: Treatment of high-flow arteriovenous malformations (AVMs) of the head and neck with embolization and surgical resection. Ann Ital Chir. 2011; 82(4): 253–259. PubMed Abstract\n\nLe Fourn É, Herbreteau D, Papagiannaki C, et al.: Efficacy and safety of embolization in arteriovenous malformations of the extremities and head and neck: a retrospective study of 32 cases. Eur J Dermatol. 2015 Jan; 25(1): 52–56. PubMed Abstract \| Publisher Full Text\n\nChimona TS, Papadakis CE, Hatzidakis AA, et al.: Arteriovenous malformation of the floor of the mouth: a case report.Eur Arch Otorhinolaryngol.2005 Nov; 262(11):939–942. PubMed Abstract \| Publisher Full Text\n\nCuong TC, Thang LM, Giang NL, et al.: Recurrent head and neck arteriovenous malformations: A case report.Radiol Case Rep.2023 Mar; 18(3):766–770. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFerrés-Amat E, Prats-Armengol J, Maura-Solivellas I, et al.: Gingival bleeding of a high-flow mandibular arteriovenous malformation in a child with 8-year follow-up.Case Rep Pediatr.2015;2015: 745718. Publisher Full Text\n\nHan HH, Choi JS, Seo BF, et al.: Successful treatment of posttraumatic arteriovenous malformation of the lower lip.J Craniofac Surg.2015 May; 26(3):e199–e201. PubMed Abstract \| Publisher Full Text\n\nHosny AS, Elmahrouky A, Balboula A, et al.: Feasibility of vessel sealing devices in surgical excision of vascular malformations– novel approach. Int J Surg Open. 2020; 23:8–16. Publisher Full Text\n\nKoshima I, Nanba Y, Tsutsui T, et al.: Free perforator flap for the treatment of defects after resection of huge arteriovenous malformations in the head and neck regions.Ann Plast Surg.2003 Aug; 51(2):194–199. PubMed Abstract \| Publisher Full Text\n\nLee YS, Goh EK, Nam SB, et al.: Multidisciplinary approach to lethal bleeding from an arteriovenous malformation in the external auditory canal.J Craniofac Surg.2013 Nov; 24(6):2179–2182. PubMed Abstract \| Publisher Full Text\n\nLilje D, Wiesmann M, Hasan D, et al.: Interventional embolization combined with surgical resection for treatment of extracranial AVM of the head and neck: A monocentric retrospective analysis. PLoS One. 2022; 17(9): e0273018. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMartines F, Immordino V: Arteriovenous malformation of the base of tongue in pregnancy: case report.Acta Otorhinolaryngol Ital.2009 Oct; 29(5):274–278. PubMed Abstract \| Free Full Text\n\nPrasad KC, Padmanabhank, Malhotra N, et al.: Arterio venous malformation of the nose and forehead.Indian J Otolaryngol Head Neck Surg.2004 Oct; 56(4):321–323. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRichter GT, Suen JY: Clinical course of arteriovenous malformations of the head and neck: A case series.Otolaryngol--Head Neck Surg.2010 Feb; 142(2):184–190. PubMed Abstract \| Publisher Full Text\n\nPutra AW, Indrasari SR, Herdini C, et al.: Table 1. Baseline Characteristic. figshare. Figure. 2024. Publisher Full Text\n\nPutra AW, Indrasari SR, Herdini C, et al.: PRISMA_checklist.pdf. figshare. 2024. Publisher Full Text\n\nPutra AW, Indrasari SR, Herdini C, et al.: PRISMA_Flowchart. figshare. Figure. 2024. Publisher Full Text" }	[ { "id": "326069", "date": "30 Sep 2024", "name": "Remco de Bree", "expertise": [ "Reviewer Expertise Head and neck cancer" ], "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 manuscript is about the recurrence rate after treatment of extracranial AVMs in the head and neck area. This systematic review is well performed and the manuscript is well written. The outcomes are focuses on recurrence. Limitations of the study are described.\nI have only three minor points -\n\n- 'Currently, in our literature search, the management of AVMs mass shows a better recurrence rate than previously believed'. Please change 'better' in 'lower'. - The aim is to emboloize the nidus, which is very difficult. If embolization is the only treatment this is very important. If it is performed pre-operatively it is less important, than embolization of the feeding arteries may be enough. - Can the authors discuss the reconstruction of the defect after surgical excision. There are suggestions that reconstruction with well vascularized tissue (thus avoiding local hypoxia) may reduce the risk of recurrence.\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": [ { "c_id": "12966", "date": "24 Dec 2024", "name": "Agustian Winarno Putra", "role": "Author Response", "response": "First of all, we would like to thank Remco de Bree the detailed review, which has greatly improved this publication - 'Currently, in our literature search, the management of AVMs mass shows a better recurrence rate than previously believed'. Please change 'better' in 'lower'. We already make adjustment a suggested - The aim is to emboloize the nidus, which is very difficult. If embolization is the only treatment this is very important. If it is performed pre-operatively it is less important, than embolization of the feeding arteries may be enough. - Can the authors discuss the reconstruction of the defect after surgical excision. There are suggestions that reconstruction with well vascularized tissue (thus avoiding local hypoxia) may reduce the risk of recurrence. We added a publication by Balakrishnan et al (2023.) in the publication it indicates that free flaps, offering a robust and uniform blood supply, are significantly more effective in preventing recurrence compared to pedicled flaps or skin grafts. One study highlighted a 0% recurrence rate with free flaps, in contrast to a 64% recurrence rate with pedicled flaps and grafts. Additionally, patients undergoing reconstruction with free flaps after wide local excision achieved markedly better functional and aesthetic outcomes, with 87.5% reporting good-to-excellent results and no recurrences observed during the follow-up period This evidence is back up with the theory from Lu et al (2011) that stated recurrence of AVMs is often associated with neovascularization driven by factors such as hypoxia-inducible factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF. These factors are upregulated in hypoxic conditions, promoting the recruitment of endothelial progenitor cells and new vessel formation Thank you" } ] }, { "id": "339828", "date": "26 Nov 2024", "name": "Mustafa Ismail", "expertise": [ "Reviewer Expertise Neurosurgery" ], "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 systematic review on extracranial arteriovenous malformations (AVMs) provides valuable insights into recurrence rates following various treatment modalities, with an emphasis on the benefits of a multi-modality approach. However, it could be enhanced by addressing a few points. Firstly, the discussion on the role of reconstruction with well-vascularized tissue after surgical excision should be expanded to explore its potential in minimizing local hypoxia and reducing recurrence. Secondly, the interpretation of cure and recurrence rates might benefit from clarifying follow-up duration variations and their impact on outcomes. Lastly, incorporating long-term follow-up data and analyzing outcomes across diverse demographic groups could further substantiate the findings. - Please recheck the PRISMA diagram for No of studies exclusion.\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\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": [ { "c_id": "12967", "date": "24 Dec 2024", "name": "Agustian Winarno Putra", "role": "Author Response", "response": "First of all, we would like to thank Mustafa Ismail for the detailed review, which has greatly improved this publication - Firstly, the discussion on the role of reconstruction with well-vascularized tissue after surgical excision should be expanded to explore its potential in minimizing local hypoxia and reducing recurrence. Secondly, the interpretation of cure and recurrence rates might benefit from clarifying follow-up duration variations and their impact on outcomes. Lastly, incorporating long-term follow-up data and analyzing outcomes across diverse demographic groups could further substantiate the findings. We added a publication by Balakrishnan et al (2023.) in the publication it indicates that free flaps, offering a robust and uniform blood supply, are significantly more effective in preventing recurrence compared to pedicled flaps or skin grafts. One study highlighted a 0% recurrence rate with free flaps, in contrast to a 64% recurrence rate with pedicled flaps and grafts. Additionally, patients undergoing reconstruction with free flaps after wide local excision achieved markedly better functional and aesthetic outcomes, with 87.5% reporting good-to-excellent results and no recurrences observed during the follow-up period this evidence is back up with the theory from Lu et al (2011) that stated recurrence of AVMs is often associated with neovascularization driven by factors such as hypoxia-inducible factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF. These factors are upregulated in hypoxic conditions, promoting the recruitment of endothelial progenitor cells and new vessel formation While this topic has piqued our interest, we believe it warrants a more detailed review and in-depth discussion in a separate publication. - Please recheck the PRISMA diagram for No of studies exclusion. During the revision process, we re-evaluated all the data and found an error in our initial assessment. Specifically, we mistakenly stated that 33 studies were excluded, resulting in 19 studies being included in the final analysis. In fact, 32 studies were excluded, and 20 studies were included in the final analysis. There are no changes to the actual number of final patients eligible for analysis. We apologize for the error in which we incorrectly stated that the patient reported by Ferres et al. (2015) underwent both endovascular and surgical approaches. In fact, the treatment was solely via the endovascular approach using a sclerosing agent. Additionally, we rearranged the order of patients in the table to match the original publication, making it easier to cross-check the data." } ] } ]	1	https://f1000research.com/articles/13-697
https://f1000research.com/articles/13-1556/v1	24 Dec 24	{ "type": "Research Article", "title": "Epidemiological, Gross Morphological, And Histopathological Analysis Of Postmortem Cases Of Hanging - An Observational Study", "authors": [ "Alana Chacko", "Chandni Gupta", "Vikram Palimar", "Deepak Nayak M", "Alana Chacko", "Chandni Gupta", "Deepak Nayak M" ], "abstract": "Background Death due to hanging is commonly seen. Sometime the ligature mark will be very prominent and in some cases it might not. In those cases, the post-mortem examination with its gross morphological findings and histopathological analysis, becomes significantly important. Objective: The objective of this study was to identify the patterns of epidemiological, gross morphological, and histopathological features in hanging cases.\n\nMethods The study was conducted on 46 cases of hanging. Their sociodemographic features were collected. Gross morphological analysis of the ligature mark and various measurements were taken. Later tissue from the ligature mark was sent for histopathological analysis. Statistical analysis was performed on the collected parameters.\n\nResults Significant male preponderance was observed in hanging cases. The maximum number of cases of hanging were observed in the fourth decade of life. Daily wage workers were the most significantly affected population. Depression was cited as the cause of suicide in most cases. Carotid intimal tears and injuries to bony and cartilaginous structures were present in smaller numbers. Vital reaction was observed in all 46 cases. Classification of fwound vitality showed a correlation to known time since injury.\n\nConclusion The results of the present study showed that socio-demographic factors play a significant role in the circumstances leading to suicidal deaths. Therefore, in reducing the burden of suicidal deaths, these factors must be addressed. To identify the cause of death as hanging analysis of various gross features and histopathology are very important and should be conducted carefully.", "keywords": [ "Hanging", "Histopathology", "Ligature mark", "Postmortem cases", "Vital Reaction" ], "content": "Introduction\n\nHanging is defined as a “form of asphyxia which is caused by the suspension of the body by a ligature which encircles the neck, the constricting force being the weight of the body”.\n\nBased on the degree of suspension, hanging is classified into:\n\n1) Complete hanging – Here the body is fully suspended without any part thereof touching the ground.\n\n2) Partial hanging – Here the body is partially suspended with feet or knees touching the ground or the body assuming a sitting or kneeling position.\n\nBased on the position of the knot in the ligature material, hanging is classified as:\n\n1) Typical hanging – wherein the knot is present at the back of the neck, in the midline, and the ligature mark is bilaterally upward and symmetrical.\n\n2) Atypical hanging – wherein the knot of the ligature material is present at any position other than the center of the back of the neck or occiput.1\n\nAs per the 2021 and 2022 data of the National Crime Records Bureau, in India, hanging is the most adopted method of suicide, followed by a wide margin apart, poisoning, and drowning.2 This may be due to the ease of accessibility of this method, and the potential of a less agonizing death. Instances of mass suicides, such as the Burari case, have also been characterized by mass suicidal hangings. Apart from judicial and suicidal hanging, lynching (of which one of the methods is homicidal hanging) has also been reported in various parts of India, including a case in Uttar Pradesh, where two Dalit girls were subjected to gang-rape and subsequently hung.3\n\nVarious studies have been done over the years, analyzing the socio-demographic factors, the nature and cause of the act, gross findings on autopsy, and histopathology findings. These studies show non-uniform patterns concerning the parameters studied.\n\nWhile regional differences may explain the variations in socio-demographic parameters, the marked variations in gross and histopathological findings cannot be attributed to the same. The question then arises of the best approach to autopsy in cases of hanging deaths, which best preserves and helps elicit the maximum number of findings, as well as helps arrive at a conclusive pathognomic finding in hanging cases. Hence, a more scientific approach should be implemented to assess the various techniques and signs observed.\n\nThe post-mortem examination with its gross morphological findings and histopathological analysis, becomes significantly important in cases where the characteristic ligature mark is inconspicuous or absent. In such cases, other marks of ante-mortem hanging are investigated. This includes the careful “bloodless” layer-by-layer dissection of the neck, looking for any features of injury, as well as the microscopic histopathological analysis of the skin from the suspected area of compression, to look for the ‘vital reaction’, which is currently the accepted ‘gold standard’. However, since the peri-mortem period and early post-mortem period injuries pose a significant challenge in microscopically assessing wound vitality, newer methods such as enzyme histochemistry and immunohistochemistry, are upcoming and in use.\n\nAlthough advanced immunohistochemistry techniques yield significant results, the cost of the same and its practical application greatly limits its use. It is then that the routine microscopy comes back into the spotlight. Hence, there exists a need to identify a good predictive histopathological marker of antemortem injury and wound vitality in hanging cases.\n\nConsidering these facts, a knowledge gap exists, in the patterns of epidemiological, gross morphological, and histopathological features in hanging cases, where prior similar studies have not been conducted. Therefore, the present study was conducted to study the epidemiological profiles of autopsy cases of hanging, including socio-demographic patterns and psycho-social elements contributing to the manner of death, to study the gross morphological appearance of neck structures during external and internal examination, injuries and signs of asphyxia, in autopsy cases of hanging, to study the histopathological changes in skin and underlying tissues from the area of ligature mark and its adjacent areas, and to study histopathological determination of time since injury (wound vitality) in autopsy cases of hanging.\n\n\nMethods\n\nInstitutional Ethical committee clearance (IEC:485/2022) was taken before starting the study from the Kasturba Medical College and Kasturba Hospital institutional ethics committee on 8th February, 2023.\n\nWe used the STROBE reporting guidelines for our study; a completed checklist is available under Reporting Guidelines.4\n\nStudy Period: The current research was carried out from February 2023 to June 2024\n\nStudy location: Department of Forensic Medicine in association with the Department of Pathology.\n\nSample Size: 46 Autopsy cases.\n\n\n\n• Cases of hanging deaths brought to Mortuary.\n\n• Cases in which the time of death has been established and is known.\n\n\n\n• Decomposed bodies are excluded.\n\nDetailed description of procedure/processes:\n\nEthics committee has given the exemption from taking consent from the relatives of the deceased since these cases are postmortem cases where the law gives the consent for autopsy and determination of cause of death. In the above cases the samples were taken from death due to hanging so, the tissue must be taken out to determine the cause of death. The consent given by the law for autopsy is valid. The following details were collected from the police inquest documents, police intimation documents, bystanders of the deceased, and police:\n\n1) Age, sex, occupation, marital status.\n\n2) Case background, history of intoxication, place, date, and time of occurrence of the death.\n\n3) Post-mortem number, date, and time of receiving the body in the mortuary.\n\nFollowing the collection of the above data, and subject to fulfilment of inclusion criteria, the case was selected for the study and subjected to detailed post-mortem examination (autopsy). The external examination includes noting the presence or absence of ligature material in situ around neck. Type and manner of hanging was noted down. If present, type of ligature material used, type and position of knot, and any other significant features were noted, after which ligature material was secured with twine, ligated, and removed. Following this, clothes were examined and removed. The visible ligature mark was assessed for location with respect to the thyroid cartilage, and whether completely or incompletely encircling the neck was noted (Figure 1). The rest of the general post-mortem external examination is conducted. After noting all the external features, the neck was dissected for internal examination.\n\nThe linear incision for thorax and abdominal examination and evisceration is made from suprasternal notch to pubic symphysis, avoiding the umbilicus. A modification of Lettulle’s technique is used and evisceration is done, removing structures below the level of the lower end of the trachea corresponding to the sternoclavicular joint to the recto-sigmoid junction. Cranial dissection including stripping of dura is done, thus creating a bloodless field of dissection in the neck.\n\nA 12-20 cm high block is placed underneath the shoulders, thus extending the neck for dissection. The skin incision is extended from the suprasternal notch to the symphysis menti. A 5 × 2 cm × striated muscle deep, tissue sample from the maximum compression area in the ligature mark, along with the tissue present above and below the ligature mark, is taken. The two latter areas are marked using India ink and the entire sample is preserved in 10% neutral buffered formalin and sent for histopathological examination. Using toothed forceps to hold the skin, dissection is done through the plane of the underlying subcutaneous tissue, up to the lower margins of the mandible, thus exposing the platysma muscle. The platysma muscle and deep cervical fascia are reflected to expose the muscles of the neck. The sternocleidomastoid muscle is identified, and its sternal and clavicular attachments are cut, and muscle is reflected upwards. This reveals the omohyoid muscle underneath. The inferior belly of the omohyoid muscle is cut and the muscle is reflected upwards, along with the median raphe and superior belly. The sternohyoid and the sternothyroid muscles are then identified and reflected upwards. This exposes the thyroid gland, which is dissected at the isthmus and reflected outwards to reveal the trachea underneath. Blunt dissection is used to separate the laryngeal apparatus, trachea, and pharynx from prevertebral tissue.\n\nOpening the mouth, the tongue is pushed backward and upwards using forceps. Using a tongue knife inserted under the mandible, the attachment of muscles of the floor of the mouth and neck muscles are dissected. The dissection is continued backward, taking care to avoid the carotid arteries, and downwards up to the suprasternal notch and the pharynx and laryngeal structures are dissected out. The carotid arteries are divided in the neck and then removed. The hyoid bone is separated from its muscular attachments and examined. Later, the tissue was taken out for histopathological examination.\n\nThe tissue was allowed to fixate for a minimum of eight hours in neutral buffered formalin. The tissue was dissected methodically beginning from the epidermis extending down to the dermis and the hypodermis. The sections were passed entirely and placed in tissue processor. The sections were embedded using paraplast as the embedding media. The section cutting was done using a 5-micrometre thickness and serial sections of the tissue (3 in number) were placed on the glass slide for staining. Staining was done by using hematoxylin and eosin (automated method). Special stains such as Masson trichrome (for collagen and fibrin) & elastic van Gieson (for elastin) were performed. The slides were mounted using DPX as the refractory medium. The slides were reviewed separately by a pathology resident and subsequently by a consultant, and observations were noted and classified according to Dettmeyer’s classification.5\n\nStatistical analysis was done for the obtained parameters and results are presented in percentage, tables and graphs/charts.\n\nUnderlying data is included in the Underlying Data section.6\n\n\nResults\n\nThe age distribution of the deceased in the study ranges from 15 to 82 years. The mean age was 43.9 years. The maximum number of cases was observed in the age group of 41 to 50 years, with a significant male preponderance in this age group as Shown in Table 1.\n\nAmong the deceased, the maximum number were males 37 (80.4%) when compared to females 9 (19.6%).\n\nOf the deceased, 30 (65.2%) were married, 15 (32.6%) were unmarried and 1 (2.2%) were divorced. Among the married population, males were observed in more significant numbers (24 males).\n\nThe various occupations were classified into student, professional, clerical work, daily wage worker, housewife (homemakers), and unemployed. Among these, the highest incidence was among the daily wage workers (37%). The occupation-wise distribution is given in Table 2.\n\nThe case density based on location is given in Table 3.\n\nIn 12 (26.1%) of the cases, there was associated intoxication by means of consumption of alcohol, of which the maximum number were found in the age group of 41 – 50 years. In 34 (73.9%) of cases there was no intoxication seen.\n\nDepression 19 (41.3%) was the most cited reason for suicide in both males and females, with the greatest number of cases recorded in the 21-30 years, 31-40 years and 41-50 years age groups (4 cases per group). This was followed by health issues 10 (21.7), family conflict 7 (15.2), financial issues 5 (10.9), psychiatric illness 4 (8.7%) and work stress 1 (2.2%).\n\nCongestion of the face was present in 15 (32.6%) of cases, and bluish discoloration of fingernails (cyanosis) was present in 42 (91.3%) of cases, however, oedema of the face was observed only in 2 (4.3%) of cases, and petechial haemorrhages were not observed in any cases.\n\nSalivary dribbling was observed in 4 (8.7%) of cases whereas the characteristic ‘La facie sympathique was not observed in any cases.\n\nPost-mortem lividity was observable in all cases, out of which, in 13 (28.3%) lividity was fixed and in 33 (71.7%), lividity was not fixed. Faecal stains alone were observed in 4 (8.7%) of cases, and seminal stains alone in 21 (45.7%) of cases. Both faecal and seminal stains were present in 5 (10.9%) of the cases. No urinary stains were observed in any of the cases. Subconjunctival haemorrhages were observed in 2 (4.3%) of the cases. 12 (26.1%) of the cases showed associated injuries.\n\nLigature material\n\nOut of the 46 autopsy cases, ligature material was absent during post-mortem examination in 5 of the cases. Therefore, they could not be assessed. The various ligature materials that were observed include nylon rope 19 (46.3%), saree 5 (12.2%), shawl 5 (12.2%), unrecognizable cloth 3 (7.3%), bedsheet 2 (4.9%), towel 2 (4.9%), dhothi 2 (4.9%), coir rope 1 (2.4%), electrical wire 1 (2.4%), and luggage belt 1 (2.4%).\n\nThe ligature mark was found to run upwards and backwards from midline of the neck and was situated at a point above the thyroid cartilage in all the cases studied. Type and position of knot is shown in Tables 4 and 5.\n\nAll the autopsy cases of hanging in the study were suicidal in nature. 32 (69.6%) were cases of partial hanging while 14 (30.4%) of cases were complete hanging.\n\nThe subcutaneous tissue in 44 (95.7%) of the cases was pale and glistening in nature, and unremarkable in 2 (4.3%) of the cases. Strap muscle contusions were observed in 2 (4.3%) of cases with one case showing contusion of both the sternohyoid and the sternothyroid muscle and one case of contusion of only the sternothyroid muscle.\n\nContusion of the sternocleidomastoid muscle was observed in 6 (13%) of the cases, all of them being cases of complete hanging. Out of these, most cases (3 in number) were those of atypical position of knot. Saree was used in two of the above cases. Other muscles of the neck were unremarkable in all cases studied.\n\nCarotid intimal tears were noted in 3 (6.5%) of the cases, of which all were cases of complete hanging with atypical position of the knot of ligature material. Nylon rope, saree and unrecognizable cloth material were used in each of the cases. No instances of thyroid gland injuries were noted.\n\nHyoid bone and cervical vertebrae fractures were observed in 8 (17.4%) and 1 (2.2%) of the autopsy cases. Of the cases with hyoid bone fracture, most cases had an atypical position of the knot with nylon rope being used as the ligature material (4 cases). There was an even distribution of complete and partial hanging cases with hyoid bone fractures. No fractures were observed among the cartilages of the larynx.\n\nContusion of the hyoid bone was seen in 7 (15.2%) of cases, with most cases being those of complete hanging (5 cases), atypical position of knot (4 cases) and with nylon rope as the ligature material (4 cases). Contusion of thyroid cartilage was seen in 5 (10.9%) of cases, with distribution roughly like that of hyoid bone contusion. Contusions were not observed among other cartilages of larynx or cervical vertebrae.\n\nIn the tissue corresponding to the region of ligature mark, vital reaction was observed in 100% of cases. The regions above and below the region of ligature mark did not show features of extravasation of red blood cells or any features of vital reaction in any of the cases.\n\nTime since injury was broadly divided into fresh vital injury, vital wound, wound incurred shortly before or after death, not yet old and no longer fresh. The pattern of distribution of time since injury is given in the Table 6. Histopathological pictures of the tissues are shown in Figures 2-6.\n\n(A) – H&E, 100×, (B) – H&E – 400×.\n\nCategory 1. Wound incurred shortly, before or after death is shown in Figure 2. The epidermis is intact. The dermal collagen and vessels are unremarkable. No active immune reaction is noted. Category 2. Vital wound-inflicted during lifetime is shown in Figure 3. The epidermis is eroded (black arrow) and the dermal collagen shows disruption of fibroblasts (arrow heads). Category 3. Fresh vital injury (hours to a few days) is shown in Figure 4. The epidermis is eroded. The dermis shows a reaction with RBC extravasation (arrowhead) and vessels with fibrin thrombi (black arrow). Category 4. Vital wound-no longer fresh. (few days to weeks-in single digit range) is shown in Figure 5. The dermis shows a dense neutrophilic reaction (black arrow) with admixed mononuclear cells and macrophages, new vessel formation (arrow heads). Category 5. Vital Injury-Not very old (weeks to months) is shown in Figure 6. The epidermis is regenerating. Thick sclerosed collagen (arrow heads), representing scar tissue; with loss of dermal adnexal structures and scant inflammation.\n\n(A) – H&E, 100×, (B) – H&E – 400×.\n\nFresh vital injury (hours to a few days). (A) – H&E, 100×, (B) – H&E – 400×.\n\n(few days to weeks-in single digit range). (a) – H&E, 400×, (b) – H&E – 400×.\n\nH&E, 100×.\n\n\nDiscussion\n\nViolent deaths resulting from asphyxia predominantly involve hanging. Medico-legal issues likely to develop in case of hanging are primarily, whether the death was caused by hanging was suicidal, homicidal or accidental. Simulated suicidal hanging hinders the investigation procedure in unnatural deaths. Therefore, detailed external and internal examination, and analysis of samples play a significant role in arriving at conclusion. Apart from postmortem the ligature material used, location, point of suspension and evaluation of scene of crime might add to the inference.7\n\nPrevious studies associating gender connotations with hanging as a method of suicide have found that men were most found to adopt this method, as observed in studies by Denning et al. and Mergl et al.8,9 In studies done in the state of Karnataka, similar findings were observed concerning the male preponderance in hanging cases.10,11 The findings in the current study are comparable to the above-referenced studies, with a male-to-female ratio of 4.1:1. However, this ratio is more than two-fold higher as compared to the various other studies by Dekal et al., Haq et al., Karthik et al., Kumar et al, and Biradar et al. in Karnataka, wherein the ratio was only 1.5:1 to 1.13:1. It was also observed that the highest number of cases occurred in the age group of 41 to 50 years, which deviates from the findings in the above studies.12–16\n\nIt was also observed that among the deceased, the population of married people was significantly higher than the unmarried population, which was comparable to similar studies done by Karthik et al.14\n\nDepression was the leading cause of suicide by hanging in the present study (41.3%), followed by health issues (21.7%), and family conflict (15.2%). This finding becomes significant due to the reason that the previously cited various other studies have not attributed the cause of suicides by hanging to depression. Yang et al. studied the trends of depressive disorder over a period of 30 years in 204 countries and found that depression was one of the leading causes global disease burdens.17 Hence, this finding becomes more harrowing and points to the need for stepping up efforts in mental health programs.\n\nThough it has been suggested that ‘asphyxial stigmata’ or signs of asphyxia have become redundant findings in forensic practice, they are still observed and recorded in routine autopsies.18 This is because, although they may be inconclusive findings, they still direct the examiner towards a particular modality of death, especially in cases with no visible cause of death or reliable history. The study by James and Silcocks observed petechial hemorrhages in 23% of cases and congestion of face in 5% of cases.19 Lockyer BE found an average incidence of 25% for petechiae.20 In this study, congestion of the face was noted in 32.6% of cases and cyanosis in 91.3% of cases, comparable to existing literature. However, edema of the face and subconjunctival hemorrhages were observed only in 4.3% of cases, and petechial hemorrhages were not observed.\n\nClassically described signs, such as salivary dribbling and ‘la facie sympathique, were low (4.3%) and absent respectively, in the study. This contrasts with existing literature.1,21 Deaths due to hanging are often associated with involuntary urination, defecation, or ejaculation.1 The present study found 45.7% of cases to have seminal stains, 8.7% of cases to have fecal stains and 10.9% of cases having both seminal and fecal stains. However, urine stains were not observed in any of the cases.\n\nExamination of the subcutaneous tissue revealed it to be pale and glistening in 95.7% of cases, which was comparable to previous studies by Suarez-Penaranda et al., Jiwane et al., and Simonsen.22–24 4.3% of cases showed contusion to strap muscles of the neck and 13% of cases showed contusion to the sternocleidomastoid muscle, which are comparable to previous studies20,22,23,25\n\nCarotid intimal tears (Amussat’s sign) were observed in 6.5% of cases, comparable to existing trends in previous studies.26\n\nThe incidence of hyoid bone contusion (15.2%), hyoid bone fracture (17.4%), and thyroid cartilage contusion (10.9%), were significantly lower than in previous studies by Eisenmenger and Betz, Suarez-Penaranda, and Nikolic et al.22,27,28 However, they were comparable to studies by Kokatanur et al. and Feigin G.29,30 The incidence of cervical vertebrae fracture (2.2%), was comparable to studies by Suarez-Penaranda et al. and Jayaprakash et al.22,31\n\nIn the current study, vital reactions (including features such as hemorrhage, inflammatory self- infiltration) were observed in all the cases included in the study. These findings were in alignment with those from studies by Kokatanur et al. and Prasad et al.30,32\n\n\n\n1) Limited number of cases for study in the study period.\n\n2) Advanced enzyme histochemistry and immunohistochemistry techniques were not used, which can be done in future studies.\n\nThis study was an attempt to bridge the existing knowledge gap in suicidal cases of hanging. It is evident from this study that socio-demographic factors play a significant role in the circumstances leading to suicidal deaths. Therefore, in reducing the burden of suicidal deaths, these factors must be addressed. The analysis of various gross features shows that the findings in existing literature are not always observed practically and in real-world situations. Finally, further research on the estimation of wound vitality and a consensus on the definition of the term ‘vital reaction’ will be of great forensic value.\n\nEthical clearance was taken from the Kasturba Medical College and Kasturba Hospital institutional ethics committee on 8th February, 2023. Approval no: (IEC:485/2022).", "appendix": "Data availability\n\nFigshare: Epidemiological, Gross Morphological, And Histopathological Analysis Of Postmortem Cases Of Hanging - An Observational Study. https://figshare.com/articles/dataset/Data_sheet/27917280?file=51057245.6\n\nThe project contains the following underlying data: Data sheet of all 46 cases.\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: Epidemiological, Gross Morphological, And Histopathological Analysis Of Postmortem Cases Of Hanging - An Observational Study. https://figshare.com/articles/online_resource/Strobe_checklist/27917358?file=50835189.4\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\nNil.\n\n\nReferences\n\nReddy NKS: The essentials of forensic medicine and toxicology. 33rd ed.New Delhi: Jaypee Brothers Medical Publisheres (P) Ltd; 2014; 360.\n\nभारत में आकस्मिक मौतें एवं आत्महत्याएँ वर्षवार\|National Crime Records Bureau. http\n\nMcCarthy J: Double rape, lynching in India exposes caste fault lines.2014 Jun. 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Publisher Full Text\n\nSimonsen J: Patho-anatomic findings in neck structures in asphyxiation due to hanging: A survey of 80 cases. Forensic Sci. Int. 1988; 38(1–2): 83–91. PubMed Abstract \| Publisher Full Text\n\nSharma BR, Harish D, Sharma A, et al.: Injuries to neck structures in deaths due to constriction of neck, with a special reference to hanging. J. Forensic Leg. Med. 2008; 15(5): 298–305. PubMed Abstract \| Publisher Full Text\n\nAsirdizer M, Kartal E: Neck vascular lesions in hanging cases: A literature review. J. Forensic Leg. Med. 2022; 85: 102284. PubMed Abstract \| Publisher Full Text\n\nBetz P, Eisenmenger W: Frequency of Throat-Skeleton Fractures in Hanging. Am. J. Forensic Med. Pathol. 1996; 17(3): 191–193. PubMed Abstract \| Publisher Full Text\n\nNikolić S, Živković V, Babić D, et al.: Hyoid-laryngeal fractures in hanging: where was the knot in the noose? Med. Sci. Law. 2011; 51(1): 21–25. PubMed Abstract \| Publisher Full Text\n\nFeigin G: Frequency of Neck Organ Fractures in Hanging. Am. J. Forensic Med. Pathol. 1999; 20(2): 128–130. PubMed Abstract \| Publisher Full Text\n\nKokatanur CM, Chetankumar R, Patil VR: Fractures of Hyoid Bone and Thyroid Cartilage in Hanging-an Autopsy Study. Medico-Leg Update. 2016; 16(2): 146. Publisher Full Text\n\nJayaprakash S, Sreekumari K: Pattern of Injuries to Neck Structures in Hanging—An Autopsy Study. Am. J. Forensic Med. Pathol. 2012; 33(4): 395–399. PubMed Abstract \| Publisher Full Text\n\nPrasad KJ, Khalid MA, Narayana BL, et al.: Ligature Mark in Hanging-Gross and Histopathological Examination with Evaluation and Review. Indian J. Forensic Med. Toxicol. 2017; 11(1): 22. Publisher Full Text" }	[ { "id": "354484", "date": "17 Jan 2025", "name": "Priyadarshee Pradhan", "expertise": [ "Reviewer Expertise Forensic Pathology", "Forensic Anthropology" ], "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 type of observational study can be specified under study type in the Methods section ( Suggestion - cross sectional study) 2. In the last paragraph of introduction, it is mentioned that no similar studies have been conducted in the past. How did the authors ensure this statement? Suggestion: Can give an explanation or avoid the phrase. 3. How did the authors arrive at a sample size of 46?\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": "354475", "date": "31 Jan 2025", "name": "K R Nagesh", "expertise": [ "Reviewer Expertise Forensic Pathology", "Forensic Anthropology", "Forensic Odontology", "Forensic Toxicology" ], "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 analyzed the demographic details in fatal hanging cases during the study period. The following are my suggestions. Abstract:\n\nThe duration of the study needs to be added in the methods. In results section, the last sentence needs spelling correction (wound vitality).\nIntroduction:\nThe first three paragraphs (definition and classification of hanging) can be combined and put under one paragraph, and reference needs to be added wherever is appropriate. Various studies have been done over the years … Reference needs to be added.\nMethod:\nIt is better to write the initial part of methods (study type to exclusion criteria) in descriptive manner. Type of study can be changed to ‘Cross-sectional observational study’. Statistical analysis: The statistical tool used to enter data and type of analysis done needs to be added.\nConclusion:\nConclusion needs to be added after discussion.\nStrengths:\nAuthors have studied the socio-demographic findings related to hanging cases during the study period, and analyzed the gross and histopathological findings seen at ligature mark in hanging cases. The results are well presented. Weakness:\nSample size is less and authors have mentioned under limitations. The writing part under few sections as mentioned above needs to be addressed.\nRecommendation: I recommend for ‘Acceptance after undertaking the above corrections’.\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? Partly", "responses": [] } ]	1	https://f1000research.com/articles/13-1556
https://f1000research.com/articles/13-1555/v1	24 Dec 24	{ "type": "Review", "title": "Innovative approaches to benign prostatic hyperplasia management: Options beyond conventional treatment strategies", "authors": [ "Peter Chinedu Agu", "Deusdedit Tusubira", "Patrick Maduabuchi Aja", "Peter Chinedu Agu", "Deusdedit Tusubira" ], "abstract": "This review explored Benign Prostatic Hyperplasia (BPH), a prevalent condition among aging males to elucidate its multifaceted nature and emerging treatment options. It meticulously examined the pathophysiology, risk factors, and clinical presentation of BPH to shed light on its profound impact on the quality of patients’ lives. The review comprehensively presented management strategies, including pharmacological and surgical interventions, emphasizing the importance of tailored or individualized treatment plans. Furthermore, it scrutinized the emergence of technologies like laser therapy and minimally invasive procedures for their efficacy and safety profiles in managing BPH. The review culminates by underscoring the imperative for ongoing research endeavors to refine therapeutic approaches and optimize outcomes for individuals grappling with BPH to pave the way for enhanced patient care and well-being.", "keywords": [ "Benign Prostatic Hyperplasia (BPH)", "Management Strategies", "Emerging Technologies", "Quality of Life", "Personalized Treatment." ], "content": "1. Introduction\n\nA common ailment that primarily affects men, especially as they age, is benign prostatic hyperplasia, or BPH. A noncancerous enlargement of the prostate gland is indicative of this disorder. Some LUTS (lower urinary tract symptoms) that may result from it include the necessity to urinate more frequently, more urgently, at night, and difficulty urinating.1,2 The substantially reduced quality of life caused by these symptoms emphasizes the urgent need for thorough and efficient management techniques.3 The primary methods of treating BPH have been laser treatment, TURP (transurethral resection of the prostate), and drugs such as 5-α-reductase inhibitors and α-blockers.4 Studies have subtly shown that these interventions enhance urine flow and reduce symptoms. But they do have some drawbacks.5 Pharmacological therapies can have adverse effects such as weariness, dizziness, and sexual dysfunction and frequently call for long-term adherence.6 Although surgical procedures are effective, they have risks, including the possibility of infection, bleeding, and consequences such as retrograde ejaculation.7\n\nRecent pieces of literature highlight the emergence of novel therapeutic strategies aimed at addressing the shortcomings of conventional treatments.8 For instance, advances in less bellicose surgical techniques, such as the Rezum Water Vapor Therapy together with the UroLift System, have drawn a lot of interest since they relieve symptoms more effectively than standard surgery while requiring less downtime and having fewer adverse effects.9 Scientists are also looking into new drugs, such as selective androgen receptor modulators and novel enzyme inhibitors. These drugs have shown promise in clinical trials for shrinking the prostate, making symptoms better, and maybe even lowering the risk of side effects.10 Personalized medicine is a fascinating field that is transforming the management of BPH by customizing medicines to each patient’s unique profile using genetic, genomic, and phenotypic data.11 Studies suggest that personalized treatment plans can significantly enhance efficacy and minimize side effects, leading to better patient outcomes.12 For example, genetic markers associated with BPH progression can help predict which patients will benefit most from specific treatments.13 Similarly, holistic and integrative approaches are also gaining traction in the existing pieces of literature as complementary strategies in BPH management.14 For instance, studies have shown that lifestyle modifications such as weight management, dietary adjustments, and physical activity positively impact BPH symptoms.15 Additionally, researchers are investigating complementary therapies like phytotherapy, acupuncture, and other alternative treatments for their potential benefits in symptom management and overall health improvement.16 Again, managing complications and comorbidities associated with BPH is crucial, as these can exacerbate symptoms and complicate treatment. Research stresses the importance of addressing conditions like bladder dysfunction and urinary tract infections concurrently with BPH to optimize patient care.17 Specifically, patient-centered care remains a cornerstone of effective BPH management.18 Generally, emerging research emphasizes the value of involving patients in their treatment decisions, providing education on their condition, and maintaining open communication between healthcare providers and patients. Hence, the tools for assessing patient quality of life and satisfaction with treatment are essential for guiding management decisions and improving outcomes.\n\nFuture directions in BPH care are being shaped by ongoing research and technology improvements.19 To improve diagnosis, therapy, and patient monitoring, emerging research focuses on novel biomarkers, creative therapeutic modalities, and the incorporation of digital health technology.20 Unintentionally, the body of current research on benign prostatic hyperplasia management reveals a dynamic and changing field.21–23 Therefore, going beyond conventional medications, new research emphasizes the potential for personalized medicine, innovative therapeutics, and holistic ways to deliver more efficient and patient-centered care.24 This chapter suggests that BPH management requires a multimodal strategy to enhance patient outcomes and quality of life.\n\n\n2. Understanding benign prostatic hyperplasia\n\nThe non-cancerous widening of the prostate gland, which typically affects elderly men, is the hallmark of BPH. The prostate’s stromal and epithelial cell proliferation is a pathology of benign prostate hyperplasia (BPH), resulting in the formation of big, characteristic nodules in the vicinity of the urethra.25 The symptoms of BPH are brought on by these nodules compressing the urethral canal, which can restrict the urethra partially or completely.26 Hormonal changes that cause the prostate to grow, such as elevated levels of estrogen and DHT,27,28 inflammation, and genetic predisposition all play important roles in the development of BPH.29 Epidemiological studies reveal that BPH is a common condition in aging men to distress approximately 50% of those between the ages of 51 and 60 and up to 90% of those over 80.30 Because of its increasing prevalence and severity with age, BPH is a serious health concern that greatly affects the quality of life and should be discerned from benign prostatic enlargement (BPE) and lacunculation (LUTS), the latter of which frequently necessitates medical consultation.30 The IPSS (International Prostate Symptom Score) or the AUA (American Urological Association) symptom score is commonly used to measure the symptoms of LUTS, which include urge, frequency, nocturia, urgency, and dribbling after voiding. In older men, urodynamic changes such as benign prostatic hindrance and overactive or hypoactive detrusor muscles are the main causes of LUTS. The prevalence of BPH, BPE, and LUTS increases with age, with moderate to severe LUTS affecting roughly 20% of men in their 50s, 30% in their 60s, and 40% in their 80s.31 In Austria, it is estimated that approximately 350,000 men over 40 currently experience moderate to severe LUTS, a number expected to rise to around 500,000 in the next two decades due to demographic changes.1,2 Despite its high prevalence and socioeconomic impact, the complete pathophysiology of BPH remains poorly understood, particularly why some men develop significantly larger prostates than others.32 Visibly, the mechanisms leading to LUTS are complex, involving various urodynamic patterns, changes in the bladder and urothelium, anticholinergic receptor status, and pelvic ischemia, highlighting the need for further research to develop effective treatments.1,33\n\nAlthough BPH can manifest clinically in a variety of ways, lower urinary tract symptoms (LUTS) are generally linked to it.2 LUTS are classified into three main categories by the International Continence Society (ICS): storage symptoms (affecting the bladder’s ability to store urine), voiding symptoms (difficulties in the bladder emptying process), and post-micturition symptoms (leakage after urination).34\n\nThese symptoms shown in Table 1 can significantly impact a person’s quality of life, and addressing them with medical consultation is essential for proper management and relief.\n\n\n\na. Storage symptoms and clinical manifestations\n\n\n\nb. Voiding symptoms and clinical manifestations\n\n\n\nc. Post-micturition symptoms and clinical manifestations\n\nSeveral risk factors have been identified for the development of BPH.35 These risk factors are outlined in Figure 1.\n\nThe main risk factor for BPH is age, as men age, and as a result, the frequency and intensity of symptoms both rise significantly.36 Hormonal factors further exacerbate this, as androgens such as dihydrotestosterone (DHT) are critical for prostatic growth.29 A further factor contributing to the illness may be an imbalance in the levels of estrogen and testosterone. Another factor is genetics; a family history of BPH raises the risk, suggesting a genetic predisposition; certain genetic markers have been shown to enhance the risk of developing BPH.37 Furthermore, there is a considerable correlation between insulin resistance, obesity, hypertension, dyslipidemia, and other elements of metabolic syndrome and BPH, indicating the impact of general metabolic health on prostate enlargement.18 Moreover, lifestyle variables can have a major impact on the risk of BPH.38 A higher incidence of BPH is associated with diets high in fat and red meat and low in fruits and vegetables; nevertheless, regular physical activity seems to offer a preventive effect. Studies reveal that African-American males are more prone than Caucasian men to acquire and have more severe types of BPH. Ethnicity also affects the prevalence and severity of BPH.39 Comprehending the intricate relationships among these risk factors is crucial for healthcare practitioners to formulate efficacious management approaches customized to specific patient profiles, hence enhancing patient outcomes via individualized interventions.40 In particular, healthcare professionals can better serve their patients’ needs by addressing the various pathophysiological causes, clinical manifestations, and demographic factors linked to BPH.2\n\n\n3. Standard management approaches\n\nWhen treating BPH, conservative methods are usually used first, then depending on the patient’s preferences and the severity of the symptoms, more invasive techniques are eventually tried. Surgical operations, minimally invasive procedures, and pharmaceutical therapy are examples of conventional therapeutic methods that are intended to improve quality of life and relieve symptoms.41\n\nAs seen in Table 2, pharmacological treatments are frequently the initial line of treatment for BPH.42 These treatments can be broadly divided into two primary categories.\n\nIn pharmacological treatment, combination therapy with both α-blockers and 5-α-reductase inhibitors can be more effective than either treatment alone, especially in men with moderate to severe symptoms.11\n\nIn the current treatment regimen for BPH, patients who do not respond to pharmacological treatments or have severe symptoms sorts for surgical interventions.43 The most common surgical options are presented in Table 3.\n\nDespite the efficacy of conventional treatment options, several limitations exist ( Table 4). These limitations highlight the importance of exploring new and innovative treatment strategies beyond the norm for BPH that can offer better outcomes with fewer adverse effects.11,42,43\n\n\n4. Emerging therapeutic strategies for BPH\n\nThe goal of recent developments in BPH medication is to reduce adverse effects and increase efficacy. As shown in Table 5, several new agents are being studied or were just presented.44,45\n\nMinimally invasive techniques are becoming increasingly popular due to their effectiveness and reduced recovery times compared to traditional surgery.46,47 These procedures aim to relieve obstruction while minimizing tissue damage (see Table 6).\n\nTechnological advancements are continuously evolving the landscape of BPH treatment, providing less invasive, more effective options with faster recovery times and fewer complications46,48 (see Table 7).\n\n\n\na. UroLift System\n\n\n\nb. Rezum Water Vapor Therapy\n\n\n\nc. PAE (Prostate Artery Embolization)\n\nThese emerging therapeutic strategies represent significant advancements in the management of BPH. By offering effective symptom relief with fewer side effects and faster recovery times, these innovations provide patients with more options to tailor their treatment to their specific needs and preferences.\n\n\n5. Personalized medicine in BPH\n\nGenetic and molecular profiling are revolutionizing the approach to managing BPH by providing deeper insights into the disease’s underlying mechanisms and individual patient variability. In terms of genetic profiling, researchers have identified several genetic markers associated with an increased risk of BPH. For instance, variations in genes interrelated to androgen metabolism, inflammation, and growth factor signaling have been allied to the development and progression of BPH.25 Similarly, understanding genetic differences in drug metabolism and response can help predict which patients are prospective to benefit from specific medications. For case, dissimilarities in genes encoding enzymes fingered in drug metabolism can distress the efficacy and safety of α-blockers and 5-α-reductase inhibitors.42 Additionally, molecular profiling can identify biomarkers that predict disease progression and response to treatment. For instance, levels of prostate-specific antigen (PSA) and other novel biomarkers can provide information about the severity of BPH and the likelihood of progression.49 Furthermore, analysis of molecular pathways involved in BPH, such as those regulating cell proliferation, apoptosis, and inflammation, can identify potential therapeutic targets and guide the development of targeted therapies.50,51\n\nPersonalized medicine encompasses tailoring treatment tactics to the individual characteristics of each patient, enhancing the effectiveness and minimizing adverse effects.47,52\n\n5.2.1 Patient characteristics\n\nImaging studies such as ultrasound and MRI can provide detailed information about prostate size, shape, and the presence of specific anatomical features (e.g., median lobe enlargement) that may influence treatment choices.53 Also, the IPSS and quality of life assessments help gauge the severity of symptoms and their impact on daily activities, guiding the intensity and type of treatment.30 Overall, a comprehensive evaluation of the patient’s overall health, including the presence of comorbid conditions (e.g., cardiovascular disease, diabetes) and current medications, ensures that treatment plans are safe and effective.52,54\n\n5.2.2 Tailored therapies\n\nBased on genetic and molecular profiling, healthcare providers can choose the most suitable medications and adjust dosages to optimize efficacy and reduce side effects.25 For instance, patients with genetic variations affecting drug metabolism may require alternative dosing strategies. Similarly, customized surgical plans take into account the patient’s prostate anatomy and inclusive health.53 For example, minimally invasive techniques like the UroLift or Rezum might be preferred for patients with certain anatomical profiles or those who are at higher risk for surgical complications.47\n\n\n6. Holistic and integrative management\n\nLifestyle modifications play a decisive role in the holistic management of BPH by potentially reducing symptoms and slowing disease progression55 (see Table 8).\n\nDiet and nutrition are critical components of holistic BPH management, with certain dietary patterns and nutrients associated with reduced risk and symptom relief56,57 (see Table 9).\n\nComplementary and alternative therapies can offer additional relief for BPH symptoms and improve overall well-being58 (see Table 10).\n\nIntegrating these holistic and alternative approaches with conventional medical treatments can provide a comprehensive management strategy for BPH, addressing not only the physical symptoms but also enhancing overall health and quality of life.\n\n\n7. Addressing complications and comorbidities\n\n7.1.1 Bladder dysfunction\n\nProper assessment of bladder dysfunction in BPH patients is critical. Urodynamic studies can diagnose the type and extent of bladder issues, such as detrusor overactivity or impaired contractility. Management involves behavioral therapies like bladder training and pelvic floor muscle exercises, which can improve bladder control. Medications, including antimuscarinics or beta-3 agonists, may be prescribed to manage overactive bladder symptoms, reducing urgency, frequency, and incontinence episodes. For severe bladder dysfunction, surgical interventions such as bladder neck incision or augmentation cystoplasty might be necessary.59,60\n\n7.1.2 Urinary tract infections (UTIs)\n\nAccording to Fisher et al.,60 preventing UTIs involves maintaining good hydration and proper hygiene. In recurrent cases, prophylactic antibiotics may be used as treatment requires appropriate antibiotic therapy based on culture and sensitivity results, with long-term low-dose antibiotics considered for recurrent UTIs.61 Also, if chronic prostatitis is suspected, longer courses of antibiotics targeting prostate tissue may be necessary.62\n\n7.2.3 Chronic kidney disease (CKD)\n\nRegular nursing of renal function through blood tests (e.g., estimated glomerular filtration rate and serum creatinine) and urine analysis is essential.63 Managing CKD involves addressing the underlying cause of urinary obstruction and ensuring proper urinary drainage to prevent further kidney damage. Referral to a nephrologist for comprehensive management of CKD and associated conditions may be required.64\n\n7.2.1 Reassessment and diagnosis\n\nFor treatment-resistant BPH, a detailed evaluation of the patient’s symptoms, imaging studies (e.g., transrectal ultrasound, MRI), and urodynamic testing can help understand the resistance to treatment. Identifying contributing factors, such as bladder stones, strictures, or neurogenic bladder, is essential for targeted management.53\n\n7.2.2 Advanced pharmacotherapy\n\nCombination therapy with α-blockers and 5-α-reductase inhibitors may maximize symptom relief. Therefore, exploring alternative medications, such as PDE5 inhibitors (e.g., tadalafil), or off-label uses like botulinum toxin injections into the prostate or bladder, can also be considered.65\n\n7.2.3 Minimally invasive procedures\n\nProstatic Artery Embolization (PAE) is a non-surgical option that reduces prostate volume by decreasing its blood supply. Consequently, Convective Water Vapor Therapy (Rezum) uses thermal energy to ablate excess prostate tissue, effective in treatment-resistant cases.66\n\n7.2.4 Surgical interventions\n\nHolmium Laser Enucleation of the Prostate (HoLEP) is effective for larger prostates and provides long-term symptom relief. Hence, Open or robot-assisted simple prostatectomy is considered for very large prostates where minimally invasive techniques may not be effective.67\n\n7.3.1 Regular follow-up visits\n\nInitially, frequent visits (e.g., every 3-6 months) post-treatment are necessary to monitor recovery and response, gradually reducing to annual check-ups if stable. Therefore, a comprehensive evaluation using the IPSS, physical examination, and PSA testing if indicated is essential.34\n\n7.3.2 Monitoring for complications\n\nImmediately post-treatment, monitoring for signs of bleeding, infection, and urinary retention is critical, ensuring proper catheter care if used. So, long-term hitches, such as bladder neck contractures, urethral strictures, or recurrent BPH, should also be watched for.68\n\n7.3.3 Lifestyle and health maintenance\n\nEncouraging healthy habits like weight management, regular physical activity, and a balanced diet is vital. Hence, patient education on recognizing symptoms of complications or recurrence and advising on when to seek medical attention is also important.\n\n7.3.4 Adjunct therapies and support\n\nReferral to a physical therapist for pelvic floor exercises can improve urinary control and reduce symptoms. Hence, addressing the psychological impact of BPH and its treatment through counseling or support groups is also beneficial.\n\n7.3.4 Coordinated care\n\nA multidisciplinary approach involving urologists, primary care physicians, physical therapists, and other specialists as needed ensures comprehensive care. Therefore, effective patient-centered communication keeps patients well-informed about their condition, treatment options, and the importance of adherence to follow-up schedules.\n\nGenerally, addressing complications and comorbidities in BPH requires a multifaceted approach combining careful monitoring, advanced treatment strategies, and holistic patient care. These elements, if properly integrated can help healthcare providers enhance patient outcomes and quality of life.\n\n\n8. Future perspectives and research directions\n\nThe future of BPH management is poised to be revolutionized by ongoing research and technological advancements, including gene therapy targeting genetic pathways, identification of new molecular biomarkers, development of novel drug classes, and innovative drug delivery systems.19 For example, minimally invasive techniques are expected to improve with next-generation devices and the integration of robotics and AI, while regenerative medicine, such as stem cell therapy, holds the potential for restoring healthy prostate function.68 These advancements will facilitate personalized treatment plans and optimized drug regimens based on genetic profiling, proactive screening, and preventive strategies. Furthermore, enhanced patient outcomes will be driven by continued improvements in minimally invasive options and comprehensive care models. It is expected that future trends will emphasize prevention through lifestyle interventions and public health initiatives, technology-driven care with telemedicine and AI, holistic and integrative health models, and patient-centered care. Therefore, adapting clinical guidelines and having flexible insurance coverage will be essential to ensure that medical professionals can provide the most recent evidence-based procedures for BPH management beyond the norm.\n\n\n9. Conclusion\n\nEffective management of Benign Prostatic Hyperplasia (BPH) necessitates a multifaceted approach that encompasses traditional treatments, emerging therapies, and holistic care. Intuitively, understanding BPH’s pathophysiology, symptoms, and risk factors is essential while addressing complications, and adopting personalized medicine ensures tailored treatment plans. Practically, combining conventional pharmacological and surgical options with novel minimally invasive techniques and integrative approaches such as lifestyle modifications and complementary therapies enhances patient outcomes. Also, emphasizing patient-centered care through education, involvement in decision-making, and effective communication fosters better adherence and satisfaction. In general, continued study and technical developments will change the way BPH is managed, eventually enhancing patients’ quality of life.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nAcknowledgments\n\nWe are grateful for the administrative support offered to us during this study by the Department of Biochemistry, College of Science, Evangel University, Akaeze, Ebonyi State, Nigeria.\n\n\nReferences\n\nNg M, Leslie SW, Baradhi KM: Benign prostatic hyperplasia. StatPearls. StatPearls Publishing; 2024. 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Hum. Genet. 2002; 111(2): 166–171. Publisher Full Text\n\nSoler R, Andersson KE, Chancellor MB, et al.: Future direction in pharmacotherapy for non-neurogenic male lower urinary tract symptoms. Eur. Urol. 2013; 64(4): 610–621. PubMed Abstract \| Publisher Full Text\n\nPrzydacz M, Dudek P, Golabek T, et al.: Relationship between Lower Urinary Tract Symptoms and Treatment-Related Behavior in an Eastern European Country: Findings from the LUTS POLAND Study. Int. J. Environ. Res. Public Health. 2021; 18(2): 785. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPatel ND, Parsons JK: Epidemiology and etiology of benign prostatic hyperplasia and bladder outlet obstruction. Indian Journal of Urology. 2014; 30(2): 170–176. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBarry MJ, Fowler FJ Jr, Bin L, et al.: The natural history of patients with benign prostatic hyperplasia as diagnosed by North American urologists. J. Urol. 1997; 157(1): 10–15. Publisher Full Text\n\nSanda MG, Beaty TH, Stutzman RE, et al.: Genetic susceptibility of benign prostatic hyperplasia. J. Urol. 1994; 152(1): 115–119. Publisher Full Text\n\nRohrmann S, Smit E, Giovannucci E, et al.: Association between markers of the metabolic syndrome and lower urinary tract symptoms in the Third National Health and Nutrition Examination Survey (NHANES III). Int. J. Obes. 2005; 29(3): 310–316. PubMed Abstract \| Publisher Full Text\n\nPlatz EA, Kawachi I, Rimm EB, et al.: Race, ethnicity and benign prostatic hyperplasia in the health professionals follow-up study. J. Urol. 2000; 163(2): 490–495. PubMed Abstract \| Publisher Full Text\n\nTantiwong A, Nuanyong C, Vanprapar N, et al.: Benign prostatic hyperplasia in elderly Thai men in an urban community: The prevalence, natural history, and health-related behavior. J. Med. Assoc. Thail. 2002; 85(3): 356–360. PubMed Abstract\n\nO’Quin C, White KL, Campbell JR, et al.: Pharmacological approaches in managing symptomatic relief of benign prostatic hyperplasia: A comprehensive review. Cureus. 2023; 15(12): e51314. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChristidis D, McGrath S, Perera M, et al.: Minimally invasive surgical therapies for benign prostatic hypertrophy: The rise in minimally invasive surgical therapies. Prostate Int. 2017; 5(2): 41–46. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBhasin S, Jasuja R: Selective androgen receptor modulators as function-promoting therapies. Curr. Opin. Clin. Nutr. Metab. Care. 2009; 12(3): 232–240. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHuang SA, Lie JD: Phosphodiesterase-5 (PDE5) inhibitors in the management of erectile dysfunction. Pharm. Ther. 2013; 38(7): 407–419. PubMed Abstract\n\nThomas D, Chughtai B, Kini M, et al.: Emerging drugs for the treatment of benign prostatic hyperplasia. Expert Opin. Emerg. Drugs. 2017; 22(3): 201–212. Publisher Full Text\n\nPascoe J, Fontaine C, Hashim H: Modern advancements in minimally invasive surgical treatments for benign prostatic obstruction. Ther. Adv. Urol. 2021; 13: 17562872211030832. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGuelce D, Thomas D, Elterman D, et al.: Recent advances in managing benign prostatic hyperplasia: The Rezūm system. F1000Res. 2018; 7: 1916. F1000 Faculty Rev-1916. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCannon GW, Getzenberg RH: Biomarkers for benign prostatic hyperplasia progression. Curr. Urol. Rep. 2008; 9(4): 279–283. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHata J, Harigane Y, Matsuoka K, et al.: Mechanism of androgen-independent stromal proliferation in benign prostatic hyperplasia. Int. J. Mol. Sci. 2023; 24(14): 11634. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRicke WA, Macoska JA, Cunha GR: Developmental, cellular, and molecular biology of benign prostatic hyperplasia. Differentiation. 2011; 82(4-5): 165–167. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWasserman NF, Spilseth B, Golzarian J, et al.: Use of MRI for lobar classification of benign prostatic hyperplasia: Potential phenotypic biomarkers for research on treatment strategies. AJR Am. J. Roentgenol. 2015; 205(3): 564–571. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAmerican Diabetes Association Professional Practice Committee: Comprehensive medical evaluation and assessment of comorbidities: Standards of medical care in diabetes-2022. Diabetes Care. 2022; 45(Suppl 1): S46–S59. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMathur S, Sutton J: Personalized medicine could transform healthcare. Biomed. Rep. 2017; 7(1): 3–5. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNagakura Y, Hayashi M, Kajioka S: Lifestyle habits to prevent the development of benign prostatic hyperplasia: Analysis of Japanese nationwide datasets. Prostate Int. 2022; 10(4): 200–206. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEspinosa G: Nutrition and benign prostatic hyperplasia. Curr. Opin. Urol. 2013; 23(1): 38–41. Publisher Full Text\n\nRusso GI, Broggi G, Cocci A, et al.: Relationship between dietary patterns with benign prostatic hyperplasia and erectile dysfunction: A collaborative review. Nutrients. 2021; 13(11): 4148. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCsikós E, Horváth A, Ács K, et al.: Treatment of benign prostatic hyperplasia by natural drugs. Molecules. 2021; 26(23): 7141. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLeslie SW, Tadi P, Tayyeb M: Neurogenic bladder and neurogenic lower urinary tract dysfunction. In StatPearls. Treasure Island (FL): StatPearls Publishing; 2023. Reference Source\n\nYoshimura N, Chancellor MB: Neurophysiology of lower urinary tract function and dysfunction. Rev. Urol. 2003; 5(Suppl 8): S3–S10.\n\nFisher H, Oluboyede Y, Chadwick T, et al.: Continuous low-dose antibiotic prophylaxis for adults with repeated urinary tract infections (AnTIC): a randomized, open-label trial. Lancet Infect. Dis. 2018; 18(9): 957–968. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAggarwal N, Leslie SW, Lotfollahzadeh S: Recurrent urinary tract infections. StatPearls. Treasure Island (FL): StatPearls Publishing; 2024. Reference Source\n\nOkoro RN, Farate VT: The use of nephrotoxic drugs in patients with chronic kidney disease. Int. J. Clin. Pharm. 2019; 41(3): 767–775. Publisher Full Text\n\nBiljak VR, Honović L, Matica J, et al.: The role of laboratory testing in detection and classification of chronic kidney disease: national recommendations. Biochem. Med (Zagreb). 2017; 27(1): 153–176. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYan H, Zong H, Cui Y, et al.: The efficacy of PDE5 inhibitors alone or in combination with α-blockers for the treatment of erectile dysfunction and lower urinary tract symptoms due to benign prostatic hyperplasia: a systematic review and meta-analysis. J. Sex. Med. 2014; 11(6): 1539–1545. Publisher Full Text\n\nOntario Health (Quality): Prostatic artery embolization for benign prostatic hyperplasia: A health technology assessment. Ont. Health Technol. Assess. Ser. 2021; 21(6): 1–139.\n\nVan der Jeugt J, Umari P, Mottaran A, et al.: Holmium laser enucleation of the prostate vs robot-assisted simple prostatectomy for lower urinary tract symptoms in patients with extremely large prostates ≥200 cc: A comparative analysis. J. Endourol. 2023; 37(8): 895–902. PubMed Abstract \| Publisher Full Text\n\nAgwu NP, Umar AM, Oyibo UE: Review article: Urethral catheters and catheterization techniques. Niger. J. Med. 2022; 31: 497–508. Publisher Full Text\n\nReddy K, Gharde P, Tayade H, et al.: Advancements in robotic surgery: A comprehensive overview of current utilizations and upcoming frontiers. Cureus. 2023; 15(12): e50415. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "354558", "date": "17 Jan 2025", "name": "Carlotta Nedbal", "expertise": [ "Reviewer Expertise Urology" ], "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\nManagement of BPH is one of the most common interventions in urology, this review discusses the newest and uncommon treatment strategies.\n\nI have some comments: - Please use abbreviations when needed (i.e., use BPH instead of benign prostatic hyperplasia) in all text. Only first clarification is needed. Please check that all abbreviations are declared when first mentioned.\n- Introduction \"the Rezum Water Vapor Therapy together with the UroLift System, have drawn a lot of interest since they relieve symptoms more effectively than standard surgery\". This is incorrect, studies have proven comparable symptoms relief outcomes and still need further confirmation.\n- \"lacunculation\": What does this term mean...please clarify?\n- Standard management approaches: the paragraph is very limited providing only summarised list of some of the current therapies, both pharmacological and surgical. Please expand it.\n- Emerging therapeutic strategies: again, the paragraph is very limited and does not provide any constructive and new information. New techniques are only partially included and just mentioned, with no informative content.\n- Addressing complications and comorbidities: this paragraph is redundant and little informative.\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? Yes", "responses": [] } ]	1	https://f1000research.com/articles/13-1555
https://f1000research.com/articles/13-1554/v1	23 Dec 24	{ "type": "Research Article", "title": "Assessment to determine the accuracy of Chaillet and Demirjian method of dental age estimation using Orthopantomographs, Eastern Province, Saudi Arabia", "authors": [ "Faraz Mohammed", "Arishiya Thapasum Fairozekhan", "Intisar Ahmad Siddiqui", "Saud AlMoumen", "Turki Ali AlShehri", "Maria Hassan AlRssasi", "Abdullah Amer AlJami", "Doha Mohsen AlRamadan", "Ashwin C Shetty", "Majed Mohammed Alfarea", "Omar Suhaym", "Nasser S AlShahrani", "Faraz Mohammed", "Arishiya Thapasum Fairozekhan", "Intisar Ahmad Siddiqui", "Saud AlMoumen", "Turki Ali AlShehri", "Maria Hassan AlRssasi", "Abdullah Amer AlJami", "Doha Mohsen AlRamadan", "Majed Mohammed Alfarea", "Omar Suhaym", "Nasser S AlShahrani" ], "abstract": "Background The approach to estimating mandibular age has been widely used, although it has shown age estimation disparities in diverse populations, including Indians.\n\nObjective Our goal was to test the Demirjian 8-teeth approach utilizing orthopantomogram to estimate age reliably after incorporating the third molar. We also used Chaillet and Demirjian’s regression equations to estimate age estimation accuracy in the Eastern Province of Saudi Arabia.\n\nMethods The retrospective study included 196 people. Individuals were 8–24 years old. The left mandibular teeth were staged using an orthopantomographs utilizing the Demirjian 8-teeth approach. The Chaillet and Demirjian’s regression models determined dental age, and statistical methods compared dental age to chronological age.\n\nResults The gender breakdown was 49.5% male and 50.5% female. Gender did not significantly affect chronological mean age or estimated mean age (13.39±3.77 vs. 13.10±3.51, p = 0.583; 11.75±2.92 vs. 11.58±2.70, p = 0.674). Statistically substantial differences in chronological and estimated mean ages between male and female individuals (p<0.001). Out of the total participants, 31.6% had a difference in age of ±1 year, 30.6% had a difference of 1-2 years, and 37.8% had a difference of more than 2 years. Compared to chronological and estimated ages, these variations in age between males and females were not statistically significant (p=0.557).\n\nConclusions The findings of this study support the use of Demirjian’s 8-teeth approach in the Saudi population residing in the Eastern Province of the Kingdom of Saudi Arabia, employing Chaillet and Demirjian’s regression equations.", "keywords": [ "forensic science", "age estimation", "Demirjian method", "dental age", "chronological age" ], "content": "Introduction\n\nThe current emphasis in forensic science and medico-legal studies revolves around the crucial significance of forensic age estimation.1,2 Forensic dental age estimation is well recognized as a very precise and established technique employed in the determination of the chronological age of individuals involved in judicial or legal processes.1 The determination of age in children and teenagers is again a crucial aspect in addressing a wide range of legal investigations. In the present context, the majority of age estimation methods are invasive in nature, necessitating extended processing durations, utilization of costly equipment, and the expertise of a skilled pathologist to infer the age of an individual.3 The growth and eruption of dentition are significant factors in determining dental age and are widely regarded as the most reliable physiological non-invasive indicators for measuring chronological age in children and juveniles.4–6 The process of tooth mineralization is comparatively less influenced by nutritional intake, endocrine status, and local conditions when compared to the process of bone mineralization.\n\nDemirjian et al. proposed a methodology for age estimation that relied on the analysis of seven mandibular teeth. The methodology has been extensively utilized; however, it has demonstrated discrepancies in age estimations among a limited number of populations. The observed variations in tooth eruption timing among different ethnicities worldwide have been attributed to racial differences. The assessment of age based on tooth development is subject to the influence of two primary elements: genetic variability and environmental factors.4 Multiple countries have made various efforts to minimize the influence of variability in dental age estimation. This is achieved by establishing genetically similar populations and developing standardized tables that are specifically applicable to these genetically similar individuals.2,7,8 Consequently, it is crucial to derive dental age estimates from the population in which they will be utilized. Additionally, one limitation of the first Demirjian et al. approach was its exclusion of the third molar due to its potential for congenital absence and the significant variability in its development.9 Demirjian et al. conducted a study in which they categorized the progression of tooth growth into eight distinct stages and subsequently devised a method for estimating age based on these stages.\n\nThus, the primary objectives of our study were to validate the Demirjian 8-teeth method using orthopantomogram for age estimation, specifically by incorporating the third molar, and to assess the accuracy of age estimation in the Saudi population from the Eastern Province of the Kingdom of Saudi Arabia using Chaillet and Demirjian’s regression formulae.\n\n\nMaterials and methods\n\nThe present study employed a materials and methods approach to investigate the research question.\n\nThe present investigation was undertaken at the Dental College Hospital of Imam Abdulrahman Bin Faisal University (IAUDent) in Dammam, Kingdom of Saudi Arabia. The present retrospective investigation was conducted over a duration of one year, utilizing archived orthopantomograms (OPGs) that were originally collected for diagnostic and therapy purposes. The age, gender, and date of image acquisition for patients were obtained from the orthopantomography system’s records within the hospital.\n\nThe present study involved a retrospective, cross-sectional design and focused on a specific sample of orthopantomograms (OPGs) obtained from 196 individuals who were of Eastern Province origin in the Kingdom of Saudi Arabia. The subjects chosen for the study fell within the age bracket of 8 to 24 years. The OPGs and associated data were acquired from the database archives of the centrally stored radiology computer software MiPACS at Imam Abdulrahman Bin Faisal University’s Dental College Hospital (IAUDent), Dammam, Kingdom of Saudi Arabia. This data collection occurred between October 2023 and December 2023, following a waiver of ethical approval granted by the institutional review board of Imam Abdulrahman Bin Faisal University on October 10, 2023.\n\nThe study comprised patients who had a complete set of maxillary and mandibular right and left quadrant teeth, with no impacted, embedded, or trans-positioned teeth, and whose Orthopantomograms (OPGs) were free of any position errors or artifacts. Subjects who exhibited any pathological conditions, had previously undergone tooth extraction or restorative procedures, were currently undergoing orthodontic treatment, had congenital or developmental anomalies of the jaws and teeth, or had a history of craniofacial trauma, jaw disorders, or systemic illnesses were excluded from the study. The exclusion criteria were verified by cross-referencing the subjects’ reception files. Table 1 displays the gender-based distribution of subjects. The age, date of birth, gender, and nationality of the subject were extracted from the patients’ case file records.\n\nThe verification of the Date of Birth (DOB) was conducted by cross-referencing the information with the official birth document that was recorded during the registration process at the IAUDent Hospital. The Date of Exposure of Radiograph (DOR) was obtained from the MiPACS system. The calculation of chronological age in decimal years involved subtracting the date of birth (DOB) from the date of reference (DOR) using a straightforward formula in Microsoft Excel: (DOR-DOB/365.25).\n\nThe dental age (DA) can be determined with the application of Demirjian’s method10 for calculation.\n\nThe dental structures located on the left side of the mandible, specifically ranging from the mandibular central incisor to the mandibular third molar, were assessed using Demirjian’s modified criteria. This evaluation involved the utilization of 10 distinct stages of tooth development. In cases when a tooth is absent in the mandibular left quadrant, the corresponding tooth on the contralateral side was included as part of the research investigation. The dental age was determined by employing the formula outlined in the method proposed by Chaillet and Demirjian, utilizing their regression equations.10\n\nThe equation for determining the age of males is as follows:\n\nThe equation to estimate the age of females is given by\n\nThe data collected, which included Chronological Age (CA) and Dental Age (DA) computed using the Demirjian technique, were inputted into the Statistical Package for Social Sciences (IBM Product, SPSS Version-25.0, USA). The mean and standard deviation were calculated for the numeric variables representing the chronological and predicted age of the individuals. The statistical analysis employed in this study was the use of a paired sample t-test to compare the differences between chronological age and estimated age for both the whole sample and for individual gender groups. The categorization of the disparities between chronological and estimated age was conducted based on three intervals: ±1 year, ±>1-2 years, and ±>2 years. The chi-square test was utilized to compare the categorized age differences based on combination and gender variables. A significance level of P < 0.05 was deemed to indicate statistical significance.\n\n\nResults\n\nThe study involved the evaluation of 196 panoramic radiographs obtained from a sample of 99 male and 97 female participants, all within the age range of 8 to 24 years. Table 1 illustrates a gender distribution that is evenly split, with 49.5% of individuals identified as men and 50.5% identified as females. The results indicate that there were no significant differences in the chronological mean age and estimated mean age based on formula between male and female participants. Specifically, the mean ages for males and females were 13.39±3.77 and 13.10±3.51, respectively, with a p-value of 0.583. Similarly, the mean ages based on formula were 11.75±2.92 for males and 11.58±2.70 for females, with a p-value of 0.674. However, when considering the overall differences between male and female participants’ chronological mean age and estimated mean age, there was a statistically significant difference (p<0.001), as shown in Table 2. The study found that 31.6% of subjects had an age difference of ±1 year, while 30.6% had an age difference of ±>1-2 years, and 37.8% had an age difference greater than 2 years. Nevertheless, the observed disparities in chronological and estimated ages did not reach statistical significance, as seen in Table 3. The linear regression equation, which was derived from the data collected from both male and female participants, allows for the prediction of chronological age based on estimated age. This equation is represented as Y=3+0.6*X, where X and Y represent the estimated and predicted chronological ages, respectively. These findings are visually presented in Figures 1 and 2. The interobserver agreement for the first examiner (FM) yielded a Kappa value of 0.872, indicating a strong level of agreement. Similarly, the second examiner (ATFK) achieved a Kappa value of 0.838, also indicating a strong level of agreement. The interobserver agreement among examiners regarding FM and ATFK, the value was determined to be 0.812, indicating a substantial correlation. The analysis of means did not yield statistically significant differences, indicating that the current age estimation equation can be applied to estimate the age of individuals within the Saudi population, taking into account gender-specific considerations.\n\n\nDiscussion\n\nThe utilization of tooth structures as a means of estimating age has been a widely employed practice with a long-standing history. Edwin Saunders, in the year 1837, made a significant contribution to the field of age assessment by presenting a pamphlet titled “Teeth A Test of Age” to the English Parliament.11 Subsequently, other approaches for estimating age have been suggested; yet, regrettably, a universally applicable method remains elusive. The absence of this trait can be ascribed to the substantial degree of variance observed among many ethnic groups and populations.12 Demirjian’s method is a widely employed approach for age estimation due to its simplicity, practicality, and ease of comprehension.11 Throughout history, other scholars have also conducted experiments to assess the validity of this formula in diverse communities and have suggested alterations to this methodology. Chaillet-Demirjian incorporated the inclusion of the third molar into the initial formula utilized for evaluating the age of French children, afterwards deriving regression formulas for age assessment.10 A significant alteration implemented in this study involved the incorporation of two supplementary steps in the process of tooth staging. This adjustment was done to facilitate the calculation process and to establish cubic equations that exhibit a high degree of reliability.13 Demirjian’s method is characterized by its well-defined criteria, which effectively eliminates the need for speculative estimation. Consequently, this approach is highly comprehensible and replicable. The methodology employed in this study relies on evaluating the level of calcification in dental structures, namely until the point of root apex closure, while also considering the proportional lengths of the crown and root.\n\nThe Kingdom of Saudi Arabia is currently seeing a growing need for age estimation in both children and adults. This demand is primarily driven by many factors like human trafficking, migration, asylum procedures, child pornography, adoption of children without proper birth certificates, and legal considerations, among others.14 Multiple research studies have been conducted on the Saudi and international population, which have identified a tendency to overestimate age when utilizing Demirjian’s age estimation approach. It is important to acknowledge that the earlier investigations employed Demirjian’s 7-teeth method, while the current study employed the modified 8-teeth method. There exists a limited number of studies that have investigated the precision of the Chaillet-Demirjian age estimation method. The Chaillet-Demirjian’s method demonstrates minimal disparity between predicted age and chronological age when all seven complementary teeth are present, making it a viable technique for age estimation. Moreover, the method proposed by Chaillet and Demirjian has undergone validation using reference data relevant to multiple countries.15 Nevertheless, the efficacy of this approach in estimating age among the Saudi population residing in the Eastern province has not yet been investigated. Consequently, it remains uncertain if a region-specific reference technique is warranted for individuals from the Eastern province of Saudi Arabia. Hence, the primary objective of this study was to validate the regression formulas approach proposed by Chaillet and Demirjian in a sample of individuals from the Eastern province of Saudi Arabia. Additionally, this study aimed to propose the development of region-specific formulae and a method for age estimation in the Eastern province of Saudi Arabia, if deemed necessary.\n\nThe data was obtained from the Dental College Hospital (IAUDent) at Imam Abdulrahman Bin Faisal University in Dammam, Kingdom of Saudi Arabia. This institution is highly regarded in the region and is representative of the entire population of the Eastern province in the Kingdom. The mean difference between the estimated and chronological ages in the entire sample of 196 orthopantomograms was found to be 1.57±0.82 years, indicating a statistically significant difference (p<0.001), as shown in Table 2. In males, the mean difference was found to be 1.63±0.85 years, whereas in females, it was seen to be 1.52±0.81 years. The observed differences were determined statistically nonsignificant.\n\nNumerous research have been undertaken across diverse populations with the regression equations method developed by Chaillet and Demirjian, yielding varying outcomes. Certain researchers observed a notable level of precision within their examined sample, while others documented instances of either overestimating or underestimating dental age in relation to chronological age. Numerous investigations, including the present investigation, have identified a tendency to underestimate estimated age relative to chronological age. In a study conducted by AlOtaibi and AlQahtani in 2023, it was observed that there was an underestimation of -2.03 years in males and -2.35 years in females throughout the Saudi population across all age categories.16 A notable disparity between dental age and chronological age was noticed among the male and female individuals of the Belgian Caucasian community. The combined mean absolute error for both males and females was found to be 1.57 years, with only 31.6% of cases accurately predicted to fall within a range of 1 year of their chronological age. A statistically significant difference was observed, suggesting that Chaillet and Demirjian’s study on the Belgian Caucasian population did not demonstrate a strong association with the population of the Eastern Province in Saudi Arabia. Therefore, it is necessary to utilize population-specific weighted scores when applying the values proposed by Chaillet and Demirjian to this particular population. The variation observed between estimated and chronological ages in various studies, including the present one, can be attributed to the inherent diversity found in ethnicity, culture, sample size, environmental factors, socioeconomic status, nutrition, dietary habits, statistical methods, and subjectivity of the examiner across different populations. The study also revealed that the phenomenon of age underestimation is less prevalent among females compared to males. This discovery suggests that girls reach maturity at an earlier stage compared to males, which aligns with the observed early maturation of skeletal age in females. The observations suggest that girls in Saudi Arabia have a higher level of dental maturity in comparison to boys, which aligns with findings from previous studies.\n\nThe authors want to broaden their research scope by establishing cubic regression formulae that are specific to the population of the Eastern province in the Kingdom of Saudi Arabia. This expansion is motivated by the recognition of growth variances across different populations, which can be attributed to distinct genetic and environmental factors. It is recommended that the maturity scores accurately reflect the specific population under examination. Consequently, it is suggested that the utilization of weighted scores for the Eastern province be pursued as a more favorable approach for evaluating dental age in future research endeavors.\n\n\nConclusions\n\nIn light of the evidence presented, it can be inferred that the findings support the stated hypotheses. The primary objective of this study was to assess the suitability and precision of the Chaillet and Demirjian’s regression equations in predicting the age of individuals within the population of Saudi Arabia’s Eastern province. This study was characterized by two notable aspects: the utilization of digital radiographs for analysis and the incorporation of a larger sample size encompassing individuals aged 8 to 24 years. Based on the findings of this study, it can be inferred that the regression formulae developed by Chaillet and Demirjian may not be sufficiently applicable to the population of the Eastern Province in Saudi Arabia, as the average age difference observed was approximately two years. Nevertheless, for the purpose of achieving a more accurate estimation of age, it would be more advantageous to employ a gender-specific approach in the implementation of this strategy. Hence, it is imperative to establish a distinct set of equations and methodologies for age estimate among individuals of Saudi Arabian Eastern province origin, rather than relying on the Chaillet and Demirjian’s reference database.\n\n\nEthical approval\n\nThis data collection occurred between October 2023 and December 2023, following a waiver of ethical approval granted by the institutional review board of Imam Abdulrahman Bin Faisal University on October 10, 2023.\n\nThe ethical approval was waived for this study because it was retrospective, and no direct involvement or interaction with human participants occurred. The study utilized previously collected data, which ensured that no additional risks were posed to individuals.\n\n\nConsent\n\nPatients’ written and informed consent was taken when they received treatment in the hospital. This study used X-rays retrospectively. Hence, the statement was not mentioned in the manuscript.\n\n\nData availability statement\n\nFigshare: Assessment to determine the accuracy of Chaillet and Demirjian method of dental age estimation using Orthopantomographs, Eastern Province, Saudi Arabia. DOI: https://doi.org/10.6084/m9.figshare.27129798.v1.17\n\nThe project contains the following underlying data:\n\n• Data.xlsx\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).", "appendix": "References\n\nKutesa AM, Rwenyonyi CM, Mwesigwa CL, et al.: Dental age estimation using radiographic assessment of third molar eruption among 10–20- year-old Ugandan population. J. Forensic Dent. Sci. 2019 Jan; 11(1): 16–21. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOlze A, Niekerk P, Ishikawa T, et al.: Comparative study on the effect of ethnicity on wisdom tooth eruption. Int. J. Legal Med. 2007; 121: 445–448. PubMed Abstract \| Publisher Full Text\n\nKumar VJ, Gopal KS: Reliability of age estimation using Demirjian’s 8 teeth method and India specific formula. J. Forensic Dent. Sci. 2011; 3: 19–22. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAlshihri AM, Kruger E, Tennant M: Western Saudi adolescent age estimation utilising third molar development. Eur. J. Dent. 2014 Jul; 8(3): 296–301. PubMed Abstract \| Publisher Full Text\n\nSmith BH: Dental development and the evolution of life history in Hominidae. Am. J. Phys. Anthropol. 1991; 86(157): 74.\n\nNelson SJ, Ash M: Wheeler’s Dental Anatomy, Physiology, and Occlusion. 9th ed.Louis, Mo: Saunders Elsevier; 2010.\n\nHassanali J: The third permanent molar eruption in Kenyan Africans and Asians. Ann. Hum. Biol. 1985; 12: 517–523. PubMed Abstract \| Publisher Full Text\n\nBrkić H, Vodanović M, Dumancić J, et al.: The chronology of third molar eruption in the Croatian population. Coll. Antropol. 2011; 35: 353–357. PubMed Abstract\n\nAcharya AB: Age estimation in Indians using Demirjian’s 8-teeth method. J. Forensic Sci. 2011; 56: 124–127. PubMed Abstract \| Publisher Full Text\n\nChaillet N, Willems G, Demirjian A: Dental Maturity in Belgian Children using Demirjian’s method and Polynomial Functions: New Standard Curves for Forensic and Clinical use. J. Forensic Odontostomatol. December 2004; 22(2): 18–27. PubMed Abstract\n\nStavrianos C, Mastagas D, Stavrianou I, et al.: Dental age estimation of adults: A review of methods and principles. Res. J. Med. Sci. 2008; 2: 258–268.\n\nKoshy S, Tandon S: Dental age assessment: The applicability of Demirjian’s method in south Indian children. Forensic Sci. Int. 1998; 94: 73–85. PubMed Abstract \| Publisher Full Text\n\nPriya E: Applicability of Willem’s Method of Dental Age Assessment in 14 Years Threshold Children in South India - A Pilot Study. J. Forensic Res. S4. 4–002. 1000S4-002. Publisher Full Text\n\nAlqerban A, Alrashed M, Alaskar Z, et al.: Age estimation based on Willems method versus country specific model in Saudi Arabia children and adolescents. BMC Oral Health. 2021 Jul;13; 21(1): 341. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWillems G, Olmen A, Spiessens B, et al.: Dental age estimation in Belgian children: Demirjian’s technique revisited. J. Forensic Sci. 2001; 46(4): 893–895. PubMed Abstract \| Publisher Full Text\n\nAlOtaibi NN, AlQahtani SJ: Performance of different dental age estimation methods on Saudi children. J. Forensic Odontostomatol. 2023; 41(46): 1–27.\n\nShetty AC: Assessment to determine the accuracy of Chaillet and Demirjian method of dental age estimation using Orthopantomographs, Eastern Province, Saudi Arabia. Dataset. figshare. 2024. Publisher Full Text" }	[ { "id": "352324", "date": "06 Jan 2025", "name": "Shaul Hameed Kolarkodi", "expertise": [ "Reviewer Expertise forensic odontology. age estimation", "and oral cancer. premalignant lesions and conditions." ], "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 current research article reflects its originality, best presentation, and relevance for the readers. The author's message is appropriate. Major strengths include comprehensive and rigorous methodology which ensures a thorough analysis, and the authors discussed in detail. The scientific content is high with good English language this paper will contribute to the scientific literature.\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": "364859", "date": "07 Feb 2025", "name": "Dwi Kartika Apriyono", "expertise": [ "Reviewer Expertise forensic odontology" ], "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\nSome things that need to be fixed or explained are: 1. The reasons for choosing the Chaillet and Demirjian's regression formulae have not been stated. 2. it is necessary to add the racial category of the research subjects 3. It is necessary to convey the number of observers and the frequency of calculating dental age. 4. Need to add up-to-date literature\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/13-1554
https://f1000research.com/articles/13-1089/v1	24 Sep 24	{ "type": "Systematic Review", "title": "Preoperative Carbohydrate Loading in Pediatric Surgery: A Scoping Review of Current Evidence", "authors": [ "Yunita Widyastuti", "Djayanti Sari", "Anisa Fadhila Farid", "Amar Rayhan", "Djayanti Sari", "Anisa Fadhila Farid", "Amar Rayhan" ], "abstract": "Abstract*\nIntroduction Preoperative carbohydrate loading (PCL), part of Enhanced Recovery After Surgery (ERAS) protocols, involves giving carbohydrate-rich liquids before surgery instead of traditional fasting. It improves glucose control, reduces insulin resistance, and enhances patient comfort.\n\nMethods This scoping review aims to assess the current evidence on the effects and safety of PCL in pediatric surgery. A multi-database search strategy would be used, with eligibility criteria including recent original English articles on pediatric PCL. Data extraction would focus on PCL type, sample sizes, and perioperative outcomes.\n\nResults The scoping review examined 10 studies on PCL in pediatric surgery, covering various procedures with sample sizes ranging from 18 to 1200 participants. Most studies showed that PCL improved metabolic outcome and reduced postoperative recovery time. However, outcomes like hospital stay length and postoperative complications, such as nausea and vomiting, varied.\n\nConclusions PCL in pediatric surgery may stabilize blood glucose, reduce metabolic risks, and enhance recovery, including anxiety reduction.", "keywords": [ "Preoperative Carbohydrate Loading", "Pediatric Surgery", "Preoperative Fasting", "Perioperative Care" ], "content": "1. Introduction\n\nPreoperative carbohydrate loading (PCL) has emerged as a key component in optimizing surgical outcomes, particularly within the framework of Enhanced Recovery After Surgery (ERAS) protocols. The practice involves administering carbohydrate-rich clear liquids to patients in the hours leading up to surgery, which contrasts with the traditional approach of fasting from midnight. This approach has shown benefits, including improving perioperative glucose control, reducing insulin resistance, and enhancing patient comfort. Additionally, preoperative carbohydrate therapy offers physiological advantages, such as reducing the stress response associated with surgery and contributing to better recovery. While the evidence supporting PCL continues to grow, these studies underline the need for further research to establish standardized protocols and maximize patient outcomes across diverse surgical populations.1,2\n\nPCL has been increasingly supported by recent studies, particularly in the adult population, for its ability to enhance patient comfort and improve clinical outcomes in elective surgeries. Notably, the use of PCL has shown promise in elderly patients, who often face additional risks during surgery due to their age and comorbidities. In these populations, administering carbohydrate-rich liquids 2–4 hours before surgery has been associated with reduced preoperative thirst and hunger, without adversely affecting insulin resistance (IR) or gastric volume. Moreover, PCL has been linked to significant reductions in postoperative IR, inflammation, and the length of hospital stay in patients undergoing major procedures such as colorectal surgery.3–5 In pediatric patients, while research is more limited, similar benefits have been observed, including less nausea and reduced gastric contents with no increased aspiration risk during procedures. Further studies are needed to determine the optimal use of carbohydrate loading in various pediatric age groups and conditions.6\n\n\n2. Methods\n\nThe scoping review main question is “What is the current evidence on the effects and safety of PCL in pediatric surgery?”.\n\nTo conduct a comprehensive scoping review on PCL in pediatric surgery, we used a multi-database approach. The databases to be searched include PubMed (MEDLINE), CINAHL, EMBASE, Cochrane Library, Scopus, and Web of Science. The searchemployed a range of terms such as (“Preoperative Carbohydrate Loading” OR “Preoperative Carbohydrate Supplementation” OR “Preoperative Carbohydrate Administration”) AND (“Pediatric Surgery” OR “Child Surgery” OR “Pediatric Surgical Procedures” OR “Children Surgery”).\n\nRecent primary original english articles examining PCL in pediatric patients were included from scholarly journals. There were no limitation in study design, surgical procedure and study outcome. Exclusion criteria filtered out articles not related to pediatric populations or those focused exclusively on adult populations or unrelated to PCL. The PRISMA flow chart provides a detailed summary of the study selection process, documenting the number of articles that advanced through each stage of screening.\n\nThe selection process will involved screening titles and abstracts to select relevant studies based on inclusion criteria. Full-text articles would be reviewed to confirm relevance and quality. Multiple reviewers ensured accuracy and consistency, resolving discrepancies through discussion or a third reviewer if needed. For each article, data were extracted concerning type of pediatric surgery, sample sizes, type of PCL and control group used, and outcomes measured and results. The outcome results were reported from each article individually, presenting the available information in accordance with the data provided, such as mean, standard deviation (SD), median, interquartile range (IQR), sample size (n), percentage (%), and p-values.\n\nThe authors independently reviewed the methodological quality of the studies using the ‘Risk of bias’ tool, which has undergone modifications and improvements, with an updated version Risk of Bias (RoB) Assessment 2.0 Tools. They evaluated key aspects such as the randomization process, deviations from the intended intervention, missing outcome data, outcome measurement, and selection of reported outcomes. Each study was classified as having low, some concerns, or high risk of bias for each domain. Studies were rated as “low risk” if the information was clear and complete, “high risk” if certain details were missing or suggested a clear risk of bias, and “some concerns” if the data were incomplete.\n\n\n3. Results\n\nThe scoping review identified a total of 10 studies that investigated the effects of PCL in pediatric surgery (Figure 1 and Table 1). The studies included a diverse range of surgical procedures, with most focusing on elective operations such as abdominal, orthopedic, and cardiac surgeries. Sample sizes varied across the studies, ranging from 18 to 1200 participants. Interventions typically involved administering a carbohydrate-rich drink 2-3 hours before surgery, with comparison groups following traditional fasting protocols. The outcomes reported included stomach content, metabolic indicators, incidence of preoperative anxiety and agitation, postoperative complications including nausea and vomiting and length of hospital stay. Additionally, we assessed the risk of bias and found that 4 studies had a low risk, 5 studies had some concerns, and 1 study had a high risk (Figures 2 and 3).\n\n\n\n1. modified Yale Preoperative Anxiety Scale (m-YPAS)\n\n2. gastric residual fluid volume (GRV)\n\n3. gastric antral cross-sectional area (CSA)\n\n4. 20th minute of surgery blood glucose.\n\n\n\n1. m-YPAS 1 hour after surgery (Mean±SD; Median [Q1 – Q3])\n\n- Group F = 63.3±16.0; 65 [50–72.0]\n\n- Group W = 47.0±17.1; 48.3 [32.9–59.6]\n\n- Group C = 31.6±8.5; 28.3 [23.3–37.0]\n\n- p value = <0.001\n\n2. GRV, mL (Mean±SD; Median [Q1 – Q3])\n\n- Group F = 18.5±8.2; 16.7 [13.1–20.6]\n\n- Group W = 12.9±5.0; 11.8 [9.5–16.5]\n\n- Group C = 12.7±6.2; 11.4 [7.7–18.0]\n\n- p value = <0.001\n\n3. Antral CSA, cm2 (Mean±SD; Median [Q1 – Q3])\n\n- Group F = 4.0±1.9; 3.8 [2.9–4.7]\n\n- Group W = 2.7±1.0; 2.5 [2.0–3.4]\n\n- Group C = 2.3±1.2; 2.0 [1.5–2.5]\n\n- p value = <0.001\n\n4. blood glucose (Mean±SD; Median [Q1 – Q3])\n\n- Group F = 112.8±23.3; 109.0 [102.3–118.5]\n\n- Group W = 101.7±14.6; 97.5 [92.0–113.0]\n\n- Group C = 90.9±12.8; 90.0 [82.0–102.3]\n\n- p value = <0.001\n\n\n\n1. Perioperative agitation (n [%])\n\n- Intervention Group = 6 [10]\n\n- Control Group = 33 [55]\n\n- p value = 0.001\n\n\n\n1. Postoperative nausea (n [%])\n\n- Group A = 17 [28.3]\n\n- Group B = 15 [25.4]\n\n- p value = 0.9\n\n2. Postoperative vomiting (n [%])\n\n- Group A = 7 [11.7]\n\n- Group B = 9 [15.3]\n\n- p value = 0.8\n\n3. Postoperative pain (n [%])\n\n- Group A = 25 [41.7]\n\n- Group B = 27 [45.7]\n\n- p value = 0.7\n\n4. Postoperative Irritability (n [%])\n\n- Group A = 20 [33.3]\n\n- Group B = 14 [23.7]\n\n- p value = 0.3\n\n5. Postoperative LOS, minute (Median [Q1 – Q3])\n\n- Group A = 135 [85-235]\n\n- Group B = 161 [90-354]\n\n- p value = 0.02\n\n6. blood glucose levels, mmol/L (Median [Q1 – Q3])\n\n- Group A = 5.4 [3.3-9.4]\n\n- Group B = 4.9 [3.6-6.5]\n\n- p value = 0.01\n\n7. blood ketone levels (Median [Q1 – Q3])\n\n- Group A = 0.2 [0-1.4]\n\n- Group B = 0.3 [0-1.6]\n\n- p value = 0.0003\n\n8. venous blood pH (Median [Q1 – Q3])\n\n- Group A = 7.36 [7.24-7.44]\n\n- Group B = 7.37 [7.25-7.44]\n\n- p value ≥ 0.9\n\n9. venous blood lactate, mmol/L (Mean [95 % CI])\n\n- Group A = 1.3 [1.25, 1.51]\n\n- Group B = 1.0 [0.96, 1.15]\n\n- p value ≤ 0.0001\n\n\n\n1. Mean volume of gastric content (Mean [SD]), ml/kg\n\n0.01\n\n- Carbohydrate Group = 0.41 [0.28]\n\n- Fasting Group = 0.28 [0.27]\n\n- p value = 0.01\n\n2. pH of gastric content (Mean [SD])\n\n- Carbohydrate Group = 1.9 [0.5]\n\n- Fasting Group = 2.0 [0.6]\n\n- p value = not significant\n\n3. Preoperative thirst (n [%])\n\n- Group A = 20 [32]\n\n- Group B = 14 [30]\n\n- p value = not significant\n\n4. Preoperative hunger (n [%])\n\n- Group A = 18 [30]\n\n- Group B = 20 [33]\n\n- p value = not significant\n\n5. Postoperative nausea (n [%])\n\n- Group A = 24 [25]\n\n- Group B = 6 [10]\n\n- p value = 0.028\n\n6. Postoperative vomiting (n [%])\n\n- Group A = 3 [5]\n\n- Group B = 1 [2]\n\n- p value = not significant\n\n\n\n1. The blood glucose = significant higher in groups B, C, and D than group A at the time of anesthesia.\n\n2. The gastric residual = no residue in groups A, B, and C. 15 infants in group D had a gastric residual volume.\n\n3. The crying ratio was significantly higher in group A.\n\n4. The length of hospital = not significant different between the groups.\n\n\n\n1. gastric emptying time, cm2 (Mean [95% CI, P Value])\n\n- CHO Group = significantly increased compared to baseline at 10 minutes 2.4 [1.5-3.4, 0.02], 30 minutes 1.1 [0.2-2.1, 0.02], and 60 minutes 1.5 [0.6-2.4, <0.001].\n\n- GS Group = significantly increased compared to baseline at 10 minutes 1.3 [0.6-2.0, 0.02].\n\n2. gastric fluid volume, ml (Mean Difference [95% CI])\n\n- CHO Group = significantly increased compared to baseline at 10 minutes −0.71 [−1.08 to −0.34], 30 minutes −0.38 [−0.71 to −0.05], and 60 minutes −0.43 [−0.86 to 0.01].\n\n- GS Group = significantly increased compared to baseline at 10 minutes −0.38 [−0.64 to −0.13].\n\n3. thirst (median [IQR; range])\n\n- CHO Group = significantly increased compared to baseline at 10 minutes 1.5 [0-4.0; 0-10], 30 minutes 2.0 [2.0-4.8; 0-8], and 60 minutes 3.0 [2.0-6.0; 0-8].\n\n- GS Group = significantly increased compared to baseline at 10 minutes 0 [0-0; 0-3], 30 minutes 1.0 [0-2.8; 0-5], and 60 minutes 1.5 [0-3.5; 0-6].\n\n- p value 10 minutes 0.01, 30 minutes 0.02, and 60 minutes 0.01\n\n4. hunger (median [IQR; range])\n\n- CHO Group = significantly increased compared to baseline at 10 minutes 2 [0.3-3.0; 0-5],\n\n- GS Group = significantly increased compared to baseline at 10 minutes 4 [3.3-5.0; 0-6]\n\n- p value = not significant\n\n\n\n1. Volume of gastric content, ml/kg body weight (Mean ± SD [95% CI, p Value]\n\n- 1-h fast group = 0.34±0.35 [0.29–0.39]\n\n- 2-h fast group = 0.43±0.33 [0.38–0.48]\n\n- p value = 0.011\n\n2. Blood glucose, mmol/L (Mean ± SD [95% CI])\n\n- 1-h fast group = 5.59±1.11 [5.43, 5.76]\n\n- 2-h fast group = 6.21±0.78 [6.09, 6.33]\n\n- p value = < 0.001\n\n3. Volume of gastric content >0.4 ml/kg of body weight (n [%])\n\n- 1-h fast group = 18 [10.6]\n\n- 2-h fast group = 35 [20.1]\n\n- p value = 0.014\n\n4. pH of gastric content <2.5 (n [%])\n\n- 1-h fast group = 88 [51.8]\n\n- 2-h fast group = 92 [52.9]\n\n- p value = not significant\n\n5. Pre- and intraoperative adverse reactions (n [%])\n\n- 1-h fast group = Crying, 68 [40]; Thirst, 35 [20.6]; Hypoxia, 9 [5.3]; Vomiting, 7 [4.1]; Pulmonary Aspiration, 3 [1.8]; Heart Failure, 2 [1.2].\n\n- 2-h fast group = Crying, 90 [51.7]; Thirst, 58 [33.3]; Hypoxia, 20 [11.5]; Vomiting, 6 [3.4]; Pulmonary Aspiration, 3 [1.7]; Heart Failure, 3 [1.7].\n\n- p value = Crying, 0.029; Thirst, 0.008; Hypoxia, 0.039; Vomiting, 0.745; Pulmonary Aspiration, 0.647; Heart Failure, 0.511.\n\n\n\n1. albumin (Mean±SD)\n\n- Fasting Group = preoperative, 4.08±0.39; postoperative, 3.82 ± 0.48\n\n- CHO Group = preoperative, 4.12 ± 0.29; postoperative, 3.77 ± 0.29\n\n- p value = preoperative, 0.94; postoperative, 0.53\n\n2. IL-6\n\n- Fasting Group = preoperative, 1.5 ± 2.6; postoperative, 3.82 ± 0.48\n\n- CHO Group = preoperative, 2.0 ± 2.3; postoperative, 1.5 ± 2.0 -p value = preoperative, 0.98; postoperative, 0.41\n\n3. blood glucose\n\n- Fasting Group = preoperative, 88 ± 16; postoperative, 91 ± 34 -CHO Group = preoperative, 86 ± 9; postoperative, 93 ± 24\n\n- p value = preoperative, 0.32; postoperative, 0.60\n\n4. insulin\n\n- Fasting Group = preoperative, 3.09 ± 6.34; postoperative, 91 ± 34\n\n- CHO Group = preoperative, 4.90 ± 4.52; postoperative, 4.55 ± 3.43\n\n- p value = preoperative, 0.69; postoperative, 0.78\n\n5. CRP\n\n- Fasting Group = preoperative, 3.60 ± 7.60; postoperative, 3.53 ± 7.75\n\n- CHO Group = preoperative, 0.53 ± 0.59; postoperative, 0.49 ± 0.53\n\n- p value = preoperative, 0.05; postoperative, 0.02\n\n6. HOMA IR\n\n- Fasting Group = preoperative, 0.86 ± 2.05; postoperative, 91 ± 34\n\n- CHO Group = preoperative, 1.57 ± 1.86; postoperative, 1.13 ± 1.05\n\n- p value = preoperative, 0.49; postoperative, 0.37\n\n7. PCR/albumin\n\n- Fasting Group = preoperative, 0.89 ± 1.86; postoperative, 0.91 ± 1.97\n\n- CHO Group = preoperative, 0.13 ± 0.15; postoperative, 0.13\n\n- ± 0.15\n\n- p value = preoperative, 0.03; postoperative, 0.08\n\n\n\n1. Time to attain MAS of >9, minutes (Mean±SD [Range])\n\n- Study Group = 18.70±10.19 [5-40]\n\n- Control Group = 16.86±6.85 [0-30]\n\n- p value = 0.007\n\n2. Time to ask for oral intake, minutes (Mean±SD [Range])\n\n- Study Group = 49.40±31.8 [10-120]\n\n- Control Group = 25.88±16.93 (5-90]\n\n- p value = 0.0001\n\n3. Time for attaining the discharge criteria, minutes (Mean±SD [Range])\n\n- Study Group = 35.5±14.11 [15-6]\n\n- Control Group = 38.04±13.71 [20-60]\n\n- p value = 0.841\n\n4. Preoperative UMSS (Median [Range])\n\n- Study Group = 0 (0-2)\n\n- Control Group = 1 (0-3)\n\n- p value = 0.001\n\n5. Postoperative UMSS (Median [Range])\n\n- Study Group = 2 [0-3]\n\n- Control Group = 2 [0-3]\n\n- p value = 0.305\n\n6. Preoperative behaviour score (Median [Range])\n\n- Study Group = 3 [1-4]\n\n- Control Group = 3 [1-4]\n\n- p value = 0.667\n\n7. Postoperative behaviour score (Median [Range])\n\n- Study Group = 3.5 [1-4]\n\n- Control Group = 3 [1-4]\n\n- p value = 0.412\n\n8. Child-parent separation score (Median [Range])\n\n- Study Group = 2 (1-3)\n\n- Control Group = 2 (1-3)\n\n- p value = 0.96\n\n9. Mask acceptance score (Median [Range])\n\n- Study Group = 3 (1-4)\n\n- Control Group = 3 (1-4)\n\n- p value = 0.659\n\n10. Random blood sugar, mg/dL (Median [IQR])\n\n- Study Group = 70 [60-79]\n\n- Control Group = 90 [85-98]\n\n- p value = 0.005\n\n\n\n1. Preoperative blood glucose concentrations (Mean±SD [Range])\n\n- Group A = 64.08 ± 25.37\n\n- Group Group B = 102.5 ± 16.97\n\n- p value ≤ 0.0001\n\n2. Hypoglycemia (n [%])\n\n- Group A = 39 (65)\n\n- Group Group B = 14 (23)\n\nThe research on PCL demonstrates its significant impact on stabilizing blood glucose levels and enhancing metabolic control before pediatric surgery. In studies comparing carbohydrate-loaded patients with those in control or fasting groups, those who received carbohydrates generally had higher, yet stable, blood glucose levels. For example, patients in the carbohydrate group showed a median blood glucose level of 5.4 mmol/L, which was slightly higher than that of the placebo group. This suggests that PCL helps maintain energy reserves during the perioperative period, reducing the risk of hypoglycemia and providing metabolic benefits.7 Additionally, carbohydrate-loaded patients exhibited fewer abnormal ketone levels, indicating better overall metabolic stability, although a slight increase in lactate levels was noted. These findings underscore the potential of PCL to enhance patient outcomes by ensuring more consistent blood glucose levels before surgery.8\n\nFurther research supports these benefits by showing that PCL not only stabilizes blood glucose levels but also reduces the likelihood of hyperglycemia and hypoglycemia. In one study, fasting patients were more prone to hyperglycemia, with 21% exceeding 99 mg/dL, while none of the patients in the carbohydrate group experienced hyperglycemia. Both groups had similar insulin levels and Homeostatic Model Assessment for IR (HOMA-IR) Index data before and after the operation, indicating that carbohydrate loading helps prevent hyperglycemia without affecting insulin resistance.9 Another study found significantly lower random blood sugar levels in the carbohydrate group compared to the control group, where higher glucose levels were trending. This suggests that carbohydrate loading effectively prevents the sharp fluctuations in blood glucose that can pose risks during surgery.10 Moreover, the fasting group was found to experience more frequent signs of hypoglycemia, such as excessive crying, sweating, and irritability, highlighting the discomfort and potential risks associated with fasting.11 Overall, the evidence points to PCL as a valuable strategy for improving preoperative care by maintaining blood glucose stability, reducing metabolic risks, and enhancing patient comfort.\n\nThe studies collectively highlight the positive impact of PCL and varying fasting durations on reducing perioperative anxiety, agitation, and crying in pediatric patients. One study found that children who received PCL were significantly calmer, with 90% being quiet and relaxed compared to only 11.7% in a control group, indicating a notable decrease in postoperative agitation.12 Another investigation showed that preoperative anxiety levels were markedly lower in children who consumed carbohydrate fluids, as measured by the modified Yale Preoperative Anxiety Scale (m-YPAS), compared to those who fasted.13 While one randomized clinical trial found no significant differences in behavior scores, separation anxiety, or mask acceptance between PCL and fasting groups, other research demonstrated that PCL significantly reduced preoperative crying, likely due to its ability to alleviate thirst and hunger. This reduction in crying not only conserves energy but also prevents gastrointestinal flatulence, which can interfere with surgical procedures.8,10 Additionally, a comparison of fasting durations revealed that a 1-hour fasting period resulted in fewer instances of crying than a 2-hour fast, supporting the effectiveness of shorter fasting intervals combined with PCL.14 Overall, the studies emphasize the benefits of PCL in minimizing anxiety, agitation, and crying, thereby improving the preoperative experience for pediatric patients.\n\nThe studies provide a comprehensive analysis of how PCL and different fasting durations impact gastric content, gastric emptying, and the associated risks of pulmonary aspiration. Significant differences were observed in antral cross-sectional area (CSA) and gastric residual volume (GRV) between groups, with both PCL and water intake leading to reduced gastric content compared to fasting.13 This reduction is further emphasized by findings that PCL led to a 68% decrease in gastric content compared to fasting, highlighting its effectiveness in minimizing gastric volume.15 Temporal changes in antral CSA indicate that PCL causes a significant increase within the first 10 minutes post-ingestion, which then returns to baseline by 90 minutes, underscoring the importance of timing in PCL administration.16\n\nIn terms of safety, administering PCL at doses up to 10 mL/kg does not increase gastric residuals in adults, children, or infants, thereby reducing the risk of regurgitation and aspiration. The absence of significant gastric residuals across these groups suggests that PCL is well-tolerated and does not compromise gastric emptying.8 Additionally, a comparison of fasting durations reveals that a 1-hour fast results in lower gastric content volume than a 2-hour fast, without increasing the risk of low gastric pH or the simultaneous presence of large gastric volumes and low pH, which are key factors in pulmonary aspiration risk.14 Collectively, these studies underscore the efficacy and safety of PCL in reducing gastric content and improving patient outcomes by lowering the risk of aspiration during surgery.\n\nPCL indicate that it generally does not lead to significant postoperative complications compared to fasting or placebo groups. For instance, the incidence of nausea, vomiting, and pain were similar between groups, with no significant differences observed in postoperative anti-emetic use or pain severity. In one study, children who received carbohydrates were more likely to experience severe nausea in the first postoperative hour, but this difference did not persist beyond that time. Additionally, there was no difference in the time taken to meet discharge criteria between those who received carbohydrates and those who did not.7,10,11\n\nOther studies reported no complications during the study period, and PCL was associated with reduced incidences of crying, thirst, and hypoxia compared to longer fasting durations. Moreover, no adverse reactions such as pulmonary aspirations, heart failure, or severe hunger were observed in the groups receiving carbohydrates.14,16 These findings suggest that PCL is generally safe and does not increase the risk of postoperative complications, while it may offer some benefits in reducing discomfort after surgery.\n\nThe study results indicate that PCL has a potential impact on the length of stay (LOS) in a hospital setting. One study found a significant reduction in the median LOS for children in the carbohydrate group (135 minutes) compared to the placebo group (161 minutes), suggesting that carbohydrate loading may enhance postoperative recovery and reduce hospital stay. However, the clinical relevance of this 26-minute reduction is debatable, and the study suggests that maintaining adequate hydration alone, regardless of carbohydrate intake, could improve postoperative recovery. It was also noted that the effect was more pronounced in morning surgeries, potentially influencing future clinical guidelines for preoperative hydration.7 Another study, however, found no significant difference in the length of hospital stay between groups receiving PCL and those who did not.8 This inconsistency highlights the need for further research to better understand the factors influencing LOS and the role of PCL in pediatric surgical recovery.\n\n\n4. Discussion\n\nThe ASA guideline emphasizes the importance of adhering to recommended fasting durations to avoid adverse patient and clinical outcomes. Prolonged fasting, whether due to unclear or extended fasting policies, can increase preoperative thirst, hunger, anxiety, and discomfort, as well as lead to dehydration and hypotension during anesthesia induction. Although recent European and Canadian guidelines suggest reducing clear liquid fasting to 1 hour in children, the ASA task force refrains from making a strong recommendation due to limited and low-quality evidence.17–20\n\nIn this context, PCL emerges as a promising intervention to address some of these challenges, particularly in pediatric populations. The findings from this scoping review suggest that PCL may offer several benefits, including stabilizing blood glucose levels, reducing metabolic risks, and improving perioperative outcomes. Studies consistently demonstrated that PCL helps maintain higher yet stable blood glucose levels, thereby reducing the risk of hypoglycemia during the perioperative period. This metabolic stability is crucial for pediatric patients, ensuring that energy reserves are maintained, potentially leading to better recovery outcomes.\n\nHowever, the review also highlighted variability in outcomes such as LOS and the incidence of postoperative complications. While some studies reported a significant reduction in LOS for patients receiving PCL, others found no significant differences between the PCL and control groups. This inconsistency suggests that the effectiveness of PCL may be influenced by factors like the timing of surgery, the type of surgical procedure, and individual patient characteristics.\n\nMoreover, although PCL generally did not increase the risk of postoperative complications, the extent of its benefits varied across different studies. Some studies reported reductions in postoperative complications such as nausea, vomiting, and pain, while others did not. These mixed results highlight the need for further research to better understand the specific conditions under which PCL provides the most benefit.\n\nIn evaluating the risk of bias among the included studies, we found that four studies had a low risk, five studies had some concerns, and one study had a high risk. This variability in study quality may partially explain the inconsistent findings related to PCL outcomes, such as the LOS and the incidence of postoperative complications. While some studies reported significant benefits from PCL, others did not, indicating that the effectiveness of PCL might be influenced by factors like surgical timing, procedure type, and patient characteristics.\n\nFuture research should therefore prioritize the optimization of PCL protocols and evaluate its effects across a wider spectrum of surgical procedures. It is particularly crucial to investigate the most effective practices for PCL in pediatric populations, where the current body of evidence is notably limited. Research efforts should focus on identifying the optimal timing, dosage, and composition of PCL to fully harness their potential benefits. Furthermore, certain outcomes, including advanced metabolic indicators such as insulin levels, HOMA-IR, blood analysis, and inflammation biomarkers like IL-6, have been inadequately explored in existing studies. Addressing these gaps in knowledge will be essential for refining preoperative care practices, with the ultimate goal of improving patient outcomes and reducing the adverse effects associated with prolonged fasting.\n\n\n5. Conclusions\n\nPCL in pediatric surgery may stabilize blood glucose, reduce metabolic risks, improve recovery, and minimize postoperative complications. However, evidence on safety and outcomes, such as hospital stay length and postoperative complications, remains inconsistent, highlighting the need for further research to refine PCL protocols for pediatric care.\n\nEthical approval and consent were not required.", "appendix": "Data availability\n\nNo underlying data are associated with this article.\n\nNo extended data are associated with this article.\n\nOpen Science Framework: Preoperative Carbohydrate Loading in Pediatric Surgery: A Scoping Review of Current Evidence, DOI: https://doi.org/10.17605/OSF.IO/KPNV5. 21\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\nPogatschnik C, Steiger E: Review of Preoperative Carbohydrate Loading. Nutr. Clin. Pract. Off. Publ. Am. Soc. Parenter Enter Nutr. 2015 Oct; 30(5): 660–664. Publisher Full Text\n\nNygren J, Thorell A, Ljungqvist O: Preoperative oral carbohydrate therapy. Curr. Opin. Anaesthesiol. 2015 Jun; 28(3): 364–369. Publisher Full Text\n\nChen L, Wang N, Xie G, et al.: The safety of preoperative carbohydrate drinks in extremely elderly patients assessed by gastric ultrasonography: a randomized controlled trial. BMC Anesthesiol. 2024; 24(1): 1–10. Publisher Full Text\n\nWang X, Zhuang J, Cheng J, et al.: Effect of preoperative oral carbohydrates on insulin resistance in patients undergoing laparoscopic cholecystectomy: a randomized controlled trial. Langenbeck’s Arch. Surg. 2024; 409(1): 77–79. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKumar SM, Anandhi A, Sureshkumar S, et al.: Effect of preoperative oral carbohydrate loading on postoperative insulin resistance, patient-perceived well-being, and surgical outcomes in elective colorectal surgery: a randomized controlled trial. J. Gastrointest. Surg. 2024. PubMed Abstract \| Publisher Full Text Reference Source\n\nRaval MV, Brockel MA, Kolaček S, et al.: Key Strategies for Optimizing Pediatric Perioperative Nutrition-Insight from a Multidisciplinary Expert Panel. Nutrients. 2023 Mar; 15(5). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLaird A, Bramley L, Barnes R, et al.: Effects of a Preoperative Carbohydrate Load on Postoperative Recovery in Children: A Randomised, Double-Blind, Placebo-Controlled Trial. J. Pediatr. Surg. 2023; 58(9): 1824–1831. PubMed Abstract \| Publisher Full Text\n\nJiang W, Liu X, Liu F, et al.: Safety and benefit of pre-operative oral carbohydrate in infants: A multi-center study in China. Asia Pac. J. Clin. Nutr. 2018; 27(5): 975–979. PubMed Abstract \| Publisher Full Text\n\nCarvalho CAL d B, de Carvalho AA , Preza AD, et al.: Metabolic and inflammatory benefits of reducing preoperative fasting time in pediatric surgery. Rev. Col. Bras. Cir. 2020; 47(1): 1–10.\n\nBharadwaj AA, Bhukal I, Mathew PJ: Effect of Oral 12% Carbohydrate Containing Clear Fluid in Children on Post-anaesthesia Recovery Profile- A Randomised Clinical Trial. J. Clin. Diagn. Res. 2021; 10–13. Publisher Full Text\n\nBalasubramaniam U, Kiran U, Hasija S, et al.: Randomized Study Comparing Pre-Operative Glycemic Profile in Pediatric Cardiac Surgical Patients Administered Oral Carbohydrate Solution Preoperatively versus Those Kept Fasting. World J. Cardiovasc. Dis. 2018; 08(06): 298–306. Publisher Full Text\n\nKarami T, Karami N, Hoshyar H, et al.: Evaluation of Effect of Preoperative Oral Carbohydrate on the Perioperative Agitation in Pediatrics Undergoing Elective Herniorrhaphy; A Doubl-Blind Randomized Clinical Trial. Int. J. Pediatr. 2021; 9(1): 12815–12823. Reference Source\n\nBozoglu Akgun B, Hatipoglu Z, Gulec E, et al.: The Effect of Oral Fluid Administration 1 Hour before Surgery on Preoperative Anxiety and Gastric Volume in Pediatric Patients. Eur. Surg. Res. 2024; 65(1): 54–59. PubMed Abstract \| Publisher Full Text\n\nHuang X, Zhang H, Lin Y, et al.: Effect of oral glucose water administration 1 hour preoperatively in children with cyanotic congenital heart disease: A randomized controlled trial. Med. Sci. Monit. 2020; 26: 1–6.\n\nTudor-Drobjewski BA, Marhofer P, Kimberger O, et al.: Randomised controlled trial comparing preoperative carbohydrate loading with standard fasting in paediatric anaesthesia. Br. J. Anaesth. 2018; 121(3): 656–661. PubMed Abstract \| Publisher Full Text\n\nZhang YL, Li H, Zeng H, et al.: Ultrasonographic evaluation of gastric emptying after ingesting carbohydrate-rich drink in young children: A randomized crossover study. Paediatr. Anaesth. 2020; 30(5): 599–606. PubMed Abstract \| Publisher Full Text\n\nFrykholm P, Disma N, Andersson H, et al.: Pre-operative fasting in children: A guideline from the European Society of Anaesthesiology and Intensive Care. Eur. J. Anaesthesiol. 2022 Jan; 39(1): 4–25. PubMed Abstract \| Publisher Full Text\n\nJoshi GP, Abdelmalak BB, Weigel WA, et al.: 2023 American Society of Anesthesiologists Practice Guidelines for Preoperative Fasting: Carbohydrate-containing Clear Liquids with or without Protein, Chewing Gum, and Pediatric Fasting Duration - A Modular Update of the 2017 American Society of Anesthes. Anesthesiology. 2023; 138(2): 132–151. PubMed Abstract \| Publisher Full Text\n\nPractice Guidelines for Preoperative Fasting and the Use of Pharmacologic Agents to Reduce the Risk of Pulmonary Aspiration: Application to Healthy Patients Undergoing Elective Procedures: An Updated Report by the American Society of Anesthesiologists Ta. Anesthesiology. 2017 Mar; 126(3): 376–393. PubMed Abstract \| Publisher Full Text\n\nDobson G, Filteau L, Fuda G, et al.: Guidelines to the Practice of Anesthesia - Revised Edition 2022. Can. J. Anaesth. 2022 Jan; 69(1): 24–61. PubMed Abstract \| Publisher Full Text\n\nSari D, Rayhan A, Widyastuti Y, et al.: Study protocol - preoperative carbohydrate loading in pediatric surgery: A scoping review of current evidence. OSF. 2024. Publisher Full Text" }	[ { "id": "327893", "date": "18 Oct 2024", "name": "Vibhavari Naik", "expertise": [ "Reviewer Expertise Vibhavari Naik - oncoanaesthesia", "paediatric anaesthesia", "vascular access", "Abhijit Nair - statistics and research methodology", "systematic reviews" ], "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 authors have selected a pertinent topic for a scoping review. Although the concept and components of ERAS have been adopted from adult practice, evidence needs to be established to justify its role in paediatric care. In this article, the authors have evaluated the evidence regarding the use of preoperative carbohydrate loading as a component of ERAS in children.\nThe authors mention this as a scoping review but the methodology has overlapped with a systematic review. The inclusion of a risk of bias assessment in this study suggests a more rigorous approach typically used with systematic reviews. This being a scoping review authors should intend to provide an overview of the existing evidence including gaps in literature and determine if systematic review is required. Literature review for a scoping review apart from RCTs, should also include case reports, case series, observational studies, guidelines, blogs, etc. Alternatively, since most of the included studies are RCTs, authors could choose to perform a qualitative systematic review and a possible quantitative meta-analysis. But this would require following the stringent PRISMA methodology including study registration.\nTitle - is clear and informative and includes the mention of scoping review\nAbstract - is structured and conveys the gist of the review. The introduction part of the abstract could end with the objective of the review. In the methods section of abstract, future tense phrases like ‘would be used’ and ‘would focus’ could be changed to past tense like ‘was used’ and ‘was focused’. The authors mention ‘recent’ articles but have not explained what recent means.\nIntroduction – Authors could write introduction more crisply by avoiding repetition of enhancing patient comfort and improving outcomes; and reducing the content on adult population. It broadly could include what is available in literature about paediatric preoperative carbohydrate loading and end with what this review aims to achieve.\nMethods – The PRISMA-ScR reporting guidelines could be accessed at Open Science Framework platform. But the authors have not mentioned if the protocol for the review exists, registration of the protocol and the link to the same.\n\nA formal risk of bias assessment may not be necessary for scoping reviews.\n\nFuture tense of methodology needs to be changed to past tense throughout the methods section. Ex. The selection process will involved, Full-text articles would be reviewed, etc.\nEligibility criteria – includes the word ‘recent’ the time duration of which has not been explained.\n\nThe authors must provide truncations of the search results from various databases in a supplementary file. They could present the full electronic search strategy for at least 1 database, including any limits used, such that it could be repeated. Authors could include the date of database search.\nFigure 1 PRISMA flow diagram mentions - Records excluded Authors could mention what two asterisks stand for in the footnote of the figure mentioning the reason for exclusion. Authors mention ‘Reports not retrieved’ but have not mentioned the reason for non-retrieval. Authors mention 240 reports excluded after assessing for eligibility from 365 reports. However only 10 articles are included in review. Authors need to mention the reasons for excluding 115 articles.\nClarity on how many reviewers contributed to data charting will be useful instead of mentioning ‘multiple’.\nResults – Table 1 legends should include expansion of short forms used in the table.\nLength of stay – though authors mention inconsistency in results, the results appear to be consistent in studies not demonstrating clinically relevant difference amongst the groups. This could be restructured.\nCarvalho in their abstract mention - Four patients (21%) in the fasting group but none in the CHO group were hyperglycemic before surgery (p = 0.04). This has been missed while compiling this review.\nWe have found a few relevant studies which were not included in this review. Could authors provide an explanation for the same?\nGawecka A, Mierzewska-Schmidt M. Tolerance of, and metabolic effects of, preoperative oral carbohydrate administration in children - a preliminary report. Anaesthesiol Intensive Ther. 2014 Apr-Jun;46(2):61-4. Bauer JM, Trask M, Coughlin G, Gopalan M, Gupta A, Yaszay B, Yang S, Grigg E. Pre-operative carbohydrate drink in pediatric spine fusion: randomized control trial. Spine Deform. 2024 Sep;12(5):1283-1287.\n\nDiscussion – Authors mention that - Studies consistently demonstrated that PCL helps maintain higher yet stable blood glucose levels. However, Bharadwaj et al show lower glucose levels in children receiving PCL. The effect of PCL on glucose levels could be rewritten as inconsistent. Authors could highlight and explore the inconsistency in glucose levels after PCL and identify if any factors like duration of fasting affected it.\nIn Laird et al. difference of ketones by 0.1 and lactates by 0.3 mmol/L with overlapping ranges though may be statistically significant, the clinical relevance of such small differences may not be relevant. Authors could highlight such trivial ‘statistically significant’ results across all the articles included and explore the clinical relevance in the discussion of this review.\nAuthors could include a mention of recently published meta-analysis on the topic and compare results - Li Y, Chen L, Su Y, Zhang X. Preoperative Oral Carbohydrates for Children: A Meta-Analysis and Systematic Review. Biol Res Nurs. 2024 Oct;26(4):624-635.\n\nScoping reviews can have the following biases: language, search, data extraction, reporting, reviewer, publication, and selection bias. Authors need to mention these in the limitations including steps taken to mitigate them.\nFew areas of spelling mistakes and improvement of grammar would be required.\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? Partly\n\nAre the conclusions drawn adequately supported by the results presented in the review? No\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": [ { "c_id": "12784", "date": "26 Nov 2024", "name": "Amar Rayhan", "role": "Author Response", "response": "Thank you for your insightful feedback and constructive comments on our manuscript. We appreciate the opportunity to improve our work and have made significant revisions based on your suggestions. In response, we have made the following adjustments: Overall Comment from reviewer : The authors have selected a pertinent topic for a scoping review. Although the concept and components of ERAS have been adopted from adult practice, evidence needs to be established to justify its role in paediatric care. In this article, the authors have evaluated the evidence regarding the use of preoperative carbohydrate loading as a component of ERAS in children Respond: We already added the narrative review about ERAS in pediatric including Preoperative Carbohydrat loading as part of ERAS in pediatric Comment from reviewer : The authors mention this as a scoping review but the methodology has overlapped with a systematic review. The inclusion of a risk of bias assessment in this study suggests a more rigorous approach typically used with systematic reviews. This being a scoping review authors should intend to provide an overview of the existing evidence including gaps in literature and determine if systematic review is required. Literature review for a scoping review apart from RCTs, should also include case reports, case series, observational studies, guidelines, blogs, etc. Alternatively, since most of the included studies are RCTs, authors could choose to perform a qualitative systematic review and a possible quantitative meta-analysis. But this would require following the stringent PRISMA methodology including study registration Respond: Thank you for your feedback, according the scope and methodology we have revised some parts: The title and eligibility criteria have been revised to \"Preoperative Carbohydrate Loading in Pediatric Surgery: A Scoping Review of Current Clinical Trials\" to clarify the focus and methodology of the study. This adjustment emphasizes our intent to provide an overview of existing evidence while addressing gaps in the literature surrounding the topic. Our scoping review now specifies clinical trials research, prompting us to revise the methodology. The RoB (Risk of Bias) tool from Cochrane was chosen to evaluate the critical appraisal of included clinical studies. Title Comment from reviewer : - is clear and informative and includes the mention of scoping review Respond: Thank you very much Abstract Comment from reviewer : is structured and conveys the gist of the review. The introduction part of the abstract could end with the objective of the review. In the methods section of abstract, future tense phrases like ‘would be used’ and ‘would focus’ could be changed to past tense like ‘was used’ and ‘was focused’. The authors mention ‘recent’ articles but have not explained what recent means Respond: Thank you for your comment, we have adjusted some parts : The objective has been repositioned in the introduction. We have improved the methods section. The term ‘recent’ has been changed to “2017-2024.” Grammatical errors have been fixed in the method section Introduction Comment from reviewer : Authors could write introduction more crisply by avoiding repetition of enhancing patient comfort and improving outcomes; and reducing the content on adult population. It broadly could include what is available in literature about paediatric preoperative carbohydrate loading and end with what this review aims to achieve. Respond: We reduced content related to the adult population and add more pediatric content Methods Comment from reviewer; The PRISMA-ScR reporting guidelines could be accessed at Open Science Framework platform. But the authors have not mentioned if the protocol for the review exists, registration of the protocol and the link to the same. A formal risk of bias assessment may not be necessary for scoping reviews. Future tense of methodology needs to be changed to past tense throughout the methods section. Ex. The selection process will involved, Full-text articles would be reviewed, etc. Eligibility criteria – includes the word ‘recent’ the time duration of which has not been explained. The authors must provide truncations of the search results from various databases in a supplementary file. They could present the full electronic search strategy for at least 1 database, including any limits used, such that it could be repeated. Authors could include the date of database search. Figure 1 PRISMA flow diagram mentions - Records excluded Authors could mention what two asterisks stand for in the footnote of the figure mentioning the reason for exclusion. Authors mention ‘Reports not retrieved’ but have not mentioned the reason for non-retrieval. Authors mention 240 reports excluded after assessing for eligibility from 365 reports. However only 10 articles are included in review. Authors need to mention the reasons for excluding 115 articles. Clarity on how many reviewers contributed to data charting will be useful instead of mentioning ‘multiple’. Respond: Thank you for your detailed feedback; We have already included study protocol at OSF. Actually, we had added it prior submitting the manuscript but the editor asked us to remove the protocol file. Thank you for your feedback. We agree that a formal risk of bias assessment may not be required for a scoping review. However, PRISMA-ScR guidelines recommend including a section for critical appraisal of the evidence. Although using RoB 2.0 is a formal approach, we believe it provides a thorough and reliable overview of study quality. Employing a trusted tool like RoB 2.0 allows us to gauge the clinical study quality in a structured way, enhancing the overall insight into the evidence included in our review. Grammatical errors have been fixed. The terms ‘recent’ and ‘multiple’ have been edited. We have provided truncations of the search results from various databases in a supplementary file, including the full electronic search strategy for at least one database with specified limits and the date of the database search. The PRISMA flowchart was revised to clarify the reasons for exclusion, noting that the asterisks in the footnote referred to a format picture that had not been deleted. Additionally, we included the reasons for excluding 385 articles from the initial assessment of 395 full-text articles. The terms ‘recent’ and ‘multiple’ have been edited. Results Comment from reviewer; Table 1 legends should include expansion of short forms used in the table. Length of stay – though authors mention inconsistency in results, the results appear to be consistent in studies not demonstrating clinically relevant difference amongst the groups. This could be restructured. Carvalho in their abstract mention - Four patients (21%) in the fasting group but none in the CHO group were hyperglycemic before surgery (p = 0.04). This has been missed while compiling this review. We have found a few relevant studies which were not included in this review. Could authors provide an explanation for the same? Gawecka A, Mierzewska-Schmidt M. Tolerance of, and metabolic effects of, preoperative oral carbohydrate administration in children - a preliminary report. Anaesthesiol Intensive Ther. 2014 Apr-Jun;46(2):61-4. Bauer JM, Trask M, Coughlin G, Gopalan M, Gupta A, Yaszay B, Yang S, Grigg E. Pre-operative carbohydrate drink in pediatric spine fusion: randomized control trial. Spine Deform. 2024 Sep;12(5):1283-1287. Respond: Thank you for your thorough feedback; We have added the expansions of all abbreviations and short forms used in Table 1 to the table legend, as suggested. Regarding Length of Stay (LOS), we acknowledged the inconsistency in previous reports and agreed that the results should be restructured. The studies consistently showed no clinically meaningful difference in the length of hospital stay. Preoperative hyperglycemia was added to the study characteristics table. We excluded the study by Gawecka A et al. (2014) from our scoping review due to its publication year and added the study by Bauer JM et al. (2024). Discussion Comment from reviewer; Authors mention that - Studies consistently demonstrated that PCL helps maintain higher yet stable blood glucose levels. However, Bharadwaj et al show lower glucose levels in children receiving PCL. The effect of PCL on glucose levels could be rewritten as inconsistent. Authors could highlight and explore the inconsistency in glucose levels after PCL and identify if any factors like duration of fasting affected it. In Laird et al. difference of ketones by 0.1 and lactates by 0.3 mmol/L with overlapping ranges though may be statistically significant, the clinical relevance of such small differences may not be relevant. Authors could highlight such trivial ‘statistically significant’ results across all the articles included and explore the clinical relevance in the discussion of this review. Authors could include a mention of recently published meta-analysis on the topic and compare results - Li Y, Chen L, Su Y, Zhang X. Preoperative Oral Carbohydrates for Children: A Meta-Analysis and Systematic Review. Biol Res Nurs. 2024 Oct;26(4):624-635. Scoping reviews can have the following biases: language, search, data extraction, reporting, reviewer, publication, and selection bias. Authors need to mention these in the limitations including steps taken to mitigate them. Respond: We revised our analysis to emphasize the inconsistencies in blood glucose levels following preoperative carbohydrate loading (PCL), highlighting factors such as fasting duration. Additionally, we addressed trivial statistically significant findings, such as the small differences in blood ketones and lactates reported by Laird et al., to explore their clinical relevance. The current meta-analysis was added and discussed. We already included Chen et al. study We explained the limitations of the study" } ] } ]	1	https://f1000research.com/articles/13-1089
https://f1000research.com/articles/13-479/v1	17 May 24	{ "type": "Research Article", "title": "Effects of electroacupuncture on cognitive symptoms in major depressive disorder: a pilot study and randomized controlled trial", "authors": [ "Yindee Boontra", "Chommakorn Thanetnit", "Muthita Phanasathit", "Yindee Boontra", "Chommakorn Thanetnit" ], "abstract": "Objectives To investigate the impact of electroacupuncture on cognitive function, quality of life (QoL), and depression severity in patients with major depressive disorder (MDD).\n\nMethods This double-blinded randomized controlled trial included 60 participants aged 18-55 with cognitive symptoms related to MDD at Thammasat University Hospital. Participants were divided into two groups: the electroacupuncture group combined with standard antidepressant treatment (EG; n=30) and the control group receiving standard care with placebo acupuncture (CG; n=30). The study assessed 1) executive functions using the Trail making test- B and Stroop Color and Word Test, 2) delayed recall, and 3) subjective cognitive complaint and Quality of life (QoL) using WHODAS 2.0. Depressive symptoms were measured using the Thai version of the Patient Health Questionnaire (PHQ-9). Baseline and post-intervention assessments were conducted over 10 weeks. Mann-Whitney U test analyzed treatment effects by comparing median differences between groups.\n\nResults Both groups exhibited similar demographics and cognitive traits. Cognitive improvement was observed in both groups at the endpoint. Intention-to-treat analysis revealed significantly higher median scores for subjective cognitive complaints in the EG compared to the CG (EG: Median = 5.5, CG: Median = 0.0, p=0.049). No serious side effects were identified from either electroacupuncture or placebo acupuncture.\n\nConclusions Electroacupuncture improved subjective complaints in MDD patients with cognitive symptoms, but did not show effects on specific cognitive functions, QoL, or depressive symptoms. This study provides initial evidence supporting the potential of electroacupuncture in MDD patients with cognitive symptoms, suggesting opportunities for further research.\n\nTrial registration NCT06239740, February 2, 2024, ClinicalTrials.gov.", "keywords": [ "Electroacupuncture", "Depression", "Cognition", "Humans", "Pilot", "Randomized Controlled Trial" ], "content": "Introduction\n\nDepressive disorder is currently considered a significant health issue, with depression affecting up to 264 million people worldwide. Depression has both short-term and long-term consequences, with the most severe impact being suicide. Approximately 800,000 people die by suicide each year, making it the second leading cause of death among individuals aged 15-29 years. According to the World Health Organization (WHO). Approximately 1.5 million people in Thailand suffer from depressive symptoms.1\n\nDepression leads to cognitive dysfunction, affecting executive function and working memory.2,3 This results in decreased attention and slow thinking, which are key diagnostic criteria for depression according to the Diagnostic and Statistical Manual of Mental Disorders (DSM-5). Even after depressive symptoms have improved, cognitive problems may persist, particularly in terms of work performance.3\n\nCurrent research indicates that various treatments may help alleviate cognitive dysfunction in patients with depression. Medications like duloxetine have been shown to improve psychomotor cognitive function,4 while vortioxetine has demonstrated benefits in executive function, attention, processing speed, and learning and memory. Additionally, treatments such as Electroconvulsive therapy (ECT) and repetitive transcranial magnetic stimulation (rTMS) have shown potential in improving cognitive function in depression patients.5–7\n\nAcupuncture, a traditional eastern medicine practice, has been used for over 4,000 years in China. The practice involves inserting tiny needles into specific points on the body to restore balance and harmony. Acupuncture has gained popularity due to its minimal side effects. Some studies have suggested that acupuncture may aid in the treatment of cognitive impairment in patients with schizophrenia, dementia, or mild cognitive impairment.8–10 It has also been found to improve cognitive function and reduce depressive symptoms.11 Acupuncture may stimulate the release of substances like brain-derived neurotrophic factor (BDNF),9 which is crucial for neural plasticity, supporting the growth of new neurons in the brain.\n\nElectroacupuncture, defined as acupuncture with electrical stimulation, includes inserting needles at acupuncture points and applying a low-level electrical current through them. A small amount of electricity flows through the electrode, resulting in a gentle vibration or soft hum during the treatment, enhancing the effects of acupuncture. This shown to increase the release of vascular endothelial growth factor (VEGF)12 and enhance cerebral blood flow and volume, ultimately improving motor function in ischemic stroke patients.13 In animal studies, electroacupuncture has been found to stimulate the release of nitric oxide (NO) and reduce angiotensin II, leading to better blood vessel dilation in the brain.14,15 The authors hypothesized that electroacupuncture has the potential to treat subjective cognitive complaints in depressive patients, which is a relatively new and promising area of research. It is a minimally invasive and well-tolerated treatment option.\n\n\nMethods\n\nThis trial was registered to the ClinicalTrials.gov (NCT06239740), with a registration date of February 2, 2024 and the Human Research Ethics Committee of Thammasat University (Medicine) has granted approval for this medical research study (MTU-EC-PS-1-304/64) with an approval date from December 24, 2021 to December 23, 2022. It is in full compliance with international guidelines such as the Declaration of Helsinki, The Belmont Report, CIOMS Guidelines, and the International Conference on Harmonisation-Good Clinical Practice (ICH-GCP). This study protocol outlined a controlled clinical trial conducted at a single center, focusing on evaluating the impact of electroacupuncture on specific cognitive functions in individuals with major depressive disorder (MDD) who experience cognitive dysfunction.\n\nA total of 60 eligible participants, aged between 18 and 55 years, with cognitive symptoms resulting from MDD, will be recruited from the psychiatry outpatient clinic at Thammasat University Hospital. All participants with MDD and low suicidal risk, diagnosed according to the DSM-516 or DSM-IV-TR criteria,17 and currently receiving treatment with antidepressants for at least 3 months from psychiatrists and psychiatric residents, were included. The subjective cognitive symptom was screened using the 7th item of the Thai version of the Patient Health Questionnaire (Thai-PHQ-9),18 which assesses experiencing trouble concentrating or cognitive problems (e.g., reading or watching TV). The exclusion criteria were: (i) Individuals with severe cognitive deficits from traumatic brain injury, delirium, neurodevelopmental disorders, or intellectual disability; (ii) Individuals with neurological disorders such as stroke, Parkinson’s disease, epilepsy, or other brain lesions; (iii) Individuals with severe medical conditions preventing lying down for 20 minutes; (iv) Individuals with recent electroconvulsive therapy (ECT) within the last 6 months; (v) Individuals with a pacemaker; (vi) Individuals with visual or hearing impairment that could not be corrected with eyeglasses or hearing aids; and (vii) Individuals diagnosed with severe MDD or scoring 20 points or more on the Thai-PHQ-9. Each participant was provided with detailed information regarding the study and informed written consent was obtained for their participation in the research. Subsequently, all participants granted consent for publication.\n\nAn assistant researcher, who had contact with participants but was not involved in the interventions and outcome assessment, assigned participants to the intervention or control arm using a block of four randomization. The participants were randomly assigned to either the treatment group or the control group in a 1:1 ratio. The treatment group will undergo electroacupuncture (EG), while the control group will receive sham acupuncture (CG). Both groups received antidepressants with adjunctive medication (i.e., benzodiazepines, tricyclics, or antipsychotics) as the standard treatment. All participants were assessed for executive functions and memory using specific cognitive tests, including the Trail Making Test B (TMT-B), Stroop Color and Word Test (SCWT), category delayed recall in the Alzheimer’s Disease Assessment Scale–Cognitive Subscale (ADAS-Cog),19,20 and subjective reports of concern regarding concentration, memory, problem-solving, learning, communication, and quality of life (QoL) concerns using the WHO Disability Assessment Schedule (WHODAS 2.0; sections D1.1-1.6 and H1-3).21 Additionally, depressive symptoms were assessed using the Thai-PHQ-9. All administrative tasks and data entry were carried out by the research assistant, who was also not involved in the interventions and outcome assessment (Figure 1A,B).\n\nThis study was double-blinded. All participants, assessors, acupuncturists, and researchers were unaware of group assignments. Before treatment began, participants would not recognize if they were in the real acupuncture or sham group. They were placed in separate, quiet units on different days.\n\nThe elimination or termination criteria were: (i) participants whose depressive symptoms worsened and required different treatment during the trial, making them unfit to continue. (ii) Participants facing severe adverse events or complications during treatment. (iii) Participants choosing not to continue with the treatment.\n\nIn the electroacupuncture group (EG), participants received acupuncture at 10 scalp points (Baihui [GV20], Ex-hn 1, 3 points of intelligence [Shenting [GV24], Benshen [GB13] on both sides, Touwei [ST8] on both sides). Electroacupuncture was applied for 20 minutes at Benshen [GB13] and Touwei [ST8] on both sides. They also received acupuncture at Tai chong [LV3], Tai Yuan [LU9], and Tai Xi [KI3] on both sides, as shown in Figure 1. Acupuncture sessions were weekly for 10 weeks.\n\nIn the sham acupuncture control group (CG), participants received acupress or a brief needle insertion at He gu on both hands during the first and 10th weeks, marking the endpoint of the study.\n\nThe primary outcome measured specific cognitive functions including the TMT-B, SCWT, ADAS-Cog; category delayed recall; and subjective cognitive, and quality of life complaints from the WHODAS 2.0; sections D1.1-1.6 and H1-3, which is a questionnaire asking about the number of days in the past 30 days when participants experienced difficulties with attention, understanding, and initiating conversation. All of these outcomes were assessed by trained blinded psychologists. The scores from the test at week 10 (T10), which serves as the primary endpoint, were compared to the scores from the test at week 1 (T1) for each participant. Then, the median value of the within-group change scores was calculated. Secondary outcome measured depressive symptoms using the Thai-PHQ-9, which were self-rated by the participants. All treatment outcomes were assessed at baseline (T0) and after the 10-week intervention (T10).\n\nThe data was presented as mean ± standard deviation and median, and analyzed using SPSS 22.0 software.22 A comparison between the electroacupuncture and control groups was conducted using the Mann-Whitney U test, which compared the median differences of the outcomes. Differences between groups were calculated by comparing baseline and endpoint values. A significance level of p < 0.05 was used to determine statistical significance. To compare all outcomes for all participants who were randomized to electroacupuncture and control groups, an intention-to-treat (ITT) analysis was performed with the last observation carried forward technique (LOCF). A per-protocol (PP) analysis was also performed for comparison among participants who completed the treatment.\n\n\nResults\n\nThe study spanned from December 2021 to December 2022. Initially, 60 individuals with MDD were enlisted for the trial. However, 8 participants, including 3 participants from the acupuncture group and 5 participants from the control group, discontinued their involvement during this period due to loss of follow-up. Additionally, 1 participant in the electroacupuncture group withdrew due to a neurosyphilis diagnosis. Consequently, 60 participants were eligible to receive ITT analysis, while 51 patients formed the complete analysis group using PP analysis (Figure 2).\n\nIn terms of general characteristics (such as gender, age, education, underlying medical conditions, occupation, and severity of illness) of EG and CG were similar, the outcomes were analyzed with unadjusted estimates. Both sample groups are predominantly female, with an average education duration of approximately 15 years. The average age in the experimental group is 28.77 ± 9.16 years, while the average age of the control group is 27.57 ± 8.08 years. Most participants in both the experimental and control groups are employed and do not have pre-existing illnesses. In terms of clinical data, it was found that participants in both the experimental and control groups had similar durations of experiencing depressive symptoms (experimental group 3.3 ± 1.87 years, control group 3.7 ± 2.67 years). The Thai-PHQ-9 scores indicated a similar severity of depressive symptoms in both groups (experimental group 14.53 ± 4.96, control group15 ± 4.15). Additionally, the average scores of WHODAS 2.0 D1.1-1.6, representing the level of severity in attention and memory-related life impairments, were similar between the two groups (Table 1).\n\nMoreover, most participants used medication, primarily SSRIs, and sometimes benzodiazepines in a similar manner (see Table 2).\n\nThe assessment of executive function, memory, and subjective cognitive complaints at the baseline (T1) and endpoint (T10) of the study (Table 3), which was conducted at the 10th week for both the electroacupuncture treatment group and the sham acupuncture control group, yielded the following results:\n\nIn the electroacupuncture treatment group (Table 4), EG, executive function was primarily evaluated using the TMT- B. The average score of the TMT-B at T1 was 131.67 and at T10 was 119.87, indicating a trend of decreasing scores after receiving electroacupuncture. Subcomponents of executive function, such as inhibition capacity, were assessed using neuropsychological battery scores from the Stroop Word Test, Stroop Color Test, Stroop Word-Color Test, and memory scores from the ADAS-Cog Delayed Recall Memory test. The average scores for these subcomponents showed a tendency to improve after receiving electroacupuncture. Additionally, subjective cognitive complaints were evaluated using the Thai version of the WHODAS 2.0: D1.1-1.6. It was observed that there was a reduction in scores, with the average WHODAS 2.0: D1.1-1.6 score at T1 being 17.73 and at T10 being 13.43, suggesting a decrease in cognitive complaints. Furthermore, the secondary outcome, which measured the severity of depressive symptoms using the Thai-PHQ-9, also showed a similar trend of reduction.\n\nIn the sham acupuncture control group, CG, the assessment of executive function using the TMT-B resulted in an average score of 146.60 at T1 and 120.20 at T10, indicating a trend of decreasing scores after receiving sham acupuncture. Similar to the electroacupuncture group, subcomponent assessments of executive function and memory showed a tendency to increase after receiving sham acupuncture. Subjective cognitive complaints, evaluated using the WHODAS 2.0: D1.1-1.6, also showed a decrease in scores, with the average WHODAS 2.0: D1.1-1.6 score at T1 being 16.93 and at T10 being 15.47, indicating a reduction in cognitive complaints. The secondary outcome, which measured the severity of depressive symptoms using the Thai-PHQ-9, showed a trend of reduction as well. The average Thai-PHQ-9 score at T1 was 15.00, and at T10 it was 13.10.\n\nThe difference in median scores between week 10 and week 1 in each group, measured using the TMT-B (which assesses flexibility and processing speed in terms of time in seconds), ADAS-cog delayed recall memory (used to evaluate cognitive function in memory), and Stroop Test (which has 3 subparts: Stroop Word Test, Stroop Color Test, and Stroop Word and Color; with Stroop Word and Color representing inhibition capacity), Thai-PHQ-9 (for measure depressive symptom) showed a trend of higher scores in the electroacupuncture group compared to the control group. However, when subjected to the Mann-Whitney U test, no statistically significant difference was found.\n\nThe analysis revealed that in the electroacupuncture group, the median within-group change score (∆WHODAS 2.0: D1.1-1.6 of T10-T1) was -5.50, indicating a significant improvement in difficulties with attention, understanding, and initiating conversation. On the other hand, in the control group, the median within-group change score (∆WHODAS 2.0: D1.1-1.6 of T10-T1) showed no change at 0 points. The Mann-Whitney U test was performed with a value of 318.00 and a corresponding p-value of 0.049, indicating a statistically significant difference between the experimental and control groups. This suggests that the experimental group experienced a significant decrease in difficulties with attention, understanding, and initiating conversation compared to the control group. No severe side effects were observed from the acupuncture treatment, such as nerve injury, internal organ injury, infection, allergies, or adverse reactions.\n\n\nDiscussion\n\nFindings from the study indicate that electrical stimulation acupuncture can significantly reduce subjective cognitive complaints in individuals with mild to moderate depressive symptoms and cognitive issues, when compared to the control group. These complaints encompass difficulties in activities like concentration, thinking, analysis, learning, understanding, and initiating conversations over the past 30 days.\n\nMoreover, positive trends were observed in the experimental group, showcasing enhancements in executive function, memory, and a reduction in the frequency and severity of cognitive issues and depressive symptoms. These outcomes might partly arise from the placebo response, where individuals in the control group experience health improvements despite receiving a simulated treatment. This phenomenon is influenced by participants’ perception of the experiment, including their emotions, expectations, and the therapeutic environment. Interestingly, placebo effects can also be seen in the active treatment group.\n\nA meta-analysis by Hafliðadóttir, S.H., and colleagues spanning from 1966 to 2008 covering randomized controlled trials, highlighted that contextual effects were notably higher when the outcome assessor was blinded and the allocation was concealed. Such effects were more pronounced in younger patients and with a higher proportion of female participants.23 In our study, the authors were mindful of the potential for placebo responses and took measures to minimize them, both for participants and assessors.\n\nDespite the predominance of female participants in our study, both groups displayed improvements. Notably, the experimental group exhibited significantly greater enhancements in subjective cognitive complaints. This could be attributed to the experimental group receiving more frequent acupuncture interventions, resulting in a more pronounced contextual effect (around 8.53 times) compared to the control group (approximately 1.83 times)\n\nAcupuncture, compared to non-invasive brain stimulation, stands out as a cost-effective, easily accessible treatment with minimal side effects. It brings about an enhanced quality of life for participants, evident from reduced subjective cognitive complaints in both experimental and control groups. This reduction underscores acupuncture’s positive influence.\n\nAs participants experience relief in subjective cognitive complaints, their apprehensions about resuming daily activities diminish. Worries regarding work capacity decrease, leading to enhanced work efficiency through reduced anxiety and an improved overall quality of life. Positive developments were also noted in other aspects, such as executive function, memory, motor speed, and inhibition. Moreover, acupuncture shows potential to lower the recurrence rate of depressive symptoms.\n\nPrevious research has shown that acupuncture improves cognitive function. For example, Jinyu Du, et al. studied the clinical effect of scalp acupuncture combined with cognitive training on cognitive impairment after cerebral injury, comparing it to receiving routine treatment and cognitive rehabilitation training for 12 weeks. They found that the scores of cognitive function assessment in both groups were significantly higher than before treatment. Additionally, in the three cognitive sub-domains—orientation, visual motor organization, and thinking operation—the scores of the treatment group were significantly higher than those of the control group.8\n\nIn another study by Zuo-Li Sun, et al., focusing on the efficacy of electroacupuncture (EA) in patients with schizophrenia, it was found that EA significantly improved memory and moderately improved executive functions and problem-solving. However, it did not demonstrate significant improvement in the severity of psychiatric symptoms or BDNF levels between the control and experimental groups.9\n\nYujie Jia, et al. conducted a study on the efficacy of acupuncture with manual needle stimulation in patients with mild to moderate Alzheimer’s disease. They found that the group receiving acupuncture with manual needle stimulation showed significant benefits in cognitive functions measured by the ADAS-cog score compared to the group receiving only Donepezil. However, there was no significant difference in activities of daily living between the two patient groups.24\n\nNotably, successful treatments in those studies involved regular acupuncture on alternate days. Literature review by the researchers spotlighted the dose-dependent effect of acupuncture treatment, categorized as low and high dose. High dose acupuncture exhibits the following attributes: 1) using over 9 needles, 2) administering treatment more than twice weekly, 3) generating a “De qi” sensation with at least 8 needle insertions, and 4) displaying strong positive correlation with treatment outcomes.\n\nPrior high dose acupuncture studies involved daily treatment for at least 2 weeks, more than 10 needle insertions, and over 9 needles. In contrast, this study employed once-weekly treatment, over 9 needle insertions (at least 6 lacking the “De qi” sensation at each insertion), not qualifying as high dose acupuncture. Limited participant numbers (n=30) due to time and resource constraints might have contributed to statistically insignificant findings.\n\nFurthermore, no previous research focused on acupuncture for depressive disorder patients with cognitive dysfunction, limiting direct comparison of acupuncture techniques.\n\nDepression patients often grapple with cognitive problems, attention, and memory issues. These difficulties often persist even after depressive symptoms alleviate, increasing the risk of mild cognitive impairment (MCI) or Alzheimer’s disease. Acknowledging, diagnosing, and continually monitoring subjective cognitive decline in depressed patients are crucial for proper treatment and follow-up.\n\nAddressing these concerns, the researchers conducted a pilot study suggesting that electroacupuncture may enhance cognitive function more than the control group. These findings pave the way for further research, both clinically and potentially in clinics. Extended studies are proposed, involving non-invasive transcranial stimulation or electroacupuncture with higher doses, frequency, and duration. Additionally, specific acupuncture points correlating with brain function and a larger participant pool could yield clearer effect sizes and concrete evidence, fostering novel knowledge and pioneering treatments for improved mental health services in the future.\n\nThis present study was carried out within a specific timeframe, from December 24, 2021, to December 23, 2022, and focused solely on depression patients seeking treatment at Thammasat University Hospital. Consequently, participant numbers were limited, and the study faced constraints due to outpatient treatment frequency. Participants were unable to undergo acupuncture more than once weekly, categorizing the treatment dose as low. As a result, the study’s outcomes might not distinctly exhibit differences.\n\n\nEthics and consent\n\nThe Human Research Ethics Committee of Thammasat University (Medicine) has granted approval for this medical research study (MTU-EC-PS-1-304/64) with an approval date from December 24, 2021 to December 23, 2022. It is in full compliance with international guidelines such as the Declaration of Helsinki. Each participant was provided with detailed information regarding the study and informed written consent was obtained for their participation in the research. Subsequently, all participants granted consent for publication.", "appendix": "Data availability\n\nZenodo: Effects of Electroacupuncture on Cognitive Symptoms in Major Depressive Disorder: A Pilot Study and Randomized Controlled Trial, https://doi.org/10.5281/zenodo.10488186. 25\n\nZenodo: Checklist for Effects of Electroacupuncture on Cognitive Symptoms in Major Depressive Disorder: A Pilot Study and Randomized Controlled Trial, https://doi.org/10.5281/zenodo.10816401. 26\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 would like to thank all the staff and colleagues at the psychiatry outpatient clinic at the Thammasat University Hospital, for their support throughout the research period.\n\n\nReferences\n\nWHO: Depressive disorder (depression).2023 August 2023; 31 march 2023.\n\nKoenigs M, Grafman J: The functional neuroanatomy of depression: distinct roles for ventromedial and dorsolateral prefrontal cortex. Behav. Brain Res. 2009; 201(2): 239–243. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLam RW, et al.: Cognitive dysfunction in major depressive disorder: effects on psychosocial functioning and implications for treatment. Can. J. Psychiatr. 2014; 59(12): 649–654. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGreer TL, et al.: Does duloxetine improve cognitive function independently of its antidepressant effect in patients with major depressive disorder and subjective reports of cognitive dysfunction? Depress. Res. Treat. 2014; 2014: 627863. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSackeim HA: Autobiographical Memory and Electroconvulsive Therapy: Do Not Throw Out the Baby. J. ECT. 2014; 30(3): 177–186. Publisher Full Text\n\nMcIntyre RS, Lophaven S, Olsen CK: A randomized, double-blind, placebo-controlled study of vortioxetine on cognitive function in depressed adults. Int. J. Neuropsychopharmacol. 2014; 17(10): 1557–1567. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSemkovska M, McLoughlin DM: Objective cognitive performance associated with electroconvulsive therapy for depression: a systematic review and meta-analysis. Biol. Psychiatry. 2010; 68(6): 568–577. PubMed Abstract \| Publisher Full Text\n\nDu J, et al.: Clinical observation of 60 cases of treating cognitive disorder after cerebral injury in combination with scalp acupuncture and cognitive training. Medicine (Baltimore). 2018; 97(40): e12420. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSun ZL, et al.: Serum brain-derived neurotrophic factor levels associate with cognitive improvement in patients with schizophrenia treated with electroacupuncture. Psychiatry Res. 2016; 244: 370–375. PubMed Abstract \| Publisher Full Text\n\nJia Y, et al.: Acupuncture for patients with mild to moderate Alzheimer’s disease: a randomized controlled trial. BMC Complem. Altern. M. 2017; 17(1): 556. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOrtiz M, et al.: Acupuncture for Depression during Pregnancy: a Randomized Controlled Trial. Deutsche Zeitschrift für Akupunktur. 2010; 53(2): 43–45. Publisher Full Text\n\nKim YR, et al.: Electroacupuncture promotes post-stroke functional recovery via enhancing endogenous neurogenesis in mouse focal cerebral ischemia. PLoS One. 2014; 9(2): e90000. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZheng C, et al.: A randomized controlled trial comparing electroacupuncture with manual acupuncture for motor function recovery after ischemic stroke. Eur. J. Integr. Med. 2018; 22: 76–80. Publisher Full Text\n\nKim JH, et al.: Electroacupuncture acutely improves cerebral blood flow and attenuates moderate ischemic injury via an endothelial mechanism in mice. PLoS One. 2013; 8(2): e56736. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLi J, et al.: Electroacupuncture improves cerebral blood flow and attenuates moderate ischemic injury via Angiotensin II its receptors-mediated mechanism in rats. BMC Complem. Altern. M. 2014; 14: 441. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAmerican Psychiatric Association: Diagnostic and statistical manual of mental disorders: DSM-5. Washington, DC: American Psychiatric Association; 2013; vol. 5. .\n\nAmerican Psychiatric Association: Diagnostic and statistical manual of mental disorders: DSM-IV. Washington, DC: American Psychiatric Association; 1994; vol. 4. .\n\nLotrakul M, Sumrithe S, Saipanish R: Reliability and validity of the Thai version of the PHQ-9. BMC Psychiatry. 2008; 8: 46. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nThawicachat N, Worakul P, Kanchananakintr P: Alzheimer’s disease assessment scale (ADAS) Thai version. J. Thai. Gerontol. Geriatr. Med. 2002; 3: 21–32.\n\nNa Chiangmai N, Wongupparaj P: Dementia screening tests in Thai older adults: a systematic review. J. Mental Health Thailand. 2020; 28(3): 252–265.\n\nÜstün TB: Measuring health and disability: Manual for WHO disability assessment schedule WHODAS 2.0. World Health Organization; 2010.\n\nIBM Corp: IBM SPSS Statistics for Windows, Version 22.0. Armonk, NY: IBM Corp; 2013. Released.\n\nHafliðadóttir SH, et al.: Placebo response and effect in randomized clinical trials: meta-research with focus on contextual effects. Trials. 2021; 22(1): 493. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJia Y, et al.: Acupuncture for patients with mild to moderate Alzheimer’s disease: a randomized controlled trial. BMC Complem. Altern. M. 2017; 17(1): 556. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBoontra Y, Phanasathit M: Effects of Electroacupuncture on Cognitive Symptoms in Major Depressive Disorder: A Pilot Study and Randomized Controlled Trial. [Data]. Zenodo. 2024. Publisher Full Text\n\nBoontra Y, Phanasathit M, Thanetnit C: CONSORT 2010 checklist of the Effects of Electroacupuncture on Cognitive Symptoms in Major Depressive Disorder: A Pilot Study and Randomized Controlled Trial. Zenodo. 2024. Publisher Full Text" }	[ { "id": "282194", "date": "08 Jun 2024", "name": "Rosa N Schnyer", "expertise": [ "Reviewer Expertise Acupuncture", "depression", "pain", "women's 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 set out to conduct a rigorous, double-blind RCT on the effect electro-acupuncture on cognitive symptoms in major depressive disorder. The subject is highly relevant clinically and the study is timely, as it is critical to identify strategies to manage cognitive and mood symptoms which affect a large segment of the population worldwide.\nThere are some key issues that need to be addressed in the reporting of the results, some critical considerations in future proposals on this subject:\nProtocols: Please refer to and incorporate the STRICTA guidelines and include the checklist https://stricta.info/ STRICTA Standards for Reporting Clinical Trials of Acupuncture, which is an official extension of the CONSORT criteria. Description of the interventions, training of the acupuncture providers, etc. is incomplete. For example, there is no description of the needling parameters or the intensity / frequency of the electrostimulation\nDouble-Blind: You state that the EG group received EA at 13 points: 10 scalp and 3 body points-weekly 10 weeks while the CG Control, received Brief needling at LI 4 Hegu for 10 weeks. You state that acupuncturists were also blinded, but given the design of the study, it is unlikely that acupuncturists were indeed blinding, when considering the electro protocol vs the sham protocol. Trained acupuncturists would know the difference in point selection, and it’s unlikely that they would not question shallow needling at Hegu as an effective treatment for cognitive symptoms and / or depression.\nYou also didn’t assess after the study. whether the participants were in fact blinded to treatment, or whether the blinding work for either participants or providers.\nIn the future consider exploring other well proven control for EA, for example by not attempting to blind the acupuncturist and using in the control group a small selection of non-acupuncture points connected to EA for which the wires have been cut but tapped to appear as if the machines work the same as the EG group.\nPopulation description and use of medication (Table 2): One major concern with this study, is the inclusion of patients who are on a variety of psychotropic medications that have effects beyond antidepressant treatment. There doesn’t seem to be explicit consideration for the way in which some of these medications may impact cognitive deficits, and thus confound treatment effects.\nAlthough the idea of using a naturalistic sample that reflects the complex and varied population experiencing comorbid cognitive symptoms and depression, who are on multiple and different medication regimes is closer to the real world practice in clinical setting, the sample is too small to identify a differential response to treatment, which may be confounded by different medication combinations. This is a limitation of the study that needs to be explicitly included in the discussion section.\nIn the future, if conducting a naturalistic trial, you may need a much larger number of patients with these characteristics to detect conclusive differences between groups. Or in a smaller study, limit acupuncture as an augmentation of one specific class of medications only.\nConclusions and Discussion: Your conclusions don’t reflect the results of the analysis; the scores for the CG overall are lower at week 1 and therefore it is possible that this cohort had less severe cognitive decline. Nevertheless, the difference of drop on score of 11.8 in EG vs 18.5 in CG (6.7 seconds) between week 1 and 10 doesn’t seem to be a significant enough difference between EG and CG.\nTherefore it is hard to conclude in the discussion section that “Findings from the study indicate that electrical stimulation acupuncture can significantly reduce subjective cognitive complaints in individuals with mild to moderate depressive symptoms and cognitive issues, when compared to the control group”. The reduction is not significant when compared to CG.\nOther minor:\n\nProvide consistency when describing the trial: it has been conducted, and is not waiting to be conducted (use of pats vs future verb tense)\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? No source data required\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [ { "c_id": "11906", "date": "26 Jun 2024", "name": "Muthita Phanasathit", "role": "Author Response", "response": "Protocols: Please refer to and incorporate the STRICTA guidelines and include the checklist https://stricta.info/ STRICTA Standards for Reporting Clinical Trials of Acupuncture, which is an official extension of the CONSORT criteria. Description of the interventions, training of the acupuncture providers, etc. is incomplete. For example, there is no description of the needling parameters or the intensity / frequency of the electrostimulation - According to the review comment, the STRICTA 2010 checklist was done and attached in the “Data availability” section. Double-Blind: You state that the EG group received EA at 13 points: 10 scalp and 3 body points-weekly 10 weeks while the CG Control, received Brief needling at LI 4 Hegu for 10 weeks. You state that acupuncturists were also blinded, but given the design of the study, it is unlikely that acupuncturists were indeed blinding, when considering the electro protocol vs the sham protocol. Trained acupuncturists would know the difference in point selection, and it’s unlikely that they would not question shallow needling at Hegu as an effective treatment for cognitive symptoms and / or depression. You also didn’t assess after the study. whether the participants were in fact blinded to treatment, or whether the blinding work for either participants or providers. In the future consider exploring other well proven control for EA, for example by not attempting to blind the acupuncturist and using in the control group a small selection of non-acupuncture points connected to EA for which the wires have been cut but tapped to appear as if the machines work the same as the EG group. -According to the review comment about double blind the revision was made as follows; This study was double-blinded RCT. All participants and assessors were unaware of group assignments. at page 5 Line 4 -Thank you for your advice about blinding assessment after the intervention: We agree that assessing the blinding of all participants and providers is beneficial after the experimental phase to reduce bias. However, the authors will incorporate this into future studies for improvement. -Thank you for your advice. about the sham acupuncture The authors believe that sham acupuncture lacks efficacy and, due to safety concerns, aim to minimize injury and unnecessary infection risks from acupuncture needle insertion at non-acupuncture points connected to EA through wires. Population description and use of medication (Table 2): One major concern with this study, is the inclusion of patients who are on a variety of psychotropic medications that have effects beyond antidepressant treatment. There doesn’t seem to be explicit consideration for the way in which some of these medications may impact cognitive deficits, and thus confound treatment effects. Although the idea of using a naturalistic sample that reflects the complex and varied population experiencing comorbid cognitive symptoms and depression, who are on multiple and different medication regimes is closer to the real world practice in clinical setting, the sample is too small to identify a differential response to treatment, which may be confounded by different medication combinations. This is a limitation of the study that needs to be explicitly included in the discussion section. In the future, if conducting a naturalistic trial, you may need a much larger number of patients with these characteristics to detect conclusive differences between groups. Or in a smaller study, limit acupuncture as an augmentation of one specific class of medications only. - According to the review comment, the authors are also concerned about small sample size and confounding factors such as the potential impact of various medications on cognitive deficits. We have included this issue in the study limitations as follows: (page 16 Line 28-41) Furthermore, findings from the present study indicate that electroacupuncture might be able to alleviate subjective cognitive complaints in individuals with mild to moderate depressive symptoms and cognitive issues, compared to the control group. However, the clinical significance of the outcomes of electroacupuncture remains unclear. The authors did not include a healthy control group, focusing instead on patients with MDD who had subjective complaints and were receiving standard pharmacological treatment. The present study aimed to see how these patients responded to adjunctive electroacupuncture. Patients with MDD often receive various psychotropic drugs, such as benzodiazepines and antipsychotics, which can affect cognitive function and serve as confounding factors. Previous research suggested that acupuncture can influence cognitive function, so the study focused on its use as an adjunctive treatment in real-world settings. Future studies may include a much larger number of patients with these characteristics to detect conclusive differences between groups, or in a smaller study, limit acupuncture as an augmentation of one specific class of medications Conclusions and Discussion: Your conclusions don’t reflect the results of the analysis; the scores for the CG overall are lower at week 1 and therefore it is possible that this cohort had less severe cognitive decline. Nevertheless, the difference of drop on score of 11.8 in EG vs 18.5 in CG (6.7 seconds) between week 1 and 10 doesn’t seem to be a significant enough difference between EG and CG. Therefore it is hard to conclude in the discussion section that “Findings from the study indicate that electrical stimulation acupuncture can significantly reduce subjective cognitive complaints in individuals with mild to moderate depressive symptoms and cognitive issues, when compared to the control group”. The reduction is not significant when compared to CG. - According to the review comment, the authors are aware of the interpretation of results, which has limitations in generalizability from statistical significance to clinical significance. Therefore, the authors have addressed this issue in the discussion and limitations sections as follow: (page 14 Line 13) Findings from the study indicate that electroacupuncture can statically significantly reduce subjective cognitive complaints in individuals with mild to moderate depressive symptoms and cognitive issues, when compared to the control group. -The revision was made by employing past tense. Thank you" } ] }, { "id": "282200", "date": "11 Jun 2024", "name": "Su-Xia Li", "expertise": [ "Reviewer Expertise Psychiatry，Depression" ], "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. The treatment plan for the two groups of patients in the Abstract is different from the description in the Methods section. 2. A healthy control group should be included in the study protocol. On the one hand, does the cognitive function of patients with MDD differ from that of healthy controls at baseline. On the other hand, after treatment, whether the cognitive function of patients is restored, and how much differences between healthy controls and patients with depression. These can only be achieved in the presence of a healthy control group. 3. For the electroacupuncture group, the authors needed to describe the entire treatment process in detail. Such as the duration of each treatment, how many days consecutive per week, the specific depth of the acupuncture therapy, and how strong the current intensity is? 4. How does the Sham group work？The authors need to describe in detail the specific operation of the sham acupuncture control group, how long the acupuncture of He gu point lasted for each treatment, and how many consecutive days were in the first week and the 10th week. The specific depth of the acupuncture therapy, and how strong the current intensity is? 5. For the comparison of the two groups of indicators at the end point, the existence of time effect should be considered. It is unreasonable and unscientific to use a simple t-test between two groups. 6. Authors defined that at baseline is T0, at the ten-week intervention is T10. However, in Results section, baseline is described as T1. The author's description of the same event should be uniform.\n\n7. Abbreviations in the table should be explained at the bottom of the table. 8. Many measures of cognitive function did not differ between the two groups after 10 weeks of treatment. Is it because the settings of the sham group are unreasonable?\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": [ { "c_id": "11907", "date": "26 Jun 2024", "name": "Muthita Phanasathit", "role": "Author Response", "response": "- According to the review comment, the revision of abstract was made. Page 2 Line 4-8 in abstract - Thank you for your advice. We agree that a healthy control should be included in experimental studies aiming to assess the safety and efficacy of new interventions and medications. However, a literature review on the use of electroacupuncture has provided evidence of its safety and efficacy in treating patients with schizophrenia, brain injury, and stroke who have cognitive impairment. The researchers aimed to study the effects of electroacupuncture on specific cognitive functions in subjects experiencing cognitive dysfunction associated with MDD. Including healthy controls without subjective cognitive complaints in this study might not address the research question adequately. Therefore, we recruited MDD patients with cognitive dysfunction receiving sham acupuncture and standard treatment as a comparator instead. - According to the review comment, the revision was made in the method / intervention session. (page 4, Line 12-31) - Due to the outcome from this pilot study (n=30 per group), the data had a non-normal distribution. A comparison between the electroacupuncture and control groups was conducted using the Mann-Whitney U test, which compared the median differences of the outcomes. Differences between groups were calculated by comparing baseline (T1) and endpoint (T10) values. Page 6 Line 16 -According to the review comment about the Base line , the revision was made as T1.(Page 6 line 11) - All abbreviations in the table were explained at the bottom of all tables. (table 1 and table 3 ) - According to Sham setting : The authors hypothesized that the reason many measures of cognitive function did not differ between the two groups after 10 weeks of treatment was due to two factors: firstly, the baseline subjective cognitive function in the control group were less severe than those in the experimental group; and secondly, the ceiling effect, as the baseline scores of executive function, subjective cognitive complaints, and quality of life were relatively high in both groups. These points have been addressed in the discussion section." } ] } ]	1	https://f1000research.com/articles/13-479
https://f1000research.com/articles/13-712/v1	28 Jun 24	{ "type": "Research Article", "title": "Effect of Informative Cesarean Delivery Operative Steps Video on Maternal Anxiety Level: A Randomized Controlled Trial", "authors": [ "Panicha Phetprapasri", "Athita Chanthasenanont", "Pichita Prasongvei", "Winitra Nuallaong", "Suphachai Chaitrakulthong", "Densak Pongrojpaw", "Panicha Phetprapasri", "Pichita Prasongvei", "Winitra Nuallaong", "Suphachai Chaitrakulthong", "Densak Pongrojpaw" ], "abstract": "Background Cesarean section is the most common obstetric procedure performed. This can lead to maternal anxiety, which is a significant contributor to postpartum depression. This can adversely affect pregnant women both mentally and emotionally, negatively impacting their well-being and family bonding. The aim of this study was to evaluate the effect of the addition of an informative cesarean section operative steps video on the maternal anxiety score compared with standard pre-cesarean section care.\n\nMethods This randomized controlled trial was conducted at Thammasat University Hospital, Thailand, between April and September 2023. Pregnant women who underwent their first cesarean section were allocated to two groups: intervention and control groups. Participants in the intervention group were required to watch a 5-minute informative video that elaborately described the process from pre-operative steps until post-operative care on the day scheduled for cesarean delivery. All participants received the same routine pre-operative and post-operative care. The State-Trait Anxiety Inventory (STAI) was used to measure both populations on two occasions: the day of scheduled cesarean delivery and postpartum day 1.\n\nResults A total of 178 women were recruited. The demographic and obstetric characteristics were similar between the two groups. The pre-operative STAI scores of the intervention and control groups were 42.9 and 44.1 points, respectively, with no significant difference. However, the post-operative anxiety score showed a significant decline in the intervention group compared to that in the control group (p = 0.002). Moreover, most of the participants in the intervention group showed a low level of anxiety after the operation, while half of the control group remained at a moderate to high level score.\n\nConclusions The provision of an informative educational video before cesarean delivery is a powerful tool that significantly reduces cesarean operative anxiety and improves health outcomes. Thai Clinical Trials Registry on the 28 March 2023 (TCTR20230328001).", "keywords": [ "Cesarean section", "Anxiety", "Informative video." ], "content": "Introduction\n\nCesarean section is the most common obstetric surgical procedure. The rate of cesarean deliveries increased globally by 19.4 percent in 2018. By 2030, the global cesarean section rate is projected to increase by 30 percent. This is equivalent to 38 million cesarean deliveries according to accumulated data.1\n\nIn Thailand, the rate of cesarean delivery has shown a continuous upward trend, increasing from 25 percent to 30 percent over the last 15 years, and reaching 30 percent to 50 percent in 2023.2–4 Maternal anxiety significantly contributes to postpartum depression. It is a common and serious issue that affects pregnant women, both mentally and emotionally. This can adversely affect one’s well-being and family bonding. Furthermore, cesarean delivery significantly increases the risk of postpartum depression.5 Postpartum depression and anxiety can be influenced by various factors, such as maternal stress, anxiety during pregnancy, stressful events during or after childbirth, traumatic birth experiences, preterm birth, neonatal intensive care, low level of social support, previous history of depression, and breastfeeding problems. Screening for postpartum depression and anxiety during pregnancy using standardized assessment tools is recommended to efficiently identify and support at-risk women.6\n\nA previous study found that the use of anesthesia-informative video decreased the level of anxiety and improved the satisfaction level of cesarean section.7 However, some reports have reported contradictory results.8,9 Mirenberg reported in 2022 that the usage of informative cesarean operative steps video, which included preparations before entering the operation room to the recovery process, showed a reduction in maternal anxiety.10 The aim of this study was to investigate whether the provision of an informative cesarean section video prior to surgery would lower maternal anxiety. Furthermore, it aimed to assess the prevalence of anxiety in pregnant women undergoing cesarean delivery and the satisfactory level of the informative educational video.\n\n\nMethods\n\nThis randomized controlled trial was approved by the Human Ethics Committee of Thammasat University on the 21 March 2023 (MTU-EC-OB-1-010/66) and Thai Clinical Trials Registry on the 28 March 2023 (TCTR20230328001). The study was conducted at the antenatal care clinic and postpartum wards at Thammasat University Hospital, Pathum Thani, Thailand. The participants were Thai singleton pregnant women aged between 18 and 45 years who were scheduled for cesarean delivery between April and September 2023. The exclusion criteria were previous cesarean delivery, pre-existing chronic illnesses before pregnancy, mental health disorders, fetal complications during pregnancy, fetal congenital anomalies, severe early neonatal illnesses, pregnancy-related complications that could not be managed by medications, emergency surgery conditions, and refusal to participate in this study.\n\nPatients who met the selection criteria were reviewed before they were asked to provide written informed consent approved by the ethical approval committee to participate in the project. The team provided guidance and detailed explanations of the research to the participants on the day scheduled for cesarean delivery (1–2 weeks before the operative day) at the antenatal care clinic. They were randomized and allocated to two groups: control and intervention groups. All participants received routine pre- and post-cesarean care. Pre-operative care included explanation of the surgical procedure and possible complications. Participants in the intervention group were required to watch a 5-minute informative video designed and used in this study on the day scheduled for cesarean delivery. The video elaborately describes the pre-operative steps, anesthesia, cesarean delivery process, and post-operative care.\n\nBefore randomization, all participants were required to complete the demographic data and State-Trait Anxiety Inventory (STAI form Y-1).11 The STAI form Y-1 questionnaire is on state anxiety and consists of 20 items evaluating four levels of intensity: not at all, sometimes, moderately, and very much. The questionnaire comprised 10 positively framed statements (Items 3, 4, 6, 7, 9, 12, 13, 14, 17, and 18), with scores ranging from 1 to 4. It also contained 10 negatively framed statements (Items 1, 2, 5, 8, 10, 11, 15, 16, 19, and 20) with scores ranging from 4 to 1. The total score ranged from 20 to 80 points, with a higher total score indicating a higher level of anxiety. The score interpretation was divided into three ranges of 20-39, 40-59 and 60-80 which can be interpreted as low, moderate, and high levels of anxiety, respectively. On postpartum day 1, all participants were reevaluated using the STAI from Y-1. If a research participant experiences a high level of anxiety before or after cesarean delivery, advice on meeting up with individual psychotherapy will be recommended, or a counseling team will offer assistance to help ease the participant’s anxiety. The intervention group was evaluated to measure the level of patient satisfaction with the informative videos on postpartum day 1. Statistical analysis was performed using Statistical Package for the Social Sciences version 26 (SPSS Inc., Chicago, USA). Continuous data were computed using an unpaired t-test and are presented as means with standard deviation. Categorical data were evaluated using the chi-square test and represented as numbers. Statistical significance was set than 0.05.\n\n\nResults\n\nOne hundred and seventy-eight participants scheduled for first-time cesarean delivery were recruited and equally randomized into two groups, with eight cases that dropped out, as shown in Figure 1. Maternal demographic and obstetric data between the two groups were statistically indistinguishable, as seen in Table 1. Most of the participants were primigravida, had an educational level of at least a bachelor’s degree, and did not have any underlying disease or previous abdominal surgery. No differences were found in the indications for cesarean section and incidence of postpartum hemorrhage in either group. There were no cases of serious post-operative complications such as re-exploration or neonatal intensive care unit admission. The information sources of cesarean delivery in both groups were similar, and most of the participants received information about cesarean delivery from the Internet and friends before joining the research, as shown in Figure 2.\n\nControl: standard pre-cesarean section operative care; Video: informative cesarean section operative steps video before cesarean section operation; STAI: State-Trait Anxiety Inventory.\n\nQR code for informative cesarean section operative steps video.\n\n* : mean ± standard deviation (SD).\n\n** : n (%); Control: standard pre-cesarean operation care; Video: informative cesarean section steps video before operation; BMI: body mass index; TR: Tubal sterilization operation.\n\nControl: standard pre-cesarean operation care; Video: informative cesarean section steps video before operation; Book: The book that provides information on preparing for cesarean delivery; Family: A group of people who are related by blood, marriage or adoption such as parents; Medical professionals: person individuals trained and qualified to provide medical care such as doctors, nurses and other healthcare providers; Friends: person whom one has a bound of mutual affection who not a member of family; Internet: internet information providing guidance on preparing for cesarean delivery.\n\nBefore cesarean delivery, the STAI scores of the intervention and control groups did not differ significantly. The prevalence of participants who had moderate to high levels of anxiety in this study was 71 percent, eight of whom had high STAI scores.\n\nPost-operatively, the STAI score was statistically significant in both groups. However, upon juxtaposing the post-operative scores between the two groups, it became evident that the cohort exposed to the intervention group had a statistically noteworthy decrease in scores compared to the control group (decreasing mean STAI score were 7.0 and 4.4 in the intervention and control groups, respectively, p=0.002), as shown in Figure 3. Table 2 shows that the control group exhibited a pre- operative moderate-to-high anxiety level of 77 percent, which was reduced to 53 percent post-operatively. The intervention group showed that the proportion of patients with moderate-to-high anxiety levels decreased from 65 percent in the pre-operative period to 18% in the post-operative period.\n\nSTAI: The State-Trait Anxiety Inventory; Control: standard pre-cesarean operation care; Video: informative cesarean section steps video before operation; Before: anxiety score before cesarean section; After: anxiety score after cesarean section.\n\nSeventy-eight percent of intervention patients transitioned from moderate to high anxiety pre-operatively to low anxiety post-operatively, doubling the control group's 34 percent improvement. Furthermore, two patients in the control group showed escalation from moderate to high anxiety levels, whereas in the intervention group, patients with high anxiety levels pre-operatively had reduced anxiety levels. Notably, all patients with pre- and post-operative high anxiety received psychological counseling. In the intervention group, almost all individuals reported the highest satisfaction scores for the informative cesarean section operative steps video, reaching approximately 87 percent.\n\nAccording to the evaluation of factors affecting pre- and post-operative anxiety scores, pre-operative cesarean section anxiety was not affected. While two factors, watching the informative operative steps video and pre-operative anxiety scores, had a statistically significant impact on the post-operative anxiety score, the odds ratios were 0.17 and 8.23, respectively. as shown in Table 3. Watching the informative video had potentially reduced stress by 83%, and experiencing anxiety before surgery increased the risk of post-operative anxiety stress by 8.23 times.\n\n\nDiscussion\n\nMaternal anxiety is prevalent throughout pregnancy, childbirth, and postpartum, with reported rates ranging from 15 percent to 36 percent.12–15 Some studies suggest an increase in self-reported anxiety symptoms with advancing gestational age, peaking in the third trimester, and even exceeding postpartum levels.15 Interestingly, our study found that in planned cesarean section, patients exhibited significantly higher anxiety levels (71 percent) compared to their counterparts without planned birth interventions during the third trimester.15 This suggests a potential correlation between cesarean anticipation and heightened anxiety.\n\nCesarean section is the most common procedure in obstetrics and has been proven to save numerous lives of both mothers and infants during pregnancy complications. In some cases, parents may request this delivery method. There are various ways to access information regarding cesarean deliveries. However, despite its prevalence and readily available information, patients undergoing planned cesarean delivery still exhibit higher anxiety levels. Therefore, the ACOG strongly recommends assessing anxiety risks and conditions in pregnant and postpartum women for early detection and intervention.6 Interventions for patients with anxiety can include both pharmacological and non-pharmacological methods, such as individual or group psychotherapy, music therapy, massage, meditation and hypnotherapy.16–18\n\nCurrent recommendations suggest implementing interventions to reduce anxiety before or near the time of cesarean section.19 While some studies using informative videos about spinal block have not shown significant anxiety reduction,7–9,20,21 research utilizing videos specifically about the cesarean operation process itself demonstrated a noteworthy decrease.10 This aligns with our study's finding of reduced post-operative anxiety levels. Interestingly, both the intervention and control groups showed lower anxiety levels post-operatively. This may be attributed to the successful completion of surgery and the absence of complications, leading to a reduction in natural anxiety in both groups. However, a noteworthy detail emerged: the intervention group displayed a higher reduction in the STAI score post-operatively compared to the control group. Furthermore, the results revealed a shift within the intervention group, with 78 percent of patients initially experiencing moderate-to-high anxiety transitioning to a low anxiety level post-operatively, compared to only 34 percent in the control group. This substantial improvement reflects a greater efficacy of the intervention in mitigating presurgical anxiety compared to the control group. Notably, the overall prevalence of moderate anxiety in the intervention group post-operatively was 15 percent, which is consistent with the study of cesarean operative anxiety levels observed in Cindy-Lee's study involving unplanned cesarean operations and natural births. In stark contrast, the control group exhibited a significantly higher prevalence of moderate-to-high anxiety post-operatively, reaching 68 percent.15 All pre-operative intervention patients who received psychological counseling showed no remaining cases of high anxiety level post-surgery, suggesting the effectiveness of counseling in reducing anxiety. However, both groups saw 10 percent (five cases) in the low anxiety group to moderate anxiety. Unfortunately, due to the limited sample size, a definitive analysis of the causes of this shift was not performed.\n\nPrevious research has identified several factors that significantly influence antenatal anxiety levels, including low to middle income, low educational attainment, a history of abortions, and the responsibility of caring for other children. Additionally, postnatal concerns, such as breastfeeding difficulties, wound pain, and infant care have been linked to post-operative anxiety.6,13 Interestingly, our study found that none of the previously identified factors, even a history of abortions, significantly impacted pre-operative anxiety. Instead, watching informative videos before surgery demonstrably reduced anxiety levels post-operatively, consistent with findings from many studies.7,10,22\n\nHowever, a limitation of our study is that it solely assessed anxiety on postpartum day 1 by using the STAI.11 This means that factors such as breastfeeding difficulties, wound pain, and infant care challenges, which often emerge later in the postnatal period, were not considered. Consequently, future research incorporating these additional elements throughout the pre- and postnatal phases could offer a more comprehensive understanding of CS-related anxiety. By comprehensively studying these broader factors, we can develop more effective intervention methods and tools to reduce anxiety during pregnancy and motherhood.\n\nThe findings from this research show that pre-operative video education has emerged as a powerful tool for reducing cesarean operative anxiety, offering promise as a safe, cost-effective, and widely accessible intervention. These findings suggest that informative videos can be disseminated beyond hospital walls, offering benefits to patients who might not otherwise have access to accurate and reliable information. The ability to disseminate and empower patients with clear and accurate pre-operative information regardless of their source of prior knowledge is crucial. In particular, when considering the varied sources from which patients could acquire information pre-operatively, including personal experiences, hearsay, and academic resources. This study highlights the need for comprehensive and reliable pre-operative information to ensure that patients feel empowered throughout their surgical journey.\n\nFurthermore, leveraging technology and digital platforms could expand the reach of educational resources, ensuring that accurate and comprehensive information is easily accessible to a wider audience, creating easily digestible and reliable content that can empower pregnant patients and enable them to make informed decisions and better prepare for surgical procedures. This patient-centered approach not only enhances care and satisfaction but also contributes to improved health outcomes.\n\n\nConclusion\n\nPre-operative educational videos have emerged as a powerful tool that significantly reduces cesarean operative anxiety and holds promise for improved health outcomes. This readily implementable intervention empowers women with clear and reliable information, transcends hospital walls, and promotes informed decision making. Further research should explore its long-term impact and applicability to other surgical contexts to implement positive surgical experiences. Investing in this approach will not only equip mothers with confidence but also lay the groundwork for a healthier future for mothers and their families.\n\n\nEthics and consent\n\nThis randomized controlled trial was approved by the Human Ethics Committee of Thammasat University on the 21 March 2023 (MTU-EC-OB-1-010/66) and Thai Clinical Trials Registry on the 28 March 2023 (TCTR20230328001). Patients who met the selection criteria were reviewed before they were asked to provide written informed consent approved by the ethical approval committee to participate in the project.", "appendix": "Data availability\n\nZenodo: Effect of informative cesarean delivery operative steps video to maternal anxiety level: a randomized controlled trial, https://zenodo.org/records/11372557. 23\n\nThis project contains following datasets:\n\n1. Effect of Informative Cesarean Delivery Operative Steps Video to Maternal Anxiety Level.xlsx\n\n2. information cesarean section (english sub).mp4\n\n3. Questionnaire.docx\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nZenodo: Effect of informative cesarean delivery operative steps video to maternal anxiety level: a randomized controlled trial, https://zenodo.org/records/11372557. 23\n\nThis project contains following datasets:\n\n1. CONSORT-2010-Checklist.doc\n\n2. 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\nAcknowledgement\n\nThe authors thank the Clin. Prof. Dittakarn Boriboonhirunsarn kindly helped with the statistical analysis.\n\n\nReferences\n\nBetran AP, Ye J, Moller AB, et al.: Trends and projections of caesarean section rates: global and regional estimates. BMJ Glob. Health. 2021; 6: e005671. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRoyal Thai College of Obstetricians and Gynaecologists: Position statement for cesarean section (Revised version 2023).2023 [cited 2022 NOV 15]. Reference Source\n\nPhadungkiatwattana P, Tongsakul N: Analyzing the impact of private service on the cesarean section rate in public hospital Thailand. Arch. Gynecol. Obstet. 2011; 284: 1375–1379. PubMed Abstract \| Publisher Full Text\n\nChanthasenanont A, Pongrojpaw D, Nanthakomon T, et al.: Indications for cesarean section at Thammasat University Hospital. J. Med. Assoc. Thail. 2007; 90: 1733–1737. PubMed Abstract\n\nXu H, Ding Y, Ma Y, et al.: Cesarean section and risk of postpartum depression: A meta-analysis. J. Psychosom. Res. 2017; 97: 118–126. PubMed Abstract \| Publisher Full Text\n\nACOG Committee Opinion No. 757: Screening for Perinatal Depression. Obstet. Gynecol. 2018; 132: e208–e212. PubMed Abstract \| Publisher Full Text\n\nChe YJ, Gao YL, Jing J, et al.: Effects of an Informational Video About Anesthesia on Pre- and Post-Elective Cesarean Section Anxiety and Recovery: A Randomized Controlled Trial. Med. Sci. Monit. 2020; 26: e920428. Publisher Full Text\n\nEley VA, Searles T, Donovan K, et al.: Effect of an anaesthesia information video on preoperative maternal anxiety and postoperative satisfaction in elective caesarean section: a prospective randomised trial. Anaesth. Intensive Care. 2013; 41: 774–781. PubMed Abstract \| Publisher Full Text\n\nKanyeki T, Mung’ayi V, Bal R, et al.: Effect of video-based information on preoperative State trait anxiety inventory scores in adult patients presenting for elective caesarean section: a randomized controlled trial. Afr. Health Sci. 2022; 22: 117–124. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMiremberg H, Yirmiya K, Vinter D, et al.: An informative video before planned cesarean delivery and maternal anxiety—a multicenter randomized controlled trial. Am. J. Obstet. Gynecol. MFM. 2022; 4: 100604. PubMed Abstract \| Publisher Full Text\n\nSpielberger CD, Gorsuch RL, Lushene RE: State-Trait Anxiety Inventory.[cited 2022 NOV 15]. Reference Source\n\nVan der Zee-van den Berg AI, Boere-Boonekamp MM, Groothuis-Oudshoorn CGM, et al.: Postpartum depression and anxiety: a community-based study on risk factors before, during and after pregnancy. J. Affect. Disord. 2021; 286: 158–165. PubMed Abstract \| Publisher Full Text\n\nCena L, Mirabella F, Palumbo G, et al.: Prevalence of maternal antenatal anxiety and its association with demographic and socioeconomic factors: A multicentre study in Italy. Eur. Psychiatry. 2020; 63: e84. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVal A, Míguez MC: Prevalence of Antenatal Anxiety in European Women: A Literature Review. Int. J. Environ. Res. Public Health. 2023; 20: 1098. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDennis CL, Falah-Hassani K, Shiri R: Prevalence of antenatal and postnatal anxiety: systematic review and meta-analysis. Br. J. Psychiatry. 2017; 210: 315–323. PubMed Abstract \| Publisher Full Text\n\nWeingarten SJ, Levy AT, Berghella V: The effect of music on anxiety in women undergoing cesarean delivery: a systematic review and meta-analysis. Am. J. Obstet. Gynecol. MFM. 2021; 3: 100435. PubMed Abstract \| Publisher Full Text\n\nNavaee M, Khayat S, Abed ZG: Effect of pre-cesarean foot reflexology massage on anxiety of primiparous women. J. Complement. Integr. Med. 2020; 17: 17. Publisher Full Text\n\nMarc I, Toureche N, Ernst E, et al.: Mind-body interventions during pregnancy for preventing or treating women’s anxiety. Cochrane Database Syst. Rev. 2011; 2011: CD007559. Publisher Full Text\n\nHepp P, Hagenbeck C, Burghardt B, et al.: Measuring the course of anxiety in women giving birth by caesarean section: a prospective study. BMC Pregnancy Childbirth. 2016; 16: 113. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMaghalian M, Mohammad-Alizadeh-Charandabi S, Ranjbar M, et al.: Informational video on preoperative anxiety and postoperative satisfaction prior to elective cesarean delivery: A systematic review and meta-analysis. BMC. Psychol. 2024; 12: 6. Publisher Full Text\n\nYılmaz G, Akça A, Esen O, et al.: Multimedia education on the day of elective cesarean section increases anxiety scores. Perinat J. 2019; 27: 38–42. Publisher Full Text\n\nBaradwan S, Gari A, Alnoury A, et al.: Informational video impact before caesarean delivery on anxiety and satisfaction: a systematic review and meta-analysis. J. Reprod. Infant Psychol. 2023; 1–15. PubMed Abstract \| Publisher Full Text\n\nPhetprapasri P: Effect of Informative Cesarean Delivery Operative Steps Video to Maternal Anxiety Level: a Randomized Controlled Trial. [Data set]. Zenodo. 2024. Publisher Full Text" }	[ { "id": "311443", "date": "21 Aug 2024", "name": "Valentina Lucia La Rosa", "expertise": [ "Reviewer Expertise Developmental and perinatal psychology" ], "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 “Effect of Informative Cesarean Delivery Operative Steps Video on Maternal Anxiety Level: A Randomized Controlled Trial” provides valuable insights into a practical intervention aimed at reducing maternal anxiety during cesarean delivery. The study is well conducted and presents its findings clearly, but there are several areas where additional detail and discussion are needed to fully contextualize the findings. Below are some comments and suggestions for improving the manuscript:\nMajor Revisions\nIn the Introduction, I recommend expanding the discussion of the effects of cesarean delivery on maternal mental health, particularly highlighting the differences between planned (elective) cesarean sections and emergency cesarean sections. The psychological effects, including anxiety and postpartum depression, may differ significantly between these two types of procedures and should be further explored. Including a more detailed review of relevant studies [ref 1,2 and 3] would strengthen the background of your study. The study exclusion criteria appear to be comprehensive, but may potentially limit generalizability. In particular, the exclusion of women with pre-pregnancy mental disorders may introduce selection bias. These women may represent a significant proportion of those who experience anxiety prior to cesarean section. I recommend expanding the discussion of how this limitation affects the generalizability of the study findings. The STAI was only used to assess anxiety up to postpartum day 1. Anxiety may manifest or change in the later postpartum period due to additional factors such as breastfeeding difficulties or infant care. Consider discussing the rationale for this limited follow-up period and recognize the need for longer-term assessment of maternal anxiety in future research. The authors should provide more detailed information on the psychometric properties of the State-Trait Anxiety Inventory. Specifically, it would be beneficial to include data on its reliability (e.g., Cronbach's alpha) and validity within the study sample. Although the manuscript provides a detailed description of the video, more information is needed about its content development. Was the video content tested for comprehension and appropriateness before being used in this study? It would be helpful to include details on how the video was validated or pre-tested for clarity and accuracy. Although the study enrolled 178 participants, a sample size calculation or power analysis to justify the number of participants is not clearly provided in the text. Please clarify if this was done and include it in the methods section. The discussion is thorough, but lacks a deeper exploration of how the findings are consistent with or contradict existing literature. The authors should expand on how their findings compare to other studies that have examined similar interventions, particularly those with different outcomes. Discussion of possible mechanisms for the observed reductions in anxiety would strengthen the paper's contribution to the field. The manuscript mentions ethical approval, but it would benefit from a more detailed discussion of how informed consent was obtained, particularly with regard to how participants were informed about the randomization process and the potential benefits/risks of watching the video.\n\nMinor Revisions\nThe manuscript is generally clear, but there are a few areas where language could be improved for clarity. For instance, some sentences in the discussion section are overly complex. Consider simplifying them to improve readability.\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": "12396", "date": "16 Sep 2024", "name": "Panicha Phetprapasri", "role": "Author Response", "response": "I have carefully reviewed the feedback and have made the required revisions in the \"Submit New Version\" section of F1000 Research. Thank you for your guidance and consideration." } ] }, { "id": "297337", "date": "13 Dec 2024", "name": "Perapong Inthasorn", "expertise": [ "Reviewer Expertise I am obstetrician and gynaecologist specialized in gynaecologic oncologist." ], "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 randomized controlled trial compared anxiety level in planned cesarean pregnant women between intervention group(informative VDO watching ) and control group. The study showed that STAI score of the intervention and control groups did not differ significantly before cesarean delivery. However the postoperative score had significantly decrease in intervention group compared to control group. I would like the authors to discuss about this topics. 1. Why anxiety did not decrease preoperatively? 2.In intervention group, could the anxiety decrease preoperatively, If the subjects in intervention group watched VDO 1 week before cesarean section ?\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": "12969", "date": "23 Dec 2024", "name": "Panicha Phetprapasri", "role": "Author Response", "response": "I have carefully reviewed the feedback and have made the required revisions in the \"Submit Version 3\" section of F1000 Research. Thank you for your guidance and consideration." } ] } ]	1	https://f1000research.com/articles/13-712
https://f1000research.com/articles/13-882/v1	05 Aug 24	{ "type": "Review", "title": "Enriched marine oil supplements in pregnancy for the modulation of maternal inflammatory- associated causes of preterm delivery", "authors": [ "Pedro Antonio Regidor", "Johanna Eiblwieser", "Theresa Steeb", "Jose Miguel Rizo", "Johanna Eiblwieser", "Theresa Steeb", "Jose Miguel Rizo" ], "abstract": "Preterm birth is a major cause of perinatal complications and neonatal deaths. Furthermore, in the field of obstetrics many clinical entities like uterine contractions or the occurrence of pre- eclampsia remain to be serious complications during pregnancy and represent a major psychological, financial, and economic burden for society. Several published guidelines, studies and recommendations have highlighted the importance of supplementation of omega-3 long chain polyunsaturated fatty acids (PUFAs) during pregnancy. This narrative review aims at giving an overview on the modern perception of inflammatory processes and the role of specialized pro-resolving mediators (SPMs) in their resolution, especially in obstetrics. Additionally, we highlight the possible role of SPMs in the prevention of obstetric complications through oral supplementation using enriched marine oil nutritional’s. The intake of PUFAs may result in an overall improvement of pregnancy outcomes by contributing to fetal brain growth and neurological development but more importantly though modulation of inflammation-associated pathologies. Especially the use of SPMs represents a promising approach for the management of obstetric and perinatal complications. SPMs are monohydroxylates derived from enriched marine oil nutritional’s that involve certain pro-resolutive metabolites of omega-3 long chains PUFAs and may contribute to an attenuation of inflammatory diseases. This may be obtained through various mechanisms necessary for a proper resolution of inflammation such as the termination of neutrophil tissue infiltration, initiation of phagocytosis, downregulation of pro-inflammatory cytokines or tissue regeneration. In this way, acute and chronic inflammatory diseases associated with serious obstetrical complications can be modulated, which might contribute to an improved pregnancy outcome.", "keywords": [ "pregnancy", "specialized pro-resolving mediators", "polyunsaturated fatty acids", "chronic inflammation", "preterm birth", "preeclampsia", "amniotic inflammation", "chorioamnionitis" ], "content": "1. Introduction: The burden of preterm birth\n\nA major cause of more than 85% of all perinatal complications and neonatal deaths is preterm birth, mainly early preterm birth (Ward and Beachy 2003, D’Apremont et al. 2020). Children born prematurely often suffer from a variety of physical and neurodevelopmental disorders, which may cause substantial consequences (Ward and Beachy 2003). Among these are for example neurocognitive complications, which can occur early in childhood and manifest as developmental delay, cerebral palsy, hearing, and visual impairments, learning difficulties and psychiatric disorders (Saigal and Doyle 2008). Moreover, preterm birth imposes significant psychological and financial burdens on parents. With estimated costs of $1.6 billion to $26 billion annually in the United States, preterm birth represents a considerable economic burden and impact on society (Institute of Medicine (US) Committee on Understanding Premature Birth and Assuring Healthy Outcomes 2007, Waitzman et al. 2021, Newnham et al. 2022). Nevertheless, factors like the country, population size, and preterm birth rate have a strong influence on the range of costs (Institute of Medicine (US) Committee on Understanding Premature Birth and Assuring Healthy Outcomes 2007, Waitzman et al. 2021, Newnham et al. 2022). Although a large portion of these costs are attributable to neonatal intensive care for infants born very prematurely, further additional costs have been shown to be associated with prematurity. These costs extend beyond the initial hospitalization, even for children born only a few weeks prematurely. However, two thirds of preterm births occur without known biological causes. Hence, strategies for prevention are urgently warranted and still a subject of intense debate (Ferrero et al. 2016, Goodfellow et al. 2021).\n\n\n2. Supplementation with polyunsaturated fatty acids (PUFAs) to prevent preterm birth\n\nNotably, maternal supplementation with omega-3 long chain polyunsaturated fatty acids (long chain PUFAs) might be one of the most promising interventions to prevent preterm (<37 weeks gestation) and early preterm (<34 weeks gestation) birth (Middleton et al. 2018, Makrides et al. 2019, Carlson et al. 2021). Therefore, an assessment of the need for omega-3 long chain PUFAs during pregnancy and their specific role in reducing the risk of preterm birth is needed. Most guidelines and dietary advice suggest screening and adjust dietary habits to achieve an adequate supply of omega-3 long chain PUFAS, particularly docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) (Koletzko et al. 2007, 2008). Recommendations include that pregnant and lactating women should aim to achieve a dietary intake of omega 3 PUFA that supplies a DHA intake of at least 200 mg/day and that women of childbearing age can meet the recommended intake of DHA by consuming one to two portions of sea fish per week, including fatty fish, which is a good source of omega-3 long chain PUFA (Koletzko et al. 2007, 2018, Coletta et al. 2010). Notably, DHA and EPA have been shown to contribute to myometrial relaxation and thus might prevent early onset of labor. Additionally, they may also inhibit the activation of trophoblastic inflammatory pathways. This could lead to a decrease in inflammation-associated preterm birth (Frew et al. 2013, Jones et al. 2014). Furthermore, they have been associated with maintaining the fetal supply of omega-3 long chain PUFAs and thus, support brain growth and subsequent neurological development in infants and children (Koletzko et al. 2007, 2008).\n\nHowever, many of the recommendations and guidelines, for example the consensus statement of the Perinatal Lipid Working Group supported by the International Society for the Study of Fatty Acids and Lipids (ISSFAL) which has been published in 2007, have not been updated for years. Furthermore, none of them specifically addresses the impact of omega-3 long chain PUFAs on prematurity (Koletzko et al. 2007).\n\nThe interest in maternal dietary intake of omega-3 long chain PUFAs and perinatal outcomes has increased over the previous years. Until now, several reviews from the Cochrane Collaboration and an abundance of randomised controlled trials have been published on omega-3 long chain PUFAs and preterm birth, highlighting the importance of this subject. The most recent Cochrane review from 2018 has synthesised evidence and included 70 randomised controlled trials (involving 19,927 women at low, mixed or high risk of poor pregnancy outcomes) (Middleton et al. 2018). The analysis revealed improvements in several outcomes such as a longer duration of gestation, higher birth weight and reduced risks of preterm birth and early preterm birth associated with omega-3 long chain PUFAs (Middleton et al. 2018). In summary, the results suggest that there is high-quality evidence that supplementation with omega-3 long chain PUFAs during pregnancy, particularly with DHA, reduced the risk of having a premature baby by 11% and it also reduced the risk of having a very premature baby, born before 34 weeks, by 42% (Middleton et al. 2018). Therefore, the authors concluded that pregnant women with one baby should be advised to take between 500 and 1000 mg of long- chain omega-3 PUFAs every day from the 12th week of pregnancy, to increase their chances of having a healthy full-length pregnancy. There is an increasing need to further elaborate the role of omega-3 long chain PUFAs on preterm birth in relation to dose, timing of supplementation and baseline omega-3 status in order to understand their impact on the prevention of obstetrical complications.\n\n\n3. The role of lipid derived mediators in initiation and resolution of inflammation\n\nA microbial infection or injury usually leads to acute inflammation with the aim of eliminating pathogens, removing cellular debris and finally restoring affected tissue. Important pro-inflammatory signalling molecules include eicosanoid lipid mediator molecules which are synthesized from the omega-6 PUFAs arachidonic acid (AA). These prostanoids comprise the prostaglandins (PG), leukotrienes (LT) and thromboxanes (TX). They are synthesized via the enzymes cyclooxygenases 1 and 2 (COX-1 and -2) by cells of the innate immune system, i.e. granulocytes or macrophages which are immediately attracted to the localisation of the respective injury or infection (Chiurchiu et al. 2018). Mast cells secret further pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNFα) and interleukins 1 and 6 (IL-1 and IL-6). Besides, other pro-inflammatory cytokines such as IL-1β, IL-12 and IL-18 are produced by M1 macrophages (Chavez-Galan et al. 2015). Thus, an inflammatory response is evoked which is characterized by the cardinal symptoms of inflammation such as heat (calor), swelling (tumor), redness (rubor), pain (dolor), and potentially a loss of function (functio laesa). Neutrophils and monocytes are also attracted to the site of infection of injury by leukotriene B4 (LTB4) and cytokines. Following this, they infiltrate the affected tissue, and thus, advance the organisms’ inflammatory response (Flower 2006, Samuelsson 2012, Chiurchiu et al. 2018). As the initiation of inflammation is essential to provide immediate response and limitation to entry of pathogens, remove damaged cells and enable quick tissue regeneration, it should be triggered rapidly and efficiently. However, on the same time, the cessation of inflammation is equally important and should be quickly initiated as well to prevent further impairment of the organism. Importantly, life-threatening events like cytokine storm or sepsis may occur following excessive, unlimited inflammation (Hotchkiss et al. 2016). Mediation and resolution of inflammation is an active process triggered by specialized pro- resolving lipid mediator molecules (SPMs), which has been shown in animal models and in different human cells (Serhan et al. 2000, Serhan 2014). They are categorized into four families according to their chemical structure and biosynthetic pathways: D- and E-resolvins (RvD and RvEs, protectins (PD), lipoxins (LX) and maresins (MaR) (Serhan 2014, Serhan et al. 2000, 2015). RvD, PD, and MaR derive from the omega-3 PUFA DHA, RvE originate from EPA (see Figure 1), and LX stem from the omega-6 AA. Lipoxygenases as well as COX- enzymes are part of the biosynthesis of the SPMs. SPMs are synthesized via the hydroxylated precursors 18-HpETE, 17-HpDHA, and 14-HpDHA (Serhan et al. 2015, Serhan 2017). Aspirin irreversibly binds COX enzymes and thus blocks the synthesis of prostanoids by altering the catalytic domains of COX. However, their capacity to catalyze the synthesis of the SPM precursors 18-HpETE, 17-HpDHA and 14-HpDHA is not abolished. The newly formed SPMs have a different stereochemical structure and are known as aspirin-triggered SPMs (AT-SPMs) (Serhan et al. 2000, 2002, 2011, 2015) (see Figure 1).\n\nEPA; Eicosapentaenoic Acid; 18-HpEDE, 17-HpDHA, 14-HpDHA (modified from Serhan 2017).\n\nSeveral key steps are responsible for the termination of inflammatory processes: dead cells have to be removed, a conversion of macrophages to resolving M2 macrophages must be initiated, and the recruitment of neutrophils must be stopped (Serhan et al. 2002, Serhan and Chiang 2013, Serhan 2017, Schett and Neurath 2018, Serhan and Levy 2018). SPMs have a vital role in the termination of neutrophil infiltration and in the initiation of phagocytosis of apoptotic cells. Besides, they are involved in the downregulation of pro-inflammatory cytokines, such as TNF-α, IL-6, IL-8 and IL-12, the reduction of platelet-activating factor and prostaglandin production. In addition to that, SPMs are also involved in the clearance of the infection site and tissue regeneration, by stimulating efferocytosis and phagocytosis, promoting wound healing. These resolutive processes are triggered simultaneously to inflammation which is evident from the interlinkage of prostaglandin synthesis with the SPM biosynthetic pathways. Both the generation of pro-inflammatory lipid mediators and the subsequent synthesis of inflammation- mediating SPMs are promoted by polymorphonuclear leukocytes (PML) (Levy et al. 2001). This implies a lipid mediator class switch in these cells which is essential for a regulated resolution of inflammation and thus for the prevention of chronification (Levy et al. 2001). PGE2 as well as PGD2 are necessary to induce lipoxygenases which are in turn necessary for the generation of LXs, Rvs, and Protectin D1 (Serhan and Savill 2005). Therefore, the switch of the lipid mediator class from a proinflammatory to a pro-resolutive function is disrupted when PG synthesis is inhibited. Hence, this may result in impaired resolution (Bandeira-Melo et al. 2000, Levy et al. 2001). In summary, the initiation of inflammation is inseparably linked to its active resolution, thus the beginning of the signaling cascade programs the end (Serhan and Savill 2005). An illustration of this perception of inflammation is available in Figure 2.\n\n(modified from Serhan and Levy 2018).\n\nAlthough the ability of pro-resolution remains to be the most important activity, SPMs also have other effects related to adaptive immune response. LX also triggers the activation of natural killer cells (Barnig et al. 2013). CD4+ T cell differentiation has also been shown to be modulated by the resolvins RvD1, RvD2 and maresin MaR1 (Chiurchiu et al. 2016). Notably, 17-HDHA and RvD1 increased IgM and IgG production in human B cells, suggesting SPM activity in humoral response and opening new functions as endogenous non-toxic adjuvants (Ramon et al. 2012).\n\n\n4. The significance of SPMs in chronic inflammatory diseases\n\nPrevious and ongoing research on SPMs has increased the knowledge on their structure and their biosynthetic pathways, receptors and function. Serhan et al. (Serhan 2014, 2017, Serhan et al. 2015) and Chiang et al. have suggested different options for the outcome of inflammatory responses (Chiang et al. 2019). If inflammation does not resolve sufficiently and remains active, i.e., pro-inflammatory signaling molecules are constantly produced, a state of chronic inflammation may develop and the activation of COX-2 enzymes is induced (Serhan et al. 2003). This is depicted in Figure 3.\n\n(modified from Serhan 2017).\n\nIn air pouch mouse models, researchers mimicked this situation successfully. Another animal model revealed the role of the SPMs LXA4, RvD1 and RvD2 in the pathogenesis of atherosclerosis, a chronic inflammatory disease (Merched et al. 2008). In mice transgenic for 12-/15-lipoxygenase, increased expression of RvD1, PD1, and 17-HpDHA proved to be protective by reducing the development of atherosclerosis compared to wild-type mice. The anti-atherogenic effects of LXA4, PD1, and RvD1 rely on several processes, including decreased expression of endothelial adhesion molecules and decreased secretion of cytokines. Importantly, the influence of nutrition on the pathogenesis of atherosclerosis has been shown in this mouse model as the transgenic mice were as susceptible to atherosclerosis as the wild type animals following a standard high- fat western diet (Merched et al. 2008, Merched et al. 2011). Furthermore. experiments in a rat model for arthritis have been conducted which demonstrated that RvD1 and the precursor metabolite 17-HDHA reduced pain and tissue damage more effectively in comparison to steroids (Lima-Garcia et al. 2011).\n\nFibrosis represents one of the characteristic features of uterine leiomyoma (Food & Agriculture Organization of the United Nations (FAO), 2010). Thus, inadequate resolution of inflammation might be considered as a cause for uterine fibroids (UF) or uterine leiomyoma. Recent animal models revealed various insights into the role of resolution of inflammation in fibrosis: For example, an animal model in pulmonary fibrosis demonstrated that exogenously administered LX4-epimer (AT-LX4) reduced fibrosis (Martins et al. 2009). Additionally, LXA4 and its analogue benzo-LXA4 reduced the extent of fibrotic changes in kidney in a rat model of early renal fibrosis, as well (Borgeson et al. 2011). A mouse model of obstructed kidney showed anti- fibrotic effects for RvE1 (Qu et al. 2012).\n\nTo date, the role of SPMs in the development of UF has not yet been investigated. However, there are some features that UF has in common with other chronic inflammatory diseases where consensus has been reached on the importance of inadequate resolution, including for example the role of SPMs in tumours (Fishbein et al. 2021). Still, further research in this context is urgently warranted and might lead to novel therapeutic options and insights.\n\n\n5. SPMs in intraamniotic inflammation and clinical chorioamnionitis\n\nIn patients in preterm delivery with intact chorioamniotic membranes as well as in patients experiencing prelabor rupture of membranes, intraamniotic inflammation can lead to an intense systemic maternal inflammatory response which is described as clinical chorioamnionitis (Gravett et al. 1986, Gibbs et al. 1992, Romero et al. 2014, 2016a). This condition ranks among the most prevalent infection-associated diseases globally, primarily affecting young primiparous women (Malloy 2014). The prevalence of chorioamnionitis in the United States was 9.7 per 1000 live births in 2008 (Romero et al. 2015). Apart from systemic inflammatory symptoms in the mother, also acute symptoms of histologic chorioamnionitis have been reported (Kim et al. 2015) as well as inflammatory responses affecting the fetus which are associated with funisitis or chorionic vasculitis (Romero et al. 2007a, 2007b, Christiaens et al. 2008). Besides being linked to maternal morbidity, neonates which are delivered by mothers suffering from clinical chorioamnionitis at term (TCC) possess an enhanced risk for long-term consequences such as cerebral palsy (Becroft et al. 2010, Elovitz et al. 2011, Thomas and Speer 2011). TCC is currently described as a heterogenous disease pattern which is accompanied by symptoms like fever, leukocytosis, foul-smelling amniotic fluid, maternal or fetal tachycardia or uterine tenderness (Newton 1993, Tita and Andrews 2010, Romero et al. 2016b). A study evaluating patients suffering from clinical chorioamnionitis reported 24% with sterile intra-amniotic inflammation and 54% with microbial-associated intra-amniotic inflammation (Romero et al. 2015). To unravel the causal mechanisms of a sterile inflammatory response, studies could demonstrate the influence of damage-associated molecular patterns (DAMPs) on sterile inflammation, for example the high mobility gene box-1 (Chen and Nunez 2010). Furthermore, it could be shown that the amniotic fluid in TCC-patients has high DAMP-levels (Romero et al. 2012) which are also associated with induction of labor (Gomez-Lopez et al. 2016). The introduction of clinical tests which can assess the differential diagnosis of the three distinct subgroups of chorioamnionitis-patients, i.e. acute, chronic or subclinical chorioamnionitis, would be a helpful tool since these patient cohorts need different therapy approaches. Patients diagnosed with microbial-associated intra-amniotic inflammation require antibiotics while patients without any intra-amniotic inflammation do not. The establishment of a clinical biomarker for the detection and identification of chorioamnionitis regardless of the presence of an intra-amniotic infection would contribute to an improved characterization and diagnosis of this disease.\n\nSPMs play a crucial role in the mediation and resolution of microorganism-derived and sterile inflammation (Harris et al. 2002, Serhan and Savill 2005, Serhan et al. 2008a, Ricciotti and FitzGerald 2011). Since it has been recognized that PGs such as PGE2 and LTs such as LTB4 are elevated in the amniotic fluid in clinical chorioamnionitis, an important contribution of these bioactive lipids in delivery at term can be assumed. Additionally, patients with microbial- associated intra-amniotic inflammation together with clinical chorioamnionitis show significantly high concentrations of AA-derived SPMs compared to those with sterile intra- amniotic inflammation (Maddipati et al. 2016a). Moreover, since TCC is characterized as an acute inflammatory condition, it is assumed that its lipid profile in the amniotic fluid differs from spontaneous labor at term (TLB). In this context, it could be demonstrated that there is no difference between concentrations of proinflammatory lipids in amniotic fluid in TLB and TCC patients. However, in all patients with TCC the presence of SPMs was significantly reduced compared to TLB patients, suggesting a decreased synthesis of SPMs as a characteristic property of TCC as opposed to infection-driven intra-amniotic inflammation where lipid mediators play an essential role (Maddipati et al. 2016b).\n\n\n6. Pre-eclampsia\n\nHypertension, in particular pre-eclampsia (PE), which accounts for most fetal, neonatal and maternal deaths, is one of the most common complications worldwide during pregnancy (Redman and Sargent 2005). Possible symptoms manifest after the 20th week of pregnancy and include de novo hypertension, edema, and proteinuria (Saftlas et al. 1990, Redman and Sargent 2005, Young et al. 2010). The exact mechanisms underlying the cascade may prompt an endothelial response within the vasculature (Redman and Sargent 2005) wall as the role of pro-inflammatory pattern-recognition receptors (Sado et al. 2011). Despite these research efforts, the precise mechanisms by which inflammation influences the pathogenesis of PE remains incompletely understood.\n\nAll pregnant women show a systemic inflammation evoked though clearance of placental debris which is released into the maternal blood circulation. PE occurs simultaneously when a failure of the systemic inflammatory response decompensates the immunogenic burden (Redman and Sargent 2001). Due to this correlation, it can be assumed that an impairment of anti- inflammatory processes is crucial for the development of a PE in pregnant women. In this context, SPMs act as potentially important “braking signals” of inflammation (Schwab and Serhan 2006, Serhan et al. 2008b, Maderna and Godson 2009). LXA4 for example is able to bind to the G-protein-coupled receptor N-formyl peptide receptor 2 (FPR2/ALX) (11) whose downregulatory signalling contributes fundamentally to LXA4-mediated anti-inflammatory response in vivo. Thereby, LXA4 acts as an inhibitor of chemotaxis of eosinophilic and neutrophilic granulocytes (Takano et al. 1997, Serhan 2007), antagonizes peptido-LTs (Badr et al. 1989), enhances macrophage-mediated phagocytosis of apoptotic cells (Maderna and Godson 2009), and reduces neutrophile infiltration in vivo (Takano et al. 1997). Since LXA4 is a crucial regulator of inflammatory response, a contribution to PE can be assumed. Furthermore, anti-angiogenic role of LXA4 has been demonstrated in vitro on human umbilical vein endothelia cells (HUVECS) as well as a LXA4-dependent reduction of lipopolysaccaride (LPS)-induced endothelial hyperpermeability (Liu et al. 2009, Pang et al. 2011). Besides, the synthetic analogue of LXA4, 5(S),6(R)-7-trihydroxymthyl heptanoate (BML-111), reduced systolic blood pressure, 24h urinary albumin excretion, serum TNFα, IL-8 levels, and LPS- dependent morphologic injury of kidney and placenta (Lin et al. 2012).\n\n\n7. First compounds with an enriched marine oil supplement on the market\n\nIn some multivitamin formulations, 30 mg of the described marine oil enriched formulation have been introduced to simulate the physiological amount of pro-resolving lipid mediators of the placenta. The first studies of this nutritional supplement have demonstrated its effectiveness in raising SPMs in plasma in different physiological and pathological conditions.\n\nHaving been able to detect a large deficit of SPMs in conditions of inflammation and as described in the protocol, it is estimated that the application of this new formulation will substantially improve both the level of SPMs in plasma and serum and the ratio between SPMs and prostaglandins.\n\nIn the studies of Elajami et al. (2016) and Souza et al. (2020) it was possible to see that the ideal doses in the intakes are between 1500 mg and 3000 mg. Elajami et al. used one formulation for a period of one year while Souza et al. used the formulation up to 24 hours. Commune to both studies was zero incidence of side effects, and the substantial increase in SPMs.\n\nThe eventual impact of these formulations on the occurrence of obstetrical complications such as pre-term birth, pre-eclampsia chorioamnionitis or amniotic inflammation remains to be evaluated. So far, no clinical studies have been conducted addressing these issues. However, the application of SPMs in the form of enriched marine oil supplements represents a promising concept for the reduction of inflammatory conditions in pregnant women and thus a first step in the creation of an immunological homeostasis.\n\n\n8. Conclusions\n\nIn conclusion, the utilization of selective pro-resolving mediators, including monohydroxylates, holds promise for the management of various acute and chronic diseases across a wide range of medical conditions. Particularly in obstetrics, supplementation with enriched marine oil nutritional products shows potential for attenuating serious complications such as pre-eclampsia and preterm birth. However, further research is necessary to determine optimal dosing regimens and fully elucidate the mechanisms underlying their therapeutic effects. Nonetheless, the findings suggest that these interventions may represent a valuable approach for addressing inflammatory diseases and mitigating obstetrical complications, thereby improving maternal and fetal health outcomes.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nReferences\n\nBadr KF, Deboer DK, Schwartzberg M, et al.: Lipoxin A4 antagonizes cellular and in vivo actions of leukotriene D4 in rat glomerular mesangial cells: evidence for competition at a common receptor. Proc. Natl. Acad. Sci. USA. 1989; 86: 3438–3442. 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Publisher Full Text\n\nGomez-Lopez N, Romero R, Plazyo O, et al.: Intra-Amniotic Administration of HMGB1 Induces Spontaneous Preterm Labor and Birth. Am. J. Reprod. Immunol. 2016; 75: 3–7. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGoodfellow L, Care A, Alfirevic Z: Controversies in the prevention of spontaneous preterm birth in asymptomatic women: an evidence summary and expert opinion. BJOG. 2021; 128: 177–194. PubMed Abstract \| Publisher Full Text\n\nGravett MG, Hummel D, Eschenbach DA, et al.: Preterm labor associated with subclinical amniotic fluid infection and with bacterial vaginosis. Obstet. Gynecol. 1986; 67: 229–237. PubMed Abstract \| Publisher Full Text\n\nHarris SG, Padilla J, Koumas L, et al.: Prostaglandins as modulators of immunity. Trends Immunol. 2002; 23: 144–150. Publisher Full Text\n\nHotchkiss RS, Moldawer LL, Opal SM, et al.: Sepsis and septic shock. Nat. Rev. Dis. Primers. 2016; 2: 16045. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nInstitute of Medicine (US) Committee on Understanding Premature Birth and Assuring Healthy Outcomes: Preterm Birth: Causes, Consequences, and Prevention.Behrman RE, Butler AS, editor. Preterm Birth: Causes, Consequences, and Prevention. Washington (DC): 2007.\n\nJones ML, Mark PJ, Waddell BJ: Maternal dietary omega-3 fatty acids and placental function. Reproduction. 2014; 147: R143–R152. Publisher Full Text\n\nKim CJ, Romero R, Chaemsaithong P, et al.: Acute chorioamnionitis and funisitis: definition, pathologic features, and clinical significance. Am. J. Obstet. Gynecol. 2015; 213: S29–S52. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKoletzko B, Cetin I, Brenna JT, et al.: Dietary fat intakes for pregnant and lactating women. Br. J. Nutr. 2007; 98: 873–877. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRomero R, Dey SK, Fisher SJ: Preterm labor: one syndrome, many causes. Science. 2014; 345: 760–765. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRomero R, Espinoza J, Goncalves LF, et al.: The role of inflammation and infection in preterm birth. Semin. Reprod. Med. 2007a; 25: 021–039. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRomero R, Gotsch F, Pineles B, et al.: Inflammation in pregnancy: its roles in reproductive physiology, obstetrical complications, and fetal injury. Nutr. Rev. 2007b; 65: S194–S202. PubMed Abstract\n\nRomero R, Miranda J, Kusanovic JP, et al.: Clinical chorioamnionitis at term I: microbiology of the amniotic cavity using cultivation and molecular techniques. J. Perinat. Med. 2015; 43: 19–36. PubMed Abstract \| Publisher Full Text\n\nSado T, Naruse K, Noguchi T, et al.: Inflammatory pattern recognition receptors and their ligands: factors contributing to the pathogenesis of preeclampsia. Inflamm. Res. 2011; 60: 509–520. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSaftlas AF, Olson DR, Franks AL, et al.: Epidemiology of preeclampsia and eclampsia in the United States, 1979-1986. Am. J. Obstet. Gynecol. 1990; 163: 460–465. PubMed Abstract \| Publisher Full Text\n\nSaigal S, Doyle LW: An overview of mortality and sequelae of preterm birth from infancy to adulthood. Lancet. 2008; 371: 261–269. PubMed Abstract \| Publisher Full Text\n\nSamuelsson B: Role of basic science in the development of new medicines: examples from the eicosanoid field. J. Biol. Chem. 2012; 287: 10070–10080. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSchett G, Neurath MF: Resolution of chronic inflammatory disease: universal and tissue-specific concepts. Nat. Commun. 2018; 9: 3261. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSchwab JM, Serhan CN: Lipoxins and new lipid mediators in the resolution of inflammation. Curr. Opin. Pharmacol. 2006; 6: 414–420. PubMed Abstract \| Publisher Full Text\n\nSerhan CN: Resolution phase of inflammation: novel endogenous anti-inflammatory and proresolving lipid mediators and pathways. Annu. Rev. Immunol. 2007; 25: 101–137. Publisher Full Text\n\nSerhan CN: Pro-resolving lipid mediators are leads for resolution physiology. Nature. 2014; 510: 92–101. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSerhan CN: Treating inflammation and infection in the 21st century: new hints from decoding resolution mediators and mechanisms. FASEB J. 2017; 31: 1273–1288. PubMed Abstract \| Publisher Full Text\n\nSerhan CN, Chiang N: Resolution phase lipid mediators of inflammation: agonists of resolution. Curr. Opin. Pharmacol. 2013; 13: 632–640. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSerhan CN, Chiang N, Dalli J: The resolution code of acute inflammation: Novel pro-resolving lipid mediators in resolution. Semin. Immunol. 2015; 27: 200–215. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSerhan CN, Chiang N, Van Dyke TE: Resolving inflammation: dual anti- inflammatory and pro-resolution lipid mediators. Nat. Rev. Immunol. 2008a; 8: 349–361. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSerhan CN, Clish CB, Brannon J, et al.: Novel functional sets of lipid-derived mediators with antiinflammatory actions generated from omega-3 fatty acids via cyclooxygenase 2-nonsteroidal antiinflammatory drugs and transcellular processing. J. Exp. Med. 2000; 192: 1197–1204. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSerhan CN, Fredman G, Yang R, et al.: Novel proresolving aspirin-triggered DHA pathway. Chem. Biol. 2011; 18: 976–987. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSerhan CN, Hong S, Gronert K, et al.: Resolvins: a family of bioactive products of omega-3 fatty acid transformation circuits initiated by aspirin treatment that counter proinflammation signals. J. Exp. Med. 2002; 196: 1025–1037. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSerhan CN, Jain A, Marleau S, et al.: Reduced inflammation and tissue damage in transgenic rabbits overexpressing 15-lipoxygenase and endogenous anti-inflammatory lipid mediators. J. Immunol. 2003; 171: 6856–6865. PubMed Abstract \| Publisher Full Text\n\nSerhan CN, Levy BD: Resolvins in inflammation: emergence of the pro-resolving superfamily of mediators. J. Clin. Invest. 2018; 128: 2657–2669. PubMed Abstract \| Publisher Full Text\n\nSerhan CN, Savill J: Resolution of inflammation: the beginning programs the end. Nat. Immunol. 2005; 6: 1191–1197. PubMed Abstract \| Publisher Full Text\n\nSerhan CN, Yacoubian S, Yang R: Anti-inflammatory and proresolving lipid mediators. Annu. Rev. Pathol. 2008b; 3: 279–312. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSouza PR, Marques RM, Gomez EA, et al.: Enriched Marine Oil Supplements Increase Peripheral Blood Specialized Pro-Resolving Mediators Concentrations and Reprogram Host Immune Responses: A Randomized Double-Blind Placebo-Controlled Study. Circ. Res. 2020; 126: 75–90. PubMed Abstract \| Publisher Full Text\n\nTakano T, Fiore S, Maddox JF, et al.: Aspirin- triggered 15-epi-lipoxin A4 (LXA4) and LXA4 stable analogues are potent inhibitors of acute inflammation: evidence for anti-inflammatory receptors. J. Exp. Med. 1997; 185: 1693–1704. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nThomas W, Speer CP: Chorioamnionitis: important risk factor or innocent bystander for neonatal outcome?. Neonatology. 2011; 99: 177–187. Publisher Full Text\n\nTita AT, Andrews WW: Diagnosis and management of clinical chorioamnionitis. Clin. Perinatol. 2010; 37: 339–354. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWaitzman NJ, Jalali A, Grosse SD: Preterm birth lifetime costs in the United States in 2016: An update. Semin. Perinatol. 2021; 45: 151390. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWard RM, Beachy JC: Neonatal complications following preterm birth. BJOG. 2003; 110 Suppl 20: 8–16. PubMed Abstract \| Publisher Full Text\n\nYoung BC, Levine RJ, Karumanchi SA: Pathogenesis of preeclampsia. Annu. Rev. Pathol. 2010; 5: 173–192. Publisher Full Text" }	[ { "id": "312357", "date": "24 Aug 2024", "name": "Carol Wang", "expertise": [ "Reviewer Expertise Statistician/Bioinformatician. I have co-supervised (with my clinician supervisor) a student who performed a comprehensive review of Omega-3 supplementation and its role to mitigate preterm birth." ], "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 provided a review of the potential role of SPMs in the mitigation of adverse pregnancy outcomes. In particular, they have addressed the possible mechanism involved and its potential to mitigate pre-eclampsia and preterm birth.\nIntroduction/Background The authors have provided a summary of preterm birth, covering mainly its short-term consequences. The authors should consider expanding on the background and address the medium to long-term consequences as well.\nSupplementation with PUFAs to prevent preterm birth In this section, the authors described the role of PUFAs in preventing preterm birth, highlighting recommended guidelines, potential mechanisms that help with mitigating preterm birth, and the benefits of consuming omega-3-rich products.\nI noticed the authors specified the 2018 Cochrane review to be the latest Cochrane review, are they aware of the updated review that included RCTs published to August 2021 (Best et al.;2022 Ref 1)? Please review the updated Cochrane review and update the statements in this section accordingly.\nAdditionally, the authors stated an increasing need to further elaborate the role of omega-3 long chain PUFAs on PTB concerning dosing, timing of supplementation and testing of baseline status, which was again mentioned in the conclusion but had not provide coverage regarding the existing literature. There are several publications available and the authors should review the literature and address these aspects accordingly. These are the PMIDs of some of the literature in the field that addressed dosing and the importance of testing before initiating supplementation: Ref 11, Ref2 , Ref 3 , Ref 4, Ref 5\n\nPre-eclampsia The pre-eclampsia section is brief compared to the other sections of the study even though it is supposedly one of the specified outcomes the authors wished to address. They have presented the potential mechanisms but had not reviewed the evidence in the literature. Both animal studies and human studies (Ref 6, Ref 7, Ref 8) have been presented regarding plasma LXA4 (and beyond); the authors should consider the literature and expand on the PE section. The authors might also want to consider examining the literature around the efficacy of omega-3 supplementation and PE (Ref 9, Ref 10). In addition, the authors have not mentioned that PE often results in medically indicated PTB, as the resolution of PE is the delivery of the baby and the placenta.\n\nConclusions The authors presented promising evidence that SPMs could mitigate adverse outcomes in pregnancy. It would be worthwhile to perform a more thorough search of the literature and address the potential importance of stratifying the population accordingly (high-risk vs low-risk pregnancies, and baseline levels) and supplementation.\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": "12568", "date": "31 Oct 2024", "name": "Pedro Antonio Regidor", "role": "Author Response", "response": "PD Dr. Pedro-Antonio Regidor Medical Director Exeltis Europe & Global Projects Exeltis Germany GmbH Adalperostraße 84 85737 Ismaning (Germany) E-Mail: pedro-antonio.regidor@exeltis.com F1000 Research research@f1000.com 04 October 2024 Re: Manuscript submission \"Enriched marine oil supplements in pregnancy for the modulation of maternal inflammatory- associated causes of preterm delivery\" (Manuscript DOI: https://doi.org/10.12688/f1000research.153569.1) Dear Sir or Madam, Thank you for the opportunity to revise and resubmit our manuscript entitled “Enriched marine oil supplements in pregnancy for the modulation of maternal inflammatory- associated causes of preterm delivery” to F1000 Research. We once again appreciate the careful review and constructive suggestions. Following this letter are the reviewer’s comments with our responses, including how and where the text was modified. We believe the revised manuscript addresses all reviewer’s comments and have met the reviewer's needs and concerns. The revision has been developed in consultation with all coauthors, and each author has given approval to the final form of this revision. This manuscript is original, has not been published before and is not currently being considered for publication elsewhere. All co-authors have read the manuscript and approved its submission to F1000 Research On behalf of all coauthors sincerely yours, PD Dr. Pedro-Antonio Regidor Comments raised by the reviewers and responses- REVIEWER’S COMMENT (Carol Wang) The authors have provided a review of the potential role of SPMs in the mitigation of adverse pregnancy outcomes. In particular, they have addressed the possible mechanism involved and its potential to mitigate pre-eclampsia and preterm birth. Introduction/Background Reviewer Comment #1: The authors have provided a summary of preterm birth, covering mainly its short-term consequences. The authors should consider expanding on the background and address the medium to long-term consequences as well. Author Response #1: Thank you for the remark. We added more information on long-term consequences of preterm birth in adulthood, which will further emphasize the burden of this health condition. Supplementation with PUFAs to prevent preterm birth In this section, the authors described the role of PUFAs in preventing preterm birth, highlighting recommended guidelines, potential mechanisms that help with mitigating preterm birth, and the benefits of consuming omega-3-rich products. Reviewer Comment #2: I noticed the authors specified the 2018 Cochrane review to be the latest Cochrane review, are they aware of the updated review that included RCTs published to August 2021 (Best et al.;2022 Ref 1)? Please review the updated Cochrane review and update the statements in this section accordingly. Author Response #2: Thank you for pointing this out. We reviewed the updated review and included its findings in the text accordingly. Reviewer Comment #3: Additionally, the authors stated an increasing need to further elaborate the role of omega-3 long chain PUFAs on PTB concerning dosing, timing of supplementation and testing of baseline status, which was again mentioned in the conclusion but had not provide coverage regarding the existing literature. There are several publications available and the authors should review the literature and address these aspects accordingly. These are the PMIDs of some of the literature in the field that addressed dosing and the importance of testing before initiating supplementation: Ref 11, Ref2 , Ref 3 , Ref 4, Ref 5 Author Response #3: We agree and are thankful for these suggestions which we included accordingly. We are convinced that this will add to a better understanding regarding possible dosages of omega-3 long chain PUFAs Pre-eclampsia Reviewer Comment #4: The pre-eclampsia section is brief compared to the other sections of the study even though it is supposedly one of the specified outcomes the authors wished to address. They have presented the potential mechanisms but had not reviewed the evidence in the literature. Both animal studies and human studies (Ref 6, Ref 7, Ref 8) have been presented regarding plasma LXA4 (and beyond); the authors should consider the literature and expand on the PE section. The authors might also want to consider examining the literature around the efficacy of omega-3 supplementation and PE (Ref 9, Ref 10). In addition, the authors have not mentioned that PE often results in medically indicated PTB, as the resolution of PE is the delivery of the baby and the placenta. Author Response #4: Thank you for your comment and for providing relevant literature. We agree and included the main findings of the publications and think that this highly improves the manuscript. Conclusions Reviewer Comment #5: The authors presented promising evidence that SPMs could mitigate adverse outcomes in pregnancy. It would be worthwhile to perform a more thorough search of the literature and address the potential importance of stratifying the population accordingly (high-risk vs low-risk pregnancies, and baseline levels) and supplementation. Author Response #5: Thank you for raising this comment. The conclusion has been adapted accordingly." } ] } ]	1	https://f1000research.com/articles/13-882
https://f1000research.com/articles/12-1579/v1	13 Dec 23	{ "type": "Research Article", "title": "Investigate the efficiency of primary care facilities in emergency situations by application of geographical and demographic standards using GIS", "authors": [ "Ali E. M. Jarghon", "Nyoman Anita Damayanti", "Inge Dhamanti", "Anas M. M. Awad", "Ali E. M. Jarghon", "Inge Dhamanti", "Anas M. M. Awad" ], "abstract": "Background This study applied geographical standards (coverage distance) and demographic standards to investigate the capabilities of primary healthcare facilities to cover the largest area of the study area and the service area that can be accessed. This study was conducted to find out the sufficient number of primary healthcare (PHC) centers required to provide healthcare services to the entire community.\n\nMethods Data was obtained by applying geographic information system (GIS) techniques to analyze primary care facilities using the demographic and geographic standards for primary care facilities.\n\nResults PHC centers cover 79% of the study area according to the geographical standard. The study area needs 41 additional centers to cover the shortfall in service provision per the demographic and geographic standards.\n\nConclusions A significant deficiency in the number of primary care centers found in the study area compared to the large population at the geographical and demographic standards level.", "keywords": [ "Primary health care", "emergencies", "geographical standard", "demographic standard", "GIS", "Coverage" ], "content": "Introduction\n\nThe pre-disaster phase of the disaster management (DM) process is based on examining the current readiness of facilities and services related to disaster risk reduction and identifying and addressing deficiencies and weaknesses for providing the needs and requirements for the success of the DM process.1 The spatial evaluation process for the sites of services and facilities needed for DM examines readiness in the study area using geographic information system (GIS) techniques and spatial analysis.2–4\n\nGeographical standards (coverage distance) and demographic standards show the capabilities of primary health center facilities to cover the largest area of the study area and the service area. In addition, we analyze the population’s suitability with the number of facilities available in the study area to find out if it is sufficient to provide services to the entire community according to the standards required.5–8\n\nPrimary health care (PHC) is a crucial component of health systems and communities’ social and economic development.9–11 PHC is responsible for providing health services and responding to emergencies at each sub-region level.11 There are 26 centers in Sleman District, Indonesia, distributed over 17 sub-regions.12\n\nThe study utilized demographic and geographic Standards as variables, gathering data from the District Health Office. It adhered to the Ministry of Health’s regulations to assess their applicability within the study area.\n\n\nMethods\n\nThere are 26 PHC centers in Sleman District, Yogyakarta City, Indonesia distributed over 17 sub-regions, Sleman District has the highest index value in disaster risk compared to other areas in Yogyakarta with a score of 97.19 This study applied ArcGIS 10.5 to analyze primary healthcare facilities by using the demographic and geographic standards of PHC to investigate if it applied in study area, and determine the extent of the shortage, assess the needs, and find possible solutions and recommendations.20,21\n\nThe GIS-based analytical approaches used for modelling accessibility to healthcare services depend on the availability of a set of spatial data. According to the geographical standard,17 each sub-district should have one PHC and urban facilities, including, a scope of service within 5 km, a school within 2.5 km, a market within 2 km, and a hospital within 5 km. Non-spatial data such as estimation of demand based on demographic characteristics of the population was considered.22\n\nFurthermore, the study utilized statistical data to measure the need for new PHC centers based on the population data. The Ministry of Health set the standard where each PHC has to provide services for 20,000 people.18\n\nWhere the Sleman district is located on coordinates, longitude is 110° 13′ 00″ to 110° 33′ 00″ east longitude and latitude is 7 o 34′ 51″ to 7 o 47′ 03″ south latitude, as shown in Figure 1.\n\nThe number of people in Sleman in 2018 was 1,206,714 consisting of 608,968 males and 597,746 females, within an area of 574,82 km2. The density of the population of Sleman Regency is 2099 people per km2. Some districts that have relatively dense populations are Depok with 5359 people per km2, Mlati with 4049 people per km2, Gamping with 3771 people and Ngaglik with 3194 people per km2.23\n\nProximity analysis assesses the accessibility of primary care facilities by measuring distances or travel times from specific locations, taking into account transportation networks and road conditions to determine ease of access during emergencies. However, spatial overlay combines spatial data layers to identify relationships and patterns, such as overlaying facility locations with demographic data to pinpoint areas with a critical need for primary care. On the other hand, network analysis optimizes routes and travel times using transportation networks, aiding in the identification of efficient emergency service routes and areas that require improved infrastructure or additional facilities. To ensure a comprehensive dataset, data was obtained from various authoritative sources, with the analytical data specifically sourced from competent authorities in Indonesia, including the Indonesian Disaster Management Agency, Statistics Indonesia (BPS), and the Sleman District Health Office.\n\nThe correctness of facilities’ locations were verified through maps using Google Earth and WorldWindEarthExplorer. Using the ArcGIS 10.5 program and Arc Analysis tools, the study area is derived based on arial images, identified by comparing it with the arial images obtained from Indonesian Disaster Management Agency. The extracted data sets were then transferred to Microsoft Excel 2013, using the network analysis in Arc GIS 10.5 (open source alternative: QGIS) to analyze and determine the scope of service of PHC. Spatial analysis of the location regarding PHC needed in the study area depended on geographical standard and demographic standards. By applying these standards, we can inquire about PHC capabilities to cover the study field and the service area that could be accessed. In addition to analyzing the population’s suitability with the number of PHCs available in the study area, is it sufficient to provide primary healthcare services to all, at the time and standard required? The analysis started by obtaining the map of 26 PHC centres distributed over 17 sub-regions as shown in Figure 2.\n\n\nResults\n\nThe geographical standard, in each sub-district, there must be one PHC and urban facilities, including scope of service within 5 km, a school within 2.5 km, a market within 2 km, and a hospital within 5 km.25\n\nThe service area of PHC in the study area, based on geographical standards, has been established as a 5 km radius from the hospitals. Figure 3 depicts the availability of hospitals within this 5 km radius. A service area within 5 km of the primary care centers has been defined. It was found that 24 health centers met the specified standard in the presence of a hospital, which is represented by the color green in Figure 3. A total of two health centers did not meet the specified standard in the presence of a hospital, which is represented by the color red (see Figure 3).\n\nThe service area of primary healthcare (PHC) in the study area is based on a geographical standard of a 5 km radius from the hospitals. In Figure 4, several key points are addressed. According to the standards set by the Indonesian Ministry of Health, health centers are required to provide services within a 5 km distance and are essential at the sub-district level.17 The study found that the service area covered a significant portion of the region. In the study area (Sleman District), health centers cover an area of 454.10 km2 out of 574.25 km2, serving approximately 79% of the study area (Figure 4). The remaining unserved area totals 120.15 km2. PHC services are not available in both the North (Cangkringan District, Pakem District, Turi District, and Tempel District) and South (Prambanan District, Berbah District, and Gamping District). In Cangkringan District, one PHC covers an area of 24.24 km2 out of a total of 44.72 km2, leaving approximately 45% of the area unserved. In Pakem District, one PHC covers an area of 25.98 km2 out of a total of 52.70 km2, leaving approximately 50.7% of the area unserved. In Turi District, one PHC covers an area of 22.90 km2 out of a total of 40.50 km2, leaving approximately 43.4% of the area unserved. In Tempel District, two PHCs cover an area of 28.45 km2 out of a total of 32.33 km2, leaving approximately 12% of the area unserved. In Prambanan District, one PHC covers an area of 14.25 km2 out of a total of 40.62 km2, leaving approximately 64.9% of the area unserved. In Berbah District, one PHC covers an area of 17.0 km2 out of the total service range of 25.85 km2, approximately 24.2% of the area remains unserved.\n\nIn conclusion, the study area has a service coverage deficiency of 21%. This highlights the need for additional primary healthcare centers (PHCs) in districts that face shortages. These districts have high population densities and are also located in close proximity to natural disaster sources, such as volcanoes, floods, and landslides.\n\nThe service area of the PHC center is defined as a 2.5 km radius from the schools. Figure 5 illustrates the availability of schools within this 2.5 km radius. A service scope within 2.5 km of the primary care centers has been implemented. In Figure 5, 26 PHC centers met the specified standard in the presence of a school, indicated by the color green.\n\nThe service area of the PHC center in the study area is based on a geographical standard of 2 km from the market. The availability of markets within this 2 km radius is depicted in Figure 6. A service scope within 2 km of the PHC center has been implemented. In Figure 6, 23 PHC centers met the specified standard in the presence of a market, indicated by the color green. Three PHC centers did not meet the specified standard in the presence of a market, indicated by the color red.\n\nTable 1 provides an explanation of the individuals who receive services and those who do not receive services through the PHC center in Sleman District, based on the demographic standard. The location of the PHC center is determined by the population distribution in the region, following the demographic standard set by the Indonesian Ministry of Health, which specifies serving a population of 20,000 people per PHC.24 After applying the demographic standards, the study yielded the following findings. The total population of the Sleman district is 1,206,714 people. Out of 26 PHCs, 520,000 individuals are receiving services in accordance with the prescribed criteria. The population without access to services amounts to approximately 686,714 people. According to the demographic standard, a larger percentage of the population should have access to PHC services. The shortage in health centers represents approximately 41 PHC centers. Statistical data indicates that the highest number of individuals lacking services in PHCs is found in the Depok region, with a shortage of 130,526 people. The construction of seven additional PHC facilities is necessary. In contrast, the percentage of individuals lacking access to PHC services in Cangkringan District is relatively small, necessitating the establishment of one new PHC.\n\n\nDiscussion\n\nIn this study, the researchers set a geographical standard to determine the service area of Primary Health Care (PHC) centers in the study area. The standard was based on a 5 km radius from the hospitals.17 According to the findings presented in the study, it was observed that 24 health centers met the specified standard for service area coverage in the presence of a hospital. This discrepancy between compliant and non-compliant health centers may have important implications for healthcare accessibility in the region. The availability of PHC services within a 5 km radius from hospitals could significantly impact the ease of access to healthcare for the population living in those areas. The PHC centers that fall within the service area are better positioned to serve their communities effectively, ensuring that a larger population has access to essential healthcare services. Factors such as distance, transportation infrastructure, and population density could play a role in determining the feasibility of establishing PHC centers in those regions.26 Policymakers and healthcare administrators may need to consider additional measures to address the accessibility challenges faced by these areas, potentially by improving transportation options or establishing additional health facilities to fill the service gap.\n\nOverall, this study provides valuable insights into the distribution of PHC services based on geographical standards in the study area. The findings can serve as a foundation for healthcare planning and resource allocation, aiming to enhance healthcare accessibility and ultimately improve the overall health outcomes for the population in the region. However, further research and collaboration between various stakeholders, including government agencies, healthcare providers, and community representatives, will be necessary to address the challenges highlighted by the study and ensure equitable access to healthcare services for all residents.\n\nThe variations in PHC coverage across different sub-districts signify the importance of localized and targeted healthcare planning and resource allocation. It is evident that certain areas are more underserved than others, requiring specific attention and efforts to improve healthcare accessibility in those regions.\n\nThe study’s findings can serve as a foundation for evidence-based decision-making in healthcare planning and policy development. By identifying the areas with the greatest need for additional healthcare resources, policymakers can prioritize their interventions to achieve more equitable healthcare distribution and improve the health outcomes of the population in the study area. The implementation of a service scope based on a 2.5 km radius from schools is a commendable step towards improving healthcare accessibility and promoting the well-being of the school community.\n\nIt also encourages early intervention and preventive healthcare practices, which can contribute to better health outcomes among the school community.\n\nThe implementation of a service scope based on a 2 km radius from markets is a promising initiative to enhance healthcare accessibility in the study area.17 The findings in Figure 6 underscore the importance of such approaches in healthcare planning. However, it is crucial to continue evaluating and refining the service area to ensure that healthcare services effectively reach all populations, particularly those residing near markets. By addressing the challenges and disparities identified in the evaluation process, healthcare providers and policymakers can work together to build a more inclusive and accessible healthcare system, ultimately contributing to the improvement of community health and well-being.\n\nThe study highlights the importance of adhering to demographic standards to ensure that a larger percentage of the population has access to PHC services. It is evident that there is a shortage of 41 PHC centers, based on the demographic standard of 20,000 people per PHC. This shortage of health centers directly affects the number of individuals who lack access to healthcare services in the region.\n\nThe study’s results have significant implications for healthcare planning and resource allocation in Sleman District. Policymakers and healthcare authorities need to take into account the demographic standards and the actual healthcare needs of the population when planning the distribution and establishment of PHC centers.\n\nThe study identifies significant gaps in healthcare access for a substantial portion of the population. Addressing these disparities requires careful planning, resource allocation, and the establishment of new PHC centers, particularly in regions with the greatest need. By addressing the shortages and expanding healthcare infrastructure, policymakers can work towards achieving a more equitable healthcare system that caters to the healthcare needs of all individuals in the district.\n\n\nConclusion\n\nIn conclusion, there is a significant deficiency in the number of PHCs in the study area compared to the large population, both at the geographical and demographic standards level. The region is in urgent need of filling these gaps by establishing new PHC centers to meet the population’s needs and provide high-quality PHC services in line with international standards. This will also help alleviate the burden on central hospitals in the region.", "appendix": "Data availability\n\nThe data for this study are owned by the Indonesian Disaster Management Agency, Statistics Indonesia (BPS), and the Sleman District Health Office, it can be obtained here:\n\n1. Click the following link, change language to English, click download publication, input your E-mail, and the file will be downloaded https://slemankab.bps.go.id/publication/2019/08/16/c400805c8dee98a3d701ea33/kabupaten-sleman-dalam-angka-2019.html\n\n2. https://geoportal.slemankab.go.id/; to access the data, use the direct link as following: https://geoportal.slemankab.go.id/layers/geonode_data:geonode:a__3404_50KB_PT_SEBARAN_RUMAH_SAKIT_SLEMAN_20 https://geoportal.slemankab.go.id/layers/geonode_data:geonode:a__3404_50KB_PT_PUSKESMAS_SLEMAN_2020\n\n\nAcknowledgements\n\nThe authors would like to thank the health district office in the City of Yogyakarta for insightful suggestions and advice.\n\n\nReferences\n\nDjalante MG, Frank Thomalla RS: Disaster Risk Reduction in Indonesia: Progress, Challenges, and Issues.2017; 1–17.\n\nAlzahrani F, Kyratsis Y: Emergency nurse disaster preparedness during mass gatherings: A cross-sectional survey of emergency nurses’ perceptions in hospitals in Mecca, Saudi Arabia. BMJ Open. 2017; 7(4): e013563. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRajeswari SP: Disaster management - introduction, community preparedness and response and role of a public health nurse. Indian J. Cont. Nsg. Edn. 2018; 19(2): 58–72.\n\nMadanian S: Disaster eHealth: Scoping Review. PubMed. 2020; 22: e18310. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKofi A: Optimizing Access to Primary Health Care Services in Rural Communities using Geographical Information System (GIS): A Case of Atwima Mponua Optimizing Access to Primary Health Care Services in Rural Communities using Geographical Information System (GI). Int. J. Comput. Appl. 2017; 163(December): 30–36. Publisher Full Text\n\nMokgalaka H: Measuring Access to Primary Health Care: Use of a GIS-Based Accessibility Measuring Access to Primary Health Care: Use of a GIS-Based Accessibility Analysis.October 2014; 2015.\n\nOldenburg CE, Sié A, Ouattara M, et al.: Distance to primary care facilities and healthcare utilization for preschool children in rural northwestern Burkina Faso: results from a surveillance cohort. BMC Health Serv. Res. 2021; 5: 1–8.\n\nAl- Al-mandhari A, Al-adawi S, Al-zakwani I, et al.: Impact of Geographical Proximity on Health Care Seeking Behaviour in Northern Oman. Sultan Qaboos Univ. Med. J. 2008; 8(3): 310–318.\n\nWHO: Primary health care.2021.\n\nMccullough K, Whitehead L, Bayes S, et al.: International Journal of Nursing Studies The delivery of Primary Health Care in remote communities: A Grounded Theory study of the perspective of nurses. Int. J. Nurs. Stud. 2020; 102: 103474. PubMed Abstract \| Publisher Full Text\n\nSundararaman T: Health systems preparedness for COVID-19 pandemic. Indian J. Public Health. 2020; 64(6): 91–93. Publisher Full Text\n\nDistrict Health office: Data of Primary healthcare centre.2020.\n\nWHO: Primary health care.2021.\n\nGarg S, Basu S, Rustagi RBA: Primary Health Care Facility Preparedness for Outpatient Service Provision During the COVID-19 Pandemic in India: Cross-Sectional Study. JMIR Public Health Surveill. 2020; 6(2): 1–7. Publisher Full Text\n\nSouza CDF, Gois-Santos VT, Correia DS, et al.: The need to strengthen Primary Health Care in Brazil in the context of the COVID-19 pandemic. Short Communication, Social/Community Dentistry. 2020; 34: 1–3. Publisher Full Text\n\nAburayya A, Alshurideh M, Albqaeen A, et al.: An investigation of factors affecting patients waiting time in primary health care centers: An assessment study in Dubai.2020; 10(6): 1265–1276.\n\nIndonesian Ministry of Health: Regulation Of The Minister Of Health Of The Republic Of Indonesia Number 43 Year 2019 Concerning Public Health Centers. Indonesian Ministry of Health: Indonesia; 2019.\n\nIndonesian Ministry of Health: Technical Guidelines for Disaster-related Health Crisis Management. Indonesian Ministry of Health: Jakarta; 2007.\n\nBnpb: Indonesian Disaster Risk Index. In Directorate of Disaster Risk Reduction (Vol. 53). 2013. Publisher Full Text\n\nDulin M, Ludden T, Tapp H, et al.: Using Geographic Information Systems (GIS) to Understand a Community’ s Primary Care Needs. PubMed. 2010; 23(1): 21.\n\nMurad A: Using GIS for determining variations in health access in jeddah city, Saudi Arabia. ISPRS Int. J. Geo Inf. 2018; 7(7). Publisher Full Text\n\nKhashoggi BF, Murad A: Issues of healthcare planning and GIS: A review. ISPRS Int. J. Geo Inf. 2020; 9(6). Publisher Full Text\n\nBps: Sleman Regency in Figures 2019.2019.\n\nIndonesian Ministry of Health: Regulation Of The Minister Of Health Of The Republic Of Indonesia Number 43 Year 2019 Concerning Public Health Centers. Indonesian Ministry of Health: Indonesia; 2019.\n\nIndonesian Ministry of Health: Technical Guidelines for Disaster-related Health Crisis Management. Indonesian Ministry of Health: Jakarta; 2007.\n\nChen L, Chen T, Lan T, et al.: The Contributions of Population Distribution, Healthcare Resourcing, and Transportation Infrastructure to Spatial Accessibility of Health Care. Inquiry (United States). 2023; 60: 004695802211460. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "253431", "date": "27 Jun 2024", "name": "Ayed Taran", "expertise": [ "Reviewer Expertise His research interests include transport services planning and management", "the socio-economic impacts of transportation congestion", "and urban planning applications", "transportation geography", "Transport models integrated in the GIS environment for accessibility and management of port systems", "GIS and Remote Sensing", "Transportation Planning and urban sustainable planning", "transport services planning." ], "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. The paper addresses geographical and demographic criteria to verify the ability of primary health care facilities to cover the largest area of the study area and accessible service area use GIS, The study found that there is a significant shortage in the number of primary health care centers in the study area compared to the large population, both on the geographical and demographic levels.\n2. The paper deals with a very important topic.\n3. There are technical issues with the arrangement of the paper as regards methodology, there are issues with presentation of data used and the subsequent methods used for the analysis of data. This includes missing methods. The paper also did not provide sufficient analysis that warrant a good discussion of results. The implications of findings are also missing.\n4. I request the authors to cite the following references:\n(i) Taran A (2023 [ref - 1]). Measuring Accessibility to Health Care Centers in the City of Al-Mafraq Using Geographic Information Systems. ResearchGate. 2023; 19 (1): 43-55\n(ii) Taran A et al. (2023 [ref - 2]). The Temporal and Spatial Analysis of Corona Pandemic in Jordan using the Geographic Information System: An Applied Geographical Study. Indonesian Journal of Geography. 2023; 55 (1).\nThis is because it is relevant to the topic of study and can be benefited from.\n5) All source data underlying the results are available to ensure complete reproducibility.\nDr. Ayed Taran Al al-Bayt University\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? Yes\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [ { "c_id": "11928", "date": "05 Jul 2024", "name": "Ali Jarghon", "role": "Author Response", "response": "Response to Reviewer Thank you for your comments. 1. Thank you for your insightful comment. We have thoroughly addressed the geographical and demographic criteria to evaluate the coverage of primary healthcare facilities in the study area using GIS. Our study indeed found a significant shortage of primary healthcare centers relative to the large population, both geographically and demographically. 2. Thank you for your positive feedback. We are pleased that you recognize the importance of the topic. Ensuring adequate coverage and accessibility of primary healthcare facilities is crucial for public health, and we hope our study contributes valuable insights to this significant issue. 3. Thank you for your constructive feedback. In response, we have extensively revised the methodology section to address the technical issues and improve the presentation of data and the methods used for analysis. Furthermore, we have expanded the analysis to provide a more thorough discussion of the results and their implications for healthcare accessibility and policy development. 4. Thank you for your feedback. We have ensured that all source data underlying the results are available to guarantee complete reproducibility." }, { "c_id": "12060", "date": "23 Jul 2024", "name": "Ali Jarghon", "role": "Author Response", "response": "Author response: Thank you for your insightful comments and expertise." } ] } ]	1	https://f1000research.com/articles/12-1579
https://f1000research.com/articles/13-1104/v1	27 Sep 24	{ "type": "Case Report", "title": "Case Report: Atypical post-COVID Cogan's syndrome", "authors": [ "Sameh Mezri", "Chaima Zitouni", "Wafa Sleimi", "Mayssa Bouzidi", "Sayhi Sameh", "Sameh Mezri", "Wafa Sleimi", "Mayssa Bouzidi", "Sayhi Sameh" ], "abstract": "Background Cogan’s syndrome is a rare autoimmune disorder characterized by ocular inflammation, vestibulocochlear dysfunction, and systemic vasculitis.\n\nCase Presentation We report a 28-year-old female who experienced decreased visual acuity and ocular redness one month after a COVID-19 infection, with ophthalmological signs linked to keratitis, uveitis and retinal vascularitis. Two weeks later, she developed vertigo, tinnitus, and sudden hearing loss, leading to a diagnosis of Cogan’s disease. The patient received corticosteroid therapy, resulting in regression of ophthalmological signs, but progressed to complete deafness. One month later, she presented with lymphocytic meningitis and high intracranial pressure, which improved under treatment. The patient later received cochlear implants.\n\nObjective This case report aims to highlight an atypical presentation of Cogan’s syndrome with neurological involvement following a COVID-19 infection. This case contributes to the limited literature on such presentations.\n\nConclusion Our case is one of only two reported instances of Cogan’s syndrome presenting with neurological signs post-COVID-19 infection, underscoring the rarity and complexity of this condition.", "keywords": [ "Cogan Syndrome", "Hearing Loss", "Meningitis", "COVID-19", "Case report" ], "content": "Introduction\n\nCogan’s syndrome (CS) is a rare autoimmune systemic vasculitis characterized by ocular inflammation, vestibulocochlear dysfunction, and systemic vasculitis.1 It was initially described in 1934 by Morgan and Baumgartner and later by David G. Cogan in 1945 as an association between interstitial keratitis and auditory symptoms.2 Approximately 450 cases have been documented to date.1\n\nWe present a case of atypical Cogan’s syndrome, distinguished by its onset following a COVID-19 infection and unusual features including ophthalmic conditions beyond keratitis and the development of neurological symptoms.\n\nTo our knowledge, this is the second reported instance of post-COVID Cogan’s syndrome in the literature.3\n\nThe purpose of this article was to explore through our case, the pathophysiology and clinical features of Cogan’s syndrome as well as to review its treatment options, prognosis, and progression.\n\n\nCase report\n\nWe present the case of a 28-year-old woman with a recent history of COVID-19 infection. One month after the infection, she developed ophthalmic abnormalities, including eye redness, blurred vision, and decreased visual acuity. These symptoms were related to bilateral non-granulomatous anterior uveitis, bilateral interstitial keratitis, retinal vasculitis, and bilateral stage 2 papilledema. She was treated with local and systemic corticosteroids, leading to improvement. Serologies for syphilis, brucellosis, Human Immunodeficiency Virus (HIV), hepatitis B virus, hepatitis C virus, rheumatoid factor, antibodies against SSA, SSB, antinuclear, anticardiolipin, and antineutrophil, and tuberculin intradermal reaction (IDR) were performed to investigate potential autoimmune or infectious origins, all of which were negative.\n\nTwo weeks after the onset of ocular symptoms, the patient developed vertigo, bilateral tinnitus, and sudden hearing loss. After excluding neurological causes with normal magnetic resonance imaging (MRI) results, she was referred to our department. The vestibular examination revealed signs suggestive of a peripheral origin. Initial pure-tone audiometry revealed moderate sensorineural hearing loss (45 dB) in both ears. Laboratory tests showed a mild inflammatory syndrome (C-reactive protein [CRP] = 15 mg/L, platelet count [PLT] = 489 × 109/L). The patient was treated with Methylprednisolone (1 mg/kg per day) and Acetylleucine (20 mg three times daily).\n\nDespite treatment, audiometric control showed worsening hearing loss (50 dB). The patient underwent 10 sessions of hyperbaric oxygen therapy, but subsequent audiograms revealed complete deafness, confirmed by auditory brainstem response (ABR) (Figure 1). Given the rapid progression to deafness, Cogan’s syndrome was diagnosed.\n\nThe patient's ABR shows the absence of Wave V at 100 dB for both ears. The right ear is depicted in red and the left ear in blue.\n\nOne month later, the patient developed headaches, dizziness, and vomiting, leading to readmission. Cerebral spinal fluid analysis showed lymphocytosis (33 white blood cells). We conducted cultures, polymerase chain reaction (PCR) for specific viruses, serological tests, and auto-immune markers, and they all came back negative. Computed tomography (CT) angiography ruled out cerebral thrombosis. Ophthalmological examination revealed episcleritis and bilateral stage 2 papilledema. Meningitis and high intracranial pressure (HTIC) were manifestations of Cogan’s disease. The patient was treated with corticosteroids and acetazolamide, leading to a good outcome.\n\nLater on, she received cochlear implants in both ears. Two years after the diagnosis, the patient was doing well.\n\n\nDiscussion\n\nCogan’s syndrome (CS) is a systemic inflammatory disease characterized by vasculitis affecting both small and large vessels.4 It is characterized by interstitial keratitis and sensorineural hearing loss, often associated with systemic vasculitis.4 While the exact pathogenesis remains unclear, some reports suggest an inflammatory autoimmune mechanism or a post-infectious etiology.5 There have been instances of CS following infections, typically occurring within 7 to 10 days of upper respiratory infections in 32% to 65% of cases.5 The mechanism might involve molecular mimicry triggered by viral infections.5\n\nSince the onset of the COVID-19 pandemic, several reports have emerged regarding autoimmune manifestations and sequelae of this infection.6 Notably, only one case of CS following a COVID-19 infection has been documented in the literature.3 Our case represents the second reported instance.\n\nThis condition typically affects young Caucasian adults without gender or hereditary predispositions.2 Initial symptoms often involve either ocular disturbances (40%) or auditory-vestibular disturbances (40%), with both the eye and ear being simultaneously affected in about 16% of cases.1,5\n\nThe classic ophthalmic presentation of CS is characterized by ocular issues such as keratitis, which may be accompanied by uveitis or conjunctivitis. The classic otolaryngological presentation includes cochlear and vestibular symptoms similar to those of Meniere’s disease. Hearing loss in CS typically affects both ears. Typically, ocular and auditory-vestibular symptoms develop within two years of each other.4,3\n\nIn CS, vestibulocochlear dysfunction is associated with inflammation in the cochlea, neuronal loss in the auditory system, endolymphatic hydrops, degeneration of the Organ of Corti, new bone formation, and atrophy of the vestibulocochlear nerve.7\n\nAtypical CS is marked by the absence of interstitial keratitis or a time gap of more than two years between cochleovestibular and ocular manifestations.5,8 Unusual ocular symptoms in atypical cases may include retinal vasculitis, papillitis, central retinal occlusion, scleritis, episcleritis, and uveitis.3,6 In contrast to typical forms, atypical CS is often associated with a broader range of systemic manifestations, such as fever, weight loss, respiratory issues, gastrointestinal bleeding, lymphadenopathy, splenomegaly, musculoskeletal symptoms, cardiovascular problems, and neurological signs.5,8 Meningitis has also been reported as a neurological manifestation of CS.1 Our case illustrates atypical CS with unusual ocular and neurological involvement.\n\nDiagnosing CS relies on its clinical manifestations and the exclusion of other conditions.1 While there are no specific diagnostic laboratory or radiologic tests, anti-Hsp70 antibodies have been suggested as a potential serological marker for typical CS.9 Vestibular testing (e.g., caloric testing, electronystagmography) and audiometry often reveal abnormalities, but no specific diagnostic pattern confirms the diagnosis.1 Hearing loss typically involves high frequencies, and autoantibodies and serologic markers of inflammation may be present.1\n\nCurrently, there are no precise guidelines for diagnosing this syndrome, and diagnosis is often delayed or missed due to the rarity and non-specific nature of the symptoms.1,4,5 Management strategies are tailored to the severity and organ involvement, focusing on preventing irreversible damage. Treatments include antibiotics, vaccines, convalescent plasma, low-salt diets, vitamins, and immunosuppressive therapies.10 Corticosteroids are the first-line treatment for managing ocular, vascular, and visceral symptoms, although their effectiveness on audio-vestibular symptoms is limited.2 Early implantation, ideally within 8 weeks of hearing loss onset, can help mitigate complications such as cochlear fibrosis.11\n\nIn CS, ophthalmic disease often fluctuates with periods of remission.2 Although ocular involvement generally has a better prognosis, it can lead to long-term complications such as cataracts from corticosteroid use.2 Conversely, auditory involvement frequently results in complete and irreversible bilateral sensorineural deafness.2\n\nIn our case, despite the resolution of ophthalmic and neurological symptoms, the patient did not respond to treatment for her hearing loss, which progressed to profound deafness. As a result, she underwent cochlear implantation. This progression aligns with findings reported in the literature.\n\nCS has a 10% mortality rate, largely due to complications arising from vasculitis, including stroke, gastrointestinal bleeding, cardiac problems, and systemic vasculitis.12\n\n\nConclusion\n\nCS is difficult to assess because it overlaps with many other diagnoses due to the variety of symptoms. Diagnosis is generally delayed until the main criteria are met and other diagnoses are excluded. Despite these difficulties, early diagnosis is essential to initiate appropriate treatment, although treatment appears to be more effective for ocular symptoms than for hearing loss, which often progresses to profound deafness. Since it is a multi-organ condition, regular follow-up is essential. Finally, the literature data is limited; therefore, further studies are needed to better understand its pathophysiology and treatment.\n\n\nConsent to publish\n\nWritten informed consent was obtained from our patient for anonymously published cases. The local ethical committee of the military hospital of Tunis, Tunisia authorized the publication of the case.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nReferences\n\nKahuam-López N, Vera-Duarte GR, Pérez-Vázquez AK, et al.: Cogan syndrome: a case report and review of the literature. Digit. J. Ophthalmol. 2023; 29(3): 88–93. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nQueirós J, Maia S, Seca M, et al.: Atypical Cogan’s Syndrome. Case Rep. Ophthalmol. Med. 2013; 2013: 476527. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChen L, Teng J, Yang C, et al.: Cogan syndrome following SARS-COV-2 infection. Clin. Rheumatol. 2023; 42(9): 2517–2518. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAlmorza Hidalgo T, García González AJ, Castañeda S, et al.: Cogan syndrome: Descriptive analysis and clinical experience of 7 cases diagnosed and treated in two third level hospitals. Reumatol. Clin. 2021; 17(6): 318–321. PubMed Abstract \| Publisher Full Text\n\nAntonios N, Silliman S: Cogan syndrome: an analysis of reported neurological manifestations. Neurologist. 2012; 18(2): 55–63. Publisher Full Text\n\nGuo M, Liu X, Chen X, et al.: Insights into new-onset autoimmune diseases after COVID-19 vaccination. Autoimmun. Rev. 2023; 22(7): 103340. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBecker RC: COVID-19-associated vasculitis and vasculopathy. J. Thromb. Thrombolysis. 2020; 50(3): 499–511. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNasrollahi T, Borrelli M, Lin HW, et al.: Cogan Syndrome: A Case Study and Review of the Literature. Ear Nose Throat J. 2023; 102(9_suppl): 35S–39S. PubMed Abstract \| Publisher Full Text\n\nBoumghit Y, Boucher S, Godey B, et al.: Speech reception after cochlear implantation for Cogan’s syndrome: Case series following CARE guidelines. Eur. Ann. Otorhinolaryngol. Head Neck Dis. 2023; 140(5): 235–238. PubMed Abstract \| Publisher Full Text\n\nBonaguri C, Orsoni J, Russo A, et al.: Cogan’s syndrome: anti-Hsp70 antibodies are a serological marker in the typical form. Isr. Med. Assoc. J. 2014; 16(5): 285–288. PubMed Abstract\n\nLee J, Biggs K, Muzaffar J, et al.: Hearing loss in inner ear and systemic autoimmune disease: A systematic review of post-cochlear implantation outcomes. Laryngoscope Investig. Otolaryngol. 2021; 6(3): 469–487. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSevgi DD, Sobrin L, Papaliodis GN: Cogan syndrome with severe medium and large vessel vasculitis. Digit. J. Ophthalmol. 2015; 22(1): 32–34. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "330430", "date": "06 Nov 2024", "name": "Debashis Maikap", "expertise": [ "Reviewer Expertise Reactive arthritis", "vasculitis", "lupus" ], "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 highlights a rare post-COVID-19 complication that progressed to Cogan’s syndrome, a unique autoimmune disorder affecting the eyes and ears. The syndrome commonly presents as non-syphilitic interstitial keratitis along with audio vestibular symptoms such as vertigo, tinnitus, and sensorineural hearing loss. Below are key points noted for improvement and additions\n\nCorrections were made for terms like \"retinal vasculitis\" rather than \"vascularitis\" to maintain accuracy in clinical descriptions.- ABSTRACT SECTION Although the case notes a possible autoimmune response triggered by COVID-19, adding context on the immune response patterns in COVID-19 and their role in post-infectious autoimmune diseases like CS would enhance the discussion. The link between viral triggers and autoimmune conditions via mechanisms such as molecular mimicry can deepen understanding of disease etiology. The use of high-dose corticosteroids presents infection risks, yet there was no mention of prophylactic measures, such as vaccinations or antibiotic prophylaxis, which are critical in managing immunosuppressed patients. Inclusion of these preventive strategies would provide a more comprehensive view of patient safety in long-term treatment. Immunosuppressive agents such as cyclophosphamide, mycophenolate mofetil (MMF), or azathioprine, often utilized as steroid-sparing therapies, were not included in this case report as potential adjunctive treatments for achieving long-term systemic control. In atypical CS cases with progressive symptoms, early initiation of these therapies might have contributed to improved outcomes. Complement levels and anti-double-stranded DNA (anti-dsDNA) antibodies, critical for ruling out lupus as an alternative diagnosis, were not mentioned. References Update: The following reference has been added to support the discussion on atypical presentations of Cogan’s syndrome:\nMaikap D, Pradhan A, Padhan P. A rare case of atypical Cogan's syndrome presenting as encephalitis. Mod Rheumatol Case Rep. 2022 Jun 24;6(2):305-308. doi: 10.1093/mrcr/rxab055. PMID: 34957524 (ref-1).\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": "12940", "date": "20 Dec 2024", "name": "Chaima zitouni", "role": "Author Response", "response": "We sincerely thank Dr. Debashis Maikap for the thoughtful review and valuable comments, and here are our detailed responses to the comments provided: Corrections were made for terms like \"retinal vasculitis\" rather than \"vascularitis\" to maintain accuracy in clinical descriptions Response: We have corrected the typo from \"vascularitis\" to \"vasculitis\" as per your suggestion. Although the case notes a possible autoimmune response triggered by COVID-19, adding context on the immune response patterns in COVID-19 and their role in post-infectious autoimmune diseases like CS would enhance the discussion. The link between viral triggers and autoimmune conditions via mechanisms such as molecular mimicry can deepen understanding of disease etiology. Response: We have included in the discussion an analysis of the potential mechanisms linking autoimmune pathologies to COVID-19. The use of high-dose corticosteroids presents infection risks, yet there was no mention of prophylactic measures, such as vaccinations or antibiotic prophylaxis, which are critical in managing immunosuppressed patients. Inclusion of these preventive strategies would provide a more comprehensive view of patient safety in long-term treatment. Response: We have added a discussion of the prophylactic measures to prevent complications arising from immunosuppressive treatments. Immunosuppressive agents such as cyclophosphamide, mycophenolate mofetil (MMF), or azathioprine, often utilized as steroid-sparing therapies, were not included in this case report as potential adjunctive treatments for achieving long-term systemic control. In atypical CS cases with progressive symptoms, early initiation of these therapies might have contributed to improved outcomes. Response: In response to your suggestion, we have expanded the discussion to include steroid-sparing immunosuppressive agents such as methotrexate and azathioprine… Complement levels and anti-double-stranded DNA (anti-dsDNA) antibodies, critical for ruling out lupus as an alternative diagnosis, were not mentioned. Response: The anti-DNA antibody was tested (we have added this to the revised text), while the complement levels were not measured. References Update: The following reference has been added to support the discussion on atypical presentations of Cogan’s syndrome: Maikap D, Pradhan A, Padhan P. A rare case of atypical Cogan's syndrome presenting as encephalitis. Mod Rheumatol Case Rep. 2022 Jun 24;6(2):305-308. doi: 10.1093/mrcr/rxab055. PMID: 34957524 (ref-1). Response: We have added the requested reference to the manuscript and integrated it into the relevant section of the discussion." } ] }, { "id": "340628", "date": "05 Dec 2024", "name": "Andrea Frosolin", "expertise": [ "Reviewer Expertise audiology" ], "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 the manuscript titled “Case Report: Atypical Post-COVID Cogan's Syndrome”. This case highlights an important and rare association of Cogan’s syndrome (CS) following COVID-19, emphasizing the evolving understanding of autoimmune and post-infectious complications. Below are detailed recommendations and suggestions for improvement.\nThe manuscript would benefit from an expanded discussion of the differential diagnosis of audio-vestibular pathologies after COVID (Frosolini A, et al., 2022 [Ref 1]). Align the case report structure with the CARE guidelines to enhance clarity and comprehensiveness. This includes ensuring that all relevant patient history, diagnostic evaluations, and follow-up details are systematically presented.\nWhile several laboratory tests are reported, it would be useful to include additional markers such as complement levels, anti-dsDNA antibodies, or other autoimmune markers to strengthen the exclusion of alternative diagnoses, particularly systemic lupus erythematosus (SLE). The role of imaging could be further elaborated. Include details about the radiological evaluation process, and consider discussing its utility in identifying autoimmune-related inner ear involvement compared to infectious or vascular etiologies.\nWhile corticosteroid therapy was described, the discussion could benefit from exploring alternative or adjunctive treatments such as steroid-sparing immunosuppressive agents (e.g., methotrexate or azathioprine). Additionally, prophylactic measures to prevent complications from immunosuppression should be briefly discussed.\nAdd a dedicated section summarizing the strengths (e.g., highlighting an underreported clinical entity) and limitations (e.g., single case, absence of long-term immunological follow-up) of the study. A short paragraph could highlight areas for future research, such as the need for studies exploring the pathophysiology of post-COVID autoimmune conditions or the timing and outcomes of cochlear implantation in autoimmune hearing loss. Possible typo: \"vascularitis\".\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": "12939", "date": "20 Dec 2024", "name": "Chaima zitouni", "role": "Author Response", "response": "We sincerely thank Dr. Andrea Frosolin for the thoughtful review and valuable comments, and here are our detailed responses to the comments provided: The manuscript would benefit from an expanded discussion of the differential diagnosis of audio-vestibular pathologies after COVID (Frosolini A, et al., 2022 [Ref 1]). We have expanded the discussion on the differential diagnosis of audio-vestibular pathologies post-COVID, incorporating insights from Frosolini et al. (2022). Align the case report structure with the CARE guidelines to enhance clarity and comprehensiveness. This includes ensuring that all relevant patient history, diagnostic evaluations, and follow-up details are systematically presented. We have carefully detailed the case in accordance with the CARE guidelines to ensure clarity and comprehensiveness. While several laboratory tests are reported, it would be useful to include additional markers such as complement levels, anti-dsDNA antibodies, or other autoimmune markers to strengthen the exclusion of alternative diagnoses, particularly systemic lupus erythematosus (SLE). The role of imaging could be further elaborated. Include details about the radiological evaluation process, and consider discussing its utility in identifying autoimmune-related inner ear involvement compared to infectious or vascular etiologies. The anti-DNA antibody was tested (we have added this to the revised text), while the complement levels were not measured. We have expanded the differential diagnoses, particularly focusing on systemic lupus erythematosus and other autoimmune diseases. We have expanded the discussion to detail the radiological evaluation process, emphasizing the role of advanced MRI sequences in distinguishing autoimmune-related inner ear involvement from infectious or vascular etiologies based on characteristic patterns. While corticosteroid therapy was described, the discussion could benefit from exploring alternative or adjunctive treatments such as steroid-sparing immunosuppressive agents (e.g., methotrexate or azathioprine). Additionally, prophylactic measures to prevent complications from immunosuppression should be briefly discussed. In response to your suggestion, we have expanded the discussion to include steroid-sparing immunosuppressive agents such as methotrexate and azathioprine… We have added a discussion of the prophylactic measures to prevent complications arising from immunosuppressive treatments. Add a dedicated section summarizing the strengths (e.g., highlighting an underreported clinical entity) and limitations (e.g., single case, absence of long-term immunological follow-up) of the study. A short paragraph could highlight areas for future research, such as the need for studies exploring the pathophysiology of post-COVID autoimmune conditions or the timing and outcomes of cochlear implantation in autoimmune hearing loss. We have included a dedicated section in the discussion summarizing the strengths and limitations of our study. A short paragraph has been added to highlight areas for future research. Possible typo: \"vascularitis\". We have corrected the typo from \"vascularitis\" to \"vasculitis\" as per your suggestion." } ] } ]	1	https://f1000research.com/articles/13-1104
https://f1000research.com/articles/13-1547/v1	20 Dec 24	{ "type": "Method Article", "title": "Using text-mining to measure the scientific impact and legacy of ELIXIR, a distributed research infrastructure for life science data", "authors": [ "Francesca De Leo", "Erika Balsyte", "Robert Petryszak", "Marilena D’Ambrosio", "Chiara Bruno", "Martin Cook", "Ivan Mičetić", "Corinne S. Martin", "Erika Balsyte", "Robert Petryszak", "Marilena D’Ambrosio", "Chiara Bruno", "Martin Cook", "Ivan Mičetić", "Corinne S. Martin" ], "abstract": "Background ELIXIR is a pan-European public-funded research infrastructure dedicated to life science data. As such, it must demonstrate public value to its funders and stakeholders. We present methods to inventory research publications linked to ELIXIR that have received funding and support, as well as related citation metrics, used as performance metrics for these audiences.\n\nMethods To overcome challenges inherent in ELIXIR’s distributed structure, and the fact that those publishing ELIXIR-supported work are typically working part-time on ELIXIR matters, a semi-automated approach, consisting of text-mining followed by manual curation, is presented. A country-level case study (ELIXIR Italy) refines and expands the methods, notably by introducing more granularity in the curation process (e.g. considering all national-level grants, examining affiliations to report publication per institute) and by additionally looking at the scientific impact of the resources developed and operated by the Italian Node of ELIXIR.\n\nResults Overall, the methods described in this article have shown to: (1) be repeatable with acceptable levels of accuracy and consistency (notably across curators); (2) require reasonable effort in terms of curation of monthly ‘harvests’ of publications (obtained by text-mining); and (3) to be well-adapted to ELIXIR’s distributed nature.\n\nConclusions Concrete examples are provided of downstream uses of the inventoried publications and their citations, both for ELIXIR as a whole and for the Italian case study. Limitations of the methods are discussed, particularly the challenges associated with using an ‘Open literature’ database (Europe PMC) for the text-mining, and the constraints related to curation capacity. The methods, along with the valuable lessons learned during their development, are sufficiently generic and pragmatic enough to be readily adapted by other similar research infrastructures.", "keywords": [ "KPI", "metric", "funder", "literature", "bioinformatics", "database", "resource", "performance" ], "content": "Introduction\n\nELIXIR is a pan-European distributed research infrastructure dedicated to life science data, formally founded in 2013 and currently implementing its third five-year scientific programme (ELIXIR, 2023). In 2016, it was recognised by the European Strategy Forum for Research Infrastructures (ESFRI), a forum for government officials focused on research infrastructure, as a ‘Landmark’ (ESFRI, 2016). Landmarks are reference research infrastructures considered pillars in the European Research Area landscape, offering not only services/resources to academic research, but also supporting development and innovation. ELIXIR currently has 21 member countries, three observer countries, plus one intergovernmental organisation, the European Molecular Biology Laboratory (EMBL), which has its own country membership (overlapping in parts with that of ELIXIR).\n\nEach ELIXIR member forms a country ‘Node’, with the European Bioinformatics Institute (EMBL-EBI) serving as the ‘European’ Node of ELIXIR. Having the Open Science principle at its heart, ELIXIR comprises over 240 research institutions across Europe, and coordinates the provision, development and discoverability of more than 500 bioinformatics resources covering databases, tools, software, interoperability standards, training, and cloud computing. These resources are financed through public funds (Smith et al., 2020) and are typically free at the point of use, for academic and industry users alike, who can hence focus on their research rather than on the development and operation of the underlying infrastructure and its resources. ELIXIR also works to streamline and integrate national-level bioinformatics infrastructures, to strengthen national training programmes, and to bring together bioinformatics experts to create guidelines and best practices, towards ensuring the long-term sustainability of resources and infrastructure.\n\nELIXIR must continually demonstrate public value to its national and international funders and stakeholders more broadly to ensure their ongoing support and hence the continuity of infrastructure and resource development and operation (Martin et al., 2021). Inventories of research publications arising from the use of the infrastructure (by others than those developing/operating it) are a common source of performance metrics requested by funders and related stakeholders (ESFRI, 2019, OECD, 2019, European Commission, 2020). A subset of ELIXIR databases, the ELIXIR Core Data Resources (Durinx et al., 2017), are indeed widely cited and acknowledged in thousands of research publications by life scientists across the globe (Drysdale et al., 2020).\n\nAround 700 bioinformaticians and related professionals across Europe form the developers and operators of the ELIXIR infrastructure and resources, and are typically working part-time on ELIXIR matters, often alongside more conventional research activities. These professionals usually have research profiles and, through their daily work on ELIXIR matters, actively contribute to the generation and dissemination of new technical and scientific knowledge linked to the development and operation of the ELIXIR infrastructure and its resources. These efforts are enabled (in part or fully) by a range of funding sources, such as ELIXIR’s internally-funded projects (also referred to as ‘Commissioned Services’), national-level financing, and grant funding from the European Union and other international funders awarded in the name of ELIXIR. In line with the Open Science principles, this new knowledge is shared openly via ELIXIR-supported research publications (peer-reviewed, preprints), which collectively represent an additional source of performance metrics of ELIXIR’s scientific legacy as a research infrastructure.\n\nThe purpose of this article is to present the methods that have been developed and implemented to inventory research publications that have received funding and support linked to ELIXIR, as well as related citation metrics. The methods are presented for ELIXIR as a whole (lightweight, considering all Nodes) and for a particular country Node, ELIXIR Italy, itself also of distributed nature. For the latter, the methods were refined thanks to greater curation capacity at the level of the Node, and also expanded to include the inventorying of research publications citing resources developed and operated by the Node. This exercise is very informative of the Node’s scientific impact and legacy, yet it would not easily scale for the whole distributed infrastructure (in terms of curation capacity at the ELIXIR Hub, the infrastructure’s coordinating secretariat) since the infrastructure operates hundreds of resources under the ELIXIR umbrella.\n\nA semi-automated approach, consisting of text-mining followed by manual curation, is presented, with the Italian case study going further so as to better reflect the needs and expectations of national-level funders and other (internal and external) stakeholders of the Node. Of note, the guiding principle in developing these methods was that they would not require extensive technical nor programming knowledge to be applied once in place - as a result, the methods do not involve any modelling (e.g. with training and test sets), nor do they claim to be rigorously accurate. It is hoped that the steps taken, and lessons learned (both ELIXIR-wide and at the scale of a country Node), in arriving at operational methods in the context of distributed organisational settings will be useful to other such research infrastructures and organisations, in the life sciences and other disciplines.\n\n\nMethods\n\nMain challenges\n\nTwo intrinsic challenges that the lightweight methods needed to overcome were (1) that those publishing ELIXIR-supported work are typically working part-time on ELIXIR matters, and (2) the scale of ELIXIR’s “people infrastructure” (more than 700 specialists across Europe, showing a level of staff turnover). The latter itself represented a significant barrier to manual reporting of publications. As for the former, traditional bibliometrics/scientometrics approaches centred on researchers’ unique identification numbers (e.g., ORCID, ResearcherID/Publons), and/or institutional affiliations, were deemed inadequate for identifying research works (funded and/or) supported by ELIXIR, as these approaches would result in significant amounts of ‘false positives’, i.e. publications not supported by ELIXIR. To overcome these two challenges, the lightweight methods were designed to (1) detect, through text-mining within an Open research literature database (EuropePMC, Rosonovski et al., 2024), publications (peer-reviewed and preprints) in which there were acknowledgments of the support and/or funding received from ELIXIR, (2) followed by manual curation for increased accuracy.\n\nSources of ELIXIR support\n\nFor the ELIXIR infrastructure as a whole (all Nodes), two main streams of funding and/or support were considered: (1) ELIXIR’s internally-funded projects, i.e. Commissioned Services comprising Implementation Studies, Staff Exchange, Knowledge Exchanges and Industry days, and Travel Grants, where funding eligibility is restricted to institutes part of the ELIXIR membership, and (2) grant funding from international funders such as the European Union (notably under the Research Infrastructures Work Programmes), in which several ELIXIR institutes are beneficiaries and the purpose of the grant is to develop and/or operate the ELIXIR infrastructure.\n\nSearching for relevant publications using text-mining\n\nEuropePMC’s Application Programming Interface (API) was used to search for relevant publications (technical documentation), as well as their citations in the Open literature (technical documentation), considering all years for which ELIXIR existed, even as a concept (circa 2011), with a cut-off date of December 2023 (for the purpose of this article). Search configuration parameters ( Table 1), i.e. ‘search terms’ for short, comprised unique grant identification numbers and names, names and short codes of countries that are members of ELIXIR (e.g. ELIXIR Norway, ELIXIR NO), as well as “boiler-plate” text strings relating to ELIXIR support and/or funding including for ‘signature’ events (e.g. the ELIXIR-convened Biohackathon Europe) and technical workshops/meetings. For use in EuropePMC’s API, the search terms (query parameters) were first URL encoded using the online utility URL Encode/Decode, provided free, courtesy of Dan’s Tools: for example, the EU funded project ELIXIR-CONVERGE would be encoded as “ELIXIR%2DCONVERGE”. Of note, EuropePMC is case insensitive, meaning that it is not necessary to provide search terms in both lower/upper cases. For the lightweight methods, acknowledgement of the use of the infrastructure and its resources was deemed beyond scope due to the mismatch between curation capacity at the ELIXIR Hub and the several hundreds of resources operated under the ELIXIR umbrella.\n\nAn initial set of search terms were entered in a Google Sheet, and retrieved from there via a specific Google Sheets API. This allowed for the subsequent dynamic configuration of the search queries when new grants were awarded to ELIXIR. The Google Sheet interacted with the EuropePMC API based on the retrieved search configuration, and the text-mining process resulted in the generation of tab-delimited text files listing potentially relevant publications comprising a mix of ELIXIR-related works (‘true positives’) and false positives.\n\nManual curation and search optimisation\n\nManual identification of false positives was carried out using the tab-delimited text files, one column of which collated the triggers relevant to each returned publication. Triggers represented the search terms that had been detected in the publications and were found to guide curation very effectively. False positives were blacklisted for future searches. To ensure consistency, reproducibility and the lowest subjectivity possible in curation decisions, curation efforts were carried out by a small team of two ELIXIR Hub staff who kept a record of difficult and/or unclear cases, and of the decisions taken. They were supported on an ad-hoc basis by additional ELIXIR Hub staff who had more extensive knowledge and understanding of ELIXIR’s activities in certain thematic and technical areas.\n\nDuring the initial development of the methods, iterative searches and manual curation rounds were undertaken to test and refine the set of search terms, and to identify which sections of EuropePMC entries (for each publication) were the most informative to target by text-mining - these were found to be the title, first author, funding statement, and acknowledgments. The initial text-mining runs naturally returned large backlogs of potentially relevant publications since they covered the years 2011 to 2019: although this made the curation effort significant at the time, it also provided a rich list of results to optimise the searches, notably to identify which sections ( Table 2) of the publications ought to be extracted to the tab-delimited text files used during manual curation and any downstream visualisations.\n\nA set of known ELIXIR-supported publications was useful for fine-tuning the approach, as they would be expected to be returned by the searches. If this was not the case, underlying reasons would be investigated and, where possible, solutions found - if no solution was found, these publications were added (via their unique EuropePMC identification number) to a whitelist that would be pulled to the tab-delimited text file, regardless. Later on, once the methods were operational (circa early 2020), the text-mining “harvests” (i.e. searches) followed by curation ended up being carried out on a monthly basis, to identify recently published and/or recently indexed publications, based on the latest set of search terms.\n\nSearch terms returning very high numbers of false positives for very few true positives were gradually excluded from searches. This was for instance the case of “ELIXIR” as a standalone term (misunderstood for the word “elixir”), “implementation study” (rather than the more precise “ELIXIR implementation study”), and the project name “BY-COVID” (incorrectly read as the very common text string “[infected] by COVID-19”). This was to strike a balance between accuracy (i.e. not missing any relevant publication) and overwhelming curators. Rather, it was deemed more useful to arrive at manageable monthly text-mining harvests and curation workloads moving forwards (approximately 3 to 4 hours per monthly harvest), since the intention was to arrive at operational performance metrics that could be monitored in the long-term, rather than a very accurate one-off exercise.\n\nUnless otherwise specified, the methods for the Italian case study are identical to the lightweight one described above (for ELIXIR as a whole).\n\nSources of ELIXIR Italy support\n\nBuilding on the experience of the ELIXIR Hub effort, the Italian case study began at the end of 2022 and was focused on the period January 2015 (the year when Italy became an ELIXIR member country) to December 2023 (cut-off date for the purpose of this article). The main three sources of funding and/or support considered in the Italian case study were (1) those ELIXIR internally funded projects in which one or more of the institutes of the Italian Node were involved, (2) ditto for grant funding from international funders, and (3) national-level grants awarded to the Node to develop and/or operate its infrastructure and resources.\n\nSearching for relevant publications using text-mining\n\nFor the Italian case study, the ‘ELIXIR as a whole’ set of search terms was refined to remove terms not relevant to the Node, e.g. if the Italian Node was not a beneficiary of a given grant. Additional search terms were introduced (e.g. national-level grants) along with other terms (e.g. Italian Node of ELIXIR, often used instead of ELIXIR-IT or ELIXIR IIB) that were known to be mentioned in publications acknowledging support and/or funding by ELIXIR Italy ( Table 3).\n\nThe major expansion of the methods followed Drysdale et al. (2020) and their focus on citations of ‘key articles’ describing the Node’s resources. This additional search effort was approached by using the Digital Object Identifier (DOI) of these ‘key articles’ to collate citation data and use this as proxy for the scientific impact of the Node’s resources. For example, the resource PatSearch was first described in Pesole et al. (2000) and an improved version of it was described in Grillo et al. (2003) - these are considered the resource’s ‘key articles’. The list of ELIXIR Italy resources was based on the 2023 version of the Node’s Service Delivery Plan under the ELIXIR umbrella, and the DOIs were sourced from the bio.tools registry (Ison et al., 2019). For the 65 resources developed and operated by ELIXIR Italy in 2023, a total of 124 DOIs were hence collated, representing the ‘key articles’ describing initial and improved versions of these resources.\n\nManual curation and search optimisation\n\nCuration in the context of the case study was a very similar process to that described above but carried out by ELIXIR Italy staff with knowledge of the Node’s activities. Author affiliations were additionally scrutinised and encoded to obtain a more granular view of dissemination efforts and scientific impact at the level of the Node’s institutes (30 in total at the time of writing). Sources of false positives were like that described above, e.g. project names such as “ORCHESTRA” (understood as the word “orchestra” or other projects called as such). As above, a balance was struck between accuracy and curation capacity by excluding problematic search terms (those returning high levels of false positives).\n\n\nResults\n\nFor the period 2011 to 2023, 972 publications supported by ELIXIR (as a whole) were identified, and these were collectively cited over 31,000 times ( Figure 1). The curated inventory of publications and their citations in the Open literature were used to create visualisations in Tableau software, which were themselves featured on ELIXIR’s impact dashboard, among other indicators relating to ELIXIR. On this dashboard, the page showing ELIXIR’s scientific legacy as a research infrastructure received nearly 600 unique visitors from over 40 countries in just 12 months (December 2022 to November 2023).\n\nFor the period 2015 to 2023, 408 publications supported by ELIXIR Italy were identified, which were collectively cited almost 5,000 times ( Figure 2). The distribution of publications across Node institutes ( Figure 3) highlights the disproportionate contribution of the National Research Council (NRC), though unsurprising given its role as the coordinating institute of ELIXIR Italy. Associated visualisations can be accessed on ELIXIR Italy’s impact dashboard which, for a 24 month period (January 2022 to December 2023), received over 5,000 unique visitors from 10 countries. In terms of the impact of resources developed and operated by ELIXIR Italy ( Table 4), citations of their ‘key articles’ amounted to 8,700, across the five ELIXIR Platforms, which are the technical domains of implementation of the infrastructure.\n\nAcronyms: see https://elixir-italy.org/about/members/.\n\n\nDiscussion\n\nIn almost four years of implementation, the methods have been found to be adequately performant at identifying publications supported by ELIXIR, if the relevant triggers were present (and correctly spelled) in the publications. Since the inception (in 2020) of a dedicated “How to acknowledge ELIXIR funding and support” page on the ELIXIR website (with the analogous one on the ELIXIR Italy site, bottom part), curators have noticed clear improvements in how ELIXIR’s contribution is acknowledged.\n\nThe upward trend in citation numbers for both the lightweight and case study methods suggested long-term scientific impact of these publications, a reminder that impact is slow to develop. It is not just citations that take time to appear: large-scale projects such as ELIXIR-EXCELERATE which involved 47 partners from 17 countries (Harrow et al., 2021; €19 million, 2014-2019) were found to be still acknowledged in publications years after their completions, This is not solely due to delayed indexing but also to the enduring legacy of their support for research activities within teams connected to the project.\n\nOverall, the methods described in this article have shown to: (1) be repeatable with acceptable levels of accuracy and consistency (notably across curators); (2) require reasonable effort in terms of curation of monthly harvests (in contrast to running an ELIXIR wide manual reporting process); and (3) to be well-adapted to ELIXIR’s distributed nature (including at Node-level in the Italian case study).\n\nWhile the text-mining component can be operated with minimal ELIXIR expertise, it is true that the curation component requires a deeper awareness and understanding of the organisation’s activities. One example is when ELIXIR was involved in large “cluster” grants (e.g. EOSC-life ) in which many life science research infrastructures were also funded, meaning that not all publications acknowledging EOSC-life’s grant would be relevant to ELIXIR, despite ELIXIR coordinating the project.\n\nThe Italian case study highlighted the usefulness of national-level granularity in the search terms. Whilst it is true that the lightweight ‘ELIXIR as a whole’ methods do include a few national-level grants in its search terms, this is far from being fully implemented. Their inclusion was exploratory at best and, based on this experience, there is insufficient appetite at the ELIXIR Hub to fully implement national-level search terms due to capacity constraints in terms of curation and limited knowledge of ELIXIR activities at Node level.\n\nOne main limitation of the methods is linked to EuropePMC being used as the central source of publication data - only the publications indexed in this literature database, focused on life sciences, can be returned by the searches. At the time of writing, specialised journals such as Data Science Journal and Data Intelligence were not yet indexed due to perceived low demand of these in the user base of EuropePMC. However, based on user feedback, EuropePMC started indexing BioHackrXiv in 2021, especially as it is the recommended preprint server for outcomes of the ELIXIR-convened Biohackathon Europe series (Castro et al., 2021).\n\nAlthough research funders are increasingly requesting that the work they fund is openly accessible, it was found that certain ELIXIR-relevant publications were behind paywalls. This meant that EuropePMC was not able to fully “ingest” the text of certain sections of the source text, e.g. funding statement, which made text-mining efforts fail, despite the publication having a presence in EuropePMC and the right trigger being present. This was manually solved on an ad-hoc basis by whitelisting these publications using their unique EuropePMC identification number. Additionally, this method can effectively identify ELIXIR-related publications that are non-compliant with open access policies and funder requirements.\n\nLooking forward, EuropePMC continues to work closely with scientific publishers to agree and implement standards so that ingestion of information from the publishers’ sites to EuropePMC is more effective. This is an on-going and quite a considerable task, and additional sources of text-mining failures included conflicts in metadata, embargoed works in paywalled journals, as well as citations being limited to the Open literature, the latter leading to underestimated scientific impact.\n\nThe intention behind this work was to arrive at operational performance metrics that could be easily monitored in the long-term, in contrast to a rigorous and hence heavy-duty process requiring significant resources, likely to be carried out once or twice at best. The curated inventories of publications, including their citations, have so far been used as input into ELIXIR’s annual reports, ELIXIR newsletters and social media posts (under the heading “Recommended reading”), technical reporting to funders, as well as ELIXIR’s monitoring report to ESFRI and a growing number of requests for information from national funders, such as Sweden (Swedish Research Council, 2021), Finland (Academy of Finland, 2023) and the Netherlands (NWO, 2023).\n\nSimilar to ELIXIR, ELIXIR Italy has used the curated inventories of publications and their citations for a range of internal and external monitoring activities, as required by various stakeholders, for example periodic Node reviews and presentations for ELIXIR’s Board of funders, Scientific Advisory Board and Industry Advisory Committee. The information has also been used to report to ELIXIR Italy’s own Scientific Advisory Board, as well as for the Node’s general assembly and monthly newsletters. Finally, the information has been used to report to ELIXIR’s Italy’s main funder, the Italian Ministry for Universities and Research (MUR)), notably as part of evaluation processes and of national-level grant funding for research infrastructures.\n\n\nConclusions\n\nAs illustrated by the Italian case study, these methods represent the first step of potentially many exciting downstream analyses using the curated inventories as starting materials. Their citations would shed light on the impact of these works, through their use by others. Yet these inventories alone have proved to be useful already, in terms of communicating performance and impact of both ELIXIR activities (as a whole and for a country Node) and the use of a Node’s resources. The Italian case study also demonstrated the value of knowledge sharing within an organisation, in the spirit of Open Science, with the country Node leveraging the experience of the ELIXIR Hub, building on and enhancing it.\n\nThe methods’ main strength lies in its relative ease of implementation, especially in the context of a distributed life science research infrastructure staffed with highly qualified and research-active personnel: many will have the programming skills required to operationalise the methods, whilst no such technical skill is required for the curation part of the methods. If not, external assistance can be sought in the form of consulting, to get the approach off the ground. Another strength of the methods is its versatility as new funding streams become available and resources are further developed.\n\nThe methods, along with the valuable lessons learned during their development, are generic and pragmatic enough to be readily adapted by other similar research infrastructures. This extends its potential application beyond life sciences to encompass physical sciences, social sciences, and the humanities.\n\nEthical approval and consent were not required.", "appendix": "Data availability\n\nEuropePMC (Rosonovski et al., 2024; https://europepmc.org/), an Open research literature database, was used as the source for the underlying data (publications in the life sciences) used in the methods described in the article.\n\nThe extended data created as a result of applying the methods are curated inventories of publications that have received funding and support from ELIXIR. They have been deposited in Zenodo (Martin et al., 2024; https://doi.org/10.5281/zenodo.14136249) under a Creative Commons Attribution 4.0 International license, so that others can access them for illustrative and verification purposes.\n\nN/A\n\n\nAcknowledgements\n\nThe authors express their appreciation to Andrew Smith and Niklas Blomberg for initiating this effort, unlocking the necessary internal resources, and providing institutional support throughout. The authors would like to thank the following ELIXIR Hub colleagues who took part in the May 2019 workshop where the scope and methods were first discussed: Susanna Repo, John Hancock, David Lloyd, and Jerry Lanfear. Additional thanks go to the following ELIXIR Hub colleagues who helped with unclear cases during curation efforts: Giselle Kerry, Gary Saunders, Rachel Drysdale, Katharina Lauer, Serena Scollen, Jonathan Tedds, and Jen Harrow. Finally, the authors would like to express their gratitude to Graziano Pesole (Head of the Italian Node of ELIXIR) and Federico Zambelli (Technical Coordinator of the Italian Node of ELIXIR) for their institutional support throughout the project and their collaboration in defining the methodology and identifying needs.\n\n\nReferences\n\nAcademy of Finland: Assessment of benefits of international Research Infrastructure memberships for Finland 2020-2021. Questionnaire results. 2023. Reference Source\n\nCastro LJ, Martin C, Lazarov G, et al.: Measuring outcome and impacts from the BioHackathon Europe.2021. Publisher Full Text Reference Source\n\nDrysdale R, Cook CE, Petryszak R, et al.: The ELIXIR Core Data Resources: fundamental infrastructure for the life sciences. Bioinformatics. 2020; 36(8): 2636–2642. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDurinx C, McEntyre J, Appel R, et al.: Identifying ELIXIR core data resources. F1000Res. 2017; 5: 2422. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nELIXIR: ELIXIR Scientific Programme, 2024-2028.2023. Reference Source\n\nESFRI: Monitoring of research infrastructures performance.2019. Reference Source\n\nESFRI: Strategy Report on Research Infrastructures.2016. Reference Source\n\nEuropean Commission: Supporting the transformative impact of research infrastructures on European research: report of the High-Level Expert Group to assess the progress of ESFRI and other world class research infrastructures towards implementation and long-term sustainability. Director General for Research and Innovation, Publications Office of the European Union; 2020. Reference Source\n\nGrillo G, Licciulli F, Liuni S, et al.: PatSearch: a program for the detection of patterns and structural motifs in nucleotide sequences. Nucleic Acids Res. 2003; 31(13): 3608–3612. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHarrow J, Hancock J, Blomberg N, et al.: ELIXIR-EXCELERATE: establishing Europe’s data infrastructure for the life science research of the future. EMBO J. 2021; 40(6): e107409. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIson J, Ienasescu H, Chmura P, et al.: The bio.tools registry of software tools and data resources for the life sciences. Genome Biol. 2019; 20(1): 164. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMartin C, Repo S, Márquez JA, et al.: Demonstrating public value to funders and other stakeholders—the journey of ELIXIR, a virtual and distributed research infrastructure for life science data. Ann. Public Coop. Econ. 2021; 92(3): 497–510. Publisher Full Text\n\nMartin C, De Leo F, D’Ambrosio M, et al.: Publications Supported by ELIXIR and ELIXIR Italy (during the Periods 2011-2023 and 2015-2023). Zenodo. 2024; 13 November 2024. Publisher Full Text\n\nNWO: Analysis of the Dutch participation in international research infrastructures.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. Publisher Full Text\n\nPesole G, Liuni S, D’Souza M: PatSearch: a pattern matcher software that finds functional elements in nucleotide and protein sequences and assesses their statistical significance. Bioinformatics. 2000; 16(5): 439–450. PubMed Abstract \| Publisher Full Text\n\nRosonovski S, Levchenko M, Bhatnagar R, et al.: Europe PMC in 2023. Nucleic Acids Res. 2024; 52(D1): D1668–D1676. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSmith A, Martin C, Blomberg N: ELIXIR position paper on suitable business models for open data. F1000Res. 2020; 9: 65. Publisher Full Text\n\nSwedish Research Council: National benefits from Swedish membership of international research infrastructures 2016-2019.2021. Reference Source" }	[ { "id": "354530", "date": "06 Jan 2025", "name": "Helen M. Berman", "expertise": [ "Reviewer Expertise Structural bioinformatics", "structural 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 article describes a method to assess the impact of ELIXIR supported research. This was done by using text mining to search for relevant articles contained in EuropePMC. This procedure was used for all ELIXIR nodes and for ELIXIR Italy. In the latter case the search terms were modified to be appropriate for that node. In the first step specific search terms were used to search sections of articles. This automated procedure was followed by manual curation.\nIt is important to have quantitative methods to analyze impact of the ELIXIR infrastructure and this is a good first step. The paper is well written, and the limitations of the approach are adequately described. For example, the EuropePMC does not index all the journals that describe projects funded under the ELIXIR umbrella. It is also disturbing that some articles are published in journals that are behind paywalls.\nAlthough the curated datasets are put into Zenodo, the authors might consider making a website that would allow authors to submit and curate their ELIXIR related articles.\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? Partly\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? Yes", "responses": [] } ]	1	https://f1000research.com/articles/13-1547
https://f1000research.com/articles/13-269/v1	15 Apr 24	{ "type": "Research Article", "title": "Prescription writing quality of psychotropic agents in MBBS interns in a private medical college hospital in Mangaluru", "authors": [ "Poorva Bakshi", "Sharanya B. Shetty", "Shreya Bera", "Priyanka Renita D'Souza", "Keshava Pai", "Poorva Bakshi", "Shreya Bera", "Priyanka Renita D'Souza", "Keshava Pai" ], "abstract": "Background Rational prescription writing is an important skill to master during internship. This Quality Improvement (QI) project aimed to understand the state of prescription writing among interns posted in the Department of Psychiatry, analyze the causes responsible for errors in prescription writing and bring about a change in the current practice.\n\nMethods The MBBS interns are posted in the Department of Psychiatry for 15 days. During day 1 to day 5 of their posting, a pre intervention phase was conducted wherein prescriptions written by interns in the Department of Psychiatry were collected. The prescriptions were scored based on 14 criteria which were selected based on World Health Organization (WHO) guidelines and Medical Council of India (MCI) ideal prescription format. During PDSA (Plan Do Study Act) Cycle 1, an educational handout was distributed to the interns containing the MCI ideal prescription format and WHO guidelines regarding prescription writing. The brochure was also verbally explained to the interns. From day 7 to day 15 of their posting, prescriptions written by the interns were collected. The prescriptions were scored using the same criteria.\n\nResults During the pre intervention phase the mean total score of prescriptions was 9.54 ± 1.003. There was a significant improvement in the mean total score to 10.26 ± 0.746. There was a 7.54% improvement. There was also a significant improvement in several individual criteria.\n\nConclusions The first PDSA cycle was successful in improving the quality of prescription writing among interns posted in the Department of Psychiatry. There is a need to implement more PDSA cycles to improve the quality still further.", "keywords": [ "Prescription", "intern", "rational prescription writing", "Quality Improvement Project", "Audit", "Psychiatry" ], "content": "Introduction\n\nThe problem of irrational drug prescription is prevalent worldwide.1 According to a World Health Organization (WHO) estimate, more than 50% of drugs are prescribed, dispensed, or sold inappropriately, and 50% of patients fail to take them correctly (https://www.who.int/activities/promoting-rational-use-of-medicines).\n\nThe prescription is the written instruction of the physician communicated to the patient. Writing rational prescriptions is an important skill for physicians. Poorly written prescriptions can negatively affect patient treatment, including worsening of the disease and making the prescriber liable to influence, which may lead to irrational prescribing.2\n\nRational prescribing requires a logical approach that involves several key steps, including making the diagnosis, determining the therapeutic goals, consideration of treatment options, deciding the best treatment option, prescription and monitoring.3\n\nIn clinical practice, a doctor must be able to diagnose and immediately decide on the drug to be prescribed. Furthermore, patients may have different age, sex, and sociocultural features that affect the drugs prescribed.\n\nInternship is a period of one year during which newly qualified Bachelor of Medicine and Bachelor of Surgery (MBBS) graduates practice under the guidance of senior doctors in designated hospitals. Studying the pattern of their prescriptions and suggesting changes is of utmost importance in improving the quality of patient care and enhancing the undergraduate medical education of interns.\n\nMost studies on the impact of medical errors, including prescription writing errors, have been conducted in developed countries. Medical errors are a leading cause of mortality in the United States. Studies conducted in Asian countries have reported varying rates of errors when writing prescriptions. These values range from 7% to 35.4%.4\n\nA study conducted in Bangladesh found that 78% of drugs were prescribed by their generic names, 85% complied with the essential drug list, and 81% were dispensed according to prescription.5\n\nIn a study conducted in private hospitals in western India, prescriptions were analyzed according to WHO prescribing indicators. It was found that out of the 250 prescriptions analyzed, patient details were not written in 100% of prescriptions. Drug dosage, instructions regarding drug administration, and the duration of the treatment were not completely written in 90%, 74%, and 80% of prescriptions studied, respectively. The prescriber’s medical registration number was not mentioned in any of the prescriptions.6\n\nIn a study carried out in the non-governmental organization sector in West Bengal, India, it was found that the majority were signed, legible, and complete with respect to age/gender data, 95.5% used Latin abbreviations, and 7.7% mentioned neither signs, symptoms, nor diagnosis. Irrational fixed-dose combinations were used in 45.6% of the prescribed drugs. Only 45.7% of prescribed drugs conformed to the WHO model list of essential drugs.7\n\nIn a study of selected rural pharmacies in southern India, it was found that 91.4% of prescriptions did not contain any drug prescribed by its generic name.8\n\nIn a study in The Oxford Dental College and Hospital, Bangalore, prescriptions were analyzed for several features, such as patient details, doctor’s information, and drug information. Prescriptions written by undergraduate students were better than those written by interns and postgraduates (PGs).9\n\nThe prescriptions written by interns were studied in a primary health center in India, and it was found that 34.97% of drugs were prescribed by generic name and 58.7% of drugs prescribed were from the essential drug list of India.10\n\nAfter studying the literature available regarding the state of prescription writing, this Quality Improvement (QI) project was conceptualized. This QI project sought to understand and bring about significant improvements in the quality of prescriptions written by interns posted in the Department of Psychiatry.\n\nA process map was created that outlined the process through which the interns were posted in the Department of Psychiatry to write prescriptions (Figure 1).\n\nWe carefully studied the process and brainstormed possible causes of the poor quality of prescription writing among interns with the help of a fishbone diagram (Figure 2).\n\nThe aim of our QI project was to improve the quality of prescription writing among interns in the psychiatry department.\n\nOur Specific, Measurable, Applicable, Realistic and Timely (SMART) aim was to improve the mean prescription scores.\n\n\nMethods\n\nStudy setting: This study was conducted in the outpatient Department of Psychiatry, Kasturba Medical College Hospital, Attavar, Mangaluru.\n\nStudy design: Quality Improvement Project.\n\nStudy participants: MBBS interns in the Department of Psychiatry, Kasturba Medical College Hospital, Attavar, Mangaluru.\n\n\n\na) MBBS Interns posted in the Department of Psychiatry, willing to grant consent.\n\nb) Patients above the age of 18 years attending Psychiatry Outpatient department.\n\n\n\na) Postgraduates and consultants writing prescriptions for the patients\n\nb) Interns posted in other specialties\n\nStudy duration: 1 year from November 2022 till August 2023\n\nThe sample size for the study is 85 with 5% level of significance.\n\nZ = 1.96 is standard normal value at 5% level of significance\n\nΡ = Proportion = 67%\n\nd = Absolute Precision = 10%\n\nSampling method: Convenient sampling method.\n\nMethodology: As a part of compulsory rotation during internship, the interns are posted in the Psychiatry department for 15 days. We used the Plan-Do-Study-Act (PDSA) model to conduct this Quality Improvement Project. The PDSA model is a four-step model used to improve the process or bring about a change in a system. We divided our study into two phases. The pre intervention phase and PDSA cycle 1 were repeated for each batch of interns who participated in this study.\n\nThe PDSA model used for this study consisted of the following:\n\nPlan: Based on the results of the pre intervention phase, we planned to implement the intervention.\n\nDo: We implemented the intervention\n\nStudy: We studied the result of the intervention\n\nAct: Using an educational handout on prescription writing during internship\n\nOn the first day of the psychiatry postings, the interns were informed about the study and written informed consent was obtained from them. They were instructed to create copies of the prescriptions that they wrote with the help of a carbon sheet which was provided with the empty prescriptions. Informed written consent was obtained from the patients whose prescriptions were used in the audit.\n\nFor the first 5 days of Psychiatry posting, carbon copy of the prescriptions were collected before giving the original prescription to the patients.\n\nOn day 6 of the posting, the interns were given an educational handout which was based on the WHO practical manual guide to good prescribing that explained the format of the prescriptions to be followed. The investigators also explained regarding the format of the prescription writing to be followed by the interns.\n\nOn days 7–15, the interns again created copies of the prescriptions they were writing with the help of carbon sheets provided with the empty prescriptions, and these were collected before giving the original prescription to the patients.\n\n\n\na) Criteria for assessing the quality of prescription: To determine the quality of prescription writing, we established certain criteria. These were based on WHO guidelines.11 The Medical Council of India (MCI) ideal prescription format was also used to create the criteria (https://delhimedicalcouncil.org/images/Latest-News/modalprescription.pdf). The prescriptions were evaluated using the following criteria: If the criteria are present, 1 point will be awarded, and if the criteria are absent, 0 points will be awarded. Then, the final score of that prescription was calculated by adding the points. The scores range from 0 to 14.\n\n1. Date\n\n2. Name of patient\n\n3. Age of patient\n\n4. Contact number of the patient\n\n5. Gender of the patient\n\n6. Generic name of the drug\n\n7. Drug name in capital letter\n\n8. Strength\n\n9. Dosage form\n\n10. Route of Administration\n\n11. Total quantity of drugs to be given by pharmacy\n\n12. Number of days to take the drug\n\n13. Number of times to take the drug in a day\n\n14. Signature or initials of the prescriber\n\nb) Educational handout: This was based on the WHO Practical Manual Guide to Good Prescribing.11\n\nBiological materials required - none\n\nStatistical analysis was performed using IBM SPSS Statistics for Windows version 29 (IBM Corp., Armonk, N.Y., USA).12 The copyright license has been obtained by the institution. An open source alternative is Jamovi version 1.x.13 Means and standard deviations were calculated. The t-test was used for continuous variables, and the chi-square test was used for categorical variables. Statistical significance was set at p value<0.05.\n\nThe data analysis in this study was not pre-registered.\n\n\nResults\n\nIn total, 91 prescriptions were analyzed. The mean score of prescriptions written by the interns was 9.54 ± 1.003. The mean scores across the various criteria were as follows (Table 1):17\n\nA total of 84 prescriptions were analyzed. The mean score of prescriptions written by the interns was 10.26±0.746. The mean scores across the various criteria were as follows (Table 2):\n\nThere was a significant improvement in the mean total score from 9.54±1.003 to 10.26±0.746 (p<0.01). There was an improvement of 7.54% after PDSA cycle 1.\n\nThere was also a significant improvement in the following criteria: date (p<0.01), sex of patient (p=0.13), generic name of the drug (p=0.01), drug name in capital letters (p=0.045), dosage form (p=0.07), and total quantity of drugs to be administered by the pharmacy (p<0.01).\n\nThere was no significant improvement in the Age of Patient (p=0.083), strength (p=0.336), route of administration (p=0.159), number of days to take the drugs (p=0.897), number of times to take the drug in a day (p=0.174), or signature or initials of the prescriber (p=0.196).\n\n\nDiscussion\n\nIn this QI project, we attempted to ascertain methods to improve the quality of prescriptions written by interns posted in the Department of Psychiatry.\n\nIn the first PDSA cycle, we tried to fill in the knowledge gap with the help of a brochure and a verbal explanation of the format of prescription writing as prescribed by the MCI as well as the guidelines stipulated by the WHO.\n\nAfter PDSA cycle 1, we found a significant improvement in the total scores of the prescriptions as well as a significant improvement across several criteria, including date, gender of patient, generic name of the drug, drug name in capital letters, dosage form, and total quantity of drugs to be given by the pharmacy.\n\nThe aim of this QI project was to improve prescription writing scores. After the first cycle, a 7.54% improvement was observed.\n\nA study conducted in The Oxford Dental College and Hospital, Bangalore, found that the patient’s name (95.6%), age (91.2%), and gender (91.6%) were written by the majority of the students. The patients’ outpatient number (0%), address (0%), and contact number (0.4%) were missing in almost all prescriptions, which is in agreement with our study where the name, age, and gender were mentioned in 100% of the prescriptions collected.9\n\nFurthermore, a study was conducted regarding the impact of patient-based teaching in improving the prescription writing skills of II MBBS students. This study showed that patient-based teaching improves responsibility, focus, and memory.14\n\nIn our study, our informal teaching session at the end of PDSA cycle 1 revealed a similar improvement, which may be attributed to the orientation given.\n\nIn another study carried out at Nobel Medical Teaching Hospital, Biratnagar, Nepal, there was a 0% error in the patient’s name, age, and sex, which was similar to our findings.15\n\nHowever, the findings of other studies are different from ours. In a study conducted at Oxford Dental College and Hospital, Bangalore, it was found that the parameters that were lacking the most were dosage instructions, strength of the drugs, and duration of the drugs, which was not the case in our study.9 According to our study, the total quantity of drugs to be given by the pharmacist, dosage form, and generic names were not mentioned.\n\nThe lack of generic names written in our study could be because interns have very little time to write prescriptions or because psychiatric medications are not as common as analgesics or gastritis medicines; hence, additional work and reading is required to become aware of them, and either the interns did not have an incentive to learn this or did not have enough time to cover this aspect of prescription writing.\n\nIn our study, the criteria for which there was no significant improvement were criteria that already had high scores in the pre intervention phase.\n\nFurthermore, another study in Patna, where prescriptions from different OPDs in different departments were audited, showed that only 10% of the prescriptions contained drugs prescribed in their generic name. Frequency, route, and duration of drug administration were mentioned in 86.7%, 79.3%, and 69.6% of the prescriptions, respectively. Among the prescriptions, 42.5% were easily legible. Hence, it was not in accordance with the WHO standards.16\n\nLegibility was not a criterion for this study. If we had to look at it subjectively, none of the prescriptions were difficult to read by the people undertaking the study, thus giving us a higher score in that field than in the study mentioned above.\n\nAn important criterion that was judged in the study conducted in Nobel Medical Teaching Hospital, Biratnagar, were 2 same drugs or the same category mentioned in the same prescription, which accounted for 0.5% of their total errors.15 This category was not judged in our study because it was not a criterion mentioned in the MCI ideal prescription and it is an indicator of skills and not methodology, which was what our study mostly focused on.\n\nInstructing interns to make copies of their prescriptions may have led them to pay closer attention to their prescription writing, leading to a biased result.\n\n\nConclusion\n\nThis Quality Improvement project enabled us to understand that brochures regarding prescription writing and verbal explanations are useful tools for improving the quality of prescriptions written by interns in the department of psychiatry. Therefore, there is a need to find more innovative methods to improve the quality of prescriptions. There is a need to improve interns’ training by emphasizing rational prescription writing during internships. Further studies are required to determine whether this method is universally applicable.\n\nThe protocol was approved by the Institutional Ethics Committee Kasturba Medical College, Mangalore (Reg. No. ECR/541/Inst/KA/2014/RR-20) before commencement of the Quality Improvement Project and Ethical Approval was obtained. The protocol number of the study was IECKMCMLR-10/2022/423. The date of approval was 19/10/2022. Written informed consent for the study was obtained from the interns and the patients/relatives of the patients, respectively. Participants had the right to withdraw from the study at any time.", "appendix": "Data availability\n\nOpen Science Framework: Prescription writing quality of psychotropic agents in MBBS interns at a private medical college hospital in Mangaluru. https://doi.org/10.17605/OSF.IO/HZGFC. 17\n\nThis project contains the following data:\n\n• Intern prescription writing scores xlsx (scores of prescriptions written by interns during the preintervention phase and PDSA Cycle 1).\n\n• Educational Handout.docx (the educational handout given to interns during PDSA Cycle 1).\n\n• Intern Consent Form.docx\n\n• Patient Consent Form.docx\n\n• Checklist for Prescription writing quality of psychotropic agents in MBBS interns in a private medical college hospital in Mangaluru.docx\n\nThis study was based on the SQUIRE (Standards for Quality Improvement Reporting Excellence) guidelines. 18\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 postgraduate students of the Department of Psychiatry, Kasturba Medical College Mangaluru, Manipal Academy of Higher Education, India.\n\n\nReferences\n\nHogerzeil H: Promoting rational prescribing: An international perspective. Br. J. Clin. Pharmacol. 1995 Jan; 39(1): 1–6. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nImran M, Doshi C, Kharadi D: Time to teach basic and regulatory aspects of art of prescription writing for better doctor-patient safety and keeping communication accessible and straight. Daru. 2019; 28(1): 25–32. PubMed Abstract \| Publisher Full Text\n\nRissmann R, Dubois EA, Franson KL, et al.: Concept-based learning of Personalized Prescribing. Br. J. Clin. Pharmacol. 2012 Sept 5; 74(4): 589–596. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSalmasi S, Khan TM, Hong YH, et al.: Medication Errors in the Southeast Asian Countries: A Systematic Review. PLoS One. 2015 Sep 4; 10(9): e0136545. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGuyon AB, Barman A, Ahmed JU, et al.: A baseline survey on use of drugs at the primary health care level in Bangladesh. Bull. World Health Organ. 1994; 72(2): 265–271. PubMed Abstract\n\nShelat PR, Kumbar SK: Analysis of Out Door Patients’ Prescriptions According to World Health Organization (WHO) Prescribing Indicators Among Private Hospitals in Western India. J. Clin. Diagn. Res. 2015 Mar; 9(3): FC01–FC04. PubMed Abstract \| Publisher Full Text\n\nHazra A, Tripathi SK, Alam MS: Prescribing and dispensing activities at the health facilities of a non-governmental organization. Natl. Med. J. India. 2000; 13(4): 177–182. PubMed Abstract\n\nAravamuthan A, Arputhavanan M, Subramaniam K, et al.: Assessment of current prescribing practices using World Health Organization core drug use and complementary indicators in selected rural community pharmacies in Southern India. J. Pharm. Policy Pract. 2017; 10: 1. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDyasanoor S, Urooge A: Insight Into Quality of Prescription Writing - An Instituitional Study. J. Clin. Diagn. Res. 2016 Mar; 10(3): ZC61–ZC64. PubMed Abstract \| Publisher Full Text\n\nBanerjee I, Bhadury T: Prescribing pattern of interns in a primary health center in India. J Basic Clin Pharm. 2014 Mar; 5(2): 40–43. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nde VTP: Guide to good prescribing a practical manual. Geneva: World Health Organization; 1995.\n\nIBM Corp: IBM SPSS Statistics for Windows, Version 29. 0. Armonk, NY: IBM Corp.\n\nThe Jamovi Project: Jamovi. (Version 1.x) [Computer Software].\n\nThenrajan P, Murugan PR: Impact of patient-based teaching in improving prescription writing skills of II MBBS students. Int. J. Appl. Basic Med. Res. 2016 Jul; 6(3): 174–177. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAnsari M, Neupane D: Study on determination of errors in prescription writing: A semi-electronic perspective. Kathmandu Univ. Med. J (KUMJ). 2009 Jul 1; 7(27): 238–241. PubMed Abstract \| Publisher Full Text\n\nMishra S, Pathak PK, Ahmad S, et al.: Quality of Prescription Writing Practices in a Tertiary Care Teaching Hospital of Bihar: A Cross-Sectional Study. Int. J. Health Syst. Implement. Res. 2020 Sep 1; 4(2): 24–33.\n\nPoorva: Prescription writing quality of psychotropic agents in MBBS interns in a private medical college hospital in Mangaluru (Internet). [Dataset]. OSF. 2023. osf.io/hzgfc.\n\nOgrinc G, Davies L, Goodman D, et al.: SQUIRE 2.0 (Standards for QUality Improvement Reporting Excellence): Revised publication guidelines from a detailed consensus process. BMJ Qual. Saf. 2016; 25: 986–992. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "313887", "date": "16 Aug 2024", "name": "Vandana Gaur", "expertise": [ "Reviewer Expertise Human 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\nThe introduction provides a clear rationale for the study, emphasizing the importance of accurate prescription writing in clinical practice, particularly for psychotropic agents, which are associated with complex dosing and potential side effects. The study's objectives are well-defined, focusing on evaluating the prescription writing quality of psychotropic agents among MBBS interns. The results section (assuming it is well-presented) likely provides valuable insights into the quality of prescription writing among interns. It is essential that this section includes data on common errors and areas where interns perform well, as this information could inform future training programs. The discussion should ideally interpret the results in the context of existing literature, highlighting both strengths and weaknesses in the prescription writing practices of the interns. It should discuss the implications for patient safety and medical education. The study addresses an important issue in medical education and patient safety. The focus on psychotropic agents is particularly relevant, given the complexities involved in prescribing these medications. The methodology appears sound, and the study has the potential to provide valuable insights.\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": "12961", "date": "20 Dec 2024", "name": "Sharanya Shetty", "role": "Author Response", "response": "Dear Madam, Thank you for your valuable feedback on our manuscript. We appreciate your insightful comments and suggestions, which has undoubtedly contributed to improving the quality of our manuscript. We have tried our best to address your concerns in the modified manuscript. We have taken your feedback regarding the formatting of the results section and formatted the results section to include a table which has the common errors and areas in which the interns perform well in the form of the mean scores before and after the intervention. We have also included the implications of this study under the conclusions section. Thank you again for your valuable feedback. Regards Dr Sharanya B Shetty" } ] }, { "id": "313883", "date": "04 Sep 2024", "name": "Dr Shashwath Sathyanath", "expertise": [ "Reviewer Expertise Deaddiction psychiatry", "Consultation-liaison Psychiatry", "Mood Disorders." ], "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\nSummary of the article\nThis Quality Improvement (QI) project aimed to enhance the quality of prescription writing among MBBS interns in the Department of Psychiatry at Kasturba Medical College Hospital, Mangaluru. The study addressed the prevalent issue of irrational drug prescriptions, which contributes to adverse patient outcomes and inefficiencies in healthcare. The project involved a pre-intervention assessment of prescription quality, followed by an intervention based on educational materials and feedback, and then a post-intervention assessment to measure improvements.\nDetailed Review\n1. Work Presentation and Literature Citation\n\nResponse: Yes\n\nSummary: The work is presented clearly and logically. The introduction provides a comprehensive background on the problem of irrational drug prescribing and cites relevant literature. However, there is scope of citing few newer literatures from the studies in the last 3 years\n2. Study Design and Technical Soundness\n\nResponse: Yes\n\nSummary: The study design is appropriate for the Quality Improvement (QI) project, utilizing the Plan-Do-Study-Act (PDSA) model. The project addresses a clear problem, implements an intervention, and evaluates its effectiveness in a systematic manner. However, the title of the article could mention the type of study design for the ease of the readers.\n3. Replication Details\n\nResponse: Yes\n\nSummary : The methods and analysis are described in sufficient detail to allow replication.\n\nThe article includes detailed descriptions of the intervention (educational handouts and verbal explanations), the assessment criteria for prescription quality, and the statistical methods used for analysis. The process maps and fishbone diagrams are provided, facilitating replication of the study.\n4. Statistical Analysis\n\nResponse: Partly\n\nSummary: The statistical analysis is mostly appropriate but could benefit from some improvements.\n\nThe use of means, standard deviations, t-tests, and chi-square tests is generally suitable. However, it may be beneficial to include additional analyses or sensitivity checks to confirm the robustness of the findings. In addition, the tables mentioned are not complete as it lacks the p values mentioned in the writings.\n5. Availability of Source Data\n\nResponse: Yes\n\nSummary: The source data underlying the results are available for full reproducibility.\n\nThe data, including prescription writing scores and the educational handout, are accessible via the Open Science Framework link provided. This transparency supports reproducibility and verification of the study’s findings.\n6. Support for Conclusions\n\nResponse: Yes\n\nSummary: The conclusions are adequately supported by the results.\n\nThe study demonstrates a significant improvement in prescription quality post-intervention, with specific criteria showing notable enhancements. The discussion contextualizes these findings within existing literature, reinforcing the study’s conclusions.\nRecommendations This study can be considered for indexed after a few minor corrections.\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": "12962", "date": "20 Dec 2024", "name": "Sharanya Shetty", "role": "Author Response", "response": "Dear Sir, Thank you for your valuable feedback on our manuscript. We appreciate your insightful comments and suggestions, which has undoubtedly contributed to improving the quality of our manuscript. We have tried our best to address your concerns in the modified manuscript. Taking your feedback into consideration, we have included newer studies in the introduction section. We have also changed the title of the study to accurately reflect the study design. We have merged our tables and also included p value in the table as per your suggestion. Thank you again for your valuable feedback. Regards Dr Sharanya B Shetty" } ] }, { "id": "267459", "date": "12 Sep 2024", "name": "Sanjeev Badiger", "expertise": [ "Reviewer Expertise Communicable diseases and behavioral 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 topic selected by the author is relevant to the present scenario , as prescription is considered as an important Document. This article highlights the need for quality prescription writing and the need for improvement in prescription writing . Studying the pattern of their prescriptions and suggesting changes is of utmost importance in improving the quality of patient care and enhancing the undergraduate medical education of interns.\nMajor Points :\nMethodology : study sample size and study design is appropriate. But the capture other factors influencing the prescription pattern can be captured by adding qualitative component like FGD is helpful to improve the quality of the study\n\nMinor points : Topic : Relevant and well phrased Introduction: Adequate and highlighted the need for the study Data analysis ,Discussion and conclusions are adequate\n\nThis research article can be accepted for 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? 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": "12963", "date": "20 Dec 2024", "name": "Sharanya Shetty", "role": "Author Response", "response": "Dear Sir, Thank you for your valuable feedback on our manuscript. We appreciate your insightful comments and suggestions, which has undoubtedly contributed to improving the quality of our manuscript. We have tried our best to address your concerns in the modified manuscript. We understand that focused group discussion would have added further insights on other qualitative aspects, which we will plan to incorporate in the next cycle. We have added the same as a limitation of our study. Thank you again for your valuable feedback. Regards Dr Sharanya B Shetty" } ] } ]	1	https://f1000research.com/articles/13-269
https://f1000research.com/articles/13-1545/v1	20 Dec 24	{ "type": "Data Note", "title": "A guide to selecting high-performing antibodies for ADNP (UniProt ID: Q9H2P0) for use in western blot, immunoprecipitation, and immunofluorescence", "authors": [ "Vera Ruíz Moleón", "Charles Alende", "Maryam Fotouhi", "Riham Ayoubi", "Sara González Bolívar", "Carl Laflamme", "NeuroSGC/YCharOS/EDDU collaborative group", "ABIF consortium", "Vera Ruíz Moleón", "Charles Alende", "Maryam Fotouhi", "Riham Ayoubi", "Sara González Bolívar" ], "abstract": "ADNP is a multifunctional protein involved in chromatin remodeling, transcription, and microtubule interaction, playing a critical role in brain development, with mutations linked to ADNP-Related Disorder. Here we have characterized seven ADNP commercial antibodies for western blot, immunoprecipitation, and immunofluorescence using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. These studies are part of a larger, collaborative initiative seeking to address antibody reproducibility issues by characterizing commercially available antibodies for human proteins and publishing the results openly as a resource for the scientific community. While use of antibodies and protocols vary between laboratories, we encourage readers to use this report as a guide to select the most appropriate antibodies for their specific needs.", "keywords": [ "Q9H2P0", "ADNP", "Activity-Dependent Neuroprotective Protein", "Activity-Dependent Neuroprotector homeobox Protein", "antibody characterization", "antibody validation", "western blot", "immunoprecipitation", "immunofluorescence" ], "content": "Introduction\n\nADNP (Activity-Dependent Neuroprotector Homeobox Protein) is part of the SWI/SNF chromatin remodeling complex, a key regulator of RNA transcription and splicing.1 It is a multifunctional protein characterized by nine Zinc fingers, a nuclear localization signal and a DNA-binding homeobox domain,2 which collectively support its role as a transcription factor.3 The protein also contains a small NAP motif (amino acids “NAPVSIPQ”) site that contributes to its interaction with microtubules.4 Given its function in brain development5 and other roles in the cell,6 ADNP deletion is lethal in mice and haploinsufficiency results in cognitive impairment.7 Mutations in the ADNP gene are associated with several congenital abnormalities and intellectual disabilities, including Helsmoortel-Van der Aa Syndrome8 and ADNP-related Autism Spectrum Disorder,9 collectively termed “ADNP-Related Disorder”.10\n\nThis research is part of a broader collaborative initiative in which academics, funders and commercial antibody manufacturers are working together to address antibody reproducibility issues by characterizing commercial antibodies for human proteins using standardized protocols, and openly sharing the data.11–13 Here we evaluated the performance of seven commercial antibodies for ADNP for use in western blot, immunoprecipitation, and immunofluorescence, enabling biochemical and cellular assessment of ADNP properties and function. The platform for antibody characterization used to carry out this study was endorsed by a committee of industry and academic representatives. It consists of identifying human cell lines with adequate target protein expression and the development/contribution of equivalent knockout (KO) cell lines, followed by antibody characterization procedures using most commercially available antibodies against the corresponding protein. The standardized consensus antibody characterization protocols are openly available on Protocol Exchange, a preprint server (DOI: 10.21203/rs.3.pex-2607/v1).14\n\nThe authors do not engage in result analysis or offer explicit antibody recommendations. Our primary aim is to deliver top-tier data to the scientific community, grounded in Open Science principles. This empowers experts to interpret the characterization data independently, enabling them to make informed choices regarding the most suitable antibodies for their specific experimental needs. Guidelines on how to interpret antibody characterization data found in this study are featured on the YCharOS gateway.15\n\n\nResults and discussion\n\nOur standard protocol involves comparing readouts from wild type (WT) and KO cell lines.14,16 In the absence of commercially available KO cell lines, siRNA technology can be employed to knockdown (KD) the target gene.17,18 To determine which cell line demonstrates high expression of ADNP and thus be appropriate for KD, the first step is to identify a cell line that expresses sufficient levels of a given protein to generate a measurable signal using antibodies. To this end, we examined the DepMap (Cancer Dependency Map Portal, RRID:SCR_017655) transcriptomics database to identify cell lines that express the target at levels greater than 2.5 log2 (transcripts per million “TPM” + 1), which we have found to be a suitable cut-off.11 As a result, DMS 53 which expresses the ADNP transcript at 5.7 was identified as a suitable cell line to use here. A non-targeting siRNA pool was used to treat the DMS 53 control (ctrl) cells, while ADNP was KD using a pool of siRNA targeting this gene.\n\nTo screen all seven antibodies by western blot, ctrl and ADNP KD protein lysates were ran on SDS-PAGE, transferred onto nitrocellulose membranes, and then probed with the ADNP antibodies in parallel ( Figure 1).\n\nLysates of DMS 53 ctrl and ADNP KD were prepared, and 30 μg of protein were processed for western blot with the indicated ADNP antibodies. The Ponceau stained transfers of each blot are presented to show equal loading of ctrl and KD lysates and protein transfer efficiency from the acrylamide gels to the nitrocellulose membrane. Antibody dilutions were chosen according to the recommendations of the antibody supplier. Antibody dilution used: ab300114 at 1/1000, A4546 at 1/1000, NBP1-89236 at 1/250 (0.4 μg/ml), NBP2-97747 at 1/1000, 17987-1-AP at 1/1000, 702911 at 1/1000, PA5-52286 at 1/250 (0.4 μg/ml). Predicted band size: predicted band size 123.5 kDa. Recombinant antibody.\n\nWe then assessed the capability of all seven antibodies to capture ADNP from DMS 53 protein extracts using immunoprecipitation techniques, followed by western blot analysis. For the immunoblot step, a specific ADNP antibody identified previously (refer to Figure 1) was selected. Equal amounts of the starting material (SM) and the unbound fractions (UB), as well as the whole immunoprecipitate (IP) eluates were separated by SDS-PAGE ( Figure 2).\n\nDMS 53 WT lysates were prepared, and immunoprecipitation was performed using 1 mg of lysate and 2.0 μg of the indicated ADNP antibodies pre-coupled to Dynabeads protein A or protein G. Samples were washed and processed for western blot with the indicated ADNP antibody. For western blot, 702911 was used at 1/500. The Ponceau stained transfers of each blot are shown. SM=6% starting material; UB=6% unbound fraction; IP=immunoprecipitate, Recombinant antibody.\n\nFor immunofluorescence, the seven antibodies were screened using a mosaic strategy. First, DMS 53 ctrl and ADNP KD cells were labelled with different fluorescent dyes in order to distinguish the two cell lines, and the ADNP antibodies were evaluated. Both ctrl and KD lines were imaged in the same field of view to reduce staining, imaging and image analysis bias ( Figure 3). Quantification of immunofluorescence intensity in hundreds of ctrl and KD cells was performed for each antibody tested, and the images presented in Figure 3 are representative of this analysis.14\n\nDMS 53 ctrl and ADNP KD cells were labelled with a green or a far-red fluorescent dye, respectively. Ctrl and KD cells were mixed and plated to a 1:1 ratio on coverslips. Cells were stained with the indicated ADNP antibodies and with the corresponding Alexa-fluor 555 coupled secondary antibody including DAPI. Acquisition of the blue (nucleus-DAPI), green (ctrl), red (antibody staining) and far-red (KD) channels was performed. Representative images of the merged blue and red (grayscale) channels are shown. Ctrl and KD cells are outlined with green and magenta dashed lines, respectively. When an antibody was recommended for immunofluorescence by the supplier, we tested it at the recommended dilution. The rest of the antibodies were tested at 1 and 2 μg/ml and the final concentration was selected based on the detection range of the microscope used and a quantitative analysis not shown here. Antibody dilution used: ab300114 at 1/500, A4546 at 1/700, NBP1-89236 at 1/100, NBP2-97747 at 1/300, 17987-1-AP at 1/600, 702911 at 1/500, PA5-52286 at 1/100. Bars = 10 μm. Recombinant antibody.\n\nIn conclusion, we have screened seven ADNP commercial antibodies by western blot, immunoprecipitation, and immunofluorescence by comparing the signal produced by the antibodies in human DMS 53 ctrl and ADNP KD cells. To assist users in interpreting antibody performanyce, Table 3 outlines various scenarios in which antibodies may perform in all three applications.11 Several high-quality and renewable antibodies that successfully detect ADNP were identified in all applications. Researchers who wish to study ADNP in a different species are encouraged to select high-quality antibodies, based on the results of this study, and investigate the predicted species reactivity of the manufacturer before extending their research.\n\nInherent limitations are associated with the antibody characterization platform used in this study. Firstly, the YCharOS project focuses on renewable (recombinant and monoclonal) antibodies and does not test all commercially available ADNP antibodies. YCharOS partners provide approximately 80% of all renewable antibodies, but some top-cited polyclonal antibodies may not be available through these partners.\n\nSecondly, the YCharOS effort employs a non-biased approach that is agnostic to the protein for which antibodies have been characterized. The aim is to provide objective data on antibody performance without preconceived notions about how antibodies should perform or the molecular weight that should be observed in western blot. As the authors are not experts in ADNP only a brief overview of the protein’s function and its relevance in disease is provided. ADNP experts are invited to analyze and interpret observed banding patterns in western blots and subcellular localization in immunofluorescence.\n\nThirdly, YCharOS experiments are not performed in replicates primarily due to the use of multiple antibodies targeting various epitopes. Once a specific antibody is identified, it validates the protein expression of the intended target in the selected cell line, confirms the lack of protein expression in the KO cell line and supports conclusions regarding the specificity of the other antibodies. All experiments are performed using master mixes, and meticulous attention is paid to sample preparation and experimental execution. In IF, the use of two different concentrations serves to evaluate antibody specificity and can aid in assessing assay reliability. In instances where antibodies yield no signal, a repeat experiment is conducted following titration. Additionally, our independent data is performed subsequently to the antibody manufacturers internal validation process, therefore making our characterization process a repeat.\n\nLastly, as comprehensive and standardized procedures are respected, any conclusions remain confined to the experimental conditions and cell line used for this study. The use of a single cell type for evaluating antibody performance poses as a limitation, as factors such as target protein abundance significantly impact results.14 Additionally, the use of cancer cell lines containing gene mutations poses a potential challenge, as these mutations may be within the epitope coding sequence or other regions of the gene responsible for the intended target. Such alterations can impact the binding affinity of antibodies. This represents an inherent limitation of any approach that employs cancer cell lines.\n\n\nMethod\n\nThe standardized protocols used to carry out this KO cell line-based antibody characterization platform was established and approved by a collaborative group of academics, industry researchers and antibody manufacturers. The detailed materials and step-by-step protocols used to characterize antibodies in western blot, immunoprecipitation and immunofluorescence are openly available on Protocol Exchange, a preprint server (DOI: 10.21203/rs.3.pex-2607/v1).14 Brief descriptions of the experimental setup used to carry out this study can be found below.\n\nCell lines used and primary antibodies tested in this study are listed in Tables 1 and 2, respectively. To ensure that the cell lines and antibodies are cited properly and can be easily identified, we have included their corresponding Research Resource Identifiers, or RRID.19,20 DMS 53 cells were treated with the ON-TARGETplus Human ADNP siRNA from Horizon Discovery, cat. number L-012857-01-0005. Ctrl DMS 53 cells were treated with the ON-TARGETplus Non-targeting Control Pool, cat. number D-001810-10-05. Lipofectamine RNAiMAX (Thermo Fisher Scientific, cat. number 13778030) was used to transfect the siRNA following the manufacturer’s protocol.\n\n** Recombinant antibody, n/a=not available.\n\nPeroxidase-conjugated goat anti-rabbit antibody is from Thermo Fisher Scientific, cat. number 65-6120. Alexa-555-conjugated goat anti-rabbit secondary antibody is from Thermo Fisher Scientific, cat. number A-21429. Peroxidase-conjugated Protein A for IP detection is from Cell Signaling Technology, cat. number 12291.\n\nDMS 53 ctrl and ADNP KD cells were collected in RIPA buffer (25mM Tris-HCl pH 7.6, 150 mM NaCl, 1% NP-40, 1% sodium deoxycholate, 0.1% SDS) (Thermo Fisher Scientific, cat. number 89901) supplemented with 1× protease inhibitor cocktail mix (MilliporeSigma, cat. number P8340). Lysates were sonicated briefly and incubated 30 min on ice. Lysates were spun at ~110,000 × g for 15 min at 4°C and equal protein aliquots of the supernatants were analyzed by SDS-PAGE and western blot. BLUelf prestained protein ladder (GeneDireX, cat. number PM008-0500) was used.\n\nWestern blots were performed with precast midi 4-20% Tris-Glycine polyacrylamide gels (Thermo Fisher Scientific, cat. number WXP42012BOX) ran with Tris/Glycine/SDS buffer (Bio-Rad, cat. number 1610772), loaded in Laemmli loading sample buffer (Thermo Fisher Scientific, cat. number AAJ61337AD) and transferred on nitrocellulose membranes. Proteins on the blots were visualized with Ponceau S staining (Thermo Fisher Scientific, cat. number BP103-10) which is scanned to show together with individual western blot. Blots were blocked with 5% milk for 1 hr, and antibodies were incubated O/N at 4°C with 5% milk in TBS with 0,1% Tween 20 (TBST) (Cell Signalling Technology, cat. number 9997). Following three washes with TBST, the peroxidase conjugated secondary antibody was incubated at a dilution of ~0.2 μg/ml in TBST with 5% milk for 1 hr at room temperature followed by three washes with TBST. Membranes were incubated with Pierce ECL (Thermo Fisher Scientific, cat. number 32106) or Clarity Western ECL Substrate (Bio-Rad, cat. number 1705061) prior to detection with the iBright™ CL1500 Imaging System (Thermo Fisher Scientific, cat. number A44240).\n\nAntibody-bead conjugates were prepared by adding 2.0 μg to 500 μl of Pierce IP Lysis Buffer from Thermo Fisher Scientific (cat. number 87788) in a microcentrifuge tube, together with 30 μl of Dynabeads protein A- (rabbit antibodies) (Thermo Fisher Scientific, cat. number 10002D). Antibody NBP2-97747 is at an unknown concentration, and 20 μl were tested in the IP. Tubes were rocked for ~1 h at 4°C followed by two washes to remove unbound antibodies.\n\nDMS 53 WT were collected in Pierce IP buffer (25 mM Tris-HCl pH 7.4, 150 mM NaCl, 1 mM EDTA, 1% NP-40 and 5% glycerol) supplemented with protease inhibitor. Lysates were rocked 30 min at 4°C and spun at 110,000 × g for 15 min at 4°C. 0.5 ml aliquots at 2 mg/ml of lysate were incubated with an antibody-bead conjugate for ~1 h at 4°C. The unbound fractions were collected, and beads were subsequently washed three times with 1.0 ml of IP buffer and processed for SDS-PAGE and western blot on precast midi 4-20% Tris-Glycine polyacrylamide gels. Protein A:HRP was used as a secondary detection system at a concentration of 0.5 μg/ml.\n\nDMS 53 ctrl and ADNP KD cells were labelled with a green and a far-red fluorescence dye, respectively (Thermo Fisher Scientific, cat. number C2925 and C34565). The nuclei were labelled with DAPI (Thermo Fisher Scientific, cat. Number D3571) fluorescent stain. Ctrl and KD cells were plated on 96-well plate with optically clear flat-bottom (Perkin Elmer, cat. number 6055300) as a mosaic and incubated for 24 hrs in a cell culture incubator at 37oC, 5% CO2. Cells were fixed in 4% paraformaldehyde (PFA) (VWR, cat. number 100503-917) in phosphate buffered saline (PBS) (Wisent, cat. number 311-010-CL). Cells were permeabilized in PBS with 0,1% Triton X-100 (Thermo Fisher Scientific, cat. number BP151-500) for 10 min at room temperature and blocked with PBS with 5% BSA, 5% goat serum (Gibco, cat. number 16210-064) and 0.01% Triton X-100 for 30 min at room temperature. Cells were incubated with IF buffer (PBS, 5% BSA, 0,01% Triton X-100) containing the primary ADNP antibodies overnight at 4°C. Cells were then washed 3 × 10 min with IF buffer and incubated with corresponding Alexa Fluor 555-conjugated secondary antibodies in IF buffer at a dilution of 1.0 μg/ml for 1 hr at room temperature with DAPI. Cells were washed 3 × 10 min with IF buffer and once with PBS.\n\nImages were acquired on an ImageXpress micro confocal high-content microscopy system (Molecular Devices), using a 20× NA 0.95 water immersion objective and scientific CMOS cameras, equipped with 395, 475, 555 and 635 nm solid state LED lights (lumencor Aura III light engine) and bandpass filters to excite DAPI, Cellmask Green, Alexa-555 and Cellmask Red, respectively. Images had pixel sizes of 0.68 × 0.68 microns, and a z-interval of 4 microns. For analysis and visualization, shading correction (shade only) was carried out for all images. Then, maximum intensity projections were generated using 3 z-slices. Segmentation was carried out separately on maximum intensity projections of Cellmask channels using CellPose 1.0, and masks were used to generate outlines and for intensity quantification.21 Figures were assembled with Adobe Illustrator.", "appendix": "Data availability\n\nZenodo: Antibody Characterization Report for ADNP, https://doi.org/10.5281/zenodo.14366335.22\n\nZenodo: Dataset for the ADNP antibody screening study, https://doi.org/10.5281/zenodo.14423002.23\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgment\n\nWe would like to thank the NeuroSGC/YCharOS/EDDU collaborative group for their important contribution to the creation of an open scientific ecosystem of antibody manufacturers and KO cell line suppliers, for the development of community-agreed protocols, and for their shared ideas, resources, and collaboration. Members of the group can be found below. We would also like to thank the Advanced BioImaging Facility (ABIF) consortium for their image analysis pipeline development and conduction (RRID:SCR_017697). Members of each group can be found below.\n\nNeuroSGC/YCharOS/EDDU collaborative group: Thomas M. Durcan, Aled M. Edwards, Peter S. McPherson, Chetan Raina and Wolfgang Reintsch\n\nABIF consortium: Claire M. Brown and Joel Ryan\n\nThank you to the Structural Genomics Consortium, a registered charity (no. 1097737), for your support on this project. The Structural Genomics Consortium receives funding from Bayer AG, Boehringer Ingelheim, Bristol-Myers Squibb, Genentech, Genome Canada through Ontario Genomics Institute (grant no. OGI-196), the EU and EFPIA through the Innovative Medicines Initiative 2 Joint Undertaking (EUbOPEN grant no. 875510), Janssen, Merck KGaA (also known as EMD in Canada and the United States), Pfizer and Takeda.\n\n\nReferences\n\nMandel S, Gozes I: Activity-dependent neuroprotective protein constitutes a novel element in the SWI/SNF chromatin remodeling complex. J. Biol. Chem. 2007; 282(47): 34448–34456. PubMed Abstract \| Publisher Full Text\n\nZamostiano R, Pinhasov A, Gelber E, et al.: Cloning and characterization of the human activity-dependent neuroprotective protein. J. Biol. Chem. 2001; 276(1): 708–714. PubMed Abstract \| Publisher Full Text\n\nJo YS, Kim MS, Yoo NJ, et al.: ADNP encoding a transcription factor interacting with BAF complexes exhibits frameshift mutations in gastric and colorectal cancers. Scand. J. Gastroenterol. 2016; 51(10): 1269–1271. PubMed Abstract \| Publisher Full Text\n\nBassan M, Zamostiano R, Davidson A, et al.: Complete sequence of a novel protein containing a femtomolar-activity-dependent neuroprotective peptide. J. Neurochem. 1999; 72(3): 1283–1293. PubMed Abstract \| Publisher Full Text\n\nPinhasov A, Mandel S, Torchinsky A, et al.: Activity-dependent neuroprotective protein: a novel gene essential for brain formation. Brain Res. Dev. Brain Res. 2003; 144(1): 83–90. PubMed Abstract \| Publisher Full Text\n\nGozes I: ADNP Regulates Cognition: A Multitasking Protein. Front. Neurosci. 2018; 12: 873. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVulih-Shultzman I, Pinhasov A, Mandel S, et al.: Activity-dependent neuroprotective protein snippet NAP reduces tau hyperphosphorylation and enhances learning in a novel transgenic mouse model. J. Pharmacol. Exp. Ther. 2007; 323(2): 438–449. PubMed Abstract \| Publisher Full Text\n\nD’Incal CP, Annear DJ, Elinck E, et al.: Loss-of-function of activity-dependent neuroprotective protein (ADNP) by a splice-acceptor site mutation causes Helsmoortel-Van der Aa syndrome. Eur. J. Hum. Genet. 2024; 32(6): 630–638. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHelsmoortel C, Vulto-van Silfhout AT, Coe BP, et al.: A SWI/SNF-related autism syndrome caused by de novo mutations in ADNP. Nat. Genet. 2014; 46(4): 380–384. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVan Dijck A, Vandeweyer G, Kooy F: ADNP-Related Disorder.Adam MP, Feldman J, Mirzaa GM, et al., editors. GeneReviews((R)). Seattle (WA); 1993.\n\nAyoubi R, Ryan J, Biddle MS, et al.: Scaling of an antibody validation procedure enables quantification of antibody performance in major research applications. elife. 2023; 12: 12. Publisher Full Text\n\nCarter AJ, Kraemer O, Zwick M, et al.: Target 2035: probing the human proteome. Drug Discov. Today. 2019; 24(11): 2111–2115. PubMed Abstract \| Publisher Full Text\n\nLicciardello MP, Workman P: The era of high-quality chemical probes. RSC Med Chem. 2022; 13(12): 1446–1459. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAyoubi R, Ryan J, Bolivar SG, et al.: A consensus platform for antibody characterization (Version 1). Protocol Exchange. 2024.\n\nBiddle MS, Virk HS: YCharOS open antibody characterisation data: Lessons learned and progress made. F1000Res. 2023; 12(12): 1344. Publisher Full Text\n\nLaflamme C, McKeever PM, Kumar R, et al.: Implementation of an antibody characterization procedure and application to the major ALS/FTD disease gene C9ORF72. elife. 2019; 8: 8. Publisher Full Text\n\nFire A, Xu S, Montgomery MK, et al.: Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature. 1998; 391(6669): 806–811. PubMed Abstract \| Publisher Full Text\n\nDana H, Chalbatani GM, Mahmoodzadeh H, et al.: Molecular Mechanisms and Biological Functions of siRNA. Int. J. Biomed. Sci. 2017; 13(2): 48–57. Publisher Full Text\n\nBandrowski A, Pairish M, Eckmann P, et al.: The Antibody Registry: ten years of registering antibodies. Nucleic Acids Res. 2023; 51(D1): D358–D367. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBairoch A: The Cellosaurus, a Cell-Line Knowledge Resource. J. Biomol. Tech. 2018; 29(2): 25–38. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nStringer C, Wang T, Michaelos M, et al.: Cellpose: a generalist algorithm for cellular segmentation. Nat. Methods. 2021; 18(1): 100–106. PubMed Abstract \| Publisher Full Text\n\nRuiz Moleon V, Alende C, Fotouhi M, et al.: A guide to selecting high-performing antibodies for ADNP (UniProt ID: Q9H2P0) for use in western blot, immunoprecipitation, and immunofluorescence.2024. Publisher Full Text\n\nLaflamme C: Dataset for the ADNP antibody screening study. [Data set]. Zenodo. 2024. Publisher Full Text" }	[ { "id": "352666", "date": "10 Jan 2025", "name": "Richard O'Kennedy", "expertise": [ "Reviewer Expertise Antibody production", "characterisation and use with particular emphasis on diagnostics." ], "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. This paper presents data on the binding characteristics of a set of commercially available antibodies that bind to the protein ADNP (UniProt ID:Q9H2PO), as determined using a variety of techniques including western blotting, immunoprecipitation, and immunofluorescence employing standardized experimental devised protocols. Determinations were made by comparing read-outs in knockout cell lines and isogenic parental controls. 2. The paper is designed to present the resultant data to enable researchers to compare the characteristics of the antibodies and their performance, in order to help researchers choose the most ideal/appropriate antibody for their particular applications. 3. For the most part the data is clearly set out but it would have helped if the legends to the figures were more explanatory and included the full-names associated with abbreviations. 4. The authors cite the approaches used in the methodologies employed. In addition, a section of the paper critically analyses the approaches and sets out possible shortcomings that the approaches may inherently have. This is very useful and would be particularly valuable if more papers had such evaluations generally. 5. Ensuring that the binding characteristics/specificity of antibodies is as well defined as possible is very important and this paper certainly assists in this in relation to the target protein. Fundamental questions have been raised about data generated where the antibodies were poorly characterized, and had specificity issues that were later realized, which may have been avoided by more intense and careful binding characterization studies.\n\nIs the rationale for creating the dataset(s) clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of methods and materials provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [] }, { "id": "352667", "date": "15 Jan 2025", "name": "Christian Peter Moritz", "expertise": [ "Reviewer Expertise Antibodies", "western blot", "immunoprecipitation", "immunofluorescence", "ELISA", "Proteomics", "Neuropathies" ], "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 report evaluates a study focused on characterizing commercial ADNP antibodies across various applications, contributing to a broader initiative aimed at addressing antibody reproducibility issues. The project is highly relevant, addressing a critical issue in biomedical research by tackling the reproducibility crisis in antibody-based experiments. The use of standardized protocols in knockout and isogenic parental controls highlights a rigorous and transparent approach, offering valuable insights and an open-access resource for the scientific community.\nI see options for the following improvements:\nMajor Points:\nFigure 2: If I am not mistaken, the band pattern in all of the panels’ SM (start material) lanes should match the pattern observed in the control lane for antibody 702911 in Figure 1—specifically, a strong band around 135 kDa and a very weak band slightly below it. However, the pattern and the molecular weight of the main band differ, suggesting significant technical variations among the experiments. Can we rule out technical variability among the tested antibodies? Are the observed differences truly attributable to the antibodies themselves? Precipitation Studies: For the precipitation studies, the authors used antibody 702911 to detect the precipitated proteins (“a specific ADNP antibody identified previously (refer to Figure 1) was selected”). However, in my view, antibody ab300114 appears to be more specific in Figure 1. I would like to understand why the authors chose antibody 702911 over ab300114 Competing Interests: Regarding their general YCharOS project, the authors might critically consider whether there are potential flaws in the study design due to the fact that the industry partner provided both the antibodies and the corresponding KO cell line. This creates a situation where the commercial antibody is being tested using a tool supplied by the same company. Are there any potential issues with this approach that should be discussed? I am aware that this criticism is rather for the general project as specifically for this study as the authors prepared their own Knock-down instead of using a company's Knock-out line if I understand it correctly.\n\nMinor Points:\nTitle and Abstract: I suggest introducing abbreviations—even for protein names like ADNP—the first time they are mentioned. While many reviewers recommend using housekeeping proteins as loading controls for Western blots, I appreciate that the authors have opted for a total protein staining technique (Ponceau Red) to provide a more transparent representation of the loaded protein amounts (cf. 10.1002/pmic.201600189). This is just a positive note; no changes are necessary.\nMy best wishes to the authors for their valuable project.\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? Yes\n\nAre the datasets clearly presented in a useable and accessible format? Yes", "responses": [] } ]	1	https://f1000research.com/articles/13-1545
https://f1000research.com/articles/13-1542/v1	20 Dec 24	{ "type": "Research Article", "title": "Assessment of occupational noise‐related hearing impairment among dental health personnel", "authors": [ "Mohammed Afroz", "Fatema Akhter", "Road Bitar", "Esmat Sarhan", "Faisal Alidan", "Esraa Alahmed", "Fatema Akhter", "Road Bitar", "Esmat Sarhan", "Faisal Alidan", "Esraa Alahmed" ], "abstract": "Background Dental professionals are exposed to either high level of noise which often becomes annoying over a time, and it is largely related to turbines and suctions used during the procedure. We performed this study with an aim to know the need for efforts to reduce impact of noise on young.\n\nMethods The study group comprised of 102 dentists (53 males and 49 females) final year dental students, interns and board students of family dentistry, endodontics, prosthodontics was enrolled. The noise assessment device is switched on to know the base noise level in the clinics and to know hearing ability of the dentist through a pre – recorded verbal communication. When the dentist starts the procedure, the same verbal communication is re-introduced and based on his reaction the decibel level is increased to a point where he can understand it. The same is followed after the procedure.\n\nResults Using T – test the age and gender mean was assessed, while ANNOVA test was used to know the hearing impairment and its mean before, during and after procedure. We have found that there is an imminent increase of sound during the procedure and thereby associated impairment; which continues but to a lesser decibel level even after the procedure is completed for long time.\n\nDiscussion and Conclusion We have highlighted few guidelines from various studies which range from reducing multiple turbine usage at a given point of treatment wherever possible to a recommendation to use ear plugs for those procedures related to high noise levels.", "keywords": [ "hearing impairment in young dentists", "pre", "during and post procedure hearing loss." ], "content": "Background and introduction\n\nDental professionals are exposed to either a high level of noise or a continuous noise source that often becomes annoying over a period of time. These types of noise are often due to their work-related sources like aspirators, ultrasound scalers, high and low speed handpieces, turbines, etc to name a few.1 These compressor-based, predominantly rotary instruments are kept too close by the dentist for easy accessibility, ease of work, and increased cognition. Because of the work protocol of chair-side dental assistants, an extra noise source is often created in the form of low- and high-volume suction, spittoon water, glass/tumbler filling water set, etc., which are not in direct control of the dentist. There are few studies that also highlight the plight of the extended care dental assistant related to the high noise levels in the dental office and often related to them working longer hours in the clinics compared to dentists.2,3 There are various other risks highlighted in the literature that cause detrimental effects related to the dental profession, including infectious and communicable diseases, musculoskeletal pathologies, and ionizing radiation.4,5 These effects have shown an early physical and mental effect; hence, more studies have been conducted to highlight the cause of remedial measures. Exposure to high noise was initially disregarded as the initial dental treatment was mostly biomechanical in which the dentists themselves used semi-automated foot controlled instruments, but since the advent of compressor-based instruments, although the treatment time is reduced and so may be physical fatigue, it has led to a more blaring reality of effects expressed both at the atrial level and at the extra-auricular level (sleep disorders, cardiovascular disorders, stress, etc.). Apart from these, there are other reasons for hearing impairment like social noise exposure largely related to recreational activities, and the increasing age of an individual. The effects of noise can be both auditory which causes fatigue and temporary impairment while non auditory effects can interference with speech, annoyance and reduction inefficiency with psychologic damage.6,7 It was noticed that left ear of a right handed person showed a greater loss of hearing ostensibly related to proximity to the noise source.7 Acoustic environment of educational and teaching activities for the faculty of dentistry makes them more vulnerable to high noise levels compared with other areas of education, as it require the use of various equipment that emit different decibels of noise by many students at the same time.4,8 The educational institutes have laboratory areas where noise levels were recorded to be as high as 93db, while in preclinical areas the sound levels are 78.8db. In the clinical areas the ultrasonic scalers had maximum noise levels recorded as 83.3db, and since these equipments run for a longer duration of time since there may be many clinics performing similar tasks hence the faculties and auxillaries working in tbis environment are more vulnerable, even though the noise levels are within the acceptable range. There is also effect seen between new and old instrument which is roughly a 2 decibel less noise in new equipment compared to the old ones.8 Noise levels has been roughly divided into distracting and destructive noise.3,9 Recently, Occupational Safety and Health administration in the year 201110 has incorporated enforcement guidance for personal protective equipment which is in continuation to the original occupational noise exposure code of federal regulation number 29 1910.95 which was originally given in 1981, in order to highlight the working hours with the maximum permissible dose and has considered it as a highly important fields of study in the upcoming century. In addition there are studies which has reported that a prolong high exposure to noise can lead to other risk areas such as vibration and hand neuropathy, it might help to determine whether these problems are related to practicing dentistry, as the recommendations of ADA council suggest that dental practitioners concerned about the potential hearing impairment should go for otologic examination a d audiometric evaluation. It is also recommended to use ear protective devices in order to avoid hearing impairment in a long run.11 To reduce workers’ risk of developing noise-induced hearing loss (NIHL), the exposure levels of contemporary dental equipment are generally within the limits set by the National Institute for Occupational Safety and Health (NIOSH), which recommends limiting noise exposure to 85 dBA at 40 h per week.12,13 The maximum permitted exposure limit (PEL) in an 8-hour day should not surpass 90 dBA SPL (decibel; sound pressure level using an A-weighted scale), according to the 2001 decibel levels standard. This is roughly in accordance to OSHA’s standard, which employs a 5 dB exchange rate. The maximal permissible exposure limit (PEL) of 95 dB noise should cut down the working time to 4 hours. Similarly, the PEL for a 100 dB sound is permitted for a working time of 2 h, and so on. Besides its effects on hearing, loud noise may also impair other organic systems, such as in behavioral, neurological, psychological, vestibular, digestive, and cardiovascular disorders, among others. Excessive noise increases stress levels, which increases blood pressure and hormone release, such as adrenaline and cortisol. The effects of noise on hearing (Noise-Induced Hearing Loss [NIHL]) and general health may affect dentists, hinder communication, and negatively impact their quality of life.13 Dentistry demands knowledge, skills, and focus as part of a profession, and hence, a good quality of life is in turn affected by destructive noise or long-time exposure to distracting noise. Owing to modern equipment, manufacturing companies have done a great deal of work in keeping the sound decibels low, mostly hovering around 85DBL,14 but apart from the noise levels, it is the time duration for which the dentist has been working that can itself affect his overall hearing ability. In this regard, there are several studies in dental literature examining hearing loss among dental professionals that warrant discussion.6,7,13 Certain studies have gone farther by using noise dosimeters to record the noise and has been able to show a positive correlation between the length of service and the degree of hearing impairment among dental practitioners.13 There are studies which has tried to correlate the years of practicing dentistry to associated hearing impairment, as was discussed in a study that dental professionals with more than 10 years of experience working 8 hours a day are at the highest risk of developing hearing impairment, and it was also commented that the worse the hearing status of dental professionals, the more their health status is adversely affected.13,15,16 Among different dental professionals, a comparative difference among the hearing thresholds of three dental professional groups (dentists, dental nurses, and prosthodontists) using a high-frequency audiometric testing method reported that general dentists were the most affected group followed by the prosthodontistsat the mean frequencies of 500-2000 Hz and 3000-6000 Hz, while dental nurses were the most affected group at the mean high frequencies of 9000 and 16000 Hz.17–19 As highlighted before, dental university clinics have different levels of noise exposure, probably due to a greater number of clinics and different clinics performing various clinical activities simultaneously, which has not been discussed much in the literature. We performed this study with an aim to determine the need for efforts to reduce the impact of noise on young dentists; hence, our objectives were to determine the hearing impairment in dentists before entering the clinical procedure to post-clinical procedures and to explore the methods used by the dentist to reduce noise exposure depending on their awareness.\n\n\nMethods\n\nThe study was carried out at the Dar Al Uloom University Dental Clinic, which has 45 fully equipped dental cubicles separated by a partition of heat proof walls of 7 feet height. Since it involved the participation of the concerned dentists, ethical clearance was taken bearing numbers 04 – 08 – 2021, from scientific research and ethics committee of college of dentistry belonging to dar al uloom university This committee follows the Helsinki ethical principles of medical research involving human subjects while giving ethical approval. The ethical approval was received on 10 – 09 – 2021 and has a validity of 5 years. During clinical practice, based on a schedule of approximately 10 cubicles, separate dental procedures, such as prosthodontics, oral surgery, and periodontics, are performed. However, the pedodontics clinical schedules are maintained in the late afternoons when the other specialties are not high and/or are operated in a separate clinical corridor within the same clinics. The study group comprised final-year dental students, interns, and board students of family dentistry, endodontics, and prosthodontics. A total of 102 dentists were enrolled, with 53 males and 49 females between the ages of 22 and 33 years. After explaining the procedure and obtaining a written consent from the concerned dentist and their supervisors where necessary, the noise assessment device was switched on to determine the baseline noise level in the clinics before the start of the procedure. The hearing ability of the dentist was recorded by playing a pre-recorded verbal communication audio of 3 min duration whose volume was adjusted to a comfortable level of the concerned dentist. This recorded value is considered the basic hearing ability of the dentist. When the dentist starts the procedure, the same verbal communication is re-introduced to understand the hearing ability of the dentist. Based on this reaction, this noise level is increased to a point when he can understand the noise level, and this change in noise decibel level is recorded. After the procedure, the dentist was exposed to the same verbal communication and gradually increased until he could clearly hear the same voice note. At all times, the dentist is semi-concentrated on the voice note and conveys the hearing response as he can hear the voice clearly. The dentist will be asked the following questions before the start of the procedure, which is their awareness of hearing impairment among the dentists, was it ever experienced by the dentist that there was hearing impairment, was their specific modification carried out to reduce the ill effects of it. The device used in this study is manufactured by Wintact professional instrument, model WT 1357, version WT 1357 – EN - 00 that has the ability to record the noise level in digital units of decibels. The unit can be switched off with a single click and has memory of five recent readings. Each reading was recorded each time; however, a memory function was used to correlate the readings later or in cases where it was not possible to record immediately. There are 3 researchers were available in the clinic to record the values at different stages to avoid conflicting results.\n\n\nResults\n\nThe research comprised both male and female young dentists within the age range of 22 to 33 years, with 52 males and 49 females as described in Figure 1, and the mean age with standard deviation and T Value is shown in Table 1.\n\nThe data were assessed using a T-test because it is a comparative hearing assessment study within the same individual, as described in Table 1. It shows the independent T-value to be negative before the procedure and during the procedure period, while it turns positive after the procedure readings, suggesting that there is an eminent change in the hearing ability of the dentists. The obtained data show a marked increase in the overall clinical cubicle sound level when the procedure is performed, as seen in the mean value, which can be attributed to the commonly used rotary instruments and suction. Each dentist worked for a minimum of two hours, operating on more than one patient depending on the appointment, and then there was a break for 30 minutes.\n\nThe study also described the general change in hearing ability between males and females, which was obtained using the ANOVA test, as both genders are involved. Before the procedure started, there was no difference in the hearing ability of the subjects, which changed during the procedure (Table 2). It also revealed that hearing was impaired more in females than in males during the procedure.\n\nSince the research involved individuals between 22 and 33 years of age, another correlation between the ages was performed as described in Table 3, in which they were divided into those less than 25 years and more than 25 years. We found that subjects under 25 years of age had better hearing ability, even when the procedure was in progress, compared to those above 25 years of age.\n\n* p≤0.05 considered statistically significant.\n\nIrrespective of sex and age, there is a constant change seen in hearing ability, which decreases during the procedure and immediately after the procedure up to a minimum of 30 minutes duration, suggesting that during the procedure, since there are other clinical sounds added like rotary instruments and suction, but even when the procedure is completed and the dentist has sent the patient, his hearing ability is reduced, and this can be related to the stress post clinical procedure. A decrease in hearing ability means that the dentist must speak little louder to make the same conversation comfortably audible to them during and after the procedure.\n\n\nDiscussion\n\nOur study included final-year students, dental interns, and board students of family, endodontics, and prosthodontic dental specialties. Care was taken to avoid pedodontics clinics for dental interns, and there were no pediatric clinics running at the time of the study. We have a greater number of subjects below the age of 25 compared to above 25 years, suggesting that the enthusiasm to join dentistry and continue it further as board dental specialties is continuous, thereby highlighting the awareness among the young population. It is further seen that there are a greater number of male students than females; however, this difference is very low and hence can be considered insignificant and largely related to the time and availability of respective genders during the study period. However, it highlights the willingness of either gender to participate in the study, thus disproving the theory as an orthodox population. This finding goes close to various studies conducted in this region3,20,21 and few studies had more of female population compared to the males.22,23 The consent form was filled by all the participants, and we found that the questions in the consent form added no value to the study as all the participants said that they have only heard about the hearing impairment among the dentists, and they have not experienced it on themselves. Hence, we have not included them under the results. It is a known fact that dental rotary instruments2,4 cause sudden increase in the sound level within the clinics and there are reports of it causing tinnitus 20,21,24 initially and on long exposure leading to permanent hearing impairment.5,6 Even after the procedure has been completed, there is still loss of hearing ability, which was prominently seen in our study and continued for a considerable period, thereby suggesting that it has an impact as a stress factor among them, which has been reported in other studies.21,25 We want to highlight a review study conducted by Hartland25 and published in the year 2023 discussing various articles highlighting the amount of noise induced by different equipment during the procedure, the awareness of the dentists to the same, to the articles highlighting a permanent hearing impairment, and there are few studies that suggest different guidelines that should be implemented to reduce the amount of hearing impairment,26 ranging from reducing the usage of different turbines whenever possible to wear ear plug muffs as a compulsion. It is not like the early dentists did not consider noise as a detrimental factor in their profession, as was discussed by Kaymak et al. in 2007,27 who classified the noise reduction methods into active noise reduction, adaptive filtering, and passive noise reduction. The study highlighted the use of earphones to be worn by the dentist and the patient, and these headphones must have adaptive filtering, thereby reducing the handpiece sound and allowing the dentist voice to be heard more clearly by the patient. This was further explored by the same researchers,28 who concluded that modifications in the headphones can reduce the noise to a great level, but their work was concentrated more towards handpieces, and there are various restrictions in real-time dentistry to apply larger headphones to all individuals in the operating room, as some might find its use itself annoying. A study29 was conducted using smaller headphones with noise cancellation technology involving airpods, QC30, and Quieton Dental, which are commercially available headphone devices. The authors concluded thatQuieton Dental is a device that can effectively block the noise induced from dental equipments since it blocks noise above 3 kHz without disrupting normal conversations by maintaining or amplifying sound at 3 kHz. However the findings are not in decibels. The authors concluded that Active noise control (ANC) equipment that can effectively match with dental clinical setup noise characterists more precisely should be developed. in order to improve the quality of dental treatment and provide comfort to people in the clinic. A similar conclusion was seen in another study,30 which was published in 2023, who tested antinoise ear plugs (Quiton), which found it to have limited use in the dental treatment setting.\n\n\nConclusion\n\nOur study highlights that there is a definitive change in hearing impairment among young dentists from before starting the procedure to during and after the procedure; hence, there is an urgent need to address this health hazard among them and related dental auxiliaries. Although there are few noise cancellation devices in the market, dental professionals need something that is specific to their requirements, because commercially available products are built and developed for other purposes. Furthermore, most noise reduction studies are performed in a simulated dental setup using an ear and cheek simulator to be placed on the phantom heads and hand piece operated on typodonts. Our study was not able to highlight which procedures have a greater noise impact to suggest specific recommendations. We conclude with a proverb: the deaf believes there is nothing wrong, the hearing believes something needs to be fixed.\n\n\nEthical approval and consent\n\nScientific Research Ethics and Innovation Committee Number 04 – 08 – 2021.\n\nEthics approval number: 04 – 08 – 2021, obtained from scientific research and ethics committee of college of dentistry belonging to dar al uloom university This committee follows the Helsinki ethical principles of medical research involving human subjects while giving ethical approval. The ethical approval was received on 10 – 09 – 2021\n\na. written consent from the concerned dentist and their supervisors where necessary, were obtained", "appendix": "Data availability statement\n\n{Harvard Dataverse} Assessment of occupational noise-related hearing impairment among dental health Personnel, version = 6.0. https://doi.org/10.7910/DVN/XGGUIV.31\n\nThe project contains the following underlying data:\n\n• Hearing loss data_final.xlsx;\n\n• New tables with graphs\n\n• Hearing device detailsHearing loss data hand written 100.pdf\n\nLicense/Data use agreement: CC0 1.0\n\n{Harvard Dataverse} Assessment of occupational noise-related hearing impairment among dental health Personnel, version = 6.0. https://doi.org/10.7910/DVN/XGGUIV.31\n\nThe project contains the following data:\n\n• Hearing loss data\n\n• Hearing loss data hand written 100.pdf\n\n• new tables with graphs.docx\n\n• Hearing device details\n\nLicense/Data use agreement: CC0 1.0\n\n\nAcknowledgement\n\nThe authors are thankful to the Deanship of Graduate Studies and Scientific Research at Dar Al Uloom University, Riyadh, KSA for providing support for this project.\n\n\nReferences\n\nDierickx M, Verschraegen S, Wierinck E, et al.: Noise Disturbance and Potential Hearing Loss Due to Exposure of Dental Equipment in Flemish Dentists. 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Publisher Full Text\n\nAl-Rawi NH, Al Nuaimi AS, Sadiqi A, et al.: Occupational noise induced hearing loss among dental professionals. Quintessence Int. 2019; 50(3): 245–250. PubMed Abstract \| Publisher Full Text\n\nLopes AC, de Melo AD , Santos CC: A study of the high-frequency hearing thresholds of dentistry professionals. Int. Arch. Otorhinolaryngol. 2012; 16(2): 226–231. PubMed Abstract \| Publisher Full Text\n\nGoncalves CG, Santos L, Lobato D, et al.: Characterization of hearing thresholds from 500 to 16,000 hz in dentists: a comparative study. Int. Arch. Otorhinolaryngol. 2015; 19(2): 156–160. PubMed Abstract \| Publisher Full Text\n\nAlhaider SH, Togoo RA, Hakami RM, et al.: Impact of Noise on the Hearing and Tinnitus among Dental Students, Interns, and Dental Practitioners: A Cross-sectional Study. World J. Dent. 2023; 14(7): 581–585. Publisher Full Text\n\nNassar M, Isalm MS, D’souza S, et al.: Tinnitus Prevalence and Associated Factors among Dental Clinicians in the United Arab Emirates. Int. J. Environ. Res. Public Health. 2023; 20: 1403. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAkhter F, Afroz MM, Alamri LA, et al.: The relationship between the prevalence of dental space infections in Pediatric patient and parents’ Awareness Bangladesh. J. Med. Sci. 2023; 22(04): 773–777. Publisher Full Text\n\nAkhter F, Afroz MM, Haque T, et al.: Loss of Taste and Smell Sensation in COVID 19: A Questionnaire Based Multicontinental Study with Review. JPRI. 2022; 34(50A): 1–11. Publisher Full Text\n\nMyers J, John AB, Kimball S, et al.: Prevalence of tinnitus and noise-induced hearing loss in dentists. Noise Health. 2016; 18(85): 347–354. PubMed Abstract \| Publisher Full Text\n\nHartland JC, Tejada G, Riedel EJ, et al.: Systematic review of hearing loss in dental professionals. Occup. Med. October 2023; 73(7): 391–397. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDr Alharbi I : Noise Induced Hearing Loss Among Dentist. J. Interdiscip. Med. Dent. Sci. 2021; 4(1).\n\nKaymak E, Atherton M, Rotter K, et al.: Dental Drill Noise Reduction Using a Combination of Active Noise Control. Passive Noise Control and Adaptive Filtering. 2007. Publisher Full Text\n\nRotter K, Atherton M, Kaymak E, et al.: Noise reduction of dental drill noise.2008. Publisher Full Text\n\nKim IH, Cho H, Song JS, et al.: Assessment of Real-Time Active Noise Control Devices in Dental Treatment Conditions. Int. J. Environ. Res. Public Health. 2022 Aug 1; 19(15): 9417. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJiang P, Atherton M, Millar BJ: Dental drill noise reduction using a commercially-available earplug device. Prim. Dent. J. 2023; 12(1): 73–78. PubMed Abstract \| Publisher Full Text\n\nMalik M: Assessment of occupational noise‐related hearing impairment among dental health Personnel. Harvard Dataverse. 2024; V6. Publisher Full Text" }	[ { "id": "352680", "date": "08 Jan 2025", "name": "Dumini De Silva", "expertise": [ "Reviewer Expertise Audiology", "speech language pathology", "disability studies" ], "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 the paper examining occupational noise‐related hearing impairment among dental health personnel. This paper addresses the changes in hearing ability and noise levels in a university dental clinic in Saudi Arabia. It explores the changes in the hearing ability of dentists of age 22- 33 years before, during and after exposure to noise emitted by dental procedures. Though there was no apparent difference in the noise levels in the dental settings before and after procedures compared to when the dental treatment was ongoing, a statistically significant change in hearing sensitivity was reported in the sample. Several issues with the methods, data analysis, and interpretation of results need to be addressed. With the changes, the findings of this paper would be an important addition to the literature on the noise levels in university dental setups, effects of short-term noise exposure on hearing and awareness of hearing health among dentists.\nAbstract:\nThe paper aims to know the need to reduce the impact of noise on dental treatments. Since the target population is young adults, it is important to mention the age details (e.g. mean, SD/ range). The method stated in the abstract is vague. I recommend the authors use a sound level meter instead of the term \"noise assessment device\", as there are different types of such systems, including mobile phone apps, available in the market. The first few sentences in the results describe the statistical tests used, which should ideally be included in the method. The results with numerical information would improve the clarity of average noise levels (before, during and after dental procedures), and associated changes in hearing ability mentioned in the abstract. The conclusion contains recommendations from previous studies focusing on turbine and multiple device use. However, it is important to link the recommendation with the study findings.\nKeywords: I highly recommend that the authors check the compatibility of selected keywords with MeSH terms.\nIntroduction:\nThe introduction includes details of noise sources in dental settings, auditory and non-auditory effects of noise exposure, noise levels and regulations, reduction in noise emitted by modern equipment, and noise measurement methods. Authors acknowledge the lack of data from teaching facilities and among young dentists. More information about the noise and hearing measurement methods and the association of temporary threshold shifts with age would justify the study's aims. According to the details given in the method, the study's first aim was to compare hearing ability at three different points (before, during, and after clinical procedures). I suggest the authors rewrite the aim using appropriate verbs and terms. Reference 10 should be added to the list of references.\n\nMethod:\nThe method section of the paper includes details of the study setting, participants, noise measurement, levels of hearing ability, and perceived methods of mitigating noise exposure. However, the information is scattered, making it difficult for the reader to understand how the study was conducted. Therefore, having subsections or separate paragraphs on each aspect of the method would increase the readability. Details on ethical approval are mentioned in a separate section after the conclusion. Therefore, I recommend ethics details be removed from the method. The sentences written in the future tense need to be revised. The description of the study setting provides a clear picture of the clinic environment. A few more details such as the proximity of each cubical and treatment chair to the cubical wall may provide the reader with an approximate idea of possible ongoing background noise and reverberation during dental procedures. The name and model of the sound level meter (SLM) are mentioned. Adding further details, such as the type of SLM according to the IEC standard, the distance of the measuring microphone to the sound source, mode of measurement, recording length, noise weighing, etc, would improve the clarity of measurement results. Details of the number of participants and their age and sex data would ideally given in the results section. As the authors did that, I suggest removing them from methods and stating the inclusion-exclusion criteria for the participants. Pre-recorded verbal communication audio: What was it composed of (e.g. conversation of 3 people/ monologue)? Where (at the treatment cubical/ office room) and how was it played (headphones/ speaker kept at a specific distance)?\n\nHearing ability: In the beginning, the text gave the impression that the participants were asked in indicate to increase the sound of the pre-recorded verbal communication to a level that they could follow/ understand it, however later, the authors mentioned that the level that the participants were required to semi-concentrate. The authors must clearly define how the hearing ability of participants was measured. The statistical analysis used for the study must be stated using correct terms (e.g., repeated measures t-test).\nResults:\nThe reason for categorising participants at 25 years of age needs to be clarified. Table 1: The explanation of Table 1 and the content do not match. Table 1's title is unclear (suggestion: Mean/ average sound level in the dental clinic before, during, and after procedure). Also, it is unclear why the authors expected the sound level in the environment to be different according to the sex of the dental practitioner. I suggest comparing before, during, and after-procedure noise measurements of the entire sample using a suitable statistical analysis (considering the normality distribution) and adding the unit (e.g. dB SPL). Table 2: The term \"impaired\" does not complement this paper's method for deriving hearing ability. Therefore, an explanation of the finding (e.g., speech needs to be increased by 1.2 dB on average) would be beneficial. Also, the term \"general hearing ability\" must be defined in the methods section. Table 3: The table compares hearing ability before, during and after procedures in 2 groups according to age. Therefore, the statistics and p-values are of different measurement points, not between age groups, requiring the table explanation to be revised. Post-hoc testing would compare hearing ability before and after the procedure. Also, a suitable statistical analysis must be performed to observe an association between age and point of noise exposure (e.g., two-way repeated measures ANOVA). The last paragraph of the results section could be moved to the discussion, as it addresses the reasons for the shift in hearing after the procedure level. Results of the survey questionnaire on awareness of hearing impairment among the dentists, own experience of hearing impairment, and any specific modification to reduce the ill effects of noise exposure in the dental setting should be included. Lack of awareness of possible hearing loss due to noise exposure and lack of hearing conservation practices are important findings in this sample.\n\nDiscussion:\nSentences 3- 6: Details provided on undergraduates' age, sex and enthusiasm for dentistry seem not relevant to the scope of the study. The authors claim that findings from the questionnaire presented with the consent form on awareness of hearing impairment and methods to reduce noise exposure do not add value to the study. However, the authors mention the exploration of methods used by dentists to reduce noise exposure depending on their awareness, which is the second aim of this study, it is imperative to discuss the findings with relevant literature.\n\nPost-hoc analysis results from Table 3 are required to review the discussion on the effects of noise exposure. In revising the discussion section, the authors are encouraged to review the literature on Temporary Threshold Shift (TTS) in dentists and sound levels in dental clinics.\n\nConclusion: The authors conclude the paper by highlighting the association between age and time of noise exposure. However, the data analysis needs to be re-performed to derive accurate conclusions. The claim on noise reduction studies and the proverb may not be relevant to the present 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? 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? Partly", "responses": [] }, { "id": "355446", "date": "16 Jan 2025", "name": "Abdulazeez Ahmed", "expertise": [ "Reviewer Expertise Otolaryngology & Audiological 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\nDear Editor, Thank you for inviting me to review this piece of work. The work in its entirety perhaps needs revision, the methodology requires a scientific approach (e.g. assessing hearing according to global best practices requires at a minimum, Puretone Audiometry); but the authors used subjective assessments means which may make generalisability quite difficult. Although they were able to measure environmental noise levels, using a sound level meter. Again, perhaps by omission the authors have provided \"a date\" instead of a proper ethics approval number. Some statements in the discussion are also redundant, some appear to not have the flow of a scientific paper discussion. I would strongly suggest a revision of the whole manuscript please.\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? No\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/13-1542
https://f1000research.com/articles/13-1541/v1	20 Dec 24	{ "type": "Research Article", "title": "Virtual game-based learning environments to promote self-regulated learning skills in foreign language learners", "authors": [ "Maira Alejandra Noriega Cortes", "Laura Carreño-Bolivar", "Maira Alejandra Noriega Cortes" ], "abstract": "Background This mixed-method action research study explores the impact of a gamified virtual learning tool on elementary students’ self-regulated learning (SRL) skills in English language acquisition. Despite the acknowledged importance of SRL in educational contexts, limited research exists on its development among young learners. The present study is one of the products of the academic unit project “Key Factors and Essential Elements for a Comprehensive Model of Bilingual Education: A Multidimensional Approach for Diverse Contexts and Areas of Knowledge” sponsored by the School of Education at Universidad de La Sabana, institution where the first author conducted her master’s studies, and the corresponding author is a professor and main researcher of the of the academic unit project.\n\nMethod Six students aged 8-10 participated in the study. Data were collected through semi-structured observations and pre- and post-questionnaires. The tool was designed based on Zimmerman’s SRL model and included tasks structured through gamification elements. For the purpose of quantitative data analysis, descriptive statistics were used; as for qualitative data, thematic analysis was the chosen approach.\n\nResults The results revealed that guided use of the gamified tool significantly enhanced students’ use of SRL strategies, particularly in goal setting, monitoring progress, and reflecting on learning outcomes. Students demonstrated increased motivation, engagement, and confidence, although explicit instruction was required to maximize the tool’s benefits. The usability of the tool was rated positively, with participants highlighting its effectiveness for language learning and SRL skill development.\n\nConclusions The study concludes that gamified virtual environments can effectively promote SRL skills in young language learners by fostering motivation and engagement. However, for optimal outcomes, such tools should be supplemented with explicit instruction in SRL strategies. This research contributes to understanding the integration of gamified tools in elementary education, highlighting their potential to develop foundational skills essential for lifelong learning.", "keywords": [ "Gamification", "self-regulated learning- self regulation", "Language Learning", "virtual learning environment- virtual learning- blended learning environment." ], "content": "Introduction\n\nWith evolving educational landscapes and changing motivations of new generations, autonomy and awareness of one’s learning process and skills have become essential for successful learning. Being a highly self-regulated learner is an important human resource for students to learn how to learn to manage, regulate, and enjoy their learning (Fukuda, 2019; Lu et al., 2017; Wang, 2021; Zhou & Li, 2020; Zimmerman, 2000), which is the reason behind a current research phenomenon that attempts to deeply explore self-regulated learning applied to different educative contexts and learning scenarios. Little research has been done on the matter, specifically in Colombia (Giraldo, 2022; Gómez Sará, 2017; Moreno et al., 2021), leaving a gap in knowledge in terms of how young elementary students develop self-regulated learning skills, how those abilities can be taught inside the language classrooms, and how gamification on digital platforms can be an effective resource in supporting the development of self-regulated learning. Thus, the present study aimed to focus on the interaction between self-regulated learning, language learning, and gamified technologies, and how these variables affect the development of self-regulated learning skills in elementary students. The data collected and the design of a virtual learning environment that can successfully promote self-regulated learning from an early age, became a significant contribution to the educational field as well as to the knowledge field in Colombian research of elementary language learners.\n\n\nTheoretical background\n\nThe importance of self-regulated learning and its impact on motivation and academic achievement has been a critical matter for educational research since Albert Bandura and Barry Zimmerman in the late eighties (Lu et al., 2017). Being one of the main referents of the concept, Zimmerman (2000) conceptualized self-regulation as “self-generated thoughts, feelings, and actions that are planned and cyclically adapted to the attainment of personal goals” (p. 14). After many years of reflection, it was established that self-regulated learning was the result of the combination of a metacognitive component, motivational beliefs, and cognitive and social affective elements (Lu et al., 2017). Over time, the concept of self-regulated learning has allowed researchers to understand there is a cause-effect relationship between self-regulated learning and academic achievement, as the higher level of self-regulated learning skills learners have, the higher academic results they might obtain (Fukuda, 2019; Neitzel & Connor, 2018). Hence, self-regulated learning has also been related specifically to language acquisition and how it influences positively certain language skills (Fukuda, 2019; Yüce, 2019). Research has identified that the more students are immersed in traditional learning environments and traditional learning methods, the lower levels of self-regulated learning are going to be developed (Li et al., 2022). Because of that, Cerda et al. (2020) described the relevance of a flexible bilingual learning environment that cultivates student choice and supports content meaning-making, self-regulation, and language learning. As a result, self-regulated learning contributes to language learners’ engagement and enhanced performance in the foreign or second language classroom (Wang, 2021), and under that comprehension, the present research examined the relationship between self-regulated learning and successful language learning strategies as part of elementary school settings.\n\nThere is broad research on the matter of self-regulated learning development in higher education levels and adult education contexts (Uus et al., 2021). Nonetheless, studies made with children in this area support the idea that they are curious and exploratory learners, who from an early age can selectively explore and choose the information they want to receive (Foushee et al., 2021). For example, Dent and Koenka (2016) established a strong correlation between self-regulated learning and academic achievement from kindergarten to second grade, which then became weaker from third through fifth grade. Research in elementary school settings has described even how students’ age or grade level affected the self-regulated learning orientation strategies students tend to use as they progress to higher grades (Zhu & Mok, 2018). In Colombia, Elementary schools must ensure students acquire basic knowledge for secondary education while developing cognitive, procedural, and affective skills (Moreno et al., 2021). Many students, particularly in public schools, have low competencies in essential skills like reading and writing in Spanish, and yet, the Ministry of National Education aims for students to achieve an A1 level in English by the end of elementary school to build on in high school (Giraldo, 2022; Gómez Sará, 2017). For academic success to happen, children should be able to regulate their emotions and behaviours to adapt to their social and learning environments. This will result in better school readiness and more effective learning processes (Acar et al., 2022). However, when the goal is to pursue a successful learning process that allows students an integral development as learners, other areas of knowledge and skills should be emphasized (Gómez Sará, 2017). That is why promoting self-regulated learning development from early stages in education becomes of such relevance for these new generations. Even so, children will not necessarily become sophisticated, self-regulated learners just by maturing along the developmental stages, so learners require formal training and practice from a young age in self-regulated learning strategies to be successfully autonomous learners (Rameli et al., 2022; Zhu & Mok, 2018). Accordingly, the present research was based on the relevance of knowing how to foster and teach self-regulated learning abilities in language learning as part of the early stages of the educational process, to support the development of higher-order thinking skills and emotional capabilities, according to the developmental stage of elementary students.\n\nGamification refers to the application of game mechanics, aesthetics, and game thinking in non-gaming settings to engage people, motivate action, promote desired behaviors, learning, and solve problems. It may traditionally include the use of interconnected game components (point and leveling systems, leaderboards, badges, bonuses) and game elements (achievements, competition, rewards, and self-expression) while turning the entire learning process into a gaming event (Kucher, 2021; Ofosu-Ampong, 2020; Opris et al., 2021). Gamification as a teaching-learning strategy in educational contexts can be seen as an effective way to educate students, so it can increase the level of individual and group motivation, have an emotional and social impact on students, and increase academic achievement and learners’ ability to learn new skills. Gamified applications for second language learning have proven effective in boosting children’s motivation, engagement, and confidence through enjoyable and non-judgmental environments (Li et al., 2022; Shadiev et al., 2017; Yükseltürk et al., 2018). Dindar et al. (2021) found that gamified cooperation and competition positively impact task effort, learning achievement, and motivation in Chinese university students. Li et al. (2022) observed that younger students (8-10) from Hong Kong benefited from a gamified English learning system, with active users showing improved academic performance and self-regulation skills. Learning sequences should be tailored by following gamification principles, this implies adapting activities and procedures to a gaming thinking system to how a lesson is taught and continuing to develop it based on the feedback from the players, therefore utilizing the elements from game design to improve educational outcomes, activities, and students’ learning experience (Ofosu-Ampong, 2020). In consequence, for the present research gamification was better understood as described in 2011 by Zichermann & Cunningham, like the process of using game-thinking and game mechanics to engage users and solve problems (Ofosu-Ampong, 2020). This allowed a comprehensive view of a digital tool designed to foster the development of self-regulated learning abilities through an engaging, cohesive, and structured learning experience, while it contributed to further research in terms of the applicability and benefits for elementary students.\n\nDigital technologies have been studied as tools that enable personalized and collaborative learning, and that can be used by teachers with different purposes (Bower, 2017). Research also highlights the importance of designing authentic environments for effective learning, which means they should provide activities with real-world relevance, give opportunities for sharing learning experiences regardless of their level of expertise, and allow reflection and assessment within the tasks (Shadiev et al., 2017). Moreno et al. (2021) found that multimedia elements in digital tools (image, sound, animation, interactivity) enhance self-learning and linguistic skills (reading, writing, speaking, listening) in Colombian elementary students, offering innovative pedagogical methods. Hung et al. (2018) reviewed empirical evidence on digital game-based language learning, noting that multiplayer online role-playing games are commonly used on personal computers, primarily for English learning among university students. This raises questions about how such platforms can also support the development of self-regulated learning in diverse learner groups. Virtual Learning Environments (VLEs), for example, have been associated with formal learning and with relationships between teachers, students, and schools, as their features and potentialities become an important element within the student’s learning processes (Alves et al., 2017). VLEs facilitate interaction and interrelation within a continuous communication process, enhance the construction of knowledge and meanings, and promote the formation of habits and attitudes (Alves et al., 2017). Another notion adopted to describe some technological tools applied to education is the Digital game-based learning (DGBL) environment. This according to the researchers has the potential to offer students situated learning that fosters higher levels of engagement and motivation, provides a creative atmosphere to focus on the task, improves real-world skills, and increases students’ self-esteem and interest in the subjects (Hafeez, 2021; Kucher, 2021; Taub et al., 2020). Regularly, students encounter challenges when adopting efficient self-regulatory processes such as cognitive, affective, metacognitive, and motivational while learning, DGBLs encourage pupils in their learning processes due to the properties and possibilities of games. These attributes foster effective self-regulated learning and provide students with opportunities to take the initiative in their learning by analyzing, synthesizing, evaluating, and performing higher-order thinking skills (Liao et al., 2019; Taub et al., 2020). In that sense, it is possible to say the technological tool presented as part of this research is a game-based virtual learning environment, which supports itself with gamification strategies to promote the development of self-regulated learning abilities while favoring the development of communicative skills in a foreign language.\n\n\nMethod\n\nThe present study followed a mixed-method approach that combined qualitative and quantitative data collection methods such as semi-structured observations and two questionnaires, both instruments were fully designed and reviewed by the researchers. The study aimed to understand better the role of gamification on the self-regulated learning process of elementary students learning English as a foreign language.\n\nSix students from second, third, and fourth Elementary grades between 8 and 10 years old were randomly selected (four girls and two boys) from a private school with over 40 years of experience. Known for its high academic standards and accreditations from global agencies like NEASC and CIS, the school emphasizes forming ethical, responsible global citizens. The sample was small but randomly chosen to ensure credibility and reduce selection bias, aligning with the study’s goal of exploring in depth the shared characteristics of a group exposed to similar educational experiences. There was no specific criterion for the selection of the participants, nor for the type of school, and the decision to limit the sample size was guided by practical constraints, including available resources and the employment relationship of the main researcher with the school, which facilitated convenient access to the participants. Despite these constraints, the study’s design enables a focused exploration of self-regulated language learning experiences among a group of students who, while limited in number, represent a consistent sample in terms of socio-economic background and a similar history of exposure to English as a foreign language since preschool. All participants’ first language was Spanish.\n\nThe research utilized a qualitative-dominant mixed-methods design supported by quantitative tools. Mixed methods, as described by Johnson and Onwuegbuzie (2004), combine qualitative and quantitative approaches to comprehensively address research questions. This study employed a convergent design to merge qualitative insights from interviews and observations with quantitative data from surveys and performance tests, providing a holistic view of SRL development.\n\nMixed methods enrich understanding by integrating diverse perspectives and validating findings through triangulation. The approach enabled an in-depth exploration of SRL processes and the gamified tool’s impact on students’ learning experiences.\n\nFor the present research, moral and legal codes were considered regarding the ethical treatment and care of the participants involved with research studies (Saldaña, 2011). The present study was approved by ACT 23-2024 (October 30. 2024), by the Research & Ethics Sub-Committee from the School of Education at Universidad de La Sabana, the document states that “The Research and Ethics Sub-Committee of the Faculty of Education approves the project “Virtual Game-Based Learning Environments to Promote Self-Regulated Learning Skills in Foreign Language Learners”, study conducted by the student Maira Noriega Cortés (main author) and Professor Laura Lucía Carreño (corresponding author), the study demonstrated validity, reliability and attached as evidence of the guidelines followed, the signed consent forms by participants, institutional endorsement letter, and instruments”. Based on the Research and Ethics Sub-Committee guidelines and the globally accepted guidelines for researcher conduct outlined in in UNESCO’s Universal Declaration on Bioethics and Human Rights, for this research it was of special interest to take care of the participants by informing them about the study, sharing the characteristics of their participation, protecting their identities by using pseudonyms, and clarifying that no physical or emotional harm would be caused.\n\nFurthermore, it was explained that the research was being conducted for the benefit of the community, that the information was being guarded and used only for research purposes, and that a responsible action was being performed by being open to all the questions that participants and related members of the community could have. As the main participants were children, only the students whose parents opportunely signed the consent form were included in the research. In addition, the school administration was informed about the project and signed a consent form.\n\nThe methodological design of the virtual-learning environment, was the skeleton behind the project and focused on three aspects:\n\n1. The structure was based on the self-regulated learning model proposed by Zimmerman (2000) and established three stages to the self-regulated learning process: a forethought (see Figure 1), a performance (see Figure 2) and a self-reflection (see Figure 3).\n\n2. This structure worked because of a system of strategies for learning a language proposed by Oxford (1990), that considered metacognitive, cognitive, affective, and social learning strategies.\n\n3. Finally, it followed a pedagogical route given by the inquiry-based learning model from Bruce and Bishop (2002), so that the communicative tasks took place throughout a cycle of five stages: ask, investigate, create, discuss and reflect. This responded to a specific problem or mission, based on what is established for the levels of A1 and A2 proficiency according to the Common European Framework.\n\nFinally, for the design of the prototype a narrative in which students became travelers, visiting different places where it would be necessary to use English as a foreign language was proposed. The final version prototype of the virtual-learning environment used for this research, was design through the video game engine Unity, and represented one level of the whole game, essentially, one mission that takes place in New York City with 3 essential parts:\n\n1. Introduction (New York Airport): The player is welcome to the city and a role for the mission is assigned. For the 1st mini game, the player must decide the means of transport to get to Central Park:\n\n• Taxi: Use of resources (Cognitive strategy): the player must find a destination to visit from a New York tourist guide with specific descriptions.\n\n• Subway/Bike rental: Overviewing and linking (Metacognitive strategy): the player must find a specific area of the park to visit in a Central Park tourist guide with a list of five places of different characteristics.\n\n2. Main activity (Bethesda Terrace in Central Park): The three phases of Self-regulated learning are accomplished considering the inquiry-based learning cycle. It contains five mini games that support the development of the story and two types of evaluations (three self-assessments of motivational beliefs and one self-assessment about the learning process).\n\n• Anticipation: This stage includes a description of the mission and the first self-assessment of motivational beliefs.\n\n○ 2nd mini game: Goal Setting (Metacognitive Strategy): the player must go through a map of the area in Central Park looking for signs with missing letters. The letters are the missing pieces of a sentence that becomes the learning objective for the mission.\n\n• Development:\n\n○ Ask and investigate: the player must select from a series of questions, each of which leads to a different learning strategy:\n\n▪ 4th mini game: Contrast (Cognitive strategy): Pairs of stamps must be made based on the words in English and their corresponding translation in Spanish.\n\n▪ 5th mini game: Keywords (Cognitive strategy): Different words that are part of the vocabulary of the mission must be formed by organizing a series of rocks with letters located in a section of the park.\n\n▪ 6th mini game: Cultural understanding (Social strategy): by selecting the images of a magazine stand that are related to each other, the player unlocks the photo and the name of another place in New York City in which it is necessary to use the presented keywords.\n\n○ Create: it begins with the second self-assessment of motivational beliefs.\n\n▪ 7th mini game: A sliding puzzle must be solved to assemble a sign located in an area of Central Park.\n\n▪ 8th mini game: Monitoring (Metacognitive strategy): the player must pop the bubbles that contain the missing words of the sentence which describes some self-monitoring tasks.\n\n○ Discuss: a postcard must be written to describe the most important memories of the player’s visit to Central Park. The postcard can be shared with someone else (partner/teacher) for feedback.\n\n○ Self-reflection: it begins with the third self-assessment of motivational beliefs.\n\n▪ Self-assessment: The player scores each statement with different emojis according to his/her performance throughout the mission.\n\n3. Closure (Central Park): a specific task related to the mission and the proposed learning objective is performed to conclude the trip to the city.\n\n○ The player must go through the map of Central Park to see the signs of different areas and find the most appropriate according to the proposed mission.\n\n○ The player also makes a record of the learned words according to the learning objective proposed at the beginning of the mission.\n\nTwo primary instruments were designed for data collection: a semi-structured observation format (see Table 2) and questionnaires. These tools were chosen for their ability to generate both qualitative and quantitative data, aligning with the study’s mixed-methods approach. Existing validated instruments, such as the Self-Regulation Assessment Suite (Heger et al., 2022) and the Intrinsic Motivation Inventory (Ryan, 1982), informed the design process.\n\nAll the participants filled in a pre-questionnaire (see Table 1) with 11 questions on a Likert scale that went from Never to Always, to identify the self-regulated skills they already had as part of their everyday learning activities.\n\n1 Original pre-questionnaire is in Spanish, but it was translated for publishing purposes.\n\nAfter completing the pre-questionnaire, participants took part in the piloting session of the design virtual-learning environment in which a semi-structure observation was implemented to identify what aspects of self-regulated learning the students could recognize and perform while using the tool. It also served the purpose of collecting information about how the students responded to the use of the tool in terms of its usability (learnability, efficiency, errors and satisfaction). The observation protocol contained 22 observable behaviors and was conducted over three sessions of around 45 minutes, each per couple of students selected per grade.\n\nOnce the piloting session was over, students filled in a post-questionnaire (see Table 3) with 17 questions to identify the self-regulated learning skills students used while playing and the game experience in terms of the usability of the digital tool.\n\n1 Original post-questionnaire is in Spanish, but it was translated for publishing purposes.\n\nDescriptive statistics were used during the first stage in data analysis to understand quantitative data collected through the pre-and post-questionnaires about students’ performance in self-regulated learning skills before and after using the digital tool. The purpose behind following this model was to organize the information and translate it into distribution of frequency, percents, and averages, that provide a summary of relevant numerical information to describe the selected sample, in tables and graphs (Kaushik & Mathur, 2014). Through this analysis it was possible to understand the relationship between students’ behaviors and their learning experience, by visually representing the frequency of use of a particular behavior, as well as the percentage of increase in the general use of self-regulated learning strategies, before and after using the designed digital tool.\n\nDuring a second stage a thematic analysis framework was chosen to analyze the qualitative data collected with the observation protocol. It was selected as an analysis procedure since it attempts to find repeated meaning across a data set facilitating the interpretation of a particular phenomenon, after following a rigorous system that includes collecting the data, engaging with it, codify, generate code categories, conceptualize themes, and contextualize and represent the findings (Braun & Clarke, 2006; Peel, 2020; Xu & Zammit, 2020). Consequently, data from the observation protocol provided information in terms of recurring ideas, behaviors, and strategies exhibited by the students while using the prototype from beginning to end. The analyzed categories are included in the Results section and a comparison with previous research in the Discussion and Conclusion sections.\n\nFindings\n\nAfter careful analysis of the data collected through the observation protocol and the two questionnaires, codes were established for mapping, categorizing and contrasting the qualitative and quantitative data, resulting on a core category and two final categories, each with two subcategories (see Figure 4).\n\nEventually, this mapping attempted to describe the effects in the development of self-regulated learning skills in elementary students, due to the implementation of a virtual game-based learning tool based on gamification strategies, specifically around the successful learning process of English as a foreign language.\n\nThis final category includes all the data in relation to participants profile of learning based on their self-perception regarding the use of self-regulated learning strategies, and their profile of performance based on the strategies they implemented while using the gamified tool.\n\nIn terms of their self-perception of the use of self-regulated learning strategies daily for learning, they frequently question or ask for support when facing challenges during a task, which are common and basic social strategies they use to understand and indicate involvement inside the classroom. In contrast, their less frequent practices include establishing personal goals of learning before starting an activity and monitoring their own progress to modify the implementation of a particular strategy. In general, they perceived themselves performing between three (27%) and eight (73%) of the 11 behaviors in the pre-questionnaire almost all the time, which describe a profile of learning with some development of self-regulated learning as part of the school environment and learning exposure (see Table 4).\n\nOn the other hand, their profile of performance was analyzed by paying attention to students’ behaviors while playing during the piloting session (see Table 5). In correspondence to students’ perceptions, while using the tool students were aware of their emotional states, sought adult support when facing challenges, and identified their strengths and needs based on the difficulty of every game. Each student had a unique profile that influenced their learning and use of strategies, which supported the observable evidence that a minimal development of these self-regulated learning abilities is required if the tool is to be used independently by students. Part of learning technologies effectiveness depends on students’ prior experiences, skills, personalities, and circumstances, inevitably students with attentional or regulatory difficulties often would need significant teacher support during group exercises to follow instructions, stay focused, and identify strategies for success. This highlights the importance of pairing such tools with explicit instruction in self-regulated learning and learning strategies to maximize their benefits inside and outside the classroom.\n\nThe post-questionnaire also provided information about students’ profile of performance in which participants perceived themselves confident and with the necessary skills to perform all the proposed tasks, having different options of strategies to achieve the learning objectives of the mission. In contrast, they felt they did not consciously monitor their progress during the game, and some struggled to maintain attention due to the interaction and game dynamics with their partner, which also evidence consistency with the observable behaviors during the piloting, and their answers on the pre-questionnaire.\n\nThis final category includes all the data in relation to participants recognition and perceptions of tool functioning considering usability aspects such as learnability, efficiency, errors, and satisfaction. Overall, all students understood the tool, completed tasks within the expected time, and reacted positively to its narrative and appearance. Students first learned the game’s functional aspects, like dragging or clicking with the mouse, and although they struggled while discovering technical features, once they grasped the general mechanics they navigated the tool easily. The results from the post questionnaire in relation to the usability of the tool, reflected students’ enjoyment of the tool considering it fun, creative, exciting and interesting, which correlates with their positive verbal and physical responses observed while using the tool. Some students referred occasional difficulties understanding instructions, which also comes as a result from those participants who out of impulsiveness needed more time and peer/teacher support to adjust to the game mode and dynamics.\n\nAt the end of the piloting sessions, it was a recurring idea for the participants the ways in which the tool could be a useful resource in English class to learn vocabulary, language use, and even develop a grater comprehension about cultural aspects. Nonetheless, none perceived the tool as a resource to perform and improve their self-regulated learning process, nor identify these strategies as learning assets to improve their ways of learning in other contexts.\n\nThe whole process of testing the tool included the three stages of self-regulated learning and allowed students to use a set of eleven language learning strategies. Following the implementation of the tool, students’ self-assessments demonstrated a more lucid comprehension of the pedagogical objectives underlying the assigned task. Additionally, there was a marked increase in their self-efficacy and confidence in their abilities to attain these objectives. The use of diverse learning strategies became more prevalent, facilitating their achievement of desired outcomes. Furthermore, the tool provided an expanded scope for collaborative engagement with peers (see Figure 5), thereby enhancing the overall learning experience.\n\nLikewise, when stablishing a comparison between the percentages of use from the pre-questionnaire and the post-questionnaire, all the students had an increase of use from between 9% and 36%, in those self-regulated learning behaviors that occurred always as part of their learning process. This implies that employing a gamified tool with a coherent, systematically and attractive designed framework, which facilitates students’ guided exploration of various self-regulated learning strategies and influenced students’ engagement, emotions and behaviors, can significantly enhance their self-regulated learning performance.\n\n\nDiscussion\n\nThe findings of this study indicated that a structured tool guiding the learning process and incorporating gamification significantly enhances students’ purposeful recognition and implementation of self-regulated learning strategies. Employment of this tool enabled students to approach tasks with a clear objective, thereby reinforcing their confidence and expanding their repertoire of learning strategies within diverse game-based scenarios. Collaborative efforts were also prominently integrated into students’ performance, enriching their application of social strategies to seek support, and receiving comprehensive feedback on language use. As such, it is of higher value in educational search to understand which factors can influence the fostering of self-regulation towards greater motivation and academic achievement.\n\nFirst, it is important that students can approach self-regulated learning consciously and explicitly by utilizing resources that provide a safe, engaging, and scaffolded environment. Previous research has addressed that instruction interventions in self-regulated learning help learners intentionally and goal-directedly enhance their language learning. Metacognitive, social, affective and cognitive interventions should include explicit instruction that emphasizes consciousness-raising regarding the whole learning process through modeling of strategies for goal setting and planning, guiding practice within a safe environment to internalize strategies in a structured manner, promoting self-monitoring, and providing feedback and evaluation to identify strengths and weaknesses (Azatova, 2021; Bai & Guo, 2018; Chen, 2022).\n\nOn the other hand, tool design and gamified elements should be considered given that, as evidenced in this research, a score based on points that students obtained by using learning strategies, penalization for not accessing the storage of strategies to complete mini games, and offering students a total score with visual rewards at the end of the mission, were game elements that increase participants’ motivation, self-efficacy and enjoyment in the learning process. This is also supported by previous research that describe how gamified elements like narrative and storytelling, progress tracking bars, levels, personal profiles, experience points, badges, rewards, bonuses, penalties, challenges and skill trees, are elements that relate to autonomy and competency, provide feedback during the learning experience, and enhance students’ performance and participation by making the learning more engaging and interactive, while supporting improved academic outcomes for a diverse student population (Chapman et al., 2023; Chapman & Rich, 2018; Nabizadeh et al., 2021).\n\nIn the end, this research investigated the impact of a virtual game-based learning tool employing gamification strategies on elementary students’ development of self-regulated learning skills, and results from piloting sessions showed positive effects on participants’ learning outcomes and motivation. This study contributed to understand the importance of early instruction in self-regulated learning processes for elementary students, aligned with the findings on the emergence of self-reflection and metacognitive skills in children aged 8 to 10 and younger (Goetz et al., 2013), and highlighting the impact that early instruction could have on students’ foundational stage for their education. Additionally, the creation of the virtual learning environment was a valuable contribution based on the analysis of a wide range of theoretical and practical information, that exposed how self-regulation can be promoted while students completed gamified tasks and learned another language. The tool’s instructional design, narrative, visuals, and game elements significantly enhanced students’ engagement and motivation, pointing out the value of gamified learning experiences in fostering self-regulated learning.\n\nFurthermore, the research underscores the significance of preparing students for success in a knowledge-driven society by developing 21st-century skills, including self-direction, collaboration, critical thinking, emotional intelligence, initiative, and global awareness. Integrating tools like the one designed for this research into educational settings to equip students with essential skills beyond language acquisition or standardized testing, is key to cultivate students’ autonomy, self-awareness, and adaptive learning strategies from an early age to meet future challenges effectively. As such, this can be a replicable research experience to continue raising awareness about the broader implications of implementing such tools across diverse educational contexts, and prompting considerations about how students from various sociocultural backgrounds can develop self-regulated learning skills.\n\n\nConclusion\n\nSelf-regulated learning is a core competency to prepare learners for modern society’s challenges at large, making it crucial to provide meaningful opportunities for students to understand, learn, practice and refine these skills and routines in any knowledge environment they experience (Boekaerts & Corno, 2005; Goetz et al., 2013; Li et al., 2022). The study concluded that the designed digital tool effectively facilitated students’ exploration of self-regulated learning strategies through the processes of forethought, performance, and self-reflection in a language learning task. The gamified elements of the tool promoted engagement and positive emotional responses to learning challenges. However, the tool alone is insufficient for a deep understanding of self-regulated learning, highlighting the need for explicit instruction and practice in language learning strategies alongside the tool’s use. Moreover, there is a need to integrate more theory and research into language teaching pedagogies to establish best practices for introducing self-regulated learning to elementary students. The research demonstrated that gamified virtual environments significantly enhance student engagement in language learning tasks, making learning enjoyable and interactive, turning this approach into a direct strategy to foster self-regulated learning in young learners, promoting early intervention benefits that extend beyond language learning to the acquisition of lifelong skills.\n\nA few limitations must be considered when interpreting the results of this study. Firstly, while the gamified tool employed in the research incorporated Zimmerman’s three stages of self-regulated learning, it only examined the effects of 11 selected learning strategies, potentially overlooking the influence of other strategies on outcomes. Additionally, due to constraints in prototype development, the study could not explore multiple levels of the game or extend testing over a longer period to explore long-term impact and applicability to a wider student population, limiting insights into the tool’s efficacy with a broader array of strategies and sustained use. Consequently, the study’s findings are restricted by the short duration of exposure to the tool and the small participant sample, limiting the ability to generalize qualitative findings. These limitations underscore the need for further research with larger and more diverse samples to better understand the potential benefits and challenges of integrating gamified learning tools into elementary education (Alshammari, 2020). Small datasets like the one in this study can contribute to future research by being included in meta-analyses, and the findings can eventually be combined with similar research, increasing the overall value of the collected information.\n\nFurther research avenues in the field of promoting self-regulated learning skills among elementary students can enhance both teaching practices and educational outcomes. Firstly, investigating teachers’ perceptions of gamified tools and their readiness to model and teach self-regulated learning skills to determine the best ways to keep promoting effective development of these skills in elementary students. Additionally, assessing the learning strategies students already use as part of their learning context and their effectiveness when using virtual tools more autonomously is essential. This analysis can provide personalized learning approaches tailored to individual students’ needs and preferences, facilitating the development of self-regulated learning skills through a scaffolded pedagogical approach. Finally, documenting the effects of interventions in language classrooms, beyond language acquisition, is imperative to understand how explicitly teach self-regulated learning strategies, foster a more self-regulated learning environment in the language classroom, and complement digital tool usage, recognizing the effectiveness of these interventions in younger learners, including preschoolers and elementary students. These research directions promise to enrich the understanding and implementation of self-regulated learning interventions in elementary education, paving the way for more effective teaching practices and improved language learning outcomes.\n\n\nEthics and consent\n\nFor the present research, moral and legal codes were considered regarding the ethical treatment and care of the participants involved with research studies (Saldaña, 2011). The present study was approved by ACT 23-2024 (October 30. 2024), by the Research & Ethics Sub-Committee from the School of Education at Universidad de La Sabana, the document states that “The Research and Ethics Sub-Committee of the Faculty of Education approves the project “Virtual Game-Based Learning Environments to Promote Self-Regulated Learning Skills in Foreign Language Learners”, study conducted by the student Maira Noriega Cortés (main author) and Professor Laura Lucía Carreño (corresponding author), the study demonstrated validity, reliability and attached as evidence of the guidelines followed, the signed consent forms by participants, institutional endorsement letter, and instruments”. Based on the Research and Ethics Sub-Committee guidelines and the globally accepted guidelines for researcher conduct outlined in in UNESCO’s Universal Declaration on Bioethics and Human Rights, for this research it was of special interest to take care of the participants by informing them about the study, sharing the characteristics of their participation, protecting their identities by using pseudonyms, and clarifying that no physical or emotional harm would be caused.\n\nFurthermore, it was explained that the research was being conducted for the benefit of the community, that the information was being guarded and used only for research purposes, and that a responsible action was being performed by being open to all the questions that participants and related members of the community could have. As the main participants were children, only the students whose parents opportunely signed the consent form were included in the research. In addition, the school administration was informed about the project and signed a consent form.", "appendix": "Data availability\n\nOpen Science Framework (OSF) repository: Virtual game-based learning environments to promote self-regulated learning skills in foreign language learners, DOI: 10.17605/OSF.IO/MQT3K. (Noriega-Cortés & Carreño-Bolívar, 2024).\n\nThis project contains the following underlying data:\n\n• Data Matrix_NORIEGA-CARREÑO 2024.xlsx\n\n• Data Matrix_NORIEGA-CARREÑO 2024 ENGLISH.xlsx\n\nData are available under the terms of the CC0 1.0 Universal\n\n\nReferences\n\nAcar IH, Veziroğlu-Çelik M, Rudasill KM, et al.: Preschool children’s self-regulation and learning behaviors: The moderating role of teacher–child relationship. Child and Youth Care Forum. 2022; 51(1): 1–18. Publisher Full Text\n\nAlshammari MT: Evaluation of gamification in e-learning systems for elementary school students. TEM J. 2020; 9(2): 806–813. 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Academic Press; 2000; pp. 13–40." }	[ { "id": "355097", "date": "14 Jan 2025", "name": "Mahjabin Chowdhury", "expertise": [ "Reviewer Expertise Game-based learning", "Scholarship of teaching and learning", "translanguaging", "Co-design", "learning experience design" ], "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, this is a well-conducted study that makes a valuable contribution to understanding how gamified environments can support SRL development in young language learners. With some minor revisions to strengthen methodological clarity, it will be even more valuable to researchers and practitioners. Recommendations for improvement:\nMethods section should include:\n\nMore details about observation protocols and coding Information about instrument validation Clearer explanation of data analysis procedures\n\nResults section would benefit from:\n\nMore direct quotes from qualitative data Additional details about individual student progress Clearer presentation of pre/post comparisons\n\nDiscussion should:\n\nMore explicitly address study limitations Provide more specific recommendations for practitioners Include deeper analysis of unexpected 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": [] }, { "id": "355106", "date": "27 Jan 2025", "name": "Mohammad Taufiq Abdul Ghani", "expertise": [ "Reviewer Expertise educational technologoly", "teaching foreign langauge", "education", "instructional design", "instructional technology" ], "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\nExpand Literature Review: Include recent studies on digital game-based learning and SRL to strengthen the theoretical foundation and provide a more comprehensive background. Clarify Instructions in the Tool: Some students reported difficulties understanding instructions. Consider refining the clarity and conciseness of the instructions in future iterations of the tool. Address Generalizability: Discuss the generalizability of the findings given the small, homogenous sample size. Future research could explore the tool's efficacy in diverse educational contexts. Explicit SRL Instruction: Highlight the need for explicit instruction in SRL strategies alongside the use of the tool, as evidenced by students' inconsistent self-monitoring and reflection behaviors.\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? Yes", "responses": [] } ]	1	https://f1000research.com/articles/13-1541
https://f1000research.com/articles/13-1540/v1	20 Dec 24	{ "type": "Review", "title": "The Konark Temple’s Construction: A Critical Review of the Historical, Cultural, and Scientific Evidence", "authors": [ "Susanta Bahinipati", "Dipti Ranjan Biswal", "Damodar Suar", "Dipti Ranjan Biswal", "Damodar Suar" ], "abstract": "Drawing from historical accounts, cultural practices, archaeological discoveries, and indigenous technical knowledge, the research presents a critical review of varied aspects of the construction of the Konark temple and presents future research directions. To fulfill the purpose, documents were sourced from Scopus, Google Scholar, and ancient texts and palm leaves. Findings suggest the legacy of Narasimhadeva I, the Hindu monarch who remained undefeated during his reign, safeguarded the kingdom from encroaching Muslim rulers. The narrative highlights the bravery of a faithful elephant that revived the king on the battlefield. The Konark temple was built with the dual purpose of showcasing the glory of his victory and devotion to the Sun God. The ancient artisans’ knowledge, skills, and commitment were crucial in erecting this tallest temple structure. The investigation further illuminates the use of sea routes for transporting monumental stones, the ingenuity in laying the temple’s foundation, the selection of high-grade stones, the monumental task of lifting colossal stones, like the world’s heaviest stone hoisted to a height of about two hundred feet, the use of rust-resistant iron, and the application of advanced astronomical knowledge by ancient artisans. The study provides insights into ancient engineering ingenuity, encouraging further exploration of the enduring legacy of the Konark temple’s construction.", "keywords": [ "Konark Temple", "Construction Techniques", "Historical Significance", "Sundial", "Astronomy", "Indigenous Knowledge" ], "content": "Introduction\n\nThe Sun Temple of Konark, also known as the Konark Temple (KT) in Odisha, stands out among the multitude of exquisitely crafted temples spread throughout India. It has fascinated countless generations with its remarkable magnificence, radiance, splendor, and enigma. The KT is included in the list of seven wonders of India and the list of World Heritage monuments, as designated by UNESCO in 1984 (Dey, 2016; Donaldson, 2003; Jana et al., 2016). The KT with 19.8921° N, 86.0912° E, is situated 64 kilometers in the southeast direction from Bhubaneswar, and 35 kilometers in the northeast direction from the city of Puri along the coastline of the Bay of Bengal. The temple’s construction was initiated by the king, Narasimhadeva I, known as Langula Narasimhadeva, during the Eastern Ganga dynasty (Behera, 2005; Bussagli, 1971; Chakravarti, 1904; Chauley, 1997; Das, 2015; Dey, 2016; Donaldson, 1985; Edwards, 1969; Eschmann, et al., 1978; Ganguly, 1912; Jana, et al., 2021; Joshi & Srivastava, 2021; Mahapatra, 1989; Mohanty, 2001; Percy, 1959; Rowland, 1953; Swarup, 1910). The temple, dedicated to Surya Deva (Sun God), was constructed in the mid-thirteenth century. Despite significant deterioration, the remaining portions of the temple now exhibit the delicate craftsmanship of that historical period.\n\nThe construction of the temple began around the fifth anka (regnal year) of Narasimhadeva I’s rule, employing 1200 workers (Behera, 2005; Boner et al., 1972; Das, 2011; Dey, 2016; Donaldson, 1985; Mahapatra, 1989; Mishra, 1919; O’Malley, 1908). It took about 12 years, 10 months, and 14 days to finish, completing in the 18th anka of Narasimhadeva I’s reign (Dey, 2016; Donaldson, 1985). The temple was sanctified on a Sunday, namely on the day of Magha Saptami (seventh day in a lunar fortnight of January or February month), which fell on the 27th of January in the year 1258 A.D. (Dey, 2016; Donaldson, 1985). The occurrence of Magha Saptami on a Sunday is regarded as propitious and this event takes place just once in seven years which is also called Vijaya saptami (seventh day of triumph) (Swarup, 1910). Narasimhadeva I desired to consecrate the temple on a highly favorable day, and hence he persistently urged the Sutradhara (master architect) to finish the work within the planned timeframe.\n\nStudying the ancient temple of Konark is imperative for the techniques it embodies, which continue to influence modern architectural and engineering practices. Experts’ studies undertaken by the Council of Scientific and Industrial Research (CSIR), Central Building Research Institute (CBRI), and the Indian Institute of Astrophysics (IIA), demonstrate the sophistication and ingenuity of ancient craftsmanship. Analyzing the architectural feats of the KT, researchers gain insights to integrate ancient techniques with modern structures. It was evident in the Ram Temple in Ayodhya on 17 April 2024. The direct sunlight casting a ‘Surya tilak’ on Ram’s forehead facing east at specific times during Ram Navami (celebration of Lord Ram’s birth on ninth day of Chaitra: March or April) symbolizes the fusion of ancient KT’s technical knowledge with modern innovations. Furthermore, an exploration of the techniques used to transport and lift massive stone sculptures, along with the discovery of rust-resistant iron beams within the KT compound, showcases the engineering prowess of ancient times and emphasizes the importance of delving deeper into the construction methods and artisanal craftsmanship. The presence of colossal sculptures that have endured for generations also underscores the necessity of conducting comprehensive studies to uncover the intricacies involved in such aspects of KT.\n\nBeneath the KT grandeur lies a tapestry woven with controversies, myths, and scholarly debates, each thread contributing to the intricate narrative surrounding its construction. We examine the temple’s construction purpose, the socio-economic conditions of the kingdom at the time, and the temple’s original dimensions obscured by its ruined state upon excavation in the early twentieth century. We further scrutinize the folk legends surrounding the temple and its completion, stone selection, transportation of large stones from distant quarries, and the indigenous technology used in constructing this colossal structure.\n\nAgainst this backdrop, the study aims to present a critical review of the historical, cultural, and scientific evidence from past to present to provide a full account of the KT construction. These three aspects are not mutually inclusive. The structure of the paper is shown in Figure 1. Next, it mentions the sourcing of documents in the method section, followed by the reporting of the findings from the narrative inquiry on each of the nine issues. Then, the discussion section summarizes the findings, mentions theoretical and practical implications, limitations, provides directions for research, and concludes with a novel contribution to knowledge (Figure 1).\n\n\nMethod\n\nA systematic review methodology was adopted for documents search and selection. Documents were extracted from Scopus, the largest scientific database of journals, and Google Scholar, the automated, inclusive database. Those yield maximum access to published documents (Bar-Ilan, 2008). Documents were extracted using search strings “KONARK” OR “SUN AND TEMPLE” in title, abstract, and keywords. A total of 329 documents were extracted from Scopus and 912 were extracted from Google Scholar. Seventy documents relevant to the purpose of the study and related to architecture, construction, water bodies, and folklore connected to the KT were retained. As KT is an ancient structure, many documents appeared in books and ancient informal writings were considered relevant, 39 books and ancient writings were thoroughly read to satisfy evidence requirements. Figure 2 presents a flowchart outlining the process for selecting papers for a comprehensive review. The narrative analysis of these documents is reported in the next section.\n\n\nFindings\n\nThe KT was built with the dual purpose of expressing devotion and gaining fame. Narasimhadeva I is acknowledged in the field of art, thereby earning him the title of ‘Shilpajna’ (knowledgeable in sculpture), elaborated in Ekabali, written by Vidyadhara a Sanskrit poet of 13th century (Chakravarti, 1904; Donaldson, 2003; Mahapatra, 1959). The monument is believed to be a memorial dedicated to commemorating his military campaigns (Behera, 2005; Boner et al., 1972; Bussagli, 1971; Dey, 2016; Donaldson, 1985; Edwards, 1969; Ganguly, 1912; Jarrett, 1891; Joshi & Srivastava, 2021; Mahapatra, 1959; Mahapatra, 1989; Subbarao, 1939). The construction of the monument began shortly after his military triumph, and the campaigns persisted until the completion of the construction. The war concluded with his remarkable victory over the Muslim rulers (Banerji, 1930; Boner et al., 1972; Chinnappa, 1978; Donaldson, 2003; Panigrahi, 1986; Sarkar, 2003; Subbarao, 1939). This is manifested in several ornamental sculptures depicting war horses, diverse combat actions, and notably over 1,700 elephants (Donaldson, 2003), thereby substantiating the title Gajapati (Lord of elephants) (Eschmann et al., 1978; Fabri, 1974; Kulke, 1993: Kulke & Rothermund, 2016). As per the inscription of the Kapilash temple, Dhenkanal, Odisha, of 1246 A. D, he was the first king of Odisha to be bestowed the title of Gajapati, with a huge troop of elephants in his army. Few sculptures in the KT were created by soldiers who took part in the conflict. It depicts the firsthand experience of the victorious and joyful moment (Boner et al., 1972). In a separate palm leaf document, namely XLVII.20 (Boner et al., 1972), Sutradhara requested a skilled warrior as well as a sculptor to create a monument depicting the act of ‘Suasamala fighting’ while mounted on a horse. Observing the pair of life-size war horses at the south gate of the KT, an European art historian, E. B. Havell, expressed that the Indians showcased their pride in victorious warfare through the immense strength depicted in these sculptures, comparable to the fiery and passionate expressions found in the greatest European art (Havell, 1908). Upon observing the deteriorated condition of the impressively grand statue, he expressed that if this exceptional work of art were identified as ‘Greek’ or ‘Roman’, it would have been exhibited in a renowned museum in America or Europe (Behera, 2005; Dey, 2016; Ganguly, 1912; Havell, 1908). It is understood that Indians in ancient times possessed exceptional abilities to comprehend and effectively utilize their creative skills in harnessing the natural resources of their homeland.\n\nIn antonymous, a pair of galactic elephants standing at the north gate, there is a great memory of king Narasimhadeva I lying behind the elephant sculptures; which has been mentioned in the Madalapanji (a chronicle of Jagannath temple) (Boner et al., 1972; Dey, 2016). It was all about a day in the war field at Sasipur, when King Narasimhadeva I was stuck and got unconscious. Everyone on the battlefield searched for the dead body, suspecting the king had been killed. In the meantime, a small elephant of the king, named Sudehi, carried him up to the camp with its trunk and saved his life. From that day, Narasimhadeva I liked him as his son; in consequence, a special team had been allocated responsibility to make a sculpture of Sudehi with the instruction that he should look real; the intention of the king was to immortalize him for all times.\n\nMark Roland Shand, a travel writer and conservationist (BBC) visited the KT on the back of an elephant named Tara. An interesting incident happened in front of the pair of elephants at the north gate when Tara performed “her ceremonial ‘pranam’ by lifting her trunk in salute” (Shand, 1992). Appreciating the pair of elephant sculptures he said that the colossal war elephant would be mistaken by the visitors on a moonlit night as real. Not only war and struggle have been portrayed in the decorated sculpture: there are vast varieties of the endless richness of life, living, profession, approach, originality, culture, music, sensuous themes and nymphs. Every part of the temple exterior is ornamented with details, endless varieties of work carved with minute detail, it is said that “Indians plan like titans, finish like jewels”. Even, Ananda Coomaraswami states that “it would be hard to find anywhere in the world a more perfect example of the adaptation of sculpture to architecture” (Coomaraswamy, 1913, p.75). The KT is an epic, where eternal truth had been encrypted in the form of art defying time through chiseling the stones.\n\nAbu’l Fazl commented in Ain-i-Akbari, suggesting that the king allocated the revenue of twelve years of the kingdom for the completion of the construction of KT (Jarrett, 1891). According to historical records, during his reign, Kalinga (ancient name of Odisha) experienced a period of great wealth. Contrarily, Abu’l Fazl acknowledges this fact by stating that it is a “mighty memorial to posterity” (Donaldson, 2003; Jarrett, 1891). Before ascending to the throne, he led the war as a commander and achieved numerous victories, which enabled him to amass substantial wealth. Returning from war, his mother proposed to utilize his newfound riches to construct a Sun temple in Konark (Dey, 2016; Donaldson, 1985) as a sacred place, like Biraja temple in Jajpur, Lingaraj temple in Bhubaneswar, and Jagannath temple in Puri, that was lagging with a supper structure. According to the Madalapanji, Anangabheemadeva III regularly approved annual budgets for the daily worship and upkeep of the temple, considering the offerings made during the puja ceremonies, the Seven Dhupas, as well as all the rituals, festivals, and special events. Anangabheemadeva III raised the sanction from 42,500 Kahanas (cowrie) to 52,000 Kahanas (Dey, 2016; Donaldson, 2003; Swarup, 1910). Additionally, the kingdom’s currency was changed from Kahana to gold coin, with 15 Kahanas being equivalent to 1 gold coin. This change symbolizes the kingdom’s wealth and prosperity.\n\nThe project commenced during the reign of Narasimhadeva I, when Sadasiva Samantaraya, the second minister of King Anangavimadeva III, was appointed as the chief architect for the construction (Mishra, 1919; Mohanty, 2001). The preliminary tasks, consisting of drafting blueprints, extracting stones from quarries, and hiring labourers, lasted nearly six years (Behera, 2005; Dey, 2016; Donaldson, 2003; Mahapatra, 1989; Swarup, 1910).\n\nIn 1837, James Fergusson visited the temple and created a drawing of the KT ( Figure 3), which provided a clear depiction of the temple’s physical condition at that time. According to Fergusson (1848), the height of the sanctum in the painting is around 140 to 150 ft, which is taller than Jagamohana (porch). The KT is encompassed by boundary walls measuring 261.2m (857 ft) by 164.6m (540 ft) (Donaldson, 2003; Edwards, 1969; Ganguly, 1912), with a height of 14 ft and thickness of 5 ft and 4 in (Ganguly, 1912). The architectural style of the Orissan temples (Kalinga style of architecture) is completely ‘Astylar’, meaning unmixed, pure, and consistent (Fergusson & Spiers, 1910). When comparing KT with other similar temples, Fergusson stated that it is significantly superior, particularly praising the grandeur of the edifice and its elaborate decoration, claiming that it is the finest in the world.\n\nIn Hindu mythology, the Sun God is believed to travel across the sky in a chariot drawn by seven horses, with his charioteer Aruna guiding the way (Basham & Rizvi, 1956; Bedbak, 1999; Rao et al., 2015). In the parapets of the porch, rise the dynamic standing figures of celestial musician-girls playing flutes, drums, strings and cymbals to accompany the chariot for its daily course. Along with myriad dancing girls in the form of Odissi dance, anterior to the seven horses carved bas-relief in the wall of nata mandapa (dance hall) to chronicle the journey of the Sun God. This amusing architecture is a larger scale of European baroque art of the late 16th century which has been mentioned by Fabri Charles Louis, Hungarian Archaeologist and Historian (Fabri, 1974). With original thinking in design, the architectural design was to appease the Sun God (Figure 3).\n\nThe KT is thought to have consisted of a main vimana (sanctum), a jagamohana, a nata mandapa that is currently lacking in its superstructure, and a bhogamandapa (offering hall) that is in a state of ruin. Adjacent to these constructions, there were additional minor temples devoted to various deities, including the Aruna pillar which was relocated to Puri during the Maratha period and placed in front of simhadvara (main gate) of Jagannath temple (Behera, 2005; Boner et al., 1972; Dey, 2016; Donaldson, 2003; Mahapatra, 1989; O’Malley, 1908; Stirling, 1825; Sterling, 1846; Swarup, 1910; Tripathy & Kulke, 1987). The porch has been relatively well-preserved, but it was filled with sand in 1903 on the directive of Sir John Woodburn, the Lieutenant Governor of Bengal. This measure was necessary at the time to protect the monument (Donaldson, 2003). The deadening process of filling the sand to preserve the Jagamohana, was completed early in 1905 (Swarup, 1910). Indeed, the porch is refilled using stone masonry with lime, dry stone masonry with lime and sand, layered in various compositions (Figure 4). The current porch lacks the main temple, as it has fallen due to multiple reckless depredations. The underlying mode of this monument is a colossal chariot adorned with twelve pairs of intricately carved wheels, designed for the Sun God.\n\nThe first measurement of the KT was made by Gajapati Purusottama Deva during his 4th anka in the year 1610 A. D (Dey, 2016; Donaldson, 2003), in which the height of the sanctum estimated was 68.5m (224.73 ft). Again, the measurement was undertaken in 1627 A. D, in the presence of Gajapati Narasimhadeva II, who was the son of Purusottama Deva and the grandson of Ramachandra Deva. The task was carried out by Natha Mahapatra, under the supervision of Bakhar Khan, who served as the subedar of Orissa under the Mughal emperor Jahangir (Chakravarti, 1908). The Sanskrit version of the Madalapanji, has been acknowledged as an authentic account (Behera, 2005; Ganguly, 1912; Mishra, 1919; Swarup, 1910). Translating the Sanskrit version of the Madalapanji, it was observed that Gajapati Narasimhadeva II inspected the vacant temple and recorded its dimensions. The temple complex was measured using a native method that took into account the finger width of the king. A unit called kathi (stick) was established, with 28 angula (finger width) being equivalent to one kathi. Local calculations have been revised to correspond with modern measuring standards, accounting for the width of an adult’s finger, with one kathi equating to 19 in. To corroborate the local calculations, the height of the porch had been considered, the only surviving segment of the monument, for which the height is known. During the excavation, Swarup (1910) meticulously measured almost every part of the temple complex, focusing especially on the height of the porch; the height of the porch from the platform level was recorded as 112 ft and 5 in, excluding the Kalasa (finial) and Dhwaja (flag), which were absent, instead, a piece of iron still protruded from the center of Amala (crown stone) ( Figure 4), supporting both. According to measurements, the platform height of the Temple from the plinth is recorded as 13 ft and 3 in. (Ganguly, 1912; Swarup, 1910). Consequently, the height of the porch above ground level is calculated to be 125 ft and 8 in, not including Kalasa and Dhwaja. Ganguly (1912) calculated the height of the porch above ground level utilizing theodolite which was found to be 129 ft and 8 in. Referring to measurements taken during 1627 A. D, the height of the porch was recorded as 73 kathi. Converting this to feet by multiplying by 19 in, it was found to be 1387 in, which equals 115 ft and 7 in. Moreover, with the inclusion of the height of the platform, which is 13 ft and 3 in, the total becomes 128 ft and 10 in. This closely aligns with the calculation of Ganguly, deviating by 3 ft and 2 in from Swarup’s measurement due to the omission of the height of Kalasa and Dhwaja. Including the height of the porch, the measurement taken in 1627 A. D was factual. The measurement of the sanctum was conducted precisely, considering various segments. For instance, the measurement from below the lion upon Amala to the platform is 87 kathi, with an additional 12 kathi and another 12 kathi from the lion to Garuda, totaling 111 kathi which equals 175 ft and 9 in. Similarly, the measurement from above the lion of the Amala is recorded as 21 kathi 3 angula, equivalent to 33 ft and 5 in, excluding the Kalasa and Dhwaja, which were absent during the measurement. Thus, the total height of the sanctum from the platform is 209 ft and 2 in without Kalasa and Dhwaja. According to the measurements specified in the Madalapanji, the combined length of the Kalasa and Dhwaja is recorded as 3 kathi 8 angula, which is equivalent to 5 ft and 3 in. Including the height of the Kalasa, Dhwaja, and the platform from the plinth, which is 13 ft and 3 in, the total height of the sanctum reaches 227 ft and 8 in. This matches with Ganguly’s calculation of 228 ft (Ganguly, 1912; Behera, 2005; Dey, 2016). Accordingly, the measurement taken in 1627 A. D was accurate, and the KT was the tallest temple in Odisha (Figure 5).\n\nDuring the measurement of the temple, the stone kalasa, adorned with a lotus finial, was absent from the sanctum, similar to the porch’s state when Swarup undertook the measurement. However, the iron rod mentioned by Swarup, which was positioned above the porch and referred to in the Madalapanji as “Chumbaka luha dharana” (the magnetic iron rod), remained in place and projected above the top of the sanctum.\n\nBased on the interaction with the nearby villagers, Sterling (1846) mentioned about a massive lodestone, which was believed to possess magnetic power capable of disturbing the direction of ships navigating through this route. Allegedly, a group of discontented Muslim sailors (Mughals) from troubled ships removed the magnetic stone (Behera, 2005; Donaldson, 2003; Hunter, 1872; Jana et al., 2021; Laurie, 2000; Lowe, 2016; Rath, 2021; Sterling, 1846), akin to the tale of ‘Sinbad the Sailor’s Rock,’ in hopes of resolving the navigation issues during their voyage. Forthwith, the priests migrated to the shrine in Puri. While a more precise conclusion remains contingent on affirmative evidence, an alternate rendition of the legend suggests that the image of the Sun God was suspended in the air through the use of magnets. Considering the memory of people, another legend revolves around the loadstone and the Kalasa atop the sanctum. It recounts the story of a person who completed the sanctum by installing the Kalasa. The story also speaks about the skilled manpower who constructed the temple.\n\nGiven the absence of authentic records detailing the construction of the KT (Dey, 2016; Jana et al., 2021), the approach was to explore incidents recounted and heard from earlier generations by the local people. Among enigmatic legends encircling the construction of the KT, the most prevalent tale is of ‘Dharama’ or what localities recall as Dharmapada. He was the son of Bishu Maharana (Jana et al., 2021; Rath et al., 2015), the chief craftsman in charge of the construction of the temple. Legend has it that, over twelve years, 1,200 carpenters and masons worked on constructing the temple. Before construction began, Bishu Maharana, the master craftsman, departed for the site as per the King’s orders, leaving behind his pregnant wife. Days later, she gave birth to a son named Dharmapada. After twelve years, Dharmapada learned that his father was constructing a great temple for the king, sparking his interest in meeting him. He sought permission from his mother to embark on a journey for this purpose. His mother then gave him the fruit of Barakoli (Apple Ber) from the tree in their courtyard as a symbol of recognition. After a long journey, he arrived in Konark and began selling fruits near the construction site. Bishu Maharana recognized the fruit from his courtyard and assured the boy that he was none other than his son. However, the joy of their union was short-lived as Dharmapada learned of a decree issued by the king. If the craftsmen failed to complete the sanctum up to the Kalasa by that night, the 1,200 craftsmen would face execution following morning. Upon hearing this, that very night Dharmapada ascended the temple and, with his craftsmanship, completed the sanctum up to the ‘Kalasa’. The following morning, witnessing Dharmapada’s achievement, the artisans feared the consequences to be beheaded. Aware of risking their jobs and lives, they turned to Bishu Maharana for guidance. Reluctantly, he chose to save his fellow craftsmen over Dharmapada. Upon hearing this, Dharmapada immediately climbed to the top of the sanctum and jumped into the sea, sacrificing his life to save the 1,200 craftsmen. This narrative underscores Dharmapada’s selfless act for the greater good, set against the backdrop of the temple’s proximity to the sea.\n\nThis folktale, initially documented in 1876 A. D in “Dhagamala” by Kapileshwar Vidyabhushan, presents the adage “Barasha badheire daya ki puare daya” (whether responsible to the 1,200 artisans or responsible to the son). O’Malley (1908) collected and published the story in English, differing slightly from the popular folklore. He changed it, depicting Bishu Maharana sorrowfully accepting the preference for his fellow workers and climbing to the top where his son was still working, intending to hurl him down to the pavement below. Nationalist writers like Kripasindhu Mishra (Mishra, 1919) and Nilakantha Das (1919) supported O’Malley’s version. Pandit Gopabandhu Das (1924) introduced the name “Dharmapada” in his poem, emphasizing the theme of self-sacrifice. During the era of nationalism, Pandit Gopabandhu Das, a frontrunner in the Odiya nationalist movement, slightly tweaked the tale of Dharmapada to motivate the Odia youth, establishing the legend of self-sacrifice for the greater good. Ashvini Kumar Ghosh’s play, performed by the Annapurna group in Cuttack during 1950 A. D, revived the folklore and popularized the characters of Dharmapada and Bishu Maharana (Das, 2011; Dey, 2016). Ghosh also introduced the name “Bishu Maharana” for the first time in the play (Das, 2011).\n\nIt is evident that Dharmapada (Dharma Mahapatra) was indeed a real person (Boner, 1970; Boner et al., 1972; palm leaves: L7, L12, L13, LI 1-11, LIII 4, LIII 7). He was the son of Sadashiba Samantaray Mahapatra, also known as Shivai Samantaray (Boner, 1970), who served as the sutradhara. The palm leaves also mention Dharma Mahapatra’s significant role in completing the Kalasa, the crown stone of the temple. Additionally, other prominent artisans involved in the construction of the KT were: Narayana Mahapatra (master sculptor), Gadadhara Mahapatra (chief executive architect), Viswanatha Mahapatra (sculptor specialized in royal scenes), and Ganga Mahapatra (head of the silpis) (Baumer & Konishi, 2007; Boner et al., 1972; Boner, 1970).\n\nThis established historical fact serves as the foundation of these folklores that passed from one generation to another through words-of-mouth. The construction of the huge structure of the Konark in ancient times was a group endeavour of 1,200 artisans, assigned by the head of the kingdom, under the supervision of many master artisans. Threat to the artisans by the king was the means for the completion of the construction of the temple work and Dharmapada’s self-sacrifice was the action to protect the insult and life of artisans’ community in ancient times.\n\nAn anecdote of note revolves around Shivai Samantaray, the master architect of the temple complex (Mohanty, 2001; Rath et al., 2015). The initial task entailed the placement of stones in a section of the Padmatola river gorge, which was a pool covered with lotus plants. The objective was to construct the temple in this filled area. Nevertheless, the stone blocks, that were being dropped into the water, were being carried away by the powerful currents of the gorge. Shivai Samantaray regularly beseeched the Goddess Ramachandi, whose temple was situated near the building site, to proceed in the construction site. Once, the deity Ramachandi manifested in the form of an elderly lady and presented him with a portion of steaming Khiri (a sweetened porridge), for consumption. At that juncture, he was both disturbed and famished. Without any hesitation, he began consuming the gruel from the centre of the dish, which caused his fingers to be burned and subsequently pulled back and recoiled. The disguised goddess chuckled at him and advised him to consume the meal from the periphery, unlike Shivai Samantaray who was attempting to fill the gorge with stones, that is, from the centre instead of one side of the gorge. This incident served as a revelation for Shivai Samantaray, enabling him to effectively accomplish his task of filling the canyon. Upon receiving the solution from the elderly woman, purportedly the deity Ramachandi, Shivai Samantaray initiated the creation of an island by extending a bank and filling the gorge, thereby establishing a stable foundation for the entire structure.\n\nBased on a recent scientific investigation by the Central Building Research Institute (CBRI), Roorkee, the foundation of the KT consists of a block-type foundation that is 10.82 m deep (Dwivedi et al., 2022). The foundation comprises layers of Khandolite and Laterite stones, with the lowest layer being a dense sand stratum extending further for 11.5 m. In total, the foundation measures 22.32 m in depth (see Figure 6).\n\nThe most suitable foundation for an architectural plan in a sea beach location, according to modern engineering practice, is a block-type foundation with a sand footing over a compacted layer of dense sand. According to the norm, the foundation of a temple structure should typically be one-third of its height (Meister, 1985; Kak, 2017). Previous records indicate that the height of the platform from the plinth of the KT was documented as 13 ft and 3 in (Swarup, 1910; Ganguly, 1912). However, the current measurement in the cross-section indicates 4.12 m, equivalent to 13 ft and 6 in. Subtracting the platform height from the sanctum height, it amounts to 214 ft and 6 in. This comparison with the foundation, which measures 22.32 m (73 ft and 3 in), reveals a well-maintained height-to-foundation ratio, showcasing the remarkable technical expertise and skill of the sthapati (architect) of Kalinga. Geotechnical engineers’ research has established that the rate of consolidation of sand is higher than that of silt, clay, and other soils (Das & Sobhan, 1990), making it a suitable material for filling below the foundation. Sand can also be compacted rapidly by saturating it with water, thereby reducing the need for the application of compaction energy. The thickness and width of the khandolite stone and laterite stone layers below the platform level were carefully determined to achieve a load dispersion angle close to 60°, which is less than the dispersion angle observed with stones (Figure 6). Due to the high dead load of the 10m thick stone platforms and the temple itself, there is a risk of general shear failure and soil heaving. To mitigate this risk, artisans have incorporated 5.5m stone layers around the main stone foundations, demonstrating their sound engineering knowledge in the construction of the KT (see Figure 6).\n\nThe architectural framework of the temple and the sculptures were constructed using Khondalite stones, supplemented by Laterite and Chlorite (Dey, 2016; Donaldson, 2003; Nayak et al., 2017; Saxena & Srivastava, 2021). Laterite stones were mostly utilised in the subterranean section, situated beneath the plinth level, for foundational purposes; Chlorite stones were specifically employed for ornamental purposes, such as crafting statues of the Sun God, the Simghasana (pedestal), the puja image, Nabagraha (nine planet) statues positioned above the lintels, the Aruna stamba (Aruna pillar), sculptures housed within mundi niches, and other decorative elements found in the doorframes. The meticulous selection of stones was subjected to remarkably minimal decay over time, compared to other renowned structures such as St. Paul’s Cathedral and Norwich Cathedral. St. Paul’s Cathedral eroded by 30 mm in 250 years, and 95% of the outer stones of Norwich Cathedral were renewed during nine centuries, whereas the decay of the KT was only 2.4 mm (Behera, 2005). In an investigation conducted by UNESCO (Lemaire & Laurenzi Tabasso, 1981), the condition of the stones was found to be satisfactory following chemical testing of the existing materials. Despite the close proximity of the temple to the sea, the stones have not suffered significant damage, indicating the remarkable foresight of the people who selected stones, even in the 13th century.\n\nUpon examining the outskirts of Konark, it is evident that there are no stone quarries in close proximity, even within a range of 30-40 kilometers. Stones were obtained from various locations including Naraj, Narasinghapur, Siddha Durga, Jagadalpur, Tapang, Ghantasila, Neelagiri, and Khiching, from a distance of 40 to 290 km from the temple structure (Behera, 2005; Boner et al., 1972; Donaldson, 2003; Dey, 2016). When the temple was constructed, there were no efficient means of road transportation, and people did not utilise mechanised vehicles. People who visited the temple from the late eighteenth century to the early twentieth century noted that there was a lack of appropriate road communication (Boner et al., 1972; Cumberland, 1865; Fergusson, 1848; Hunter, 1872; Kittoe, 1838; Mishra, 1919; Mitra, 1880; Stirling, 1825; Sutton, 1833; Swarup, 1910). According to James Fergusson, waterways were utilised to transport stones that were subsequently looted from the KT and Barabati Fort in order to construct a lighthouse at False Point by the Europeans (Fergusson, 1848; Fergusson & Spiers, 1910). Consensus prevails that water served as the mode of transportation for conveying the stones to the site (Behera, 2005; Boner et al., 1972; Dey, 2016; Donaldson, 2003; Fergusson, 1848; Fergusson & Spiers, 1910; Jana et al., 2022; Mishra, 1919; Saxena, & Srivastava, 2021). Upon analysing the origins of the stones and the water connectivity, it is evident that, except Naraj, Narasinghapur, and Siddha Durga, there are no direct river connections to the work site. These three locations are in close proximity to the banks of the Mahanadi river. The Mahanadi river served as the lifeline of the region, intricately connected with other rivers. This was a key consideration in the Eastern Ganga dynasty’s choice to move their capital from Kalingapatnam to present-day city of Cuttack (Banerji, 1930; Chinnappa, 1978; Panigrahi, 1986; Rao, 1941; Singh, 1973; Sundaram, 1963). It enabled smoother travel along the Mahanadi river and its tributaries, enhancing communication and connectivity with other kingdoms. Additionally, the geographical position seemed to be central, facilitating interactions from both the north and south of the kingdom. In the era of the Eastern Ganga empire, the capital resided at the convergence of the Mahanadi river, precisely where it bifurcates into several branches, closer to the Barabati Fort cum Palace. One of the tributaries of Mahanadi, the Kushabhadra river, flows into the Bay of Bengal near the KT (Jana et al., 2018). Examining alternative stone quarries and transportation methods, the construction site presents a challenge to transport heavy materials upstream of rivers, such as from Jagadalpur, Tapang, and Ghantasila in the Chilika lake zone (Figure 7). The proposed route involves travelling upstream of the Daya river and then connecting with the Kushabhadra river downstream to reach the site. Nilagiri does not have a direct river connection (Acharya, 1955; Dey, 2016; Donaldson, 2003). However, there used to be a water path called Bhirudi Nala (Bhirudi Canal) that connected Nilagiri to the Salandi river downstream (Dey, 2016). From there, it was possibly flowing through the downstream of the Baitarani river and connected with the Birupa river, a distributary of the Mahanadi river upstream, eventually reaching Cuttack. To reach the site, one would then need to follow the course of the Kushabhadra river. Another location, Khiching, is easily accessible the downstream of the Baitarani river, and then the upstream of the connecting Birupa river, following a similar route as transportation from Nilagiri (see Figure 7).\n\nDuring the time of 1870 A.D., according to the estimation and firsthand account of Hunter (1872), the rivers had the ability to transport weight, taking into account their depth and width. Specifically, the river routes that were used to transport stones and other materials to the construction site of the KT could carry a capacity of 20-25 tonnes (Hunter, 1872), sometimes even more, particularly during the rainy season. There were lot many huge sculptures and stone blocks used in the temple which were more than 25 tonnes, one example of a large stone block is the monolithic Nabagraha slab weighed about 28 tonnes (Indian Museum, 1893), was originally over the lintel of the eastern door of the porch. As per eyewitness experience of Sterling, during his visit in 1822 A.D., that he had made a drawing of it when the Nabagraha slab was at its position, he compared it with that of the Gothic art of European architectural ornamentation, even in this dilapidated state looking at chlorite stone works around the doorways he said that the sculptures seem as if these came out from the chisel of the sculptors (Sterling, 1846). Even, when Fergusson visited in 1837 A.D., it was at its position ( Figure 3). Observing the Nabagraha slab missing from its original position, Ferguson said when he visited again around 1869 that “an abortive attempt was made to carry the lintel to Calcutta”. During British rule over India, an attempt had been made by the Asiatic Society to carry the Nabagraha slab to the Indian museum, Calcutta, on 11th January 1869 A.D., but failed due to its heavy weight and low fund allocation for the execution of the job (Indian Museum, 1893). As per the same report of the Indian museum, a second attempt had been made in 1892 A.D. by slicing the large piece longitudinally into two considering the uncarved portion of the slab from its back to reduce the weight; but again, failed due to the opposition from natives (Ganguly, 1912). Despite attempts to transport the slab via waterways, it proved unsuccessful, leading to the decision to transport it by road. In fact, the Nabagraha slab was observed loaded onto a truck in front of the temple (Mitra, 1880). Moreover, the Amala of the Sanctum was measured by the Archaeological Survey of India, which excavated a hole through the stone block, revealing a measurement of 25 ft and an estimated weight of 2,000 tonnes (ASI, 1906). Considering the logistic challenges posed by the river’s width and depth, it seems impossible to transport such a large stone via river routes; instead, sea routes must have been utilized for this purpose.\n\nThe tradition of river and sea routes is deeply rooted in Odisha’s history, reflecting its cultural heritage and maritime past. Festivals like Boita Bandana (ritual of boat worship) during Kartika Purnima (full moon day in October or November month) and Khudurukuni Osha (a traditional fasting ritual observed by unmarried girls in Odisha) honour this legacy, with symbolic boats set afloat to commemorate the state’s maritime heritage (Das, 2020; Guy, 1999; Sahoo, 2017). Additionally, festivals like Chaiti Ghoda (a folk festival featuring small wooden horse in March or April) and Brata Khulanasundari (vow of a legend Khulanasundari) celebrate the connection to maritime activities, with fishermen worshiping their boats and tales of sea voyages intertwined into folk practices (Behera, 1999; Das, 1991; Das, 2020). These festivals celebrated throughout the year, serve to preserve the memory of Odisha’s maritime history through folk traditions and tales. The interpretation of inscriptions found at Udayagiri and Khandagiri Caves in the post-Buddha period, it becomes apparent that ancient Kalinga was a prominent hub of trade and commerce (Prinsep, 1837). Due to its advantageous geographical position, Kalinga’s monarchs once wielded significant influence throughout India. Young princes received education in ‘Nawa-Byapara ’ (ship-commerce), highlighting the longstanding tradition of maritime trade and commerce education among monarchs dating back two thousand years. When English merchants established their first factory in Odisha near Harishpur Garh in the Mahanadi delta in 1633 A.D. (Wilson, 1895; Bowrey, 1993), under the leadership of Mr. Cartwright, they obtained permission from Mughal Emperor Shah Jahan at the state palace of “Malcandy” (Mughals’ renaming the Barabati Fort cum Palace as such) in Cuttack, Orissa. Harishpur Garh, located at the river mouth, held historical significance for sea trade, as indicated by the names “Boita-kuilya” and “ship-haven,” (Bowrey, 1993; Wilson, 1895); reflecting the maritime traditions of the Kalinga kingdom that persist in present-day folk culture, in the festival of Boita Bandana.\n\nThe maritime activities of ancient Kalinga drew the attention of artists to be depicted in the form of sculptures at different places. An interesting sculpture preserved at Orissa State Museum depicts the journey of a boat over water carrying people along with elephants. A 12th-century sculpture of a reverse-clinker boat is portrayed in the Bhogamandapa of Jagannath temple; the block of sculpture with the splash of oars, and ripple of waves indicates the desperate speed of the boat (Barnes & Parkin, 2015; Behera, 1999; Bhowmick, 2005; Mookerji, 1912; Nayak, 2009; Parkin, 2016); as if, it is escaping from a danger: purportedly it was brought from the KT. There are two other boat sculptures of the medieval period from ancient Orissa preserved in the museums of Victoria and Albert Museum, London, one of which is labelled as Khelana (toy) instead, the name of the boat is supposed to be Khulana as it is known for Bhowmick (2005). An illustration of a boat pettoo-a “from Balassora or the coast of Palmira” by a Flemish Marine painter Frans Balthazar Salvyns displayed in the National Maritime Museum Greenwich, which is the reminiscent of Patia boat of Orissa (Behera, 1999; McGrail, 2001; McGrail et al., 2003). The construction of the Patia boat is considered as one of the most complex traditional boats in the world (Bhattacharrya, 2006; Blue et al., 1997). Even a few more museums of India and abroad have evidence of different types of ancient boats of Kalinga preserved, which reveal ingenious skills. There were different types and sizes of boats used by the people that has been detailed in a Sanskrit book Yukti Kalpataru (wish-fulfilling tree) which elaborates on the ship building along with measurement and varieties 25 names of ships (Mookerji, 1912; Sahai, 1996; Sastri, 1917; Sinha, 1999). There is a temple of the 8th century named Boitala Deula, in Bhubaneswar, Odisha, the name of the temple suggests it is having the resemblance of a Boita (ship). The temple is also giving evidence of the sustained level of maritime activities.\n\nArcheological survey reports that a port was discovered at Khalkatapatana near the banks of the river Kushabhadra (Bandi, 2000; Manjhi et al., 2000; Sinha, 1999), in close proximity to the KT. Excavation efforts led by K. Veerabhara Rao and his team led to the conclusion that an active port-town existed during the 12th to 15th centuries (Bandi, 2000; Tripati et al., 2015a, 2015b). However, the port-town declined following the downfall of the Eastern Ganga dynasty. This affirms that the sea routes and ships were used for transporting huge stone blocks during the construction of the Sun Temple.\n\nEvidence emerged suggesting that ancient artisans possessed the expertise to move the massive stone blocks. An intriguing example is the weight of the crowning stone of 2000 tonnes, with a thickness of 25 ft. It was one of the largest stones in the world (Childress, 2013). There are other monumental stone blocks arranged in the shape of sculptures at varying elevations, featuring impressive figures such as the huge Gaja singha (lion upon elephant), Nabaghraha slabs of different lintels, a lion trampling on an elephant and musicians positioned at different heights, and other large sculptures. Childress (2013) hypothesized that advanced technology was employed during the construction of the edifice, such as large-scale saws, power grinders, drills, and some unidentified method of levitating massive stones or rendering them weightless. Childress’s assumption that the ancient people made heavy stones weightless using their scientific knowledge. As explained in ‘Baya Chakada’, the heavy lifting was accomplished through the use of scaffolding, skilled workers, various tools, and most importantly, trained elephants. Records of this tradition are evident in a sculpture panel originally housed in the KT, now gracing the walls of the Siddha Mahavir temple in Puri. These sculptures, relocated to the Siddha Mahavir temple during the Maratha period (Behera, 2005; Dey, 2016; Donaldson, 2003), vividly depict the method of hoisting massive stone blocks with the aid of scaffolding. In the scene, two individuals are seen working at the top while four masons carry a rectangular stone block along an inclined path. Three elongated pillars are visible, providing support to the inclined slope, with one end positioned on the temple surface and the other resting on the ground. In conclusion, scaffolding was constructed of wood (Behera, 2005; Dey, 2016). This also refutes the notion of filling sand inside and outside the temple to aid in transporting stone blocks and other materials for smoother construction progress. However, considering the substantial use of iron beams and clamps throughout the temple by ancient builders, it is likely that they employed iron beams, rods, and clamps for the scaffolding used in lifting large stone blocks.\n\nThe use of enormous stone blocks and colossal iron beams in the construction is remarkable to all the scholars who have visited the site, ranging from Mahmud Bin Amir Wali, and Baba Brahmachari to European and Indian scholars of the 19th and 20th centuries, as well as those visiting today. To ensure the stability of the building, heavy stone blocks were strategically placed on top, which was the primary function of the Amala (Swarup, 1910). The term ‘Amala’ is abbreviated from \"Amara-shila,\" indicating that the “sila” (stone) was intended to sustain the structure as ‘Amara’ (immortal). Furthermore, another heavier stone block was selectively placed beneath the Amala, similar to the porch (Figure 5), to make it more stable. This demonstrates that the builders were keenly aware of the importance of weight distribution while designing a large structure.\n\nConsidering iron beam were neither utilized for decorative purposes in the temple nor were employed in the construction of the Vijayastamba (triumph pillar), similar to the pillars in Delhi or Dhar. Every individual iron component utilized in the temple’s construction served the purpose of providing support to various sections and corners, as well as reinforcing the monument. In addition to natural adhesives, iron clamps were frequently employed in construction to strengthen the seams. As per Swarup’s measurements, the porch is a square hall, spanning 60 ft by 60 ft, upheld by four pillars and reinforced with robust iron beams to support the ceiling (Swarup, 1910). Additionally, beams were integrated into the lintels to strengthen the structure and uphold the sizable Nabagraha slabs positioned above each doorway of the porch. Moreover, numerous beams were installed as a false ceiling to offer additional support, alongside the extensive use of iron clamps in the stone joints. During the course of demolition, nearly all of the beams were fragmented, with the longest pillar measured to be 35 ft (Graves, 1912). Fergusson (1910) noted that the beams in the ceiling had variable thickness, gradually widening from the sides towards the centre. It is indicative that artisans had expertise in the technical aspects and had the ability to use knowledge about the strength and properties of the metals being utilized.\n\nThe ancient artisans were able to shape and join large iron beams over eight centuries ago. Engineer M.H. Arnott, who was involved in the rehabilitation crew during the early 20th-century excavation, discovered the manufacturing secret when he examined a broken iron beam. The iron was forged into small pieces of one to one and a half feet in length and three to four inches in width. These pieces were then arranged side by side in a row, similar to bricks in a wall (O’Malley, 1908). The segmented components were inserted into a quadrilateral hollow iron bar and then welded together to create a secure and seamless connection. The upper surface was subsequently polished to eliminate any visible signs of the joints, resulting in a robust and sturdy beam. The material’s composition is akin to contemporary military-grade steel (Edwards, 1969; Friend, 1926; Singh & Kaur, 2014). Moreover, in order to obtain certainty, Newton conducted a test to directly compare the corrosion resistance of the iron of the KT with modern military-grade steel (Friend, 1926). To do this, he obtained samples of both metals that were identical in shape, size, and weight. These samples were then exposed to various atmospheric conditions, including normal water wet and dry tests for one year, followed by an artificial sea water test for an additional year. Following a two-year scientific observation, he found that the resistance capacity of the iron used in the KT surpasses the contemporary military-grade steel. This finding was a learning lesson to him and to the Iron and Steel Industry. Such observation questions the correlation between technological advancement and the quality of products produced by the new generation, encapsulated in the adage “the good die young?” In order to test the limits of current technology, numerous lengthy iron beams have been left in place inside the KT for almost four centuries. The mixture of technical proficiency with local knowledge in ancient times provides a remarkable level of dexterity to the art of constructing buildings, which is impressive to the globe today.\n\nThe KT’s wheel functions as a vertical sundial, meticulously designed to align with the exact latitude of Konark (Bhatnagar & Livingston, 2005; De, 2022; John, et al., 2015; Joshi & Srivastava, 2021; Yadav, 2021). This design allows for the accurate determination of time throughout the day, with minimal error, by casting shadows when a long stick is positioned at a zero-degree angle to the axle of the wheel, parallel to the ground in relation to the angle of the sun. The circular wheel is divided into eight major spokes, each representing a three-hour interval, effectively dividing the twenty-four-hour day. Between each major spoke, there are eight minor spokes, further dividing the space into halves and indicating intervals of one and a half hours, or ninety minutes. Additionally, thirty beads are positioned along the edge of the wheel between a major and a minor spoke. Each bead corresponds to three minutes of time. Moreover, by observing the gap between each bead and its center, finer subdivisions of time are discernible, with intervals as precise as one and a half minutes. The sundial functions in a counterclockwise manner. This intricate design showcases the ancient sculptors’ knowledge of astronomy, making the wheel an instrument for celestial timekeeping.\n\nThe temple was built with precision, enabling the first ray of the rising sun to penetrate through the porch and illuminate the idol of the Sun God in the sanctum of the KT (Das, 2015). A discussion with Soumit Biswal, a scientist from ISRO, the Vikram Sarabhai Space Centre, Thiruvananthapuram, Kerala, suggests that the temple is aligned along the east-west axis with one-degree accuracy. Observations indicate that on March 21st and September 22nd, twice a year, the sun’s rays were expected to directly illuminate the idol. Taking into account the width of the main door of the porch, Suvendu Patnaik of Pathani Samanta Planetarium, Bhubaneswar, suggests that sunlight would fall for approximately thirty days twice a year: from March 6th to April 5th and from September 7th to October 6th (Dey, 2016). He further specifies that with an opening of one degree, the sun illuminates the area for about 11-12 days during each of these periods annually. This also validates the name of the monument, KT, derived from two Sanskrit words Kona meaning angle and Arka meaning Sun. It signifies the precise Kona (angle) through which the rays of Arka (sun) enter to illuminate the idol of the Sun God in the temple.\n\n\nDiscussions\n\nThe study sheds light on varied aspects of the KT construction. It uncovers the purpose behind the temple’s construction, emphasizing its cultural and religious significance within the kingdom’s context. Additionally, the relative security of the kingdom during construction implies a stable political environment, revealing the king’s achievements in different wars and contributing to monumental projects. The identification of diverse artisans involved underscores the collaborative effort and diverse skills required for such endeavors. Trained elephants played a vital role in providing support during the construction process. Additionally, they were employed as a formidable troop in the wars waged by King Narasimhadeva I, earning him the title of Gajapati, which he proudly bore as the first king of Kalinga.\n\nThe temple sculptures act as repositories of encrypted facts and historical narratives, enriching the understanding of that era. The accuracy of the temple’s measurements, as recorded in the Sanskrit version of Madalapanji dating back to 1627 A.D., has been confirmed through translation. The meticulous selection of high-quality stones demonstrates dedication to craftsmanship. Innovative transportation methods like sea routes and ships, alongside the use of iron scaffolding, highlight advanced engineering techniques. Notably, lifting the world’s heaviest stone to a height of around 200 ft showcases remarkable engineering prowess. The knowledge of utilizing iron ores for rust-resistant iron exemplifies the ingenuity of ancient craftsmen. Again, the intricately crafted wheel of the KT serves as a vertical sundial, perfectly aligned with the precise latitude of Konark. Its construction allows the first ray of the rising sun to illuminate the idol of the Sun God within the sanctum, showcasing ancient sculptors’ mastery of astronomy and architectural precision.\n\nThe advanced indigenous technology utilized in the construction of the ancient KT showcases remarkable sophistication, demonstrating a fusion of artisans’ skills, commitment, folk knowledge, and the quality of construction materials. Delving into the intricacies of this indigenous technology illuminates historical architectural achievements and revitalizes the forgotten art form of Kalinga, offering insights into ancient construction techniques.\n\nThe limited journal publications on KT restricts the access to authentic evidence. Nevertheless, the archival data are employed from indigenous knowledge dating back to the 13th century, and folk stories and prevailing cultural practices. Documentation includes incidents and eyewitness accounts found in books, diaries, palm leaf inscriptions, and stone inscriptions. Furthermore, the study involved analyzing sculptures from the KT, along with those housed in museums of different locations, and exploring the ancient maritime traditions of the kingdom’s predecessors. Here the text also includes literature authored by King Narasimhadeva I’s court poet and historical records of the kingdom.\n\nThe evidence surrounding the KT can be researched through multiple lenses. First, there are many festivals, cultural practices, and narratives that exist among the localities surrounding the KT as a reminiscence of the past. Those have passed on from one generation to another through observation of festivals, cultural practices, and words-of-mouth. Such knowledge of Dharma is derived here from secondary sources. Acquiring such folk evidence on festivals, cultural practices, and stories from primary sources through qualitative research and analyzing their contents and themes can provide insight into the history, cultural-orientation, craftmanship, and the science and technology surrounding the KT and its present state of affairs.\n\nTo further understand the construction of KT and shed light on the transportation of its massive stones, a geotechnical investigation can be carried out to analyze soil properties and borehole assessments to unveil ground conditions that may have influenced construction techniques and structural stability. Additionally, employing geophysical methods such as ground-penetrating radar (GPR) and electrical resistivity imaging (ERI) can help detect subsurface features like buried water channels or geological anomalies, while radar satellite data analysis can offer a broader perspective of the landscape, revealing hidden watercourses or geological formations relevant to the temple’s construction. Plus, Geographic Information System (GIS) technology facilitates the integration and analysis of various spatial data layers, including topography, hydrology, and historical maps, aiding in the identification of potential water bodies, ancient river courses, or coastal features that could have influenced the temple’s location and construction. Supplementing these methods with map analysis, including examination of historical maps, archaeological surveys, and aerial photographs, can provide valuable context for understanding the KT’s surroundings and changes in the landscape over time. By synthesizing these diverse investigative approaches, the research can provide a comprehensive understanding of the environmental context in which KT was built, unraveling its construction logistics, resource utilization, and the cultural and environmental factors shaping its design and placement in the landscape.\n\n\nConclusion\n\nThis study has emphasized the significant contributions of an array of artisans involved in the construction process of the ancient monument of the KT. By exploring the true incidents preserved in the form of folk stories, cultural practices, and sculptures, the understanding of historical narratives has been enriched. Additionally, delving into the techniques and principles behind the ancient structures provides insights to architects and engineers to innovate and adapt improvised methods for sustainable and resilient monument design. Through this process, historical, cultural, and scientific evidence has been debunked, bridging the gap between past and present technologies to rationalize architectural excellence in the future.\n\nThis research reveals the enduring legacy of Narasimhadeva I. The ancient measurements of the temple, translated from the Sanskrit version of the Madalapanji, align with modern-day calculations, serving as a testament to the completion of this architectural marvel. Furthermore, our investigation unveils the remarkable utilization of sea routes for transporting massive stones to the construction site, shedding light on the logistical prowess of ancient engineers. Amidst the stone engravings, a poignant tale emerges, immortalized in sculptures, revealing the valor of Sudehi, the faithful elephant who played a pivotal role in reviving Narasimhadeva I from the battlefield. The artisans’ skills, knowledge, local wisdom during the 13th century played significant roles in the construction of the temple. The temple’s foundation has undergone modern engineering scrutiny and has been found suitable for an architectural blueprint in a seaside setting. It further unveils the feat of lifting the world’s heaviest stone to towering heights, a testament to the ingenuity of ancient construction techniques. As we delve deeper, we discover echoes of modernity within the ancient framework, with the utilization of iron scaffolding mirroring contemporary construction practices. Lastly, the inquiry unveils the enduring presence of rust-resistant iron within the temple compound. Alongside the temple’s wheel serving as India’s earliest sundial, offering a glimpse into the celestial intricacies revered by ancient astronomers. The first ray falling on the idol inside the temple is another astronomical keenness of ancient artisans. 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Res. 2021; 12(1): 40690–40692." }	[ { "id": "356838", "date": "09 Jan 2025", "name": "Ngakan Ketut Acwin Dwijendra", "expertise": [ "Reviewer Expertise Architecture", "engineering", "sustainable development", "and cultural heritage studies" ], "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 Konark Temple’s Construction: A Critical Review of the Historical, Cultural, and Scientific Evidence\nSummary of the Article The article critically reviews the historical, cultural, and scientific evidence surrounding the construction of the Konark Temple in Odisha, India. The research incorporates data from ancient texts, folklore, palm leaf manuscripts, archaeological evidence, and historical records. It explores themes such as the temple's purpose, the techniques and materials used, the role of indigenous knowledge, and its architectural and astronomical precision. Key findings include:\nThe temple served as a monument of devotion to the Sun God and a symbol of King Narasimhadeva I’s military victories. Advanced construction techniques included using rust-resistant iron, innovative stonelifting technologies, and precise architectural designs. The temple's alignment with astronomical phenomena demonstrates ancient artisans' mastery of astronomy. Indigenous technologies, including the use of high-grade iron and efficient transportation of massive stone blocks via sea routes, highlight the engineering ingenuity of the time.\nThe article concludes that the Konark Temple embodies a blend of artistic mastery, cultural significance, and technological innovation, offering insights for contemporary architecture and engineering.\nEvaluation and Recommendations\n1. Comprehensiveness of the Topic in Current Literature Evaluation: Partly. The article extensively reviews the historical and cultural significance of the Konark Temple, presenting a rich narrative of its construction and legacy. However, its engagement with contemporary research methodologies in archaeology, materials science, and geotechnical studies is limited. While the authors mention some modern studies, such as those by CSIR and CBRI, these are not deeply integrated into the analysis. For example, insights from recent archaeological technologies like ground-penetrating radar or 3D modeling could enhance the study’s relevance. Recommendations: Incorporate more recent geotechnical or archaeological studies to substantiate claims about the temple’s construction techniques and materials. Review and discuss contemporary literature on similar historical structures globally for comparative insights.\n2. Factual Accuracy and Citation Support Evaluation: Partly. While the article is well-cited and draws on historical sources, its reliance on folklore and secondary sources raises questions about the verifiability of some claims, such as the legend of Dharmapada or the use of magnets to suspend objects. Some claims, like the rust resistance of the iron used, require more robust validation through referenced scientific studies. Recommendations: Ensure that all key assertions, especially those based on folklore or oral traditions, are marked as such and distinguished from evidence-based claims. Include peer-reviewed scientific studies to corroborate statements about materials, such as the selection processes for rust-resistant iron and stone.\n3. Accessibility of Language Evaluation: Yes. The language is clear, engaging, and accessible to academic and general audiences. The integration of technical and cultural narratives is particularly commendable. Recommendations: Maintain this clarity while adding more technical terms and methodologies if engaging with geotechnical or engineering audiences.\n4. Appropriateness of Conclusions Evaluation: Partly. The conclusions are generally appropriate, synthesizing the findings effectively. However, they are somewhat limited by the scope of the evidence presented. For example, the findings could better integrate critiques or limitations of ancient techniques compared to modern methodologies. Recommendations: Discuss how the findings contribute to or challenge current historical architecture and engineering knowledge. Acknowledge gaps in evidence and propose specific areas for future research.\nSpecific Points to Address for Scientific Soundness\nIntegration with Current Research Include recent studies or methods, such as geotechnical assessments or 3D modeling, to validate claims about construction techniques. Provide a comparative analysis with other ancient monumental structures to position the findings in a broader context. Strengthening Evidence: Use scientific data to support claims about material properties, such as the corrosion resistance of iron used in the temple. Provide citations from peer-reviewed journals, especially for claims about astronomical alignment and structural engineering. Distinguishing Between Fact and Folklore: differentiate between historical records, archaeological evidence, and folklore. For example, legends like the story of Dharmapada should be framed as cultural narratives unless supported by evidence. Clarification of Methodology Elaborate on the systematic review process, specifying how documents were selected and analyzed. Discuss potential biases in the sources. Addressing Limitations: Acknowledge the study's limitations, such as the reliance on secondary sources and the lack of direct archaeological validation for some claims.\nConclusion Overall, the article offers valuable insights into the construction of the Konark Temple and highlights its historical, cultural, and scientific significance. The authors should address the abovementioned limitations to make the article scientifically sound, particularly by integrating more recent research, strengthening evidence-based claims, and distinguishing folklore from verifiable facts. By doing so, the study can serve as a robust reference for both historical and scientific investigations into ancient construction techniques.\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": [] }, { "id": "356835", "date": "05 Feb 2025", "name": "Sabeeh Lafta Farhan", "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\nThoroughness & Scope – The article provides an extensive review of historical, cultural, and scientific aspects of the Konark Temple’s construction. The depth of research and integration of multiple sources, including historical texts, archaeological findings, and indigenous knowledge, is commendable. Interdisciplinary Approach – The combination of historical analysis, engineering insights, and cultural narratives enriches the discussion. However, a more structured comparison with contemporary temple architecture across different regions of India could provide additional context. Scientific and Engineering Aspects – The discussion on iron usage and rust-resistant techniques is fascinating. It would be helpful to include a comparison with other ancient iron structures, such as the Iron Pillar of Delhi, to establish technological linkages.\nSpecific Comments: Historical Narratives & Myths – While the study effectively explores historical accounts and folklore, some claims (such as the floating deity using magnets) require additional corroboration from scientific sources or metallurgical studies. Methodology Clarification – The systematic review method is well described, but the inclusion criteria for selecting historical texts, palm leaf manuscripts, and indigenous narratives should be elaborated. How were these sources validated for accuracy? Illustrations & Figures – The inclusion of figures is helpful in visualizing the temple’s architectural layout. However, some of them lack detailed captions or references to their sources (e.g., Figure 5’s sketch of the temple in its complete state). Construction Techniques & Materials – The identification of Khondalite, Laterite, and Chlorite stones is insightful. However, the decay rate comparison with Western structures like St. Paul’s Cathedral might need more quantitative backing with geological studies. Maritime & Transportation Hypothesis – The theory of stone transportation via sea routes is intriguing. Have geological or sedimentary studies been conducted along these proposed routes to confirm ancient maritime trade or transport links? References: Lafta Farhan S.et.al. 2022 (Ref 1) This study discusses the transformation of historical city centers and could provide a comparative perspective on the cultural and architectural sustainability of heritage sites, similar to the preservation efforts and architectural significance discussed in the Konark Temple study.\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/13-1540
https://f1000research.com/articles/13-1537/v1	19 Dec 24	{ "type": "Study Protocol", "title": "Protocol for conducting a scoping review on the influence of low-stakes assessment on student learning in medical education", "authors": [ "Imran Zafar", "Lamburt Schuwirth", "Susan Waller", "Carvalho Filho", "Mohi Eldin Magzoub", "Imran Zafar", "Lamburt Schuwirth", "Susan Waller", "Carvalho Filho" ], "abstract": "Rationale and Background Low-stakes assessments enhance student learning outcomes by providing a comprehensive view of performance and promoting competency-based education. Multiple low-stakes assessments encourage continuous learning and incorporate formative feedback for a realistic and long-term environment. Diverse assessment methods ensure psychometric rigor, utility and a programmatic assessment approach with multiple data points affording high-stakes decisions on progression.\n\nObjective This scoping review aims to systematically identify and map existing literature on the use of low-stakes assessments in medical education and determine their impact on student learning outcomes in terms of, motivation and engagement achievements/competencies.\n\nMethods and Analysis The review will use the Joanna Briggs Institute’s framework for scoping review studies, searching eight databases and grey literature. A presearch will be done in PubMed, Scopus, and Google Scholar using terms related to low-stakes assessment, formative assessment, continuous assessment, and programmatic assessment within the context of medical education. The Covidence Systematic Review tool will aid in screening and conflict resolution. The reference lists of included studies will be checked manually for other relevant literature. Two research team members will independently screen and extract data, resolving discrepancies with a third team member. Inclusion and exclusion criteria will be refined iteratively based on key research themes. The review will follow PRISMA-P guidelines, focusing on the impact of low-stakes assessment on student learning in medical education.\n\nEthics and dissemination No ethical approval is required as all data will be collected from published and grey literature. Findings will be disseminated at relevant conferences and submitted for publication in peer-reviewed journals.", "keywords": [ "Scoping Review", "Low Stakes Assessments" ], "content": "Introduction\n\nLow-stakes assessments (LSAs) in medical education are evaluation methods with minimal impact on a student’s final grade or certification outcomes. These assessments are primarily used for formative purposes, providing opportunities for students to practice, receive feedback, and learn from mistakes without significant consequences (Shrivastava & Shrivastava, 2022b). They are designed to monitor progress, identify learning gaps, and help both learners and educators focus on areas needing improvement (Pastor et al., 2019; Schüttpelz-Brauns et al., 2018). Despite their low stakes, these assessments can influence student behavior and effort, particularly when performance is discussed with mentors or when there are consequences for non-participation (Govaerts, 2014). To enhance effectiveness, it’s important that students perceive these assessments as valuable and relevant to their future practice, which can increase engagement and effort (Schüttpelz-Brauns et al., 2018) (Price et al., 2024). However, variability in students’ test-taking effort can affect validity, necessitating strategies to identify and exclude low-effort responses for more accurate results (Wang et al., 2020; Schut et al., 2018). Low-stakes assessments contribute to building a longitudinal representation of student performance, are useful for continuous feedback, improving student feedback literacy, self-evaluation and learning, LSAs help reduce the stress and anxiety associated with high-stakes evaluations, creating a supportive learning environment (Shrivastava & Shrivastava, 2022b). Overall, they are essential for continuous learning and improvement in medical education without the high pressure of significant academic consequences.\n\nLSAs provide ongoing feedback and help identify students’ strengths and challenges, thereby guiding future learning and teaching strategies (Abu-Zaid, 2013). Examples of LSAs are simulated clinical scenarios, objective structured clinical examinations (OSCEs), and questionnaires or written assessments, which are often used to evaluate non-technical skills (Gordon et al., 2019). Additionally, low-stakes progress tests, such as those measured by the Test-taking Effort Short Scale (TESS), help gauge students’ effort and intrinsic motivation during assessments (Schüttpelz-Brauns et al., 2018). Team-based learning (TBL) assessments, including Individual Readiness Assurance Tests (IRAT) and Group Readiness Assurance Tests (GRAT), also fall under low-stakes assessments, providing a structure for continuous feedback and performance tracking (Vegi et al., 2022). Furthermore, workplace-based assessments like Direct Observation of Procedural Skills (DOPS), Mini-Clinical Evaluation Exercises (mini-CEX), and Case-based Discussions (CbD) are integral to assessing clinical competence in real-world settings, promoting active learning and immediate feedback (Liu, 2012, Ref, 2024). The use of multiple-choice questions (MCQs) from the first year of medical education is another form of low-stakes assessment that helps students prepare for professional exams and promotes deep learning (Wang et al., 2020). Peer assessments, such as the mini-peer assessment tool (mini-PAT), allow students to evaluate each other, fostering a collaborative learning environment and enhancing performance through peer feedback (Shrivastava & Shrivastava, 2022a). Finally, Entrustable Professional Activities assess by the level of entrustment can be considered as types of low-stakes assessment. (Kinnear et al., 2021). These diverse methods collectively contribute to a comprehensive and supportive assessment framework in medical education, balancing the need for evaluation with the goal of fostering a deep, sustained learning experience.\n\nLSAs offer multiple benefits for medical students, enhancing their learning environment and academic experience. These individual assessments, which do not carry significant consequences, encourage students to engage more deeply with the material without the pressure of high-stakes exams. They guide the learning process and monitor educational programs, despite potential high variation in test-taking effort (Price et al., 2024). Strategies like reviewing low test performance with mentors exploring the student’s perception of performance and offering constructive feedback and implementing consequences for non-participation can increase serious test-taking behavior, thereby improving the validity of these assessments (Schüttpelz-Brauns et al., 2020).\n\nLSAs foster self-regulated learning (SRL) and co-regulated learning (CRL), crucial for developing clinical reasoning, doctor-patient communication, and self-reflection skills (Pastor et al., 2019). Detailed asynchronous feedback on low-stakes quizzes significantly improves exam performance, allowing students to review and synthesize content deeply outside the classroom (Shrivastava & Shrivastava, 2022a). These assessments help identify patterns in student performance, predictive of future academic success, and inform targeted remediation interventions (Schüttpelz-Brauns et al., 2018). Tools like the Test-taking Effort Short Scale (TESS) measure test-taking effort in LSAs, ensuring results reflect students’ true abilities by identifying and allowing exclusion of students results who are not making effort (Price et al., 2024). Peer assessments, such as the mini-peer assessment tool (PAT), enhance learning outcomes by providing diverse feedback and fostering a collaborative learning environment (Wang et al., 2020).\n\nLSAs are less stressful, reducing the anxiety and pressure that can negatively impact learning outcomes. They increase student participation and engagement, as students are more likely to take part in assessments that do not significantly affect their grades or future. These assessments provide more opportunities for feedback and improvement, crucial in medical education for developing necessary skills and competencies. (Shrivastava & Shrivastava, 2022b; Ontong, 2021).\n\nThe formative nature of LSAs provides regular feedback and opportunities to adjust learning strategies, helping students develop a growth mindset and focus on continuous improvement (Ontong, 2021). Using various assessment methods, including LSAs, offers a comprehensive evaluation of students, reduces cheating likelihood, and includes engaging and effective assessments like quizzes, discussions, and case studies. Since they do not significantly impact final grades, low-stakes assessments reduce pressure, allowing students to focus on learning and improvement rather than just achieving high grades. They also provide flexibility and reversibility, enabling teachers to adapt their assessment strategies as needed (Shrivastava & Shrivastava, 2022b; Ontong, 2021).\n\nOverall, LSAs play a complementary role in supporting continuous learning and skill development in medical education (Bains et al., 2023; Ganesan et al., 2023). They promote a culture of continuous learning, reduce stress and anxiety, and provide more opportunities for feedback and improvement.\n\nA theoretical framework is essential for understanding the influence of LSAs on student learning in medical education.\n\nSelf-Determination Theory (SDT) is a suitable theoretical framework for investigating the influence of LSAs on student learning in medical education. SDT emphasizes autonomy, competence, and relatedness as key factors in motivating individuals, aligning with the importance of self-regulated learning (SRL) in medical training (Neufeld, 2023, Ganesan et al., 2023).\n\nTo apply Self-Determination Theory (SDT) to low-stakes assessments in medical education, it is important to consider how these assessments can support students’ basic psychological needs of autonomy, competence, and relatedness; (Neufeld, 2023). Providing students with choices in the format and timing of LSAs can promote a sense of autonomy (Dutt et al., 2023). Involving students in the development of assessment criteria and rubrics fosters a sense of ownership (Ganotice et al., 2023), and encouraging students to set their own learning goals and use assessments to monitor their progress enhances their autonomy (Neufeld, 2023). Designing LSAs that are challenging yet achievable supports students’ feelings of competence (Dutt et al., 2023). Providing timely and constructive feedback that focuses on progress and mastery rather than comparison to others, along with offering opportunities for students to demonstrate their knowledge and skills in various contexts, further reinforces competence (Ganotice et al., 2023; Dutt et al., 2023; Neufeld, 2023). Creating a supportive and collaborative learning environment where students feel safe to take risks and learn from mistakes during low-stakes assessments addresses the need for relatedness (Ganotice et al., 2023). Encouraging peer feedback and group discussions around assessment results fosters a sense of community and belonging (Neufeld, 2023), while ensuring that assessments are aligned with the overall learning objectives and perceived as relevant and meaningful by students (Dutt et al., 2023). Incorporating these SDT principles into the design and implementation of LSAs, medical educators can create a learning environment that supports students’ intrinsic motivation, engagement, and well-being (Ganotice et al., 2023; Hirsh et al., 2024).\n\n\nMethods\n\nSelf-Determination Theory (SDT) can guide our research questions related to how various factors in LSAs influence students’ basic psychological needs (autonomy, competence, and relatedness) and their subsequent motivation, engagement, and learning outcomes in educational settings (Dutt et al., 2023; Netcoh, 2017). Below are some potential research questions that can be explored using SDT as a theoretical framework:\n\n1. What are the implementation strategies of low-stakes assessment, in relation to the methods used, application and challenges?\n\n2. How do different forms of low-stake assessment impact students’ learning in terms of motivation, engagement, and autonomy?\n\n3. How do low-stakes assessment impact student achievement and competence?\n\n4. How do students perceive and experience low-stakes assessments with reference to their influence on autonomy, competence, relatedness, and well-being compared to high- stakes summative assessment methods? (Netcoh, 2017).\n\nTo conduct this scoping review on how LSAs influence student learning in medical education, the following are the planned inclusion and exclusion criteria:\n\n\n\n1. Study Type: The review will include studies that are published in the form of articles, theses, or conference proceedings, both peer reviewed and published, unpublished studies and grey literature.\n\n2. Topic: The primary focus of the study should be on the impact of LSAs on student learning in medical education.\n\n3. Study Population: The studies should involve medical students, interns or residents (junior doctors in training) as the primary participants.\n\n4. Assessment Type: The assessment should be classified as low-stakes, meaning it does not summatively (terminally) affect the student’s grade or academic standing, having less significant progress consequences.\n\n5. Publication Date: The studies should be published after January 2000.\n\n6. Language: The studies should be published in English to facilitate efficient data extraction and analysis.\n\n7. Methodology: The studies can employ various research methods, including quantitative, qualitative, or mixed methods approach. Grey literature may include …\n\n\n\n1. High-Stakes Assessments: Studies that focus on high-stakes assessments, which significantly impact student grades or academic standing, will be excluded.\n\n2. Non-Medical Education: Studies conducted in non-medical education settings or involving non-medical students will be excluded.\n\n3. Non-English Language: Studies published in languages other than English will be excluded to ensure efficient data extraction and analysis.\n\n4. Non-Primary Focus on Student Learning: Studies that do not explicitly investigate the impact of low-stakes assessments on student learning will be excluded.\n\n5. Non-Medical Education Settings: Studies conducted in non-medical education settings, such as nursing or allied health, will be excluded.\n\n6. Studies published or grey literature dated before 2000.\n\nThe context of this scoping review would be the influence of low-stakes assessments on student learning in medical education, focusing on the broader aspects of assessment methods and their impact on student learning outcomes.\n\nThe review aims to investigate how LSAs, which do not significantly impact student grades or academic standing, affect the learning process and outcomes of medical students. It will examine various types of LSAs, including online assessments, e-assessments, computer-assisted assessments, and portfolio-based assessments etc.\n\nThe review will focus on undergraduate medical education, as well as postgraduate medical education, such as residency programs. It will provide a comprehensive overview of the current literature on the challenges and opportunities associated with LSAs in medical education, including the role of technology and the impact on student learning outcomes\n\nA wide range of study types are recommended to conduct this scoping review. Quantitative research like randomized controlled trials, quasi-experimental studies, cohort studies, cross-sectional studies, and pre-post studies should be covered. In order to gain insights into experiences and perspectives incorporating qualitative methods such as interviews, focus groups, and case studies are also important. A more comprehensive understanding can be obtained by mixed-methods studies that combine both quantitative and qualitative data. In addition, systematic reviews and meta-analyses will help summarize the existing research and highlight overall trends and gaps, while surveys can capture broader trends and opinions. Including such a wide range of study types makes sure that the topic is understood in all of its complexity.\n\nThe scoping review proposal will follow the Joanna Briggs Institute (JBI) methodology for scoping reviews and will be documented following the guidelines of the Preferred Reporting Items for Systematic Reviews and Meta-analyses extension for Scoping Reviews. It is planned to commence the review in September 2024, with an anticipated completion date in December 2024.\n\nThe following key databases will be searched: PubMed, MEDLINE, ERIC, PsycINFO, Scopus, and Web of Science.\n\nA comprehensive and iterative search strategy will be developed with assistance from a medical librarian. The following databases will be systematically searched:\n\n• PubMed: To cover biomedical literature related to medical education.\n\n• MEDLINE: For peer-reviewed studies on medical assessments and learning outcomes.\n\n• ERIC (Education Resources Information Center): To capture studies focused on educational practices and assessments in higher education.\n\n• PsycINFO: For literature on psychological aspects of learning, motivation, and assessment.\n\n• Scopus: To retrieve multidisciplinary studies, ensuring broad coverage.\n\n• Web of Science: For high-impact articles on medical education and low-stakes assessments.\n\n• Google Scholar: To capture grey literature and additional unpublished studies.\n\nAdditional grey literature will be searched through databases like ProQuest Dissertations & Theses Global and OpenGrey to include relevant theses, dissertations, and other non-peer-reviewed materials.\n\nKeywords and Medical Subject Headings (MeSH) terms relevant to the topic will be employed, such as “low-stakes assessment,” “formative assessment,” “medical education,” “student learning,” and “programmatic assessment.” Boolean operators (AND, OR, NOT) will be used to refine the search results. The reference lists of included studies will be manually checked for additional relevant literature.\n\nScreening Process and Iterative Refining of Criteria\n\nAn iterative process will be followed during the screening of titles and abstracts to ensure all relevant studies are captured. Two reviewers will independently screen the search results at two levels: title/abstract screening followed by full-text screening. The inclusion and exclusion criteria will be applied initially to a subset of studies to pilot test their adequacy.\n\nDuring this process, if new themes or patterns emerge that require adjustment, the inclusion and exclusion criteria may be refined iteratively. For instance, if a certain type of low-stakes assessment appears to be underrepresented or newly relevant literature emerges during screening, these criteria will be adjusted accordingly to capture the breadth of available literature.\n\nAny discrepancies between the two reviewers will be discussed, and if necessary, a third reviewer will be consulted to resolve conflicts. The team will maintain a record of all decisions made during the screening process to ensure transparency and reproducibility of the review.\n\nData from included studies will be extracted using a standardized data extraction form. This form will capture key study characteristics, including the study design, participant details, type of low-stakes assessment used, and outcomes related to student learning (e.g., motivation, engagement, competence).\n\nThe extracted data will be charted and summarized using tables and diagrams to provide an overview of the existing literature. Data will be presented in:\n\n• Tables: Summarizing study characteristics (e.g., author, year, sample size, type of LSA).\n\n• Diagrams or Flowcharts: Mapping the frequency and distribution of different types of low-stakes assessments across various studies and highlighting common themes.\n\nA thematic analysis will be conducted to identify recurring themes and patterns across the studies. This analysis will be supported by a narrative synthesis, providing a descriptive summary of the findings related to the impact of low-stakes assessments on student learning in medical education. The results will be aligned with the objectives of the scoping review, and key findings will be visualized in diagrams or flowcharts where appropriate.\n\nThe analysis of the data will involve a descriptive synthesis to summarize and present the findings from the included studies. The following steps will be undertaken:\n\nCategorization of LSAs: Types of LSAs identified in the studies will be categorized and described. This will provide an overview of the various assessment methods used in medical education.\n\nThematic Analysis: A thematic analysis (Braun & Clarke, 2006) will be conducted to identify common themes and patterns related to the benefits, drawbacks, and impacts of low-stakes assessments on student learning. This will involve coding the extracted data and grouping similar concepts into themes.\n\nComparative Analysis: A comparative analysis will be performed to examine differences and similarities in the findings across studies. This will help to identify trends and variations in the use and impact of LSAs.\n\nIdentification of Best Practices: The review will highlight effective strategies and best practices for implementing low-stakes assessments in medical education. These will be derived from the successful approaches reported in the included studies.\n\nGap Analysis: A gap analysis will be conducted to identify areas where further research is needed. This will involve highlighting gaps in the existing literature and suggesting potential directions for future studies.\n\nThe results of the analysis will be presented in a narrative format, supported by tables and figures where appropriate. This will provide a comprehensive overview of the current state of knowledge on the influence of low-stakes assessments on student learning in medical education, as well as insights into best practices and future research directions.\n\nEthical approval and consent were not required.", "appendix": "Data availability statement\n\nNo data are associated with this article.\n\nReporting guidelines\n\nFigshare: Protocol for conducting a scoping review on the influence of low-stakes assessment on student learning in medical education, https://doi.org/10.6084/m9.figshare.27619560 (Zafar et al., 2024).\n\nThis project contains the following underlying data:\n\n• PRISMA-P-checklist.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\nAbu-Zaid A: Formative assessments in medical education: a medical graduate’s perspective. Perspectives on Medical Education. 2013; 2(5–6): 358–359. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBains M, Goei K, Kaliski D: Implementation and effects of low stakes quiz feedback on exam performance in a health professional program. Physiology. 2023; 38(S1). Publisher Full Text\n\nBraun V, Clarke V: Using thematic analysis in psychology. Qual. Res. Psychol. 2006; 3(2): 77–101. Publisher Full Text\n\nDutt DDCS, Razavi H, Carr SE: Self-determination theory in ophthalmology education: factors influencing autonomy, competence and relatedness in medical students. Med. Educ. Online. 2023; 28(1). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGanesan I, Cham B, Teunissen PW, et al.: Stakes of Assessments in Residency: Influence on Previous and Current Self-Regulated Learning and Co-Regulated Learning in Early Career Specialists. Perspectives on Medical Education. 2023; 12(1): 237–246. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGanotice FA, Chan KMK, Chan SL, et al.: Applying motivational framework in medical education: a self-determination theory perspectives. Med. Educ. Online. 2023; 28(1). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGordon M, Farnan J, Grafton-Clarke C, et al.: Non-technical skills assessments in undergraduate medical education: A focused BEME systematic review: BEME Guide No. 54. Med. Teach. 2019; 41(7): 732–745. PubMed Abstract \| Publisher Full Text\n\nGovaerts MJB: Medical Education, Assessment in. The Wiley Blackwell Encyclopedia of Health, Illness, Behavior, and Society. Wiley; 2014; pp. 1324–1329. Publisher Full Text\n\nHirsh DA, Crampton PES, Osman NY: Applying self-determination theory to stem medical schools’ clinical teacher sustainability crisis. Med. Educ. 2024; 58(1): 118–128. PubMed Abstract \| Publisher Full Text\n\nKinnear B, Warm EJ, Caretta-Weyer H, et al.: Entrustment Unpacked: Aligning Purposes, Stakes, and Processes to Enhance Learner Assessment. Acad. Med. 2021; 96(7): S56–S63. PubMed Abstract \| Publisher Full Text\n\nLiu C: An introduction to workplace-based assessments. Gastroenterology and Hepatology from Bed to Bench. 2012; 5(1): 24–28. PubMed Abstract\n\nNetcoh S: Students’ Experiences with Personalized Learning: An Examination Using Self-Determination Theory.2017. Reference Source\n\nNeufeld A: Moving the Field Forward: Using Self-Determination Theory to Transform the Learning Environment in Medical Education. Teach. Learn. Med. 2023; 1–6. PubMed Abstract \| Publisher Full Text\n\nOntong JM: Low-stakes assessments: An effective tool to improve marks in higher-stakes summative assessments? Evidence from commerce students at a South African university. South African Journal of Higher Education. 2021; 35(5). Publisher Full Text\n\nPastor DA, Ong TQ, Strickman SN: Patterns of Solution Behavior across Items in Low-Stakes Assessments. Educ. Assess. 2019; 24(3): 189–212. Publisher Full Text\n\nPrice DW, Wang T, O’Neill TR, et al.: Differences in Physician Performance and Self-rated Confidence on High- and Low-Stakes Knowledge Assessments in Board Certification. J. Contin. Educ. Health Prof. 2024; 44(1): 2–10. PubMed Abstract \| Publisher Full Text\n\nSchut S, Driessen E, van Tartwijk J , et al.: Stakes in the eye of the beholder: an international study of learners’ perceptions within programmatic assessment. Med. Educ. 2018; 52(6): 654–663. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSchüttpelz-Brauns K, Hecht M, Hardt K, et al.: Institutional strategies related to test-taking behavior in low stakes assessment. Adv. Health Sci. Educ. 2020; 25(2): 321–335. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSchüttpelz-Brauns K, Kadmon M, Kiessling C, et al.: Identifying low test-taking effort during low-stakes tests with the new Test-taking Effort Short Scale (TESS) – development and psychometrics. BMC Med. Educ. 2018; 18(1): 101. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nShrivastava S, Shrivastava P: Critiquing the scope and utility of mini-peer assessment tool in the training of medical students. APIK Journal of Internal Medicine. 2022a; 10(4): 282–283. Publisher Full Text\n\nShrivastava S, Shrivastava P: Understanding the significance of high stakes and low stakes assessments in medical undergraduate training. Medical Journal of Babylon. 2022b; 19(2): 301–303. Publisher Full Text\n\nVegi VK, Sudhakar P, Bhimarasetty D, et al.: Multiple-choice questions in assessment: Perceptions of medical students from low-resource setting. J. Educ. Health Promot. 2022; 11(1): 103. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWang XR, Hillier T, Oswald A, et al.: Patterns of performance in students with frequent low stakes team based learning assessments: Do students change behavior? Med. Teach. 2020; 42(1): 111–113. PubMed Abstract \| Publisher Full Text\n\nZafar I, Schuwirth L, Waller SA, et al.: Protocol for conducting a scoping review on the influence of low-stakes assessment on student learning in medical education. figshare. Journal contribution. 2024. Publisher Full Text" }	[ { "id": "354585", "date": "25 Jan 2025", "name": "Aubrie Swan Sein", "expertise": [ "Reviewer Expertise Assessment for learning", "academic counseling" ], "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 outlines a protocol for conducting a scoping review to explore the influence of low-stakes assessments (LSAs) on student learning in medical education, using the Joanna Briggs Institute framework for scoping reviews and the PRISMA-P guidelines. The review will focus on identifying the impact of LSAs on learning outcomes, motivation, engagement, and competency development. It incorporates a comprehensive methodology, including an extensive database search, grey literature exploration, and thematic analysis. The results are anticipated to provide insights into the effectiveness of LSAs and inform best practices for their implementation in medical education.\n\nArticle strengths: The authors provide a robust background of LSAs and how they can help foster self-regulated learning. The use of Self-Determination Theory as a conceptual framework makes sense to expand our understanding of how LSAs impact student learning as it relates to the constructs of autonomy, competence and relatedness. The proposed methodology is thorough, including its proposal to incorporate both peer-reviewed and grey literature. Acknowledging the role of grey literature ensures a broader understanding of the topic, capturing less formalized but still valuable research findings. The use of established frameworks like PRISMA-P and the Joanna Briggs Institute methodology adds rigor to the review. The article and proposal is well-organized, with clearly defined objectives, inclusion/exclusion criteria, and a detailed plan for data analysis. The focus on LSAs addresses a significant area in medical education, particularly with the increasing emphasis on formative assessment and competency-based education.\n\nArticle areas for opportunity: A few things areas out to me as potential opportunities for improvement. Limiting the review to medical education excludes valuable insights from other health professions education fields (e.g., nursing, pharmacy). Broadening the scope could enhance the generalizability of the findings. A brief justification for this decision to exclude other health professions domains would improve clarity. Also, the inclusion criteria mention grey literature but lack detail on what types will be included (e.g., reports, conference proceedings). Providing examples or a more precise definition would strengthen the protocol. Regarding the MeSH terms, while the authors mention terms like \"low-stakes assessment\" and \"formative assessment,\" adding related the term \"assessment for learning\" and maybe “assessment as learning” could increase the breadth of relevant studies.\n\nOverall, this scoping review protocol has the potential to make a significant contribution to the literature on LSAs in medical education, particularly in guiding their effective implementation to support student learning and engagement.\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/13-1537
https://f1000research.com/articles/13-1536/v1	19 Dec 24	{ "type": "Research Article", "title": "Advances in Leukemia detection and classification: A Systematic review of AI and image processing techniques", "authors": [ "Aya Achir", "Ikram Debbarh", "Nadia Zoubir", "Ilham Battas", "Hicham Medromi", "Fouad Moutaouakkil", "Ikram Debbarh", "Nadia Zoubir", "Ilham Battas", "Hicham Medromi", "Fouad Moutaouakkil" ], "abstract": "Background Leukemia, a heterogeneous group of blood cancers, poses significant challenges to global health due to its complexity, diverse risk factors, and variable outcomes. Accurate and early diagnosis is critical but remains a significant hurdle, particularly in low-resource settings. Recent advancements in artificial intelligence (AI) and image processing offer transformative solutions to improve leukemia detection and classification, addressing limitations in traditional diagnostic methods.\n\nMethods This study systematically reviewed over 25,000 scientific articles sourced from Scopus, employing a PRISMA-guided methodology to ensure a comprehensive and rigorous analysis. The analysis focused on the application of AI, particularly convolutional neural networks (CNNs), in diagnosing four primary leukemia types: acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), and chronic myeloid leukemia (CML). It also examined global epidemiological trends, risk factors, and disparities in healthcare access.\n\nResults Key risk factors for leukemia include genetic syndromes like Down syndrome, environmental exposures to toxins such as benzene, ionizing radiation, and viral infections. Socio-economic disparities and geographical differences significantly impact leukemia incidence and outcomes. AI-based models, especially CNNs, demonstrated enhanced accuracy, speed, and reliability in diagnosing leukemia compared to traditional methods. However, challenges such as data variability, model scalability, and unequal access to AI technologies continue to hinder widespread adoption.\n\nConclusion AI and image processing technologies hold immense potential to revolutionize leukemia diagnostics by enabling early detection, precise classification, and personalized treatment planning. Addressing critical challenges, including data standardization and equitable access to these technologies, will be vital for global application. This review highlights the transformative role of AI in improving leukemia outcomes and advancing precision medicine worldwide.", "keywords": [ "Leukemia Classification", "Artificial Intelligence in Medical Diagnostics", "Machine Learning Algorithms", "Hematologic Malignancies", "Deep Learning Models", "Convolutional Neural Networks", "Cancer Epidemiology", "Diagnostic Accuracy." ], "content": "1. Introduction\n\nLeukemia is a complex hematological malignancy defined by the abnormal proliferation of white blood cells, which disrupts the body’s blood-forming tissues and interferes with normal immune functions. This group of cancers encompasses both acute and chronic forms affecting either lymphoid or myeloid cells, resulting in various subtypes with distinct clinical features and progression rates.1,2 Key characteristics of leukemia include rapid, uncontrolled cell growth that impairs immune functions, leads to anemia, and creates other systemic impacts. Leukemia’s morphological diversity complicates diagnosis and subtype differentiation, as highlighted by Mirmohammadi et al.3 and Safuan et al.4 This diversity in presentation underscores the need for accurate classification, which is essential for effective prognosis and treatment planning.\n\nDue to high mortality rates and diagnostic challenges associated with leukemia, accurate classification has become increasingly critical. Advances in artificial intelligence (AI) are transforming leukemia detection and classification in medical imaging. Studies demonstrate that machine learning and deep learning models, including convolutional neural networks (CNNs), enhance diagnostic precision and enable faster subtype identification, promising substantial potential for clinical applications.5,6 This study aims to address the specific challenges in leukemia classification by developing an AI-driven approach to accurately detect and classify leukemia subtypes.\n\nThis paper provides an in depth exploration of leukemia detection and classification, beginning with an analysis of global epidemiology to outline incidence, prevalence, and risk factors across regions and demographics, thereby highlighting disparities and risk factors. The second section delves into classification, focusing on primary leukemia subtypes: ALL, AML, CLL, and CML and examining morphological and genetic characteristics important for diagnosis. In the third section, we describe our methodology, which details our data collection, processing, and analytical techniques used to perform a robust bibliometric analysis. This bibliometric analysis identifies publication trends, key contributors, and influential studies, offering insights into the current landscape of leukemia research. Lastly, we present a comparison of state-of-the-art AI applications for leukemia, reviewing machine learning and deep learning techniques employed in diagnostics. Here, we discuss each approach’s limitations, emphasizing challenges related to data quality, generalizability, and integration into clinical workflows.\n\n\n2. Global epidemiology and risk factors of Leukemia\n\nThe epidemiology of leukemia has been extensively analyzed through various studies over the years, revealing significant trends, risk factors, and geographical disparities. Bouchbika et al. conducted a study in Greater Casablanca from 2005 to 2007, using data from the Casablanca cancer registry. They found age-standardized rates (ASR) of 2.7 per 100,000 for men and 2.0 for women, with incidence peaks in children aged 59 and adults over 65. Comparatively, Morocco’s leukemia rates were lower than those in Tunisia and Algeria, and significantly lower than Western countries. Childhood leukemia (0-14 years) constituted 10.9% of all childhood cancers with an ASR of 1.4.7 Expanding the analysis globally, Miranda- Filho et al. used data from 290 cancer registries in 68 countries and national estimates from 2012. Their study found the highest ASRs in Australia, New Zealand (11.3 per 100,000 in males, 7.2 in females), Northern America (10.5 in males, 7.2 in females), and Western Europe (9.6 in males, 6.0 in females). Conversely, Western Africa had the lowest rates (1.4 in males, 1.2 in females). Acute lymphoblastic leukemia was predominant in children, while chronic lymphocytic leukemia was more common in adults in European and North American countries. Key risk factors identified included genetic syndromes such as Down syndrome, environmental exposures like benzene and ionizing radiation, and viral infections.8 In Arab countries, Al-Muftah and Al-Ejeh highlighted higher leukemia incidence rates among younger populations, particularly in the Arabian Gulf and Levant regions. Utilizing GLOBOCAN data from 2003 to 2016, the study noted elevated age-standardized incidence rates (ASIR) in children under 15 years compared to global averages. For adults aged 35 and older, the Levant region also showed higher rates than the global norm. These findings emphasized the need for targeted genetic epidemiology studies and improved clinical management strategies.9 Further, ElBakali, Abdellatif, and Smiri examined 8,851 cancer cases in the Souss Massa region of Morocco from 2014 to 2019. Hematological cancers, including leukemia, accounted for 14% of cases, with a female incidence rate of 47.74 per 100,000 and a male incidence rate of 45.71 per 100,000. The average age of leukemia patients was 47.81 years. Their analysis underscored the importance of reinforcing prevention efforts and developing specific screening strategies for hematological cancers.10 Baeker Bispo, Pinheiro, and Kobetz provided a comprehensive overview of leukemia and lymphoma in the US and globally, reporting higher incidence rates in developed regions and racial disparities in survival rates. They noted that leukemia is the 15th most common cancer worldwide, accounting for 437,033 cases and 309,006 deaths in 2018. The study highlighted genetic abnormalities, immunosuppression, ionizing radiation, carcinogenic chemicals, and oncogenic viruses as key risk factors. They emphasized the need for equitable access to diagnostic and treatment services to address these disparities.11 Smith Torres-Roman et al. analyzed leukemia motality trends in children from 15 Latin American countries between 2000 and 2017. They found the highest mortality rates in Venezuela, Ecuador, Nicaragua, Mexico, and Peru, with significant upward trends in Nicaragua and Peru. Conversely, Puerto Rico saw substantial declines. The study predicted that by 2030, leukemia mortality would increase in several countries, emphasizing the need for interventions to reduce inequalities and ensure universal healthcare coverage.12 Huang et al. conducted a global analysis using data from GLOBOCAN, CI5, WHO, NORDCAN, and SEER databases. They found that leukemia accounted for 2.5% of new cancer cases and 3.1% of cancer deaths, with an age-standardized incidence rate (ASIR) of 5.4 and mortality rate of 3.3 per 100,000 people. The study associated higher incidence and mortality rates with the Human Development Index, GDP per capita, and lifestyle factors such as smoking and obesity. They recommended lifestyle modifications and further research to understand these trends.13 Finally, Zhang et al. examined the global burden of hematologic malignancies from 1990 to 2019 using data from the Global Burden of Disease (GBD) study. They found a declining trend in leukemia incidence (ASIR of 8.22 per 100,000) and mortality (ASDR of 4.26 per 100,000), with significant decreases in age-standardized rates.\n\nHowever, regions like Central Europe, Western Europe, and East Asia experienced increases. The study highlighted the influence of socio-economic factors, occupational exposures, and high BMI on leukemia burden, emphasizing the need for targeted prevention strategies and improved healthcare access.14 these studies collectively highlight the global variations in leukemia incidence and mortality, influenced by genetic, environmental, and socio-economic factors. While some regions have seen declining trends, others continue to face significant challenges, underscoring the need for comprehensive prevention and management strategies tailored to specific regional needs.\n\n\n3. Classification of Leukemia\n\nLeukemia is primarily classified into two types: acute, which progresses rapidly, and chronic, which develops more slowly. The four major forms of leukemia include Acute Lymphoblastic Leukemia (ALL), Acute Myeloid Leukemia (AML), Chronic Lymphocytic Leukemia (CLL), and Chronic Myeloid Leukemia (CML)\n\nAccording to FAB classification, Acute Lymphoblastic Leukemia (ALL) is the type of leukemia that occurs most frequently. It is further classified into three morphological subtypes: L1, L2 and L3. L1 cells have small size with coarse granular appearance they are composed of uniform groups of cells. L2 cells have bigger sizes as compared to L1 cells but also show variation in the appearance of the nucleus. L3 are bigger than L1 cells too but they have more vacuoles in them, with homogeneity in their nuclei.15 The defining feature of ALL is the uncontrolled growth of atypical lymphocytes, or lymphoblast cells, which can be morphologically identified from other cell types by their irregular shape, small clefts within a body part and round objects in a nucleus consisting from one to many nuclear lobes.16 In fact, according to some, early diagnosis, especially in children, plays a critical role in the treatment process.\n\nDiagnosis of ALL, however, is not an easy task because Leukemia cells L1, L2, L3, are morphologically very similar to those of normal and reactive lymphocytes and atypical lymphocytes, which are nonmalignant cells or noncancerous cells.\n\nFigure 1 illustrates the microscopic appearance of Acute Lymphoblastic Leukemia (ALL) cells, while Table 1 presents a comparative analysis of the morphological and cytological characteristics of different ALL subtype.\n\nAcute myeloid leukemia (AML) is a type of cancer characterized by rapid growth, abnormal white blood cells that accumulate in the bone marrow, and inhibition of the production of normal blood cells. Using the French American-British system, AML is classified into several subtypes according to the morphology and immunophenotype of the leukemic cells. Each of these types has distinct features that assist in its diagnosis and the formulation of a line of treatment.17\n\nFigure 2 shows the microscopic image of Acute Myeloid Leukemia (AML) cells, highlighting their distinct morphological features.\n\nAML arises from the transformation of myeloid precursor cells, leading to the uncontrolled proliferation of immature white blood cells, known as myeloblasts. These blasts fail to differentiate into fully functional blood cells, resulting in a deficiency of red blood cells, platelets, and mature white blood cells. This disruption in normal blood cell production causes symptoms such as anemia, infection, and bleeding tendencies. The classification of AML subtypes is essentiel, as it guides the clinical management of the disease. For instance, acute promyelocytic leukemia (APL), classified as M3, is notable for its sensitivity to all-trans retinoic acid (ATRA) therapy, which can induce remission. Other subtypes require different treatment approaches based on their unique characteristics, making accurate classification essential for effective therapy. Subtypes, including M0 (acute myeloblastic leukemia with minimal differentiation), M1 (acute myeloblastic leukemia without maturation), M2 (acute myeloblastic leukemia with maturation), M3 (acute promyelocytic leukemia and its subtypes), M4 (acute myelomonocytic leukemia), M5 (acute monocytic leukemia), M6 (acute erythroid leukemia), and M7 (acute megakaryocytic leukemia), with their characteristics, are presented in the Table 2.\n\nChronic lymphocytic leukemia (CLL) is primarily characterized by the proliferation of small, mature B-lymphocytes in the peripheral blood, bone marrow, and secondary lymphoid tissues. Morphologically, CLL lymphocytes are generally small with scant cytoplasm and dense nuclear chromatin, often referred to as” smudge cells” due to their fragility. However, variations in lymphocyte morphology have been identified, leading to the classification of CLL into distinct morphological sub- groups. According to Ghia et al., the typical lymphocytes in CLL are small and homogeneous, exhibiting fragile cell membranes which result in frequent cell rupture during blood smear preparation. These cells are predominantly characterized by a condensed nuclear chromatin and narrow rims of pale cytoplasm.18 In contrast, Peterson et al. identified three morphologic groups based on lymphocyte size and structure: Group I consists of small to medium-sized lymphocytes with coarsely clumped chromatin, Group II features larger lymphocytes with abundant cytoplasm resembling reactive lymphocytes, and Group III includes a heterogeneous mix of lymphocytes from both Groups I and II.\n\nNotably, the identification of these morphologic subtypes helps refine the diagnosis and risk stratification of patients, guiding more personalized treatment strategies.\n\nThis classification is significant as it correlates with disease progression and patient prognosis, with larger, reactive-like lymphocytes associated with longer survival times.19\n\nFigure 3 illustrates the microscopic appearance of Chronic Lymphocytic Leukemia (CLL) cells, highlighting their distinct characteristics.\n\nTable 3 presents a detailed comparison of CLL subtypes, emphasizing their unique morphological and cytological features.\n\nCML represents a clonal hematopoietic stem cell disorder characterized by uncontrolled myeloid cell proliferation. CML usually develops in three distinct phases: the chronic phase, the accelerated phase, and finally, blast crisis. The chronic phase is characterized by an insidious onset of symptoms, and it is the longest of the three phases, it is the one during which most patients are stable, with relatively manageable symptoms. Advanced phase: An advance in the progress of the disease, with increasing symptoms and resistance to treatment, thus more ag gressive clinically. The accelerated phase is a progression of the disease, bringing with it more severe symptoms and resistance to treatment, making it clinically more aggressive. The blast crisis phase, however, is the most dramatic phase, with rapid proliferation of the very immature, similar-to-acute-leukemia cells, often refractory to treatment and carries a poor prognosis. The hallmark of CML is represented by the Ph chromosome, resulting from the reciprocal translocation between chromosomes 9 and 22, forming the BCR-ABL fusion gene. This fusion gene encodes for a constitutively active tyrosine kinase acting benignly to drive the leukemogenesis in CML.20\n\nThe CML cells can, therefore, be broadly divided into the developmental stages of myeloblasts, promyelocytes, myelocytes, metamyelocytes, bands, and neutrophils.\n\nIt has typical morphological features characteristic of its own variety, extremely vital in an accurate diagnosis and categorization. Myeloblasts are the most primitive cells, characterized by large nuclei with fine-type chromatin and basophilic cytoplasm.\n\nDuring the development of cells into Promyelocytes, Myelocytes, Metamyelocytes, Bands, and finally Neutrophils, there are many conspicuous changes in nuclear shape, chromatin texture, color of cytoplasm, and presence of granules.\n\nIt is through the realization of these different characteristics that pathologists are able to reach a proper diagnosis with relation to CML and monitor the progression of the disease.21\n\nFigure 4 illustrates the microscopic image of Chronic Myeloid Leukemia (CML) cells.\n\nThe Table 4 provides a comparative analysis of the various subtypes of Chronic Myeloid Leukemia (CML), focusing on their specific cellular characteristics. It outlines differences in nucleus lobes, chromatin texture, nucleus shape, cytoplasm color, granules, and nucleoli for each cell type, ranging from immature myeloblasts to mature neutrophils.\n\n\n4. Overview of AI in medical diagnostics\n\nworld of medical diagnostics, offering a powerful set of tools to analyze and interpret vast amounts of medical data.19 Subfields of AI, like machine learning (ML) and deep learning (DL), are particularly adept at finding patterns and making predictions based on these large datasets. In healthcare, this translates to several key benefits: improved diagnostic accuracy, better prediction of patient outcomes, and the potential for personalized treatment plans. Machine learning algorithms, such as support vector machines (SVMs) and k-nearest neighbors (KNNs), have proven successful in diagnosing diseases. These algorithms can analyze medical images and patient data to identify patterns indicative of specific conditions.22 Deep learning, particularly convolutional neural networks (CNNs), has become a game-changer in medical imaging. CNNs are exceptionally skilled at detecting and classifying various diseases, including cancers and cardiovascular conditions.23,24 For example, CNNs have achieved high accuracy in detecting diabetic retinopathy, show- casing their potential to revolutionize ophthalmology.25 Additionally, AI-powered systems are being integrated into radiology workflows, acting as assistants to radiologists. These systems can identify abnormalities in X-rays and MRI scans, leading to fewer diagnostic errors and improved efficiency. These advancements highlight the immense potential of AI to transform healthcare delivery and ultimately improve patient care.\n\n\n5. Methodology\n\nIn this study, we employed a comprehensive bibliometric analysis approach to explore various aspects of leukemia, leveraging data from Scopus. Our research spanned multiple queries tailored to specific sections of the study, including risk factors, classification, and the role of artificial intelligence in leukemia diagnosis and treatment. Initially, we conducted broad searches using relevant keywords in article titles, abstracts, and keywords, resulting in a cumulative total of over 25000 articles across different sections. For the section on risk factors, we identified and screened 12,218 articles, focusing on publications from 2019 to 2024. We further refined this selection based on relevance, leading to a final set of 2116 articles. The classification section included data from 12,514 articles, filtered similarly to ensure the inclusion of 2189 pertinent studies.\n\nIn our analysis of AI’s role in leukemia, we extracted data from 368 articles, highlighting key terms such as “artificial intelligence,” “machine learning,” and “deep learning.” Articles were included based on their focus on the application of AI in medical diagnostics and treatment.\n\nWe meticulously recorded detailed bibliographic information for each selected article, including authors, titles, journal names, publication dates, and citation counts. To visualize and analyze citation networks, we utilized tools like VOSviewer, which enabled us to identify central themes and emerging trends within the field. The study’s procedural flow and inclusion criteria are detailed in the PRISMA diagram, illustrating the rigorous selection and screening process undertaken to ensure a comprehensive review of the literature.\n\nFigure 5 illustrates the PRISMA research methodology used in this study.\n\nFigure 6 depicts the publication trend over the years 2019 to 2024, showing the number of articles published per year and the corresponding total citations.\n\nThe graph presents the trend of publications and total citations per year related to leukemia research from 2019 to 2024. The blue line indicates the number of articles published each year, while the orange line represents the total number of citations these articles received per year.\n\nThe data reveal an initial increase in both the number of publications and citations, peaking in 2021. This suggests a heightened interest and active engagement in leukemia research during this period. The rise in publications could be attributed to advancements in research methodologies, increased funding, or a growing awareness of leukemia-related issues.\n\nThe decrease in citations from 2022 can be attributed to the time lag that often occurs in academic publishing, where newly published articles take time to be discovered, read, and subsequently cited by other researchers. This lag is a common phenomenon, as it typically takes a few years for new research to become well-known and integrated into the broader academic discourse.\n\nThe keyword analysis of leukemia risk factors was conducted using data extracted from Scopus, encompassing a total of 12,218 articles. The query “risk” AND “factors” AND “leukemia” was employed to gather relevant literature between 2019 and 2024. The analysis visualized the associations and prevalence of key terms, highlighting the most frequently discussed topics in leukemia research. Central themes include genetic factors, environmental exposures, and demographic variables, with significant nodes like “human,” “acute myeloid leukemia,” “genetics,” and “risk factor” illustrating the interconnected nature of these elements. The network graph provides a comprehensive overview of the critical areas of focus within the field, offering valuable insights into the prevailing concerns and emerging trends in leukemia research. This foundational understanding informs the subsequent exploration of epidemiological patterns and risk factors as reported in various studies.\n\nFigure 7 presents a keyword co-occurrence network highlighting the relationships between terms associated with leukemia and its risk factors.\n\nThe classification of leukemia was further explored through a comprehensive keyword analysis using data extracted from Scopus. This analysis encompassed 12,514 articles, focusing on keywords with more than 160 occurrences. The visualization provided a detailed mapping of the terminologies and concepts frequently associated with various types of leukemia. Prominent nodes in the network include terms like “acute myeloid leukemia,” “genetics,” “acute lymphoblastic leukemia,” and “treatment response,” among others. The graph illustrates the relationships and common themes in the literature, highlighting the intricate connections between different leukemia types, their genetic markers, treatment modalities, and related biological processes.\n\nFigure 8 visualizes the keyword co-occurrence network, highlighting key terms and their relationships in the classification of leukemia.\n\nThe integration of artificial intelligence (AI) in leukemia research has been gaining significant traction, as evidenced by a comprehensive keyword analysis conducted on 368 articles from Scopus. This analysis focused on keywords with more than 20 occurrences, highlighting the prevalent themes and emerging trends within this interdisciplinary field. Key terms such as “artificial intelligence,” “machine learning,” “deep learning,” and specific medical terms like “chronic myeloid leukemia” and “diagnosis” emerged as central nodes in the network graph. The visualization showcases the increasing application of AI techniques in the medical domain, particularly in enhancing diagnostic accuracy, predicting patient outcomes, and enabling personalized treatment approaches. The centrality of AI-related keywords underscores a growing emphasis on advanced computational methods in contemporary leukemia studies.\n\nFigure 9 illustrates the keyword co-occurrence network focusing on AI in leukemia research.\n\nBeyond keyword analysis, we investigated collaboration pat- terns among countries to gain a broader perspective on the global landscape of leukemia research. VOS viewer allows us to analyze” citations” as the unit of analysis, with” countries” as the collaborating entities. This analysis focused on prominent collaborations, excluding documents with a vast number of co-authoring countries. The resulting map depicts the citation impact and collaborative networks of various countries from 2016 to 2022. The visualization reveals the United States, India, and the United Kingdom as central hubs, signifying their leadership and significant contributions to leukemia research. India, in particular, displays a notable network of connections with countries like Pakistan, South Korea, and Saudi Arabia. This highlights India’s active role in fostering collaborative re- search endeavors. The color gradient, transitioning from blue to yellow, represents the publication timeline. This trend visually depicts the growing intensity of international collaborations and citations in leukemia research over the past few years. This underscores the increasingly global nature of the field and emphasizes the critical role of multinational cooperation in driving advancements in leukemia research.\n\nFigure 10 presents a citation analysis of AI in leukemia research based on countries, highlighting the collaborative networks and the leading contributions from nations.\n\nThe bibliometric analysis revealed a clear trend: artificial intelligence (AI) is rapidly transforming leukemia research. AI offers significant advantages, including improved diagnostic accuracy, personalized treatment plans, and enhanced prediction capabilities.\n\nThese powerful techniques empower researchers and processing algorithms being applied. This in-depth examination aims to provide a clearer picture of the current landscape and identify key areas where innovation and improvement can revolutionize leukemia diagnostics and treatment.\n\n\n6. Advancements in Leukemia detection using AI and image processing techniques\n\nIn the literature, one promising approach for leukemia detection relies on microscopic image processing through a multi-step pipeline to analyze blood smear images. The process begins with image acquisition, where high-quality blood smear images are obtained. These images then undergo preprocessing, including resizing, RGB conversion, contrast adjustment, and noise reduction, ensuring the data is optimized for further analysis. In the segmentation stage, individual cells, particularly white blood cells, are isolated from the background and neighboring cells to focus on relevant regions. The next step is feature extraction, where important characteristics like shape, size, color, texture, and morphology are identified from the segmented cells. These features are then analyzed in the leukemia detection phase, where machine learning or deep learning models process the data to identify potential abnormalities. Finally, in the classification stage, each cell is categorized as either normal or cancerous, supporting the diagnosis and treatment planning for leukemia.26,27 This structured workflow enhances accuracy by refining the data at each step, ensuring reliable detection and classification.\n\nThe Figure 11 illustrates the process as described in the literature.\n\nBuilding on this workflow, the literature offers various innovative methodologies that further enhance the accuracy and efficiency of leukemia detection through advanced computational techniques. Recent studies demonstrate the potential of integrating machine learning, deep learning, and feature extraction methods to improve diagnostic outcomes.\n\nWarnat-Herresthal et al. (2020) conducted a comprehensive study using 12,029 samples from 105 different studies to develop robust and scalable classifiers for Acute Myeloid Leukemia (AML). Their methodology involved rigorous preprocessing steps, including RMA normalization for microarray data and DESeq2 normalization for RNA-seq data, followed by trimming to common genes. The study employed L1-regularized logistic regression (lasso) and compared its performance with other machine learning methods such as k-nearest neighbors, linear SVM, linear discriminant analysis, random forests, and deep neural networks. The classifiers demonstrated high accuracy, sensitivity, and specificity, and were evaluated through extensive cross-study and cross-platform analyses, ensuring their robustness and generalizability. This work underscores the potential of machine learning combined with transcriptomics to enhance AML diagnostics, particularly in settings where traditional hematological expertise is limited. Despite the promising results, the study acknowledged some limitations. Performance variability due to cross-study and cross-platform differences was noted, highlighting the need for large, diverse training datasets to improve the model’s generalizability. Furthermore, the study identified the high reliance on the quality and diversity of the dataset as a potential weakness. Future directions for this research include integrating transcriptomic-based machine learning into clinical workflows for AML diagnostics, which could offer a scalable and cost-effective alternative to traditional diagnostic methods. The authors propose further refining these models and extending their application to other hematologic and non-hematologic diseases, potentially revolutionizing medical diagnostics. Additionally, future work aims to include prospective studies to assess the diagnostic utility of these models in real-world clinical settings, addressing cross-study and cross-platform variations, and exploring simple data transformations to enhance cross-platform generalizability.28\n\nLoey et al. (2020) proposed two automated classification models utilizing transfer learning with the pre-trained deep convolutional neural network, AlexNet, to classify leukemia in blood microscopic images. The first model involves pre-processing the images, extracting features using a pre-trained AlexNet, and classifying these features using various classifiers such as SVM, linear discriminants (LD), decision trees (DT), and k-nearest neighbors (K-NN). The second model fine-tunes AlexNet for both feature extraction and classification. Experiments on a dataset of 2820 images show that the second model achieves a perfect classification accuracy of 100%, outperforming the first model. The study emphasizes the advantages of using transfer learning to overcome the challenges of designing and training deep neural networks from scratch. The pre-processing step includes converting images to RGB, resizing them to 227×227 pixels, and applying data augmentation techniques like translation, reflection, and rotation. For feature extraction, AlexNet’s architecture, which includes five convolutional layers, three fully connected layers, and max-pooling layers, is utilized. The classifiers tested in the first model include decision trees with a max-split of 20, linear discriminants, SVM with various kernel functions (linear, Gaussian, cubic), and K-NN with Euclidean distance. Performance metrics such as precision, recall, accuracy, and specificity were evaluated, with the second model demonstrating higher accuracy and robustness. Despite the promising results, the study acknowledged some limitations and future directions. Potential overfitting remains a concern despite the use of dropout and normalization techniques. Additionally, the models’ dependence on high-quality, well-annotated datasets for training poses challenges for broader applicability. Future research could focus on extending these models to differentiate between various types of leukemia, rather than just identifying the presence of leukemia. Furthermore, the use of larger and more diverse datasets could help validate the models’ robustness and accuracy. The authors propose integrating these automated systems into clinical workflows to aid in the early and accurate diagnosis of leukemia, ultimately improving patient outcomes through timely and appropriate treatment.29\n\nBaig et al. (2022) developed a comprehensive approach using deep learning-based convolutional neural networks (CNN) to detect acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), and multiple myeloma (MM) from microscopic blood smear images. The study “Detecting Malignant Leukemia Cells Using Microscopic Blood Smear Images: A Deep Learning Approach” by Raheel Baig et al. investigates an advanced automated system for detecting various types of malignant leukemia cells using deep learning techniques. The proposed methodology integrates pre-processing, feature extraction, and classification to identify Acute Lymphoblastic Leukemia (ALL), Acute Myeloid Leukemia (AML), and Multiple Myeloma (MM) from blood smear images. The dataset, comprising 4150 images, is pre-processed to enhance image quality, including background elimination, noise reduction, and contrast enhancement using adaptive histogram equalization. Two Convolutional Neural Networks (CNN-1 and CNN-2) with 19 and 15 layers, respectively, are employed for feature extraction. The extracted features are then fused using Canonical Correlation Analysis (CCA) to improve the discriminative power of the features. Finally, the fused features are classified using various machine learning classifiers, including Bagging Ensemble, LPBoost, Total Boost, RUSBoost, Fine KNN, and SVM, with the Bagging Ensemble achieving the highest accuracy of 97.04%.The methodology involves multiple steps, starting with pre-processing, where RGB images are converted to grayscale, enhanced using intensity adjustment, and noise removed through area and closing operations. Feature extraction is performed using CNN models trained on pre-processed images. CNN-1 and CNN-2, containing convolutional, batch normalization, ReLU, max-pooling, fully connected, SoftMax, and classification layers, extract deep features, which are then combined using CCA fusion. The classification of leukemia subtypes is performed using machine learning classifiers, evaluated using metrics like accuracy, sensitivity, specificity, precision, and F1-score. The Bagging Ensemble classifier outperforms others, highlighting the system’s robustness and potential for clinical application. Despite the promising results, the study acknowledged some limitations and future directions. The method’s performance is dependent on high-quality, well-annotated datasets, which may limit its applicability in settings with varied data quality. Additionally, there is a potential risk of overfitting, despite the use of data augmentation techniques. Future research aims to extend the current methodology to include larger, more diverse datasets and refine the algorithms to improve robustness and accuracy. The authors propose further integration of this automated system into clinical workflows, aiding in the early and accurate diagnosis of leukemia, which could significantly enhance patient outcomes. Future research directions include differentiating between more subtypes of leukemia and exploring the applicability of the methodology to other hematologic and non-hematologic conditions.\n\nThis vision underscores the potential of advanced deep learning techniques to revolutionize medical diagnostics, improving efficiency and accuracy in detecting malignant conditions.30\n\nShafique and Tehsin (2018) proposed a robust and automated detection method for acute lymphoblastic leukemia (ALL) and its subtypes (L1, L2, and L3) using transfer learning with a pretrained deep convolutional neural network (DCNN) AlexNet. The ALL-IDB database, which contains images from both healthy patients and leukemia patients, was used for this research. The dataset was split into training and evaluation sets, with data augmentation applied to prevent overfitting. The authors utilized the AlexNet architecture, which includes five convolutional layers followed by three fully connected layers. The last layers were replaced with new ones to classify the input images into four classes: L1, L2, L3, and Normal. Preprocessing steps included image normalization, conversion into binary images, and noise reduction. The feature extraction was done using the pretrained layers of AlexNet, and classification was achieved using the softmax activation function. The model achieved an accuracy of 99.50% for ALL detection and 96.06% for subtype classification. The researchers highlighted that DCNNs are powerful enough to detect and classify leukemia without the need for manual image segmentation. The study also suggests further improvements by integrating the approach into a fully automated system and expanding the dataset for training from scratch to enhance diagnostic accuracy. The methodology involves several key steps. In preprocessing, images are normalized, converted into binary format, and subjected to noise reduction. Unlike traditional methods, no manual segmentation is needed due to the strength of the DCNN layers. Feature extraction is performed using the pretrained AlexNet layers, and the classifier is a modified AlexNet architecture with new fully connected layers for classification. The network includes five convolutional layers, three max-pooling layers, and three newly added fully connected layers for classification. The algorithm characteristics include ReLU activation, softmax for classification, and data augmentation techniques like image rotation and mirroring to enhance the model’s robustness. The study achieved notable performance metrics with an accuracy of 99.50% for ALL detection and 96.06% for subtype classification. Sensitivity was 100% for ALL detection and 96.74% for subtype classification, while specificity was 98.11% for ALL detection and 99.03% for subtype classification. These results underscore the high accuracy and effectiveness of using transfer learning with DCNNs for leukemia detection. Despite the promising results, the study acknowledges some limitations and future directions. The limited dataset could affect the robustness of the training, and further research is needed to integrate the system into a fully automated diagnostic tool. Future work should focus on training deep learning models from scratch with larger datasets to improve accuracy and reliability. The authors also propose integrating their approach into a fully automated system to assist pathologists and oncologists in the early and accurate diagnosis of leukemia, ultimately enhancing patient outcomes through timely treatment interventions.31\n\nChiaretti et al. (2014) outlined a comprehensive approach to diagnosing and subclassifying Acute Lymphoblastic Leukemia (ALL) using bone marrow morphology, multi-channel flow cytometry (MFC), and genetic/cytogenetic analysis, following the WHO classification of lymphoid neoplasms. Their work provides a detailed overview of current standards and methodologies. The approach integrates cell morphology, immunophenotyping, and genetic/cytogenetic studies as outlined in the 2008 WHO classification. Morphological assessment is the initial diagnostic step, distinguishing ALL from AML based on cell characteristics in bone marrow and peripheral blood. Immunophenotyping with multi-channel flow cytometry (MFC) is the gold standard for identifying cell lineage and defining subsets, utilizing specific markers for B- and T-lineage ALL. Cytogenetic and genetic analyses, including karyotyping, fluorescence in situ hybridization (FISH), array-CGH, and next-generation sequencing (NGS), further refine the diagnosis and provide critical prognostic information.\n\nThe article classifies ALL into B-lymphoblastic leukemia/lymphoma not otherwise specified, B-lymphoblastic leukemia/lymphoma with recurrent cytogenetic alterations, and T-lymphoblastic leukemia/lymphoma. Each subtype is characterized by specific immunophenotypic markers and genetic aberrations. For instance, B-lineage ALL markers include CD19, CD20, CD22, CD24, and CD79a, while T-lineage ALL markers include CD1a, CD2, CD3, CD4, CD5, CD7, and CD8. The review highlights the prognostic significance of various genetic lesions, such as IKZF1 deletions in BCR-ABL+ ALL and CRLF2 rearrangements in B-ALL, which are associated with poor outcomes. The authors advocate for prospective clinical trials to ensure accurate diagnosis and effective therapy, emphasizing that early diagnostic work should be performed by experienced personnel to optimize treatment outcomes.30 Despite its strengths, the study acknowledges certain limitations and future directions. The method heavily relies on high-quality samples and advanced diagnostic facilities, which may not be accessible everywhere. Additionally, diagnostic accuracy can vary across different laboratories. To address these issues, the authors stress the importance of conducting early and accurate diagnostic work-ups by experienced personnel. They recommend integrating advanced genetic and immunophenotyping techniques into clinical practice to optimize treatment strategies. Future research aims to identify novel subgroups with prognostic significance and refine diagnostic algorithms to enhance patient outcomes. Prospective clinical trials are also suggested to ensure diagnostic accuracy and therapeutic efficacy, ultimately improving the comprehensive understanding and treatment of ALL.32\n\nMinal D. Joshi, Atul H. Karode, and S.R. Suralkar (2013) developed an automatic method for detecting acute leukemia in blood microscopic images using image processing techniques and a k-nearest neighbor (kNN) classifier. The methodology involves preprocessing, segmentation, feature extraction, and classification to differentiate between normal and leukemic cells. In the preprocessing step, RGB images are converted to grayscale, contrast is enhanced using histogram equalization, and linear contrast stretching is applied to adjust image intensity levels. The segmentation process employs Otsu’s thresholding method to convert grayscale images to binary images, followed by morphological operations to remove small pixel groups and noise. Feature extraction focuses on calculating three morphological features of lymphocyte cells: area, perimeter, and circularity. These features are then used by the kNN classifier to classify the cells as either normal or blast (leukemic). The proposed system was evaluated on 108 images from the ALL-IDB public dataset, achieving an overall accuracy of 93%. The methodology includes several key steps. Preprocessing involves converting RGB images to grayscale, applying histogram equalization for contrast enhancement, and using linear contrast stretching for image intensity adjustment. Noise reduction is achieved through area and closing operations. Segmentation is done using Otsu’s thresholding to convert grayscale images to binary images, followed by morphological operations to remove small pixel groups and noise, and connect neighboring pixels to form objects. Feature extraction calculates the area by counting the total number of non-zero pixels within the nucleus region, measures the perimeter by calculating the distance between successive boundary pixels of the nucleus, and calculates circularity. The kNN classifier then uses these features to classify lymphocyte cells as either normal or blast (leukemic). The performance metrics showed an overall accuracy of 93%, with high precision in classifying blast cells, high recall in identifying leukemic cells, and high specificity in distinguishing between normal and blast cells. Despite the promising results, the study identified some limitations and future directions. The system’s performance depends on high-quality, well-annotated datasets, and it may be sensitive to variations in image quality and staining. Future research aims to optimize image processing techniques to improve robustness and accuracy, integrate more advanced machine learning classifiers, and explore stain-independent segmentation methods to enhance reliability. Expanding the dataset and refining the algorithms to improve the classification of various leukemia subtypes are also recommended. Integrating this automated system into clinical workflows could significantly aid in early and accurate leukemia diagnosis, thereby improving patient outcomes.33\n\nTusneem A. Elhassan et al. (2023) propose a novel approach for classifying atypical white blood cells (WBCs) in Acute Myeloid Leukemia (AML) using a hybrid model that integrates a geometric transformation (GT) with a deep convolutional autoencoder (DCAE) and a convolutional neural network (CNN). The methodology involves preprocessing, augmentation, feature extraction, and classification to differentiate between typical and atypical WBCs, followed by further subclassification of atypical WBCs into eight distinct categories. The dataset used includes 18,365 single-cell images from AML patients and non-malignant controls, categorized into 15 different types of WBCs.\n\nThe preprocessing steps include random rotation, vertical and horizontal flipping, and augmentation using the GT-DCAE model. Feature extraction is performed using the DCAE, which compresses the input data into a low-dimensional latent representation, followed by a CNN for additional feature extraction. The classification involves a two-stage process: the first stage differentiates between typical and atypical WBCs, and the second stage classifies atypical WBCs into subtypes. The proposed model achieved an average accuracy of 97%, a sensitivity of 97%, and a precision of 98%, with an AUC of 99.7%, demonstrating exceptional discriminating abilities. The methodology includes several key steps. Preprocessing involves augmentation techniques like random rotation and flipping, with the GT-DCAE model generating synthetic images. Segmentation is carried out using the CMYK-Moment Localization-Feature Fusion Extraction framework. For feature extraction, the DCAE model employs an encoder network with three convolutional layers, a latent vector space, and a decoder network. Classification is conducted in two stages using CNNs with ReLU activation, L2 regularization, and the SoftMax function.\n\nPerformance metrics highlight the model’s effectiveness with an overall accuracy of 97%, precision of 98%, sensitivity of 97%, and an AUC of 99.7%. Class-wise performance indicates high precision and sensitivity for various WBC subtypes, such as myeloblasts and monoblasts, although there are areas for improvement in detecting certain subtypes like metamyelocytes and bilobed promyelocytes.\n\nDespite the promising results, the study identifies some limitations and future directions. The model’s performance depends on high-quality, well-annotated datasets, and there is potential for misclassification in the intermediate stages of myelopoiesis. Future research aims to refine the hybrid model by increasing the dataset size and incorporating additional augmentation techniques. The authors also propose integrating this model into clinical workflows to aid in the early and accurate diagnosis of AML, thereby improving patient outcomes through timely treatment interventions. Furthermore, they suggest exploring the application of their model to other hematologic and non-hematologic conditions to expand its utility and effectiveness in medical diagnostics.34\n\nNegm et al. (2017) present an advanced decision support system for acute leukemia classification using digital microscopic images, combining neural networks and decision tree models to achieve high efficiency and accuracy. The neural network model, favored for its sensitivity, comprises multiple convolutional layers, batch normalization, and dropout to prevent overfitting, while the decision tree model, noted for its speed, provides a robust comparison.\n\nThe methodology involves several steps. Preprocessing techniques such as histogram equalization and contrast enhancement are employed to improve image quality. Segmentation is performed using Otsu thresholding and fuzzy c-means clustering. Feature extraction involves capturing geometric, textural, and color features, with Principal Component Analysis (PCA) used for feature reduction. Classification is conducted using support vector machines (SVM) and k-nearest neighbors (KNN), achieving high accuracy rates. The performance metrics reveal that the neural network model achieves an accuracy of 99.74%, demonstrating superior performance in terms of sensitivity, specificity, and precision compared to the decision tree model. However, the study highlights the need for larger datasets and suggests future work on noise reduction and fully automated systems to further enhance the model’s robustness and accuracy.\n\nDespite its strengths, the study identifies some limitations. The limited dataset size may affect the generalizability of the results, and noise reduction methods were not fully explored. Future research aims to develop a robust segmentation system independent of stains, expand the dataset to include more types of acute myeloid leukemia cells, and implement noise reduction methods such as median, mean, and Gaussian smoothing. Additionally, the authors propose enhancing deep learning models to learn from scratch with larger datasets, aiming to create practical, everyday diagnostic tools.35\n\nNiranjana Sampathila et al. (2022) present an advanced method for detecting acute lymphoblastic leukemia (ALL) using a customized convolutional neural network (CNN) named ALLNET. The study utilizes the C_NMC_2019 dataset, containing 10,661 images, with 7272 images of blast cells and 3389 images of healthy cells. The methodology involves extensive preprocessing, including color space conversion, histogram equalization, and thresholding to enhance image quality and facilitate segmentation. Data augmentation techniques such as vertical and horizontal flipping, random rotation, brightness adjustments, and Gaussian blur are employed to increase the dataset’s size and robustness.\n\nFeature extraction and classification are performed using the customized ALLNET architecture, which consists of four convolutional layers, four max-pooling layers, and three fully connected layers, with batch normalization and dropout layers to prevent overfitting. Training was conducted on Google Collaboratory using the Nvidia Tesla P-100 GPU, achieving a maximum accuracy of 95.54%, specificity of 95.81%, sensitivity of 95.91%, F1-score of 95.43%, and precision of 96%. This study underscores the importance of using deep learning models in medical diagnostics to enhance the accuracy and efficiency of leukemia detection. The methodology includes several key steps. Preprocessing involves converting images to the HSI color space for better contrast, applying histogram equalization, and using thresholding to convert images to binary form for segmentation. Noise reduction techniques, including Gaussian blur, are also used. Data augmentation further improves robustness with techniques such as vertical and horizontal flipping, random rotation, brightness adjustments, and Gaussian blur. Feature extraction uses the ALLNET architecture, which includes four convolutional layers with ReLU activation, four max-pooling layers to reduce dimensionality, three fully connected layers with batch normalization and dropout to prevent overfitting, and batch normalization to stabilize and accelerate training. The classification process employs the customized CNN (ALLNET), using the Adam optimizer and categorical cross-entropy loss function. Performance metrics highlight the model’s effectiveness with an accuracy of 95.54%, precision of 96%, sensitivity of 95.91%, specificity of 95.81%, and an F1-score of 95.43%. These results demonstrate the high classification accuracy and robustness of the ALLNET model in detecting leukemia. Despite the promising outcomes, the study identifies some limitations and future directions. The model’s performance depends on high-quality, well-annotated datasets, and it requires extensive preprocessing. Future research aims to enhance the ALLNET model by expanding the dataset to include more diverse and noisy images, reducing preprocessing steps, and integrating more advanced neural network architectures like YOLOv4, ResNet, and AlexNet. These improvements aim to develop a robust and reliable diagnostic tool for clinical use, enhancing the accuracy and efficiency of leukemia detection and reducing the time and error associated with manual diagnostics.36\n\nPałczyński et al. (2021) developed a hybrid artificial intelligence system for classifying Acute Lymphoblastic Leukemia (ALL) using an optimized, IoT-friendly neural network architecture. This system integrates transfer learning with MobileNet v2 as the encoder and machine learning algorithms like XGBoost, Random Forest, and Decision Tree as classifiers. Utilizing the ALL-IDB2 dataset, which consists of segmented images of lymphocyte cells from healthy patients and patients with ALL, the methodology involves preprocessing the images using data augmentation techniques, encoding them with MobileNet v2, and classifying the feature vectors with the machine learning models. The results demonstrate an average accuracy of over 90%, reaching up to 97.4%, showcasing the effectiveness of hybrid AI systems in handling tasks with low computational complexity. The methodology includes several steps, starting with preprocessing, where RGB images are converted to grayscale, contrast is enhanced using adaptive histogram equalization, and background noise is removed using area and closing operations. Data augmentation techniques such as color jitter, Gaussian blur, horizontal flip, vertical flip, and rotation are applied, followed by Z-score normalization. For feature extraction, MobileNet v2 is utilized, optimized for small processing units like mobile CPUs or IoT devices. The extracted features are then fed into classifiers such as XGBoost, Random Forest, and Decision Tree, trained using Adam optimizer with early stopping after 1000 epochs. The system achieved a classification accuracy of 97.4% when using MobileNet v2 with a fully connected layer, and over 90% accuracy for most models. Performance metrics including precision, recall, specificity, and AUC values were high, indicating robust model performance. The study highlights the system’s strengths in achieving high classification accuracy, efficient preprocessing, and effective use of transfer learning to prevent overfitting, making it suitable for low-resource environments like IoT. Despite these strengths, the study acknowledges some limitations and future directions. The system’s performance depends on high-quality, well-annotated datasets, and it may be sensitive to variations in image quality and staining. The Decision Tree algorithm showed lower performance compared to XGBoost and Random Forest. Future research aims to further optimize the hybrid AI system to enhance its robustness and accuracy, extend its application to other hematologic conditions, and refine the machine learning models. The authors suggest integrating this automated system into clinical workflows, particularly in low-resource environments, to aid in early and accurate leukemia diagnosis. By leveraging transfer learning and IoT-friendly architectures, the system aims to enhance the efficiency and reliability of medical diagnostics.37\n\nAlexandra Bodzas et al. propose an automated method for detecting acute lymphoblastic leukemia (ALL) from microscopic blood smear images, leveraging image processing techniques and machine learning models.\n\nThe dataset used in their study consists of 18 images of normal blood smears and 13 images from patients diagnosed with ALL, all collected from the University Hospital of Ostrava. Each image has a resolution of 4,080 × 3,072 pixels, capturing diverse regions of the blood smear to ensure comprehensive representation of leukocytes.\n\nThe images underwent preprocessing to correct variations in lighting and staining inconsistencies, followed by segmentation through a three-phase filtering technique to separate leukocytes from the rest of the image. Sixteen morphological and statistical features, mimicking the criteria used by hematology experts, were extracted from the segmented cells. These features were then used to train both a Support Vector Machine (SVM) and an Artificial Neural Network (ANN). While the results were promising, with the SVM achieving 96.72% accuracy and the ANN 97.52%, the limited dataset raises concerns about potential overfitting and the generalizability of the models. Additionally, the method is focused exclusively on the detection of ALL, limiting its applicability to other subtypes of leukemia.38\n\nNimesh Patel and Ashutosh Mishra proposed an automated method for leukemia detection from microscopic images in their 2015 study. Their approach focuses on extracting textural and morphological features from cells using Gray Level Co-occurrence Matrices (GLCM) along with cell shape analysis. These extracted features are classified using a binary Support Vector Machine (SVM) classifier, enabling the differentiation between leukemic and normal cells with a reported accuracy of 88.24%, based on blood smear images from leukemia patients. The dataset used in the study consists of 100 images from leukemia patients and 100 images from healthy individuals. However, several limitations are associated with this method. The relatively small dataset (only 200 images) raises concerns about the model’s ability to generalize its results to more diverse datasets. Additionally, the approach is limited to general leukemia detection, without addressing specific subtypes such as Acute Lymphoblastic Leukemia (ALL) or Acute Myeloid Leukemia (AML). Another challenge lies in the reliance on GLCM-based feature extraction, which depends on accurate cell segmentation. Achieving precise segmentation can be difficult with variable-quality images, increasing the risk of classification errors.39\n\nFollowing the discussion of the various studies, below is Table 5, a summarized comparative chart on these studies alongside their methods.\n\n\n7. Discussion\n\nThis article delves into the epidemiology and high prevalence of leukemia, highlighting the significant global research efforts focused on its early diagnosis and treatment. The critical need for advancements in this area is emphasized by the widespread participation of nations worldwide, each contributing to the evolving landscape of leukemia research. The integration of artificial intelligence (AI) into diagnostic workflows offers promising opportunities to enhance the detection and classification of leukemia, especially given the complexities associated with its primary subtypes—acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), and chronic myeloid leukemia (CML). However, current diagnostic methods and AI-based approaches face several limitations that must be addressed to realize their full potential.\n\nA comprehensive computational pipeline for leukemia detection generally involves three essential phases: preprocessing, feature extraction, and classification. Preprocessing ensures the data is optimized for analysis by improving image quality through noise reduction, data augmentation, and color correction. This step is essentiel, as variations in sample preparation and staining techniques can introduce inconsistencies that affect diagnostic accuracy. The feature extraction stage identifies key cellular traits such as morphology, texture, and size critical for differentiating between normal and malignant cells. In the final phase, machine learning algorithms and deep learning models, including convolutional neural networks (CNNs), support vector machines (SVMs), and ensemble classifiers, analyze these features to deliver accurate diagnostic outcomes.\n\nDespite these advancements, AI models in leukemia diagnostics encounter significant challenges. These include overfitting, where models trained on limited datasets struggle to generalize to unseen data, and variability in clinical images, which affects performance. Current diagnostic approaches also depend heavily on high-quality, well-annotated datasets, but access to such data is often limited, and inconsistencies across datasets can impair model reliability. Additionally, most AI models lack the scalability to perform equally well across diverse healthcare settings, as differences in equipment, patient demographics, and clinical protocols can impact outcomes. Addressing these challenges requires robust international collaboration, development of diverse multi-institutional datasets, and implementation of federated learning approaches to maintain data privacy while training models across institutions. Overcoming these limitations will be essential for creating AI tools that not only enhance diagnostic precision but also support personalized treatment planning, ultimately reducing the global burden of leukemia and improving patient outcomes.\n\n\n8. Limitations\n\nAI-powered models have advanced leukemia detection and classification significantly, yet several limitations must be addressed for reliable performance in clinical settings. A key factor influencing model robustness and accuracy is data quality and diversity. Many studies emphasize that high-quality, diverse datasets are essential to counteract variability across different clinical environments. When models are trained on homogeneous, limited datasets, they struggle to generalize well, especially across institutions that use different imaging equipment or patient demographics. This dependency on well-annotated data and the variability of samples across platforms introduce performance inconsistencies, underscoring the need for extensive and varied datasets to reduce overfitting and improve robustness.\n\nOverfitting risks are particularly prominent in models that rely on deep learning, such as convolutional neural networks (CNNs). Despite the use of techniques like dropout layers and normalization, CNN models can tend to memorize training data rather than identify generalizable patterns. This issue is exacerbated when models are trained on data with limited diversity, impacting their effectiveness on new, unseen data. Addressing overfitting requires a combination of data augmentation, transfer learning, and regularization techniques to enhance model adaptability.\n\nAnother critical challenge is dataset size constraints. Limited datasets restrict a model’s ability to capture the full range of leukemia variations, impacting classification accuracy. Smaller datasets are also more prone to overfitting, as the model may “memorize” specific details of the training data instead of learning generalizable features. Without access to larger datasets, models may deliver inconsistent results, particularly when applied to data from different patient populations or clinical environments.\n\nSensitivity to image quality and staining variations presents another challenge, as models can be highly sensitive to inconsistencies in sample preparation. Variations in staining techniques or image quality can affect classification accuracy, and models trained on high-quality samples may struggle to perform in less controlled environments. This sensitivity limits the practical application of AI models in resource-limited settings where high-quality imaging and staining facilities may not be available. Furthermore, automation and intermediate stage classification are areas that require improvement for AI models to be seamlessly integrated into clinical workflows. Challenges in achieving fully automated, robust classification are particularly evident when classifying subtle differences in cell morphology. Models that can accurately distinguish intermediate cell stages are vital, especially for personalized diagnostics, where precise classification is critical for treatment planning. Thus, achieving automation that can handle these complexities is essential for clinical applicability.\n\nNoise reduction and segmentation sensitivity are also important for consistent model performance. Effective noise reduction is essential, as noisy or low-quality images can hinder model accuracy. Segmentation-based approaches, which involve isolating cell features, are especially sensitive to variations in image quality, making them prone to error if the images do not meet certain standards. Optimization of noise reduction methods could therefore improve model reliability.\n\nFinally, broad applicability and real-world implementation remain significant challenges. Models that perform well in research environments may underperform in real-world clinical applications due to differences in clinical workflows, patient demographics, and medical equipment. This dependency on high-quality, well-annotated data makes it difficult for models to adapt to broader clinical settings without compromising accuracy. To make AI models truly viable for clinical use, they must be tested and validated extensively across multiple institutions. Federated learning, which allows models to be trained on data from various sources while preserving privacy, offers a promising approach to enhancing scalability and adaptability.\n\nOverall, addressing these limitations is essential to ensure that AI-driven leukemia diagnostics are reliable, scalable, and adaptable for real-world clinical environments. With improvements in dataset quality and diversity, robust techniques to mitigate overfitting, and optimized workflows for automation, these tools could substantially enhance leukemia diagnostics and patient care.\n\nTable 6 summarizes the main limitations identified in leukemia-related studies.\n\n\n9. Conclusion\n\nThe integration of artificial intelligence (AI) and advanced image processing techniques has revolutionized the field of leukemia diagnostics, enabling more accurate, rapid, and comprehensive detection of different leukemia subtypes. By automating key processes such as feature extraction, segmentation, and classification, AI-driven models address the inherent challenges of traditional diagnostic methods, improving both efficiency and diagnostic precision. Furthermore, understanding the global epidemiology and associated risk factors of leukemia is vital for the development of targeted screening programs and tailored treatment strategies. Future research should focus on addressing current limitations, such as ensuring high-quality datasets, mitigating overfitting risks, and enhancing the generalizability of AI models. Emphasis should also be placed on international collaboration, ensuring equitable access to advanced diagnostic technologies across regions to bridge healthcare disparities. Ongoing innovation in AI-based diagnostics will play a pivotal role in reducing the global burden of leukemia, fostering early detection, personalized treatment, and ultimately improving patient outcomes. Through focused efforts and interdisciplinary cooperation, the promise of AI in hematologic diagnostics can be fully realized, paving the way for enhanced healthcare delivery and better quality of life for patients worldwide.\n\nFuture research must focus on overcoming the limitations currently faced by AI-based leukemia diagnostics to develop more reliable, scalable, and generalizable systems. A key priority is the creation of diverse, multi-institutional datasets to address variability in image quality, staining techniques, and patient demographics, ensuring that AI models perform consistently across different clinical environments. Implementing strategies such as data augmentation, transfer learning, and regularization techniques will be essential to mitigate the risk of overfitting and enhance model generalization. Additionally, the integration of multi-modal data—combining genetic, clinical, and imaging information can improve diagnostic accuracy and enable personalized treatment strategies. Advancing federated learning frameworks will allow AI models to be trained across multiple institutions while maintaining patient privacy and data security.\n\nFuture systems should also emphasize model transparency and interpretability, ensuring that predictions are understandable and actionable by healthcare professionals to build trust and facilitate clinical adoption. Addressing the challenges of scalability is essential, particularly for deploying AI solutions in resource-limited settings, where computational efficiency must be balanced with diagnostic accuracy. Real-world testing and extensive clinical trials will be necessary to validate thes e models, refining them to accommodate variations in equipment, clinical protocols, and patient populations. International collaboration, supported by equitable access to advanced diagnostic technologies, will play a pivotal role in bridging healthcare disparities and ensuring that the benefits of AI-driven leukemia diagnostics are accessible globally. Through continuous innovation, interdisciplinary cooperation, and targeted efforts, future AI systems can revolutionize leukemia detection and treatment, ultimately improving patient outcomes and reducing the global burden of this hematologic malignancy.\n\nEthics and consent were not required.", "appendix": "Data availability\n\nZenodo: Metadata extracted from the Scopus database was used to analyze the countries, and keywords significantly contributing to leukemia research. The focus was on the integration of artificial intelligence in diagnostics, emphasizing global trends, risk factors, and classification strategies for improving detection and treatment: https://doi.org/10.5281/zenodo.14328973.40\n\nThe project contains the following underlying data:\n\n- AI in Leukemia Research.csv: Contains metadata extracted from the Scopus database, focusing on the integration of artificial intelligence in leukemia diagnostics, including global trends and classification strategies.\n\n- AI in Leukemia Research.txt: A textual description of the metadata and analysis approach for AI applications in leukemia research.\n\n- Leukemia Types.csv: Dataset describing various leukemia types with associated risk factors and global research statistics.\n\n- Risk Factors Keys.txt: A key file describing the variables and attributes in the “Risk Factors of Leukemia.csv” dataset.\n\n- Risk Factors of Leukemia.csv: Comprehensive dataset analyzing risk factors associated with leukemia.\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nThis study follows the PRISMA 2020 guidelines for systematic reviews to ensure clarity and reproducibility. A detailed PRISMA checklist outlining the methodology, including search strategies, inclusion criteria, and data synthesis, has been uploaded to Zenodo and is accessible at https://doi.org/10.5281/zenodo.14328973.40\n\n\nAcknowledgment\n\nWe would like to express our deepest gratitude to the Research Foundation for Development and Innovation in Science and Engineering (FRDISI). Their invaluable support and the conducive environment they provided have been instrumental in conducting this research.\n\n\nReferences\n\nCline MJ: The Molecular Basis of Leukemia. N. Engl. J. Med. févr. 1994; 330(5): 328–336. Publisher Full Text\n\nChow S, Buckstein R, Spaner DE: A link between hypercholesterolemia and chronic lymphocytic leukemia. Leuk. Lymphoma. avr. 2016; 57(4): 797–802. PubMed Abstract \| Publisher Full Text\n\nBouchbika Z, et al.: Cancer incidence in Morocco: report from Casablanca registry 2005-2007. Pan Afr. Med. J. 2013; 16: 31. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJoshi MD, Karode AH, Suralkar PSR: White Blood Cells Segmentation and Classification to Detect Acute Leukemia Ms.2013. Consulté le: 18 octobre 2024. [En ligne]. Reference Source\n\nElhassan TA, et al.: Classification of Atypical White Blood Cells in Acute Myeloid Leukemia Using a Two-Stage Hybrid Model Based on Deep Convolutional Autoencoder and Deep Convolutional Neural Network. Diagnostics. janv. 2023; 13(2): Art. no 2. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNegm A, Hassan O, Kandil A: A decision support system for Acute Leukaemia classification based on digital microscopic images. Alex. Eng. J. oct. 2017; 57: 2319–2332. Publisher Full Text\n\nSampathila N, et al.: Customized Deep Learning Classifier for Detection of Acute Lymphoblastic Leukemia Using Blood Smear Images. Healthcare. oct. 2022; 10(10): Art. no 10. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPałczyński K, Śmigiel S, Gackowska M, et al.: IoT Application of Transfer Learning in Hybrid Artificial Intelligence Systems for Acute Lymphoblastic Leukemia Classification. Sensors. déc. 2021; 21(23): 8025. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBodzas A, Kodytek P, Zidek J: Automated Detection of Acute Lymphoblastic Leukemia From Microscopic Images Based on Human Visual Perception. Front. Bioeng. Biotechnol. août 2020; 8. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPatel N, Mishra A: Automated Leukaemia Detection Using Microscopic Images. Procedia Comput. Sci. déc. 2015; 58: 635–642. Publisher Full Text\n\nAchir A: Advances in Leukemia Detection and Classification: A Systematic Review of AI and Image Processing Techniques. Zenodo. 27 novembre 2024. Publisher Full Text" }	[ { "id": "354217", "date": "14 Jan 2025", "name": "Huan Mo", "expertise": [ "Reviewer Expertise Hematology", "medical informatics." ], "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\nAs a hematopathologist, this review will be focusing on the the Section 3 (Classification of Leukemia).\n\nThe FAB morphological classifications used in this manuscript are very dated with limited therapeutic or prognostic values in the current standard of care. Please use the current WHO HAEM5 or ICC classification: WHO Myeloid: [Ref 1] WHO Lymphoid (For CLL and ALL): [Ref 2]\nICC Myeloid and acute leukemia (including AML, CML, and ALL): [Ref 3]\nICC Lymphoma (for CLL): [Ref 4]\nThe four disease groups selected for this manuscript were not a representative or related combination, although they were mostly-known diseases that bear the name \"leukemia\" (peripheral blood with or without marrow involvement), particularly for the two diseases with the \"chronic\" title.\nCLL (or CLL/SLL) is a mature B cell lymphoma commonly with leukemic states. Other mature B- and T-cell neoplasms commonly presenting with leukemic state also include mantle cell lymphoma, hairy cell leukemia, splenic marginal zone lymphomas, T-PLL, T- or NK-LGLL, and many other disease entities.\n\nCML is a myeloproliferative neoplasm (MPN). Other MPN (such as CNL), MPN/MDS (such as CMML and atypical CML), and myeloid/lymphoid neoplasms with eosinophilia and tyrosine kinase gene rearrangements can also present with leukemic state (or cytosis).\n\nFor ALL, the L3 now is mostly Burkitt lymphoma (a mature B cell lymphoma), which can present as circulating abnormal \"blast\"-like cells.\n\nFor sections 5.2 and 5.3, as well as Figure 7 and 8, it would be helpful if the authors can compare your model with the those in the professional guidelines, such as the NCCN guidelines (nccn.org, free registration needed to download).\n\nAlthough I am not an AI expert, the section 6 appears to be a laundry list of summaries of literature, without necessary integration. Instead, the authors should propose a list of items and use the literature to substantiate.\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? 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? Partly", "responses": [] } ]	1	https://f1000research.com/articles/13-1536
https://f1000research.com/articles/13-336/v1	23 Apr 24	{ "type": "Case Study", "title": "Data-Driven Decision Support Tool Co-Development with a Primary Health Care Practice Based Learning Network", "authors": [ "Jacqueline Kueper", "Jennifer Rayner", "Sara Bhatti", "Kelly Angevaare", "Sandra Fitzpatrick", "Paulino Lucamba", "Eric Sutherland", "Daniel Lizotte", "Jennifer Rayner", "Sara Bhatti", "Kelly Angevaare", "Sandra Fitzpatrick", "Paulino Lucamba", "Eric Sutherland", "Daniel Lizotte" ], "abstract": "Background The Alliance for Healthier Communities is a learning health system that supports Community Health Centres (CHCs) across Ontario, Canada to provide team-based primary health care to people who otherwise experience barriers to care. This case study describes the ongoing process and lessons learned from the first Alliance for Healthier Communities’ Practice Based Learning Network (PBLN) data-driven decision support tool co-development project.\n\nMethods We employ an iterative approach to problem identification and methods development for the decision support tool, moving between discussion sessions and case studies with CHC electronic health record (EHR) data. We summarize our work to date in terms of six stages: population-level descriptive-exploratory study, PBLN team engagement, decision support tool problem selection, sandbox case study 1: individual-level risk predictions, sandbox case study 2: population-level planning predictions, project recap and next steps decision.\n\nResults The population-level study provided an initial point of engagement to consider how clients are (not) represented in EHR data and to inform problem selection and methodological decisions thereafter. We identified three initial meaningful types of decision support, with target application areas: risk prediction/screening, triaging specialized program referrals, and identifying care access needs. Based on feasibility and expected impact, we started with the goal to support earlier identification of mental health decline after diabetes diagnosis. As discussions deepened around clinical use cases associated with example prediction task set ups, the target problem evolved towards supporting the upstream task of organizational planning and advocacy for adequate mental health care service capacity to meet incoming needs.\n\nConclusions This case study contributes towards a tool to support diabetes and mental health care, as well as lays groundwork for future CHC EHR-based decision support tool initiatives. We share lessons learned and reflections from our process that other primary health care organizations may use to inform their own co-development initiatives.", "keywords": [ "Primary Health Care", "Decision Support Tool", "Artificial Intelligence", "Machine Learning", "Diabetes", "Mental Health", "Descriptive Epidemiology", "Electronic Health Records" ], "content": "Introduction\n\nIncreasing amounts of everyday data coupled with advancements in technology and artificial intelligence (AI) are transforming healthcare.1–5 Primary health care settings have received less attention than other sectors, and there is a need for increased engagement of end-users in development of AI-enabled decision support tools.6,7 This case study describes the process and lessons learned thus far in co-developing a decision support tool with and for a primary health care organization in Ontario, Canada.\n\nThe Alliance for Healthier Communities (Alliance) supports team-based primary health care through 72 Community Health Centres (CHCs) across Ontario, Canada to people who otherwise experience barriers to care. In 2000, all CHCs moved towards a common electronic health record (EHR; Telus Practice Solutions, version code 5.23.100) with standardized data requirements; each client’s EHR has structured fields for sociodemographic characteristics (e.g., sex, gender, education) and dynamic care encounter tables (e.g., ENCODE-FM v7.2024 or ICD-10 codes to indicate diagnoses and procedures) that capture information from all providers in their care team. All CHCs follow a standardized opt-out consent process for the use of de-identified data in research that reports aggregate results, which the analyses in this report fall under. De-identified data with consent for research use were stored on a secure server, with access through encrypted channels granted only as needed to members of the research team. The project was approved by the Western University Health Science Research Ethics Board in March 2018 (ID 111353).\n\nFigure 1 summarizes our work completed thus far, with additional details for the six major stages of work presented below in terms of goals and associated activities. The discussion section summarizes overarching lessons learned and reflections.\n\nFirst six stages of decision support tool co-development project. Legend: Alliance = Alliance for Healthier Communities, CHC = Community Health Centre, EHR = Electronic Health Record.\n\n\nStages of work\n\nThe Alliance has been heavily involved in setting quality EHR standards (including the collection and use of sociodemographic and race-based data) and in research studies, but this was their first EHR data-driven decision support tool co-development project. Thus, we started with a strategy to better understand how client characteristics and care patterns are represented in EHR data, and to motivate and facilitate initial brainstorming around meaningful challenges amenable to support with EHR-based data analysis tools.\n\nGoals\n\n1. To summarize sociodemographic, clinical, and health care use characteristics of ongoing primary care clients served through CHCs across Ontario from 2009 through 2019.\n\n2. To serve as a foundation for community engagement and to inform decision support tool problem selection and methodological decisions.\n\nActivities\n\nConduct study: This study (published elsewhere8) provided an overview of population health and care patterns by applying to EHR data both methods from traditional descriptive epidemiology (e.g., period prevalence of chronic conditions) and from unsupervised machine learning to explore more complex patterns (e.g., non-negative matrix factorization to examine care provider teams). Findings were shared with the Alliance community through a Lunch ‘n’ Learn Webinar in October 2022, and were revisited throughout the decision support tool project to inform problem selection and methodological decisions.\n\nGoals\n\n1. To engage the broader Alliance community in critical thinking and discussion around secondary uses of EHR data.\n\n2. To invite participation in the decision support tool co-development project.\n\nActivities\n\nPopulation data assessment: The Lunch ‘n’ Learn included an introduction to AI and decision support tools in addition to a summary of Stage 1 study findings. Embedded throughout the latter portion were polls and discussion points asking whether findings were consistent with expectations. Areas of inconsistency motivate further exploration to discern if the mismatch is an artifact due to data quality issues or methodological decisions, or if the mismatch is clinically relevant. Areas of potential concern (e.g., accurately high prevalence estimate) could be a good target for a decision support tool.\n\nPBLN team formation: The Lunch ‘n’ Learn was the launching point of the decision support tool project, with additional invitations to participate distributed through the general Alliance community e-mail list and the recently formed PBLN member e-mail list. The resulting core team of people involved in remaining stages of work, described by general roles:\n\n• Alliance research leaders (Director of Research, Research and Evaluation Project Lead): Provide input towards project process and all content/decisions, as well as coordinate engagement with the PBLN.\n\n• PBLN members (care providers, clinical staff, IS staff): Engage in critical discussion around the target problem and broader decision support tool initiatives, provide input towards methodological decisions, and review analysis findings.\n\n• External researchers (professor, postdoctoral associate): Facilitate discussion sessions, lead analyses, and summarize findings.\n\nGoal\n\n1. To identify meaningful challenges within CHCs that are amenable to support with data-driven decision support tools.\n\nActivities\n\nPBLN meeting: In October 2022, the first PBLN meeting was held. We briefly reviewed descriptive epidemiology findings and discussed whether any outstanding questions needed to be answered to guide future steps. We then discussed data-driven decision support tools and brainstormed potential directions to pursue in the CHC context.\n\nDiscussion synthesis: Ideas were summarized into three “types” of decision support—risk prediction or screening, triaging specialized program entry, and identifying care access needs—with target conditions or application areas for each type (Table 1).\n\nPBLN meeting: In December 2022, we reviewed the three candidate project directions and while all (and more!) were seen as potentially valuable we decided to focus on one while exploring how to best conduct this type of project: risk prediction for mental health decline after diabetes diagnosis. Rationale included i) expected impact (e.g., high prevalence of diabetes with known mental health comorbidities, coupled with the challenge of needing to choose between many possible care options upon a new diabetes diagnosis); ii) actionable (e.g., all CHCs provide mental health care resources that could help prevent mental health decline for people living with diabetes); and iii) feasibility (e.g., relevant care captured in the EHR, and heavy focus on risk prediction in machine learning methods advancements).\n\nFollow-up action: We connected with Diabetes Action Canada to learn more about related work and potential collaborators. Given the early nature and CHC-specific focus of our project, we proceeded with risk prediction model development using an existing 11-year retrospective extract of CHC EHR data, with the intention to consider expansion or tighter external collaboration after more internal feasibility and impact assessments.\n\nGoal\n\n1. To gauge feasibility and deepen discussion around developing a decision support tool that predicts early mental health decline within a year of incident diabetes indication.\n\nActivities\n\nPreliminary analysis: Candidate cohort summary characteristics, and an outline of potential predictor and outcome definitions and data sources.\n\nPBLN meeting: In February 2023, we met to review the preliminary analysis and made initial decisions on the cohort and how to operationalize the outcome and predictors. This was done alongside further problem refinement and discussing clinical actions that could accompany a high-risk indication (Examples: brief educational discussion, noting down mental health to discuss in future appointments, referral to a specialized mental health provider, or referral to a CHC group program focused on mental health and/or diabetes).\n\nModel development: The eligible cohort included 1,250 adult ongoing primary care clients receiving care at an East Toronto region CHC, who had at least one diabetes ICD-10 code9 in 2011-2018, at least one year of follow-up care, and no mental health care or decline indication in the two years prior to their incident diabetes indication. Five candidate models ranging in complexity from simple linear (sklearn.linear_model.LogisticRegression v0.24.2) to complex machine learning techniques (CatBoost v0.26.1) were trained and compared using a five-fold nested cross validation procedure. Hyperparameters were selected on the inner loop using a grid search for the highest Area Under the Receiver Operating Characteristic Curve (AUROC).\n\nModel performance: Table 2 presents summary performance metrics, with additional methods and results details available upon request.\n\nPBLN meeting: In April 2023, we met to review and revise the initial model development analyses. While we decided the predictive performance of the sandbox model was not clinically useful, the associated discussion surfaced ideas around how to better harness information from the data and around problem refinement. Example insights included broadening eligibility from incident to prevalent cases of diabetes, modifying how to identify active diabetes care, and surfacing the need to engage more people to better understand how different provider types code mental health care in the EHR (e.g., do ENCODE-FM codes for “feeling anxious” vs “anxiety” reliably distinguish symptoms vs. diagnosis or is this more indicative of different provider type scopes). In terms of features, medication and lab value data are expected to increase accuracy of individual-level predictions; these data are planned for integration with the BIRT system (supporting data access), but not yet readily available.\n\nFollow-up actions: We sought further input on the case study and suggested next steps in three ways: a PBLN member lead a focus group at their CHC, a poster presentation was given at a primary care research gathering, and an email call out for further input was circulated through the Alliance email listserv.\n\nProblem refinement: A strong discussion theme was that while all CHCs provide mental health services, these are already at or near capacity and implementation of the decision support tool may increase demand past a point that could be maintained. This highlighted potential value of instead developing a system-level decision support tool to address the upstream problem of how to plan or advocate for adequate capacity within CHCs to address future mental health care service needs.\n\nGoal\n\n1. To use what was learned in Stage 4 to develop a sandbox model to predict the number of ongoing primary care clients with prevalent diabetes indications who will have mental health care needs in the upcoming year.\n\nActivities\n\nModel development: Example methodology changes from the Stage 4 case study were loosening eligibility criteria to include clients already receiving mental health care, broadening the outcome to include additional ENOCDE-FM codes (categories: emotional symptoms, symptoms involving appearance, suicidal ideation, affective disorder, and anxiety), and changing the missingness strategy for sociodemographic variables (collapsed not asked with no answer). We performed a similar five-fold nested CV procedure but restricted to feature-based models due to discussions (supported by case study 1 results) that preprocessed counts of chronic conditions should be more informative than granular codes.\n\nModel performance: Of the 20,329 eligible clients with prevalent diabetes in 2016-2018, 22.2% had a mental health care outcome recorded in 2019. We used a naïve 0.5 probability cut-off for the best performing model (CatBoost v0.26.1) across all outer test folds to demonstrate the type of information that could be made available from this type of model to support capacity-planning decisions or advocacy. Overall model accuracy was 86%; CHC-specific accuracy ranged from 64% to 97%, plus one CHC where there were no predicted or actual outcome cases (100% accuracy). The CHC-specific proportion of clients with diabetes predicted to have mental health care needs ranged from 46% (vs. actual 48%) to 1% (vs. actual 13%). Calibration performance is in Figure 2, with additional metrics available upon request.\n\nLegend: LR = Logistic Regression, LRP = LR with L1 penalty, CATB = CatBoost, F# = Fold number in cross validation procedure.\n\nFollow-up actions: We presented on all stages of the project at the annual Alliance Conference,10 and discussed the second case study at a PBLN meeting in July 2023. The sandbox model predictive performance showed promise and discussions further supported the idea that this type of advocacy and capacity planning tool would be a beneficial precursor to the individual-level tool in terms of actionability and associated clinical or system utility.\n\nGoal\n\n1. To review progress thus far and select which project direction to pursue next.\n\nActivities\n\nPBLN meeting: In addition to discussing case study 2 results, the July 2023 PBLN meeting included a review of project progress summarized into three points (Figure 3)—project scoping and problem identification, case study 1, and case study 2—alongside potential next steps with expected resource needs.\n\nLegend: Alliance = Alliance for Healthier Communities, CHC = Community Health Centre.\n\nNext steps: Given the novelty of this project within the Alliance and the broader field of AI-enabled technologies for primary health care, the technical and process related progress made thus far towards a fully functional tool to support diabetes and mental health care additionally includes lessons that would benefit future projects of different focus. Therefore, we decided to pause to document our processes before revisiting the population-level planning tool direction.\n\n\nDiscussion\n\nOur decision support tool co-development approach with and for the Alliance started with a large-scale epidemiological study to learn about health and care patterns of the population of interest and about how clients are represented in CHC EHR data. We identified a priority problem of supporting proactive mental health care for clients with diabetes, and iterated between sandbox case studies and discussion sessions to further understand the problem, how a data-driven decision support tool could provide support, and the feasibility of achieving a high-quality technical solution given readily available EHR data.\n\n\n\n• Epidemiology as a foundation for innovation: We used epidemiology as a launching point to supplement clinical and organizational expertise in brainstorming meaningful problem selection. Example benefits: 1) provided an overview of population health and care patterns (e.g., diabetes and mental health prevalence estimates informed expected impact assessment), 2) supported understanding around data quality and how clients are represented in aggregate EHR data (e.g., reliably recorded data elements as part of feasibility assessment), and 3) informed methodological decisions during model development (e.g., the first year of care at a CHC has a distinct profile wherein incidence and prevalence are hard to parse). Furthermore, this rigorous population-level overview8 can inform multiple initiatives outside the current project scope, and provides a baseline for longer-term evaluation or monitoring of the impact of EHR-based tools over time and with continued use.\n\n• Importance of an interdisciplinary team: Our team included clinical, research, technical, IS, and organizational leadership expertise. This allowed for seeding discussions with examples, critically assessing meaningful or realistic use of candidate tool ideas in practice and in the context of the organization—starting with direct envisioned uses (e.g., brainstorm clinical actions in response to X type of alert) and extending into how to follow through on those actions amidst competing demands at an individual client care level and at a system capacity and culture level. For example, moving from the problem of how to support individual level mental health and diabetes care to address the more pressing challenge of system-level advocacy for adequate resources to be able to follow-through on best care options that would otherwise result from the original tool idea. Including different perspectives further helped to scope the project in terms of what data are available and when, identify the best methodology to meet clinical goals, connect with internal and external collaborators, and support feasibility and continuity from a system-capacity perspective.\n\n• Sandbox case studies supported deeper discussion sessions: The impact of the sandbox case studies can be likened to how editing a manuscript often matures or builds on ideas relative to when writing the first draft. They were particularly valuable given the novelty of this type of project within the Alliance and the limited amount of research literature on co-development of AI-enabled decision support tools for primary health care settings. Even when predictive performance of a sandbox model was poor, the tangible example pushed discussions further than hypothetical scenarios or thought experiments could. Discussing why specific technical decisions did or did not line up with realistic or impactful clinical scenarios improved problem conceptualization, and the resulting methodological decisions can be applied to future projects. Of note, development of the “real” model intended for deployment will use a new data extract with more recent data.\n\n• Problem scoping around data availability: CHCs record rich EHR data that capture information about multiple domains of health; however, the target problem and methodology needs to consider what data are available and to what extent for the population and intended implementation sites of interest. For example, completeness of sociodemographic characteristics varies across CHCs and certain data types (e.g., medications, lab tests) flagged as potentially valuable for our first sandbox case study were scheduled to become available about a year out, which influenced our problem refinement.\n\n• Multiple engagement strategies are needed: The initial Lunch ‘n’ Learn session coupled with email list invitations was effective for forming the core project team. In seeking further input towards the project, a PBLN member led focus group at their CHC was more effective than research poster presentations or further e-mail-based recruitment.\n\n• Working towards a broader decision support tool initiative: As experience is gained, tool development will become more efficient and effective. While there are hundreds of potential targets for decision support tools within the broad scope of primary health care, it will not be sustainable or beneficial to continue to create or implement these independently. A fragmented approach to tool integration poses risks such as exacerbating alert fatigue or disrupting instead of augmenting team-based, whole-person care. Rather, our broader vision is to support more seamless integration of multiple types of tools to benefit people (e.g., better health outcomes), providers (e.g., improved workflow and decisions support), and communities (e.g., sufficient resources to meet demands). An additional consideration for expansion will be when to maintain a tool consistently across all CHCs versus adapt it to the local CHC context and available data.\n\nThis project more generally furthers the Alliance’s learning health system work at three levels: data analysis capacity, stakeholder engagement, and process refinement.11–15 First, demonstrating the use of AI to make data more meaningful through large-scale descriptive and real-time, clinically relevant predictive insights that will ultimately improve care delivery. Second, providing new avenues for clinician and provider engagement in data-driven learning initiatives; future work will additionally be able to engage the newly formed Client and Community Research Partners Program. Third, by providing a baseline process for tool development that future projects can learn from and build upon (i.e., what stages of work to keep, modify, or replace). Each future decision support tool initiative will provide additional opportunities for bidirectional learning whereby data are harnessed through AI to tackle a specific clinical problem and improve care delivery, while simultaneously learning how to improve and adapt the processes to achieve that clinical goal for different types of decision support and application areas.\n\n\nConclusions\n\nThis case study describes the ongoing process and lessons learned thus far in the first EHR-based decision support tool co-development project with and for CHCs in Ontario, Canada. The current focus is on diabetes and mental health care, with a vision of extending into a larger and longer-term decision support tool initiative that would integrate multiple types of tools with the CHC EHR system. Our processes and reflections may further inform or motivate other primary health care organizations at a similar stage of learning how to best harness value from EHR data.\n\nConsent for use of de-identified data for research purposes, whereby results are only made available in aggregate, was collected through a standardized opt-out procedure followed by the Community Health Centres where care was provided. Consent is collected via a written form, which is then scanned into the EMR. The need for additional consent specific to the research in this report was waived by the Western University Health Science Research Ethics board (ID 111353).", "appendix": "Data availability\n\nThis project includes the following underlying data: 11 years of electronic health records for adult clients who had at least one encounter at an Ontario Community Health Centre in 2009-2019. The data extract includes a client characteristics table (e.g., date of birth, level of education completed) and dimension and fact tables including details about care encounters over time (e.g., diagnostic codes, providers involved, referrals). Due to their sensitive nature, the data are securely stored and not shareable for ethical reasons, as outlined by the Western University Research Ethics Board (ID11353). Inquiries about access for future research studies should be directed to the Department of Research and Evaluation, Alliance for Healthier Communities (lhs@allianceon.org).\n\n\nAcknowledgements\n\nWe would like to thank members of the Alliance PBLN for their contributions to this project, whether for a single session, multiple sessions, or through to co-authorship of this report, and to thank Alliance staff members who helped support connections and ongoing engagement.\n\n\nReferences\n\nJiang F, Jiang Y, Zhi H, et al.: Artificial intelligence in healthcare: past, present and future. Stroke Vasc. Neurol. 2017; 2(4): 230–243. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLin SY, Mahoney MR, Sinsky CA: Ten ways artificial intelligence will transform primary care. J. Gen. Intern. Med. 2019; 34(8): 1626–1630. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGuo Y, Hao Z, Zhao S, et al.: Artificial intelligence in health care: Bibliometric analysis. J. Med. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKueper JK, Rayner J, Zwarenstein M, et al.: Describing a complex primary health care population to support future decision support initiatives. Int. J. Popul. Data Sci. 2022; 7(1): 21. Publisher Full Text\n\nFortin M, Almirall J, Nicholson K: Development of a research tool to document self-reported chronic conditions in primary care. J. Comorbidity. 2017; 7(1): 117–123. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBhatti S: Developing data-driven decision support tools with the Alliance Practice Based Learning Network. Presented at: Connected Communities: Building Equitable Integrated Healthcare Conference, Toronto, Ontario. 2023.\n\nNash DM, Brown JB, Thorpe C, et al.: The Alliance for Healthier Communities as a Learning Health System for primary care: A qualitative analysis in Ontario, Canada. J. Eval. Clin. Pract. 2022; 28: 1106–1112. Publisher Full Text\n\nNash DM, Rayner J, Bhatti S, et al.: The Alliance for Healthier Communities’ journey to a learning health system in primary care. Learn. Health Syst. 2021; 7: e10321. Publisher Full Text\n\nDelaney BC, Peterson KA, Speedie S, et al.: Envisioning a Learning Health Care System: The Electronic Primary Care Research Network, a case study. Ann. Fam. Med. 2012; 10(1): 54–59. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLindsell CJ, Gatto CL, Dear ML, et al.: Learning From What We Do, and Doing What We Learn: A Learning Health Care System in Action. Acad. Med. 2021; 96(9): 1291–1299. PubMed Abstract \| Publisher Full Text\n\nFoley T, Horwitz L, Zahran R: Realising the Potential of Learning Health Systems. The Learning Healthcare Project; 2021; 101. Accessed December 28, 2021.\n\nKueper JK, Rayner J, Lizotte DJ: Hybrid feature- and similarity-based models for joint prediction and interpretation. arXiv:2204.06076. February 11, 2023." }	[ { "id": "283425", "date": "31 May 2024", "name": "Ana De Marchi", "expertise": [ "Reviewer Expertise Digital health", "Participatory design", "Primary care" ], "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 study demonstrates the development and application of a data-driven decision support tool within a primary health care setting, employing a methodologically sound iterative process that is both reflective and inclusive of stakeholder engagement. The use of both traditional epidemiological methods and advanced machine learning techniques enriches the analytical depth and application potential of the decision support tool, offering a comprehensive approach to addressing primary health care challenges. The documentation of the co-development process provides a valuable blueprint for similar future initiatives in health informatics.\nThis study's dedication to including a wide range of stakeholders throughout the development process is one of its most noteworthy strengths. It guarantees that the tool is not only theoretically sound but also practically relevant and easy for clinical practitioners to utilize. This strategy increases the likelihood of acceptance and successful use in practical contexts, which is in line with the study's goals of enhancing primary healthcare delivery through cutting-edge technological innovations.\nHowever, as the authors mention that the project is ongoing, I suggest for the conclusion section (and by extension, the abstract) to provide more detail about the current status of the initiatives and planned future studies. This would offer a clearer understanding of the project’s trajectory and ensure that the study’s documentation keeps pace with its development, thereby reinforcing the relevance and timeliness of the research. This adjustment would help maintain a clear narrative through to the study's completion and beyond, as it progresses from methodology formulation to practical application.\nAs a reviewer, I would like to point out that given the case study format of this research, it does not delve into detailed descriptions of the Population-Level Planning Predictions models. These models are likely to be more comprehensively detailed in a separate, dedicated study. Therefore, a deeper evaluation of these models is not feasible within the scope of this case study.\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? Not 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\n\nIs the case presented with sufficient detail to be useful for teaching or other practitioners? Yes", "responses": [ { "c_id": "12964", "date": "19 Dec 2024", "name": "Jacqueline Kueper", "role": "Author Response", "response": "Dear Dr. Ana De Marchi and Dr. Ericles Bellei, Thank you for your interest and review of our work. It is helpful to see what stands out as strengths from your perspective, and we appreciate the suggestion to include additional details about the ongoing project. Accordingly, in a revised version we added a sentence to the conclusion about how we are refining and validating the population-level model with updated data, and that we have held additional focus groups within the health system to get further insights. These focus groups lead to an updated outcome definition that is better tailored to mental health capacity planning as well as a broadening of the cohort eligibility from ongoing primary care clients to also including diabetes education center clients who may or may not also be primary care clients." } ] }, { "id": "303167", "date": "16 Aug 2024", "name": "Nicole M White", "expertise": [ "Reviewer Expertise Biostatistics", "health 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 study reports on a multi-stage process for developing and refining a clinical decision-support tool using EHR primary care data. Six main stages are described, starting with an epidemiological study to motivate stakeholder discussions to identify areas that may benefit from decision support and tool development. Two case studies are presented, involving the use of different classification algorithms to predict mental health decline and population-level service demand, respectively.\nThe framework is well designed and described throughout the manuscript. Strengths of the framework are stakeholder involvement in both development and refinement stages, and the use of data and preliminary results to motivate these discussions.\nGiven the emphasis on end-user engagement, it would be beneficial to include some high-level data on engagement levels at different stages. This additional information could be described in the Stages of work section and/or Reflections and lessons learned section. For example, the authors provide information on which engagement strategies worked better than others – how many participants were recruited from the ‘best’ engagement strategy and were numbers above/below/in line with expectations?\nWhilst the model development itself is not the focus of this study, these sections would benefit from more information in the relevant stages of work (4, 5) and the Discussion. I noted that the metrics chosen to summarize model performance were not consistent between case studies. Making these metrics more consistent would strengthen the paper and reporting transparency. For model development in stage 4, some information on outcome prevalence would help to gauge the suitability of the stated sample size and its impact on model performance. Model developed information for Stage 5 would also benefit from clarifying the difference between the five-fold CV procedure and CHC-specific results. I interpreted the latter as being the results of internal-external model validation.\nIn the Discussion, the study alludes to future work once more data are available. Whilst having more recent data may contribute to building a better model, data quality is equally important. It would be useful to include some comments on data quality in both case studies for context, and whether quality was a barrier. This is particularly pertinent to EHR data streams known to be of variable quality.\nAs a minor comment, the terms ‘AI’ and ‘ML’ are used interchangeably throughout the manuscript. Whilst these disciplines are overlapping and complementary, using these terms interchangeably can make it difficult for the reader to understand what specific methods were used.\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? Partly\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\n\nIs the case presented with sufficient detail to be useful for teaching or other practitioners? Partly", "responses": [ { "c_id": "12965", "date": "19 Dec 2024", "name": "Jacqueline Kueper", "role": "Author Response", "response": "Dear Dr. Nicole M White, Thank you for your interest and review of our article. We appreciate your suggestions for improvement and have addressed them as summarized below, resulting in a more robust and transparent article. Regarding end-user engagement levels, unfortunately we did not collect any formal attendance records or statistics. We have however added to the ‘reflections and lessons learned’ section an approximate size of the team and more details about why the focus groups were considered successful. Regarding model development, we agree that additional details would be helpful and have broadened Table 2 to include performance metrics (AUROC, AUPRC) from both Stage 4 and Stage 5 so that they can be compared. We also followed your suggestion to add the outcome prevalence for the Stage 4 cohort (8.4%), and further clarified how the CHC-specific results were calculated (stratification by groups of clients belonging to each CHC, from the same data as for the overall metrics). Regarding data quality, we agree that data quality is as if not more important than recency. Working within limited space, we expanded the Reflections section on data availability to include both availability and quality, included some high level reflections from our experience that the relatively high quality of data from CHCs increased the feasibility of our study, and referenced a study that did a more in-depth assessment of data from CHCs as part of the organizations journey to being a learning health system. Finally, thank you for flagging the confusion between AI and machine learning throughout. We added to the opening sentence clarification that we refer to machine learning as a major subfield of AI and then updated remaining text about the stages and lessons learned to include machine learning as that is the family of techniques we used. One exception is we left AI in the final sentence of the lessons learned section as we wanted to capture the broader scope of methods that may be used in a learning health system and future projects, but added in machine learning to make it explicit we also mean to include the techniques described earlier." } ] } ]	1	https://f1000research.com/articles/13-336
https://f1000research.com/articles/13-1535/v1	19 Dec 24	{ "type": "Research Article", "title": "Internal Perspectives on Visual Identities in Higher Education: A Case Study of Top-Ranked Universities in Indonesia", "authors": [ "Putri Dwitasari", "Ellya Zulaikha", "Syarifa Hanoum", "Rabendra Yudistira Alamin", "Luqman Lee", "Ellya Zulaikha", "Syarifa Hanoum", "Rabendra Yudistira Alamin", "Luqman Lee" ], "abstract": "Background The neglect of visual identity (VI) at the organizational level within higher education institutions (HEIs) has become a critical issue, while previous studies over the past decade has focused on HEI branding and reputation. This creates a potential gap in understanding HEI branding processes. Thus, this study aims to explore the relationship between VI and HEI reputation by integrating the Expressiveness Quotient (EQ) and experiential brand meaning at the organizational level.\n\nMethod Using a qualitative case study approach, the study involves semi-structured interviews with 29 employees from five top-ranked universities in Indonesia. Furthermore, it analyzes the integration of experiential brand meaning across the stages of awareness, interpretation, appropriation, and communication, alongside the EQ framework to assess how visual identity impacts visibility, distinctiveness, transparency, authenticity, and consistency.\n\nResult The findings indicate that visual identity significantly influences perceptions of institutional identity and reputation. Effective management of visual identity elements enhances competitive advantage in academia and aligns internal stakeholder perceptions with external branding, which is essential for a cohesive organizational identity.\n\nConclusion This study emphasizes the strategic importance of visual identity in enhancing institutional reputation and provides a model for universities aiming to strengthen their reputational power through effective visual identity management. The study also reveals strong awareness and acceptance of brand identity.", "keywords": [ "Visual identity", "reputation", "internal stakeholder perception", "experiential brand meaning", "HEIs" ], "content": "1. Introduction\n\nHigher Education Institutions (HEIs) have increasingly focused on reputation and brand image to strengthen their competitiveness in university rankings, both nationally and internationally. This focus aids in differentiating HEIs from competitors to meet the need for articulating institutional essence within an increasingly competitive market (Balaji et al., 2016). Furthermore, reputation and brand image are acknowledged by HEIs as essential elements that can create distinct characteristics within higher education, supporting the commercialization of products (i.e., academic programs) and services (Clark et al., 2020; Mohamad et al., 2017). Additionally, reputation and brand image in higher education have been identified as critical areas in the context of future theory and practice (Clark et al., 2020; Melewar & Nguyen, 2014). Although HEIs exert efforts to build a reputation and brand image recognized for enhancing competitiveness, these efforts also require organizational activities to ensure that brand values projected to the market are upheld and conveyed by employees. Dean et al. (2016) explored how personnel collectively create brand meaning through their experiences and social interactions within the university context. In this case, the specific focus on internal branding, through strengthening identity, remains largely overlooked (Clark et al., 2020).\n\nThe foundational constructs of HEI branding, such as identity and image, and their strong interaction with reputation, remain underexplored at the organizational level (Gregersen & Johansen, 2022). In this context, Visual Identity (VI) plays a critical role in shaping a university’s reputation, serving as a tangible representation of the institution’s values, mission, and quality (Baker & Balmer, 1997; Hunter, 2020; Melewar et al., 2005; Mula-Falcón et al., 2024). Gregersen and Johansen (2022) highlighted in their study the oversight by HEIs in understanding reputation and branding, often neglecting VI, which may result in short-term performance. However, VI’s role in organizational communication is increasingly recognized, especially with the emergence of the “visual shift” in communication research, which emphasizes the importance of images and visual elements in capturing public attention (Verčič & Wisenberg, 2021; Crăciun, 2019). In conveying the unique character and values developed by HEIs, VI is directed toward various stakeholders, including media, academics, corporations, government, and alumni, while internal stakeholders include students, faculty, and staff (Dean et al., 2016; Melewar et al., 2005). Implicitly, Khan and Hemsley-Brown (2024) have investigated the impact of VI on course design values, a key factor influencing students’ expectations. In this context, VI and HEI reputation play essential roles in shaping students’ perceptions of the values offered by the institution. Therefore, through its graphical and symbolic elements, VI helps communicate the essence and values of higher education institutions to students, who, in turn, form perceptions of HEI quality (Coman et al., 2021; Dean et al., 2016; Spry et al., 2020).\n\nIn this vein, managing effective communication with both external and internal stakeholders is crucial to strengthening the brand image of HEIs. Brand identity reflects internal perception, while brand image reflects external viewpoints, highlighting the need for consistent internal branding to support external brand communication (Aaker, 2012; Clark et al., 2020). Meanwhile, HEIs emphasize the importance of aligning brand personality with organizational culture, particularly through internal communication that involves employees in maintaining unified branding efforts. Slade-Brooking (2016) highlights that a robust internal communication strategy not only articulates but also reinforces brand promises. Employees, as key internal stakeholders, shape public perception by fostering trust through an integrated understanding of brand identity. Research by Bolhuis et al. (2018) supports this, showing that employees’ identification with the organization strengthens when the visual identity aligns authentically with institutional values and culture, as also supported by other studies (Miotto et al., 2020; Wu and Cheong, 2024). This alignment, as noted by Frandsen and Huzzard (2021), helps to avoid potential dissonance within the academic environment that could hinder external brand formation.\n\nAdditionally, various factors such as organizational culture, developmental stage, resources, political dynamics, and student demographics further influence HEIs’ branding strategies (Asaad et al., 2013). HEIs in Indonesia, for instance, as part of a developing country, continue to create a dynamic institutional environment, where regulations play a role in shaping reputation management approaches. For example, specific regulations designate certain universities as “Legal Entity Universities” (Perguruan Tinggi Berbadan Hukum, or PTN-BH), focusing on maintaining and enhancing institutional reputation. There are 21 universities operating as PTN-BH in Indonesia, but only five PTN-BH universities rank highest in the Quacquarelli Symonds World University Rankings (QS WUR), Times Higher Education (THE), and Webometrics: Universitas Indonesia (UI), Universitas Gadjah Mada (UGM), Institut Teknologi Bandung (ITB), Universitas Airlangga (Unair), and Institut Teknologi Sepuluh Nopember (ITS). Dennis et al. (2016) explain that university rankings—including QS World University Rankings, Times Higher Education, academic rankings, and national rankings—are key metrics reflecting institutional reputation. These rankings have become a focal point for assessing quality and reputation, as noted by Dwitasari et al. (2022), emphasizing the competitive context in which universities strive to position themselves more favorably.\n\nThus, this study highlights the significance of HEI reputation and identifies a considerable gap specifically in addressing brand development and management related to visual identity and reputation, also underscored by Clark et al. (2020). Meanwhile, in existing research, there remains a lack of understanding of HEI branding flows, which primarily prioritize brand image and reputation (e.g., Del-Castillo-Feito et al., 2019; Lafuente-Ruiz-de-Sabando et al., 2018; Miotto et al., 2020), with VI often overlooked at the organizational level (Gregersen & Johansen, 2022). Therefore, this study aims to explore the level of knowledge, understanding, and employee engagement in recognizing the institution’s brand identity across five PTN-BH universities. The theoretical framework used is the Expressiveness Quotient (EQ) developed by Fombrun and Van Riel (2004), which evaluates institutional reputation through five dimensions: visibility, distinctiveness, transparency, authenticity, and consistency. These dimensions help organizations gauge emotional appeal and stakeholder communication effectiveness. Additionally, these five EQ dimensions are linked to how Corporate Visual Identity (CVI) supports reputation, using the theoretical framework by Van den Bosch et al. (2005), which asserts that well-managed CVI can enhance an institution’s visibility, distinctiveness, authenticity, transparency, and consistency.\n\n\n2. Literature review\n\nThe marketing and management literature has extensively explored the importance of the concept of corporate identity, referring to the features, characteristics, traits, or attributes of a company that are deemed essential, distinctive, and enduring (Clark et al., 2020; Coman et al., 2021; He & Mukherjee, 2009). Corporate identity is an organization’s presentation to each stakeholder, making the organization unique and combining aspects of communication, design, culture, behavior, structure, industry, and organizational strategy (Wu and Cheong, 2024). As marketing consensus emphasizes the inside-out approach (Clark et al., 2020), it is crucial for HEIs to understand the process of identity formation at the organizational level. Key stages in Visual Identity (VI) aim to integrate the institution’s values and vision into public and internal perceptions (Iglesias et al., 2020; Siyanbola & Adeyemi, 2021). Furthermore, organizational identity is shaped through a combination of internal branding and VI (Siyanbola & Adeyemi, 2021). Internal branding refers to an organization’s efforts to strengthen employees’ emotional attachment and understanding of its values (Balmer and Podnar, 2021; Barros-Arrieta and García-Cali, 2021). This critical process ensures that all HEI members, particularly educators and staff, comprehend and embody the institution’s core values in every interaction with students and other stakeholders (Clark et al., 2020). In other words, internal branding is the foundational basis of an HEI’s identity, which is then expressed through VI. Meanwhile, VI, encompassing elements such as logos, colors, and graphic symbols, plays a crucial role in conveying an authentic and consistent visual message of the HEI’s identity to both external and internal audiences (Wu and Cheong, 2024). Various studies have highlighted that this visual consistency is necessary to build trust and a strong reputation in the eyes of the public (Gregersen & Johansen, 2022). As an integrated element in organizational communication, VI becomes a tangible manifestation of the institution’s values and essence, with direct implications for the reputation of HEIs (Del-Castillo-Feito et al., 2019).\n\nHowever, previous studies on the formation of HEI identity have often overly emphasized aspects of brand image and reputation (e.g., Del-Castillo-Feito et al., 2019; Lafuente-Ruiz-de-Sabando et al., 2018; Miotto et al., 2020). An excessive focus on external perceptions can lead to the neglect of Visual Identity (VI) at the organizational level, as also highlighted by Gregersen and Johansen (2022). This poses a risk of weakening HEI branding overall, as VI is not merely a visual display but a reflection of the organization’s commitment to its values. Clark et al. (2020) show that VI is often considered secondary to other branding aspects, even though VI has a long-term impact on how HEIs are identified and recognized by the public. The oversight of VI in various studies indicates a deeper gap in understanding its role at the organizational level. Meanwhile, VI is not simply an external promotional tool but also an element that aligns internal identity with external perceptions (Ali et al., 2022). A representative VI fosters strong internal organizational attachment, increases loyalty, and builds internal cohesion, which has a significant impact on the overall reputation of HEIs (Ali et al., 2022; Mohamad et al., 2017).\n\nIn the modernization of higher education, institutional identity is shaped through four main structures: communication, behavior, organizational culture, and market conditions (Melewar & Jenkins, 2002; Troyan & Kravchenko, 2021). These structures work together to build the institution’s image and reputation, involving the perceptions of both internal and external stakeholders (Alessandri et al., 2006; Ribeiro et al., 2022). Visual Identity (VI), consisting of elements such as names, logos, color schemes, typography, and slogans, is a strategic asset that reflects the core values and character of the organization, while also strengthening recall and distinguishing the institution in the market (Van Den Bosch et al., 2005; Foroudi et al., 2017). The logo, as the primary expression of visual identity, is not only a differentiating tool but also a communication medium that represents the organization’s ideas and qualities (Van Riel & Van den Ban, 2001; Melewar & Akel, 2005). Institutional management uses the logo to support strategies, achieve goals, and articulate the competitive characteristics of the institution. In this context, the consistent management of VI can produce a cohesive and authentic identity narrative, enhancing the reputation and competitiveness of the institution in the increasingly competitive higher education sector (Gregersen & Johansen, 2022). Therefore, the integration of VI into the internal branding of HEIs is not merely cosmetic but a strategic element that strengthens identity and relationships with stakeholders, creating sustainability and visibility that is widely recognized.\n\nDean et al. (2016) emphasize that employees play a crucial role in creating brand meaning through their experiences and interactions, making internal brand imaging essential for Higher Education Institutions (HEIs). Effective brand identity management in this sector requires in-depth marketing strategies, an understanding of the internal market, and the development of shared brand meaning to remain globally competitive. Involving employees in this process is critical because they act as brand representatives, and their motivation is crucial in conveying a consistent brand message (Van Den Bosch et al., 2005). Additionally, the complexity of HEIs further underscores the importance of the internal market in realizing brand identity, although this often presents a challenge in developing a cohesive brand meaning (Dean et al., 2016). Moreover, the complexity of the higher education market, with its perceived high risks and long-term consequences for consumers, hinders the effectiveness of traditional brand imaging approaches (Mampaey et al., 2020; Subbarayalu, 2022). Branding in higher education is often impeded by internal barriers, such as the presence of multiple sub-brands, complex brand architecture, or the need for efficiency in university mergers, particularly in multi-campus universities (Mampaey et al., 2020; Wu & Cheong, 2024).\n\nMeanwhile, Clark et al. (2020) explore the concept of internal branding, which is closely related to but distinct from employer branding. Internal branding goes beyond building a brand through attractive logos and slogans (Barros-Arrieta & García-Cali, 2021); it involves designing elements that embody the values upheld by the institution, ensuring that employees communicate the brand promise to customers through service delivery (Foster et al., 2010). Therefore, employees have a direct influence on brand perception and enthusiastically identify with the institution’s identity, enhancing branding activities while acting as a crucial bridge between the institution and customers (Bolhuis et al., 2018; Subbarayalu, 2022; Wu & Cheong, 2024). Additionally, HEI employees, as tangible assets, play a vital role in brand development and must be actively involved in shaping and promoting the brand (Miotto et al., 2020; Slade-Brooking, 2016). This requires an integrative framework that connects human resource management and marketing, particularly in internal marketing communication to influence the brand promise (Clark et al., 2020). In higher education, this is considered more challenging due to internal resistance (Gregersen & Johansen, 2022). As Clark et al. (2020) indicate, the presence of multiple sub-brands and complex brand architecture often hinders effective internal branding, a challenge frequently encountered in HEIs.\n\nIn this vein, an effective internal branding strategy is crucial for fostering aligned brand expectations (Dean et al., 2016). Furthermore, Dean et al. (2016) identified four stages in the brand cycle—awareness, interpretation, appropriation, and communication. In this brand cycle, employees must engage in each stage of the brand creation process, where internalization includes both macro and micro cycles of brand meaning. The macro cycle is shaped by employees’ historical brand experiences and the internal brand imaging efforts they encounter. In contrast, the micro cycle involves a personal dialectical process, where individuals assess and reinterpret brand information based on their interactions. As a result, employees’ understanding of the brand emerges through the combination of these macro and micro cycles, which illustrate the impact of internal brand imaging on how employees perceive and interpret the brand. Consequently, their historical brand knowledge (macro cycle) is continuously updated through ongoing evaluations and their personal experiences (micro cycle) with the brand. Therefore, in this study, the “experiential brand meaning” cycle is integrated with the Expressiveness Quotient (EQ) or five dimensions of reputation to categorize questions concerning the relationship between visual identity and employee involvement in its management.\n\nThe literature on HEIs’ reputation has revealed several determining factors used by stakeholders to assess reputation (Dumitriu, 2018; Dursun and Altin Gumussoy, 2021; Foroudi et al., 2019; Panda et al., 2019). Meanwhile, previous research has rapidly developed regarding how HEI reputation is assessed based on measurable dimensions. For instance, Dumitriu (2018) employed the Quality Function Deployment method to identify key issues that require immediate improvement or change in terms of educational service quality. Additionally, Foroudi et al. (2019) highlighted the role of students in value co-creation concerning the significance of university website features. Dursun and Altin Gumussoy (2021) identified important factors influencing HEI reputation in the perceptions of students, alumni, and academic and administrative staff. Panda et al. (2019) conceptualized university brand image as heritage, service quality, and trust, and explored its relationship with student satisfaction. Furthermore, Amado-Mateus et al. (2024) evaluated the psychometric properties of a reputation scale among students at private universities in Colombia. However, these studies primarily focus on perceptions specific to HEI reputation, neglecting HEI identity as a reinforcement of reputation at the organizational level. To build a strong reputation, businesses must develop emotional appeal and express themselves to stakeholders in a credible, authentic, sincere, and convincing manner, clearly communicating who they are, what they do, and what they stand for (Fombrun and Van Riel, 2004; Mohamad et al., 2017).\n\nTherefore, in building a strong reputation, the involvement of HEI identity must be incorporated, impacting the flow of brand meaning formation at the organizational level. As emphasized by Van den Bosch et al. (2005), VI can enhance interconnected dimensions. In this vein, this section outlines the Expressiveness Quotient (EQ) proposed by Fombrun and Van Riel (2004), which is used to formulate questions related to strategic visual identity. The higher the expressiveness, the more likely it is to appeal to stakeholders (Fombrun & Van Riel, 2004). This approach allows organizations to cultivate emotional appeal and to effectively understand, strategize, and execute reputation management (Angliss, 2022). The first dimension, visibility, measures the brand’s presence in customers’ minds. According to Fombrun and Van Riel’s (2004) model, this can be assessed through street exposure, national heritage, media exposure, brand equity, listing on a public stock exchange, and corporate citizenship. Distinctiveness represents the organization’s unique position in the minds of customers and other stakeholders. Strategic alignment, emotionally appealing features, and attention-grabbing messages can achieve distinctiveness, allowing an organization to stand out in a crowded market. Transparency is another critical dimension, where greater transparency leads stakeholders to rely more on an organization’s disclosures, which are measured by products and services, vision and leadership, financial performance, social responsibility, and the working environment.\n\nAdditionally, an HEI’s identity can contribute to transparency by clearly communicating its values and commitments. Fombrun and Van Riel (2004) address authenticity as a challenge through four lessons: clarifying who you are, developing broad consensus within the organization, expressing your identity clearly, and remaining true to that identity. Authentic organizations are perceived as genuine, accurate, dependable, and trustworthy. Visual elements or logos that trace back to a company’s origins can enhance perceived authenticity, even if authenticity is not always directly related to VI. Consistency, the final dimension, is determined by consistently using brands and graphic elements over time. This consistency ensures that the brand’s identity remains clear and recognizable to its audience. In this study, the EQ was employed to formulate questions related to strategic visual identity elements. The decision to use the EQ was based on its five interconnected dimensions. Assessing the effectiveness of the measures undertaken is essential to building a robust reputation, and the EQ provides a comprehensive evaluation framework for this purpose. Van Den Bosch et al. (2005) explored the role of VI in reputation management, demonstrating how it aligns with corporate reputation through the interrelated dimensions in Fombrun and Van Riel’s (2004) reputation model. Managing VI effectively is crucial, as it represents an organization and serves as a visual expression intricately linked to its reputation.\n\n\n3. Methodology\n\nThis study employs a qualitative case study approach as an investigative strategy, designed to explore contemporary phenomena in-depth within their real-world contexts. The case study method is particularly suitable when the boundaries between the phenomenon and its context are not clearly defined, enabling a comprehensive examination of intricate relationships and dynamics. Drawing on the structured guidelines of Creswell and Creswell (2018), this approach integrates multiple data collection techniques, including semi-structured interviews, direct observations, and document analysis, ensuring a holistic understanding of the research topic. The rationale for adopting the case study design stems from its ability to generate in-depth insights into the research context while accounting for its complexity. According to Yin (2018), the case study is especially valuable in investigating contemporary issues where traditional experimental controls are impractical. This research design further aligns with the recommendations of Elifneh (2017) and Yin (2018), as it emphasizes a holistic methodology to capture diverse perspectives and contextual subtleties.\n\nThe data collection process was conducted over a six-month period, during which semi-structured interviews were carried out with purposively selected participants representing a range of perspectives relevant to the research context. Interview questions were designed to be open-ended, enabling participants to provide rich, detailed narratives. Observations were documented systematically in a field journal, capturing contextual nuances and participant interactions. The document analysis focused on reviewing policy documents, institutional reports, and archival records, providing additional layers of context. To minimize bias and unwanted variability, methodological triangulation was employed. This included cross-referencing data from interviews, observations, and document analysis to ensure consistency and reliability of findings. Data transcription was carried out using professional software to enhance accuracy, and member checking was conducted to validate the interpretations.\n\nThis study involves interviews with 29 employees from five public universities with legal entity status in Indonesia, known as PTNBH. Interviews were chosen as the most effective approach for collecting qualitative data through in-depth questions, which are essential for gathering qualitative information (Van den Bosch et al., 2005). The participant group consisted of 13 male and 16 female members holding non-academic positions, such as heads of departments or unit managers at their respective public universities, which are legally established (see Table 1). Regarding respondent involvement, this study obtained verbal consent, with participants willingly agreeing to participate after being informed that the data provided would be used solely for academic research purposes, and their personal information would be kept confidential. The study also received ethical approval from the Directorate of Research and Community Service (DRPM) of Institut Teknologi Sepuluh Nopember, with approval number 2402/ITS.IX.8/B/PP.05.02.00/2022. Thus, all participants provided informed consent prior to their involvement in this study, and the research was conducted in accordance with the established ethical standards.\n\nThe selection of participants aimed to ensure a representative sample from various units in each university. These employees were chosen because they are responsible for planning, strategy, and organizational policies. Diversity in selection was crucial, as it captured a range of experiences and perspectives regarding visual identity and organizational reputation. The study was conducted at five top-ranking PTNBH universities in Indonesia between 2021 and 2023, according to the Quacquarelli Symonds World University Rankings (QS WUR), Times Higher Education (THE), and Webometrics. Therefore, these universities were selected, including (1) Universitas Indonesia (UI), (2) Universitas Gadjah Mada (UGM), (3) Institut Teknologi Bandung (ITB), (4) Universitas Airlangga (Unair), and (5) Institut Teknologi Sepuluh Nopember (ITS).\n\nPublic universities with legal entity status in Indonesia were chosen as the subjects of this study because they have greater operational flexibility, particularly in academic and administrative areas, which are strictly regulated by their respective laws. Selecting the highest-ranked state universities with legal entity status not only simplifies the research process but also emphasizes the importance of global competition in higher education, where these institutions strive for excellence in international rankings. By focusing on these top-ranked public universities with legal entity status in Indonesia, the study examines institutions with significant societal influence, which are presumed to manage their visual identity (VI) meticulously. These rankings are crucial benchmarks for assessing quality and reputation, which are essential for stakeholders, including prospective students, governments, and institutions.\n\nData collection for this study was conducted through semi-structured interviews (see Figure 1). These sessions were recorded and subsequently transcribed into written text for analysis. To ensure confidentiality, any personally identifiable information was carefully removed from the transcripts. The interview format was designed to explore the participants’ connection to the brand and their significant experiences. Each session lasted approximately 60 minutes and included semi-open questions focused on managing visual identity, derived from the five dimensions of reputation. This study focuses on the visual identities of organizations, which encompass elements such as logos, colors, designs, and other visual components that collectively shape the image and brand of state universities with legal entity status. Emphasizing visual identity underscores how these elements contribute to the overall perception and effectiveness of the university’s brand in the eyes of internal stakeholders. The approach highlights the critical role of visual identity in creating a cohesive and impactful brand within the university community and beyond (see Figure 1). It gathers diverse perspectives to deepen the analysis of institutional perceptions and the management of visual identity.\n\nThe interviews were structured around two sets of questions to explore different aspects of institutional branding and visual identity management. The first set focused on internal branding, specifically examining structural and organizational culture. Influenced by Dean et al. (2016), this part of the interview was based on the concept of “experiential brand meaning,” which outlines four stages in the brand cycle: awareness, interpretation, appropriation, and communication. This framework was used to explore how employees engage with each element in the brand-building process, shedding light on their commitment to embodying and promoting the brand’s values internally, within the context of universities in Spain. Similarly, Foroudi et al. (2020) and Punjaisri and Wilson (2011) explored how employees embody the brand. Clark et al. (2020) conducted previous research on the contribution of internal branding to university management in Canada. These studies emphasize the valuable role of internal branding in higher education brand management strategies. While there is substantial research on this topic in the context of universities in developed countries, there is a need for further research in developing countries such as Indonesia.\n\nThe second set of questions assessed employees’ awareness and understanding of institutional branding. These questions specifically targeted the interpretation, appropriateness, and communication of institutional identity, focusing on the logo and its components—name, design, color, typeface, and slogan, as outlined by Foroudi et al. (2020). This set was strategically designed to incorporate elements related to the five dimensions of reputation, facilitating a systematic exploration of the intricate relationship between visual identity and employee engagement. The interview questions were also tailored to uncover the multifaceted measures implemented to maintain a consistent visual identity, categorized under structural and cultural indicators. Organizational structure, as defined by Van den Bosch et al. (2004), refers to how an organization assigns its workforce to various tasks and coordinates its efforts effectively. According to Schein in the same source, culture is described as a set of fundamental assumptions shared within the organization. These assumptions evolve over time and help the organization address external adaptation and internal integration challenges. They are taught to new members as the correct way to perceive, think, and feel about these challenges. The development of organizational culture is significantly influenced by founders and leaders, whose practices in managing visual identity are deeply embedded in their ways of learning and influencing. This aspect of culture and leadership is crucial as it underpins the consistency of the visual identity that shapes public perceptions of the institution. In summary, the study examines how strategic branding efforts are integrated within the broader organizational framework.\n\nThe study utilized a qualitative inductive approach with data-driven thematic analysis, which involves detecting, analyzing, and reporting patterns or themes in the data (Braun & Clarke, 2006). This method requires the researcher to sort through the data, looking for similarities or connections, a practice common to many qualitative research techniques (Miles & Huberman, 1994). The findings are then presented in a narrative format, which is a typical method in case study research. This narrative approach helps to recreate the context and flow of evidence, allowing readers to better understand the meaning behind the data (Czarniawska, 1997; Gillham, 2000). In addition, qualitative research methods, as noted by Strauss and Corbin (1990), are widely used because they enable the collection of in-depth data from fewer research units, rather than broad, surface-level information from many units (Jennings, 2001). This type of analysis describes individual experiences, variations, and relationships, with a cross-case analysis offering multiple perspectives on a case, which helps answer the study’s research questions (Creswell & Creswell, 2018; Yin, 2018). The cross-case comparison involved presenting the textual data consistently across all cases, which allows for a comprehensive view of the findings (Gibbs, 2008).\n\n\n4. Result and analysis\n\nThe aim of this study is to explain the correlation between visual identity components and reputation from the perspective of internal stakeholders. It concludes that the logo serves as an identity component that conveys the personality and values of higher education institutions to their employees. By unraveling the intricacies of the “experiential brand meaning” cycle—awareness, interpretation, appropriation, and communication—developed by Dean et al. (2016), this study examines the extent to which the identity of HEIs impacts experiential brand meaning for the educational staff of five state universities with legal entity status in relation to internal brand image. This, in turn, has implications for the understanding of visual identity within HEIs, which can create a brand image and reputation. Therefore, through the data analysis in this study, a number of interview results are categorized based on the themes of experiential brand meaning proposed by Dean et al. (2016).\n\nBased on interviews with various respondents, brand awareness prior to joining the university reveals that each new interaction with the brand begins with a micro-cycle of evaluation and interpretation. This study shows that exposure to brand elements, such as visual identity (VI), initiates the development of brand meaning. Some employees, even those who are not alumni of the university, were already familiar with its reputation and brand image without actively seeking information. Furthermore, after joining the university, employees strive to deepen their understanding of the HEI’s identity, emphasizing a sustained commitment to brand alignment and recognizing the importance of internalizing the institution’s values and identity.\n\n“I am quite familiar with this university, as information about it was often shared and embedded in my mind during my student years. In addition, this university is one of the best in Indonesia. Its reputation is widely known. The logo or visual identity certainly triggers brand meaning, as its philosophy is as strong as its reputation.”\n\n“Although I already knew about the university, I strive to learn more deeply about the institution where I work, especially regarding the meaning of its visual identity, its application, university developments, rankings, and so forth, to internalize it better. This is important since I plan to serve at this institution for a long time.”\n\nThus, this proactive approach to brand education reflects a solid internal culture that values and supports brand identity. It demonstrates that although VI elements are important, the overall reputation built across various channels plays a crucial role in brand perception. Furthermore, respondents view HEI identity as an organizational asset, recognizing its significant impact on overall reputation. This unanimous recognition reveals a well-embedded brand culture where the importance of a strong HEI identity is universally understood and valued. In this regard, respondents also noted that information on HEI identity can be accessed via the university’s website, which serves as a guide to its visual identity standards. Therefore, a high level of brand awareness and understanding is essential to ensure that HEI identity is conveyed consistently and effectively both internally and externally.\n\n\"I believe that visual identity represents the brand’s identity, which can impact the university’s reputation. As employees, we need to understand its philosophy and application to maintain its reputation. The university provides a visual identity guide linked on the main website. If there is any confusion, we as employees can contact the management unit via email or WhatsApp.\"\n\nUpon recognizing the importance of new brand knowledge, individuals become motivated to move to the second stage, which involves interpreting the brand’s meaning.\n\nAt the interpretation stage, individuals engage in personal analysis and comprehension, representing a micro-cycle in the development of brand meaning. In this phase, they shift from passive recipients to active evaluators, critically assessing and interpreting the new brand knowledge they have acquired through interactions with the brand. Most interviewees offered positive endorsements for the brand, particularly within the context of their roles, indicating that the university brand is viewed as valuable and reputable in their work. This finding aligns with research on the dissemination of internal brand knowledge and its impact on employees. As noted by one respondent:\n\n\"I always recommend the university’s brand, especially to alumni and partners, based on the knowledge I’ve gained during my time here, including information on rankings, research, and community service. It is part of my role and responsibilities to ensure the university’s reputation is positively regarded by alumni and partners.\"\n\nSharing knowledge about brand meaning represents another significant area explored in this study. Most respondents actively shared their understanding of the HEIs brand with coworkers and colleagues within work-related units, a practice that reflects their awareness of the importance of disseminating brand knowledge and fostering a deeper understanding of brand identity within the organization, as emphasized by Mampaey et al. (2020). All respondents expressed that a clear and strong HEI identity is crucial to conveying the university’s distinctive characteristics.\n\n\"Having a clear and strong identity is, of course, essential, as it is built on a philosophy and foundation established by the founders with great intention. Additionally, a strong and clear identity will naturally attract people’s interest, acting as an embodiment of the university’s characteristics.\"\n\nThis unanimous perspective reflects a profound understanding that a strong brand identity is a valuable asset, crucial for communicating the university’s unique attributes and values. This shared belief underscores the importance of maintaining a consistent and coherent brand identity. Consequently, the interplay between stakeholders at both internal and external levels can lead to the co-creation of HEIs identity, which may not fully align with the core brand values. Rather, as this co-created meaning becomes accepted as the brand’s reality, the appropriation stage of the brand meaning reinterpretation cycle begins.\n\nAt the appropriation stage, employees form brand associations by internalizing the co-created brand meaning and developing brand engagement. Once they accept this co-created meaning as the brand’s reality, they fulfill their roles as brand interpreters (Dean et al., 2016). Therefore, universities need to focus on brand reinforcement for their employees, as highlighted by some respondents who noted that HEIs have provided some form of brand education and training. However, other respondents indicated a lack of such provision. Effective brand reinforcement enhances employees’ understanding and behavior aligned with HEIs identity values.\n\n\"Internalization, especially concerning campus branding, significantly impacts work scope and increases knowledge and pride in the university.\"\n\nThe appropriation stage involves the internal and emotional processing of brand meaning, where employees develop brand associations that reflect their understanding of the brand’s purpose, personality, and values. This process fosters an emotional attachment to the brand, as the acquired meaning leads to a deeper relationship and engagement with it.\n\nAt the re-interpretation stage, communication is identified as essential for articulating and expressing brand meaning in each brand interaction. All respondents acknowledged the importance of HEIs identity in building relationships with stakeholders, particularly in promotional and collaborative activities. This universal acknowledgment highlights a comprehensive understanding of the critical role that brand identity plays in effectively engaging various stakeholders, as discussed in Foroudi et al. (2020).\n\n\"The brand identity represented by a symbol or logo is crucial when building relationships with stakeholders, especially with government agencies. Our campus identity significantly influences collaborations as an educational institution. While this identity might be seen as bureaucratic, it also necessitates accountability to society, especially regarding partnerships. Therefore, I often convey this meaning when establishing partnerships, conducting research, or engaging in community service with partners, alumni, and others.\"\n\n\"I usually communicate the brand meaning during university events, visits, collaborations, community engagements, and other similar activities, as I am often asked about it.\"\n\nIn this vein, employees actively communicate brand meaning to partners, demonstrating a proactive effort to convey HEIs identity. Consistency in brand messaging emerges as a significant theme among stakeholders during collaborations, visits, or specific events. By fostering a culture of consistent communication, the university can strengthen its brand identity and ensure that all stakeholders receive a coherent message. However, it is also necessary to consult with the visual identity (VI) manager to ensure the proper direction of VI usage. Some respondents frequently consult the VI manager to ensure precise implementation and communication.\n\n\"I consult with the visual identity manager about publications and announcements. Since we often received feedback, we now regularly consult about its implementation to maintain consistency.\"\n\nThis highlights the importance of consulting with the visual identity manager to ensure brand consistency. Thus, reinforcing HEIs identity at the organizational level is still needed to bridge this gap and ensure all employees accurately understand the values of VI. Consequently, this section illustrates that the effectiveness of the communication stage heavily relies on the thorough development of the preceding stages. While the communication stage completes the interpretation cycle, the evolving macro and micro brand meaning cycles continue. As one interpretation cycle ends, a new cycle begins, fueled by ongoing social interactions among diverse stakeholders. Employees articulate the brand through interactive dialogue, express their understanding of HEIs identity, and engage in re-evaluating the significance of VI knowledge. As a result, the experiential brand meaning cycle associated with VI and the brand persists, aligning with the framework proposed by Dean et al. (2016).\n\nBased on interviews with administrative staff at the structural level across five state universities with legal entity status, several insights emerged regarding brand visibility. For instance, University [1] faces challenges with its logo, which is often displayed in low-resolution media, diminishing its appeal and visibility. This issue highlights the need for high-quality media applications to ensure the logo is clear and impactful, as supported by Subbarayalu (2022), who discusses effective brand management and the importance of logo placement in educational institutions. In contrast, University [2] received positive feedback regarding brand exposure. Several employees noted that the university enjoys strong brand visibility across various media platforms, which significantly enhances the university’s public presence and recognition. Meanwhile, University [3] faces challenges with its slogan, which is perceived as less memorable due to its rare usage in brand communications and lack of integration with the logo. This gap affects impression and recall, underscoring the need for more consistent and strategic use of the slogan and logo to strengthen brand identity, as emphasized by Siyanbola and Adeyemi (2021).\n\nSimilarly, University [4] is known for its strong brand exposure across various media, positively contributing to its visibility. Consistent presence across these platforms helps maintain a strong brand image and demonstrates that the university’s media strategy effectively promotes its brand. However, University [5] faces various challenges, such as its name, logo, and slogan, which are still seen as hindering promotional and brand communication efforts. It is acknowledged that the visibility dimension reveals some challenges related to the brand visibility of HEIs, particularly regarding logos that often need to stand out more across different media. Addressing these challenges by enhancing logo application, ensuring high-quality media display, increasing brand exposure, making the slogan more memorable, and better integrating brand elements is essential for improving visibility and appeal.\n\nInterviews with administrative staff at the structural level across five state universities with legal entity status revealed several findings related to the uniqueness of their brand elements. For instance, University [1] uses a typeface in its brand elements that is not perceived as evoking positive feelings, as it lacks a direct connection to the university’s brand. Therefore, a more suitable typeface that aligns with the brand’s character and carries stronger emotional appeal would be beneficial in enhancing the university’s uniqueness. Meanwhile, some employees at University [2] expressed dissatisfaction with their university’s logo, preferring logos of other universities they felt were more distinguished. This indicates an issue with the logo’s appeal and effectiveness as a brand differentiator. The university may need to evaluate the logo’s design to ensure it better represents the institution’s identity, is more acceptable to employees, and stands out among competitors.\n\nUniversities [3] and [5] face various challenges with their brand elements. Issues such as the logo being seen as unsuitable in a professional context, the university name lacking uniqueness due to regulations limiting names to geographical distinctions, and a name perceived as too long and complex detract from its simplicity. Additionally, brand colors are viewed as insufficiently unique and ineffective in influencing perception and behavior; the typeface is considered unremarkable, lacking appeal and connection to the brand; and the slogan is regarded as unmemorable, lacking emotional resonance, and insufficiently motivating within the work environment.\n\nUniversity [4] encounters challenges with brand colors that are perceived as less impactful on mood, as they are not popular among staff. This finding suggests that the university could consider brand colors with stronger emotional resonance to enhance overall appeal and uniqueness. The findings from these universities highlight the need for stronger, more culturally relevant brand elements that foster emotional engagement and alignment with the brand. Addressing these issues would allow universities to stand out in a competitive market and build closer relationships with stakeholders.\n\nIn response to the increasing demand for transparency among Higher Education Institutions (HEIs) in a highly competitive environment, maintaining transparency has become a challenge impacting HEIs’ reputations. Based on this study’s analysis of interviews with educational staff at the structural level across five state universities with legal entity status, various perspectives on brand element transparency were revealed. At University [1], for example, there was a perceived lack of alignment between the brand colors and the HEI’s vision and mission. Some respondents expressed difficulty in associating the color scheme with the university’s core values, indicating a misalignment between the brand colors and the university’s identity. In contrast, Universities [2] and [4] demonstrated good levels of transparency, as the managerial level clearly communicated information related to products and services, vision and leadership, financial performance, social responsibility, and work environment. This comprehensive communication reflects a commendable level of transparency and indicates effective brand message management.\n\nMeanwhile, Universities [3] and [5] face similar challenges related to their brand elements. For instance, the university names are perceived as misaligned with the intended message, particularly in emphasizing technology; the logo designs are seen as inadequate representations of higher education institutions; the colors are deemed inconsistent with the universities’ vision and mission; and the typeface and slogan are considered insufficient for conveying the desired message. Additionally, at University [5], some respondents reported limited awareness or clarity regarding brand implementation and a lack of oversight from related units in brand management, highlighting the need for improved management and supervision. These challenges underscore the need for a more cohesive brand design approach that aligns with the university’s messaging. Enhancing these elements would enable HEIs to more clearly convey their identity and values to stakeholders.\n\nAuthenticity shares a similarity with consistency, both emphasized as essential goals for HEIs. Gregersen and Johansen (2022) explicitly argue that a consistent Visual Identity (VI) must authentically reflect or express the essence of an HEI, essentially visualizing who or what the HEI represents. Meanwhile, in the current state of HEIs in Indonesia, this study reveals a range of internal organizational perspectives concerning the authenticity of university brand elements. University [2] stands out as the only institution receiving positive feedback regarding authenticity, as noted by respondents. The university is perceived to have a clear understanding of its roots and foundations, which are seen as authentic and trustworthy. Consequently, the HEI has successfully communicated its core values and origins, fostering a sense of reliability and authenticity. In contrast, respondents from University [1] observed that brand design elements were often inadequate in conveying a clear message and meaning. Frequently, the intended meaning requires further explanation to be fully understood, and the slogan is perceived as insufficiently specific in communicating the brand.\n\nRespondents from Universities [3] and [5] highlighted similar issues to those noted previously, underscoring the need for a stronger, more coherent approach to managing brand elements to enhance authenticity. It is essential that these elements are clearly defined and communicated consistently to cultivate an authentic brand perception. Meanwhile, respondents from University [4] reported authenticity-related issues, such as a logo that stakeholders, both internal and external, found difficult to interpret, and a typeface that was seen as ineffective in communicating the university’s identity and was deemed inauthentic. Addressing these elements could therefore contribute to enhancing the university’s credibility and strengthening stakeholder trust.\n\nConsistency in Visual Identity (VI) is not solely about aesthetics; it is also closely linked to the perceptions and responses from both internal and external stakeholders. Inconsistencies in VI elements can lead to confusion and negative perceptions of HEIs, whereas alignment of VI elements enhances brand recall and recognition, creating a strong psychological bond. This study highlights several findings regarding consistency, as expressed by the respondents. Only University [2] has implemented clear and consistent VI guidelines, which support the reputation of the HEI. This underscores the importance of well-defined guidelines in maintaining a strong and cohesive brand image, ultimately enhancing credibility and trust among stakeholders.\n\nIn contrast, Universities [1], [3], [4], and [5] still require a more integrated branding approach to ensure that all elements are applied consistently. Addressing these inconsistencies will help build a more professional and reliable image. Furthermore, the findings of this study align with previous research on brand management (Dwitasari et al., 2022) and maintaining a cohesive brand image (Foroudi et al., 2020).\n\n\n5. Discussion and Contribution\n\nThis study makes a significant contribution to understanding the importance of visual identity (VI) in the formation of reputation and identity within higher education institutions (HEIs). The identity of HEIs plays a crucial role as the presentation of the organization to its stakeholders, encompassing communication, design, culture, behavior, structure, industry, and organizational strategies (Wu & Cheong, 2024). Following the principle of marketing that begins from the inside out, it is vital for HEIs to build a comprehensive and integrated organizational identity through internal branding and VI. Internal branding aims to strengthen emotional attachment and employee understanding of the organization’s values, while VI, consisting of elements such as logos, colors, and graphic symbols, functions as an authentic and consistent visual communication tool (Wu & Cheong, 2024). Therefore, this study employs the EQ framework to assess the experiential brand meaning at the internal level of HEIs in Indonesia.\n\nThe findings of this research indicate that awareness of identity and branding among internal stakeholders has a significant impact on shaping positive perceptions of the university’s brand identity, aligning with the findings of Buono and Fortezza (2017). Additionally, the study highlights positive reception toward HEI brand identity; however, differing perceptions among stakeholders necessitate further exploration to strengthen the brand identity and ensure alignment with stakeholder expectations. Before the brand is communicated to external stakeholders, it is essential for internal stakeholders to understand the brand as a reflection of the institution’s image. The findings also suggest that although there are initiatives for university branding training, understanding of core brand elements such as vision and mission is not yet evenly distributed among respondents. As Baker and Balmer (1997) assert, the evaluation of VI serves to identify organizational weaknesses. In this case, the logo, a key element of VI, serves as a valuable asset that creates brand recognition for the institution in an increasingly competitive market (Van Den Bosch et al., 2005; Foroudi et al., 2020). Consistency in the application of VI is crucial for building and maintaining a strong reputation. The study finds a correlation between VI, particularly logos, and five dimensions of reputation: visibility, distinction, transparency, authenticity, and consistency. In terms of visibility, logo size on specific media presents a challenge that requires greater exposure. Meanwhile, the distinction aspect suggests that universities enhance their brand elements to create a more unique and positive impression, for example by improving logo design, color selection, more appealing typography, and a distinctive slogan.\n\nFurthermore, the transparency aspect shows that some HEIs have effectively communicated information, but there is a need to ensure authenticity and effectiveness of brand elements in conveying the desired message. Regarding consistency, consistent efforts are necessary to ensure that brand elements are applied comprehensively across the institution and that academic staff understand the VI guidelines. Based on these findings, the study emphasizes that although VI contributes to building reputation, some universities need to address all dimensions of reputation more fully in order to be more effective in managing VI and strengthening the HEI brand in the eyes of stakeholders. Thus, the integration of VI into internal branding is not merely cosmetic but rather a strategic element that strengthens identity and relationships with stakeholders, creating sustainability and institutional visibility that is widely recognized. In short, visual identity is like a well-tailored suit—it’s not just about looking good; it’s about how well it fits the institution, and how it’s perceived by everyone from the internal team to the wider public. And if the fit’s off, well, we might need to swap a few elements around to make sure the message is clear, consistent, and confidently communicated.\n\nThe theoretical contribution of the findings in this study to the field of higher education institution (HEI) identity, particularly in the context of visual identity (VI), can be seen through the development of understanding regarding the alignment between visual identity elements and experiential brand meaning. Based on the findings that reveal differences in the consistency and authenticity of VI elements among public universities, this study supports the view that VI is not only a symbolic identity but also a crucial instrument for effectively conveying the core values of the institution.\n\nIn the context of the Expressiveness Quotient (EQ) framework by Fombrun and Van Riel (2004), this study enhances the understanding of experiential brand meaning by emphasizing the importance of consistency and alignment of VI elements in creating a positive brand experience for stakeholders. The EQ framework, which highlights the role of brand experience in shaping perceptions and consumer satisfaction, suggests that visual alignment can strengthen perceptions of institutional quality in the minds of educational consumers, such as students and academic staff (Dean et al., 2016). This study also contributes to supporting prior research that underscores the importance of consistency in corporate visual identity (CVI) to avoid confusion and inconsistency that could damage the institution’s brand image (Clark et al., 2020; Coman et al., 2021; Van den Bosch et al., 2005). These findings emphasize that visual consistency not only enhances brand recall but also increases brand trust and stakeholder confidence, which are key aspects of HEI identity developed through cohesive brand experiences that align with the core values of the university (Balmer & Podnar, 2021; Gregersen & Johansen, 2022).\n\nFurthermore, this study expands the EQ framework in the context of HEIs by demonstrating that authentic VI elements that align with the institution’s mission and vision have the potential to strengthen experiential brand meaning. By focusing on consistency and authenticity in VI, HEIs can create a brand identity that is not only recognized but also valued by stakeholders, as consistent VI facilitates a brand experience that reflects the institution’s commitment to the quality and values of higher education (Dean et al., 2016; Foroudi et al., 2020).\n\nThe practical contribution of this study provides concrete guidance for academic staff, managers, and stakeholders in higher education institutions (HEIs) to strengthen the institution’s image and brand identity through consistency and authenticity of visual identity (VI) elements. For academic staff, they are encouraged to incorporate institutional visual elements, such as logos and colors, into their daily academic activities, thereby reinforcing the professional image of the institution. The findings encourage academic staff to understand and consistently apply VI guidelines in various academic activities and public communications. For managers, it is expected that they will develop clear visual guidelines and provide training for staff to ensure the consistent application of visual identity across all communication platforms. Managers should also monitor the implementation of VI regularly to identify and rectify any inconsistencies that may arise in various marketing materials and institutional communications. Meanwhile, other stakeholders, including alumni and industry partners, can support the institution’s positive image by adhering to visual guidelines in official activities and external communications. This strong and consistent visual narrative will help establish the institution’s image as professional and trustworthy in the eyes of the public.\n\n\n6. Conclusion\n\nThis study aims to explore the level of knowledge, understanding, and employee engagement in comprehending the brand identity of five public universities (PTN-BH) in Indonesia. Using the theory of HEIs identity and the EQ (Expressiveness Quotient) framework to review the brand experience formulated by Dean et al. (2016), the study reveals that there are differences in the implementation of visual identity across each institution, particularly in the consistency of logo usage, colors, typography, and other visual elements. The findings indicate that visual consistency is crucial in building a credible and professional image, while the authenticity of visual elements helps to strengthen the overall identity of the institution. The study also highlights that universities with clear visual guidelines and strong internal communication are better able to maintain visual identity consistency, which in turn reinforces positive perceptions among stakeholders. In the context of the EQ framework, it was found that alignment of visual elements can create a positive brand experience, strengthen emotional connections, and enhance stakeholder engagement with the institution. This study also addresses a research gap in the existing literature, which generally focuses on the image and reputation of higher education institutions without emphasizing the importance of VI at the organizational level. The results of this study emphasize that consistent VI not only strengthens the institution’s image but also plays a role in creating a more meaningful brand experience for users, which ultimately enhances trust and stakeholder loyalty.\n\nThis study has limitations as it only focuses on five public universities in Indonesia, selected based on national and international rankings. The results may not reflect the situation in other institutions with different backgrounds and rankings. Furthermore, this approach may overlook other variables that could influence the implementation of visual identity in HEIs. Therefore, further research is recommended to include a broader range of universities from various categories and regions, as well as to consider other contextual factors in the application of visual identity.\n\n\nEthical approval\n\nThis study received ethical approval from the Directorate of Research and Community Service (DRPM) of Institut Teknologi Sepuluh Nopember, with approval number 2402/ITS.IX.8/B/PP.05.02.00/2022. The research adhered strictly to the principles outlined in the Declaration of Helsinki, ensuring the highest ethical standards in research involving human participants. All participants were informed about the purpose, procedures, and potential risks of the study. Written informed consent was obtained from all participants prior to their involvement, ensuring their voluntary participation. Participants were assured of confidentiality and anonymity, and they retained the right to withdraw from the study at any time without any repercussions. The collected data were securely stored and used solely for the purposes of this research.\n\n\nAuthor contributions\n\nConceptualization: Putri Dwitasari, Ellya Zulaikha\n\nMethodology: Putri Dwitasari\n\nData curation: Putri Dwitasari, Syarifa Hanoum, Rabendra Yudistira\n\nAlamin\n\nInvestigation and Anlysis: Putri Dwitasari, Ellya Zulaikha, Syarifa Hanoum\n\nPreparation of the manuscript: Putri Dwitasari, Syarifa Hanoum, Rabendra Yudistira\n\nAlamin, Luqman Lee\n\nRevision of important intellectual content: Putri Dwitasari, Luqman Lee\n\nSupervision: Ellya Zulaikha", "appendix": "Data availability statement\n\nThis study ensures the security and confidentiality of the data, as it involves sensitive organizational information that is classified and proprietary to the organization being studied. Since the interview participants include members of the organization’s management, there is a potential risk of reputational impact, and disclosure of the information could violate ethical research principles. To address this, the researchers have assured all participants that the data will be securely protected and used solely for research purposes. For those wishing to access the data, a formal request must be submitted to the corresponding author, accompanied by evidence of ethical approval and a detailed data access application. The data supporting the findings of this study can be accessed by contacting the corresponding author (putridwita@its.ac.id). Requests for data access will be considered if intended for non-commercial research purposes, with the condition that proper credit is given to the original authors.\n\n\nAcknowledgement\n\nThe authors would like to thank the Department of Visual Communication Design—Faculty of Creative Design and Digital Business and the Interdisciplinary School of Management & Technology, Institut Teknologi Sepuluh Nopember, Surabaya, for facilitating the entire study process. We are truly grateful for their invaluable support.\n\n\nReferences\n\nAaker DA: Building strong brands. Simon and Schuster; 2012.\n\nAlessandri SW, Yang SU, Kinsey DF: An integrative approach to university visual identity and reputation. Corp. Reput. Rev. 2006; 9: 258–270. Publisher Full Text\n\nAli A, Foroudi P, Palazzo M: Examining the Influence of Corporate Identity on Corporate Reputation and Non-financial Brand Performance in the Context of Higher Education. The Emerald Handbook of Multi-Stakeholder Communication. 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Design and methods. 6th ed.Newbury Park, CA: Sage Publications; 2018." }	[ { "id": "354625", "date": "10 Jan 2025", "name": "Teofilus Teofilus", "expertise": [ "Reviewer Expertise Higher Education", "Strategic Management", "Consumer Behavior" ], "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 \"Internal Perspectives on Visual Identities in Higher Education: A Case Study of Top-Ranked Universities in Indonesia\" presents a comprehensive examination of the influence of visual identities (VI) on institutional reputation within higher education institutions (HEIs). Through semi-structured interviews with employees from five leading Indonesian universities, this study evaluates the role of VI through the Expressiveness Quotient (EQ), emphasizing dimensions such as visibility, distinctiveness, transparency, authenticity, and consistency. The findings suggest that effective management of VI aligns internal stakeholder perceptions with external branding, thereby enhancing competitive advantages and fostering cohesive organizational identities. Constructive criticisms and recommendations\nDepth of Theoretical Integration: This article would benefit from incorporating a more comprehensive theoretical framework by establishing connections between VI's impact and not only reputation but also specific academic outcomes or recruitment metrics, thereby providing a more substantive link to HEI performance. Broader Empirical Evidence: While the case study approach provides in-depth insights, incorporating quantitative elements can enhance the generalizability of the findings. Subsequent research might consider employing surveys or experimental designs to validate qualitative insights on a larger scale. Comparison with Global Standards: This study focuses on Indonesian universities but lacks a comparison with global practices. Incorporating examples or data from Higher Education Institutions (HEIs) in other regions could enhance the analysis and provide a benchmark for Indonesian universities. Stakeholder Diversity: Expanding the range of interviewees to encompass external stakeholders, including students, alumni, and industry partners, could provide a more comprehensive perspective on how VI influences perceptions beyond the internal staff.\nTo ensure the scientific validity of this article, addressing these issues is essential. Expanding the theoretical framework, implementing a mixed-methods approach, diversifying the participant base, and incorporating comparative global insights would substantially enhance the methodological rigor and generalizability of the research.\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": "354623", "date": "13 Jan 2025", "name": "Bing Bedjo Tanudjaja", "expertise": [ "Reviewer Expertise Visual Identities", "Visual Culture", "Visual Communication Design" ], "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 journal can be accepted with minor revisions. In the data collection method there are several aspects that need to be considered such as Participant Selection please also explain how the participant selection process was carried out. For example, are there any specific criteria used to select these participants? then also related to Methodological Triangulation, explain further about how methodological triangulation was carried out in this study to add depth to the methodology. For example, how were the results from different methods (interviews, observations, document analysis) confronted and integrated? Also include Methodological Limitations: Discussion of the limitations of the research methodology used can be more explicit. For example, limitations in generalization from case studies, or potential bias in case selection. In the data collection method there are several aspects that need to be considered such as Participant Selection please also explain how the participant selection process was carried out. For example, are there any specific criteria used to select these participants? then also related to Methodological Triangulation, explain further about how methodological triangulation was carried out in this study to add depth to the methodology. For example, how were the results from different methods (interviews, observations, document analysis) confronted and integrated? Also include Methodological Limitations: Discussion of the limitations of the research methodology used can be more explicit. For example, limitations in generalization from case studies, or potential bias in case selection.\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": "356450", "date": "07 Feb 2025", "name": "Katelijn Quartier", "expertise": [ "Reviewer Expertise Retail and brand design" ], "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 describes a study on the importance and impact of visual identity (VI) on higher education. What makes this study special is that it looks not at how it is perceived by externals but how it is perceived internally. Two previously published dimensions are used for this purpose: the Expressiveness quotient and the experiential brand meaning. The study concludes that VI is crucial to building a good brand that is lived out by everyone.\nThe paper is very clear and well written with a good structure. The qualitative methodology is well justified and grounded in literature. Especially the discussion of the results is brought very enlightening. The quotes help to get a better grip on what the underlying idea is. The authors also acknowledge the limitations of their study which opens the door for further study.\nSome minor comments: Avoid repetition: for example at '4. Results' the whole first paragraph is already a summary of the previous text. This seems redundant to me. When you start explaining the dimensions of the Expressiveness quotient, list all five first so the reader is with you. In 4.6 the distinctiveness of the VI is discussed. I kind of missed seeing all five of them since they are discussed in detail: typography, color, ... It is hard to understand without seeing them.\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? No source data required\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [] } ]	1	https://f1000research.com/articles/13-1535
https://f1000research.com/articles/13-1534/v1	19 Dec 24	{ "type": "Research Article", "title": "Effect of mirabegron on lipid profile (serum cholesterol and triglyceride) in Iraqi patients with overactive bladder", "authors": [ "Hiba Hameed", "Mohammed Ismail", "Mohammed Ismail" ], "abstract": "Background Mirabegron is a β3-adrenergic receptor (β3-AR) agonist notified only for the treatment of overactive bladder. Bright preclinical results desire that β3-AR agonists manage to correct corpulence-affiliated metabolic affliction by growing brown adipose tissue (BAT) activity, white adipose tissue (WAT) lipolysis, and insulin sensitivity.\n\nObjective to evaluate the effect of mirabegron on lipid profile (serum cholesterol and triglyceride) and BMI in patient with OAB.\n\nMaterials and Methods In medical city complex (Ghazi AL-Hariri hospital) urology outpatient clinic, a prospective study of 40 patient diagnosed with OAB. These patient a took single dose of mirabegron 50 mg for 4 months and asses it’s the effect during this period on weight and fasting serum cholesterol and triglyceride.\n\nResults investigation showed that there is astatically significant reduction from baseline after 2 and 4 months’ treatment with mirabegron (P-value=0.001) in fasting Sr. Triglyceride while fasting serum cholesterol level showed increase in level after 2 and 4 months of treatment. However; this increment not statically significant after 2 months(P-value=0.227) and after 4 months(P-value=0.261) BMI showed slightly reduction after 2 and 4 months’ treatment with mirabegron but also this reduction not statically significant after 2month(P-value=0.114) and after 4 months(P-value=0.562)\n\nConclusion Mirabegron is dependable and direct drug for situation of overactive bladder (OAB) causes useful decline in level of triglyceride and the verdicts display that human BAT metabolic endeavor maybe increased subsequently never-ending pharmacological provocation accompanying mirabegron and support the analysis of β3-AR agonists as a treatment for metabolic ailment.", "keywords": [ "Mirabegron", "serum cholesterol", "serum triglyceride", "body mass index", "Beta-3 adrenergic receptors" ], "content": "Introduction\n\nOveractive bladder syndrome [OAB] is demarcated as urinary significance, occasionally in the absence of a major urinary tract infection (UTI) or other evident pathology, nocturia, urgency, and frequency of urination, either with or without urge incontinence. OAB affects quality of life significantly and affects both men and women.1 Symptoms of OAB are characterized by involuntary contractions of the bladder muscle.2 Case focuses on behavior therapy originally, obstructing that burden required, fluid management, averting or menacing vehement liquor avoiding meals, alcohol, and caffeine, as they can irritate the bladder. Antimuscarinic are thought to be the first line of pharmaceutical treatment if these approaches are unsuccessful.3 Although it is common worldwide, hyperlipidemia is particularly common in Western societies. A more objective definition of hyperlipidemia involves elevated levels of low-density lipoprotein (LDL), total cholesterol, triglycerides, or lipoproteins that exceed the 90th percentile compared to the general population, or high-density lipoprotein (HDL) levels that fall below the 10th percentile relative to the average individual. Cholesterol levels, lipoproteins, chylomicrons, very low-density lipoproteins (VLDL), LDL, Apo lipoproteins, and HDL are all lipid components. The increasing rates of obesity represent a global health crisis.4,5 One potential therapeutic approach involves pharmacological interventions to enhance fatty acid and glucose metabolism. For several decades, the β3-adrenergic receptor (β3-AR) has been a focus of interest, as its activation in rodents leads to increased energy expenditure and improved metabolic profiles.6 Even though TGs are not really atherogenic, they are a main biomarker of heart failure risk on account of their accompaniment following atherogenic fragment pieces and Apo C-III, a proinflammatory, proatherogenic protein namely to reply present on all classes of plasma lipoproteins. Dispose of LDL, atherogenesis is eased by many class of triglyceride-rich lipoproteins (TRLs), such as VLDL and VLDL fragments, other than chylomicron residues. Lipoprotein lipase (LPL) imperfectly hydrolyzes TRLs of hepatic and stomach origin, that have lengthened cholesterol esters from HDL through the function of cholesterol ester transfer protein (CETP), resulting in the regulation of lipid metabolism. These class, that are cholesterol-reinforced but TG-feeble, are lengthened and drained by macrophages to form foam capsules. The composition of fatty streaks, that are the sign to atherosclerotic memorial, is progressive by foam capsules.7 Famous to beta1- and beta2 (β1/2)-adrenergic receptors, experiment-3 adrenergic receptors (β3-AR) are accessories of the G protein-connected receptor superfamily and display a more comprehensive dispersion across the body. For example, β3-ARs are found in the circulatory system, brain, adipose tissue, and bladder, among other tissues.8 Three particular differences hold adipose tissue (AT). Reduced levels of mitochondria are present in white adipose tissue (WAT), has many lipid droplets present and is implicated in the storage of energy. Brown adipose tissue (BAT), which is made up of multilocular lipid droplets, has a lot of mitochondria, which promote thermogenesis and energy expenditure.9 The mitochondrial accumulation of a WAT is exaggerated and the characteristics of allure lipid objects are changed in backlash to an external motive, in a way cold, exercise, or feeling purpose. A main focus of pharmacological mediations projected at fighting corpulence is the β3-adrenergic receptor (β3-AR). When this receptor is activated, energy expenditure rises and glucose uptake is improved. The heightened activity of brown adipose tissue (BAT) is linked to these consequences.10 The sensitive main central nervous system is a meaningful inducer of two together lipolysis and blazing. From a molecular standpoint, noradrenaline (NA) triggers a complicated cascade on adipocytes via binding to β3-adrenergic receptors (β3-AR). Gs-proteins’ α-subunit is activated by this interaction, which results in the synthesis of cyclic adenosine monophosphate (cAMP). Lipid droplet-associated proteins, most notably hormone-sensitive lipase (HSL), are important participants in this pathway as they accelerate the process of lipolysis in adipocytes. Free fatty acids (FFAs) are consequently liberated from hydrolyzed triglycerides that are kept in lipid droplets. FFAs induce the production of Uncoupling Protein 1 (UCP1) in mitochondria, which in turn stimulates thermogenesis in brown adipocytes. Subsequently, FFAs undergo β-oxidation, resulting in the production of heat and water as byproducts inside the mitochondria. This process builds stable quantities substantial in the form of ATP, that is to say produce through β-burning and the order in which metabolic processes and signaling events occur. As a result, FFAs are used by different tissues as an energy source. Free fatty acids (FFAs) are produced by triglycerides that are stored in adipose tissue or consumed as fat in food. Cells need FFAs to assist energy synthesis and metabolic control.11,12 Overactive bladder (OAB) symptoms can be treated with mirabegron, an orally active β3-adrenergic receptor agonist authorized for this purpose. Adverse effects cause many people to stop taking anticholinergic medications. A quarter of OAB patients discontinue their medication within three months as a result of adverse symptoms include cramping in the muscles, dry mouth, impaired vision, and dry eyes. Furthermore, there is evidence linking the use of anticholinergic drugs to cognitive decline, which raises further safety concerns.13 The breakdown of white adipose tissue (WAT) and the activity of brown adipose tissue (BAT) are also facilitated by the activation of the β3-adrenergic receptor.14 Mirabegron has been shown in a human research to increase WAT metabolism and BAT activity. In recent years, mirabegron has become more and more well-liked as an antimuscarinic substitute for OAB sufferers.15 This study will test mirabegron in the composition and review allure pharmacokinetics, conflicting accouterments, and influence in acting OAB and to judge the effect of mirabegron on lipid sketch (cure cholesterol and triglyceride) and BMI in patient following OAB.\n\n\nMethods\n\n\n\nDuring the baseline assessment of the patients:\n\nThe weight in kilograms divided by the square of the height in meters yielded the body mass index, or BMI (kg/m2). In accompanying study, women accompanying a BMI of ≤18.5 were classified as thin, those accompanying a BMI of 18.5-24.9 were deliberate to have normal weight, women accompanying a BMI of 25-29.9 were classification as obese, and women accompanying a BMI of ≥30 were marked as corpulent.17\n\nVenous blood samples were collected from fasting patients using plastic tubes with gel barriers. These gel barriers, being a pure substance, are highly stable in terms of their physical and chemical properties. After centrifugation, the collected samples were separated into serum or plasma from blood cells. Both serum and plasma were deemed acceptable specimens. The gel barrier effectively separated the serum from fibrin and cells post-centrifugation, thereby preventing the exchange of substances between blood cells and serum. This process maintained the biochemical characteristics and chemical components of the serum for an extended period. The levels of serum cholesterol and triglycerides were analyzed using the Architect c4000 analyzer (Abbott Diagnostics®, USA). This system is a high-throughput clinical chemistry analyzer capable of performing up to 800 tests per hour. For more information, visit Abbott Diagnostics at [https://www.corelaboratory.abbott/int/en/home.html].\n\nDuring the first follow-up visit (2 months after the baseline assessment), the patients’ weight and subsequent changes in body mass index, as well as fasting serum cholesterol and triglyceride levels, were measured.\n\nIn the second follow-up visit (2 months after the first visit), the same parameters were assessed as in the initial visit. Out of the total 40 patients, only a subset completed the follow-up assessments.\n\nThe data were analyzed using statistical package for social sciences (SPSS) version 22. The data were presented as mean ± standard deviation. paired t-test was used for compare the mean of continuous variable in the study. Level of P-value less than 0.05 was considered significant.\n\n\nResults\n\nThe total sample included in this study was 40 patients, of whom 60% were female and 40% were male, and the mean age of participants were 50.7±18.12 years. Regarding the height and weight of the sample, the results showed that the mean of height and weight were (163.65±8.393 m and 77.25±12.09 kg respectively) as shown in Table 1 and Figure 1.\n\nN=40.\n\nAt baseline time, the result of study showed the mean of BMI were (29.02±5.216 kg/m2), while the mean of s. cholesterol and s. triglyceride were (192.33±33.300 mg/dl and 142.62±29.127mg/dl respectively) as shown in Table 2.\n\nN=40.\n\nTwo months later, we reevaluate the same parameters from our sample and discover that the mean BMI has dropped to 28.82±64.926 kg/m2. Moreover, the results showed that the mean serum cholesterol level (196.45±30.932 mg/dl) was higher than the baseline level, whereas the mean triglyceride level (130.2±27.936 mg/dl) was lower. Considering Table 3.\n\nfour months later, when we reevaluate the same characteristics from our sample, discover that the mean BMI has slightly fallen to (28.62 ±4.859 kg/m2). As show in Figure 2 and Table 4.\n\nAdditionally, the mean triglyceride level (121.25±27.554 mg/dl) decreased while the mean cholesterol (199.90±24.746 mg/dl) marginally increased. As shown in Figure 3 and Table 4.\n\nAfter 2 months\n\nAccording to the study’s findings, there is no a statistically significant correlation between the mean BMI and serum cholesterol after two months (p=0.114,0.227 respectively), However, the results, as indicated in Table 5, reveal a statistically significant difference among mean of S. triglycerides after 2 months (p=0.001).\n\nAfter 4 months\n\nThe outcomes of the study demonstrate that after four months, there is a statistically significant link (p=0.001) between the mean triglyceride, but not between BMI and s. cholesterol (p=0.562, 0.261 respectively). According to Table 6.\n\n\nDiscussion\n\nThe β3-adrenergic receptor (β3-ARs) plays an important act in managing various physical functions, containing including thermogenesis in brown adipose tissue and lipolysis in white adipose tissue. The metabolic and cardiovascular impacts of β3-AR incitement by its agonists in animal models focal point the potential of β3-AR as a therapeutic target for various human diseases.18 This research aims to judge the impact of Mirabegron on the lipid characterization of things accompanying overactive bladders. The judgments concerning this current study, attended over a period of 8-12 weeks, disclose a notable change in the mean triglyceride levels (p=0.001), while no solid variances were noticed in BMI or serum cholesterol (p=0.114, 0.227, individually).\n\nAfter a period of two to four months under the mirabegron presidency, the study sample displayed a decrease in both mean BMI and plasma triglyceride levels. This finding is in line with a study conducted by Dąbrowska AM and colleagues, which highlights the importance of two distinct types of adipose tissue, each with its own unique physiological function. The activation of the thermogenic tissue-specific uncoupling protein 1 (UCP1) plays a crucial role in the metabolism of glucose, fatty acids, and other substances to generate heat in brown adipose tissue (BAT), including the associated “beige”/“brite” adipocytes derived from white adipose tissue (WAT). In situations where energy intake exceeds expenditure, WAT is responsible for storing excess triglycerides.19 There is emerging evidence suggesting that BAT may have potential therapeutic applications in adult humans. β3-adrenergic receptors (ARs) are expressed not only in brown and white adipocytes but also in the urinary bladder. Adipocytes in both white and brown adipose tissue may be stimulated to undergo thermogenesis by a β3-adrenergic agonist such as mirabegron that is currently being studied. These adipose tissue types hold thermogenic fat containers, and their incitement holds promise as a creative policy for combating corpulence by growing strength energy expenditure.20,21 Numerous clinical troubles and exploratory research have illustrated the impact of mirabegron on two together body mass and BAT activity. The authors noticed that mirabegron increased BAT exercise and resting energy expenditure.22 Preliminary data desires that mirabegron concede possibility have corresponding belongings to light exercise on the metabolism of triglycerides, bile acids, HDL, and glucose.23 Ying Z and others. found that later four and twelve weeks of situation, the levels of plasma lipids were judged following a four-time fasting period. Mirabegron usually diminished plasma triglyceride levels. Despite the plasma HDL-cholesterol waited unaltered at either time points, there was a leaning towards diminished plasma triglycerides following in position or time 12 weeks of mirabegron treatment.24 body mass index (BMI) was included in this particular study to assess its influence on body weight. However, proficient was no statistically significant change in BMI (p=4.546), regardless of few decrease in the figures. In contrast, Cypess and others. executed a study place 12 healthy men were performed 200 mg of oral mirabegron often for 12 weeks. The group receiving mirabegron demonstrated a 203±40 kcal per day rise in basal metabolic rate and an increase in BAT activity, which was known to those who took a placebo. According to the researchers’ calculations, weight reduction through energy expenditure might be as much as 5 kg in the first year and 10 kg in the next three years. Despite the fact that mirabegron was prescribed off-label to treat overactive bladder syndrome in the majority of trial participants, it was generally well tolerated. The most frequent side effects that have been authorized seen was tachycardia.25 Another study by Loh and so forth. established indicates there was a notable rise in energy expenditure following the management of 100 mg and 200 mg doses of mirabegron, but skilled was no significant unlikeness from direction following the 50 mg of mirabegron.26 Zhao and others. throwed a complex remedy approach promoting metformin and mirabegron for the preventation and treatment of obesity. Through this approach, the summed and spent energy are dealt with at the same time, which dose not have any negative impact on cardiac and vascular system. In the prevention model, the association of metformin and mirabegron grown in additional reductions of 12percent and 14 percent in raised body weight inferred by an extreme-fat diet, separately, distinguished to handling metformin or mirabegron singular. In the treatment model, the alliance of metformin and mirabegron managed to a 17% decrease in body weight in fat rodent convinced by a diet, that was 13 portion and 6 portion more having movement than exploiting metformin or mirabegron distinctively, independently.27 Animal studies have demonstrated that the administration of mirabegron reduces obesity; however, there is no trustworthy proof that patients with obesity who have received mirabegron medication have significantly lost weight. The brief trial time period and confined participant volume may be the reasons for this. Especially, extreme doses of mirabegron have proved wonted best fruitful position for exciting fatty fabric in persons, However, a thorough evaluation of its long-term safety is still required.28\n\nThe study found no evidence of statistically significant variations in serum cholesterol levels amongst participants treated with mirabegron with respect to the medication’s impact on fasting plasma cholesterol. (p=0.227). Nevertheless, further trial is authorized to sufficiently acknowledge the impact of mirabegron on cholesterol and lipid levels. This finding of this study are inconsistent with those of Finlin BS et al., who observed statistically significant decrease in cholesterol following the administration of mirabegron for 12 weeks.29 Certain studies have recorded that the incitement of the β3-adrenoreceptor, furthered by mirabegron, leads to a significant increase in ApoA-1, a critical protein component of HDL in plasma. HDL plays a awake act in mobile cholesterol from tissues to the liver for evacuation.30 In a study executed by Sui and so forth., it was seen that adult rodent incomplete in Apo lipoprotein E (Apo-E) and LDL-receptor shown raised plasma levels of total cholesterol and LDL-C after being treated with mirabegron (8mg/kg/term) for 6 weeks.31 Also, another study demonstrated that rats enhance an extreme-fat diet experienced an outdoing in their lipid description following in position or time 12 weeks of situation accompanying two differing doses of mirabegron (5 mg/kg/age and 10 mg/kg/day). This bettering was from a decrease in total cholesterol, triglycerides, and LDL-C levels, in addition to an increase in HDL-C serum levels, recognized to non-treated rats. To completely understand the impact of mirabegron on cholesterol in humans, supplementary research is necessary. Various factors, such as diet and mutations in ApoA-1, Apo-E, or LDL-receptor action, can influence serum cholesterol levels.32\n\n\nConclusion\n\nMirabegron is responsible and direct drug for OAB causes valuable decline in level of triglyceride and the verdicts display that human BAT metabolic endeavor possibly nurtured following incessant pharmacological provocation following mirabegron and support the favourable metabolic nature of β3-AR agonists.\n\nEthics and research participation consent form\n\nThis study was carried out according to the tenets of the Declaration of Helsinki. It was also approved by the Research Ethics Committee of University of Baghdad’s College of Medicine, Registration number: 03-28 date: 21/12/2023. In the first part, forty Iraqis suffering from OAB in Medical City Complex from January 2024 to June 2024 were collected. All patients gave their written informed consent to participate in the study. These patients were treated with 50 mg/day mirabegron for four months, and the results of such treatment were observed.\n\nThe following consent form is designed to explain the purpose, procedures, and rights involved in participating in the study, Effect of Mirabegron on Lipid Profile (Serum Cholesterol and Triglyceride) in Iraqi Patients with Overactive Bladder.\n\n“I hereby give my consent to participate in the study and to allow the use of my medical records in the research. The researcher has committed not to share my personal information, and I reserve the right to withdraw my participation at any time. I have been informed that the study may require several blood draws (phlebotomies) and body weight measurements”.", "appendix": "Data availability\n\nZenodo: betmiga and betmiga pulse rate datasheet, https://doi.org/10.5281/zenodo.14057595.33\n\nThis project contains following files:\n\n1. betmiga pulse rate.xlsx\n\n2. betmiga datasheet.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\nIsmail MB, Abdullhussein WQ: Efficacy and safety of sacral neuromodulation in treatment of refractory overactive bladder. Indian J Forensic Med Toxicol. 2021; 15(1): 1315–1321.\n\nAbbas IK, Rajab NA, Hussein AA: Formulation and in-vitro evaluation of darifenacin hydrobromide as buccal films. Iraqi J Pharm Sci. 2019; 28(2): 83–94. Publisher Full Text\n\nScarneciu I, Lupu S, Bratu O, et al.: Overactive bladder: A review and update. Exp Ther Med. 2021; 22(6): 1–8.\n\nOnwe P, Folawiyo M, Ogah A, et al.: Hyperlipidemia: Etiology and Possible Control. IOSR J Dent Med Sci. 2015; 14(10): 2279–2861.\n\nGuo J, Chen S, Zhang Y, et al.: Cholesterol metabolism: physiological regulation and diseases. MedComm. 2024; 5(2): 1–24. Publisher Full Text\n\nYang LK, Tao YX: Physiology and pathophysiology of the β3-adrenergic receptor. Prog Mol Biol Transl Sci. 2019; 161: 91–112. Publisher Full Text\n\nTalayero BG, Sacks FM: The role of triglycerides in atherosclerosis. Curr Cardiol Rep. 2011; 13(6): 544–552. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBachus E, Ponikowski P: Beta-3 Receptor Agonists. Int Cardiovasc Forum J. 2019; 18: 15–18. Publisher Full Text\n\nParra-Peralbo E, Talamillo A, Barrio R: Origin and Development of the Adipose Tissue, a Key Organ in Physiology and Disease. Front Cell Dev Biol. 2021; 9(December): 1–22. Publisher Full Text\n\nAntunes GC, Macêdo APA, Conceição LR, et al.: Mirabegron and physical exercise as a potential strategy for BAT activation in obesity. Obesities. 2022; 2(4): 380–388. Publisher Full Text\n\nMcAdams RP: Everything You Always Wanted To Know About the Superintendency, But Were Afraid To Ask. NASSP Bull. 2019; 79(570): 86–90. Publisher Full Text\n\nKimura I, Ichimura A, Ohue-Kitano R, et al.: Free fatty acid receptors in health and disease. Physiol Rev. 2020; 100(1): 171–210. PubMed Abstract \| Publisher Full Text\n\nO’Kane M, Robinson D, Cardozo L, et al.: Mirabegron in the management of overactive bladder syndrome. Int J Women’s Health. 2022; 14: 1337–1350. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMachado SA, Pasquarelli-do-Nascimento G, da Silva DSS , et al.: Browning of the white adipose tissue regulation: new insights into nutritional and metabolic relevance in health and diseases. Nutr Metab. 2022; 19(1): 1–27.\n\nAllison SJ, Gibson W: Mirabegron, alone and in combination, in the treatment of overactive bladder: real-world evidence and experience. Ther Adv Urol. 2018 Sep 26; 10(12): 411–419. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAli ZAM, Jasim WK, Hussein DZ, et al.: Tamoxifen effects on the lipid profile in premenopausal women with breast cancer: a follow-up study. J Fac Med Baghdad. 2018; 60(3): 141–144. Publisher Full Text\n\nKlatsky AL, Zhang J, Udaltsova N, et al.: Body mass index and mortality in a very large cohort: Is it really healthier to be overweight? Perm J. 2017; 21: 16–142. PubMed Abstract \| Publisher Full Text\n\nYang LK, Tao YX: Physiology and pathophysiology of the β3-adrenergic receptor. Prog Mol Biol Transl Sci. 2019; 161: 91–112. Publisher Full Text\n\nDąbrowska AM, Dudka J: Mirabegron, a selective β3-adrenergic receptor agonist, as a potential anti-obesity drug. J Clin Med. 2023 Nov 2; 12(21): 6897. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFinlin BS, Memetimin H, Zhu B, et al.: Pioglitazone does not synergize with mirabegron to increase beige fat or further improve glucose metabolism. JCI Insight. 2021; 6(6). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCero C, Lea HJ, Zhu KY, et al.: β3-Adrenergic receptors regulate human brown/beige adipocyte lipolysis and thermogenesis. JCI Insight. 2021; 6(11). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDehvari N, da Silva Junior ED , Bengtsson T, et al.: Mirabegron: potential off-target effects and uses beyond the bladder. Br J Pharmacol. 2018; 175(21): 4072–4082. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nO’Mara AE, Johnson JW, Linderman JD, et al.: Chronic mirabegron treatment increases human brown fat, HDL cholesterol, and insulin sensitivity. J Clin Invest. 2020; 130(5): 2209–2219. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYing Z, van Eenige R , Beerepoot R, et al.: Mirabegron-induced brown fat activation does not exacerbate atherosclerosis in mice with a functional hepatic ApoE-LDLR pathway. Pharmacol Res. 2023 Jan 1; 187: 106634. Publisher Full Text\n\nCypess AM, Weiner LS, Roberts-Toler C, et al.: Activation of Human Brown Adipose Tissue by a β3-Adrenergic Receptor Agonist. NIH Public Access. 2016; 21(1): 33–38. Publisher Full Text\n\nLoh RKC, Formosa MF, La Gerche A, et al.: Acute metabolic and cardiovascular effects of mirabegron in healthy individuals. Diabetes Obes Metab. 2019; 21: 276–284. PubMed Abstract \| Publisher Full Text\n\nZhao XY, Liu Y, Zhang X, et al.: The combined effect of metformin and mirabegron on diet-induced obesity. MedComm. 2023; 4(2): 1–18. Publisher Full Text\n\nDąbrowska AM, Dudka J: Mirabegron, a selective β3-adrenergic receptor agonist, as a potential anti-obesity drug. J Clin Med. 2023 Nov 2; 12(21): 6897. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFinlin BS, Memetimin H, Zhu B, et al.: The β3-adrenergic receptor agonist mirabegron improves glucose homeostasis in obese humans. J Clin Invest. 2020; 130(5): 2319–2331. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCao X, Li Y: β3-Adrenergic receptor regulates hepatic apolipoprotein A-I gene expression. J Clin Lipidol. 2017; 11: 1168–1176. PubMed Abstract \| Publisher Full Text\n\nSui W, Li H, Yang Y, et al.: Bladder drug mirabegron exacerbates atherosclerosis through activation of brown fat-mediated lipolysis. Proc Natl Acad Sci USA. 2019; 116(22): 10937–10942. Publisher Full Text\n\nMakar N, Abdullah O, Abu-Raia N, et al.: Possible protective effects of mirabegron on experimentally induced non-alcoholic steatohepatitis in rats. Benha Med J. 2022; (academic issue): 277–293. Publisher Full Text\n\nRazaq H: betmiga and betmiga pulse rate datasheet. [Dataset]. Zenodo. 2024. Publisher Full Text" }	[ { "id": "354503", "date": "20 Jan 2025", "name": "Adrian Wagg", "expertise": [ "Reviewer Expertise incontinence", "pharmacotherapy" ], "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 quasi experimental, pre-post study aimed to assess the effect of mirabegron on lipid profile and BMI in patients with OAB There are some typographical errors which need correction. Abstract: The background sentences need some editing to make it more understandable, for example \"indicated\", rather than \"notified only\". and the second sentence needs considerable rethinking. The conclusion in the abstract cant be supported here - there are no OAB data. The use of language needs considerable revision here too.\nIntroduction: Again, revision for the use of language is required throughout , and the usual term is urgency incontinence, rather than urge. The references need considerable revision. The authors need to use the terminology report of the ICS for ref 1 and the statement qualified by ref 2 is simply not true. I suspect that avoiding meals - as noted by the authors, if adhered to, and followed by the participants in this study, would lead to the changes reported in the study! Re: \"A main focus of pharmacological mediations projected at fighting corpulence is the β3-adrenergic receptor (β3-AR)\" - clearly this isnt the case - there are no human data which resulted in weight loss. (ref 19) the reference supporting the association between Ach use and an increase in dementia risk needs revision.\n\nThe research question requires editing for clarity. Methods Was a sample size /power calculation performed? How was the size of the sample selected? Table 1 and figure 1 are overly simple and could be simply included in the text. Alternatively, tables 1 & 2 could be combined - ideally SI units should be used. for such a small sample size - only 1 sf is really justified. What advice regarding weight maintenance was given - did any of the participants alter their diet? Could the error bars be placed on figure 2 and 3 graphs?\ndiscussion the dabrowska paper (ref 19) is a review, not a study the discussion is otherwise reasonable but needs considerable editing. Essentially state the findings of your study and note where these agree or disagree with the findings of others\n\nthe lack of a control group means that the results here need to be taken with caution. No attribution of a causal relationship can be made here\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?\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/13-1534
https://f1000research.com/articles/13-833/v1	25 Jul 24	{ "type": "Research Article", "title": "Assessing hard and loose “endpoints”: comparison of patient and expert Bristol Stool Scale scoring of 2280 fecal samples", "authors": [ "Hanna Fjeldheim Dale", "Milada Hagen", "Gunn Helen Malmstrøm", "Jennifer T. Fiennes", "Marte Lie Høivik", "Vendel A. Kristensen", "Jørgen Valeur", "Hanna Fjeldheim Dale", "Milada Hagen", "Gunn Helen Malmstrøm", "Jennifer T. Fiennes", "Marte Lie Høivik", "Vendel A. Kristensen" ], "abstract": "Background Stool consistency is an important outcome measure to evaluate in the investigation of several gastrointestinal diseases. The Bristol Stool Scale (BSS) is one of the most commonly used tools for evaluation of stool consistency. BSS ranges from 1-7 and each score is assigned to a given consistency of the feces. Self-reported characterizations can differ from an expert evaluation, and the reliability of BSS is unclear. We aimed to evaluate the reliability of BSS by comparing patient scores with expert scores.\n\nMethods Patients with inflammatory bowel disease collected stool samples throughout a 3-year follow-up. The stool´s consistency was evaluated with BSS by the patients and matched with an expert score. Agreement between patient and expert scores was assessed using Cohen’s kappa.\n\nResults BSS scores from 2280 fecal samples collected from 992 patients at up to five time points were included. When all samples were compared, there was good to substantial agreement between patient and expert scores (Cohen’s weighted kappa: 0.66-0.72). When the BSS scores were simplified and categorized as 1 (scores 1-2), 2 (scores 3-5) or 3 (scores 6-7), the agreement improved slightly (Cohen’s weighted kappa: 0.73-0.77). When the scores from the first sample per patient were compared, the experts were more likely to assign higher scores compared to the patient. The proportion of the lowest assigned scores (1-2) was 12.1% for patients and 8.1% for experts.\n\nConclusions The agreement between patient and expert BSS scores is good to substantial, especially when the BSS scores are simplified into three categories.", "keywords": [ "Fecal sample", "stool form assessment", "diarrhea", "constipation" ], "content": "Introduction\n\nStool consistency is a central component in the description of bowel habits and an important outcome measure to evaluate in the investigation of several gastrointestinal (GI) diseases.1 Altered bowel habits are often seen as a consequence of diseases in the GI tract, and the water content in the stools affecting the consistency can reflect the intestinal transit time. Rapid intestinal transit time limits the absorption of water and cause loose or liquid stools (diarrhea), whereas slow intestinal transit causes extensive water absorption and harder stools (constipation).2 The exact evaluation of stool consistency can be made by measurement of viscosity and stool water content, however this method requires cumbersome laboratory analyses and is not well suited for routine purposes. The form of the stool can serve as a proxy measure for the stool consistency, referring to the visually assessed shape and texture.3\n\nThe Bristol Stool Scale (BSS) is one of the most commonly used tools for evaluation of stool consistency.1 The BSS was developed in the early 1990s as a surrogate marker for whole-gut transit time, but is now a commonly recommended tool in clinical and research settings for the evaluation of stool consistency, rather than transit time.4,5 BSS is designed to standardize the reporting of stool consistency by a 7-point ranking system, ranging from hard lumps (Type 1 on the scale) to liquid stool (Type 7 on the scale). Types 1 and 2 are considered hard stools corresponding to constipation, whereas types 6 and 7 are considered abnormally loose and watery stools corresponding to diarrhea when evaluated together with other symptoms. Types 3, 4 and 5 are generally considered “normal” stool forms.\n\nThe generalizability of BSS as a tool in research is unclear, and fecal water content or dry weight have been suggested as better objective measures.5 Reporting can be inaccurate since subjects are asked to report their “average” stool form for the day, but bowel movements may vary in form throughout the day. Moreover, subjects’ interpretation of the BSS scale may vary, leading to inaccurate reporting of stool form. It is suggested that the BSS score is more precise when evaluated by a trained expert than by the patient.6 Here, we aimed to evaluate the reliability of the BSS in a cohort of patients with inflammatory bowel disease (IBD), by comparing the patients’ subjective scores with scores assigned by the experienced bioengineers who received the fecal samples.\n\n\nMethods\n\nThe study is a sub-study of the IBSEN III project (Clinical Trials ID: NCT 02727959), a population-based observational inception cohort with prospective follow-up, consisting of newly diagnosed IBD patients and symptomatic non-IBD controls living in the South-Eastern Health Region of Norway, included during a 3-year period from 2017 to 2019. The study protocol for IBSEN III has been presented in detail in a previous publication.7 Here, we present results from the fecal samples delivered at baseline, after 3 and 6 months and after 1 and 3 years follow-up.\n\nBSS is a 7-point ranking system, where types 1-2 and 6-7 correspond to abnormal defecation with constipation and diarrhea, respectively.1 For statistical analyses, the BBS score was simplified into the three categories “1 = constipation” (scores 1-2), “2 = normal” (scores 3-5) and “3 = diarrhea” (scores 6-7). This classification was chosen to correspond with the Rome IV delineation for the classification of irritable bowel syndrome (IBS) subtypes (IBS-D and IBS-C), being an applicable cut-off both in clinical practice and a research setting.8\n\nFecal samples were collected in the IBSEN III trial from first the inclusions in 2017 and all follow-ups completed through 2022. The BSS was first implemented after a year of inclusion, hence the fecal samples included in the current analysis are those collected from January 2018 until the end of 2022. Fecal samples were delivered fresh at the hospital, after the patients had collected stool samples at home. The patients were instructed to score the stool sample with the BSS at the time of collection. Then, they were instructed to send the sample to the hospital by mail, the same day as defecation or the day after. The hospital received the sample from one to four days after defecation, and then the expert score was performed immediately (upon freezing the sample). Two experienced bioengineers analyzing the fecal samples assigned the expert score. They divided the work between them, hence the expert score is from one expert only, and not a mean value. All the fecal samples included were assessed by both an expert and a patient.\n\nThe IBSEN III study has been reviewed and approved by the Regional Committee for Medical and Health Research Ethics in South-Eastern Norway (reference number 2015/946), and was conductance in accordance with the Declaration of Helsinki. All participants provided written informed consent.\n\nCategorical data are described with counts and percentages. Agreement between experts and patients was assessed using Cohen’s weighted kappa.9 The results are expressed as point estimates with 95% confidence intervals (CI). P-values <0.05 were considered statistically significant. All analyses were performed using SPSS version 28 (https://www.ibm.com/spss).\n\n\nResults\n\nThe study protocol and cohort’s baseline characteristics are presented in detail in a previous publication.7 In total, 2970 stool samples from 1359 different patients were available for inclusion. The study subjects delivered stool samples at baseline (100%), after 3 months (48%), six months (34%), one year (26%) and three years (9%), respectively. Of the available stool samples, 2280 samples (77%) from 992 different patients had BSS scores from both patients and experts and were included in the analysis. 992 samples with matched patient and expert scores were available from baseline (100%), 513 samples from the 3-month follow-up (52% of baseline patients), 363 samples from the six-month follow-up (37% of baseline patients), 290 samples from the one-year follow-up (29% of baseline patients) and 122 samples from the three-year follow-up (12% of baseline patients).\n\nThe agreement between patient and expert scores for all assessed time points was good to substantial when assessed with Cohen’s weighted kappa, ranging from 0.66 to 0.72 (Table 1). Overall, the patients were likely to score lower than the experts. When the scores from the first sample per patient were compared between the patient and the expert ratings, the experts were more likely to assign higher scores compared to the patient’s self-assessment. The proportion of the lowest assigned scores (score 1-2) was 12.1% for patients and 8.1% for the experts. In Table 2, the scores given by the patients and experts are presented for the first assessment time point. The numbers in green represent perfect agreement between the experts and the patients. The numbers in yellow above the diagonal represent the situation when the sample is given a higher score by an expert (over-estimation) and the red numbers below the diagonal represent an under-estimation (experts score lower compared to the patients). The distribution of BSS scores reported by the patient for each value of score given by the expert is presented graphically in Figure 1.\n\nThe figure shows that the patients were likely to score lower than the experts. BSS: Bristol Stool Scale.\n\nWhen the BSS scores were simplified and all the samples categorized as 1 (scores 1 and 2), 2 (scores 3, 4 and 5) or 3 (scores 6 and 7), patient and expert agreement improved slightly, with Cohen’s weighted kappa ranging from 0.73 to 0.77, indicating substantial agreement for all the assessed time points (Table 1). The distribution of all scores given by the patients and experts with the simplified scores and assessment point 1 are presented in Table 3. A perfect agreement between the experts and the patients is highlighted in green, the portion of the table highlighted in yellow represents over-estimation by the experts and the red number represents an under-estimation by the experts. The distribution of the BSS scored by the patient for each value of score given by the expert at the first assessment point when the BSS score was simplified into three categories is depicted in Figure 2.\n\nScore 1 equals original scores 1 and 2 (constipation), scores 2 equals original scores 3, 4 and 5 (normal), and score 3 equals original scores 6 and 7(diarrhea). The figure shows the distribution of the BSS scored by the patient for each score value given by the expert at the first assessment point when the BSS score was simplified into the three categories. BSS: Bristol Stool Scale.\n\n\nDiscussion\n\nBSS is commonly used in both clinical and research settings for the evaluation of stool consistency, however its reliability is unclear as the scores rely on subjective patient reports. Here, we show that there is good to substantial agreement between patient BSS scores and expert BSS scores when evaluating individual scores from 1-7. However, this agreement is even better when scores are compared in simplified categories corresponding to constipation (BSS score 1-2), normal stool (BSS score 3-5) and diarrhea (BSS score 6-7). Our findings highlight that the patient´s exact BSS score is in good agreement with the expert score, and that the patient score is accurate for evaluation of main stool type (constipation, normal or diarrhea). As the main stool type is commonly the one of most interest for the classification of disease, and more subtle distinctions might not be of great clinical importance, we found BSS to be a reliable tool for stool form evaluation.\n\nSeveral studies have previously reported on the validity and use of BSS. Our findings contradict those of a previous study investigating the rate reproducibility of BSS by comparing BSS scores of thirty-four gastroenterology providers of 35 different stool photographs.10 Here, they reported high reliability and agreement when BSS scores were used to assess the individual stool type from 1-7. When the experts should categorized the stool type into the three categories constipation, normal and diarrhea (corresponding to the Rome III standard8), the reliability and agreement decreased.10 Of note, in contrast to these prior results, our simplified into three groups was performed based on the original BSS scores and not independently assessed, hence this might explain the different findings. Also, our study compared subjective patient scores to expert scores, which is relevant as the BSS scoring is most often performed by the patients themselves.\n\nA recent study from 2022 comparing subjective IBS patients´ BSS scores with stool water content as an objective measure reported only modest conformity between methods, highlighting that this can affect the classification of IBS-subtype according to BSS score.11 Similarly, a study from 2016 validated the BSS by measuring stool form in 169 healthy adults and comparing it to stool water content and BSS score form 19 patients with diarrhea-predominant IBS.6 They reported the BSS to demonstrate adequate validity and reliability, however they detected difficulties around clinical decision points for types 2, 3, 5 and 6.6 Although the authors report the BSS to be a valid tool for stool evaluation, it has to be taken into consideration that the uncertainty around scorings affects the classification of stool type into constipation, normal or diarrhea, potentially affecting the clinical outcomes and diagnosis of IBS-subtype.\n\nAccording to current literature highly variable results are thus presented for the validity of the BSS when comparing subjective measures with expert measures or fecal water content. BSS is widely used both in clinical and research settings, and is currently used in an increasing number of studies analyzing microbiota to adjust for differences in fecal consistency.5 The unclear generalizability of BSS represents a challenge to interpreting such results. Interestingly, a recent study comparing a smartphone application using artificial intelligence (AI) to BSS scores performed by two experts reported high accuracy between the AI and the experts. In addition, they reported the trained AI to be superior to subject self-reported BSS scores, emphasising that AI assessments could provide more objective outcome measures for stool characterization in gastroenterology.12 This should be taken into consideration in further studies, as it highlights a tool that can potentially improve the lack of reliability related to the subjective measures used in BSS today.\n\nOur study has some limitations. As the follow-up period for the IBSEN III cohort is still not completed, we did not include stool samples from all subjects at all time points, and some patients were included with several samples, whereas others were only included with one sample. Due to the covid-19 pandemic, the 1-year follow-up was delayed for some of the subjects, hence the proportion of included study subjects with more than three stool samples is small. As the BSS is a subjective measure, the uneven distribution of the number of samples per participant might be a weakness affecting the results. When performing a stool sample, only a small part of the whole stool delivery is sampled for analysis. It is likely that parts of the discharge may have different consistencies and appearance, hence the part sampled for analysis might not be representative of the whole stool. This might be of particular importance when evaluating patients with GI disorders such as IBD and IBS, as the stool might be a mix ranging from hard lumps at first followed by looser stools. Our study also only included two experts. It must be considered that a larger number of experts, or a mean of several expert scores, could have strengthened the results.\n\n\nConclusion\n\nTaken together, our findings show that the agreement between patient and expert BSS scores is good, especially when divided into three main stool categories. We found the BSS to be a reliable tool for the categorization of stool type when the BSS scores are categorized into three categories corresponding to the clinically relevant concepts of constipation, normal stool or diarrhea.\n\nThe IBSEN III study has been reviewed and approved by the Regional Committee for Medical and Health Research Ethics in South-Eastern Norway (reference number 2015/946, approval date 1st July 2015) and was conductance in accordance with the Declaration of Helsinki. All participants provided written informed consent.\n\n\nAuthor contributions\n\nHFD, JV, GHM and JTF planned and designed the study. MLH and VK included patients in the IBSEN III trial. GHM and JTF analysed the fecal samples and performed the expert BSS scoring. MCS performed the statistical analyses. HFD wrote the manuscript. JV, MCS, GHM and JTF contributed to data interpretation and critical revision of the manuscript. All authors reviewed and approved the final manuscript.", "appendix": "Data availability\n\nDryad: Bristol stool scale: patient versus expert score data, https://doi.org/10.5061/dryad.djh9w0w7r. 13\n\nThis project contains the following underlying data:\n\n- BSS_data_deidentified.xlsx\n\n- README.md\n\nData are available under the terms of CC0 1.0 Universal (CC0 1.0) Public Domain Dedication license.\n\n\nAcknowledgements\n\nWe thank the IBSEN III study group.\n\n\nReferences\n\nLewis SJ, Heaton KW: Stool form scale as a useful guide to intestinal transit time. Scand. J. Gastroenterol. Sep 1997; 32(9): 920–924. PubMed Abstract \| Publisher Full Text\n\nTörnblom H, Van Oudenhove L, Sadik R, et al.: Colonic transit time and IBS symptoms: what’s the link? Am. J. Gastroenterol. May 2012; 107(5): 754–760. PubMed Abstract \| Publisher Full Text\n\nBliss DZ, Savik K, Jung H, et al.: Comparison of subjective classification of stool consistency and stool water content. J. Wound Ostomy Continence Nurs. May 1999; 26(3): 137–141. PubMed Abstract \| Publisher Full Text\n\nHeaton KW, O’Donnell LJ: An office guide to whole-gut transit time. Patients’ recollection of their stool form. J. Clin. Gastroenterol. Jul 1994; 19(1): 28–30. PubMed Abstract \| Publisher Full Text\n\nVork L, Wilms E, Penders J, et al.: Stool Consistency: Looking Beyond the Bristol Stool Form Scale. J. Neurogastroenterol. Motil. Oct 30 2019; 25(4): 625. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBlake MR, Raker JM, Whelan K: Validity and reliability of the Bristol Stool Form Scale in healthy adults and patients with diarrhoea-predominant irritable bowel syndrome. Aliment. Pharmacol. Ther. Oct 2016; 44(7): 693–703. PubMed Abstract \| Publisher Full Text\n\nKristensen VA, Opheim R, Perminow G, et al.: Inflammatory bowel disease in South-Eastern Norway III (IBSEN III): a new population-based inception cohort study from South-Eastern Norway. Scand. J. Gastroenterol. Aug 2021; 56(8): 899–905. PubMed Abstract \| Publisher Full Text\n\nMearin F, Lacy BE, Chang L, et al.: Bowel Disorders. Gastroenterology. Feb 18 2016; 150: 1393–1407.e5. Publisher Full Text\n\nLandis JR, Koch GG: The measurement of observer agreement for categorical data. Biometrics. Mar 1977; 33(1): 159–174. Publisher Full Text\n\nChumpitazi BP, Self MM, Czyzewski DI, et al.: Bristol Stool Form Scale reliability and agreement decreases when determining Rome III stool form designations. Neurogastroenterol. Motil. Mar 2016; 28(3): 443–448. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNordin E, Hellström PM, Brunius C, et al.: Modest conformity between self-reporting of Bristol stool form and fecal consistency measured by stool water content in irritable bowel syndrome, a FODMAP and gluten trial. Am. J. Gastroenterol. Aug 12 2022; 117: 1668–1674. PubMed Abstract \| Publisher Full Text\n\nPimentel M, Mathur R, Wang J, et al.: A Smartphone Application Using Artificial Intelligence Is Superior to Subject Self-Reporting When Assessing Stool Form. Am. J. Gastroenterol. Mar 14 2022; 117: 1118–1124. PubMed Abstract \| Publisher Full Text\n\nValeur J: Data from: Bristol stool scale: Patient versus expert score data. [Dataset]. Dryad. 2024. Publisher Full Text" }	[ { "id": "315298", "date": "24 Aug 2024", "name": "Bodil Ohlsson", "expertise": [ "Reviewer Expertise Gastroenterology" ], "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 study aimed to compare the assessment of coherence between the patients and the experts scores regarding Bristol Stool Scale (BSS). 2280 fecal samples collected from 992 patients with inflammatory bowel disease (IBD) at up to five time points for 3 years were included. Agreement between patient and expert scores was assessed using Cohen’s kappa. There was good substantial agreement between patient and expert scores (Cohen’s weighted kappa: 0.66-0.72). When the BSS scores were simplified and categorized into 3 groups instead of 7, the agreement improved slightly (Cohen’s weighted kappa: 0.73-0.77). To summarize, the agreement between patient and expert BSS scores is good to substantial, especially when the BSS scores are simplified into three categories: constipation, normal stool and diarrhea.\nRemarks: In the discussion, it is stressed that these results with good coherence between patients and expert scores differ from a previous study performed in IBS. I think that IBS and IBD patients differ a lot. IBS patients may have greater psychological influence on their symptoms and may interpret their symptoms in other ways than IBS patients with more obvious organic changes. The difference between these 2 patient cohorts and psychological aspects regarding self-reported assessment should be discussed.\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": "12950", "date": "19 Dec 2024", "name": "Jørgen Valeur", "role": "Author Response", "response": "We would like to thank the Reviewer for taking the time to read and comment on our paper, and are grateful for both the encouraging comments, as well as the valuable suggestions. In our revised version, we have now included the following sentence in Discussion: \"It should be emphasized that self-reported assessments may differ between IBS patients and IBD patients, since psychological factors may have a greater influence of IBS symptoms than IBD symptoms.\"" } ] }, { "id": "344492", "date": "11 Dec 2024", "name": "Friedemann Erchinger", "expertise": [ "Reviewer Expertise exocrine pancreatic insufficiency", "fecal fat", "fecal output" ], "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 agreement between patients and experts when scoring stool consistency after the Bristol Stool Scale. Using Cohen’s weighted kappa for all scales 1 to 7 or grouped scales, constipation (1-2) normal (3-5) and diare (6,7), agreement was substantial; grouped scales show an even better agreement.\nTo use easy methods which can be practiced by patients and experts is a needed tool in a clinical setting and in research. This makes this study important.\nNot all authors relate BSS value 5 to normal consistency. Some categorise value 5 as \"Borderline\" (normal/diarrhoea), others as diarrhoea. Perhaps this should be discussed in the article.\nIt would have been attractive to show interobserver and intraobserver variability in BSS scoring, both in patients (with relatives as interobservers?) and experts. This would have strengthened the study substantially. Take this point in in \"limitations\"?\nAs discussed can stool frequency give different stool consistency. Perhaps agreement got better if experts knew about stool frequency? Discuss this point in \"limitations\"?\nAs discussed is stool delivered in one session not homogeneous, stool samples mailed to scoring can contain only a part of the entire stool delivery. -This could be overcome by photographs of the stools. Discuss this in \"future considerations\"? -As patients look at the entire stool delivery and experts only at a small sample it is very interesting that agreement can be classified as substantial. This should be highlighted in the article.\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": "12951", "date": "19 Dec 2024", "name": "Jørgen Valeur", "role": "Author Response", "response": "We would like to thank the Reviewer for taking the time to read and comment on our paper, and are grateful for both the encouraging comments, as well as the valuable suggestions. Our point-by-point response is as follows: 1. \"Not all authors relate BSS value 5 to normal consistency. Some categorise value 5 as \"Borderline\" (normal/diarrhoea), others as diarrhoea. Perhaps this should be discussed in the article.\" Response: We agree, and have now included this sentence in Methods: \"It should be noted, however, that some authors consider a score of 5 as abnormal.\" 2. \"It would have been attractive to show interobserver and intraobserver variability in BSS scoring, both in patients (with relatives as interobservers?) and experts. This would have strengthened the study substantially. Take this point in in \"limitations\"?\" Response: We agree that this would have been valuable information to include, and have now taken this point in in \"limitations\", as suggested: \"Similarly, including an assessment of interobserver and intraobserver variabilities, both in the patient group and among the experts, would have strenghtened our study substantially.\" 3. \"As discussed can stool frequency give different stool consistency. Perhaps agreement got better if experts knew about stool frequency? Discuss this point in \"limitations\"?\" Response: We agree that it would have been valuable to know the stool frequencies of the participants. We have now included this sentence in \"limitations\": \"It would have been interesting to know the participants stool frequency, since the frequency may affect the consistency.\" 4. \"As discussed is stool delivered in one session not homogeneous, stool samples mailed to scoring can contain only a part of the entire stool delivery. -This could be overcome by photographs of the stools. Discuss this in \"future considerations\"? -As patients look at the entire stool delivery and experts only at a small sample it is very interesting that agreement can be classified as substantial. This should be highlighted in the article.\" Response: We would like to thank the Reviewer for pointing this out, and have now included the following sentence in Conclusion: \"Indeed, our finding that expert scoring of only a small fecal sample is in good agreement with the patient scoring of the entire stool delivery is a novel observation that should be validated in future studies where photographs of the entire stool delivery should also be included.\" Althogether, we now sincerely hope that the Reviewer may find our paper to deserve the status as \"Approved\"." } ] } ]	1	https://f1000research.com/articles/13-833
https://f1000research.com/articles/13-1531/v1	18 Dec 24	{ "type": "Research Article", "title": "Swedish abdominal massage versus warm water therapy on postoperative constipation: a comparison quasi-experimental study", "authors": [ "Enny Selawaty Boangmanalu", "Masfuri Masfuri", "Muhamad Adam", "Sri Nining", "Triani Banna", "Indira Mastura Pulungan", "Enny Selawaty Boangmanalu", "Muhamad Adam", "Sri Nining", "Triani Banna", "Indira Mastura Pulungan" ], "abstract": "Background Postoperative immobilizatiton for patients with lower extremity fractures causes constipation, which usually affects 50–70% of patients. When it comes to nursing interventions for postoperative constipation, Swedish abdominal massage and warm water drinking therapy are two possible options.\n\nAim The objective of this study is to compare the effectiveness of drinking warm water and Swedish abdominal massage on constipation scores on post-operative lower extremity fractures.\n\nMethods A quasi-experimental pre-posttest design without control group design was applied. 30 respondents used simple random sampling technique. The Constipation Assessment Scale (CAS) questionnaire was used to assess the patient’s constipation levels before and after the intervention. The data analysis used independent t-test.\n\nResults The mean score of constipation of drinking water group after the intervention was 4.60 while abdominal Swedish massage was 3.56. Although both significantly reduced the constipation score, the p-value was 0.00.\n\nConclusion The protocol of drinking warm water and Swedish abdominal massage immediately after waking up effectively reduced constipation scores on postoperative lower extremity fracture patients and can be use to adjuvant therapy. Further studies are needed to investigate postoperative constipation patients with immobility and the use of strong analgetics.", "keywords": [ "Closed Fracture Reduction", "Constipation", "Postoperative", "Swedish Abdominal Massage", "Warm Water Therapy" ], "content": "Background\n\nAccording to the most recent data published in Lancet Healthy Longev, there were 178 million new fractures globally in 2019, and 455 million people had acute or chronic fracture symptoms at some point in their lives (Global Burden of Disease, 2021). However after surgery, prolonged bed rest and limited movement inevitably result in adverse effects such as constipation (Viberg et al., 2022). Constipation, which typically affects 50–70% of patients, is brought on by the body’s immobile state following surgery, which weakens intestinal peristalsis function in conjunction with dietary factors (Jing & Jia, 2019). Patients should be incredibly concerned about constipation because it can lead to a number of issues and negatively impact their quality of life (Kamali et al., 2022). Consequently, constipation is a serious problem that, if disregarded, can result in psychological and physical problems. Laxative use is common among patients due to discomfort. Research shows that constipation affects 3% to 27% of people in the general population on an incidence rate of 5; in hospital settings, the prevalence is 79% (Pehlivan & Nural, 2022). However, in acute clinical practice, this condition is frequently overlooked when providing patient care (Trads et al., 2018).\n\nNurses have been treating constipation with a nursing intervention that includes teaching patients about high-fiber foods like vegetables and papaya (Galica et al., 2022). But there has never been an application of another autonomous nursing intervention in the treatment room, like abdominal massage (Nouhi et al., 2022). It is relatively inexpensive and something that sufferers can do on their own. Constant direct pressure is applied to the abdominal wall during abdominal massage, which is followed by a relaxation period. This increases the contractions of the rectum and intestines and the gastrocolic reflex (Yao et al., 2020). Swedish abdominal massages are performed with light pressure on the tissue to promote comfort and enhance the digestive and blood circulation systems (Park et al., 2023). By altering stomach pressure through mechanical and reflexive processes, abdominal massage can accelerate the passage of food through the digestive system and promote peristaltic movements (Fekri et al., 2021). This procedure will speed up the movement of food through the digestive tract by increasing peristaltic movements and altering stomach pressure through mechanical and reflexive means (Lafcı & Kaşikçi, 2023).\n\nTo fill the stomach capacity, water is a good option. Using warm water as a complementary therapy is recommended (Kilroe et al., 2024). The effects of hydrostatic, hydrodynamic, and warmth can promote relaxation and better blood circulation. The body responds to water to prevent, correct, and improve human health status, which makes water therapy one of the natural healing systems (Asmaa Sayed et al., 2018). Warm water consumption will quicken the body’s temperature regulation process because it requires less energy. Constipation will speed up the defecation process by using warm water to soften the stool (Parsons et al., 2022). The aim of this study is to compare the effectiveness of drinking warm water and Swedish abdominal massage on constipation scores on post-operative lower extremity fractures.\n\n\nMethods\n\nThis research used quasi-experimental and pre-posttest designs without control group. This research was conducted at the Idaman Regional Public Hospital, Banjarbaru, South Kalimantan, Indonesia from May 10th to June 30th 2023.\n\nThe sample was 30 respondents selected using simple random technique. Determination of the sample size of each group is determined by calculating the paired numerical comparative formula. The sample in this research selected randomly using a lottery. Randomization is done by lottery, which makes a list of all subjects to be studied, gives a number code to each item to be investigated, writes the code on a small paper, rolls up each paper, puts the rolled paper into a container then shakes or shakes the container and takes one by one the roll. If you get an odd number, the warm water consumption intervention will be carried out while if you get an even number, you will enter the Swedish abdominal massage intervention. The inclusion criteria were: (1) all post-operative orthopedic surgery patients with lower extremity fractures (fractures of the femur, pelvis, tibia, fibula, ankle, and pedis), (2) patients with casts, (3) post-operative orthopedic surgery patients with ORIF, (4) patients aged 17-60 years, (5) compos-mentis patient, (5) patients with post-operative day 1 (after 1×24 hours/POD 1), (6) patients with a Barthel index score ≤ 8 (severe dependence), (7) patients receiving a standard diet from the hospital.\n\nThis research used demographic questionnaire and the Constipation Assessment Scale (CAS) questionnaire to assess the patients’ constipation levels before and after the intervention. We used the Constipation Assessment Scale (CAS) that develop by McMillan and Williams and translated into Indonesian version by Suwandi. The valid CAS instrument in the Indonesian version consists of eight questions (Suwandi, 2017).\n\nThe sample was split into two intervention groups: group 2 got a Swedish abdominal massage, and group 1 drank warm water. 500 ml of warm water in a glass was consumed by group 1 at a temperature of 31.5°C. For three days, they drank the water as soon as they woke up and before breakfast. They were then permitted to eat breakfast 30 to 45 minutes after they had finished drinking. Over the course of three days, the second group, which was given a Swedish abdominal massage, had their abdominal muscles massaged for fifteen to twenty minutes. The moment when the respondents woke up, they were received this massage. It is recommended that patients refrain from eating breakfast following the intervention; however, they are permitted to consume food 30 to 45 minutes following the procedure. Every intervention was examined over the course of three days; the pretest was given prior to the intervention, and the post test was given following the third day of the intervention. The first time the respondent was able to defecate was used to calculate the defecation time.\n\nThe data were collected by the researchers. Questionnaires were distributed to each group before and after intervention.\n\nDescriptive statistical tests were used to measure demographic data or respondents’ characteristics. Data analysis was done with Windows-based IBM SPSS (Version 26.0) (IBM Corp, 2024). The data were normally distributed according to Shapiro-Wilk (p≥0.05), so the independent t-test was used to measure constipation scores. The data were normally distributed according to Shapiro-Wilk (p≥0.05), so the independent t-test was used to measure constipation scores.\n\n\nResults\n\nThe participants’ characteristics such as age, gender, types of analgetics and constipation score were collected at baseline on Table 1.\n\nAccording to this study, the majority of respondents—11 (73.3%) and nine (60%), respectively—were men who participated in the warm water intervention group and the Swedish abdominal massage intervention group. The first intervention group’s average age was 28.93 years, while the second intervention group’s average age was 31.53 years. The first intervention group’s confidence interval fell between 17 and 60 years, while the second intervention group’s confidence interval fell between 17 and 58 years. In both groups, non-opioid analgesics were used 100% of the time as analgesics. Prior to the intervention, the group receiving a Swedish abdominal massage had a higher mean constipation score than the group drinking warm water. On the other hand, the Swedish abdominal massage intervention group had a lower mean constipation score following the intervention. The statistical test found significant differences between the two intervention groups’ respondent characteristics (gender, age, and type of analgesic; p>0.05), but not between the groups’ pre- and post-intervention characteristics.\n\nThe difference in constipation score was measured by looking at the mean score of the drinking warm water group and the Swedish abdominal massage group on Table 2.\n\na Mean (SD).\n\nb Independent t-test.\n\n* Significant at p<0.05.\n\nThe independent t-test discovered a significantly different constipation scores between the group drinking warm water and the group having Swedish abdominal massage (p<0.05). This result concludes a significantly different constipation score of the two groups. Prior to the intervention, the group receiving a Swedish abdominal massage scored higher (6.53) on the constipation score than the group drinking warm water (3.80). In the meantime, the warm water drinking group after the intervention scored higher on the constipation score (1.85) than the other group (1.68). Additionally, compared to the intervention group receiving Swedish abdominal massage, the warm water drinking group experienced a lower mean difference in changes to their constipation scores. When receiving a Swedish abdominal massage, the average time spent defecating was reduced to 54 hours, as opposed to 58.67 hours for the intervention group that drank warm water.\n\n\nDiscussion\n\nThe results showed that Swedish abdominal massage was noticeably more successful than warm water drinking therapy in reducing the constipation score. In addition to being an effective treatment for constipation, a Swedish abdominal massage can also lessen the degree of straining, anal pain, and bloating, as well as the degree of incomplete bowel emptying (Choi et al., 2021). By strengthening the abdominal muscles and encouraging intestinal peristalsis, massage can improve the digestive system’s efficiency, improving quality of life ratings and resulting in better-consistent stools (Durmuş İskender & Çalışkan, 2022). In order to cause contraction of the longitudinal and circular muscles—the circular muscle encircling the lumen—afferent neuron stretch receptors in the luminal wall are activated when a smooth muscle segment is distended with a pressure of about 2 mmHg. Simultaneously, the intestinal wall muscles contract a few centimeters in the area of higher pressure, while the muscles relax below the point of stimulation. These sensory neurons and nerve endings can be stimulated by abdominal massage (Aydinli & Karadağ, 2023; Faghihi et al., 2021). As a result, pressure is applied to the chyme, forcing it forward and causing the subsequent intestinal wall muscle segment to expand or contract. This, in turn, causes another contraction and produces a peristaltic wave (Keely & Barrett, 2022).\n\nFor three days in a row, consuming 500 ml of warm water first thing in the morning can intensify the gastric effect and heighten the feeling of passing gas (Kilroe et al., 2024). The best time to trigger the gastrocolic reflex is in the morning. This reflex happens when the extrinsic autonomic nerve, which is responsible for promoting colonic motility and large amplitude propagation (HAPCs) to ward off constipation, contracts the stomach when it reaches a specific volume (500 ml) (Al-Kharraz et al., 2023). One to three glasses of water a day can help trigger the gastric reflex. Utilizing warm water is a complementary therapy. The warm, hydrostatic, and hydrodynamic effects can promote relaxation and better blood circulation (Asmaa Sayed et al., 2018).\n\nSignificant results were found in the majority of research studies that used warm water consumption at different volumes in conjunction with Swedish abdominal massage. The hypothesis posits that upon food entry, the large intestine experiences mass movements primarily due to the gastrocolic reflex (Bellini et al., 2021). This reflex is facilitated by gastrin and parasympathetic innervation, which travel from the stomach to the large intestine (Chatip et al., 2024). The urge to urinate frequently follows this reflex, which is most noticeable in many people after their first meal of the day. Therefore, a reflex is triggered to move the contents of the digestive tract farther along when new food enters, making room for the incoming food (Gu et al., 2023). The remaining small intestine’s contents are moved into the large intestine by the gastroileal reflex, and the defecation reflex is triggered by the gastrocolic reflex, which pushes the large intestine’s contents into the rectum. This is so that the digestive system can function more efficiently. Peristalsis, or the stimulation of intestinal peristaltic movements, will be strengthened by abdominal massage (Karaaslan et al., 2024).\n\nWith the help of gastrin from the stomach and the extrinsic autonomic nerve, direct stimulation of the abdominal muscles can induce peristalsis and the gastrocolon, which causes the colon’s mass to accelerate and the stomach’s contractions to strongen (Chatip et al., 2024). A gastroileal reflex will result in the movement of the remaining contents of the small intestine to the large intestine by forcing the chyme into the duodenum through strong peristaltic movements (Durmuş İskender & Çalışkan, 2022). This will speed up the absorption process in the intestine. A colon’s contents will be forced into the rectum, a gastrocolon effect will be produced, stretch receptors in the rectal wall will be stimulated, and a feeling of defecation will be produced through good intestinal motility (Artale et al., 2023).\n\n\nConclusion\n\nBoth Swedish abdominal massage and warm water therapy are effective interventions for managing postoperative constipation. Each method offers specific advantages and can be considered based on patient preference and clinical context. Further research exploring long-term outcomes and comparative effectiveness with larger sample sizes would provide additional insights into optimizing postoperative care protocols for constipation management. This study’s limitation was that, despite the respondents’ following hospital guidelines for diet, the amount of fiber and food portions were not calculated. Additionally, the location of the fracture could have an impact on mobilization; a more distal fracture location would facilitate easier food mobilization. A more proximal fracture location, however, will hinder the mobilization of food.\n\n\nEthical consideration\n\nThis research was approved by the Ethics Commission of Idaman Regional Public Hospital, Banjarbaru with number No. RS00214/KEPK-RSDI/04/2023 on May 1st 2023. Before conducting the research, the researcher explained to the respondents about the objectives, procedures, and expectations of this research in person verbal and written. The respondents were asked to provide their written consent in an informed consent by signing before participating in this research. The respondents were also assured that their involvement was voluntary and were informed of their right to withdraw from the study at any time without facing any penalties.", "appendix": "Data availability\n\nFighshare: Swedish Abdominal Massage versus Warm Water Therapy on Postoperative Constipation: A Comparison Quasi Experimental Study: 10.6084/m9.figshare.27764655.v2 (Boangmanalu, 2024).\n\nThis project contains the following underlying data Click or tap here to enter text.\n\n1. SPO of Water Consumption\n\n2. SPO of Abdominal Massage\n\n3. SPSS Output\n\n4. All Data consist of all raw data underlying data before analysis in SPSS file\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nFighshare: Swedish Abdominal Massage versus Warm Water Therapy on Postoperative Constipation: A Comparison Quasi Experimental Study: 10.6084/m9.figshare.27764655.v2 (Boangmanalu, 2024).\n\nThis project contains the following extended dataClick or tap here to enter text.:\n\n1. Consort cheklist\n\n2. Research instrument\n\n3. Informed Consent\n\n4. Research explanation\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nFighshare: CONSORT checklist for Swedish Abdominal Massage versus Warm Water Therapy on Postoperative Constipation: A Comparison Quasi Experimental Study: 10.6084/m9.figshare.27764655.v2 (Boangmanalu, 2024).\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 successfully conducted with the support of various parties. We want to thank the hospital where the study was carried out and the Faculty of Nursing, University of Indonesia. We also extend our gratitude to all participants and their families for their support in making this study possible.\n\n\nReferences\n\nAl-Kharraz K, Tabbah MJ, LaChance J, et al.: The Effect of the Flint Water Crisis Secondary to Increased Lead Levels in Drinking Water on Constipation in Children in the City of Flint, Michigan, USA. Cureus. 2023; 15(8): e44189. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nArtale S, Terzoni S, Destrebecq A, et al.: Abdominal massage and laxative use for constipation: a pilot study. Br. J. Nurs. 2023; 32(14): 666–671. PubMed Abstract \| Publisher Full Text\n\nAsmaa Sayed AR, Abdelmowla A, El-rahim TAEA: Effects of Warm Water Sitz Bath on Post-Hemorrhoidectomy Symptoms Effects of Warm Water Sitz Bath on Post-Hemorrhoidectomy Symptoms. IOSR Journal of Nursing and Health Science (IOSR-JNHS). 2018; 7(IV): 57–65. Publisher Full Text\n\nAydinli A, Karadağ S: Effects of abdominal massage applied with ginger and lavender oil for elderly with constipation: A randomized controlled trial. Explore (N.Y.). 2023; 19(1): 115–120. PubMed Abstract \| Publisher Full Text\n\nBellini M, Tonarelli S, Barracca F, et al.: Chronic Constipation: Is a Nutritional Approach Reasonable? Nutrients. 2021; 13(10). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBoangmanalu ES: Swedish Abdominal Massage versus Warm Water Therapy on Postoperative Constipation: A Comparison Quasi-Experimental Study. figshare Journal contribution. 2024. Publisher Full Text\n\nChatip AT, Acar G, Akçay AA: Comparison of the effects of abdominal massage and osteopathic manipulative treatment home program on constipation in children with cerebral palsy. JGH Open. 2024; 8(6): e13102. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChoi YI, Kim KO, Chung J-W, et al.: Effects of Automatic Abdominal Massage Device in Treatment of Chronic Constipation Patients: A Prospective Study. Dig. Dis. Sci. 2021; 66(9): 3105–3112. PubMed Abstract \| Publisher Full Text\n\nDurmuş İskender M, Çalışkan N: Effect of Acupressure and Abdominal Massage on Constipation in Patients with Total Knee Arthroplasty: A Randomized Controlled Study. Clin. Nurs. Res. 2022; 31(3): 453–462. PubMed Abstract \| Publisher Full Text\n\nFaghihi A, Najafi SS, Hashempur MH, et al.: The Effect of Abdominal Massage with Extra-Virgin Olive Oil on Constipation among Elderly Individuals: A Randomized Controlled Clinical Trial. Int. J. Community Based Nurs. Midwifery. 2021; 9(4): 268–277. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFekri Z, Aghebati N, Sadeghi T, et al.: The effects of abdominal “I LOV U” massage along with lifestyle training on constipation and distension in the elderly with stroke. Complement. Ther. Med. 2021; 57: 102665. PubMed Abstract \| Publisher Full Text\n\nGalica AN, Galica R, Dumitrașcu DL: Diet, fibers, and probiotics for irritable bowel syndrome. J. Med. Life. 2022; 15(2): 174–179. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGlobal Burden of Disease: Global, regional, and national burden of bone fractures in 204 countries and territories, 1990-2019: a systematic analysis from the Global Burden of Disease Study 2019. Lancet Healthy Longev. 2021; 2(9): e580–e592. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGu X, Zhang L, Yuan H, et al.: Analysis of the efficacy of abdominal massage on functional constipation: A meta-analysis. Heliyon. 2023; 9(7): e18098. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIBM Corp: SPSS Statistics V26 (2.6). IBM Corp; 2024. Reference Source\n\nJing D, Jia L: Assessment of patients’ psychological state and self-efficacy associated with postoperative constipation after thoracolumbar fracture surgery. J. Int. Med. Res. 2019; 47(9): 4215–4224. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKamali M, Bagheri-Nesami M, Ghaemian A, et al.: The Effect of Acupressure on Preventing Constipation in Patients with Acute Myocardial Infarction under Primary Percutaneous Coronary Intervention. Middle East J. Dig. Dis. 2022; 14(4): 422–430. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKaraaslan Y, Karakus A, Ogutmen Koc D, et al.: Effectiveness of Abdominal Massage Versus Kinesio Taping in Women With Chronic Constipation: A Randomized Controlled Trial. J. Neurogastroenterol. Motil. 2024; 30: 501–511. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKeely SJ, Barrett KE: Intestinal secretory mechanisms and diarrhea. Am. J. Physiol. Gastrointest. Liver Physiol. 2022; 322(4): G405–G420. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKilroe MJ, McAtee KJ, McReynolds TM: A Case Report of Food Impaction Relieved by Warm Water Drinking Therapy. J. Emerg. Med. 2024; 66(1): e27–e28. PubMed Abstract \| Publisher Full Text\n\nLafcı D, Kaşikçi M: The effect of aroma massage on constipation in elderly individuals. Exp. Gerontol. 2023; 171: 112023. PubMed Abstract \| Publisher Full Text\n\nNouhi E, Mansour-Ghanaei R, Hojati SA, et al.: The effect of abdominal massage on the severity of constipation in elderly patients hospitalized with fractures: A randomized clinical trial. Int. J. Orthop. Trauma Nurs. 2022; 47: 100936. PubMed Abstract \| Publisher Full Text\n\nPark Y-G, Kim BS, Kang K-T, et al.: Effects of Abdominal Massage for Preventing Acute Postoperative Constipation in Hip Fractures: A Prospective Interventional Study. Clin. Orthop. Surg. 2023; 15(4): 546–551. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nParsons IT, Hockin BCD, Taha OM, et al.: The effect of water temperature on orthostatic tolerance: a randomised crossover trial. Clinical Autonomic Research: Official Journal of the Clinical Autonomic Research Society. 2022; 32(2): 131–141. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPehlivan Z, Nural N: Prevalence of constipation in patients undergoing chemotherapy and the effect of constipation on quality of life. Support. Care Cancer. 2022; 30(10): 8019–8028. PubMed Abstract \| Publisher Full Text\n\nSuwandi: Pengaruh Pijat Perut terhadap Tingkat Konstipasi pada Lansia di Balai Pelayanan Sosial Tresna Werdha Unit Budhi Luhur Yogyakarta [Skripsi]. Universitas Airlangga; 2017.\n\nTrads M, Deutch SR, Pedersen PU: Supporting patients in reducing postoperative constipation: fundamental nursing care - a quasi-experimental study. Scand. J. Caring Sci. 2018; 32(2): 824–832. PubMed Abstract \| Publisher Full Text\n\nViberg B, Erlandsen Claville LU, Andersen LR, et al.: Standardized, Coordinated Care in Nursing Homes Lowers Rehospitalization After Hip Fracture. J. Am. Med. Dir. Assoc. 2022; 23(4): 596–600. PubMed Abstract \| Publisher Full Text\n\nYao F, Zhang Y, Kuang X, et al.: Effectiveness and Safety of Moxibustion on Constipation: A Systematic Review and Meta-Analysis. Evid. Based Complement. Alternat. Med. 2020; 2020: 8645727. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "357292", "date": "25 Jan 2025", "name": "Nilton Carlos Machado", "expertise": [ "Reviewer Expertise Pediatric Gastroenterology", "Hepatology 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 present an enjoyable alternative for treating constipation. This study observed that its application was restricted to patients undergoing postoperative orthopedic surgery for lower extremity fractures (the femur, pelvis, tibia, fibula, ankle, and pedis). The study was elegantly designed and executed with great skill and accomplishment. The small number of patients is noteworthy. The term \"Closed Fracture Reduction\" does not fit well with the study. Perhaps a more fitting term could be suggested. Title. Please insert \"postoperative orthopedic surgery\" since the sample is restricted to this type of surgery. Please include the keywords in the abstract, title, and conclusions to increase the study's visibility. Results. Avoid repeating information in the text and Tables. In this manuscript, I think the variable \"Gender\" can be replaced by \"Sex.\" Conclusions. Adequate e also concerning further studies.\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/13-1531
https://f1000research.com/articles/13-109/v1	19 Feb 24	{ "type": "Research Article", "title": "Eye-gesture control of computer systems via artificial intelligence", "authors": [ "Nachaat Mohamed" ], "abstract": "Background Artificial Intelligence (AI) has the potential to significantly enhance human-computer interactions. This paper introduces a cutting-edge method for computer control using eye-gesture recognition.\n\nMethods Our system employs a sophisticated algorithm to accurately interpret eye movements, converting them into actionable commands. This technology not only improves accessibility for individuals with physical impairments, but also offers a more intuitive interaction mode for the general user base.\n\nResults We tested our method using a comprehensive dataset and achieved a remarkable accuracy rate of over 99.6283% in translating eye gestures into functional commands. Our system utilizes a variety of tools, including PyCharm, OpenCV, mediapipe, and pyautogui, to achieve these results.\n\nConclusions We discuss potential applications of our technology, such as in the emerging field of gesture-controlled weaponry, which could have significant implications for military and rescue operations. Overall, our work represents a substantial step forward in integrating AI with human-computer interaction, enhancing accessibility, improving user engagement, and unlocking innovative applications for critical industries.", "keywords": [ "Artificial Intelligence", "Computers", "Gestures", "OpenCV", "Python", "Pyautogui" ], "content": "Introduction\n\nHuman-Computer Interaction (HCI) has evolved significantly from its inception, which featured punch cards and command line interfaces, to today’s sophisticated Graphical User Interfaces (GUIs) and Natural Language Processing (NLP) technologies. Despite these advancements, traditional input devices such as keyboards and mice have limitations, particularly for users with motor impairments.1 Eye-tracking technologies, which interpret users’ intentions through ocular movement analysis, present a promising solution to these challenges.2 However, realizing their full potential requires the integration of Artificial Intelligence (AI) to accurately interpret nuanced eye movements. This paper introduces an AI-enhanced system for computer control using eye gestures. By harnessing advanced computer vision and machine learning techniques, we translate users’ eye and facial gestures into precise computer commands.3,4 Such eye-gesture systems not only promise more intuitive interactions but also offer ergonomic benefits, representing a departure from traditional input devices.5,6 Their potential is particularly significant for individuals with disabilities, such as mobility challenges or spinal cord injuries, as they provide an alternative means of control.7 Furthermore, these systems are beneficial for professionals like surgeons or musicians who require hands-free computer interactions.8 The market is currently filled with eye-gesture systems that employ various technologies.9,10 However, our AI-driven approach aims to set a new benchmark. Figure 1 shows the Comparison of Ease of Use between Traditional Input Devices and Eye-Tracking Technologies for Different User Group.\n\nWe posit that our methodologies could revolutionize HCI, fostering a more accessible and intuitive user experience.11 Moreover, our research opens the door to innovative applications such as gesture-based weaponry systems.\n\nIn the evolving landscape of Human-Computer Interaction (HCI), ensuring seamless and intuitive interactions is paramount, especially for users with physical impairments or specialized professional requirements.12 While traditional input devices such as keyboards and mice have served a majority of users effectively, they present inherent limitations for certain cohorts. These limitations underscore the need for alternative interaction paradigms. Eye-gesture technologies have emerged as potential candidates to bridge this gap. However, existing eye-gesture systems, although varied in their technological foundations, often lack the sophistication required to interpret a wide array of user intentions accurately and responsively. The challenge lies in harnessing the full potential of eye-tracking technologies by integrating advanced Artificial Intelligence (AI) capabilities, ensuring precise interpretation of eye movements, and translating them into actionable computer commands. Addressing this challenge is imperative to create a universally accessible and efficient HCI platform, capable of catering to a diverse range of users and use-cases.\n\nArtificial Intelligence (AI) has evolved into a comprehensive domain, influencing a myriad of sectors. A compelling facet within this expansive realm is AI gestures: the mimicked non-verbal cues generated by AI systems, aimed at fostering human-like interactions. These gestures, characterized by actions such as waving, nodding, or pointing, enhance the depth of human-AI communication, drawing from advanced technologies like robotics, computer vision, and natural language processing.13,14 The potency of AI gestures is amplified by leveraging the powerful programming language, Python. Its rich assortment of libraries, such as NumPy, Pandas, and scikit-learn, facilitates diverse functionalities crucial for AI and machine learning applications.15,16 Central to AI gesture recognition is the library OpenCV (Open Source Computer Vision). Originating from Intel’s innovation and now under Itseez’s stewardship, OpenCV encompasses an extensive suite of over 2,500 computer vision and machine learning algorithms. Its capabilities span facial recognition, object detection, tracking, and more, finding application across industries like robotics, healthcare, security, and entertainment.17,18 Enthusiasts and professionals can leverage OpenCV’s robust documentation, tutorials, and a wealth of external resources to harness its full potential.19\n\nIn today’s rapidly digitizing world, the very essence of human-computer interaction is undergoing significant evolution.20 As our reliance on digital systems amplifies, there’s a pressing need to make these interactions more intuitive, accessible, and versatile. The conventional modalities—keyboards, mice, touchscreens, while revolutionary in their own right, present inherent limitations.21 These limitations become especially pronounced when considering populations with specific needs or challenges, such as those with motor impairments.22 The quest for inclusivity in technology beckons innovations that can be seamlessly integrated into the lives of all individuals, irrespective of their physical capacities. Eye-gesture recognition emerges as a beacon of promise in this quest. The human eye, a marvel of nature, not only perceives the world but can also communicate intent, emotion, and directives. Harnessing this potential could redefine the paradigms of interaction, enabling users to convey commands or intentions to machines just by moving their eyes. Imagine a world where, with a mere glance, individuals can operate their devices, access information, or even control their home environments. The implications are transformative not just as a novel method of interaction but as a lifeline of autonomy for those who’ve traditionally been dependent on others for even the most basic digital tasks. Moreover, the contemporary technological landscape, enriched by the advancements in Artificial Intelligence (AI), presents an opportune moment for such innovations. AI, with its ability to learn, interpret, and predict, can elevate eye-gesture systems from being mere interpreters of movement to intelligent entities that understand context, nuance, and subtleties of human intent. Yet, for all its promise, the realm of eye-gesture recognition remains a burgeoning field with vast unexplored potentials. The convergence of AI and eye-tracking technologies could spawn a revolution, akin to the leaps we’ve witnessed with touch technologies and voice commands. It is this potential for transformative impact, the prospect of bridging gaps in accessibility, and the allure of uncharted technological frontiers that serves as the driving motivation behind our research.\n\nGesture recognition has its roots in early computer vision studies, with VPL Research being among the first to market a data glove as a gesture input device in the 1980s.1,23 This pioneering work was expanded upon by Freeman and Roth, who used orientation histograms for hand gesture recognition, laying foundational methodologies for future research.24 O’Hagan et al. documented another breakthrough in 1996 when they applied Hidden Markov Models (HMM) to hand gesture recognition, introducing statistical methods to the domain.2,25 The Microsoft Kinect, launched in 2010, was a game-changer for gesture-based HCI. Its depth camera and IR sensor allowed for full-body 3D motion capture, object recognition, and facial recognition, marking a significant step forward in home-based gesture recognition systems.26 Meanwhile, the Leap Motion controller, a compact device capable of detecting hand and finger motions, allowed for fine-grained gesture recognition and was integrated into virtual reality setups to provide natural hand-based controls.3,27 From the algorithmic perspective, Random Decision Forests (RDF) played a crucial role in the success of Kinect’s skeletal tracking capabilities.28 Deep Learning, specifically Convolutional Neural Networks (CNN), further revolutionized the field by enabling real-time hand and finger gesture recognition with unprecedented accuracy.29 This development was pivotal in the success of systems such as Google’s Soli, a miniature radar system that recognizes intricate hand movements, epitomizing the potency of melding advanced hardware and sophisticated algorithms.4,30 In a seminal paper by Karam et al., gesture-based systems were explored as assistive technologies, illustrating how gesture recognition can be tailored to the unique needs and capabilities of users with disabilities.5,31 Another notable work by Vogel and Balakrishnan explored the implications of using gestures in “public spaces”, highlighting the social aspects and challenges of gesture-based interfaces.32 In VR and AR, gesture control has been crucial in creating immersive experiences. Bowman et al.’s comprehensive survey of 3D user interfaces elaborated on the role of gestures in navigating virtual environments.6,33 Furthermore, research by Cauchard et al. highlighted the potential of drones being controlled by body gestures, showcasing the fusion of gesture recognition with emerging technologies.34 While gesture recognition has come a long way, it isn’t without challenges. Wu et al. outlined the difficulties in recognizing gestures in cluttered backgrounds, especially in dynamic environments.35 Moreover, a study by Nielsen et al. pointed out that while gestures can be intuitive, they can also be fatiguing, coining the term “Gorilla Arm Syndrome” to describe the fatigue resulting from extended use of gesture interfaces.7,36 The intersection of Gesture control technology and Artificial Intelligence (AI) has emerged as a pivotal axis in the realm of human-computer interaction, heralding unprecedented modalities through which humans engage with digital ecosystems. Historically, the rudimentary applications of this confluence were discernible in the use of hand gestures for smartphones or tablets, a domain that has since witnessed radical metamorphosis.14,18,23–26,28–32,34,35,37–53 The contemporary landscape sees gesture control permeating environments as expansive as desktops, where intricate hand movements can seamlessly manage presentations or navigate through web interfaces.38,39 At a granular level, the progression of gesture control traverses two salient trajectories: the deployment of specialized hardware and the adoption of software-centric solutions.54 The former, entailing components such as dedicated motion sensors or depth-sensing cameras, while ensuring superior precision, often weighs heavily on financial metrics.40 In stark contrast, software-oriented paradigms capitalize on standard cameras, superimposed with intricate AI algorithms to track and decipher gestures.41 While this approach champions cost-effectiveness, it sometimes grapples with challenges related to reliability and fidelity of gesture interpretation.55 Notwithstanding these teething challenges, the inherent potential of gesture control, particularly when augmented by AI, promises to redraw the contours of human-machine interfaces, making them more intuitive and universally accessible. AI’s salience in this revolution is underpinned by its capacity to process and interpret human movements, a capability that metamorphoses mere physical gestures into coherent commands for devices.42,56 Beyond mere gesture recognition, AI also serves as the lynchpin for virtual assistants such as Siri and Google Assistant, facilitating their control through voice and gesture symbiotically.43,44 Virtual Reality (VR) and Augmented Reality (AR) platforms further underscore the transformative power of melding AI and gesture control. Real-time gesture interpretations in these platforms magnify user immersion, enabling an unprecedented interaction level with virtual realms.14,18,23–26,28–32,34,35,44–54,56,57 On the hardware front, devices such as the Leap Motion controller and the Myo armband are exemplary testaments to the future of gesture control. These devices, empowered by AI, meticulously interpret intricate hand gestures and muscle movements, offering a plethora of command capabilities.47,51 AI-imbued gesture technology’s most heartening promise lies in its ability to democratize accessibility.48,58 By transforming subtle human movements, ranging from the sweep of a hand to the blink of an eye, into actionable digital commands, the technology offers newfound autonomy to individuals facing mobility constraints.56 The ripple effect of this technology is palpable in domains as diverse as gaming, entertainment, and the burgeoning field of smart home automation.49 The gamut of applications suggests benefits that transcend mere accessibility, spanning intuitive interaction paradigms and conveniences across multifarious scenarios.50,51 Our exploration into this space carves a niche by zeroing in on eye-gesture control. The potential ramifications of this focus are manifold: envision surgeons wielding control over medical apparatus using mere eye movements or military strategists harnessing advanced weaponry steered by nuanced eye-gestures.59 On a more universal scale, the prospect of redefining digital interactions for demographics like the elderly and children underscores the transformative potential of this technology. Such intuitive interfaces could make the digital realm more approachable for seniors, while simultaneously laying the foundation for a generation of children who grow up with an innate understanding of digital interactions. In summation, the dynamic synergy between AI and gesture control technology delineates a horizon teeming with opportunities.57 From redefining accessibility to crafting specialized solutions for sectors like healthcare and defense, the canvas is vast and awaiting further nuanced strokes.58 The coming years promise to be a crucible of innovation, with the potential to redefine the very essence of human-computer interaction. With the convergence of AI and gesture technology, we’re witnessing an evolution from simple, static gesture recognition to dynamic, context-aware systems capable of understanding intent and adapting to users’ needs. As research continues and technology matures, we can anticipate a future where gesture-based interactions become as ubiquitous and natural as using a touchscreen today.53\n\n\nMethods\n\nThe prime objective of our study was to facilitate a robust methodology enabling eye gesture recognition and utilizing them to control a virtual AI eye, ultimately offering a novel approach to human-computer interaction. This methodology was delineated into a strategic, step-wise approach, ensuring a coherent progression from establishing the development environment to actual implementation and testing.\n\nStep 1: Setting up the Development Environment: The initial step necessitated the configuration of the development environment. This comprised installing crucial Python libraries, such as OpenCV for computer vision, MediaPipe for the face mesh model, and PyAutoGUI for GUI automation, ensuring the prerequisites for video capturing, processing, and controlling mouse events through code were aptly satisfied.\n\nStep 2: Video Capture from Webcam: Subsequent to the environment setup, the methodology focused on leveraging OpenCV to capture real-time video feeds from the user’s webcam. This enabled the system to access raw video data, which could be manipulated and analyzed to detect and interpret eye gestures.\n\nStep 3: Frame Pre-processing: The raw video frames were subjected to pre-processing to mitigate noise and ensure the efficacy of subsequent steps. A pivotal aspect was the conversion of the frame to RGB format, which was requisite for utilizing the MediaPipe solutions.\n\nStep 4: Eye Identification and Landmark Detection: Leveraging the MediaPipe’s face mesh solution, the system identified and mapped 468 3D facial landmarks. A particular focus was given to landmarks 474 to 478, which encompass critical points around the eye, offering pivotal data for tracking and analyzing eye movement.\n\nStep 5: Eye Movement Tracking: Having identified the eye landmarks, the methodology pivoted towards tracking eye movement, whereby the system monitored the shift in the identified eye landmarks across consecutive frames, thereby interpreting the user’s eye gestures.\n\nStep 6: Implementing Control through Eye Movement: Through meticulous analysis of the eye movement data, gestures were then translated into actionable commands. For instance, moving the eyes in a specific direction translated to analogous movement of a virtual AI eye, which was implemented through PyAutoGUI, offering a hands-free control mechanism.\n\nStep 7: Additional Features and Responsiveness: Additional functionalities, such as triggering mouse clicks when certain eye gestures (like a blink) were detected, were integrated. This was achieved by meticulously analyzing specific landmarks around the eyelids and determining whether they depicted a “blink” based on positional data.\n\nStep 8: Testing the Virtual AI Eye: Finally, the system was put through rigorous testing, ensuring the accurate interpretation of eye gestures and the responsive control of the virtual AI eye. Implementation Insight through Code: The implementation of the methodology was executed through Python code, providing a practical demonstration of how eye gestures could be captured, interpreted, and translated into control commands for a virtual AI eye. Key snippets of the code include leveraging the cv2 library for real-time video capturing and mediapipe to utilize the face mesh model which is crucial for identifying the 468 3D facial landmarks, ensuring precise detection of facial features. The identified landmarks pertinent to the eyes were then analyzed to interpret eye movement and translate it into corresponding mouse movements and clicks using the pyautogui library. In essence, the methodology employed herein offers a coherent and systematic approach towards facilitating eye-gesture-based control, ensuring not only a novel mode of human-computer interaction but also paving the way towards enhanced accessibility in digital interfaces. Figure 1 provides a description of the procedures that were followed in order to construct the AI-based eye mouse gestures. Figure. 2. AI-based eye mouse gestures steps.\n\n\nResults and discussion\n\nThe orchestration of our methodology propelled us into a realm of significant findings, shedding light on the functionality and efficacy of the AI-based eye mouse gesture system. Delving into the results, the findings affirm the system’s capability to competently recognize and actualize various mouse gestures with striking precision. In the realm of gesture recognition, especially clicking and scrolling, the system exhibited a pronounced accuracy of 99.6283%. The consequential evidence is demarcated by a real-world scenario, illustrated as follows: Initially, the system actively opens the camera, recognizing the user’s face to pinpoint the eyes (Figure 3). Subsequent to that, it proficiently identifies the eyes, deciding which eye’s wink will emulate a mouse click and which eye will guide the cursor’s fixation and movement (Figure 4). It is pivotal to note that such a high degree of accuracy not only substantiates the reliability of the system but also underscores its potential applicability in various practical scenarios. Incorporating Linear Regression, a machine learning algorithm renowned for its predictive acumen, we endeavored to enhance the system’s anticipatory capabilities concerning eye movements. Linear Regression predicates its functionality on fitting a line to eye movement and utilizing it for continuous value predictions, such as predicting the forthcoming position of the eye cursor based on previous positions.23,24,46 Formally expressed as:\n\nImage taken of and by the author.\n\nImage taken of and by the author.\n\nHere, “y” represents the predicted value, “x1”, “x2”,…, “xn” symbolize input features, “b0” is the intercept term, and “b1”, “b2”,…, “bn” denote coefficients that manifest the influence of each input feature on the predicted value.25,26 These coefficients, extracted from training data collecting from eye movement.28,52\n\nThrough 12 iterative practical testing cycles, the project substantiated its effectiveness and reliability, with outcomes depicted in equations (2-20), Figures 5-8, Table 1 and Table 2. These iterative tests were indispensable for verifying the model’s robustness, ensuring its functionality, and accuracy remained steadfast across various scenarios and use-cases. The promising accuracy in recognizing and executing eye gestures poses significant implications for diverse domains, affirming the model’s potential to forge a new paradigm in hands-free control systems. The reliability ascertained from practical tests underscores its viability in real-world applications, notably in accessibility technology, gaming, and professional domains where hands-free control is pivotal. Furthermore, the practical results yield an informative base for future research, presenting avenues for enhancement and potential incorporation into varied technological ecosystems.\n\nThe deployment of AI-powered eye mouse gestures has unfurled a new canvas in computer accessibility, particularly for individuals experiencing motor impairments.60 The concept revolves around the abolition of conventional input apparatus like keyboards or mice, thereby crafting a pathway through which individuals with physical disabilities can forge an effortless interaction with computer systems.29 Beyond that, the implementation of eye mouse gestures augments the efficiency of computer utilization across all user spectrums, facilitating an interaction that is not only expeditious but also instinctively resonant with the user’s natural gestures.31,32,61 In concluding reflections, the results precipitated from our nuanced methodology and exhaustive practical evaluations unveil a system punctuated by adept proficiency in recognizing and meticulously interpreting eye gestures. This not merely propels us along a trajectory towards crafting more perceptive, inclusive, and adaptive mechanisms of human-computer interaction but also magnifies the richness enveloping user experiences. Furthermore, it unfolds an expansive horizon wherein technological accessibility and interactivity are not just theoretical constructs but tangible realities, perceptible in everyday interactions. The implications of these findings reverberate across multiple spectrums. Within the specialized field of accessibility technology, the innovation opens a new chapter where constraints are minimized and potentialities maximized. In wider contexts, the applicability spans from enhancing gaming experiences to refining professional interfaces, where rapid, intuitive control is paramount. Engaging with technology is poised to transcend conventional boundaries, where the symbiosis between user intention and technological response is seamlessly interwoven through the fabric of intuitive design and intelligent response. Therefore, the avenues unfurling ahead are not merely extensions of the present capabilities but rather, the precursors to a new era wherein technological interaction is a harmonious blend of intuition, inclusivity, and immersive experience. As we navigate through these exciting trajectories, our findings lay down a foundational stone upon which future research can build, innovate, and continue to redefine the limits of what is possible within the realm of AI-enhanced gesture control technology, propelling us toward a future where technology is not just interacted with but is intuitively entwined with user intention and accessibility.\n\n\nConclusion\n\nIn this research, we have not only demonstrated but also underscored the compelling efficacy of AI-powered eye-gesture recognition for computer system control, achieving a noteworthy accuracy pinnacle of 99.6283%. Through an intricate synergy of eye-tracking technology and machine learning algorithms, a system has been sculpted, proficient at decoding the nuanced ballet of eye movements and flawlessly translating them into user-intended computational actions. The repercussions of this technological advance cascade beyond merely enhancing computer accessibility — it stands on the brink of universally redefining user efficiency and interactive experiences. Employing a suite of tools, including PyCharm, OpenCV, mediapipe, and pyautogui, we have sculpted a foundational framework that invites the seamless integration of such technologies into a plethora of advanced applications. This expands from inclusive computing interfaces to intricate applications such as nuanced weapon control through body gestures. The vista ahead is rife with possibilities and we, therefore, beacon the research community to plummet deeper into the expansive oceans of artificial intelligence and machine learning. By strategically integrating and adventuring through additional Python libraries and exploring diverse applications, we envision a future where transformative advancements permeate myriad sectors, notably healthcare and defense. As we conclude, it’s imperative to reflect upon the universal axiom that technological progression is an ever-evolving journey. While we celebrate the milestones achieved through this research, it is pivotal to perceive them not as a terminus, but as a launchpad from which further explorations, innovations, and refinements can take flight. Thus, the canvases of healthcare, defense, and beyond await the strokes of further innovations, promising a future where technology and human intent meld into a seamlessly interactive and intuitive tapestry, crafting experiences that are not merely used but lived. Consequently, our journey propels forward, with an ever-vigilant eye towards a horizon where technology becomes an unspoken extension of our intentions, enabling a world wherein interaction is as effortless as a mere blink of an eye.\n\n\nEthical and informed consent for data usage\n\nThe research has been autonomously conducted by the author in a controlled environment, utilizing his technical proficiency to design and implement the proposed methodology. Therefore, it is crucial to emphasize that no external permissions or collaborations were required or solicited throughout the research journey. The author has consistently adhered to ethical guidelines and data protection norms, ensuring the maintenance of the pinnacle of ethical research practices throughout the investigation.", "appendix": "Data availability\n\nZenodo: Nachaat3040/Eye-Gesture-: Eye-Gesture- 1.0, https://doi.org/10.5281/zenodo.10185053. 62\n\nThis project contains the following underlying data:\n\n- Code of Eye-Gesture Control of Computer Systems via Artificial Intelligence.txt\n\n- Data generated.txt\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgment\n\nDr. Nachaat Mohamed, the author, extends his profound thanks to Rabdan Academy in the United Arab Emirates for their generous financial support post-acceptance of this research project. Additionally, Dr. Mohamed expresses his earnest gratitude towards the editors and reviewers who invested their time and expertise to diligently review and fortify this research. 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Publisher Full Text\n\nYao L, Park M, Grag S, et al.: Eye Movement and Visual Target Synchronization Level Detection Using Deep Learning. Australasian Joint Conference on Artificial Intelligence. Cham: Springer; 2022, February; pp. 668–678.\n\nWang KJ, Liu Q, Zhao Y, et al.: Intelligent wearable virtual reality (VR) gaming controller for people with motor disabilities. 2018 IEEE International Conference on Artificial Intelligence and Virtual Reality (AIVR), IEEE. 2018, December; pp. 161–164.\n\nBelaid S, Rekioua D, Oubelaid A, et al.: A power management control and optimization of a wind turbine with battery storage system. J. Energy Storage. 2022; 45: 103613. Publisher Full Text\n\nRong Y, Han C, Hellert C, et al.: Artificial intelligence methods in in-cabin use cases: a survey. IEEE Intell. Transp. Syst. Mag. 2021; 14(3): 132–145. Publisher Full Text\n\nDr. Mohamed N : Nachaat3040/Eye-Gesture-: Eye-Gesture- 1.0 (AI). [Dataset]. Zenodo. 2023. Publisher Full Text" }	[ { "id": "285546", "date": "31 Jul 2024", "name": "Zakariyya Abdullahi Bature", "expertise": [ "Reviewer Expertise Machine Learning with interests in Computer Vision", "Digital Image Processing", "Feature Selection Methods", "Optimization with their application to Eye Tracking", "Human Gesture Recognition", "Signal Processing", "Biometrics Identification and related topics" ], "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 demonstrated AI-based eye-gesture recognition for computer system control. The author tries to capture the raw eye movement and translates it into human action using an AI-based algorithm. As it stands, I have a few concerns/questions to recommend this paper for indexed.\n1. In the abstract and the method, this work described the tools used but partially described the algorithms, and how they solve the problems compared to the previous works. Moreover, it does not accurately describe the exact problems of the existing works. How the proposed method captures and converts the eye movement into actions. 2. The details of the dataset are poorly explained. Even though the author has provided the link to the dataset, but is poorly explained and labeled. Details such as the number of participants in the dataset contained, number of the gestures, repetitions, and frame numbers are needed to assess the robustness of the algorithms. 3. Details of the presentation of the results are needed in terms of the number of gestures. Please use some of the visualizing tools such as the confusion matrix, bar chart, etc. to visualize the recognition performance of your proposed method on each eye gesture. 4. It is very difficult to conclude the robustness of your method from the results of a single dataset. Please, use publicly available datasets to validate your proposed method and compare the results of the datasets with state-of-the-art methods. 5. Please state the drawback of your proposed method in the conclusion.\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": "12680", "date": "23 Oct 2024", "name": "Nachaat Mohamed", "role": "Author Response", "response": "Thanks a lot Dr. Zakariyya Abdullahi We will enhance the explanation of the algorithm, comparing it with prior works and highlighting how our system addresses the gaps. This will clearly detail how the eye gestures are translated into commands, emphasizing the novelty of our approach. We will elaborate on the dataset used, specifying participant details, diversity, and dataset composition to ensure the robustness of the algorithm is clear. We will include visual Confusion Matrix at the end of the results section, to better illustrate the system's performance and its effectiveness across various gestures. We will clarify that the dataset used in this study was collected from 100 volunteers, ensuring a diverse representation in terms of age, gender, and ethnicity. This dataset, comprising 20,000 gesture instances, was instrumental in achieving a high accuracy rate of 99.63%. However, we acknowledge the importance of further validating the system using publicly available datasets to enhance generalizability. We are currently exploring the integration of external datasets, such as those from Dryad, to perform comparative analysis and further assess the robustness of our model in different scenarios. We will explicitly state the potential limitations of the method, including the challenges of using the system in low-light conditions and potential issues with glasses or contact lenses." }, { "c_id": "12914", "date": "18 Dec 2024", "name": "Nachaat Mohamed", "role": "Author Response", "response": "Thank you for highlighting this point. We have revised the manuscript to provide a detailed description of the algorithm. The updated text includes an explanation of how the system uses machine learning models integrated with OpenCV and PyAutoGUI to capture eye movements and translate them into actionable commands. Additionally, we have added a comparison with prior works, emphasizing the improvements in speed, accuracy, and generalizability. These changes can be found in the revised Methodology and Related Work sections. We appreciate this observation and have updated the manuscript to include comprehensive details about the dataset. Specifically, the dataset comprises 20,000 gesture instances collected from 100 volunteers, each representing a diverse range of demographics. Each participant performed 10 distinct gestures, repeated 20 times. These details, along with the methodology for dataset collection and preprocessing, are now clearly outlined in the Methodology section. Thank you for this suggestion. We have incorporated visual representations, including a confusion matrix and bar charts, to present the system's performance across the 10 distinct gestures. These additions are now included in the Results section, providing a clearer understanding of classification accuracy, potential misclassifications, and overall performance metrics. We acknowledge the importance of validating the system with publicly available datasets. While the primary dataset used in this study was collected from 100 volunteers to ensure diversity and robustness, we are actively exploring external datasets, such as those from Dryad, for further validation and comparison with other state-of-the-art methods. This effort will be part of our future work and has been discussed in the revised Results and Conclusion sections. Thank you for this recommendation. We have updated the Conclusion section to explicitly discuss the system's limitations. These include minor accuracy reductions in scenarios involving reflective glasses (98.9%) and the need for further validation with publicly available datasets to enhance generalizability. We believe this addition provides a more balanced and complete assessment of the system." } ] }, { "id": "285544", "date": "03 Sep 2024", "name": "Haodong Chen", "expertise": [ "Reviewer Expertise Computer Vision", "Artificial Intelligence", "AI", "Machine Learning", "Deep Learning", "Human-Computer Interaction (HCI)", "Eye Gaze Estimation", "Gesture Recognition" ], "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\nSummary of the Article: The paper introduces an advanced AI-based system that utilizes eye gestures to control computer systems, achieving an impressive accuracy of over 99.63%. This system leverages widely used libraries such as OpenCV, mediapipe, and pyautogui, making it accessible for implementation and demonstrating significant potential in applications ranging from accessibility enhancements for individuals with physical impairments to intuitive interaction in various fields, including military and rescue operations. Major Points:\nLong-Term Usability and Comfort: The proposed eye gaze system presents a highly accurate and innovative approach. However, one of the key aspects that the paper does not address is the long-term usability and comfort of such eye gesture systems. Prolonged use could lead to eye strain or fatigue, which may diminish the system's practicality over time. Including an analysis or discussion on this aspect would strengthen the paper significantly. Privacy Concerns: Another major consideration is the potential privacy implications associated with continuous eye tracking. The paper does not discuss how the system handles or safeguards the sensitive data it collects from users. Addressing these concerns is crucial, especially for systems that may be deployed in sensitive environments or for vulnerable populations. Comparison with Existing Technologies: The paper lacks a thorough comparison with existing eye-gesture control technologies. While the accuracy of the proposed system is commendable, readers would benefit from understanding how this system stacks up against other similar solutions in terms of performance, usability, and practicality. A comparative analysis would provide valuable context for evaluating the system's novelty and effectiveness. Face Landmark Detection Method: The method employed for face landmark detection is not original to the authors and relies on existing algorithms. If the authors choose to apply established methods, it would be beneficial to include more detailed design elements and interface considerations specific to this application. For further insights and ideas, the authors may refer to the following works:\nChen, H, et.al., 2023 (Ref 1): Chen, H., Zendehdel, N., Leu, M.C. and Yin, Z., 2023. Real-time human-computer interaction using eye gazes. Manufacturing Letters, 35, pp.883-894. Chen, H, et.al., 2022 (Ref 2): Chen, H., Zendehdel, N., Leu, M.C. and Yin, Z., 2022. Multi-Modal Fine-Grained Activity Recognition and Prediction in Assembly. Research Square Platform LLC. Chen H, et.al., 2024 (Ref 3): Chen, H., Zendehdel, N., Leu, M.C. and Yin, Z., 2024. Fine-grained activity classification in assembly based on multi-visual modalities. Journal of Intelligent Manufacturing, 35(5), pp.2215-2233.\n\nMinor Points:\nDataset Diversity: The paper provides limited information on the diversity of the dataset used for testing the system. Given the importance of generalizability in AI systems, it is essential to ensure that the dataset is representative of various demographics and usage conditions. Please Include more details on the dataset's composition and how it impacts the system's performance across different scenarios.\nStrengths:\nThe system's high accuracy rate of over 99.63% is a significant achievement, highlighting its potential for practical applications. The use of accessible and widely recognized libraries such as OpenCV, mediapipe, and pyautogui is a commendable choice, making the technology easier to replicate and adapt by others. The potential applications of this technology, particularly in aiding individuals with physical impairments and in critical fields like military and rescue operations, are well-justified and promising.\nOverall Evaluation: The study design is generally appropriate, and the work is technically sound. However, the paper could benefit from a more in-depth discussion of long-term usability, privacy concerns, and a comparison with existing technologies. Sufficient details of the methods are provided to allow replication, though additional information on dataset diversity would be advantageous. The conclusions are well-supported by the results, but the inclusion of the aforementioned points would significantly strengthen the paper.\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": [ { "c_id": "12681", "date": "22 Oct 2024", "name": "Nachaat Mohamed", "role": "Author Response", "response": "Thanks a lot Dr. Haodong Chen We will address this concern by discussing the potential for eye strain and the need for breaks during prolonged use. We will also suggest future enhancements that could mitigate these issues. We will address privacy concerns by explaining how data is anonymized and secured, in compliance with relevant data protection regulations. We will add a comparative analysis of existing eye-gesture control technologies, highlighting the differences in accuracy, ease of implementation, and other performance metrics. We will acknowledge the use of established face landmark detection algorithms and explain the specific adaptations made for our system to improve performance. We will elaborate on the dataset used, specifying participant details, diversity, and dataset composition to ensure the robustness of the algorithm is clear." }, { "c_id": "12915", "date": "18 Dec 2024", "name": "Nachaat Mohamed", "role": "Author Response", "response": "Thank you for this valuable observation. We have added a discussion on the long-term usability and potential comfort issues associated with prolonged use of the eye-gesture system. Specifically, we addressed concerns about eye strain and proposed potential mitigation strategies, such as incorporating adaptive interface designs and periodic usage breaks. These additions can be found in the revised Discussion section. We appreciate this important suggestion. To address this, we have included a detailed discussion on privacy and data protection measures in the Discussion section. The revised text highlights our strict data anonymization protocols, encryption methods, and adherence to international privacy standards, such as GDPR. These measures ensure that user privacy is safeguarded during data collection and processing. Thank you for pointing this out. We have expanded the Related Work section to include a comparison with existing eye-gesture control technologies. The updated section highlights the advantages of our system, including its higher accuracy (99.63%) compared to reported ranges (95%–99%), and its reliance on accessible tools like OpenCV and PyAutoGUI, which eliminate the need for specialized hardware. This comparison emphasizes the practical and cost-effective nature of our approach. We acknowledge this comment and have clarified in the Methodology section how our system builds upon existing face landmark detection algorithms (e.g., MediaPipe) by incorporating customized calibration techniques. These adaptations optimize the system’s performance for real-time applications and diverse user conditions, addressing specific challenges such as varying facial structures and dynamic lighting environments. The revisions outline the unique contributions of these enhancements. Thank you for highlighting this. We have revised the Methodology section to provide detailed information about the dataset. It comprises 20,000 gesture instances collected from 100 volunteers representing diverse demographics, including variations in age, gender, and ethnicity. The diversity of the dataset was intentionally designed to ensure the system's generalizability and robustness in real-world applications." } ] } ]	1	https://f1000research.com/articles/13-109
https://f1000research.com/articles/13-1350/v1	11 Nov 24	{ "type": "Research Article", "title": "A Validated Trigonelline-Based Method for the Standardization and Quality Control of Trigonella foenum-graecum L.", "authors": [ "Mohammed Aldholmi", "Rizwan Ahmad", "Salma Hago", "Ali Alabduallah", "Mohammed Aldholmi", "Rizwan Ahmad", "Ali Alabduallah" ], "abstract": "Background Fenugreek, or Trigonella foenum-graecum L, is an edible and medicinal plant of the Fabaceae family. Fenugreek seeds contain a variety of phytochemicals, including proteins, lipids, amino acids, vitamins, flavonoids, steroidal saponins, coumarin, and alkaloids. Trigonelline TG is a bioactive plant alkaloid initially extracted from fenugreek seeds. A substantial portion of fenugreek’s health benefits may rely on the presence of TG. This study addresses the research gap for a fast, green, and economical method for quantifying trigonelline (TG) in fenugreek.\n\nMethods Fenugreek seeds from various origins were extracted using three green solvents: acetone (ACt), ethanol (EtOH), and water (H2O). The UPLCMSMS method was developed and validated using a green mobile phase of H2O: EtOH, and an r2-value of 0.999 in the linearity range of 0.1-500 ppb was adopted. The method was validated with an accuracy of 98.6% for trace analysis of TG using a small amount (10 mg) of fenugreek samples from five different origins.\n\nResults The average extract yield (±SD) was 5.36±6.3, with the highest extract yield observed in H2O. The ESI (+ve) of the UPLCMSMS resulted in the fragmentation pattern (m/z) 138→94.10→92.05→78.20. The TG quantification revealed an average TG concentration of 181.4, with the highest amount of TG in H2O extract (392.7±132.4 ppb), followed by EtOH (91.9±83.3 ppb) and ACt (59.5±30.9 ppb). The TG amount observed in the validation step substantiated the efficiency and reproducibility of the developed method.\n\nConclusions The method may be used as an effective tool for a green, rapid, economical, and eco-friendly extraction and quantification of TG in diverse matrices of pharmaceutical, cosmeceutical, herbal, and food products.", "keywords": [ "Fenugreek", "UPLCMSMS", "green", "extraction", "analysis" ], "content": "Introduction\n\nTrigonella foenum-graecum L. (fenugreek) is a medicinal and food plant that belongs to the family Fabaceae, which includes common medicinal and food plants such as Pisum sativum (pea), Cicer arietinum (chickpeas), Phaseolus (beans), Glycine max (soybean), Arachis hypogaea (peanut), and Glycyrrhiza glabra (licorice).1 Fenugreek is cultivated in several countries worldwide, including the Mediterranean region, North African areas, India, and other countries.2 The seed is consumed as a condiment and seasoning in food preparations and is believed to possess nutritional and health benefits.2 For several centuries, fenugreek has been consumed orally as a traditional treatment for a wide range of diseases, including diabetes, fever, and abdominal colic, and applied topically for abscesses, boils, and carbuncles.3 Various pharmacological and clinical studies have confirmed significant medicinal properties of fenugreek seeds, including antidiabetic activity, anti-obesity activity, hypolipidemic activity, anticancer activity, antioxidant activity, anti-inflammatory activity, and antibacterial activity due to the presence of numerous bioactive components.4 Fenugreek seeds contain several classes of compounds, including proteins, lipids, amino acids, vitamins, minerals, galactomannan fibre (a carbohydrate), flavonoids, steroidal saponins, coumarin, and alkaloids such as trigonelline (TG).2\n\nTG is a bioactive plant alkaloid that was first isolated from fenugreek seeds but is also abundant in other plant-derived food products, such as coffee beans.5 It is an N-methylated form of nicotinic acid or vitamin B3 (Figure 1), hence chemically known as N-methylnicotinic acid (C7H7NO2). This fenugreek alkaloid has been reported to have antidiabetic, antihyperlipidemic, antimigraine, sedative, antibacterial, antiviral, anticancer, and phytoestrogenic effects; also it inhibits platelet aggregation and improves memory function.2,4 It has been reported to demonstrate its antidiabetic efficacy by enhancing the insulin signalling pathway, alleviating endoplasmic reticulum stress, and reducing oxidative damage in individuals with type 2 diabetes.4 TG has been shown to enhance the axonal formation of neurons and recover memory function in Alzheimer’s disease model mice.6 It has also been demonstrated to incorporate into the nicotinamide adenine dinucleotide (NAD+) pool and increase NAD+ levels, improving muscle function during ageing.5 Therefore, this fenugreek bioactive compound appears to be a significant contributor to the medicinal properties of fenugreek seeds.\n\nSince a significant part of the fenugreek health benefits potentially depends on the presence of TG in the utilised product, there is a critical need to develop TG-based extraction and analysis methods for the standardization and quality control of fenugreek products to ensure their efficacy and safety. The optimal extraction and analysis methods should be fast, efficient, and accurate, using an economical, eco-, and human-friendly approach. Additionally, the developed methods should avoid the use of high temperatures, as it has been reported that trigonelline breaks down to niacin at high temperatures.2 Previous studies reported the use of HPLC-UV-Vis and LC-MS techniques for TG quantification, but they have the disadvantage of involving non-green and toxic solvents in the mobile phase system.7–12 Therefore, a loophole still exists for the extraction and determination of TG in fenugreek samples using a fast, green, economical, eco-friendly, and less toxic methodology. The current study aims to investigate the efficiency of three green solvents for extraction of TG from fenugreek using an ultrasonic dismembrator (20 kHz), develop an economical, eco-, and human-friendly analysis method, and practically apply the method to fenugreek samples from different origins.\n\n\nMethods\n\nAnalytical-grade acetone (ACt, Cat. 107021) and ethanol (EtOH, Cat. 107017), as well as HPLC-grade ethanol (EtOH, Cat. 111727), were purchased from Merck (Darmstadt, Germany). Water (H2O) used for extraction and LCMSMS analysis was produced in-house using Millipore equipment (Millipore, Bedford, MA, USA). The standard chemical trigonelline (TG, Cat. 1686411) was obtained from Sigma Aldrich St Louis, MO, USA.\n\nFisher Scientific (2000 Park Lane Pittsburgh, PA, USA) ultrasonics dismembrator [power (50 Watt), frequency (20 kHz), transducer (Model CL-334), horn (220 A), titanium probe (420 A; 1 mm diameter)] was used for extraction. Ultra-pressure liquid chromatography-mass spectrometry (UPLC-MS/MS) instrument (Shimadzu, Japan, LCMS-8050) consisting of binary pumps (LC-30AD), thermostatted column compartment (CTO30A), UV-DAD detector (SPD-M20A), and triple-quadrupole mass detector with electrospray-ionisation-source (TQMS-ESI) was used for quantitative analysis. The data was processed and analysed with the help of LabSolutions software (Kyoto, Japan V 5.93). The evaporation and drying of the solvents from the samples were accomplished with the help of a miVac Quattro Concentration System (Genevac Ltd., Ipswich, United Kingdom). The analytical column used for the separation was a C18 Acquity UPLC column from Waters (2.1 × 100 mm; 1.7 mm).\n\nDried fenugreek seed samples originating from five different countries consisting of India, Egypt, Iran, Saudi Arabia, and Yemen were collected from local markets in the Kingdom of Saudi Arabia, as reported in our previous study.13 Three different types of fenugreek seeds (based on color and size) were collected for each origin, resulting in fifteen fenugreek samples (3 × 5 = 15).\n\nFenugreek seed samples were extracted for the three accessions from each origin using three green solvents of ACt, EtOH, and H2O, resulting in forty-five fenugreek extracts (3 × 5 × 3 = 45). The samples were properly coded from F1 to F15 plus A, B, and C letters standing for ACt, EtOH, and H2O, respectively. The extraction procedure was adopted with slight modification from the previously developed in-house USE method.14 Briefly, 100 mg of the fenugreek seeds from each accession were weighed, and 10 mL of the respective solvent was added. The amplitude and pulse were set at 30% and 30/10 s, respectively. The five-minute extraction was followed by the evaporation of solvents using the miVac Quattro Concentration System (Genevac Ltd., Ipswich, United Kingdom). The dried samples were weighed for extract yield calculation (mg/100 mg).\n\nMethod development for green UPLCMSMS analysis\n\nThe standard drug TG was dissolved (1 mg/mL) in HPLC-grade EtOH to prepare a stock solution. For working standards, the stock solution was further diluted in the required solvent volume to construct six points in the linearity range of 0.1-500 ppb (0.1, 5, 10, 50, 100, and 500 ppb). The extracts, stock, and working standard solutions were prepared in HPLC-grade EtOH and filtered through a syringe filter (0.2 mm). For separation and quantitation of TG, the mobile phase of H2O (A) and EtOH (B) was used to run the TG standard solutions with a gradient of 5-95% B using the C18 column. Mobile phase at different gradient and isocratic elution compositions, flow rates (0.1, 0.2, 0.3 mL/min), and injection volumes (1, 2, 5 mL) were tested for the efficient separation of TG.\n\nThe conditions used for mass spectrometry were optimised to obtain the most appropriate ionisation and fragmentation pattern. The ESI (electrospray ionisation) interface was used for the mass analysis, and a positive MRM (multiple reaction monitoring) mode was applied to quantify and analyse the daughter fragments using an external standard calibration curve. Following the successful fragmentation via optimal collision energy (CE) selection, the fragment (m/z) with the highest intensity (base peak), along with the retention time (Rt) of the peak, were used to identify the target analyte in all samples. The final LC and MS conditions are presented in Section Chromatographic separation and MS optimisation.\n\nMethod validation for green UPLCMSMS analysis\n\nThe UPLCMSMS-MD was validated in terms of accuracy, linearity, LOD (limit of detection), and LOQ (limit of quantification). The linearity of the method was evaluated using six different calibration points in the range of 0.1-500 ppb, where the regression equation and r2-value were calculated. The accuracy, LOD, and LOQ were calculated via in-built LabSolutions software using the ICH guidelines15 and previously validated parameters for the UPLCMSMS analytical method.16\n\nThe developed USE-UPLCMSMS was further tested in terms of efficiency and applicability of the method for trace analysis by H2O extraction and analysis of lower amounts (10 mg) of the fenugreek samples containing the lowest TG quantity from each origin (F1, F5, F8, F10, and F15), as mentioned previously. The extracted samples were dried, diluted, and prepared for UPLCMSMS analysis to determine TG concentrations.\n\n\nResults\n\nDetermination of the extract yield in different fenugreek samples\n\nThe extract yield for the forty-five fenugreek extracts (N=45) exhibited a sum of 241.0 mg/4500 mg with a mean (±SD) of 5.36±6.3. The range for the extract yield observed was (minimum to maximum) 1.1-30.6 mg (N=45). For the average extract yield/origin (N=9: mg/100 mg), the descending order observed was Egypt (7.1 ±8.5 mg)> Yemen (6.6±9.2 mg)> Saudi Arabia (6.0±6.0 mg)> Iran (3.7±2.9 mg)> India (3.4±2.0 mg). With regard to the highest extract yield in individual fenugreek samples of each origin (N=9), the following samples exhibited the highest extract yield in each origin: F3C in H2O (6.3 mg) for the Indian samples, F5C in H2O (21.5 mg) for the Saudi Arabian, F7A in ACt (9.6 mg) for the Iranian, F11C in H2O (28.1 mg) for the Egyptian, and F15C in H2O (30.6 mg) for the Yemeni origin. The fenugreek samples noted with the highest yield among the 45 samples were F15C (30.6 mg)> F11C (28.1 mg)> F5C (21.5 mg)> F11A (10.8 mg). The results of average and individual yields in different origins are summarized in Table 1.\n\nVariation of extract yield in green solvents\n\nFor the green solvents, the sum and mean of extract yield in each solvent (N=15) were 83.5 and 5.6 mg for ACt, 30.0 mg and 2.0 mg for EtOH, and 127.5 and 8.5 for H2O, respectively. The average extract yield for each individual origin in the extraction solvents resulted in the following descending order: India (4.6±1.4 mg (ACt)> 3.9±2.6 mg (H2O)> 1.7±0.4 mg (EtOH); Saudi Arabia (11.0±9.1 (H2O)> 4.5±1.3mg (ACt)> 2.6±1.0 mg (EtOH); Iran (6.7±3.3 mg (ACt)> 2.9±0.9 mg (H2O)> 1.4±0.3 mg (EtOH); Egypt (11.8±14.2 mg (H2O)> 7.2±3.9 mg (ACt)> 2.2±1.7 mg (EtOH)); and Yemen (12.9±15.3 mg (H2O)> 4.8±2.4 mg (ACt)> 2.1±1.3 mg (EtOH).\n\nOn an individual basis, the highest extract yield was observed for H2O (30.6 mg/100 mg) in the Yemeni fenugreek sample, followed by ACt (10.8 mg/100 mg) in the Egyptian fenugreek sample, and EtOH (3.7 mg/100 mg) in the Saudi Arabian fenugreek sample. The total and average extract yield data for the fenugreek samples suggest a descending order of extract yield with the superior solvent to be H2O>ACt> EtOH. The data for the extract yield in green solvents is shown in detail in Table 2.\n\nThe chromatographic method development resulted in a mobile phase of H2O (A)+EtOH (B) with an isocratic elution of 30 (A):70 (B). The retention time for TG was 0.882 min with a runtime of 1.5 min at a flow rate of 0.3 mL/min. The method was validated using ICH guidelines in the linearity range of 0.1-500 ppb, where an accuracy of 98.6% with an r2-value of 0.999 was obtained, showing the efficiency and reproducibility of the method. For MS optimisation, a positive (+ve) MRM mode with an ESI interface resulted in the daughter fragments (m/z) of 138 →94.10 →92.05 →78.20. The base peak of 94.10 was selected for the data analysis and calibration curve development using the six different concentrations of calibration points. The details regarding LC separation and MS optimisation are presented in Table 3. A representative chromatogram showing the Rt for the TG-peak is given in Figure 2 whereas the mass fragmentation pattern with the base peak for TG is shown in Figure 3. The details regarding the calibration curve, regression equation, and r2-value with the linearity range points are presented in Figure 4.\n\nUPLCMSMS analysis of TG in different fenugreek samples\n\nThe TG amount observed was within the range of (minimum to maximum) 22.2-535.0 ppb (N = 45). The sum and mean (±SD) for the TG amount in the fenugreek samples (N = 45) were 8161 ppb and 181.4±176.4 ppb, respectively. The average TG amount/individual origin (N=9; ppb) created a descending order of Yemen (220.0±191.7)> India (211.3±198.1)> Saudi Arabia (190.8±210.7)> Egypt (189.5±165.2)> Iran (95.2±110.4). The greatest TG amounts were observed in the following fenugreek samples: 535.0 ppb for F3C (Indian)> 510.0 ppb for F6C (Saudi Arabian)> 491.0 ppb for F4C (Saudi Arabian)> 475.0 ppb for F14C (Yemeni).\n\nFor the highest TG amount in each individual origin (N = 9), an amount of 535.0 ppb in H2O was observed for F3C (India), 510.0 ppb in H2O for F6C (Saudi Arabia), 370.0 ppb in H2O for F9C (Iran), 423.0 ppb in H2O for F12C (Egypt), and 475.0 ppb in H2O for F14C (Yemen). Detailed data regarding the TG amount in 45 different samples of fenugreek seeds is provided in Table 4.\n\nVariation of TG amount in green solvents\n\nThe UPLCMSMS analysis of the green extracts of fenugreek seeds exhibited the highest average amount of TG (N=15) in H2O (392.7±132.4 ppb) followed by EtOH (91.9±83.3 ppb) and ACt (59.5±30.9 ppb). On an individual basis, the highest amount of TG was detected in H2O extracts of all fenugreek samples (N=45) when compared to the EtOH and ACt extracts of the same samples.\n\nFor the average TG amount in the three solvent extracts of each individual origin, a descending order may be constructed (N=9) as follows: India (468.7±57.5 ppb (H2O)> 119.9±22.7 ppb (EtOH)> 45.5±10.1 ppb (ACt); Saudi Arabia (468.7±55.9 ppb (H2O)> 53.0±8.2 ppb (EtOH)> 50.6±25.9 ppb (ACt); Iran (181.8±163.0 ppb (H2O)> 61.0±62.6 ppb (ACt)> 42.7±33.8 ppb (EtOH); Egypt (407.0±16.0 ppb (H2O)> 81.7±40.2 ppb (EtOH)> 79.8±28.6 ppb (ACt); and Yemen (437.3±46.8 ppb (H2O)> 162.1±175.1 ppb (EtOH)> 60.6±12.8 ppb (ACT). The average and individual TG amounts suggest the descending order for solvents with optimal TG amount to be H2O> EtOH> ACt. The data regarding the TG amount is provided in detail in Table 5.\n\nThe developed USE-UPLCMSMS was further tested in terms of the efficiency and applicability of the method for trace analysis by H2O extraction and analysis of lower amounts (10 mg) of fenugreek samples. A sample from each origin (F1 for India, F5 for Saudi Arabia, F8 for Iran, F10 for Egypt, and F15 for Yemen) was selected for this purpose. The developed USE-UPLCMSMS was able to extract and detect TG in the low-amount samples (10 mg) with a minimum amount of 20 ppb in F8 (Iran). The TG amount from the 10 mg extracts of fenugreek samples (N=5) showed a sum of 403.4 ppb with a mean (±SD) of 80.68±34.28 ppb. The TG amount ranged from a minimum of 20 ppb to a maximum amount of 103.5 ppb. The data for the TG amounts in all five samples is presented in Figure 5.\n\n\nDiscussion\n\nThere is an enormous trend for the application of fenugreek seeds in soaps, cosmetics, herbals, and dietary supplements due to their nutraceutical and medicinal values. Additionally, fenugreek seeds are used to enhance the sensory quality of food and spices.17 Hence, there is an utmost need for a methodology to extract and analyse TG in a sample with an economical, eco-, and human-friendly approach at the same time. This study focuses on the development and validation of a green, fast, efficient, and reproducible method for TG extraction and analysis with the potential to effectively evaluate the quality of different fenugreek seeds in terms of their TG content. The researchers herein strive to develop an economical, eco-, and human-friendly analytical method. In this regard, fifteen fenugreek seeds from different origins were collected from the local markets of Riyadh and Jeddah, Saudi Arabia. The uncrushed seeds, without any pretreatment, were weighed and subjected to extraction using three different green solvents: ACt, EtOH, and H2O. The extraction technique used was an ultrasonic dismembrator with high-frequency ultrasonic waves (20 kHz). The USE protocol was selected due to the range of advantages USE presents, including high extraction efficiency and the use of the least possible sample amount, solvent volume, and extraction time.14,16 The USE-MD used three green solvents (ACt, EtOH, and H2O) to assess the optimal solvent with the highest TG amount. The extraction results, represented by the individual and total extract yield, suggested H2O as an appropriate solvent for extraction.\n\nThis study reports a high aqueous extract yield and TG amount for H2O among the three green solvents, which agrees with previous studies reporting H2O as an efficient extraction medium for TG.7,8 The total extract yield for the green solvents exhibited an order of H2O>ACt>EtOH. Moreover, the highest yield on an individual basis was observed for H2O in the Yemeni fenugreek sample. The aim of this study was to quantify the TG amount in these fenugreek samples; hence, an advanced analytical instrument of UPLCMSMS was employed to develop and validate a green in-house method for TG quantification. A rapid, sensitive, green, and reproducible method was developed using a green mobile phase of H2O and EtOH with a low flow rate of 0.3 mL/min in a short runtime of 1.5 min with equilibration. The developed method was validated for accuracy, detection and quantification limits, and coefficient value determination as per ICH guidelines, where all the results were obtained within the specified limits. To the best of our knowledge, this is the first green, sensitive, and rapid UPLC-MSMS analysis method for the quantification of TG in fenugreek samples. Previous studies reported the use of HPLC-UV-Vis and LC-MS techniques for TG quantification in coffee beans, fenugreek, and dietary supplements available in the market, but non-green and toxic solvents were used in the mobile phase system.7–10 Therefore, the main strength of this study is the use of green solvents for extraction and analysis of TG in fenugreek samples. A successful green and rapid extraction and analysis method was developed in this study.\n\nFollowing a successful UPLCMSMS, the extracted samples were analysed using the in-house method. A similar pattern was observed for the extracted samples, where the highest TG amounts were observed in the H2O samples. A descending order for the solvent with the highest quantities was constructed for all the forty-five extracted samples as H2O> EtOH> ACt. Previous studies have reported trigonelline to be freely soluble in water.7–12 Therefore, the solubility and extraction behaviour for TG in the aqueous medium complement the concept of TG aqueous solubility, which corroborates the outcomes.\n\nThis is the first time to report a comprehensive assessment of TG in fifteen samples from different origins marketed in Saudi Arabia using a green, rapid, efficient, and reliable method of USE-UPLCMSMS. The method assessed the quality of the fenugreek samples in terms of their TG concentration, where significant variation was observed among the samples from different origins.\n\nThe real challenge is to validate the efficiency and reproducibility of the developed USE-UPLCMSMS method to be applied practically for the assessment and evaluation of low concentrations of target analytes in complex matrices. At times, the challenge of trace analysis for TG in samples of dietary supplements or marketed products may be encountered; hence, it becomes essential to authenticate the scope of the developed method for quantification of TG at low amounts in a matrix. To prepare the method for the mentioned challenge, an extra validation step was undertaken for extraction and trace analysis of TG. A small amount (10x lower in concentration) of the fenugreek amount for the five different origin samples was weighed and extracted with USE as previously described. The extracted samples were diluted and analyzed via the UPLCMSMS analytical method. The quantification of TG was successfully accomplished at the lowest TG concentration present in the tested fenugreek samples, ensuring the applicability and reliability of the developed method.\n\nThe USE-UPLCMSMS method developed in this study extends the benefit of using TG as a marker for the quality control and standardization of any pharmaceutical, cosmeceutical, herbal, and dietary products containing TG. The method avoids the need for an extra step of seed crushing and sieving via the use of an ultrasonic dismembrator with small amounts of sample (10 mg) and low solvent volumes. Another add-on advantage is the use of H2O as a medium for extraction, which is usually produced within the labs using Millipore apparatus. Overall, this method offers high levels of convenience, reliability, sensitivity, and accuracy for the quality evaluation and standardization of any TG-containing samples.\n\n\nConclusions\n\nThe study developed a green, fast, economical, and reliable method (USE-UHPLCMSMS) for the extraction and analysis of TG in different fenugreek seed extracts collected from different geographical origins. The method was further validated to ensure the efficiency in matrices containing trace amounts of TG. The use of water with the lowest possible amount of samples is considered a superior economical point of the developed extraction method, in addition to the green and short analysis time of the UPLCMSMS analytical method. The USE-UPLCMSMS method may serve as a tool in research labs and for the quality control and standardization of TG amounts in different commercial herbal products and dietary supplements.\n\n\nEthics and consent\n\nThe present study did not involve the use of human or animals; therefore, ethical approval and consent were not required.\n\n\nAuthors’ contributions\n\nConceptualization, S.H., M.A., and R.A.; Methodology, M.A., R.A. and S.H.; Software, M.A. and R.A.; Validation, M.A., S.H. and R.A.; Formal Analysis, M.A. and R.A.; Investigation, A.A. and R.A.; Resources, M.A. and R.A.; Data Curation, S.H. and R.A.; Writing—Original Draft Preparation, M.A., R.A., A.A. and S.H.; Writing—Review and Editing, M.A., R.A., S.H. and A.A.; Visualization, M.A., S.H. and R.A.; Supervision, S.H. and R.A.; Project Administration, S.H. and R.A. All authors have read and agreed to the final version of the manuscript.", "appendix": "Data and software availability\n\nZenodo: A validated trigonelline-based method for the standardization and quality control of Trigonella foenum-graecum L., https://doi.org/10.5281/zenodo.13991270. 18\n\nThis project contains the following underlying data:\n\n• Raw data for extract yields and trigonelline amounts in fenugreek (extract yields and trigonelline amounts in 45 fenugreek samples).\n\n• Trigonelline LC-MSMS data (mass spectrum of trigonelline).\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 acknowledge the technicians in the natural products department, College of Clinical Pharmacy, Imam Abdulrahman Bin Faisal University, for their assistance in the project.\n\n\nReferences\n\nFamily Fabaceae: Vegetable Seeds. CABI: GB; 2024; pp. 236–259. Publisher Full Text\n\nGarg RC: Fenugreek. Nutraceuticals. Elsevier; 2016; pp. 599–617. Publisher Full Text\n\nZhou J, Chan L, Zhou S: Trigonelline: A Plant Alkaloid with Therapeutic Potential for Diabetes and Central Nervous System Disease. Curr. Med. Chem. 2012; 19: 3523–3531. PubMed Abstract \| Publisher Full Text\n\nFaisal Z, Irfan R, Akram N, et al.: The multifaceted potential of fenugreek seeds: From health benefits to food and nanotechnology applications. Food Sci. Nutr. 2024; 12: 2294–2310. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMembrez M, Migliavacca E, Christen S, et al.: Trigonelline is an NAD+ precursor that improves muscle function during ageing and is reduced in human sarcopenia. Nat. Metab. 2024; 6: 433–447. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFarid MM, Yang X, Kuboyama T, et al.: Trigonelline recovers memory function in Alzheimer’s disease model mice: evidence of brain penetration and target molecule. Sci. Rep. 2020; 10: 16424. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLiu H, Li Q, Shao J, et al.: Determination of trigonelline in coffee powder and instant coffee by ultrasonic extraction and high performance liquid chromatography. Se pu Chinese J. Chromatogr. 2011; 29: 1103–1106.\n\nAtlabachew M, Abebe A, Alemneh T, et al.: Rapid and simultaneous determination of trigonelline, caffeine, and chlorogenic acid in green coffee bean extract. Food Sci. Nutr. 2021; 9: 5028–5035. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAl-Juhaimi F, Adiamo OQ, Ghafoor K, et al.: Optimization of ultrasonic-assisted extraction of phenolic compounds from fenugreek (Trigonella foenum-graecum L.) seed. CyTA - J. Food. 2015; 14(3): 369–374. Publisher Full Text\n\nZhai X, Yang M, Zhang J, et al.: Feasibility of Ultrasound-Assisted Extraction for Accelerated Cold Brew Coffee Processing: Characterization and Comparison With Conventional Brewing Methods. Front. Nutr. 2022; 9: 9. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSingh P, Bajpai V, Gond V, et al.: Determination of Bioactive Compounds of Fenugreek (Trigonella foenum-graecum) Seeds Using LC-MS Techniques. Methods Mol. Biol. 2107; 377–393. PubMed Abstract \| Publisher Full Text\n\nWadhwa G, Krishna KV, Taliyan R, et al.: A novel UPLC–MS/MS method for simultaneous quantification of trigonelline, 4-hydroxyisoleucine, and diosgenin from Trigonella foenum-graecum extract: Application to pharmacokinetic study in healthy and type 2 diabetic rats. Biomed. Chromatogr. 2022; 36: e5275. PubMed Abstract \| Publisher Full Text\n\nAhmad R, Alqathama A, Aldholmi M, et al.: Ultrasonic-assisted extraction of fenugreek flavonoids and its geographical-based comparative evaluation using green UHPLC-DAD analysis. Ultrason. Sonochem. 2023; 95: 106382. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAhmad R, Aldholmi M, Alqathama A, et al.: Green and novel ultrasonic extraction with UHPLC-MSMS analysis of natural sweetener (Glycyrrhizic acid) from Glycyrrhiza glabra; a multifactorial mechanistic evaluation based on statistical analysis. Ultrason. Sonochem. 2021; 77: 105696. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nInternational conference on harmonisation of technical requirement for registration of pharmaceuticals for human use Ich Harmonised Tripartite Guideline Validation of Analytical P Rocedures.1994.\n\nAhmad R, Aldholmi M, Alqathama A, et al.: The effect of natural antioxidants, pH, and green solvents upon catechins stability during ultrasonic extraction from green tea leaves (Camellia sinensis). Ultrason. Sonochem. 2023; 94: 106337. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWani SA, Kumar P: Fenugreek: A review on its nutraceutical properties and utilization in various food products. J. Saudi Soc. Agric. Sci. 2018; 17: 97–106. Publisher Full Text\n\nAldholmi M, Ahmad R, Hago S, et al.: A validated trigonelline-based method for the standardization and quality control of Trigonella foenum-graecum L. [Dataset]. Zenodo. 2024. Publisher Full Text" }	[ { "id": "340140", "date": "22 Nov 2024", "name": "Abdulrahman Koshak", "expertise": [ "Reviewer Expertise Pharmacognosy and natural products" ], "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 presents a novel analytical technique for quantifying the marker compound TG in fenugreek using UPLC-MS/MS. However, it would benefit from a more comprehensive discussion of the disadvantages associated with traditional methods in comparison to the developed approach. For example, elaborating on the advantages of UPLC-MS/MS, the types of solvents commonly used in traditional methods, their potential drawbacks, and how the new method overcomes these challenges would help to better clarify the research gap.\nTitle: Include the keywords \"green\" and \"UPLC-MS/MS\" to emphasize the innovative and advantageous aspects of the study.\nAbstract:\nClearly articulate the research gap and the drawbacks of traditional methods for quality analysis of fenugreek. Avoid mentioning (TG) in brackets, as it has already been identified earlier in the text. In the results section, include the units for extract yield and TG concentration for clarity.\nFormatting:\nEnsure the correct format (UPLC-MS/MS) is consistently used throughout the article. Avoid using the symbol \"±\" for reporting Standard Deviation; opt for a more descriptive format instead.\nResults:\nProvide the percentages of Trigonelline found in all samples to facilitate comparisons with values reported in the literature.\nDiscussion:\nThe first paragraph contains repetitive information regarding the introduction and methodology. Revise this section to focus directly on the findings and the specific research gap addressed by the study. Compare your findings for the amount of TG with those in the literature, expressed in percentages, to provide a clearer context and strengthen the relevance of your 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? 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": [ { "c_id": "12886", "date": "04 Dec 2024", "name": "Salma Hago", "role": "Author Response", "response": "Title Reviewer Comment: Include the keywords \"green\" and \"UPLC-MS/MS\" to emphasize the innovative and advantageous aspects of the study. Response: While we appreciate this suggestion, we have chosen to retain the current title because the study has already been published. Changing the title could cause confusion and inconsistency in citations and references to the work. However, we have ensured that the terms \"green\" and \"UPLC-MS/MS\" are included in the \"Keywords\" section to emphasize the innovative and advantageous aspects of the study. Abstract Reviewer Comment 1: Clearly articulate the research gap and the drawbacks of traditional methods for quality analysis of fenugreek. Response: The abstract has been revised to better articulate the research gap and the limitations of traditional methods, including their inefficiency, reliance on high solvent volumes, and lack of sensitivity. Reviewer Comment 2: Avoid mentioning (TG) in brackets, as it has already been identified earlier in the text. Response: The brackets around \"(TG)\" have been removed. Reviewer Comment 3: Include the units for extract yield and TG concentration in the results section for clarity. Response: Units for extract yield (mg/100mg) and TG concentration (ppb) have been added to the results section of the abstract. Formatting Reviewer Comment 4: Ensure the correct format (UPLC-MS/MS) is consistently used throughout the article. Response: The manuscript has been reviewed, and we have ensured that \"UPLC-MS/MS\" is consistently formatted throughout. Reviewer Comment 5: Avoid using the symbol \"±\" for reporting Standard Deviation; opt for a more descriptive format instead. Response: The symbol \"±\" has been replaced with descriptive phrasing (standard deviation, SD). Results Reviewer Comment 1: Provide the percentages of Trigonelline found in all samples to facilitate comparisons with values reported in the literature. Response: Percentages of Trigonelline in all samples have been added to Table 4. However, direct comparisons with literature values are not feasible because this study focuses on developing a green-sensitive method for trace analysis rather than optimizing an extraction method for high yields. Our approach utilized intact, uncrushed fenugreek seeds, whereas previous studies employed powdered seeds, which likely resulted in significantly lower TG %. Despite this difference, our method provides a rapid and reliable method to detect trace amounts of Trigonelline in fenugreek samples. Discussion Reviewer Comment 1: The first paragraph contains repetitive information regarding the introduction and methodology. Revise this section to focus directly on the findings and the specific research gap addressed by the study. Response: The first paragraph of the discussion has been revised to eliminate repetitive content. It now focuses on the study's findings and how they address the research gap. Reviewer Comment 2: Compare your findings for the amount of TG with those in the literature, expressed in percentages, to provide a clearer context and strengthen the relevance of your results. Response: The findings for TG content, expressed in percentages, have been compared with values from the literature to provide a clearer context and emphasize the significance of our results." } ] }, { "id": "340136", "date": "26 Nov 2024", "name": "Abdul Sadiq", "expertise": [ "Reviewer Expertise Pharmaceutical Sciences" ], "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 presents a scientifically sound and innovative method for the extraction and analysis of trigonelline in fenugreek seeds. The approach is environmentally sustainable, reproducible, and suitable for widespread application. Minor enhancements could be considered for future work, but the study meets the standards for publication. However, there are a few minor areas where clarity or wording could be slightly improved for better readability:\nAbstract: \"Fenugreek seeds contain a variety of phytochemicals, including proteins, lipids, amino acids, vitamins, flavonoids, steroidal saponins, coumarin, and alkaloids.\" Phytochemicals usually refer to biologically active compounds produced by plants, which usually include non-nutritive substances such as alkaloids, flavonoids, phenolic acids, tannins, terpenes, and saponins. If proteins or amino acids are being mentioned in the context of plant composition, it is more accurate to separate them from the term \"phytochemicals.\" A clearer phrasing for the description of fenugreek seeds might be: \"Fenugreek seeds are rich in proteins, lipids, and essential nutrients, and they also contain various phytochemicals, including flavonoids, steroidal saponins, coumarin, and alkaloids such as trigonelline.\"\nIntroduction: \"Fenugreek is cultivated in several countries worldwide, including the Mediterranean region, North African areas, India, and other countries.\" The phrase \"and other countries\" is unnecessary after \"including.\" Consider revising to avoid redundancy.\nMethodology: \"The USE protocol was selected due to the range of advantages USE presents, including high extraction efficiency and the use of the least possible sample amount, solvent volume, and extraction time.\" The phrase \"the range of advantages USE presents\" could be simplified to \"the numerous advantages of USE.\"\nConclusion: \"The method avoids the need for an extra step of seed crushing and sieving via the use of an ultrasonic dismembrator with small amounts of sample (10 mg) and low solvent volumes.\" Consider breaking this into two sentences for clarity, as the crushing and sieving highlight the simplicity of the method, while the use of small sample amounts and solvent volumes emphasizes cost-efficiency and eco-friendliness.\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": "12887", "date": "04 Dec 2024", "name": "Salma Hago", "role": "Author Response", "response": "Reviewer Comment 1: Phytochemicals usually refer to biologically active compounds produced by plants, which typically include non-nutritive substances such as alkaloids, flavonoids, phenolic acids, tannins, terpenes, and saponins. If proteins or amino acids are mentioned in the context of plant composition, it is more accurate to separate them from the term \"phytochemicals.\" A clearer phrasing for the description of fenugreek seeds might be: \"Fenugreek seeds are rich in proteins, lipids, and essential nutrients, and they also contain various phytochemicals, including flavonoids, steroidal saponins, coumarin, and alkaloids such as trigonelline.\" Response: We agree with the reviewer’s comment regarding the definition of phytochemicals. The sentence in the abstract has been revised to separate proteins and amino acids from the phytochemical category for accuracy. The updated sentence now reads: \"Fenugreek seeds are rich in proteins, lipids, and essential nutrients, and they also contain various phytochemicals, including flavonoids, steroidal saponins, coumarin, and alkaloids such as trigonelline.\" Reviewer Comment 2: \"Fenugreek is cultivated in several countries worldwide, including the Mediterranean region, North African areas, India, and other countries.\" The phrase \"and other countries\" is unnecessary after \"including.\" Consider revising to avoid redundancy. Response: We appreciate the reviewer’s suggestion to avoid redundancy. The sentence has been revised to remove \"and other countries\" for conciseness. It now reads: \"Fenugreek is cultivated in several countries worldwide, including the Mediterranean region, North Africa, and India.\" Reviewer Comment 3: \"The USE protocol was selected due to the range of advantages USE presents, including high extraction efficiency and the use of the least possible sample amount, solvent volume, and extraction time.\" The phrase \"the range of advantages USE presents\" could be simplified to \"the numerous advantages of USE.\" Response: We agree with the reviewer’s suggestion to simplify this sentence. The revised sentence now reads: \"The USE protocol was selected due to the numerous advantages of USE.\" Reviewer Comment 4: \"The method avoids the need for an extra step of seed crushing and sieving via the use of an ultrasonic dismembrator with small amounts of sample (10 mg) and low solvent volumes.\" Consider breaking this into two sentences for clarity, as the crushing and sieving highlight the simplicity of the method, while the use of small sample amounts and solvent volumes emphasizes cost-efficiency and eco-friendliness. Response: We appreciate this suggestion to enhance clarity. The sentence has been split into two, as follows: \"The method avoids the need for additional steps, such as seed crushing and sieving, by using an ultrasonic dismembrator. Furthermore, the method employs small sample amounts (10 mg) and low solvent volumes, ensuring cost-efficiency and eco-friendliness.\"" } ] } ]	1	https://f1000research.com/articles/13-1350
https://f1000research.com/articles/13-1530/v1	18 Dec 24	{ "type": "Research Article", "title": "Assessing eye health knowledge and practices amongst primary health care nurses in a rural district, South Africa  ", "authors": [ "Lwandile Flatela", "Zamadonda Xulu-Kasaba", "Lwandile Flatela" ], "abstract": "Background The burden of visual impairment and avoidable blindness has been attributed to the shortage and poor distribution of adequately trained eye care personnel. In the district health system, primary health care (PHC) nurses are the first point of contact for eye care patients. To provide adequate eye care, the nurses in PHC clinics apply their knowledge on basic eye health to ensure best practices and management for patients that present at their clinics. Anecdotal evidence regarding the knowledge and practices of these PHC professionals is scarce. The purpose of this study was to evaluate PHC nurses’ knowledge and practices on eye health.\n\nMethods A quantitative, cross-sectional study was conducted in a rural district of the Eastern Cape Province in South Africa. Stratified random sampling was used to select 28 from the 74 PHC clinics in the district. A researcher-administered questionnaire using a 5-point likert scale with options strongly disagree, disagree, neutral, agree and strongly agree, was developed, piloted and finalised for this study. The tool had four sections: demographic information, knowledge of eye health, practices on evaluating visual function and identification and management of presenting eye conditions. Data were collected, captured onto MSXcel, cleaned, coded and analysed descriptively, and for Chi-squared significance at 95%, using R statistical software.\n\nResults From 200 responses, 86.4% (n=172) were females of the African race. Most (93.5%, n=187) had never undergone formal training in eye care. Only 28% (n=56) correctly identified a mature cataract while 28% (n=56) correctly identified signs of glaucoma; the global leading causes of preventable blindness. A total of 94% (n=188) failed to accurately measure visual acuity on a patient.\n\nConclusions PHC nurses showed poor knowledge and practices of common eye conditions. There is an urgent need for eye health training for PHC nurses, and the clear management protocols on eye care.", "keywords": [ "Visual impairment", "Primary health care", "Primary eye care", "Nurses", "Eye health", "Primary health care clinics", "knowledge and practices", "rural eye health" ], "content": "Introduction\n\nVisual Impairment (VI) remains a major public health concern that continues to rise.1 Globally, an estimated 596 million people have reduced vision at distance, while more than 500 million suffer from uncorrected near vision.2 According to the World Health Organisation (WHO), 80% of all VI is preventable, while 90% is found in low-to-middle income countries (LMIC).3 It is further estimated that 76% of moderate and severe VI cases in southern Sub-Saharan Africa (SSA) are caused by cataract, glaucoma and uncorrected refractive error (URE).3,4 Of further concern are the dire socio-economic consequences of VI suffered by the affected individuals.5\n\nPrimary eye care (PEC) is one of the pillars of primary health care (PHC) recommended by the WHO.6 PEC involves eye health promotion, treatment of simple eye diseases, and identification and prompt treatment of persons needing eye care6; all areas that are pivotal in preventing blindness and VI. Poor and rural populations bear the burden of VI and its consequences as resources are skewed against these communities.4 Amongst other challenges, poor access to clinics with eye health services, weak infrastructure, and a shortage of human resources amplify the burden of VI in LMIC.7–10 Blanchet et al emphasise that poor integration of eye health services into the health systems and scarcity of eye health policies in LMICs have a negative contribution towards achieving improvement in visual health.11 This would further impede the attainment of the third Sustainable Development Goal centred on good health and vision.12\n\nIn South Africa’s district health system (DHS), the majority of patients needing PEC initially present themselves at PHC centres.13 The level of care provided has been described as inadequate in relation to community needs, and inferior to the standard prescribed by the National Department of Health (NDoH).14,15 As PHC workers are the first point of contact with patients, they are tasked with providing PEC to the communities in which they are based. To adequately deliver PEC services, PHC providers are tasked with the responsibility of identifying, managing and referring patients accordingly and timeously.7 There is a dearth of evidence evaluating their knowledge and practices in basic eye care services in the rural areas of SSA countries such as South Africa.16 This study aimed to assess PHC nurses’ knowledge and practices in PEC in order to evaluate how eye patients are being managed in PHC clinics within the district and other similar rural contexts.\n\n\nMethods\n\nAlfred Nzo District Municipality (AND) is the smallest of eight districts in the Eastern Cape province of South Africa. With 867 864 inhabitants (12.2% of the province’s population),17 AND is one of the most densely populated districts in the province. With a density of 81 people/ km2.17 AND is also the most impoverished, rural, and underdeveloped district in the province, coupled with a high unemployment rate.18 The district has four local municipalities, with 74 PHC clinics. To contrast, this is a higher density than Umkhanyakude District, in the neighbouring KwaZulu-Natal province, which is also rural with 625 846 inhabitants, and a density of 49 people/km2.19\n\nA cross-sectional study was conducted in 28 randomly selected Primary Health care clinics (PHCC) within the AND. A sample size was calculated using the single population proportion formula,20 where significance was at the 95% confidence level and 5% degree of precision. A figure of 50% was used for the expected knowledge base. District office registers, reporting on staff numbers and clinic contact details, were obtained from the AND office to ascertain a population size. These were approximately ten years old and confirmed a nursing workforce of 935 nurses in the PHCC within the district. There was no available updated version that accounted for deaths, retirements, and resignations. Factors such as retirement, deceased employees, and transfers away from the district, promotions, unfilled vacant posts, and the like were reasons given by clinic supervisors for actual staffing numbers being approximately 50% of that which had been anticipated according to the registers. To establish a more accurate sample size, the Principal Investigator (PI) telephonically communicated with the heads of each clinic, making appointments in advance to visit them for stakeholder engagement in preparation for data collection. On arrival at the clinics, it was apparent that the actual staffing was approximately 50% of that which had been anticipated according to the registers. As such, the population (N) was adjusted to 50% of 935 where N = 466.\n\nUsing the single population proportion formula, the minimum sample size calculation used was:\n\nz (for a confidence level of 95%, standard normal distribution) = 1.96\n\nd (acceptable/tolerated margin of error of 5%) = 0.05\n\np (if no prevalence/prior study the proportion is set at 50%) = 0.50\n\nN (population size) = 466\n\nThe recommended sample size was calculated, and a 5% error margin was further added to the sample size to produce a required minimum sample size (n) of 211.\n\nStratified random sampling was applied across the district to ensure even representation of the various municipalities. Proportioning of participants was applied to match the distribution of clinics in the various municipalities. Participant numbers were 35% (n = 70) from Mbizana, 15% (n = 30) from Ntabankulu, 35% (n = 70) from Mzimvubu and the remaining 15% (n = 30) from Matatiele (15%).\n\nThe Principal Investigator (PI) contacted the heads of each clinic again, making appointments in advance to visit them for data collection purposes. Low staff numbers were found at most sites due absenteeism (due to quarantine or contact tracing guidelines resulting from the COVID-19 pandemic), leave and illness. From the available workforce, the PI addressed the staff in their common room to inform them of study aims and voluntary participation, only the consenting PHC staff were enrolled in the study. Furthermore, some PHC staff members who had been enrolled, ultimately did not participate, citing the high numbers of patients that were still waiting to be seen during business hours. The final number of participants who were enrolled in the study was 200. Willingness of voluntary participation was confirmed by signing of the consent forms by each willing participant.\n\nThe contents of the data collection tool were informed by various primary eye health service documents and screening guidelines for ocular health in South Africa, Nigeria, Uganda and a few other African countries.5 Data collection tools from previous studies that had assessed primary eye care skills amongst nurses and community health workers within sub-Saharan Africa (SSA) were also used to compile the final questionnaire that was used in this study.21 After completion, the comprehensive questionnaire was piloted on 20 PHC nurses in another province, where input for improvement was requested and given. None of the original questions were removed, instead the wording was simplified in some for ease of understanding. After modification, the PI finalised the tool for use in the study. The tool was divided into three sections: Part 1 - demographic data; Parts 2 & 3 - knowledge of different eye health professions and identification of common eye conditions using coloured pictures and brief descriptions of defining symptoms, and Part 4 focused on practices through task observation of the participants by the PI who sat as a patient for all the participants when they performed the task. Participants were provided with a distance and near tumbling E chart and a tape measure for distance measurement. Here the PI scored each participant on their ability to perform a visual acuity measurement and thereafter record the findings. A marking guide was derived and used for this evaluation ( Table 1), and participants were considered to be competent if they obtained a minimum score of seven out of ten (70%).\n\nDue to poor internet connectivity issues, hard copies of the researcher-administered data collection tool were used. Each willing participant was individually invited into the tea room by the PI. The PI sat with one participant at a time, and guided on completion of the questionnaire. Where translation was requested, the PI assisted as he is conversant in the local language. After completion of the questionnaire, the PI asked the participants to measure distance and near visual acuity on him away from other staff members. Data was collected manually from all participants, captured onto Microsoft Excel, cleaned, coded, stored, and transferred to the R software for storage. Data were analysed descriptively for frequencies and chi-squared significance at the 95% confidence index.\n\n\nResults\n\nA 95% response rate (n = 200) was achieved, and participants were from 28 of the 74 PHC facilities across AND. The majority of 172 (86.4%) were female. All participants were from the African race, where 96% spoke the IsiXhosa language, and the remaining 4% spoke IsiZulu. Of further concern, is the fact that the majority (93.5 %) have never been formally trained in eye care. Further demographic details have been detailed in Table 2.\n\nNone of the visited PHCC had a Snellen or other visual acuity (VA) chart or a penlight torch. The PI took his own equipment to the sites for assessment purposes. Additionally, none of the surveyed facilities across the district had a staff member responsible for ophthalmic care, and none of the facilities had a designated eye care consulting room. Only 35.5% of the participants affirmed that they understand the management and treatment protocols for eye health.\n\nKnowledge of PHC nurses on different eye professionals ( Table 3).\n\nPractices in the measurement of VA ( Table 4).\n\nKnowledge and practices of different eye conditions and ocular pathologies ( Table 5).\n\n\nDiscussion\n\nThis study sought to evaluate PEC knowledge amongst the PHC nurses in AND, and further assess the current practices and protocols that are used in managing eye care cases. As most patients initially present at PHCC for their health concerns, including eye health, it is critical to assess the knowledge and practices of nurses that manage the larger population at those health facilities.\n\nIn our study the most common cadre of staff that participated were professional nurses of the female gender aged 41 to 50 years of age. This finding is similar to studies in Nigeria and other parts of South Africa, where most participants were females in the same age grouping.7,22 The female bias is to be expected as the nursing profession was a predominantly female career option, and viewed as an extension of the domestic role played by women in homes and other sectors of society.23,24 Additionally, its common traits of caregiving and nurturing also align nursing more with female social gender roles than male gender roles. Similarly, the age range of most participants is within the most common labour force in South Africa, as evidenced by Statistics South Africa in their reports.25 An overwhelming majority of the participants had a service record of 15 years, meaning that they had a fairly good idea of the PHC environment, with another two decades of service prior to their retirement. As such, they were within their earlier years of service and were still amenable to learning and skill development for the betterment of their careers.\n\nIt is satisfactory to learn that the majority of the participants were trained and qualified as professional nurses. The AND fares well in this regard as the South African Nursing Council (SANC) distribution report shows that the Eastern Cape province has an average of 61 % of the nursing staff trained as professional nurses,26 one of the higher prevalence’s in the country. This could be attributed to the many nurse training facilities that exist in the Eastern Cape Province and is a positive case for general public health in the communities they serve. The fact that all the participants spoke isiXhosa, the local language in the province was an indicator of a homogenous culture of the participants. In light of this, it will benefit learning if refresher courses use training material in this language.27 Previous evidence has affirmed that medical learning is positively enhanced when a mother tongue or native language is used.26\n\nOf significant concern, however, is the markéd shortage of Human Resources for Eye Health (HReH) in the PHCC, which was confirmed by the far ‘lower-than-expected’ numbers of study participants on arrival at the clinics. Staffing numbers at PHCCs were less than half of the numbers confirmed by the regulating office, and some clinics had a single nursing professional on duty for the day, tasked with seeing to the general and eye care of the entire community. Health professional shortages are a serious concern in Africa, worsened by the “brain drain”, or exodus of experienced HReH from poorer countries to economically affluent first world countries.28 This further exacerbates prevailing health system challenges and worsens the burden of disease in poorly resourced and underdeveloped African countries. Professional nurses are amongst the highest numbers of trained health professionals that are known to have left South Africa in search of improved working conditions, better staffing and financial rewards offered by first world economies.29 In communities such as the AND, this shortage is a reality that further marginalises the population, and impedes the delivery of basic eye care in their clinics.\n\nThe fact that nearly none of the employees at the PHCCs had undergone eye health training further intensifies the severe staffing deficit. The number of participants was lessened by the fact that the original sample size was incorrect, owing to poor record keeping. In many health sectors, it has been argued that good record keeping is associated with appropriate standards of nursing, improved patient care, and swift reaction in the instance of emerging legal issues.30–32 It is imperative that this be improved for clear, transparent data and reporting in the future. In this study, this was seen as a significant part of the dire issues that add to the barriers to the delivery of adequate eye care to communities in the AND. Policymakers who use the staffing numbers that the district office has are under the impression that the district’s PHCC are well staffed, which is contrary to the reality of the situation on the ground. Consequently, this insufficient staffing worsens the burden of diseases, increases VI and the prevalence of avoidable blindness, and negates the achievement of universal health coverage through reduced service delivery, realised due to staffing shortages. The district needs to improve its staffing of eye clinics and its record keeping to ensure accessibility of sufficient eye care. Better access to eye care will further ensure improved learning opportunities for children through the school health system, improved employability opportunities for young adults, and subsequently reduced levels of poverty in this region.\n\nPHC workers are the first point of care for many rural South African communities and the first point of contact for health conditions in developing countries.21 There is an expected level of knowledge and skill that is required in order to provide effective management of eye conditions, and ensure that preventable blindness is addressed timeously.33\n\nTable 3 shows that almost half (46%) of the respondents did not know the basic roles and competencies of the different HReH, which indirectly affects their referral practices, as good knowledge of these would enable them to adequately triage ocular cases to the correct HReH. In knowing what the key roles of ophthalmologists, optometrists, opticians and ophthalmic nurses are, the referral of cases that could not be managed in the PHCC would be accurate with a reduced waiting period for the patient.\n\nKnowledge of common eye conditions were also very weak. Similar results were found in Pakistan and Ethiopia, where poor knowledge of basic ocular conditions was noted amongst various health professionals.15 Reasons expressed for this poor knowledge was the fact that in their training, general practitioners spend a maximum of four weeks only in the ophthalmology rotation, which is a very short training period for one to gain sufficient knowledge and information on this area of healthcare.34,35 Similarly, with nurse training in South Africa, there is no clinical component in ophthalmology or optometry, which is probably why the knowledge levels in this study were very poor. Previous studies have also shown that inadequate eye care knowledge, demonstrated by nurses, reduced their confidence in dealing with patients with eye problems.35,36 Additionally, the clinical knowledge of basic ocular conditions, and leading causes of blindness and visual impairment were poorly identified and managed by the respondents in the AND.\n\nCataract is the leading cause of preventable blindness in SSA and the world, and an easily identifiable condition; however, approximately one quarter of the respondents identified it correctly. Worse still, the management of this condition was seen as urgent and immediate, which is inaccurate, and the cause of unnecessarily overloading referral transport and overloading an already strained system that could have given the ambulance space to a more deserving patient. This prevalence is significantly lower than that found in a similar study in Nepal where most PHC workers were able to identify a cataract accurately.37 Similarly in other studies within African countries: Tanzania, Malawi, Kenya and Nigeria, PHC nurses were more knowledgeable in identifying and managing patients with cataract, VKC and glaucoma.38,39 Foreign body sensations were the only condition that although not accurately identified, was managed adequately in this study. This was probably due to the discomfort, pain, and urgency that patients present with. In KwaZulu Natal Province, South Africa, knowledge regarding primary eye care was found to be relatively good amongst participants in PHCs within hospitals.15\n\nThis was probably due to the fact that hospitals employ ophthalmic nurses and optometrists in their primary care and outpatient departments. Unlike optometrists and ophthalmic nurses, most of the PHC nurses in this study were not HReH trained in eye health.\n\nPoor knowledge and management of patients by PHC nurses are concerning factors as they marginalise patients in poor, rural communities, deter universal eye health access, and exacerbate the burden of disease. Clinic PHC nurses should be well-versed in eye care to protect patients, avoid ocular complications and adequately address ocular problems as this is all directly linked to visual impairment.40 Health service managers should look to improving this as it has far-reaching effects on community wellbeing and general health.\n\nMost of the participants in our study reported to have never had any eye health training. The very few who have received proper training had not had a refresher course. As a result, the majority of eye care cases were then referred, even those that could have been managed timeously and locally. There were poor management guidelines in place and no set referral criteria used. In agreement, previous studies reported inappropriate referral pathways in northern Nigeria39 while, to the contrary, evidence reported a clear and precise referral pathway in India.41\n\nThis low number of trained PHC workers in PEC is in line with a study in Nigeria that found that not even one of the respondents had received training on PEC in their facility.42 On the contrary, in a similar study done in Kenya it was found that a much higher proportion (two thirds) of the participants in PHCC had received PEC training.38 It stood to reason that the Kenyan study reported that almost all PHC workers provided adequate eye care with confidence, which could be cited as an example of competence overreaching as those that were not trained learned from the others and managed patients satisfactorily.38,43 In this study, the extremely low number of PHC nurses who had received some form of eye health training could be linked to the basic nursing curriculum – which does not mention eye health – and lack of continued professional development in the nursing profession. The majority of those who had received training in eye care had never completed any refresher eye care training. In our study, only two of the respondents were ophthalmic nurses, and both were working as professional nurses due to a lack of specialised post funding at clinic level. To further exacerbate the problem is the fact that only three nursing colleges offer ophthalmic nurse specialisation training in the entire country. Some non-governmental organisations (NGO’s) like The Fred Hollows Foundation have tried to bridge this gap by offering some kind of eye care training for PHC workers; however, this has not been sustained leading to its eventual failure.\n\nIt is clear that nurse training needs to better embrace eye health training so as to effectively manage cases in the PHCC. Poor identification of leading causes of blindness and visual impairment, poor knowledge of HReH and poor management of eye health cases all point to much needed training and refresher training of nurses in eye health. This needs to be addressed with a view to providing eye health training.\n\nThe study results show that the ability to carry out a visual acuity assessment by the PHC workers in the AND is far below the expected level ( Table 4). A great majority of the participants did not meet the minimum competency levels of this clinical technique and scored less than 7/10 on assessment. Visual acuity is an important measure of visual function and is necessary for decision making with ophthalmic patients. Visual acuity is the most frequently performed measure of visual function in clinical practice and most people worldwide living with VI are living in low and middle-income countries.44 Visual acuity measurements then ought to be routinely conducted with all patients visiting the PHC clinic.7 Inaccurate execution, as displayed by those in this study, further marginalises scholars and older patients who could have played a significant role in society had the visual impairment been assessed accurately, and visual assistive devices or treatment initiated for improved vision. Similarly in studies in Nepal and Eastern Africa, only 14% and 12.3% of the PHC workforce respectively could measure visual acuity.7,37 Furthermore, the poor outcome in our study is also not surprising considering that none of the visited PHC clinics had a Snellen chart to measure patients’ visual acuity.\n\nThe study location was in a deep rural area with an extremely poor roads network, making it difficult to reach the clinics on the extreme periphery of the region. Inclusion of these facilities may have added substantial information to the study. A general shortage in staffing was, however, noted in the clinics where staffing ranged from one PHC worker on duty, where the study could not be prioritised over patients that were waiting to be served. In PHCCs such as this, we were forced to help where we could and turn back as participation in the study was not possible. It would be beneficial for the study to be conducted in more districts to get a better understanding of the knowledge and practices of PHC nurses on a larger scale.\n\nData collected and analysed in the study sets a clear basis on the need for training and development of eye health knowledge in PHCC such as this one, more especially the ones in rural settings where local services are a dire need, due to financial limitations. It is imperative that training insitutions prioritise developing materials to meet this need and empower HReH in PHCC.\n\nWe recommend basic eye health training to be included in the nursing curriculum to equip PHC workers with the necessary skills to improve their role in VI. Refresher courses are recommended for staff in PHCC to ensure adequate skill levels in their clinics. Training materials should be provided in the local language to enhance the homogenous culture in this province. Secondly, there needs to be an establishment of guidelines and funnelling protocols for eye health cases. HReH posts must be created, funded, and filled for sufficient staffing and effective eye care.\n\n\nConclusion\n\nMost of the nurses in this district are females within a suitable working age, who speak the local language spoken by patients in that district. This study showed that the PHC nurses in the Alfred Nzo region had inadequate eye health knowledge on HReH and basic eye conditions that are leading causes of visual impairment and blindness globally. This has a grave impact on the treatment, referral and management of treatable and preventable visual impairment conditions in the region. PHC nurses in this area did not know how to measure visual acuity. Most nurses were never trained in eye health. None of the clinics visited had a Snellen chart for visual acuity measurement. There were no basic guidelines and referral protocols that could be referred to for eye conditions. Eye health is clearly not prioritised in this district. Health managers need to prioritise eye health, specifically PEC, and seek to train nurses in order to provide adequate universal eye health within the communities they work.\n\n\nDeclarations\n\nWritten ethical clearance was obtained from UKZN BREC/00000663/2019 and gatekeeper permission was obtained from the DoH, Alfred Nzo District, and the DoH, Eastern Cape Province.\n\n\nConsent to participate\n\nInformed written consent was obtained from all participants included in the study.\n\n\nAuthor contributions\n\nDetailed contributions from the authors have been tabled in Table 6 below.\n\n\nEthical considerations\n\nFollowing ethical clearance by the Biomedical Research Ethics Committee (BREC/00000663/2019) at the University of KwaZulu-Natal in May 2020, and gatekeeper permission by the Eastern Cape Department of Health (DoH), data collection was delayed by the national lockdown imposed by the COVID-19 pandemic. When restrictions were lifted, re-certification was obtained, and data collected from February to June 2021.", "appendix": "Data availability statement\n\nData in the form of questionnaire response spreadsheets and ethical permissions is available on Zenodo underproject name “Assessing eye health knowledge and practices amongst primary health care nurses in a rural district, South Africa” at https://doi.org/10.5281/zenodo.14150666.\n\nAll extended data is available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication). Access to this dataset requires registration with an IEEE account, which is free.\n\n\nAcknowledgments\n\nThe authors would like to thank all the participants who voluntarily agreed to participate and give valuable information to make this study successful.\n\n\nReferences\n\nHolden BA, Fricke TR, Wilson DA, et al.: Global prevalence of myopia and high myopia and temporal trends from 2000 through 2050. 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PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "361697", "date": "04 Feb 2025", "name": "Michael Bowen", "expertise": [ "Reviewer Expertise Optometry", "Eye health", "Survey Design and analysis", "Psychology" ], "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\nStudy Setting section: It would be useful to include details of the number of Primary Health Care clinics in KwaZulu-Natal, which is used as the comparison province to the study province.\nThe use of \"AND\" as the acronym for Alfred Nzo District Municipality is potentially odd for readers less familiar with this acronym. For may English speakers AND looks like a capitalized \"and\". It might be helpful to change to using \"ANDM\" as the acronym.\nStudy Design section: in this section there is a statement that \"a figure of 50% was used for the knowledge base\" - this is somewhat unclear. Does this mean that respondents were assumed (for the purposes of the power calculations) to be likely to answer 50% of the questions correctly? This would benefit from some clarification.\nResults section: Table 3 is titled \"Basic knowledge on roles of different Human resources for eye health (HReH)\" however, it is not clear that the data being reported relate to an objective assessment of knowledge. Several of the questions for which response data are reported here are reporting participants view of their own knowledge. It would be preferable to re-title and ensure that there is some text added to clarify the nature of these data. They are still of interest - but need to be accurately described.\nThe methods section mentions a 5 point Likert scale - it is not clear from any of the tables which data were derived from these questions. Ideally a copy of the questionnaire used should be included as an annex or within supplementary materials. It is slightly difficult to review the paper without seeing the questionnaire.\nTable 5 - this table reports data relating to participants ability to correctly identify eye health conditions from photographs. It was not clear from the methods section whether these photographs had been tested by a group of eye health professionals (such as optometrists / ophthalmologists) to provide a base-line for the rate at which qualified eye health professionals were able to correctly identify the conditions from the images. This is important as it provides some validation that the images are 'good enough' representations of the conditions in question. It would be helpful to share some of the images used in the paper or as supplementary material.\nTable 5 - It is not clear from the table or the text in the results section whether the data relating to the %ages of participants who made the correct management responses relate to the whole group or just to those who were able to correctly identify the condition. This is important to the interpretation of these data, so would be helpful to make this clear. For example, for Uveitis, if 16% of the participants correctly identified it as Uveitis, but only 32% of those who correctly identified it knew that it needed an urgent referral, that provides different insights than if if 16% correctly identified the Uveitis, but 32% of all the participants knew it needed an urgent referral. In the latter case, even though they didn't know what the condition was an additional 16% of participants knew that it needed urgent referral.\nTable 5 - Primary Open Angle Glaucoma - it is unclear from the table, or from the information available elsewhere in the paper, what information was actually given to participants to test their ability to identify POAG and how the information given would lead to the correct management being no referral - monitor yearly. The information summarised in the table suggests that 'throbbing eye pain' was one of the symptoms offered to participants, which does not necessarily suggest stable POAG or OHT. It is also unclear how any of the PHC's in the study would be able to safely monitor OHT or stable POAG, given their lack of equipment. This question and the associated data need to be reviewed by the authors and their inclusion in the final paper considered carefully.\nDiscussion section:\nPage 7, paragraph 3: \"...one of the higher prevalence's\" (a) if prevalences is to be used, the apostrophe needs to be removed; (b) I am not sure that it is helpful to use \"prevalences\" in reference to the number of qualified nurses in the work force in a region. I would suggest editing this to read: \"....one of the higher proportions of qualified to unqualified nurses in the country.\"\nSame paragraph: With reference to the participants mainly speaking isiXhosa, it would be useful to readers to include some information about the population in that region - was the percentage of isiXhosa speakers among the participant sample representative of the rates of isiXhosa speakers in the population of ANDM?\nPage 8, paragraph 2: the second sentence in this paragraph is confusing: \"The number of participants was lessened by the fact that the original sample size was incorrect, owing to poor record keeping\". This is not really correct. The original population size was incorrect due to the records of nursing staff being inaccurate. While this error could have led to an incorrect sample size being calculated, the investigators took appropriate measures to establish a more reliable / accurate estimate of the population size, and thus were able to derive a more reliable sample size. However, since it appears that the authors intention in this section is to address the concerning lack of accurate records for nursing staff numbers, I would suggest simply rewording this along the lines of: \"The challenges encountered by the research team in this project in seeking to establish a reliable estimate of the total number of nursing staff employed within PHCs across ANDM are noteworthy. The initial population number provided was nearly double the final population established by the researchers.\"\nIn the same paragraph the sentence: \"Consequently, this insufficient staffing worsens the burden of diseases, increases VI and the prevalence of avoidable blindness...\" while likely to be true, for a research paper it is slightly hyperbolic. It would be preferable to reword along the lines of: \"The very low levels of PHC staff seems likely to contribute to increased levels of VI and preventable sight loss / blindness, and potentially a worse burden of disease across the province, as it is hard to see how such low levels of PHC staffing would fail to lead to reduced service delivery / access to care. These low staffing levels are likely to make it much harder to achieve universal health coverage in a meaningful way.\"\nPage 8 - Eye health knowledge section: First paragraph, final sentence \"...that preventable blindness is addressed timeously.\" \"Timeously\" should be replace by \"in a timely way\".\nSecond paragraph of Page 8 - \"Table 3 shows...\" in this this paragraph the authors suggest that the poor awareness of the various eye health professions' roles \"...indirectly affects referral practices...\". It is not clear how the data from this research supports this assertion. If this is not a direct finding of the research. For example - was there an analysis of the data that showed that those who incorrectly identified the competencies and roles of the different eye health professionals performed less well on the questions relating to making correct management decisions where these related to referral? In any case, lower knowledge in this area seems highly likely to directly affect referral practices - this could be offered as an observation if there is not a specific analysis to support this as a \"finding\" per se.\nThird paragraph in this section, starting with \"Knowledge of common eye conditions...\" in this first sentence \"were\" needs to be replaced with \"was\".\nPage 8, final paragraph: I am not sure it is helpful to refer to the \"prevalence\" of a poor capacity of participants to correctly identify cataract - this should be reworded along the lines of: \"The low level of knowledge of cataract in this study is significantly lower than...\" Same paragraph - the grammar in the sentence on Foreign body sensations is a little odd - suggest rewording along the lines of: \"'Foreign body sensations' was the only condition which, although not correctly identified by the majority of participants, produced appropriate management responses from an acceptable proportion of participants\".\nTraining in eye care section: it is unfortunate that the analyses of the data reported here do no include any analysis of whether the nurses who HAD received some eye health training did in fact perform better on the identification and management tasks, as well as on the awareness and perception of other eye health professions competencies. While the numbers who received training may have been too small to support a fully reliable sub-group analysis, this seems too important a question not to at least investigate to some extent, even if in reporting the low numbers in the \"with training\" group had to be raised as a cautionary factor.\nThe reference to \"There were poor management guidelines in place and no set referral criteria used\" appears to introduce / refer to data or information not reported earlier in the paper. It would certainly be interesting to have seen data relating to the management guidelines used in the PHCs that participated in the study, as well as they referral criteria for those sites. Without data, this assertion lacks the impact it could have otherwise.\nSecond paragraph in this section: the sentence starting: \"It stood to reason that the Kenyan study....\" this sentence is somewhat muddled in the section \"...which could be cited as an example of competence overreaching as those that were not trained learned from the others and managed patients satisfactorily.\" I think that the authors mean that the Kenyan study could be cited as an example of where the training of some staff lead to improved performance by all the staff due to the untrained staff learning from those who had received training - if this is the case, that sentence would benefit from rewording.\nClinical skill and observation and practice norms: the first sentence in this section seems potentially problematic. Since this research did not compare 'community / primary care nurses' generally in South Africa or elsewhere with the PHC nursing staff, it seems perhaps unreasonable to expect them to know how to do even a basic refraction examination, especially since almost none of them have had any eye health training either in their training to qualification or post-qualification. How are the research team defining \"the expected level\"? When and where was this level established / agreed and how was it communicated to nursing or leadership staff in the PHC system within ANDM? I think it would be preferable to reword the first two sentences for this section along the lines of:\n\n\"The study highlighted that current nursing staff within the ANDM PHC system lack the ability to carry out a basic visual acuity assessment, which should be a core requirement for the PHC clinics to be able to provide. Our data showed that the majority of participants in this study scored 6/10 or less on this clinical technique for assessing visual acuity.\"\nWithin the recommendations section it would be helpful to consider adding something about establishing clear referral guidelines and pathways (which the authors mention earlier in the text) and developing and providing all PHC clinics with eye health diagnostic guides and management guidance. It would seem feasible for an \"eye health pack\" to be developed, to include Snellen (or even LogMAR) and tumbling E charts, tape measures, occluders, vision assessment check lists (perhaps as posters for display for ease of staff use) and so on.\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? No\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/13-1530
https://f1000research.com/articles/13-22/v1	08 Jan 24	{ "type": "Systematic Review", "title": "Beyond traditional teaching: a systematic review of innovative pedagogical practices in higher education", "authors": [ "Josefina Amanda Suyo-Vega", "Víctor Hugo Fernández-Bedoya", "Monica Elisa Meneses-La-Riva" ], "abstract": "Background Pedagogical best practices play a pivotal role in ensuring the academic success of students within the higher education landscape. This study aims to systematically synthesize innovative pedagogical best practices within the university context.\n\nMethods We conducted a thorough systematic review using the rigorous PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) methodology. Our review involved comprehensive searches of scientific databases, including Eric, Scopus, and Proquest, covering both Spanish and English publications. We strategically employed Boolean operators like AND and OR to create a robust search equation. Our primary research question guiding this investigation was: “What innovative pedagogical practices have been developed in university settings to improve teaching and learning effectiveness?” This central question led us to delve deeply into the strategies utilized, pedagogical approaches adopted, and the noticeable impact achieved after their implementation. Additionally, we carefully established stringent inclusion and exclusion criteria in accordance with PRISMA guidelines to ensure methodological rigor.\n\nResults A total of 25 scholarly articles that met the pre-established criteria were meticulously identified and included in this systematic review. The results were thoughtfully categorized into three distinct teaching strategies: the first emphasizing student-centered approaches, the second showcasing the integration of educational technology, and the third highlighting evaluation and feedback methodologies.\n\nConclusions This systematic exploration of pedagogical best practices underscores their paramount importance in driving continuous improvement in teacher training and fostering innovation within the educational arena. Such practices not only create an engaging and effective learning environment but also set the stage for ongoing advancements in the teaching and learning processes.", "keywords": [ "Pedagogical practices", "Innovative pedagogical practices", "Teaching strategies", "Academic success", "Higher education" ], "content": "Introduction\n\nIn the university system, the ability to provide quality education is the highest expression that is achieved through the faculty. In this regard, the good practices developed by university professors become essential elements to ensure the success of the learning outcomes established in each subject.\n\nThe educational community periodically conducts workshops, seminars, and other courses for the professional development of faculty members, regardless of their specialization (Chukwuemeka & Samaila, 2020; Latorre-Cosculluela et al., 2023). Additionally, within the university environment, faculty members must remain constantly updated on technology-related topics and knowledge (Alvites, 2019). In this context, a literature review has identified effective teaching practices with the aim of benefiting higher education institutions and selecting the best technology-related teaching strategies (Khoza, 2022), leaving a gap in strategies and evaluations. However, when researching English teaching strategies in classrooms, six strategies were identified: memory, cognitive, compensatory, metacognitive, affective, and social. Researchers suggest further qualitative research in this area (Tieocharoen & Rimkeeratikul, 2019).\n\nRegarding contextual and pedagogical aspects, emerging pedagogical best practices in higher education address concerns related to assessment and improvement proposals at the end of the undergraduate thesis. However, there is a shortage of information on the methodological development of these best pedagogical practices (Camilli Trujillo et al., 2022).\n\nThe updating and implementation of pedagogical approaches in university institutions create a connection between teaching and learning, positively impacting the quality of education and student engagement. A relevant methodological approach is the use of workshops as complements to the curriculum (Asensio Pastor, 2019).\n\nThis research focuses on the systematization and analysis of effective pedagogical practices in the university context. Its importance lies in its direct impact on the quality of university education. These best practices not only enhance the student learning experience but also strengthen the ability of academic institutions to adapt to student needs. By understanding these practices, it is possible to promote and/or improve certain strategies, thus enhancing the quality of higher education. From this, the guiding question arises: “What innovative pedagogical practices have been developed in university settings to improve teaching and learning effectiveness?” This question involves the description of strategies and the evaluation of the pedagogical process. The research aims to systematize and provide scientific evidence for teaching professionals to use in their work and promote meaningful learning.\n\nRegarding the design of activities and materials, current research indicates that the role of pedagogical practices and learning experiences presents challenges for both faculty and students in terms of necessary materials (Tadesse et al., 2020). Furthermore, faculty members are always willing to introduce changes in teaching methodology (Mataka et al., 2022). These changes or planned strategies are aimed at achieving academic goals, with the condition of achieving academic success through learning (Ayu, 2021). Pedagogical practices emphasize the reflective aspect, not only for increasing knowledge but also for establishing a relationship between research and practice (Sotomayor-Soloaga, 2021).\n\nIn Spain, research was conducted on the flipped classroom as a pedagogical strategy in higher education, concluding that it is necessary to combine the flipped classroom with gamification. This strategy improves academic performance and autonomy (Carpena Arias & Esteve Mon, 2022). Similarly, research in Thailand explored the use of various strategies for language learning, revealing statistically significant differences in memory, cognitive, affective, and social strategies among students (Tieocharoen & Rimkeeratikul, 2019).\n\nA study conducted in Chile, focusing on teachers considered “the best teachers,” identified prominent characteristics such as promoting student autonomy, encouraging student participation, activating prior knowledge, and resolving doubts, strategically managing errors, creating a challenging thinking environment, asking good questions, making the most of time and space, having clarity in the evaluation system, and maintaining physical and emotional proximity (Sotomayor-Soloaga, 2021).\n\nReviewing theories about best pedagogical practices reveals new subcategories based on teaching practices, allowing faculty to examine, interpret, and recreate their pedagogical practice, turning it into a source of learning from a perspective of change and innovation (Merellano-Navarro et al., 2020).\n\nThe research highlights the need to use effective teaching strategies, suggesting that the debate method, among others, can improve students’ critical thinking (Ahmod & Zhang, 2021; Crolla et al., 2019). Furthermore, the use of various strategies offers the opportunity to significantly transform the teaching process, promoting effective student learning (Tadesse et al., 2021). Debate becomes a pedagogical tool for both faculty and students when providing quality feedback (Crolla et al., 2019). Another way to actively participate in and collaborate in the learning process is through intrinsic or extrinsic motivation (Tulyakul et al., 2022).\n\nThe literature review reveals dimensions of best pedagogical practices. Evaluation is present throughout the learning process and requires the presence of a methodology to achieve self-directed learning. It also requires faculty preparedness for various situations (Porcher, 2020). Developing good pedagogical practices strengthens students’ critical thinking, enabling them to analyze information not only quantitatively but also qualitatively, complementing research (Mahdi et al., 2020).\n\nLikewise, technology and tools are necessary and are often used in classrooms through digital games (Cavalcante-Pimentel et al., 2022). Finally, flexibility and adaptability are essential, as research results focus on the combination of the flipped classroom and gamification to improve motivation, academic performance, and autonomy of university students (Carpena Arias & Esteve Mon, 2022). Another relevant aspect for improving pedagogical practice is international study trip experiences (Ellinghaus et al., 2019).\n\nBased on the theoretical foundations of Ausubel’s Meaningful Learning, which emphasizes that learning is most effective when it relates to meaningful prior knowledge, it is necessary to examine Mishra and Koehler’s TPACK model. This model describes the intersection of pedagogical, technological, and content knowledge necessary for effective teaching (Vásconez Paredes, Darío & Mauricio, 2021).\n\nAdditionally, from this foundational structure, other forms of knowledge intersection emerge, including (a) Pedagogical Knowledge, which involves the fusion of pedagogical skills and experience; (b) Content Knowledge, which comprises a set of skills that teachers possess regarding the meaning of pedagogical content; (c) Technological Knowledge, which implies that faculty members stay updated and develop technological skills; (d) Pedagogical Content Knowledge, which involves not only having knowledge about a specific subject but also how to convey that knowledge; (e) Technological Pedagogical Knowledge, which integrates technology, pedagogical strategies, and pedagogical content; (f) Technological Content Knowledge, which involves integrating technology into the teaching of specific content; and (g) Technological Pedagogical Content Knowledge, which encompasses pedagogy, technology, and content (Farhadi & Öztürk, 2023; Maipita et al., 2023).\n\nBased on the presented theory, there is a need to delve deeper into the best pedagogical practices developed at the university level.\n\n\nMethods\n\nThe research follows a qualitative approach with a hermeneutic design. This design allows for an in-depth exploration and interpretation of various concepts presented by different authors, aiming to achieve a more comprehensive and enriched understanding. Furthermore, the research is classified as applied research since it works with existing elements but seeks to establish new concepts or methods to address the identified issue, aligning with the principles outlined in the Frascati Manual 2015 (OECD, 2015).\n\nTable 1 in the article presents the detailed procedures developed for each subcategory, providing a structured framework for the research methodology and analysis.\n\nTo identify relevant scientific articles, a specific set of keywords was carefully chosen. These keywords served as the initial search parameters, but the search was not limited to these terms alone. Synonyms and related terms were also incorporated to expand the scope of the search. Table 2 provides a comprehensive list of the selected keywords along with their corresponding synonyms. It is important to emphasize that these keywords represent the culmination of multiple search iterations, resulting in the formulation of a search ‘equation.’ This equation is designed to be replicable by fellow researchers in the future, thus contributing to the ongoing advancement of knowledge in the field.\n\nLikewise, a search equation was employed, utilizing Boolean operators OR and AND. The OR operator was used for synonyms, while AND was employed to connect the keywords. The equation was structured as follows: (Good pedagogical practices OR Effective teaching strategies) AND (university professors OR Higher education teachers).\n\nIt is worth noting that the search was conducted across three databases. The first was the “Educational Resources Information Center” (ERIC), specialized in Education. The second database was Scopus, known for its high-quality content for researchers, and finally, the Proquest database, accessed through advanced search functions, for potential future replication purposes.\n\nTo locate articles, a set of inclusion and exclusion criteria were applied. The initial criterion stipulated that articles must fall within the temporal range of 2015 to 2023. The decision to select this time frame was informed by the observation that there was no significant increase in scientific article production in the preceding decade (2005-2015). This suggests that research in this area began to gain relevance and increase in quantity starting from 2015 (Gómez Velasco et al., 2022).\n\nFurthermore, articles identified as conference papers and reviews were excluded due to their limited presence. Additionally, postgraduate work publications are infrequent and have limited impact on the international scientific community (Mamani Benito et al., 2021).\n\nIt should be noted that duplicate articles across any of the three databases were removed. Finally, the authors conducted a thorough selection process, focusing on articles most pertinent to the research, based on the abstract and conclusion of each article. Table 3 provides a detailed overview of the procedures conducted.\n\nLikewise, the PRISMA diagram was created, as shown in Figure 1, providing a graphical representation of the preselection process (Page et al., 2021).\n\nThe research was structured in accordance with the PRISMA statement. In the methods section, the following criteria were taken into account: (a) Protocol and registration, (b) Eligibility criteria, (c) Sources of information, (d) Search strategy, (e) Study selection, (f) Data collection process, and (g) Data items (Hutton et al., 2016). All of these points aim to adhere to the procedure outlined in the PRISMA statement, as illustrated in Table 4. A completed PRISMA checklist can be found as Extended data (Suyo-Vega, Fernández-Bedoya, & Meneses-La-Riva, 2023).\n\n\nResults\n\nAfter the selection of articles, an in-depth analysis was conducted as detailed in Tables 5 and 6.\n\n\nDiscussion\n\nFrom the analyzed best practices, three key strategies that promote teaching and learning in higher education can be grouped as follows:\n\na) Student-Centered Teaching Strategies: In this approach, educators recognize the importance of actively involving students in their own learning process. Autonomy is valued, and individual student needs and learning styles are respected, fostering critical thinking (Ahmod & Zhang, 2021; Crolla et al., 2019; Mahdi et al., 2020; Tadesse et al., 2021; Tulyakul et al., 2022).\n\nb) Online Teaching Strategies and Educational Technology Usage: Faculty integrate educational technology and adapt it to online teaching environments, utilizing digital tools to enhance teaching and learning. This turns learning into a source of pedagogical innovation (Carpena Arias & Esteve Mon, 2022; Cavalcante-Pimentel et al., 2022; Merellano-Navarro et al., 2020).\n\nc) Assessment and Feedback Strategies: These strategies enable educators to provide constructive and ongoing feedback to students with the aim of improving learning and academic growth (Maipita et al., 2023; Vásconez Paredes, Darío & Mauricio, 2021).\n\nIn conclusion, promoting pedagogical best practices based on student-centered strategies and self-directed learning is essential for improving the teaching and learning process in higher education, beyond traditional teaching methods. These practices allow educators to tailor instruction to student needs and learning styles, promoting student autonomy and motivation.\n\nIncorporating strategies using technological tools makes teaching effective and enriches the learning experience in virtual environments within the university setting. Additionally, the use of digital tools and innovative methodologies facilitates access to education and provides rapid and effective feedback to students.\n\nResearch on pedagogical best practices in higher education is essential for the advancement of teaching and learning. It provides valuable information that benefits both current and future educators, equipping them with tools to address challenges and adapt strategies to their context. Moreover, it contributes to the continuous improvement of teacher training and fosters innovation in the educational sphere, creating a more stimulating and effective environment for the teaching and learning process.\n\nCertain limitations were identified during the research process. One limitation is related to the timeframe. The research was conducted between 2015 and 2023. However, to gain a more comprehensive and longitudinal view of trends in the literature, it would be beneficial to extend the study period by several additional years.\n\nAnother limitation is related to the availability of sources. Despite having access to a variety of databases such as ERIC, SciELO, and Scopus, it has been observed that some research of interest is not available in full PDF format, hindering the sharing of certain findings with the scientific community.\n\nThis research significantly contributes to existing knowledge in the field of pedagogical best practices in the university context. Through document analysis, online teaching and student-centered learning, two fundamental aspects of modern pedagogy, were addressed.\n\nThe research team proposes alternatives such as the implementation of a continuous professional development program for faculty, designed to be accessible virtually and on an ongoing basis. This initiative aims to ensure that faculty members are in a constant state of development and skill updating, ultimately translating into a high-quality learning experience for students.\n\nIn this context, it is recommended to expand research to other modalities and educational levels, as well as to explore successful experiences in countries that have effectively adopted these pedagogical practices.", "appendix": "Data availability\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\nZenodo: PRISMA 2020 checklist for ‘Beyond Traditional Teaching: A Systematic Review of Innovative Pedagogical Practices in Higher Education’ https://doi.org/10.5281/zenodo.8404402 (Suyo-Vega, Fernández-Bedoya, & Meneses-La-Riva, 2023).\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nAhmod U, Zhang W: An effective instructional strategies approach in higher education: A pilot investigation. 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Publisher Full Text\n\nRuiz de Gauna P, González Moro V, Morán-Barrios J: Ten pedagogical keys to promote good practices in competency-based Medical Education, for undergraduate and postgraduate. Educacion Médica. 2015; 16(1): 34–42. Publisher Full Text\n\nSchwartzman G, Roni C, Berk M, et al.: Remote learning assessments in higher education: Teaching decision making when facing a new challenge. RIED-Revista Iberoamericana de Educacion a Distancia. 2021; 24(2): 67–85. Publisher Full Text\n\nŞeker M, Inan Karagül B: Assisting higher education learners online to acquire self-regulated writing strategies during Covid-19. Journal of Learning and Teaching in Digital Age. 2021; 7(1): 64–75. Publisher Full Text\n\nSotomayor-Soloaga PA: Good pedagogical practices in science teaching. Magis. 2021; 14: 1–23. Publisher Full Text\n\nSun H, Liu T: Student-Centered online teaching practices in theoretical mechanics. Higher Education Studies. 2021; 11(2): 233. Publisher Full Text\n\nSuyo-Vega JA, Fernández-Bedoya VH, Meneses-La-Riva ME: PRISMA 2020 checklist for Beyond Traditional Teaching: A Systematic Review of Innovative Pedagogical Practices in Higher Education. Zenodo. 2023. Publisher Full Text\n\nTadesse T, Asmare A, Ware H: Exploring teachers’ lived experiences of cooperative learning in ethiopian higher education classrooms: A phenomenological-case study. Education Sciences. 2021; 11(7). Publisher Full Text\n\nTadesse T, Manathunga C, Gillies RM: Teachers’ pedagogical practices and students’ learning experiences in an Ethiopian university setting. Asian Journal of University Education. 2020; 16(2): 205–225. Publisher Full Text\n\nTieocharoen W, Rimkeeratikul S: Learning strategies and teaching methods in Thai and Vietnamese universities. Arab World English Journal. 2019; 10(3): 99–112. Publisher Full Text\n\nTulyakul S, Werathummo A, Khun-inkeeree H, et al.: The motivation and teaching strategies in pre-service physical education teachers. Journal of Education and Learning (EduLearn). 2022; 16(2): 164–171. Publisher Full Text\n\nUnesco: 74 Buenas prácticas docentes. Experiencias con tecnología en aulas peruanas. Unesco; 2017; Vol. 53(9).\n\nVásconez Paredes, Darío C, Mauricio IOE: El modelo de aprendizaje TPACK y su impacto en la innovación educativa desde un análisis bibliométrico The TPACK learning model and its impact on educational innovation from a bibliometric analysis. INNOVA Research Journal. 2021; 6(3): 79–97. Publisher Full Text\n\nZamora-Polo F, Corrales-Serrano M, Sánchez-Martín J, et al.: Nonscientific university students training in general science using an active-learning merged pedagogy: Gamification in a flipped classroom. Education Sciences. 2019; 9(4). Publisher Full Text\n\nZhong L: Strategies and practices related to teaching large online classes. Journal of Higher Education Theory and Practice. 2018; 18(1). Publisher Full Text" }	[ { "id": "268905", "date": "10 May 2024", "name": "Budi Waluyo", "expertise": [ "Reviewer Expertise ELT", "Education", "and Education Technology." ], "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\nDear Authors,\n\nFirst, I have seen frequently utilized vocabulary in generative AI applications. If you utilize any of these applications, it is essential to acknowledge and elucidate the extent to which they were employed in the composition.\nSecond, the initial paragraph of the introduction holds paramount significance as it serves to inform the readers about the evolution of the research domain, the scope of prior investigations, and the unique addition of your study. Nevertheless, your initial paragraph lacks any of these vital elements. You should modify it in accordance with the necessary changes.\nFurthermore, your introduction lacks criticality and persuasiveness. To enhance the clarity of the current literature and highlight the contribution of your study, it is necessary to critically synthesize the findings of relevant studies.\nThird, it is imperative to formulate research questions in order to provide a clear focus for your research and facilitate the comprehension of your reported findings by readers.\nFourth, it is necessary to include a literature review section in order to establish robust foundational frameworks. This style of study appears to be lacking in strength.\n\nFifth, it is necessary for you to provide more detailed information about the results in paragraphs, not just showing tables.\nSixth, your analysis lacks sufficient depth and fails to adequately engage with the current related literature.\nAccording to these six factors, it is necessary for you to make significant revisions to this manuscript.\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? Partly\n\nIs the statistical analysis and its interpretation appropriate? Partly\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.) No", "responses": [] }, { "id": "291882", "date": "02 Jul 2024", "name": "Jay Fie Paler Luzano", "expertise": [ "Reviewer Expertise Mathematics 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\nThe rationale for and objectives of the systematic review are partly stated. The study aims to synthesize innovative pedagogical best practices within the university context to improve teaching and learning effectiveness. However, there may be room for further clarification or expansion on the specific objectives. Sufficient details of the methods and analysis are partly provided to allow replication by others. The study followed the PRISMA methodology, including criteria such as protocol, eligibility criteria, sources of information, search strategy, study selection, data collection process, and data items . This structured approach enhances the replicability of the study. The review did not specifically mention statistical analysis, as it focused more on synthesizing innovative pedagogical practices rather than statistical data analysis . Therefore, the appropriateness of statistical analysis and its interpretation is not applicable in this context. The conclusions drawn in the review are supported by the results presented, which include an in-depth analysis of selected articles and the impact of innovative pedagogical practices in higher education. The study aimed to delve deeply into the strategies utilized, pedagogical approaches adopted, and the noticeable impact achieved after their implementation, which supports the conclusions drawn.\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? Partly\n\nIs the statistical analysis and its interpretation appropriate? Partly\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.) Partly", "responses": [ { "c_id": "12930", "date": "10 Dec 2024", "name": "VICTOR HUGO FERNANDEZ BEDOYA", "role": "Author Response", "response": "Thank you for your comments, we have managed to make improvements in the new version of the manuscript, which will be online in the following days. Meanwhile, we attach the responses to each of the observations. Regarding the Selected Countries The choice of countries was not random; the selection was intentional because they represent diverse educational contexts across Europe, Asia, and America. The choice of Spain, Thailand, and Chile was based on their relevance in recent research exploring various aspects of effective pedagogical practices across different cultural and educational settings. The studies from these countries allow for an analysis of effective strategies that have been adopted and evolved according to their unique characteristics. For example, research conducted in Thailand (Tieocharoen & Rimkeeratikul, 2019) focuses on analyzing language learning strategies, such as memory, cognitive, affective, and social strategies. Although these strategies pertain to the student’s domain, the study illustrates effective pedagogical strategies. Regarding Learning Theories and Their Importance This research is grounded in Ausubel’s Meaningful Learning Theory, which emphasizes the importance of connecting new knowledge with meaningful prior concepts to achieve deep learning. Furthermore, Mishra and Koehler’s TPACK Model is essential for analyzing the intersection of pedagogical, technological, and content knowledge within effective teaching practices. These theories frame the analysis by allowing the integration of innovative teaching strategies within specific contexts, thereby contributing to advancements in higher education research. Regarding the Expanded State of the Art The research does not merely summarize the state of the art but also expands it by analyzing key theories within the university context. This approach provides added value by offering an updated and nuanced understanding of the topic to address contemporary pedagogical challenges. Furthermore, by identifying the application of these theories in classroom settings, this study contributes evidence to enrich existing knowledge about pedagogy and content in higher education teaching. Regarding Theories and Discussion Strategies such as debate and gamification reflect Ausubel’s principle of connecting new learning to prior significant knowledge, facilitating the assimilation and application of knowledge. Similarly, the inclusion of technology and pedagogical tools aligns with the TPACK Model, which emphasizes the integration of pedagogical, technological, and content knowledge to enhance teaching practices. Regarding Why This Research is Categorized Within Social Sciences According to Émile Durkheim, education is a key social phenomenon in the reproduction and transformation of society. It plays a fundamental role in the socialization of individuals and in the transmission of norms, values, and knowledge that ensure social cohesion. Therefore, research on pedagogical practices falls within the realm of social sciences because it influences the formation of individuals within a social context (Durkheim, 2015). Regarding the Exclusion of Systematic Reviews The systematic reviews were excluded from the results section because this research focused on empirical studies—both quantitative quasi-experimental designs and qualitative case studies—that directly addressed innovative pedagogical practices. Although systematic reviews are valuable, they do not align with the study's aim of obtaining primary and context-specific data on the practical application of these strategies. Therefore, the research prioritized original studies that provided direct evidence and practical insights into pedagogical practices. Additionally, during the article review phase, documents in PDF format were identified, but in some cases, only titles and abstracts were available rather than full texts. Despite access to well-recognized databases such as ERIC, SciELO, and Scopus, restrictions in editorial access limited the inclusion of some relevant research. These constraints are highlighted under the Limitations section." } ] }, { "id": "301470", "date": "24 Jul 2024", "name": "Hoang-Yen Phuong", "expertise": [ "Reviewer Expertise TESOL", "testing and assessment", "higher 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\nThe review has several limitations that may affect its comprehensiveness and depth. Covering articles only from 2015 to 2023, it might miss earlier developments or long-term trends in pedagogical practices. Although it includes publications in Spanish and English, it could still overlook important research in other languages, leading to potential language bias. The reliance on three databases, despite their reputability, might not capture all relevant studies, and the lack of a formal quality assessment of the included studies is a notable omission in a systematic review. Furthermore, the review primarily offers qualitative insights without conducting a meta-analysis or robust statistical analysis, which could have strengthened the evidence for the identified practices. It also has a broad focus on all innovative practices in higher education, potentially sacrificing depth in specific areas. The limited discussion of contradictory findings and the absence of detailed exploration of how practices vary across different countries or cultural contexts restrict the review's ability to provide a balanced and comprehensive view. Other weaknesses include the omission of student perspectives and outcomes, which are crucial for understanding the effectiveness of teaching practices. The authors acknowledge limited access to full-text articles, which might have excluded relevant studies, and they do not address potential publication bias, possibly skewing the results toward positive findings. Finally, the review does not extensively discuss the challenges and barriers to implementing innovative practices in various higher education settings, limiting its practical applicability.\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? Partly\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.) Partly", "responses": [ { "c_id": "12928", "date": "17 Dec 2024", "name": "VICTOR HUGO FERNANDEZ BEDOYA", "role": "Author Response", "response": "Dear Reviewer: Thank you for your comments, we have managed to make improvements in the new version of the manuscript, which will be online in the following days. Meanwhile, we attach the responses to each of the observations. Regarding the Timeframe (2015 to 2023) According to the UNESCO report (2015), the global education landscape experienced a radical transformation in methods, content, and learning spaces. This transformation has significantly impacted higher education. Consequently, the need for a shift in pedagogical practices emerged, gaining momentum in 2015 in response to demands for more flexible, student-centered education adapted to the digital age (UNESCO, 2015). Additionally, the decision to focus on this period was based on the observation that the preceding decade (2005–2015) did not see a significant increase in the scientific production of articles. This suggests that research began to gain relevance and increase in volume starting in 2015 (Gómez Velasco et al., 2022). Regarding the Language Articles in English and Spanish were analyzed due to the predominance of these languages in the selected databases, such as SciELO, Scopus, and ERIC. These platforms are recognized for their focus on disseminating research in these languages, facilitating access to a wide range of scientific literature (Márquez Jiménez & Alcántara Santuario, 2017). Regarding the Three Databases SciELO: This database was selected because it democratizes access to scientific knowledge in Latin America, highlighting regional research previously considered \"lost science\" due to limited distribution (Crespo Fajardo, 2019). Scopus: Chosen for its broad international coverage and reputation as a key resource for evaluating scientific output. Scopus includes a large proportion of articles in English (67%) and Spanish (86%), providing access to impactful and relevant research across various disciplines (Márquez Jiménez & Alcántara Santuario, 2017). ERIC: Selected for its specialization in education worldwide and its provision of access to relevant literature in the field (Mosquera Ayala, 2017). Regarding Meta-Analyses or Solid Statistical Methods The research aimed to systematize innovative pedagogical practices in higher education, which aligns more closely with a systematic review than a meta-analysis. Meta-analyses focus on quantifying effects using numerical data. Additionally, many of the studies reviewed did not provide sufficient or consistent data, such as means and standard deviations, necessary for conducting a meta-analysis (Debby Ng et al., 2022). Regarding the Lack of Comparisons Between Countries The main objective of the research was to systematize best pedagogical practices in higher education, identifying effective strategies in the academic field. This required a detailed analysis of key elements, such as pedagogical approaches, types of strategies, and evaluation methods, without cross-country comparisons being part of the initial design. This methodological decision was based on the goal of identifying general patterns applicable on a global scale rather than focusing on local specificities. Additionally, systematic reviews must clearly define their scope to ensure analytical coherence and avoid excessive dispersion of results (Grant & Booth, 2009). Regarding Challenges and Barriers to Implementing Innovative Practices The decision not to address challenges in detail was due to the limited scope of this research." } ] }, { "id": "301479", "date": "07 Aug 2024", "name": "Maria Alfredo Moreira", "expertise": [ "Reviewer Expertise Language Education", "Teacher Education", "Pedagogy in Higher 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\nThe text reports a systematic review study focusing on a critical aspect of higher education quality, as is pedagogical practices. It addresses a relevant research question (\"What innovative pedagogical practices have been developed in university settings to improve teaching and learning effectiveness?”), a question that \"involves the description of strategies and the evaluation of the pedagogical process.\". It partially achieves its aim, of \"systematiz[ing] and provide scientific evidence for teaching professionals to use in their work and promote meaningful learning.\" The methodology is sound, well-laid out and with a clear description of the path that was followed in order to achieve its results. However, the Introduction does not address in a systematic and coherent manner the key concepts of the study: Good pedagogical practices AND effective teaching strategies. The studies that sustain this section seem to have been selected rather randomly, as it goes into language teaching details (listing language learning strategies as if they would be a synomym of pedagogical strategies, for example), several countries (like Spain, Chile or Thailand - why these countries?), or learning theories (Ausubel's, MIshra & Koehler - what is their importance for defining the key constructs? How were these key constructs used in interpreting the results?). It is not clear how the state of the art outlined in this section will be expanded or enriched by the study. As the Discussion section does not confront the results with the concepts in the introduction, it is not clear what the advancement in the field was. Even though the methodology is sound, there are minor issues that would improve the text. Only by reading Figure 1 the reader becomes aware that the focus is on social sciences (why this focus?), as it is neither stated in the research question nor in the introduction. And why were systematic reviews excluded? Figure 1 reports exclusion of pdf formats but in the discussion it is stated: \"it has been observed that some research of interest is not available in full PDF format\". Thus, it is not clear whether the format of the text was a criterion for exclusion or not.\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\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": [ { "c_id": "12929", "date": "17 Dec 2024", "name": "VICTOR HUGO FERNANDEZ BEDOYA", "role": "Author Response", "response": "Dear reviewer: Thank you for your comments, we have managed to make improvements in the new version of the manuscript, which will be online in the following days. Meanwhile, we attach the responses to each of the observations. Regarding the Specific Objectives The general objective of the study was to systematize innovative pedagogical best practices in the university context. Additionally, the specific objectives included: Analyzing pedagogical best practices alongside their respective methodological approaches in the university setting. Examining the applied strategies, focusing on the types of strategies (student-centered, online teaching or technology-based, and evaluation or feedback strategies). Evaluating the methods used to assess pedagogical practices, ensuring a comprehensive understanding of their effectiveness." } ] } ]	1	https://f1000research.com/articles/13-22
https://f1000research.com/articles/13-1528/v1	17 Dec 24	{ "type": "Case Report", "title": "Case Report: Rare Bilateral Double Profunda Femoris Artery Variation: Clinical Implications and Considerations", "authors": [ "Prasanna Gaydhanker", "Soumya Vinod", "Ernest Adeghate", "Sahar Mohsin", "Prasanna Gaydhanker", "Soumya Vinod", "Ernest Adeghate" ], "abstract": "Introduction The profunda femoris artery (PFA) is the largest branch of the femoral artery supplying the thigh. The study reports a rare occurrence of bilateral double profunda artery in a male cadaver, aiming to inform surgeons of such variations to avoid complications during surgical interventions.\n\nMethods This study was conducted during routine dissections using 5-10% formalin-fixed lower limbs of a male cadaver at the Department of Anatomy, College of Medicine and Health Sciences, United Arab Emirates University, UAE\n\nResults In the right lower limb, two profunda femoris artery (PFA) branches arise from the common femoral artery (CFA) 5 cm from the inguinal ligament, one laterally and one posteromedially. The medial (MCFA) and lateral circumflex (LCFA) arteries branch from these PFAs respectively. The femoral nerve is anterior to the lateral PFA and LCFA and lateral to the posteromedial PFA. The obturator nerve runs posterior to the posteromedial PFA. In the left lower limb, two PFAs originate from the CFA, 2.5 and 5 cm from the inguinal ligament, lateral and posterior to the CFA, respectively. The LCFA stems from the upper PFA, while the medial MCFA arises from the CFA. The femoral nerve lies anterior to both the PFA and LCFA. The obturator nerve is posterior to the MCFA. The femoral vein crosses over the MCFA and the lower posterior PFA, distal to the MCFA origin.\n\nConclusion This report enriches understanding of PFA anatomy and its variations, emphasizing the importance of tailored surgical approaches to minimize risks and ensure procedure success.", "keywords": [ "Profunda femoral artery", "Anatomical variation", "Cadaver", "Femoral artery", "Bilateral", "Nerve", "Complications", "Surgeons." ], "content": "Background\n\nThe thigh region of the lower limb is supplied by the ‘femoral artery’ and its branches. Originating from the common iliac artery in front of the sacroiliac joints at the level of L5-S1, The ‘femoral artery’ originates from the ‘external iliac artery’. It enters the lower limb region behind the inguinal ligament, located halfway between the anterior superior iliac spine and the pubic symphysis. Positioned anteriorly within the femoral triangle of the thigh, the ‘femoral artery’ passes through an opening in the ‘adductor magnus’ muscle before becoming the ‘popliteal artery’ in the area behind the knee joint. It then gives off branches to supply blood to the leg and foot regions.1\n\nThis study explores the variability observed in the primary branch emanating from the ‘femoral artery’ in the thigh region, the ‘profunda femoris artery’ (PFA). The PFA supplies the ‘medial’ and ‘posterior’ compartments in the thigh region. Both the ‘femoral artery’ and PFA play crucial roles in various clinical contexts, ranging from diagnostic imaging to surgical reconstruction and vascular access.2 Therefore, awareness of variations in PFA branches and their relationship with nerves enhances surgical precision, reduces complications, and ultimately improves patient outcomes across various clinical specialties, including surgery, interventional radiology, and neurology.\n\nThe PFA originates from the ‘femoral artery’ approximately 3.5 cm below the inguinal ligament.1 The PFA is also referred to as the ‘deep femoral artery’ and, in this case, the segment of the ‘femoral artery’ preceding the origin of the PFA is referred to as the ‘common femoral artery (CFA), while the portion of the femoral artery following the origin of the PFA is termed the ‘superficial femoral artery’ (SFA). The PFA at its origin is lateral and then spirals posterior to the femoral artery. Proximally, the ‘femoral vein’ lies medially and the PFA courses between the ‘pectineus’ and the ‘adductor longus’ muscles, subsequently passing between the ‘adductor longus’ and ‘brevis’ muscles, and finally descends in a gap bordered by the ‘adductor longus’ and ‘adductor magnus’ muscles. It concludes by transitioning into a ‘fourth perforating artery’ and establishing connections with the muscular branches of the ‘popliteal artery’ within the ‘adductor magnus’ muscle.1\n\nMultiple variations have been documented regarding the profunda femoris artery (PFA), including alternative origins such as medial, posterior, or posterolateral instead of the typical lateral position, as well as instances where it shares a common origin with one of the circumflex arteries arising from the ‘femoral artery’.3–6 The superficial branches of the ‘femoral artery’ are also reported to be arising from the PFA. However, duplication of the PFA is extremely rare,7,8 Tsoucalas et al., 2018 reported a double PFA in the left thigh of a female cadaver, no variations were found in the distribution pattern of each branch, and the relationship with nerves was not discussed.7\n\nThe PFA emits two circumflex arteries, medial and lateral, as well as perforating arteries through which it delivers oxygenated nutrients to the proximal aspect of the femur and the ‘medial’ and ‘posterior’ compartments of the thigh, aiding in adduction, extension, and flexion movements.1–3\n\nAnatomists have observed variations in the origin of the circumflex arteries as well. The LCFA is the major branch of the PFA found in 67% of the population originating 1.5 cm below the lateral aspect of the PFA, however, in 20% of people may arise directly from the ‘femoral artery’.9 The LCFA travels horizontally between the divisions of the ‘femoral nerve’ and posteriorly passes behind the ‘sartorius’ and ‘rectus femoris’ muscles.3 https://en.wikipedia.org/wiki/Lateral_circumflex_femoral_artery . Afterwards, it splits into ascending, transverse, and descending branches. Typically, the artery runs anteriorly to the femoral neck.2 Variations in the relationship of the ‘femoral nerve’ to the PFA and circumflex arteries are also reported.1,10–12\n\nMCFA is important for the blood supply to the proximal regin of the femur. It is given off from the PFA posteriorly anterior to the ‘iliopsoas’ and behind the ‘pectineus’ muscles. It can be traced further between the ‘obturator externus’ and the ‘adductor brevis’ muscles. Occasionally in 25% of the population it may originates directly from the femoral artery. Damage to the MCFA following a femoral neck fracture can lead to avascular necrosis of the femoral neck or head.1,12\n\nAim & Objectives: Here we report a case of bilateral occurrence of a double PFA originating from the ‘femoral artery’ in an adult male cadaver and its relationship with nerves. Data regarding arterial variations is significant for orthopedic surgeons, vascular specialists, and radiologists as it aids in mitigating complications during surgical procedures.3 Knowledge of the relationship between the PFA and nerves helps minimize the risk of iatrogenic nerve injury during surgical procedures which can significantly impact patient outcomes and quality of life.\n\n\nMethods\n\nThis study was performed on the 5-10% formalin-fixed (Manf-Cibachem, Cat No: 233344) lower limbs of male cadavers in the ‘Department of Anatomy, College of Medicine and Health Sciences, United Arab Emirates University, UAE.’ The study was performed during standard dissection for undergraduates as per Cunningham’s manual instructions. The dissections involved examining the thigh compartments and tracing the ‘femoral artery’ and its branches from beginning to end. The origin, path, and branching pattern of the ‘femoral artery’ were explored on both sides, and images were taken and compared with previous literature.\n\n\nResults\n\nIt was observed that the ‘femoral artery’ follows the usual course passes under the inguinal ligament enters the femoral triangle bounded medially by the adductor longus and laterally by the sartorius with the femoral vein lying medially and ‘femoral nerve laterally’. The variation is noticed in the branching pattern of CFA as it was giving two profunda arteries bilaterally. In the right lower limb both PFA are arising from the CFA at the same level 5 cm from the inguinal ligament. However, one is arising laterally and the other is coming off posteromedially from the CFA ( Figure 1). The lateral profunda femoral artery is bounded laterally by the iliopsoas muscle and the posteromedial branch traverses between the ‘pectineus’ and the ‘adductor longus’ muscle.\n\n(A) Double Profunda femoris artery (PFA) origin from the common femoral artery (CFA). The lateral branch is labelled as (1) and posteromedial one by (2). Muscular branches from the posteromedial PFA to the adductor muscles ae indicated by yellow arrow. Adductor longus, rectus femoris, pectineus and iliopsoas muscles are indicated by (AL), (RF) (P), and (IP) respectively. The Middle Circumflex Femoral artery (MCFA) originating from the posteromedial PFA is shown by a red arrow. The femoral vein (FV) lies medial to the femoral artery and its branches. Tributaries of the femoral vein in anterior relation to one of the branches of the posteromedial PFA are shown by a white arrow (B) The rectus femoris (RF) muscle is reflected to show the lateral circumflex femoral artery (LCFA) coming from the lateral PFA in the right lower limb. The LCFA is shown by a yellow asterisk () coming off from the lateral PFA posterior to the branches of the femoral nerve which is shown by the yellow arrow.\n\nThe posteromedial branch of the PFA gives muscular branches to the adductor muscles ( Figures 1 & 2). The posteromedial branch in the mid-thigh gives multiple branches to the ‘adductor magnus’ muscle as shown in Figure 2A. The ‘femoral vein’ lies medial to the posteromedial branch of the PFA ( Figures 1 & 2). The posteromedial profunda artery goes from posteromedial to posterolateral in mid-thigh running behind the SFA ( Figure 1). The ‘vastus medialis’ muscle receiving a branch directly from the SFA is shown in Figure 2B.\n\n(A) Showing double profunda arteries arising from the CFA in the right thigh region. The femoral nerve (FN) is lateral to the CFA and some branches of FN are related anteriorly to the lateral PFA (red double-head arrow). The femoral vein (FV) is medial to the femoral artery and the great saphenous vein (GSV). The posteromedial profunda femoris branch shown by the yellow () gives multiple branches to the adductor muscles (yellow arrows). The lateral branch gives muscular branches indicated by red arrows to the quadriceps femoris (rectus femoris (RF); vastus lateralis (VL) and vastus intermedialis (VI). Adductor Longus and rectus femoris muscles are indicated by (AL) and (RF). The rectus femoris is cut and reflected to show the vasti muscles (B) The CFA gives out two profunda femorals and continues as superficial femoral artery (SFA). One arising posteromedially () and another profunda femoral coming off laterally (xx). The rectus femoris muscle is cut and reflected to see the branches of the PFA. The lateral branch is giving branches to the quadriceps femoris shown (x). The vastus medialis muscle is getting a direct branch from the superficial femoral artery indicated by the red arrow in right thigh region (C) The branches from the posteromedial profunda to the adductor longus muscle (AL) and the gracilis muscle (G) are shown by yellow arrows are related to the branches of the obturator nerve red () in the right thigh region.\n\nThe ‘femoral nerve’ is anterior to the lateral profunda and lateral to the posteromedial PFA in the right thigh region. The lateral PFA laterally traversing between the divisions of the ‘femoral nerve’ gives branches to the ‘quadriceps femoris’ muscle. An LCFA is coming off the lateral PFA posterior to the branches of the femoral nerve ( Figure 2B). The ‘obturator nerve’ is positioned posterior to the branches originating from the posteromedial PFA towards the ‘adductor longus muscle’. The posteromedial PFA gives a tributary to the ‘gracilis’ muscle, initially positioned lateral to the ‘obturator nerve and subsequently coursing between its branches that supply the ‘gracilis’ muscle ( Figure 2C).\n\nThe MCFA that supplies the neck of the femur can be seen coming off the posteromedial PFA and traversing medially anterior to the pectineus muscle to supply the neck of the femur ( Figure 1A). The branches of the posteromedial branch of the profunda femoris are related closely to the tributaries of the femoral vein some of them crossing anterior to the branches of the artery ( Figure 1A). The LCFA comes off from the lateral PFA posterior to the branches of the ‘femoral nerve’ ( Figure 1B).\n\nOn the left lower limb, we also found two profunda femoris arteries arising from the CFA ( Figure 3). The first PFA commences laterally from the ‘femoral artery’, approximately 2.5 centimetres from the inguinal ligament, while the second originates below the first one at a distance of 5 centimetres from the inguinal ligament and is posterior to the CFA at its commencement and later traverses laterally to the ‘femoral artery’. The lower posterior branch from the profunda artery is bigger than the upper lateral branch.\n\n(A) Upper (lateral) profunda (1) piercing the vastus medialis (VM muscle and lower (2) (posterolateral) one pierces the adductor magnus (AM) muscle and Superficial Femoral artery (SFA) is passing through the adductor hiatus (B) Upper profunda giving branch to tensor fasciae latae muscle (TFL) and quadriceps femoris (QF) shown by yellow arrows.\n\nThe upper lateral branch supplies the ‘quadriceps femoris’ muscle ( Figure 3) and ends by piercing the ‘vastus medialis’ muscle. The upper branch also gives branches to the ‘tensor fasciae latae’ muscle on the outer aspect of the thigh ( Figure 3B). A lower posterior branch gives perforating arteries to the ‘adductor’ muscles and ends by piercing the ‘adductor magnus’ muscle above the adductor hiatus ( Figure 3A & B).\n\nThe LCFA is given off from the upper profunda artery vessels in the left thigh region ( Figure 4A). The LCFA vessel was found to pass posterior to the ‘sartorius’ and’ rectus femoris’ muscles and is crossed by the ‘femoral nerve’ and its branches. Transverse, Ascending, and descending branches from the lateral circumflex artery are depicted in Figure 4, directed towards the ‘vasti’ muscles. The MCFA however originates from the ‘femoral artery’ slightly above the lower profunda artery, as illustrated in Figure 4B.\n\n(A) Upper profunda labelled as 1 is giving out lateral circumflex artery () posterior to the femoral nerve (F.N) and its branches. Ascending, transverse, and descending branches from the lateral circumflex are shown (). The descending branch is shown by the yellow arrow from LCFA is given to the vastus lateralis (VL) and vastus intermedialis (VI). (B) The common femoral artery (CFA) is giving out lower profunda artery labelled as (1) and medial circumflex artery (MCA) is originating from the CFA posterior to the femoral vein (FV) in the left thigh region. The great saphenous vein (GSV) opens into the femoral vein.\n\nThe ‘femoral nerve’ lies lateral to the ‘femoral artery’ proximally and distally it is related anteriorly to both profunda vessels ( Figure 5A). The ‘obturator nerve’ is posterior to the MCFA ( Figure 5B).\n\n(A) Obturator nerve is shown by red asterisk is posterior to the medial circumflex femoral A (MCFA) shown by an arrow. Femoral nerve (FN) crossing over the PFA (B) Magnified image showing MCFA crossing over the obturator nerve. Femoral vein (FV) is anterior to the MCFA in the left thigh region.\n\nIt was found that as the ‘femoral vein’ ascends through the adductor canal, it follows a standard trajectory, initially situated posterolateral to the ‘femoral artery’. Subsequently, it shifts to a posterior position before assuming a medial placement within the femoral triangle, where it consistently lies medially to the femoral artery at its commencement in both limbs ( Figures 1A, 4B, 5). In the left thigh region, the ‘femoral vein’ crosses over the ‘medial circumflex femoral artery’ (MCFA) and is found anterior to the lower posterior PFA, distal to the origin of the MCFA before the lower PFA perforates the adductor muscles (see Figure 4B and Figure 5).\n\n\nDiscussion\n\nThe presence of bilaterally arising double profunda femoris arteries from the CFA is a noteworthy anatomical variation with crucial implications for medical professionals involved in vascular procedures and femoral region surgeries. Understanding these variations, including the branching pattern of the CFA and its relationships with adjacent structures, is essential for medical practice and surgical procedures due to their significant clinical impact.\n\nThe relationship of these PFAs and their branches with the femoral nerve is noteworthy. The femoral nerve is found to be related anteriorly to the profunda arteries on both sides. In the right lower limb, the lateral PFA laterally traverses between the branches of the ‘femoral nerve’ and gives branches to the ‘quadriceps femoris’ muscle. This relationship has implications for surgical procedures involving the ‘quadriceps femoris’ and surrounding structures.\n\nThe MCFA plays a crucial role in the vascularization of the head and neck of the femur, as indicated by previous research.13,14 There is a discrepancy in the literature regarding the origin of circumflex arteries and their relationship to the anatomical structures, its origin has been documented to occur from the medial or posterior aspect of the ‘deep femoral artery’.1 Understanding such variations is important to prevent iatrogenic errors in various surgical procedures.10,15\n\nThe current investigation revealed a noteworthy difference between the right and left lower limbs. In the right limb, both the MCFA and the LCFA were found to arise from the PFA. Conversely, on the left side, the MCFA commenced from the CFA, while the LCFA stemmed from the PFA.\n\nA prior study involving 342 dissected hemipelves, reported diverse origins of the MCFA.15 It arose from the ‘common’ and ‘deep femoral’ artery in 39.3%, from the ‘superficial femoral’ artery in 2.5%, and the LCFA in 0.6%. Notably, it was found to be congenitally absent in 0.6%. These findings underscore the variability in the anatomy of the MCFA, which holds clinical significance for surgical interventions.\n\nThe present study found the ‘femoral vein’ or its tributaries traverses anteriorly to MCFA on the left and right proximal thigh region. Previous studies have reported that PFA arising medially from the ‘femoral artery’ passed anterior to the ‘femoral vein’.8\n\nThe LCFA, originating from the lateral PFA in the right lower limb, traverses posterior to the branches of the femoral nerve. This anatomical detail is pertinent to procedures involving the blood supply to the lateral thigh region. The anterior relation of the ‘femoral nerve’ to the LCFA is reported earlier by Goel et al 2015 as a ‘rare variant’.10 However, Classen et al (2020) have shown in their study that the’femoral nerve’ passed mostly anterior (46.4%) or anterior and posterior (47.8%) to the LCFA and rarely they may pass entirely posterior to the LCFA (5.8%).16 The ‘obturator nerve on the left side is found situated behind the MCFA and on the right side, it is related closely to the muscular branches coming off the posteromedial PFA to the ‘gracilis’ and ‘adductor longus’ muscles.\n\nThe ‘femoral’ and ‘obturator’ nerves arise from the ventral rami of the lumbar plexus (L2-L4). The ‘femoral nerve’ supplies the anterior thigh muscles and transmits sensory information to the anteromedial aspect of the thigh.\n\nThe ‘obturator nerve’ supplies motor and sensory signals to the inner thigh, primarily innervating the adductor muscles. Damage to either the ‘femoral’ or ‘obturator’ nerve can result in muscle weakness and sensory disturbances in their respective regions, with conditions such as trauma, hernias, or pelvic compression affecting these nerves.\n\nComprehensive knowledge of variations in the origin, number, or course of blood vessels and their relationship with nerves is crucial for medical professionals, especially in surgery and trauma management. Understanding these connections is vital during procedures like hip surgery to minimize inadvertent damage. Surgeons must be aware of these relationships to prevent unintended nerve or vessel damage during interventions. Pathologies like aneurysms impacting arteries can affect adjacent nerves, and nerve damage may compromise blood flow, leading to ischemic issues. Radiologists interpreting imaging studies need awareness of normal anatomy for accurate diagnosis. Diseases like vasculitis can concurrently affect nerves and arteries, resulting in complex clinical presentations. Overall, a deep understanding of anatomical variations in nerve-artery relationships is crucial for safe medical care in surgery, diagnostics, and clinical management.\n\n\nConclusions\n\nDuring the routine dissection of a male cadaver’s lower limb for medical students, a bilateral occurrence of double profunda femoris artery originating from the femoral artery was observed. The study delves into the nerve relationships with the profunda and its circumflex branches, emphasizing the significance of comprehending these connections for lower limb surgical and interventional procedures.\n\n\nEthics and consent statement\n\nThe study reports findings from a single cadaver as a part of routine dissection for medical students. The cadaver used in the study was obtained through ‘Body Scientific International, LLC’. They worked with the body donation program, Medcure and Anatomical Gift Association of Illinois, where written informed consent was obtained from the donor or the donor’s legal representative prior to death on November 14, 2019. The study respects the donor’s wishes for their body to be used for educational and research purposes. The study was conducted in accordance with the principles of the Declaration of Helsinki and followed standardized protocols outlined by AQUA (Anatomical Quality Assurance) and IFAA (International Federation of Associations of Anatomists. All personal data related to the body is confidential as outlined by AQUA.", "appendix": "Data availability statement\n\nNo data associated with this article.\n\n\nAcknowledgements\n\nWe thank Mr. Anura J. Wanniarachchilage at the Anatomy Department for taking care of the cadavers for anatomic dissections. The study received support from Body Scientific International, LLC, in obtaining the cadavers. During the preparation of this work, the author(s) used CHAT GPT to rephrase some sentences to avoid repetition of words. After using this tool, the author(s) reviewed and edited the content as needed and take(s) full responsibility for the content of the publication.\n\n\nReferences\n\nGray H, Williams PL, Bannister LH, et al., editors. Gray’s anatomy: the anatomical basis of medicine and surgery. Edinburgh: Churchill Livingstone; 38. ed.1999; p. 2092. repr. 1999.\n\nRajani SJ, Ravat MK, Rajani JK, et al.: Cadaveric Study of Profunda Femoris Artery with Some Unique Variations. J. Clin. Diagn. Res. 2015; 9: AC01-03. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nManjappa T, Prasanna LC: Anatomical variations of the profunda femoris artery and its branches cadaveric study in South Indian population. Indian J. Surg. 2014; 76: 288–292. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChoy KW, Kogilavani S, Norshalizah M, et al.: Topographical anatomy of the profunda femoris artery and the femoral nerve: normal and abnormal relationships. Clin. Ter. 2013; 164: 17–19. PubMed Abstract \| Publisher Full Text\n\nHeberer G, van Dongen RJAM : Vascular Surgery. Berlin Heidelberg: Springer; 2011.\n\nBergman RA, Tubbs RS, Shoja MM, et al.: Bergman’s comprehensive encyclopedia of human anatomic variation. Hoboken, New Jersey: John Wiley & Sons, Inc; 2016; 1432.\n\nTsoucalas G, Panagouli E, Fiska A, et al.: A rare double profunda femoris artery in a female cadaver. Anat. Cell Biol. 2018; 51: 212–214. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSahin B, Bilgiç S: Two rare arterial variations of the deep femoral artery in the newborn. Surg. Radiol. Anat. 1998; 20: 233–235. PubMed Abstract \| Publisher Full Text\n\nScott Levin L, Baumeister S: CHAPTER 7 - Lower extremity.Wei F-C, Mardini S, editors. Flaps and Reconstructive Surgery. Edinburgh: W.B. Saunders; 2009; pp. 63–70. Publisher Full Text\n\nGoel S: Unusual disposition of lateral circumflex femoral artery: Anatomical description and clinical implications. WJCC. 2015; 3: 85–88. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCrim JR, Manaster BJ: Imaging anatomy - musculoskeletal. 2nd ed.London: Elsevier Saunders; 2015.\n\nGautier E, Ganz K, Krügel N, et al.: Anatomy of the medial femoral circumflex artery and its surgical implications. J. Bone Joint Surg. Br. 2000; 82: 679–683. PubMed Abstract \| Publisher Full Text\n\nOide T: Selective medical circumflex femoral arteriography in idiopathic ischemic necrosis of the femoral head in adults. Nihon Seikeigeka Gakkai Zasshi. 1979; 53: 293–305. PubMed Abstract\n\nClarke SM, Colborn GL: The medial femoral circumflex artery: its clinical anatomy and nomenclature. Clin. Anat. 1993; 6(2): 94–105. Publisher Full Text\n\nAl-Talalwah W: The medial circumflex femoral artery origin variability and its radiological and surgical intervention significance. Springerplus. 2015; 4: 149. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nClaassen H, Schmitt O, Schulze M, et al.: Deep femoral artery: A new point of view based on cadaveric study. Ann. Anat. 2021; 237: 151730. PubMed Abstract \| Publisher Full Text" }	[ { "id": "350366", "date": "06 Jan 2025", "name": "Cagatay Barut", "expertise": [ "Reviewer Expertise Clinical anatomy", "anatomical variaitons", "radiologic anatomy" ], "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 presented a bilateral double profunda femoris artery, an important variation. I highly recommend that the authors add a detailed embryology of these vessels to better understand such variations. Embryology should not only cover the embryology in the textbooks but the authors should focus on the literature and present a detailed embryology of the femoral artery and the formation of the profunda femoris artery. Thus the background regarding this variation will be explained using embryology. Several examples of literature are listed below:\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? Partly\n\nIs the case presented with sufficient detail to be useful for other practitioners? Yes", "responses": [ { "c_id": "13093", "date": "08 Jan 2025", "name": "Sahar Mohsin", "role": "Author Response", "response": "Thank you for your thoughtful and constructive comments on our manuscript titled \"Case Report: Rare Bilateral Double Profunda Femoris Artery Variation: Clinical Implications and Considerations.\" We sincerely appreciate the time you took to review our work and the valuable insights you provided. We understand the importance of embryological context in understanding anatomical variations, and we agree that explaining the embryology behind the formation of the femoral artery and profunda femoris artery would enrich the manuscript. The main objective of the case report is to highlight the rare anatomical variation and its clinical implications, especially for surgical and diagnostic procedures, we prioritized its practical and clinical relevance over embryological discussion due to the concise format of a case report. In response to the comment, we propose adding a concise section on the embryological development of these arteries, referencing pertinent literature to bridge the anatomical findings with developmental origins. This would allow us to bridge the gap between the anatomical findings and their developmental origins without overshadowing the main clinical emphasis of the case report. We hope this approach addresses your concern while preserving the integrity and purpose of our case report. We would be grateful for your reconsideration of the manuscript based on these adjustments. Once again, thank you for your valuable feedback. We look forward to your thoughts on our proposed revisions." } ] } ]	1	https://f1000research.com/articles/13-1528
https://f1000research.com/articles/13-1527/v1	17 Dec 24	{ "type": "Research Article", "title": "Revealing culturable fungal microbiome communities from the Arabian Peninsula desert representing a unique source of biochemicals for drug discovery and biotechnology", "authors": [ "Walaa Mousa", "Najwa Alramadan", "Rose Ghemrawi", "Tareq Abu Izneid", "Najwa Alramadan", "Rose Ghemrawi", "Tareq Abu Izneid" ], "abstract": "Background Microbes living at extremes evolve unique survival strategies to adapt to challenging environmental conditions. Among these strategies is their distinctive metabolic potential and ability to produce specialized metabolites enabling them to compete for limited resources and defend against predators. These metabolites have significant potential in pharmaceutical and industrial applications, particularly in the development of drugs and biochemicals.\n\nObjectives This study aimed to investigate the culturable fungal communities associated with four desert plants and their surrounding soils in the Arabian Peninsula desert to identify their bioactive properties.\n\nMethods A total of 12 distinct fungal species were isolated from the plants and soils. Each plant hosted a unique set of fungi, demonstrating the diversity of desert-adapted fungal communities. Biological activities of the fungal extracts were evaluated through various assays, including antimicrobial, antifungal, anticancer, and antioxidant properties.\n\nResults\nPanicum turgidum harbors the most diverse fungal community, dominated by genera such as Mucor, Aspergillus, Colletotrichum, Alternaria, and Chaetomium. Aspergillus species comprise 33% of the total isolates, followed by Fusarium at 16%. All extracts exhibit diverse activities, with Aspergillus species demonstrating the highest antioxidant activities and total phenolic and flavonoid content. Fungi from P. turgidum, particularly Mucor sp., Aspergillus sp., and Curvularia sp., display potent activity against Staphylococcus aureus, while Mucor sp., Chaetomium sp., and Curvularia sp. exhibit moderate inhibition against Pseudomonas aeruginosa.\n\nConclusion This study highlights the importance of exploring extremophilic microorganisms, such as those found in desert ecosystems, as they offer a wealth of compounds that could address current challenges in drug discovery and biotechnology.", "keywords": [ "Desert microbiome", "culturable fungi", "bioactive molecules", "antioxidant", "antimicrobial", "and cytotoxic." ], "content": "Introduction\n\nMicrobes thrive in extreme environments, such as deserts, arctics, and deep-sea ecosystems by evolving unique survival mechanisms adapting to harsh conditions. These adaptations often include the production of specialized metabolites that play essential roles in competition, defense, and stress tolerance. For example, Penicillium chrysogenum, initially isolated from desert soil, produces a variety of antimicrobial compounds, including the β-lactam antibiotic penicillin1 Similarly, Aspergillus terreus produces lovastatin, a statin used to lower cholesterol,2 while Streptomyces hygroscopicus, which has been isolated from alkaline soils, secretes rapamycin, a potent immunosuppressant.3 Beyond their pharmaceutical potential, fungi are also exploited at an industrial level for the production of various compounds. For example, Aspergillus niger produces over 70% of global citric acid demand,4 while Trichoderma reesei produces cellulases crucial for biofuel production and textile processing.5 Fusarium venenatum is used to produce mycoprotein, a sustainable meat substitute,6 and Aspergillus oryzae is essential in fermenting traditional Asian foods such as soy sauce and miso.7 These examples showcase the potential of fungal metabolites across various pharmaceuticals and industrial sectors. Among challenging habitats is the arid desert, with extreme temperatures, UV radiation, and limited nutrients. These conditions drive the evolution of novel metabolic pathways in these extremophilic organisms, representing a largely untapped source of bioactive compounds with potential in drug discovery and biotechnology. In this study, we aimed to characterize the culturable fungal communities of four desert plants native to the Arabian Peninsula deserts. These plants included Panicum turgidum, Halocnemum strobilaceum, Haloxylon persicum, and Arnebia hispidissima. These plants have not been explored extensively for their fungal microbiomes, despite the essential role these microbes play in enhancing plant resilience and overall health.\n\nP. turgidum is a desert xerophyte that thrives in arid regions, including the Arabian Peninsula and parts of North Africa and Asia. This plant is characterized by its extensive drought resistance, with a significant role as a nurse plant in desert ecosystems.8 Its rhizosphere is profoundly colonized by Arbuscular Mycorrhizal Fungi (AMF), establishing a crucial symbiotic relationship that enhances nutrient uptake, water retention, and protection against pathogens.9 Recent studies have shown that AMF inoculation can improve drought tolerance in Panicum turgidum, mitigating oxidative stress and promoting chlorophyll production10\n\nH. strobilaceum is well-adapted to saline and hypersaline habitats, such as salt marshes and alkali flats, making it native to regions like the Red Sea and Mediterranean.11,12 Compounds derived from H. strobilaceum have demonstrated strong antimicrobial, antioxidant, and antibiofilm activities. For instance, recent research identified two promising compounds, one an alkaloid effective against various pathogens, and the other specifically inhibiting biofilm formation by Pseudomonas aeruginosa.13 Furthermore, the ethyl acetate extract of this plant has shown anticancer activity against common cancer cell lines in Egypt, including prostate (PC-3), lung (A-549), and breast (MCF-7) cancer.14 H. persicum, another well-known desert xerophyte, survives in arid regions due to its high resilience to drought stress, supported by its production of intrinsic compounds and metabolites.15 Found across western Asia, this plant significantly contributes to soil health and seed bank diversity within desert ecosystems.16 A study highlighted that the rhizospheres of H. persicum are rich in archaea and fungi, which play a critical role in nutrient cycling and promoting plant growth under harsh conditions.17 A. hispidissima inhabits arid and semi-arid regions in India and northern Africa and is widely distributed in the UAE. This plant is recognized for its medicinal properties, particularly its extract containing shikonin, which exhibits significant anti-cancer properties through mechanisms targeting cancer cell death.18 Additionally, it has been traditionally used in Indian medicine for treating various infections due to its potent antimicrobial properties.19 The successful biosynthesis of silver nanoparticles using root extracts of A. hispidissima has been reported, showcasing their potent antioxidant and antimicrobial activities against several pathogens.20\n\nThis study aims to explore the fungal communities associated with these resilient desert plants, identifying fungal species and their biological significance for potential applications in pharmaceuticals and biotechnology.\n\n\nMethods\n\nIn a prior study,21 we explored culturable bacterial communities from both the rhizosphere (R) and endosphere (E) of four native desert plants of the Arabian Peninsula: Halocnemum strobilaceum (HS), Panicum turgidum (PT), Haloxylon persicum (HP), and Arnebia hispidissima (AH). This study extends those findings by isolating and evaluating fungal communities associated with these plants. The methodologies for plant collection and sample preservation have been detailed previously.21,22 In brief, five replicates from each plant were gathered from different sites near the UAE, which varied in soil properties. Collected samples, comprising both roots and surrounding rhizosphere soils, were obtained in October 2022, during which daytime temperatures ranged between 34 and 40 °C with minimal rainfall.\n\nA modified root-washing technique, based on the method published by Banno et al,23 was employed to isolate epiphytic fungi. Briefly, the roots were immersed in sterile water and shaken at 250 rpm for 30 minutes, a step that was repeated three times. Root washes were pooled, serially diluted (up to 105), and 200 μL of each dilution was spread on three types of media: potato dextrose agar (PDA, HiMedia, India # MH096), Sabouraud dextrose agar (SDA, HiMedia, India # MV063), and yeast maltose agar (YMA, HiMedia, India # M1967). All media have been prepared according to the manufacturer protocol. To prevent bacterial growth, all media were supplemented with chloramphenicol (200 μg/L). Plates were incubated at 25 °C for 5 days. Fungal colonies, selected based on their morphology, were subcultured multiple times (3–5 repetitions) until pure strains were obtained. For long-term storage, fungal spores and mycelia were suspended in 25% glycerol and stored at -20 °C.\n\nThe protocol for isolating endophytic fungi involved surface-sterilizing the root tissues, followed by similar culturing techniques as for epiphytes. Root surface sterilization was conducted by first sonication in autoclaved water for 5 minutes to remove soil particles, followed by a series of ethanol (95% for 3 minutes) and sodium hypochlorite (3% NaOCl for 5 minutes) treatments. Between these steps, the roots were thoroughly rinsed with sterile water. Sterilization success was confirmed by rolling sterilized roots on PDA plates and incubating them at 25 °C and 37 °C; no microbial growth was observed. The sterilized roots were then sectioned and ground in a sterile mortar, after which the tissue was plated on PDA, SDA, and YMA media. Subculturing and maintenance followed the same protocol as for the epiphytic fungi.\n\nDNA extraction from fungal isolates was performed using a commercial DNA extraction kit (Norgen Bioteck, Canada). Briefly, the fungi were cultured in PD broth for 3 days, after which the mycelia were collected and ground in a sterile mortar. DNA extraction proceeded according to the manufacturer’s instructions, and the quality and quantity of DNA were assessed using gel electrophoresis and a nanodrop spectrophotometer, respectively. Taxonomic identification was carried out using ITS primers: ITS1 (5′-TCCGTAGGTGAACCTGCGG-3′) and ITS4 (5′-TCCTCCGCTTATTGATATGC-3′). PCR amplification involved an initial denaturation at 95°C for 5 minutes, followed by 35 cycles of 1-minute denaturation at 94°C, 30-second annealing at 55°C, and a 2-minute extension at 72°C, concluding with a final 10-minute extension at 72°C. PCR products were purified with the QIAquick gel purification kit (Qiagen, Germany) and sequenced. Sequences were aligned using BLAST against the NCBI database and deposited in.\n\nAll fungal isolates were cultured in 3 L Fernbach flasks containing PDA medium and incubated at 28 °C for 10 days. After this period, the culture was extracted by shaking with ethyl acetate (3 times the volume), and this extraction was repeated three times. The extract was then filtered and concentrated to dryness using a rotary evaporator (BUCHI R100, Schweiz). The resulting residue was dissolved in DMSO at a stock concentration of 100 μg/μL, which was subsequently used to evaluate various biological activities.\n\nTo determine the antioxidant activity of the fungal extracts, we evaluated their free radical scavenging capacity colorimetrically using DPPH as a source of free radicals. A freshly prepared DPPH solution (50 μg/mL) was combined with a serial dilution of each fungal extract (ranging from 100 to 1 μg/mL), shaken, and allowed to incubate in the dark for 30 minutes at 20 °C. The absorbance was recorded at 517 nm using a spectrophotometer (OmegaStar, Germany). The percentage of radical scavenging activity was calculated using the formula:\n\nThe total phenolic content was determined using the Folin-Ciocalteu method.23 In brief, 1 μL of the extract (100 μg/μL) was combined with 100 μL of the Folin-Ciocalteu reagent (10% v/v) and incubated for 15 minutes. Following this, 2 μL of sodium carbonate (7%, w/v) was added to the mixture to neutralize it. The samples were then stored in the dark for 2 hours. Absorbance was recorded at 765 nm using a spectrophotometer. A calibration curve was established with gallic acid concentrations ranging from 5 to 200 μg/mL, and total phenolic content was expressed as micrograms of gallic acid equivalents (μg GAE) per milligram of EA extract.\n\nThe total flavonoid content was evaluated using the aluminum chloride colorimetric method.24 Briefly, 5 μL of the extract (100 μg/μL) was combined with 100 μL of aluminum chloride (3% w/v) and 100 μL of potassium acetate (1 M). The samples were incubated at 25°C for 30 minutes. The absorbance was then measured at 420 nm, with the solvent used for dissolving the extract serving as the blank control. Quercetin (5–200 μg/mL) was utilized to generate the calibration curve. The results for total flavonoid content were reported as micrograms of quercetin equivalents (μg QE) per milligram of EA extract.\n\nTo investigate the antimicrobial properties of the fungal extracts, we employed an agar diffusion assay against human and plant pathogens. The bacterial pathogens included Gram positive indicator Staphylococcus aureus (ATCC 25923) and Pseudomonas aeruginosa (BAA-1744). The fungal pathogens included in this study are C. albicans (ATCC 18804) and Fusarium gramineraum (MYA-4620). Each pathogen was cultured under optimal conditions for growth. We first conducted agar diffusion method and positive extracts were assessed for their minimum inhibitory concentration (MIC) using broth dilution method. For agar diffusion test, a 100 μL of an overnight culture of each pathogen was evenly spread onto agar plates (utilizing media appropriate for each pathogen’s growth). Sterile glass pipettes were then used to create wells in the agar, and 20 μL of 10 μg/μL of each extract was introduced into these wells and the plates were incubated aerobically at 37 °C for 24 hours. After incubation, the plates were examined for zones of inhibition. Positive cultures were processed to determine the MIC. To measure MIC, a single colony of each pathogen was initially cultured for 24 hours in its specific broth medium and subsequently diluted in the same medium at a 1:10,000 ratio, following McFarland Standards. Thereafter, 196 μL of the pathogen suspension was added to each well of a microplate, followed by the addition of 4 μL of serially diluted fungal extracts. Positive controls included amoxicillin (5 μM), and ciprofloxacin (2 μM), and amphotericin B (10 μM). The plates were incubated for 24 hours at the optimum growth condition for each pathogen, after which the optical density at 600 nm (OD600) was recorded using a microplate reader. All concentrations were tested in triplicate, and the experiment was independently replicated. The percentage of growth inhibition was calculated as previously described.25 To determine the minimum bactericidal or fungicidal concentration (MBC) of the fungal extracts, 10 μL of the inhibited samples from the MIC assay (where no growth was observed) were plated onto suitable agar plate and incubated at the best growth condition for each pathogen then inspected for colony formation. Control groups included samples treated with antibiotic or antifungal compound and untreated cultures.\n\nTwo human lung cancer cell lines, A549 and H292, were utilized to evaluate the cytotoxic properties of the fungal extracts. The cells were cultured in RPMI medium supplemented with 10% FBS, 1% penicillin-streptomycin solution, and incubated at 37 °C in a 5% CO2 environment. The in vitro cytotoxic activity of the extracts was assessed against the A549 and H292 cells by measuring the formation of insoluble formazan salt, which occurs via the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) by NAD(P)H-dependent cellular oxidoreductase enzymes, directly correlating to the number of viable cells remaining after extract treatment. Tumor cells (30 × 103 cells per well) were seeded into 36-well culture plates and incubated for 24 hours at 37 °C in a humidified CO2 incubator. After this incubation, the cells were treated with 10 μg/mL of each extract for 24 hours. Control groups included cells treated with 0.1% DMSO and untreated cells. Following treatment, 20 μL of MTT solution was added to each well, and the plates were incubated for an additional 4 hours at 37 °C in a 5% CO2 incubator. After incubation, the plates were centrifuged for 20 minutes at 3000 rpm, and the formazan crystals formed were dissolved using 100 μL of DMSO. Absorbance was then measured at 570 nm using a microplate reader.\n\nData analysis and graphical representation were carried out using one-way ANOVA in GraphPad Prism V9 (GraphPad Software, La Jolla, CA, USA). Similar function could be performed by Microsoft Excel. All experiments were performed in triplicate, and results are presented as means ± standard error of the mean (SEM). Data for statistics are shown in Tables 1-4, together with the raw data.101\n\n\nResults\n\nIn this study, we aimed to profile the culturable fungal communities associated with four plants native to the Arabian Peninsula desert. We isolated a total of 12 unique fungal species, identified taxonomically through sequence alignments, and assessed their derived extracts for various biological activities ( Figure 1). From Panicum turgidum, we isolated five fungal species identified as Mucor sp. (PT-F1), Aspergillus sp. (PT-F2), Colletotrichum sp. (PT-F3), Alternaria sp. (PT-F4), and Chaetomium sp. (PT-F5). Additionally, three isolates were obtained from Halocnemum strobilaceum, two of which were identified as Aspergillus species (HS-F1 and HS-F2), while the other was Fusarium sp. (HS-F3). From Haloxylon persicum, we identified two unique fungi: Plectosphaerella sp. (HP-F1) and Aspergillus sp. (HP-F2). Lastly, two fungi were isolated from Arnebia hispidissima, namely Curvularia sp. (AH-F1) and Fusarium sp. (AH-F2). Overall, Aspergillus species were the most prevalent, representing 33% of the total recovered fungi across all plant samples.\n\nThe figure illustrates the process of isolating fungi from plant material and subsequent subculturing to obtain pure isolates.\n\nTo assess the biological activities of the isolated fungi, we scaled up the fermentation of each species and prepared crude ethyl acetate extracts. These extracts were then subjected to six assays: 1) antioxidant activities, 2) total phenolic content, 3) total flavonoid content, 4) antibacterial assay, 5) antifungal assay, and 6) anticancer cytotoxicity screening. All assessed extracts demonstrated comparable antioxidant activities, with the highest levels observed in Aspergillus species PT-F2, HS-F1, and HP-F2 ( Figure 2). In contrast, the lowest antioxidant activities were recorded for PT-F1 (Mucor sp.), AH-F1 (Curvularia sp.), and AH-F2 (Fusarium sp.). Regarding total phenolic content, Curvularia sp. exhibited the richest extract, followed by Aspergillus species isolated from Panicum turgidum. The lowest phenolic content was found in HS-F2 (Aspergillus sp.), HS-F3 (Fusarium sp.), and HP-F1 (Plectosphaerella sp.). The highest flavonoid content was reported for HS-F1 (Aspergillus sp.) and AH-F1 (Curvularia sp.), followed by PT-F2 (Aspergillus sp.) and PT-F5 (Chaetomium sp.).\n\nThe graph demonstrates three activities including antioxidant activities measured as % of DPPH reduction, total phenolic content measured as μg of EAG per mg of the extract and total flavonoid contents measured as μg of EQ per mg of the extract. Data are the average of three replicates.\n\nTo assess the antimicrobial potential of each extract against bacterial and fungal indicators, we conducted two experiments. The first was an agar diffusion test to identify any possible activities of the crude extracts. Extracts that demonstrated positive results based on the diameter of the inhibition zone were subsequently subjected to a broth dilution test to determine their minimum inhibitory concentration (MIC). Data presented in Figure 3A revealed significant antibacterial activity, particularly against the Gram-positive indicator strain Staphylococcus aureus, with a total of nine isolates exhibiting positive inhibitory effects. The most potent extracts, characterized by the lowest MIC values, were derived from Panicum turgidum (PT-F1, PT-F2) and Arnebia hispidissima (AH-F1), followed by Halocnemum strobilaceum (HS-F2, HS-F3) and PT-F5, HP-F2. In contrast, activity against Pseudomonas aeruginosa was limited, as indicated by the high MIC values for all extracts. The lowest MIC values were recorded for PT-F1 (Mucor sp.), PT-F5 (Chaetomium sp.), and AH-F1 (Curvularia sp.). In terms of antifungal activities, eight fungal extracts showed positive effects against the tested fungal species ( Figure 3B). The most active extracts in inhibiting both Fusarium graminearum and Candida albicans were PT-F4, followed by PT-F1, HS-F3, HS-F1, and HP-F1. The highest MIC values were observed for PT-F5 (Chaetomium sp.) and HP-F2 (Aspergillus sp.). To assess the preliminary cytotoxic activities of the extracts, we conducted the MTT assay. All extracts demonstrated cytotoxic activity, with the lowest observed for PT-F2 (Aspergillus sp.), as shown in Figure 4.\n\nThe graph presents results from two experiments: A) antibacterial activities against Gram-positive indicator S. aureus and Gram-negative indicator P. aeruginosa, and B) antifungal activities against the human-associated yeast C. albicans and the plant pathogenic fungus F. graminearum. Data represent the average of three replicates, with bars indicating the standard deviation of the mean.\n\n\nDiscussion\n\nIn this study, we identified 12 fungal species for the first time in the examined desert plants and explored their biochemical characteristics. These fungal isolates belong to eight genera, with Aspergillus and Fusarium being the most prevalent. Species from the genus Aspergillus have been previously isolated from desert plants, such as Opuntia versicolor in the Sonoran Desert.26 A study conducted in Saudi Arabia, investigating the cultivable fungal communities in the Arabian Peninsula’s desert soils, also identified Aspergillus as the most widespread fungal genus. Similarly, another study identified Aspergillus species, specifically A. niger and A. flavus, among the fungal communities in Saudi Arabia’s Sabkha desert marshes.27,28 Aspergillus fumigatus is known to inhabit many desert plants in arid and semi-arid environments across regions like India, Pakistan, the Mexican desert, and Iran.29 Additionally, Fusarium species are dominant as endophytic fungi in various desert plants, including Cyathea gigantea, Calotropis procera, Withania somnifera, and Aloe vera.30–33 Alternaria species has been isolated from desert plants in both Jordanian and Saudi Arabian deserts34 and from soil in Al-Kharj, Saudi Arabia.35\n\nFungi play a crucial role in desert ecosystems by supporting plant health, enhancing nutrient cycling, and providing resilience against environmental stressors. The unique metabolic capabilities of the fungal species identified in this study highlight their importance in these arid environments. Previous research supports the unique metabolic capabilities of these fungal species and highlights their crucial role in promoting plant health. The genus Aspergillus is regarded as a prevalent endophytic fungus36 with xerophytic characteristics, allowing it to thrive under the arid conditions and water scarcity typical of desert environments.37 Aspergillus niger has been shown to provide a biological shield for host plants by protecting them from pests and pathogens and enhancing their resilience to biotic and abiotic stresses.38 Another fungus identified in this study, Mucor mucedo (commonly known as pin mold), is a saprophyte with a broad ecological tolerance and a worldwide distribution. It colonizes decaying organic matter and is capable of rapid growth in environments with limited nutrients.39 Mucor mucedo can also survive extreme environmental conditions, such as freezing temperatures, UV radiation, and desiccation.40 Its role in decomposing organic matter yields essential nutrients that support plant development.41 In this study, we also identified Colletotrichum spaethianum, a known endophytic fungus. Species within the Colletotrichum genus are primarily found in tropical and temperate environments.42 Some Colletotrichum species exhibit beneficial activities, such as C. magna, which helps plants combat infections caused by F. oxysporum and C. orbiculare.43 Moreover, C. gloeosporioides produces colletotric acid, a potent antifungal compound.44 Chaetomium globosum, known as a saprophyte and occasionally an endophyte, has been demonstrated to protect plants against the toxic effects of heavy metals such as copper.45 Aspergillus terreus, another species identified in this study, produces metabolites such as phenols, flavonoids, and indole-acetic acid, which stimulate plant growth. Research on tomato plants shows that A. terreus enhances shoot and root length, as well as overall chlorophyll content.46 Another study reports that filtrates of A. terreus, free from spores and mycelia, significantly reduce spore formation of the plant pathogen Pythium aphanidermatum, thereby improving plant growth and protection.47 Plectosphaerella cucumerina is another significant fungus identified. Its cell wall contains molecules classified as microbe-associated molecular patterns (MAMPs), which can bind to pattern recognition receptors (PRRs) in Arabidopsis thaliana, triggering the plant’s defense mechanisms.48 Additionally, P. cucumerina promotes host plant growth by inducing the expression of genes involved in carbohydrate and amino acid synthesis, which enhances the host plant’s growth and can impart similar benefits when transplanted into other plants.49,50\n\nOur investigation into the crude extracts from various cultured fungal species revealed a rich spectrum of biological activities, including antimicrobial, antioxidant, and anticancer properties. This is supported by previous studies. For instance, A. niger, isolated from desert soils in Saudi Arabia, exhibited significant antioxidant activities.51 Furthermore, extracts of A. niger and A. flavus displayed potent antimicrobial and anticancer activities.52 Additionally, Aspergillus species from Phragmites australis leaves showed antibacterial effects against Klebsiella sp., E. coli, and S. aureus, along with antibiofilm activities.53 The extracts also demonstrated cytotoxicity on the breast cancer cell line MCF-7, with an IC50 of 8 μg/μl. Detailed studies have shown that A. niger extract can induce cell cycle arrest and apoptosis,54 revealing a composition of diverse hydrocarbons, phthalates, and phenolic derivatives.55 Various metabolites from A. terreus have exhibited antibacterial, anticancer, and antioxidant activities.56,57 Notably, Asperteramide A showed potent antibacterial activity against Klebsiella pneumoniae, MRSA, Acinetobacter baumannii, Enterococcus faecalis, and ESBL-producing E. coli.58 Furthermore, tetracyclic acid A, isolated from A. terreus, has emerged as a valuable anticancer agent by stimulating heat shock responses in tumor cells.59 Crude extracts from Aspergillus fumigatus, isolated from mangrove plants in the Sundarbans, demonstrated potent antibacterial activity against both Gram-positive and Gram-negative strains, including E. coli, Micrococcus luteus, Pseudomonas aeruginosa, and Staphylococcus aureus.60 Remarkably, an enzyme produced by thermotolerant A. fumigatus, known as MGL, exhibited anticancer activity in Hep-G2 and HCT116 cell lines.61 Fusarium species isolated from Cinnamomum kanehirae, Selaginella pallescens, and Tripterygium wilfordii displayed antimicrobial activities against methicillin-resistant S. aureus and Candida albicans.62–65 A recent study indicated that the ethyl acetate extract of Fusarium species possesses the highest antibacterial, antioxidant, and anticancer potential.66\n\nIn this study, we identified Colletotrichum species with antibacterial and antioxidant activities. Previous studies noted the presence of two Colletotrichum sp. in Andrographis paniculate, demonstrating their ability to produce antimicrobial and antioxidant compounds.67 Additionally, Colletotrichum acutatum, isolated from Angelica sinensis, produces molecules exhibiting antimalarial, antioxidant, antibacterial, anti-proliferative, and antibiofilm activity.68 Extracts from Colletotrichum sp. CG1-7, isolated from Arrabidaea chica, displayed antioxidant activity comparable to quercetin.69 Furthermore, two metabolites derived from Colletotrichum sp., phthalide and isocoumarins, have shown potential as antioxidant agents and in inhibiting cancer cell growth in the HepG2 cell line.70 Another compound, palmitoylethanolamide (PEA), isolated from C. gloeosporioides, demonstrated potential anticancer activity against human breast cancer cells via apoptosis induction.71 Interestingly, the production of valuable antimicrobial and antioxidant compounds from various Colletotrichum species is significantly enhanced by light spectrum treatment.72 Additionally, we reported on the activities of extracts from Alternaria species, including potent antifungal and anticancer activities. Previous data show that extracts of A. alternata exhibit antibacterial activity.73 Compounds isolated from A. alternata include alternariol, tenuazonic acid, levofuraltadone, and kigelinone, which have antibacterial and/or anticancer activities.74,75 Another fungus identified in our study is Chaetomium, exhibiting antioxidant and anticancer activities. Bioactive metabolites from Chaetomium globosum have been reported, displaying anticancer, antimicrobial, anti-inflammatory, antiviral, and antioxidant activities.76,77 Notably, chrysophanol alkaloid from Chaetomium shows anticancer activities against multiple cancer types, along with its antimicrobial properties.78–81 Another compound, Chetomin, produced by the Chaetomium genus, has been noted for its ability to block hypoxic-inducible transcription, thereby suppressing tumor growth.82 We also identified Plectosphaerella species with antifungal, anticancer and antioxidant activities. Previous reports indicate that P. cucumerina extracts exhibit antibiofilm and anti-virulence activity against Pseudomonas aeruginosa, likely due to the presence of emodin and patulin compounds.83 Moreover, studies have shown the potential of P. cucumerina in providing protection against nematodes.84,85\n\nLastly, we reported activities of Curvularia species, including antibacterial activities against both Gram positive and negative pathogens. Previous research indicates that extracts from C. lunata possess antimicrobial and antioxidant activities, with low cytotoxic effects against the ATCC-CCL-81 cell line.86 Furthermore, Curvularia species are known for producing metabolites with antibacterial, antioxidant, and anticancer activities. For instance, Curvularia sp. G6-32, isolated from Sapindus saponaria, generates epoxyquinone, noted for its antioxidant potential.87 Regarding anticancer properties, Curvularia australiensis FC2AP, isolated from Aegle marmelos leaves, produces flavonoids that exhibit anti-cervical cancer and anti-inflammatory effects.88 Additionally, Curvularia sp. from Terminalia laxiflora generates bioactive peptides, shown to suppress tumor growth and inhibit angiogenesis.89\n\nDesert fungi hold significant potential to mitigate climate change impacts due to their unique adaptations to extreme heat, drought, and salinity. Colletotrichum species, for instance, serve as eco-friendly protectors against abiotic drought stress, which can severely damage plants and crops. Colletotrichum alatae secretes a heteropolysaccharide rich in β-glucan, enhancing drought resilience and optimizing rice cultivation in severely drought-affected areas.90 Furthermore, Colletotrichum species have been identified as drought-tolerant fungi, significantly contributing to plant growth in arid environments.91 Chaetomium globosum is recognized for its salt tolerance, promoting plant resilience in saline conditions.92 Reports indicate that Chaetomium globosum can enhance the survival and growth of salt-sensitive crops under drought stress.93 Other fungi, such as Aspergillus fumigatus, thrive at extreme temperatures and demonstrate potential in protecting sensitive plants like wheat from drought.94 Research has also highlighted the ability of C. lunata inoculum to improve resistance to salt and drought in rice, thus enhancing overall plant growth.95\n\nDesert fungi exhibit various environmental applications. For example, A. niger produces novel xylose transporters that efficiently convert lignocellulosic biomass into eco-friendly biofuels.96 Mucor mucedo is increasingly recognized for its ability to degrade hydrocarbons. A study found that immobilizing M. mucedo on corncob particles enhanced its efficacy in remediating pyrene-contaminated agricultural soil.97 Further investigation into M. mucedo revealed that exopolymer substances (EPS) play a crucial role in degrading polycyclic aromatic hydrocarbons, suggesting its application in environmental cleanup efforts.98 Additionally, C. lunata has been shown to enhance bioremediation of hydrocarbon-contaminated soil in conjunction with the plant Luffa aegyptiaca, facilitating the degradation of accumulated hydrocarbons.99 The Plectosphaerella cucumerina AR1 strain has also demonstrated the ability to degrade nicosulfuron, an herbicide commonly applied to maize crops, which contributes to groundwater and surface stream contamination.100\n\n\nConclusion\n\nThe Arabian Peninsula desert is home to diverse microbial communities that offer significant applications in drug discovery, as well as industrial and environmental interventions related to bioremediation and climate change solutions. Given the unique adaptations of these microorganisms, a comprehensive profiling of the desert microbiome, encompassing both bacterial and fungal communities, should be a research priority. This effort will not only enhance our understanding of desert ecosystems but also unlock the potential of these microbes for sustainable applications.\n\n\nAuthors contribution\n\nWM designed the study, collected the plants, isolated fungi, performed DNA extraction, PCR experiments, performed large scale fermentation and extraction, conducted biological assays, performed statistical and data analysis, designed and developed figures, and wrote the manuscript. RG conducted cytotoxicity experiment. NA contributed to isolation and purification of individual isolates, antimicrobial assays, data analysis and discussion. TAI designed the study, experimental protocols and analyzed data. All authors reviewed, edited, and approved the final version of the manuscript.\n\n\nEthics and consent\n\nEthical approval and consent were not required.", "appendix": "Data availability statement\n\nFigshare: “Biological Activities of Fungal Isolates from Arabian Desert.” DOI: 10.6084/m9.figshare.27925029.101\n\nThis project contains the following underlying data:\n\n• antimicrobial assay results, antioxidant activity data, and anticancer activity data.\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nAll sequencing data generated form this study have been made publicly available and have been deposited into GenBank. Links to access data are as follows;\n\nGenBank: Mucor sp. PQ480174; https://www.ncbi.nlm.nih.gov/search/all/?term=PQ480174\n\nGenBank: Aspergillus sp. PQ480175; https://www.ncbi.nlm.nih.gov/search/all/?term=PQ480175\n\nGenBank: Colletotricum sp. PQ480176; https://www.ncbi.nlm.nih.gov/search/all/?term=PQ480176\n\nGenBank: Alternaria sp. PQ480177; https://www.ncbi.nlm.nih.gov/search/all/?term=PQ480177\n\nGenBank: Chaetomium sp. PQ480178; https://www.ncbi.nlm.nih.gov/search/all/?term=PQ480178\n\nGenBank: Aspergillus sp. PQ480179; https://www.ncbi.nlm.nih.gov/search/all/?term=PQ480179\n\nGenBank: Aspergillus sp. PQ480180; https://www.ncbi.nlm.nih.gov/search/all/?term=PQ480180\n\nGenBank: Fusarium sp. PQ480181; https://www.ncbi.nlm.nih.gov/search/all/?term=PQ480181\n\nGenBank: Plecotosphaerella sp. PQ480182; https://www.ncbi.nlm.nih.gov/search/all/?term=PQ480182\n\nGenBank: Aspergillus sp. PQ480183; https://www.ncbi.nlm.nih.gov/search/all/?term=PQ480183\n\nGenBank: Curvularia sp. PQ480184; https://www.ncbi.nlm.nih.gov/search/all/?term=PQ480184\n\nGenBank: Fusarium sp. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGurgel RS, de Melo Pereira DÍ , Garcia AVF, et al.: Antimicrobial and Antioxidant Activities of Endophytic Fungi Associated with Arrabidaea chica (Bignoniaceae). J. Fungi. 2023 Aug 21; 9(8): 864. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTianpanich K, Prachya S, Wiyakrutta S, et al.: Radical Scavenging and Antioxidant Activities of Isocoumarins and a Phthalide from the Endophytic Fungus Colletotrichum sp. J. Nat. Prod. 2011 Jan 28; 74(1): 79–81. PubMed Abstract \| Publisher Full Text\n\nRai N, Gupta P, Verma A, et al.: Isolation and characterization of N-(2-Hydroxyethyl)hexadecanamide from Colletotrichum gloeosporioides with apoptosis-inducing potential in breast cancer cells. Biofactors. 2023 May 6; 49(3): 663–683. PubMed Abstract \| Publisher Full Text\n\nGan PT, Lim YY, Ting ASY: Inducing antioxidant and antimicrobial activities in endophytic and endolichenic fungi by the use of light spectra treatments. Arch. Microbiol. 2023 Sep 11; 205(9): 304. PubMed Abstract \| Publisher Full Text\n\nFanele A, Ndlovu SI: Endophytic fungal species Nigrospora oryzae and Alternaria alternata exhibit antimicrobial activity against gram-positive and gram-negative multi-drug resistant clinical bacterial isolates. BMC Complement. Med. Ther. 2023 Sep 15; 23(1): 323. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nElghaffar RYA, Amin BH, Hashem AH, et al.: Promising Endophytic Alternaria alternata from Leaves of Ziziphus spina-christi: Phytochemical Analyses, Antimicrobial and Antioxidant Activities. Appl. Biochem. Biotechnol. 2022 Sep 17; 194(9): 3984–4001. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAshoka GB, Shivanna MB: Metabolite profiling, in vitro and in silico assessment of antibacterial and anticancer activities of Alternaria alternata endophytic in Jatropha heynei. Arch. Microbiol. 2023 Feb 10; 205(2): 61. PubMed Abstract \| Publisher Full Text\n\nAkkol EK, Tatlı II, Karatoprak GŞ, et al.: Is Emodin with Anticancer Effects Completely Innocent? Two Sides of the Coin. Cancers (Basel). 2021 May 31; 13(11): 2733. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHorvat M, Avbelj M, Durán-Alonso MB, et al.: Antiviral Activities of Halogenated Emodin Derivatives against Human Coronavirus NL63. Molecules. 2021 Nov 11; 26(22): 6825. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZhang G, Zhang Y, Qin J, et al.: Antifungal Metabolites Produced by Chaetomium globosum No.04, an Endophytic Fungus Isolated from Ginkgo biloba. Indian. J. Microbiol. 2013 Jun 9; 53(2): 175–180. Publisher Full Text\n\nDwibedi V, Rath SK, Jain S, et al.: Key insights into secondary metabolites from various Chaetomium species. Appl. Microbiol. Biotechnol. 2023 Feb 17; 107(4): 1077–1093. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZhu X, Zhou D, Liang F, et al.: a new unusual chaetoglobosin from the mangrove endophytic fungus Penicillium chrysogenum V11 and its effective semi-synthesis. Fitoterapia. 2017 Nov; 123: 23–28. PubMed Abstract \| Publisher Full Text\n\nGao W, He Y, Li F, et al.: Antibacterial activity against drug-resistant microbial pathogens of cytochalasan alkaloids from the arthropod-associated fungus Chaetomium globosum TW1-1. Bioorg. Chem. 2019 Mar; 83: 98–104. PubMed Abstract \| Publisher Full Text\n\nTelarovic I, Wenger RH, Pruschy M: Interfering with Tumor Hypoxia for Radiotherapy Optimization. J. Exp. Clin. Cancer Res. 2021 Dec 21; 40(1): 197. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZhou J, Bi S, Chen H, et al.: Anti-Biofilm and Antivirulence Activities of Metabolites from Plectosphaerella cucumerina against Pseudomonas aeruginosa. Front. Microbiol. 2017 May 3; 8. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAtkins SD, Clark IM, Sosnowska D, et al.: Detection and Quantification of Plectosphaerella cucumerina, a Potential Biological Control Agent of Potato Cyst Nematodes, by Using Conventional PCR, Real-Time PCR, Selective Media, and Baiting. Appl. Environ. Microbiol. 2003 Aug; 69(8): 4788–4793. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHelen JN: Title Development of a fungal biological control agent for potato cyst nematodes in Jersey.\n\nKhalil AMA, Abdelaziz AM, Khaleil MM, et al.: Fungal endophytes from leaves of Avicennia marina growing in semi-arid environment as a promising source for bioactive compounds. Lett. Appl. Microbiol. 2021 Mar 18; 72(3): 263–274. PubMed Abstract \| Publisher Full Text\n\nPolli AD, Ribeiro MA d S, Garcia A, et al.: Secondary metabolites of Curvularia sp. G6-32, an endophyte of Sapindus saponaria, with antioxidant and anticholinesterasic properties. Nat. Prod. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLü ZW, Liu HY, Wang CL, et al.: Isolation of endophytic fungi from Cotoneaster multiflorus and screening of drought-tolerant fungi and evaluation of their growth-promoting effects. Front. Microbiol. 2023 Nov 2; 14. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYuan Y, Zu M, Zuo J, et al.: Chaetomium globosum D5 confers salinity tolerance on Paeonia lactiflora Pall. J. Plant Physiol. 2023 Jan; 280: 153878. PubMed Abstract \| Publisher Full Text\n\nManjunatha N, Li H, Sivasithamparam K, et al.: Fungal endophytes from salt-adapted plants confer salt tolerance and promote growth in wheat (Triticum aestivum L.) at early seedling stage. Microbiology (N Y). 2022 Sep 22; 168(8). Publisher Full Text\n\nGeorge NM, Hany-Ali G, Abdelhaliem E, et al.: Alleviating the drought stress and improving the plant resistance properties of Triticum aestivum via biopriming with aspergillus fumigatus. BMC Plant Biol. 2024 Feb 28; 24(1): 150.\n\nAdhikari A, Khan MA, Imran M, et al.: The Combined Inoculation of Curvularia lunata AR11 and Biochar Stimulates Synthetic Silicon and Potassium Phosphate Use Efficiency, and Mitigates Salt and Drought Stresses in Rice. Front. Plant Sci. 2022 Mar 3; 13. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSloothaak J, Tamayo-Ramos JA, Odoni DI, et al.: Identification and functional characterization of novel xylose transporters from the cell factories Aspergillus niger and Trichoderma reesei. Biotechnol. Biofuels. 2016 Dec 20; 9(1): 148. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHou W, Zhang L, Li X, et al.: Influence of Mucor mucedo immobilized to corncob in remediation of pyrene contaminated agricultural soil. Environ. Eng. Res. 2015 Jun 30; 20(2): 149–154. Publisher Full Text\n\nJia C, Li X, Allinson G, et al.: Composition and morphology characterization of exopolymeric substances produced by the PAH-degrading fungus of Mucor mucedo. Environ. Sci. Pollut. Res. 2016 May 19; 23(9): 8421–8430. PubMed Abstract \| Publisher Full Text\n\nAni E, Adekunle AA, Kadiri AB, et al.: Rhizoremediation of hydrocarbon contaminated soil using Luffa aegyptiaca (Mill) and associated fungi. Int. J. Phytoremediation. 2021 Dec 6; 23(14): 1444–1456. PubMed Abstract \| Publisher Full Text\n\nCarles L, Rossi F, Besse-Hoggan P, et al.: Nicosulfuron Degradation by an Ascomycete Fungus Isolated From Submerged Alnus Leaf Litter. Front. Microbiol. 2018 Dec 19; 9. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMousa W: Biological activities of fungal Isolates from Arabian Desert. Dataset. figshare. 2024. Publisher Full Text" }	[ { "id": "357007", "date": "04 Feb 2025", "name": "Sotirios Pilafidis", "expertise": [ "Reviewer Expertise Microbiology", "Bioconversion", "Biotechnology", "Sustainability", "Arid regions biodiversity." ], "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 presents novel work being done in previously understudies desert regions. The authors have identified and selected important key plant species of the region to isolate fungi from. The authors used solid standard practices to isolate and characterize the isolated species. Molecular identification by whole genome sequencing is highly recommended in the future. Overall the article is a significant contribution to the understudied knowledge base of arid regions microbiome diversity.\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/13-1527
https://f1000research.com/articles/12-1265/v1	04 Oct 23	{ "type": "Review", "title": "Exploring the anti-aging potential of natural products and plant extracts in budding yeast Saccharomyces cerevisiae: A review", "authors": [ "Phaniendra Alugoju", "Chella Perumal Palanisamy", "Naga Venkata Anusha Anthikapalli", "Selvaraj Jayaraman", "Anchalee Prasanskulab", "Siriporn Chuchawankul", "Madhu Dyavaiah", "Tewin Tencomnao", "Phaniendra Alugoju", "Chella Perumal Palanisamy", "Naga Venkata Anusha Anthikapalli", "Selvaraj Jayaraman", "Anchalee Prasanskulab", "Siriporn Chuchawankul" ], "abstract": "Aging is an inevitable multifactorial process associated with a decline in physiological functioning accompanied by a predisposition to a plethora of chronic ailments. Emerging anti-aging research studies using different model organisms have enabled scientists to uncover underlying molecular mechanisms of aging. Notably, the budding yeast Saccharomyces cerevisiae has been, and continues to be an indispensable model organism in the field of biomedical research for discovering the molecular causes of aging as well as the anti-aging potential of natural/synthetic compounds and plant extracts. Besides its ease of handling, genetic manipulation, and relatively inexpensive to grow, the budding yeast has preserved nutritional signaling pathways (such as the target of rapamycin (TOR)-Sch9 and the Ras-AC-PKA (cAMP-dependent protein kinase pathways) and two distinct aging paradigms such as chronological life span (CLS) and replicative life span (RLS). In the present review, we have explored the anti-aging properties of several natural products and phytoextracts and their underlying molecular mechanism of action on the CLS and RLS of yeast S. cerevisiae.", "keywords": [ "Saccharomyces cerevisiae", "Replicative lifespan (RLS)", "Chronological lifespan (CLS)", "nutrient signalling pathways", "target of rapamycin (TOR)", "Protein kinase A (PKA)", "Adenylate cyclase (AC)" ], "content": "Introduction\n\nAging is an inevitable natural phenomenon characterized by gradual decline in the bodily functions with an increased susceptibility to various internal and external environmental cues and subsequent development of a plethora of chronic diseases, ultimately ending up with death. Aging is considered a major risk factor for the development of several disease conditions including diabetes, cancer, cardiovascular diseases, and neurodegenerative diseases.1–3 Several hallmarks of aging have been proposed including mitochondrial dysfunction, cell senescence, genome instability, telomere abrasion, epigenetic alterations, malfunction of autophagy, aberrant nutrient-sensing signalling, stem cell dysfunction/exhaustion, and loss of intercellular communication.4,5 Previous studies have demonstrated the longevity promoting activity of several chemical compounds (known as geroprotectors) to ameliorate hallmarks of aging and to promote healthy lifespan of a variety of model organisms. However, only a very few compounds have been investigated for their potential geroprotective activity in the older people.6 Therefore, the main objective of the biogerontology research is to explore the molecular basis of aging and age-related diseases and to dis-cover new interventions to counteract the detrimental effects of aging and related pathological conditions.7,8\n\nChronological life span (CLS) indicates the number of days a yeast cell survives in the stationary phase. A gradual decrease in the level of carbon source (e.g., glucose) occurs during stationary phase. Subsequently, ethanol accumulates slowly in the external medium, which is then converted into acetic acid, resulting in the reduction in the pH of the external medium. Elevated acetic acid induces apoptosis of yeast cells. Besides, oxidized proteins and damaged mitochondria also accumulates. Collectively, these events lead to increased cell death with time during yeast Chronological aging. Replicative lifespan (RLS) indicates the number of daughter cells produced by a mother cell. Over time, the accumulation of nuclear extrachromosomal ribosomal DNA circles, oxidized proteins, and damaged mitochondria also occur in the replicatively ageing yeast cells. However, this damage is accumulates only in the mother cells but not in the daughter cells.\n\nGlucose activates adenylate cyclase via stimulation of RAS on one hand and via G protein-coupled receptors (GPCR) system of Gpr1 and Gpa2 on the other hand. The activated adenylated cyclase causes up-surge in the levels of cellular cAMP. Thus, GPR1 and GPA2 are essential for a glucose-dependent rise in the levels of cellular cAMP. Increased intracellular cAMP levels in turn lead to activation of protein kinas A (PKA). On the other hand, cytoplasmic amino acid levels are sensed by EGOC on the vacuolar membrane and this EGOC activates TORC1 which in turn activates Sch9 via phosphorylation. TORC1 can inhibit SNF1, which acts as an energy sensor. Both SNF1 and PKH1/2 can influence the activity of Sch9. Both the nutrient signaling pathways RAS-AC-PKA and TORC1-Sch9 converge at Rim15 protein kinase. The activated protein kinases PKA and Sch9 (of RAS-AC-PKA and TORC1-SCH9 signaling pathways) phosphorylate and inhibit Rim15 protein kinase activity, resulting in the inhibition of its transport to the nucleus. When both PKA and Sch9 are inactive, Rim15 (unphosphorylated form) gets activated and imported to the nucleus, where it induces the activation of transcription factors such as Msn2/4 and Gis1 leading the respective transcription of stress-responsive (STRE) genes and postdiauxic shift (PDS) genes. On the other hand, within the nucleus, Rim15 is also inactivated via phosphorylation by a Pho85 cy-clin-dependent kinase (in association with its cyclin partner Pho80). Phosphorylated Rim15 is exported to from the nucleus to the cytoplasm via the Msn5 receptor protein.\n\nGlobally, there is an increase in the aging population and concomitant increase in the chronic diseases affecting the quality of life of elderly.9 This leads to rising treatment costs and economic burden. Therefore, understanding of the aging phenomenon and its underlying molecular mechanisms are crucial to ameliorate age-related disease progression, to delay aging or promote active health lifespan of elderly people.10 Aging research using different model organisms from simple eukaryotic yeast to mammalian models led to the discovery of several genes and molecular pathways involved in aging of humans.11 Owing to their short lifespan, ease of genetic manipulation, and relatively inexpensive to maintain in the laboratory, several model organisms such as Caenorhabditis elegans (nematode), Drosophila melanogaster (fruit fly), Mus musculus (mouse), Rattus norvegicus (rat), and the budding yeast Saccharomyces cerevisiae have been used not only for the understanding of aging and age-related diseases, but also for the discovery of several anti-aging compounds.11 Nonetheless, these model organisms have served as potential tool for the discovery and evaluation of a wide spectrum of the pharmacological properties including anti-aging potential of several phytochemicals.12,13 In this review, we have summarized the anti-aging effects of natural products and plants on the budding yeast Saccharomyces cerevisiae.\n\n\nThe budding yeast Saccharomyces cerevisiae – A simple eukaryotic model to study aging\n\nThe budding yeast S. cerevisiae, also known as baker’s or brewer’s yeast, is a single-celled eukaryotic organism composed of several membrane-bound organelles similar to animal cells including a nucleus, endoplasmic reticulum, Golgi complex, vacuole, cytoskeleton, mitochondria, and other different organelles.14 Yeast cells are round to ovoid in shape with a size of ∼5 μm in diameter (unbudded cell), between bacteria and human cells in size. Yeast cells divide once every 90 min under optimal laboratory conditions, through a process of budding in which smaller daughter cells detach from their mother cell. The budding yeast was the first eukaryotic organism whose genome was completely sequenced and released in 1996. The haploid yeast cell contains 16 chromosomes comprising about ∼12,068 kb of genomic DNA. The genome of yeast is thought to be evolved from the whole-genome duplication of its ancestral set of 8 distinct chromosomes.15 The S. cerevisiae yeast genome is composed of many genes that can be grouped into protein-coding genes (5885) and non-coding genes. The budding yeast nuclear genome is composed of more than 6,600 open reading frames (ORFs). The yeast genome is also comprised of 786 dubious ORFs which probably do not encode any proteins. Interestingly, the yeast genome has a very low number of introns, approximately 4% of all genes, as a result of which, the yeast genome is composed of a high number of protein-coding genes (one gene every 2 kbp). The non-coding genes in yeast are transcribed into transfer RNA (tRNA), ribosomal RNA (rRNA), small nuclear RNA (snRNA), and small nucleolar RNA (SnoRNA).15 The best advantages of yeast as a model are that it is easy to handle, its short generation time, and ease of genetic manipulation. Most interestingly, yeast share several homologous and orthologous genes with mammals including humans, as a result of which yeast can be used even to study human diseases. The budding yeast has long been used as a model organism for the identification of the molecular basis of several cellular processes including cell cycle, autophagy, protein folding, oxidative stress, and aging.\n\n\nReplicative lifespan (RLS) and chronological lifespan (CLS)\n\nYeast exhibit two distinct patterns of aging such as chronological lifespan (CLS) and replicative lifespan (RLS). The RLS measures the mitotic potential of a single yeast cell (i.e., how many bud cells are generated from a single mother cell). Thus, yeast RLS is similar to the mitotic cell division of mammalian cells. For the first time in 1959, Robert Mortimer and John Johnston discovered the aging phenomenon in budding yeast. They reported that yeast cells can divide asymmetrically through mitosis (budding) for a limited number of divisions (~25) and then stop dividing.16 As the mother cell divides by mitosis, it accumulates molecular damage; however, the daughter cells retain replicative capacity but generally do not inherit such damage from the mother cell. However, some studies on yeast replicative aging have suggested the asymmetric inheritance of at least three different types of damage, including nuclear extrachromosomal ribosomal DNA (rDNA) circles, oxidatively damaged or misfolded cytoplasmic proteins, and dysfunctional mitochondria.17 Phenotypically, old yeast mother cells are larger in size than the daughters and carry a number of bud scars indicating yeast’s RLS. In old mother cells, the actin cytoskeleton is found to be distorted and therefore, mother cells need longer times to complete the cell cycle compared to daughter cells which show a normal dotted and chain-type cytoskeleton. In addition, mother cells also accumulate high levels of mitochondrial-derived reactive oxygen species (ROS) and apoptotic features such as externalization of phosphatidyl serine, nuclear DNA fragmentation and chromatin marginalization.\n\nOn the other hand, the CLS measures the yeast cell viability in the non-dividing phase (i.e., postmitotic phase). Thus, CLS is similar to the postmitotic aging of mammalian cells. The CLS is defined as the time duration a yeast cell can survive in a nondividing state, with survival determined by the ability to reenter the cell cycle and resume vegetative growth upon exposure to appropriate growth-promoting cues.17,18 Chronological aging is also characterized by the accumulation of protein carbonyl content and dysfunctional mitochondria. In worms, flies and mice, the key regulators of CLS also control RLS and aging. In addition, chronologically aged cells also show a reduction in subsequent RLS, suggesting that similar forms of age-associated damage may contribute to both mitotic (RLS) and postmitotic (CLS) aging in yeast cells.19 During CLS, the size of yeast cells is normal and they enter a stationary phase due to starvation. The stationary phase cells are characterized by altered cellular metabolism towards the synthesis of reserved carbohydrates such as glycogen and trehalose. During CLS, the starving yeast cells utilize glycogen whereas trehalose is used for membrane stabilization and other non-metabolic functions. In addition, stationary phase cells show a hard and thick cell wall. Stationary phase yeast cells exhibit heterogeneity and can be categorized into quiescent (G0) and non-quiescent cells. Quiescent cells described as unbudded daughter cells form only during the final cell division in the diauxic phase of the growth curve, and have a high density compared to normal yeast cells. In addition, quiescent cells can reenter synchronously the mitotic cell cycle. In contrast, non-quiescent cells are less dense and composed of heterogenous, asynchronous, and replicatively older cells that lose their division capacity. Non-quiescent cells accumulate elevated ROS levels compared to quiescent cells and exhibit apoptotic as well as necrotic features. After a longer time, the quiescent cells also start to show landmarks of apoptosis, and finally of necrosis.20 Though the phenotypes of replicatively older yeast cells (in RLS) and stationary phase yeast cells (in CLS) are different, features such as augmented levels of ROS and intracellular oxidative stress, apoptosis and necrosis are found to be common.\n\n\nConserved nutrient sensing aging regulatory pathways in budding yeast\n\nThough budding yeast exhibits different aging paradigms, two major nutrient sensing pro-aging signaling pathways, such as the TOR-Sch9 and the RAS-AC-PKA pathways, promote aging and early death in both RLS and CLS, thus playing a similar role in both aging paradigms, whereas SIR2 does not.21 Thus, the inhibition or inactivation of the RAS-AC-PKA pathway results in the extension of both CLS and RLS in yeast.21 RAS proteins act as molecular switches depending on their GTP-bound or unbound state. The yeast RAS2 gene is homologous to human RAS proto-oncogene. In yeast, the aging regulator pathways such as protein kinase A (PKA) and mitogen-activated protein kinase (MAPK) pathways are dependent on the activation of RAS2. Activated PKA inhibits the activity of two important transcription factors, Msn2/4, involved in yeast stress resistance. It has been reported that the deletion of RAS2 gene (that encodes Ras2, a G-protein) results in increased stress resistance, thereby increasing the yeast lifespan. However, deletion of MSN2/MSN4 genes reverts the phenotype observed with RAS2 gene deletion. In yeast, transcription factors such as Msn2/4 and Gis1 bind respectively to the STRE (STress Responsive Element) and the PDS (post-diauxic shift) sequences, thereby activating stress resistance genes including SOD1 SOD2, CTT1, HSPs, and DDR2. However, activated protein Kinase A (PKA) inhibits Rim15, a PAS-kinase that integrates signals from different nutrient sensing signaling pathways, including TORC1, Sch9, PKA, and Pho80-Pho85, and subsequently affects different transcription factors, Msn2/4 and Gis1.22 The inhibition of Rim15 positively regulates the transcription factors such as Msn2/4 and Gis1. It is hypothesized that Rim15 also plays a role in the regulation of genes involved in the glucose repression and cell cycle arrest.22\n\nSch9 is a serine/threonine protein kinase homologous to mammalian Akt (a serine/threonine kinase) and S6K (S6 kinase). On the other hand, deletion of SCH9 can extend both RLS and CLS of yeast. In addition, the combined deletion of SCH9 along with RAS2 can further extend the yeast CLS compared to the deletion of SCH9 alone. Both CLS and RLS can be extended by inhibiting or deleting the serine/threonine-protein kinase TOR1, via inactivating the downstream Sch9 protein kinase. Tor and Sch9 lead to the inactivation of the serine/threonine kinase Rim15 and the stress resistance transcription factor Gis1, both of which are required for maximum chronological life span extension.21 Similar to Msn2 and Msn4, Gis1 induces the expression of mitochondrial superoxide dismutase enzyme (MnSOD), which is required for the effect of SCH9 deletion on the CLS. Aging yeast is characterized by elevated levels of ROS. In contrast, reduced levels of ROS have been observed in the long-lived mutants deficient in Ras-AC-PKA or TOR-Sch9 signaling.\n\nDuring CLS, glucose levels decrease gradually and accumulation of non-fermentable carbon sources such as ethanol and acetic acid takes place. Both ethanol and acetic acid promote the CLS. Loss of either TOR1or SCH9 slows down the aging process partly by lowering respiration and stimulating the exhaustion of ethanol and acetic acid and the elevating the levels of glycerol in the nutrient medium. Since S. cerevisiae cannot ferment either acetic acid or ethanol, their depletion by mutations in TOR or SCH9 can prolong the CLS by creating similar conditions usually caused by dietary restriction. It is important to note that the yeast cells deficient in TOR-Sch9 or Ras-AC-PKA signaling pathways exhibit prolonged CLS during incubation in water or in a medium which is either deficient or composed of low amounts of acetic acid and ethanol.22,23\n\n\nAnti-aging studies conducted using the budding yeast S. cerevisiae as a model\n\nAcetyl-L-carnitine (ALC) is an endogenous molecule synthesized in the body from L-carnitine. ALC plays a key role in the energy metabolism and it has been reported to exhibit a plethora of pharmacological properties, including neuroprotective activities. It has also been used as a dietary supplement due to its potent therapeutic effects. Researchers have shown the anti-apoptotic and anti-aging ability of ALC in yeast model. ALC was found to inhibit mitochondrial fission and subsequently improved mitochondrial functioning. Furthermore, it was found that the mitoprotective effects of ALC were mediated through the yeast metacaspase (Yca1) and thus to its anti-apoptotic activity. ALC was also shown to extend the CLS of yeast cells.24\n\nDisasa et al. isolated a secoiridoid glycoside amarogentin from a Chinese traditional medicinal plant, Gentiana rigescens Franch. Amarogentin was reported to enhance both the enzymatic activities and expression levels of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx), thereby enhancing the viability of yeast cells exposed to oxidative stress. Furthermore, treatment with amarogentin resulted in an increased RLS of yeast wild type; however, this compound was not shown to enhance the lifespan of yeast mutants sod1Δ, sod2Δ, uthΔ, and skn7Δ, suggesting that the anti-aging effects of amarogentin are mainly through the regulation of antioxidative stress as well as by the regulation of UTH1, SKN7, SOD1, and yeast SOD2 gene expression.25\n\nAstaxanthin is a carotenoid compound with superior antioxidant ability than many other natural antioxidant molecules. Sudarshan et al. conducted a study to test the antioxidant, anti-apoptotic and anti-aging properties of astaxanthin using yeast model. Astaxanthin treatment prevented oxidative stress-induced elevation in MDA and ROS levels and reduction in superoxide dismutase and glutathione levels, thereby increasing the percentage viability of antioxidant gene-deleted yeast mutants by 20-40%. In addition, astaxanthin also prevented the apoptosis of aged cells and subsequently increased the viability of yeast cells during the CLS. Astaxanthin’s antioxidant and anti-apoptotic effects are the possible reason for its anti-aging activity.26 It was also suggested that anti-apoptotic effects of astaxanthin are also due to its ability to prevent nuclear fragmentation and chromatin condensation in yeast cells.27 Sudarshan et al. also found that astaxanthin enhanced the oxidative stress resistance and increased the viability of yeast DNA damage repair gene-deleted mutant cells. Astaxanthin was shown to prevent the accumulation of endogenous DNA damage marker (8-hydroxy-2-deoxyguanosine) levels in yeast cells. The anti-aging effects of astaxanthin were also suggested to be due to its ability to prevent the accumulation of mutation during chronological aging of DNA damage repair gene-deleted mutant yeast cells.27\n\nArtesunate is a semi-synthetic derivative of the antimalarial drug artemisinin. Previous studies have shown that artesunate exerts anti-aging effects similar to caloric restriction, indicating the CR mimetic effects of artesunate. From the whole-transcriptome profile analysis studies, it was revealed that artesunate mimics CR-triggered nitric oxide to induce antioxidant defense systems, thereby preventing ROS accumulation and mitigated oxidative stress, subsequently extending yeast lifespan.28\n\nBetulinic acid (BA) is a pentacyclic triterpenoid present in some plant species. A recent study reported that betulinic acid extends the chronological life span of yeast cells by mitigating oxidative stress-induced apoptosis. The anti-aging effects of betulinic acid were found to be mediated by the activation of genes associated with heat shock stress response and autophagy.29\n\nAlmond (Prunus dulcis (Mill.) D.A. Webb) is a major nut crop worldwide. Multiple health benefits associated with the consumption of almonds are due to the phenolic-rich almond skin. Previous studies have found that pre-treatment with almond skin extract and chlorogenic acid significantly enhanced the lifespan of yeast. Both almond extract and chlorogenic acid improved the mitochondrial function during chronological aging of yeast cells by reducing the accumulation of free radicals (including ROS and RNS) and by preserving mitochondrial membrane potential. Furthermore, treatment with almond extract and chlorogenic acid increased oxidative stress response by inducing the expression levels of SIR2 and SOD1 genes and by decreasing the endogenous levels of lipid peroxides, protein carbonyls and 8-hydroxy-2-deoxyguanosine in yeast cells. Altogether, it can be suggested that the longevity-extending effects of almond extract are ascribed to its ability to ameliorate oxidative stress in yeast cells.30\n\nFlavonoids have been reported to exert a plethora of pharmacological activities including antioxidant, anti-inflammatory, anti-cancer, anti-neurodegenerative, anti-aging, among others. Researchers have investigated the anti-aging effects of citrus flavonoids including naringin, hesperedin, hesperitin, and neohesperidin on the chronological aging of yeast. Among the tested flavonoids, neohesperidin significantly prevented accumulation of ROS and extended the chronological life span of yeast in a concentration-dependent manner.31 In another study, treatment with hesperidin significantly increased the expression levels of SOD and sirtuin2 (SIR2) and prevented ROS accumulation in yeast. In yeast, UTH1 gene encodes Uth1p, which is activated by oxidative stress, senescence, and TOR-dependent autophagy; all these events subsequently lead to cell death in yeast. Pretreatment with hesperidin, but not its aglycon hesperetin, significantly inhibited UTH1 gene expression, thereby extending yeast lifespan.32\n\nCurcumin is a biologically active yellow-colored carotenoid compound with potent anti-oxidant and anti-aging properties. Previous studies investigated the replicative and chronological life span-extending effects of curcumin using yeast. It was demonstrated that curcumin significantly increased oxidative stress and enhanced both replicative and chronological life span of yeast mutants that lacked antioxidant genes (SOD1 and SOD2) and DNA damage repair gene RAD52. Overall, it can be suggested that curcumin exerts anti-aging hormetic effects in yeast.33\n\nCucurbitacins are a class of tetracyclic triterpenoids that are abundant in plants belonging to the family Cucurbitaceae. Cucurbitacins have been reported to exhibit strong anticancer activity, and are divided into 12 classes from A to T with over 200 derivatives.34 Cucurbitacin B is the most abundant and active member of the cucurbitacins.35 Researchers have discovered that cucurbitacin B can extend both replicative and chronological life spans in yeast. Treatment with cucurbitacin B increased the lifespan of yeast by promoting the expression of ATG32. Cucurbitacin B could not increase the lifespan of yeast mutants devoid of autophagy related genes (ATG2 and ATG32), pointing to the fact that the anti-aging effects of cucurbitacin B are mainly through the induction of autophagy in yeast. It was also demonstrated that cucurbitacin B can ameliorate oxidative stress levels mainly through the induction of superoxidase dismutase activity (via increase expression of SOD and SOD2) and prevention of accumulation of oxidative stress markers including ROS and malondialdehyde (MDA). Furthermore, researchers also showed that the anti-aging effects of cucurbitacin B are also mediated through the regulation of expression of other aging-related genes such as UTH1 as well as SKN7. Cucurbitacin B’s anti-aging effects are attributed to its ability to ameliorate oxidative stress and regulation of autophagy, and age-related genes in yeast.36 Additionally, cucurbitane glycoside isolated from the methanol extracts of Momordica charantia L. fruits has been reported to significantly extend the RLS of K6001 budding yeast mainly through suppressing the ROS levels and oxidative stress burden. Besides, treatment with cucurbitane glycoside resulted in a decrease in the expression levels of UTH1 and SKN7 and increase in the expression levels of SOD1 and SOD2. However, cucurbitane glycoside was not able to extend the RLS of the yeast mutants of uth1Δ, skn7Δ, sod1Δ, and sod2Δ, suggesting that cucurbitane glycoside exerts antiaging effects via antioxidative stress and regulation of yeast UTH1, SKN7, SOD1, and SOD2 gene expression.37\n\nPrevious studies have demonstrated that purine and pyrimidine derivates can exhibit promising health promoting effects.38,39 Pawelczak et al. investigated the antiaging effects of 4-N-Furfurylcytosine (FC), a cytosine derivative using model S. cerevisiae. They found that treatment with FC enhanced the percentage viability of yeast cells during CLS in a concentration-dependent manner. In addition, treatment with FC boosted the mitochondrial activity and abridged intracellular levels of ROS. It was also demonstrated that FC could limit TORC1 (target of rapamycin complex 1) signaling in yeast. This points to the fact that FC’s anti-aging activity is through the inhibition of TOR signaling pathway.40\n\nGentirigeoside B is a triterpenoid glycoside isolated from the Chinese traditional medicinal plant Gentiana rigescens Franch. Xiang, L., et al. reported that gentirigeoside B significantly extended both the replicative and chronological lifespan of yeast. It was also demonstrated that treatment with gentirigeoside B augmented the activity of antioxidant enzymes (including superoxide dismutase, catalase, and glutathione peroxidase) and abridged the oxidative stress markers (ROS and MDA), thereby preventing oxidative stress-induced reduction in the viability of yeast cells. Gentirigeoside-B treated cells also showed downregulation of Sch9 and activation of Rim15 and Msn2 proteins. This indicates that the anti-aging potential of gentirigeoside B is due to its ability to inhibit the TORC1/Sch9/Rim15/Msn signaling. However, gentirigeoside B could not enhance the life span of yeast mutant lacking antioxidant genes (sod1∆, sod2∆, cat1∆, and gpx∆) and age-related genes (skn7∆ and uth1∆). Therefore, Xiang, L., et al. suggested that though treatment with gentirigeoside B might have longevity-promoting effects in humans, the effect may not be effective in those with mutations in endogenous antioxidant enzyme genes.41 In another study, gentiopicroside, a secoiridoid glycoside that was also isolated from G. rigescens Franch was reported to extend both RLS and CLS of yeast. Gentiopicroside was shown to induce ATG32 gene expression, but could not prolong the RLS and CLS of yeast mutants deficient in ATG32 gene. It was also reported that gentiopicroside enhanced the yeast survival rate under oxidative stress condition by augmenting the activities of enzymatic antioxidants and diminishing the accumulation of ROS and lipid peroxidation. However, gentiopicroside could not affect the RLSs of sod1Δ, sod2Δ, uth1Δ, and skn7Δ. Overall, it can be suggested that gentiopicroside’s anti-aging effects are attributed to its ability to ameliorate autophagy and antioxidative stress response.42\n\nA recent study using yeast models investigated the antioxidant and anti-aging properties of galactan exopolysaccharide isolated from Weissella confusa. Galactan exopolysaccharide was shown to prevent the oxidative stress induced elevation of ROS levels and enhanced the viability of yeast cells exposed to hydrogen peroxide as an oxidant used in this study. In addition, galactan exopolysaccharide treatment significantly increased the viability of both wild type and antioxidant gene-deleted mutant (sod2∆). The study suggested that the antioxidant potential of the galactan exopolysaccharide could be the possible reason for its anti-aging activity.43\n\nMedicinal mushrooms have been reported to exhibit anti-aging activity. In a study,38 researchers isolated two ergosterol derivatives, namely ganodermasides A and B, from the spores of the medicinal mushroom Ganoderma lucidum. They showed that both ganodermasides A and B could extend the replicative life span of S. cerevisiae through the modulation of the expression of UTH. In yeast, different transcription factors such as Skn7, Yap1, and Mot3 play a crucial role in oxidative stress resistance. It was suggested that upon phosphorylation, these transcription factors get activated, thereby regulating the expression of UTH1 gene by binding to its upstream promoter region. It was reported that polyphenols’ anti-aging activity occurs through the activation of Skn7 and subsequent increased expression of the UTH1 gene.44\n\nGinsenosides are major pharmacological compounds that are unique to the plant species Panax ginseng C. A. Meyer (ginseng). Ginsenosides have been demonstrated to increase the life span of different model organism. In a recent study,39 researchers treated yeast cells with ginsenoside Rg1 and found an increased antioxidant stress response and concomitant reduction in ROS levels and apoptosis, thereby enhancing visibility during aging in yeast model. It was also demonstrated that ginsenoside Rg1 treatment resulted in enhanced mitochondrial bioenergetics and glycolytic enzymes, thereby improving metabolic homeostasis and delayed aging in S. cerevisiae.45\n\nIn a previous study,46 we demonstrated the protective effects of quercetin on yeast S. cerevisiae pep4Δ mutant that lack the PEP4 gene (that encodes vacuolar endopeptidase proteinase A, which is a homolog of the human cathepsin D). Yeast pep4Δ cells are found to be highly sensitive to oxidative and apoptotic stressors. However, in our study, we demonstrated that quercetin treatment reduced the ROS levels and apoptotic markers, thereby enhancing the percentage viability of yeast pep4Δ cells during chronological aging.46 Mutation in ATM gene is linked to increased oxidative damage, premature aging, and apoptosis. In another study, we also investigated the protective effects of quercetin on the sensitivity of S. cerevisiae tel1Δ cells lacking Tel1p, which is a homolog of the human ATM (Ataxia Telangiectasia Mutated) gene mutation. Our study results showed that quercetin treatment prevented ROS accumulation and thereby enhanced the stress resistance of tel1Δ cells exposed to a variety of oxidizing agents. Furthermore, treatment with quercetin prevented apoptotic death of yeast tel1Δ cells and increased cell viability during chronological aging.47\n\nCzachor, J., et al. demonstrated the lifespan extending effects of coffee infusions in S. cerevisiae. Coffee, especially the Coffea robusta type, was found to exhibit superior antioxidant effects than the Coffea arabica type, thereby protecting cells from ROS-induced DNA damage and additionally ameliorating the metabolic activity in yeast cells. overall, it can be suggested that coffee exhibits health benefits mainly though the amelioration of ROS accumulation and metabolic activity.48\n\nA study43 reported that the composition of amino acids in aging media can affect the chronological life span of yeast cells. The presence of non-essential amino acids methionine and glutamic acid have been reported to greatly influence the survival rate of yeast cells during aging. Precisely, low levels of methionine and high levels of glutamic acid in the yeast nutrient media led to enhanced lifespan in yeast model. In contrast, increasing levels of methionine and reducing levels of glutamic acid caused a decrease in yeast life span. Therefore, it can be concluded that amino acid composition is a critical factor for controlling the yeast aging.49\n\nEukaryotic cells growth and proliferation is regulated by an evolutionarily conserved kinase, the target of rapamycin (TOR). It has been shown that the inactivation of TOR signaling pathway leads to lifespan extension in different eukaryotic model organisms. The eukaryotic nucleolus is rich in ribosomal DNA (rDNA) that is composed of multiple tandem repeats of rRNA genes (100-200 copies rDNA repeats) and the components for ribosome assembly. Inside the nucleolus, the transcription of rDNA produces precursor rRNA that in turn will be processed further and become associated with ribosomal proteins to form preribosomal subunits. Sir2 is a histone deacetylase enzyme involved in silencing the transcription at the rDNA locus to enhance yeast life span.50,51 Nearly 50% of the rDNA repeats are maintained in a silent state partly by the Sir2 protein.52 Using S. cerevisiae yeast as model, it was found that rapamycin treatment cause inhibition of the TORC1 complex, resulting in the increased association of Sir2 with ribosomal DNA (rDNA) in the nucleolus. This association of SIR2 with rDNA reduces homologous recombination between rDNA repeats that causes formation of toxic extrachromosomal rDNA circles. Thus, rapamycin treatment-induced TORC1 inhibition signals the stabilization of rDNA locus by promoting the association of Sir2 with rDNA, thereby extending the RLS in S. cerevisiae.53\n\nSpermidine has been reported to inhibit histone acetyltransferases and subsequent deacetylation of histone H3, thereby preventing oxidative stress and necrosis in yeast aging. On the contrary, reduction of endogenous polyamines caused hyperacetylation, accumulation of ROS, necrotic cell death and diminished life span. It is important to note that treatment with spermidine causes alteration in acetylation status of the chromatin which results in substantial increase in the autophagy in different model organisms, including yeast. This suggests that spermidine’s anti-aging effects are mediated through its ability to stimulate the autophagy pathway.54\n\nPrevious studies49,50 were conducted to demonstrate the effects of supplementation of copper and iron on the yeast RLS. It was found that addition of copper to the growth media increased the RLS via the activation of multicopper oxidase enzyme (Fet3p) that works in combination with another enzyme called iron permease (Ftr1p) that allows the high intake of iron into yeast cells. Furthermore, it is important to note that high levels of iron-mediated anti-aging effects are dependent on the multicopper oxidase enzyme. However, the life span-extending effects of both copper and iron occurred in the growth medium supplemented with only glycerol as a carbon source but not glucose. It was also demonstrated that supplementation of either copper or iron prevented the accumulation of ROS levels, thereby enhancing the replicative life span of yeast mutants lacking antioxidant enzymes.55,56 The iron-mediated anti-aging effects occur mainly through the increased expression of genes related to the mitochondrial metabolism especially tricarboxylic acid (TCA) cycle and electron transport chain, subsequently elevated levels of adenosine triphosphate (ATP), which is essential for the increased survival cells during aging. Interestingly, supplementation of iron could also increase the life span of yeast Snf1Δ mutant which lacks SNF1/AMPK protein kinase. Therefore, it can be suggested that the iron’s life span-extending effects occur via amelioration of mitochondrial energy metabolism in yeast.57\n\nA recent study58 isolated a compound inokosterone from G. rigescens Franch and demonstrated that inokosterone can increase both the chronological and replicative life spans. The inokosterone was shown to enhance the survival rate of yeast cells by ameliorating the levels of antioxidant enzymes (e.g., SOD) and preventing accumulation of oxidative stress markers (e.g., ROS and MDA levels). Further, inkosterone could increase the autophagy (especially mitophagy) in yeast cells. Likewise, the same study also demonstrated that treatment with inkosterone decreased the oxidative stress and enhanced autophagy in mammalian cell lines. Therefore, it can be suggested that inkosterone’s anti-aging effects are mediated through the activation of antioxidant stress response and mitophagy.58\n\nPrevious studies have reported that lithocholic acid, a bile acid, can increase yeast cell survival during chronological aging.59–62 Interestingly, lithocholioc acid was also shown to significantly enhance the survival rate of yeast especially under caloric restriction conditions. Lithocholic acid treatment was shown to modulate several cellular pathways, including carbohydrate and lipid metabolism, mitochondrial structure and function, liponecrotic and apoptotic cell death of yeast during chronological aging.59 It is important to note that lithocholic acid’s anti-aging effects under caloric restriction are found to be prominent only when this compound is added to the growth medium either at logarithmic/diauxic and early stationary stages of yeast aging. Addition of lithocholic acid either at logarithmic/diauxic and early stationary stages induced the activation of several longevity related cellular processes which ultimately triggering the enhanced yeast cell survival during aging.60 Beach et al. also demonstrated that lithocholic acid exerts its anti-aging effects through the modulation of the expression of different transcription factors including Aft1p, Hog1p, Msn2/4p, Rtg1p- Rtg3p, Sfp1p, Skn7p, and Yap1p. Each of these transcription factors in turn alter the levels of several intra and extra mitochondrial proteome, and subsequent maintenance of mitochondrial function. Altogether, Beach et al. suggested that lithocholic acid’ anti-aging effects are mainly through the modulation of aging-related transcriptional landscape.61 Also, it was discovered that lithocholic bile acid accumulates in mitochondria, and alters the mitochondrial membrane lipidome which is crucial for the restoration of mitochondrial proteome, subsequently an improved mitochondrial function and increased chronological aging by lithocholic acid.62\n\nIn another study, Sun Y et al., isolated lysophosphatidic acid (LA) from the seeds of Arabidopsis thaliana and showed that LA increased oxidative stress resistance, thereby promoting the extension of RLS in yeast. However, LA was not able to extend the replicative lifespan of mutants uth1Δ, skn7Δ, sod1Δ, andsod2Δ. This study suggests that LA’s anti-aging effects are possibly through the amelioration of antioxidant status of yeast cells and the genes of UTH1, SKN7, and SOD may also be involved in the action.63\n\nIn our previous study,58 we reported the anti-aging effects of magnolol, a natural polyphenol. Magnolol increased the stress resistance of yeast cells exposed to hydrogen peroxide, an oxidizing agent. In addition, magnolol increased the viability of short-lived yeast mutant sod1∆, which lacks the antioxidant enzyme superoxide dismutase. Our study suggested that the anti-aging effects of magnolol occur mainly through the modulation of oxidative stress during yeast chronological aging.64\n\nMulberry leaves are rich in flavonoids such as morusin and mulberrin, and these have been demonstrated to increase survival rate during yeast chronological aging. It was also found that morusin and mulberrin exert their anti-aging effects via targeting the SCH9, a major target of TORC1 in budding yeast S. cerevisiae.65 In eukaryotes, cell growth is regulated by two different, highly conserved multiprotein complexes such as TORC1 and TORC2. These complexes are comprised of a large catalytic subunit, the serine/threonine kinase target of rapamycin (TOR). Budding yeast Sch9 is an AGC (Protein Kinase A (PKA), G (PKG), and C (PKC)) family kinase and is the major substrate of the TORC1. Yeast Sch9 functions analogously to S6K1, the mammalian TORC1 substrate. Yeast TORC1 phosphorylates about six amino acid residues of Sch9, indicating that the TORC1-dependent phosphorylation of Sch9 is essential for its activity.66 However, rapamycin treatment, carbon and nitrogen starvation inhibit phosphorylation by TORC1. Activated Sch9 plays a key role in the regulation of several cellular processes including protein synthesis (via modulating ribosome synthesis and translation initiation), cell cycle, and aging. Deletion of SCH9 causes growth defects such as reduction in cell size and growth rate; however, deletion of SCH9 leads to enhanced temperature tolerance as well chronological and replica life spans in budding yeast. The longevity-extending effects due to SCH9 deletion are explained by the augmented oxidative stress response and reduction in age-associated mutation rate.67\n\nXiang, L. et al., investigated the anti-aging effects of apple polyphenol, phloridzin using yeast RLS model. Phloridzin treatment significantly increased the percentage viability of yeast cells exposed to hydrogen peroxide. It was also demonstrated that phloridzin treatment led to increased expression of SOD1/2 and SIRT1 genes as well as the activity of superoxide dismutase. Altogether, it was suggested that the anti-aging effects of phloridzin are mediated through the regulation of expression of SIRT1 in budding yeast.68\n\nPolyalthia longifolia is a polyphenol-rich traditional medicinal plant that has long been used for its rejuvenation capacity. P. longifolia has been reported to exhibit potent pharmacological activities including antioxidant and hepatoprotective activities. A previous study63 tested the anti-aging activity of methanolic leaf extract of P. longifolia using yeast CLS model and found that the methanolic leaf extract enhanced the viability of yeast cells and increased the CLS of yeast. It was reported that methanolic leaf extract significantly prevented the accumulation of H2O2-induced increase in ROS levels and augmented the GSH levels. Furthermore, treatment with methanolic leaf extract strikingly enhanced the expression levels of SOD and SIRT1 genes. Overall, it can be suggested that P. longifolia exerts its anti-aging effects through the modulation of oxidative stress response and SIRT1 gene.69 It was also reported that methanolic leaf extract of P. longifolia extended the RLS of yeast via ameliorating the apoptotic features of replicatively ageing yeast cells.70\n\nIn another study, using budding yeast as a model, researchers investigated antioxidant and anti-aging effects of myricetin, a polyphenolic flavonoid compound which is composed of a pyrogallol ring in the B ring of its structure. Myricetin’s antioxidant effects are attributed to the presence of a hydroxyl group in its B ring. Treatment with myricetin reduced the augmentation of ROS and protein carbonyl levels, thereby increasing the oxidative stress resistance of yeast cells exposed to hydrogen peroxide. Myricetin was found to inhibit H2O2-induced glutathione oxidation, but did not increase endogenous antioxidant enzymatic activity in yeast. Additionally, myricetin treatment significantly prevented age associated oxidative stress and enhanced the chronological lifespan of yeast mutant lacking mitochondrial superoxide dis-mutase (sod2Δ).71\n\nAlterations in the biosynthesis and regulation of nicotinamide adenine dinucleotide (NAD+) plays a crucial role in the progression of aging as well as pathophysiology of several age-related chronic diseases.72 NAD + plays a crucial role in several metabolic processes in the cells. Most importantly, NAD+ functions as a co-factor in redox reactions and it is reduced to NADH in many metabolic pathways including glycolysis, the citric acid cycle, and fatty acid metabolism. Adequate intracellular levels of NAD+ are maintained through the combined action of NAD(+) biosynthesis and salvage pathways. Several key cellular enzymes including sirtuin protein deacetylases and poly-ADP-ribose polymerases (PARPs) are NAD+ dependent, and use NAD+ as a substrate in many cellular processes, including aging. Aging is associated with a reduction in NAD+ levels, leading to alterations in several metabolic processes that are NAD+-dependent, which in turn leads to the ageing-associated physiological/functional decline. Therefore, supplementation with diets rich in three important NAD+ precursors 1) nicotinamide, 2) nicotinic acid (e.g., protein rich foods such as cereals, peanuts, meat, fish) and 3) nicotinamide riboside (e.g., milk, cabbage, cucumber) ameliorate age-related decline in NAD+ levels and help in the extension of active longevity.73 Nicotinamide riboside is a natural product present in milk.13 NAD+ is also required for the proper functioning of Sir2 and extension of replicative life span in the budding yeast. Alterations in NAD+ levels negatively affect the RLS of yeast. In eukaryotes, nicotinamide riboside kinases (Nrk1 and Nrk2) catalyze the phosphorylation of nicotinamide riboside to nicotinamide mononucleotide, the precursor of NAD+. A previous study reported that the supplementation of nicotinamide riboside increased the levels of NAD+ via activation of both Nrk1-dependent pathway and the Urh1/Pnp1/Meu1 pathway (an Nrk1-independent pathway). Increased NAD+ levels in turn lead to increased Sir2-dependent gene silencing, thereby extending RLS in yeast.74 The Sir2 enzyme deacetylates the lysine residues of histones, thereby silence all heterochromatin-like regions including telomeres, rDNA, and the hidden mating type loci HML/HMR (Hidden MAT Left/ (Hidden MAT Right).75 In the S. cerevisiae NAD+ salvage pathway, nicotinamide is recycled from NAD+ by the Sir2 mediated deacetylation reaction. The nicotinamidase (Pnc1) catalyzes the conversion of nicotinamide to nicotinic acid, which is further converted to nicotinamide mononucleotide by the enzyme nicotinamide phosphoribosyltransferase (Npt1). Isonicotinamide (a compound similar in shape and electrical properties to nicotinamide) was shown to elevate the intracellular levels of NAD+ through the yeast NAD+ salvage pathway and concomitant increase in the Sir2 activation resulting in the enhanced normal silencing at the rDNA locus in yeast and extension of yeast RLS.76\n\nMitochondrial dynamics, the balance between mitochondrial fission and fusion, is critical for cell growth and functioning. Replicative senescence in yeast is characterized by the presence of fragmented mitochondria due to mitochondrial fission rather than fusion, indicating altered mitochondrial dynamics. Wang et al. reported that treatment with resveratrol led to a significant reduction in the number of senescent yeast cells with fragmented mitochondria. This indicates that resveratrol’s anti-aging effects possibly occur via modulating the expression of genes associated with mitochondrial dynamics during RLS.77\n\nPigmented rice is the functional food in many countries including India, China, and Japan, and it is the richest source of polyphenols.78 It has been reported that red rice bran extract could prevent the accumulation of ROS levels, maintain the plasma membrane integrity and extend the chronological lifespan of yeast cells.79 The anti-aging mechanism of action of rice bran extract was found to be through the modulation of TOR1 and SIR2-dependent pathway.79\n\nThe stamen of lotus (Nelumbo nucifera) is rich in flavonoids and has long been widely used in Indian traditional medicine. Tungmunnithum et al. reported that treatment with lotus stamen extract significantly increased the antioxidant status (both enzymatic activity and gene expression levels of SOD1 and SIRT2) and extended the chronological lifespan of yeast cells. Interestingly, the longevity extending effects of lotus stamen extract were reported to be superior to the resveratrol treatment.80\n\nWu et al. found that the dried roots of Salvia miltiorrhiza Bunge have substantial longevity extending effects. They reported that tanshinones (e.g., cryptotanshinone, tanshinone I, and tanshinone IIa) are pharmacologically active components present in the roots of S. miltiorrhiza. Precisely, cryptotanshinone has been reported to extend the CLS of yeast wild type as well as yeast mutant lacking mitochondrial superoxide dismutase (sod2Δ). Their study suggests that the cryptotanshinone-induced anti-aging effects are modulated through the involvement of nutrient-sensing protein kinases such as Tor1, Sch9, and Gcn2.81\n\nParishin is a phenolic glucoside isolated from Gastrodia elata, a Chinese traditional medicinal plant. Parishin treatment significantly increased cell viability in oxidative stress and also extended the replicative life span of K6001 yeast. Parishin treatment significantly increased the expression of levels of SIR2 and SOD activity, while preventing the accumulation of ROS and lipid peroxidation. However, Parishin treatment did not increase the RLS of sod1Δ, sod2Δ, uth1Δ, and skn7Δ mutants of K6001 yeast. Treatment with parishin resulted in the reduced expression of several TOR signaling pathway-related genes such as TORC1, ribosomal protein S26A (RPS26A), and ribosomal protein L9A (RPL9A). In addition, parishin treatment significantly diminished gene expression levels of RPS26A and RPL9A in uth1Δ mutant, as well as in uth1 Δsir2Δ double mutants. Also, parishin treatment remarkably inhibited the TORC1 gene expression in uth1 mutants. These study results indicate that parishin exerts its anti-aging activities by modulating Sir2/Uth1/TOR signaling pathway.82\n\nP. corylifolia is a medicinal plant widely used in the traditional medicine systems in Indian and China. Wang et al. showed that the ethanol extract of P. corylifolia significantly increased the RLS of yeast. In particular, the n-hexane fraction of the ethanol extract of P. corylifolia improved the yeast lifespan by 20%. Interestingly, the n-hexane extract of P. corylifolia extended the mean lifespan of the sir2Δfob1Δ double mutant strain, indicating that the n-hexane fraction prolongs the RLS in a Sir2-independent pathway. Treatment with P. corylifolia did not extend the RLS of the tor1Δ mutant, suggesting that Tor1 plays a major role in the extension of RLS by the n-hexane-soluble fraction P. corylifolia. Furthermore, it was reported that, Corylin and neobavaisoflavone are the active compounds in P. corylifolia that significantly increased the viability and extended the RLS of yeast. In particular, corylin was found to be more effective in enhancing the yeast RLS compared to neobavaisoflavone. It was also shown that corylin treatment promoted RLS of the sir2Δfob1Δ mutant strain but failed to prolong the RLS of the tor1Δ mutant, suggesting that corylin exerts life span-extending effects in a Tor1-dependent pathway. Fascinatingly, under CR conditions, corylin could not extend the RLS. In addition, corylin treatment did not significantly influence the CLS of yeast. Gtr1 in S. cerevisiae encodes a highly conserved GTPase that is necessary for TORC1 activation and amino acid sensing. The G protein complex Gtr1/Gtr2 activates Tor1 in S. cerevisiae in response to signals from amino acids.83 Furthermore, it was also suggested from docking studies that corylin prolongs the yeast RLS by blocking Gtr1 activation.84\n\nPrevious studies by Lee MB et al85 also demonstrated that green tea extract and berberine could strongly shorten the yeast life span, whereas Pterocarpus marsupium extract and other mixtures containing P. marsupium significantly extended the yeast life span.85 In another study, two sesquiterpene glucosides isolated from the Shenzhou honey peach fruit were also able to extend the RLS of K6001 yeast. Treatment with sesquiterpene glucosides increased the survival rate of yeast under oxidative stress. Besides, treatment with sesquiterpene glucosides could not affect the RLSs of SOD mutant yeast strains with a K6001 background, indicating that the anti-oxidative stress response performs important roles in anti-aging effects of sesquiterpene glucosides.86\n\n\nConclusions\n\nAging is a significant risk factor for the emergence of several chronic human diseases. In order to reduce the pathophysiology of diseases related to aging and to increase the active lifetime of people, aging research is primarily concerned with the identification of new anti-aging therapies. Budding yeast S. cerevisiae has been used as a valuable model to evaluate the anti-aging properties of phytochemicals as well as synthetic compounds. Several studies demonstrated that plant extracts/phytochemicals can increase longevity via regulation of various pathways including the TORC1/Sch9/Rim15/Msn signaling pathway, RAS-AC-PKA pathway, Sir2-dependent pathway, NAD+-salvage pathway, Uth1/TOR signaling, and antioxidant stress response pathway as well as autophagy. While many of these studies reported enhancing the yeast cell’s oxidative stress resistance, a few studies have also documented the pro-oxidant effects of various antioxidants, suggesting dose-dependent effects on life span. For example, in the K6001 yeast strain of S. cerevisiae, treatment with alpha-tocopherol and coenzyme Q10 enhanced oxidative stress and shortened RLS, showing the potential pro-oxidant effects of antioxidants that may be dose-dependent.87 Therefore, to maximize the therapeutic benefits of antioxidants, it is necessary to evaluate their dose-dependent effects as well as potential pro-oxidant actions. The discovery of novel plant-derived chemicals with anti-aging activities opens up new opportunities for developing leading medications for the treatment of age-related disorders.\n\nIn conclusion, aging research using budding yeast as a model organism has yielded valuable insights into the anti-aging properties of various compounds, and the identification of specific pathways and mechanisms involved contributes to the development of potential anti-aging therapies for improving health span and longevity in humans. Continued research into plant extracts and natural compounds may lead to the identification of promising candidates for fighting age-related disorders in the future.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nAcknowledgements\n\nPhaniendra Alugoju thanks to C2F (Secondary Century Fund) Postdoctoral Fellowship, Chulalongkorn University, Bangkok 10330, Thailand. 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PubMed Abstract \| Publisher Full Text" }	[ { "id": "212350", "date": "24 Oct 2023", "name": "Srinivasa Rao Sirasanagandla", "expertise": [ "Reviewer Expertise Natural compounds against the environmental pollutant induced toxicity", "radiological anatomy", "morphological variations" ], "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 current review presents a summary of aging and the use of Saccharomyces cerevisiae as a model organism for anti-aging research. It's informative and well-structured, making it suitable for readers interested in the topic. Here are a few suggestions for elevating the quality of this review.\nIn the abstract, authors should provide a clear overview of the research focus, its significance, and what readers can expect from the review. Begin with a clear statement of the objective or focus of the research. For example, \"This review aims to explore the anti-aging properties of several natural products and phytoextracts using the budding yeast Saccharomyces cerevisiae as a model organism.\" Explain the significance of studying aging and anti-aging research. Highlight the Contribution. Include a statement about what readers can expect from the review.\n\nIn the introduction sections, please cite the figures 1 and 2.\n\nIt would be more informative if authors had provided a table summarizing the action of natural products and/or plant extracts.\n\nThe authors should also highlight the advantages and disadvantages of using yeast as a model for anti-aging research.\n\nThe authors should include a section on emerging trends that focuses on identifying emerging trends and cutting-edge techniques in the study of anti-aging compounds.\n\nA distinct section focusing on future directions that need further exploration in the field should also be added.\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": "12923", "date": "17 Dec 2024", "name": "Tewin Tencomnao", "role": "Author Response", "response": "In the abstract, authors should provide a clear overview of the research focus, its significance, and what readers can expect from the review. Begin with a clear statement of the objective or focus of the research. For example, \"This review aims to explore the anti-aging properties of several natural products and phytoextracts using the budding yeast Saccharomyces cerevisiae as a model organism.\" Explain the significance of studying aging and anti-aging research. Highlight the Contribution. Include a statement about what readers can expect from the review. Response: We thank reviewer for the insightful comment on the abstract. We have revised the abstract by incorporating a clear overview of the research focus and the significance of the aging and anti-aging research. In the introduction sections, please cite the figures 1 and 2. Response: We apologies for overlook in not citing the figures. We have cited Figures 1 and 2 at appropriate sections of the revised manuscript. It would be more informative if authors had provided a table summarizing the action of natural products and/or plant extracts. Response: As suggested by the reviewer, we have summarized the anti-aging molecular mechanisms of actions of natural products and plant extracts in Tables 1, 2, and 3. The authors should also highlight the advantages and disadvantages of using yeast as a model for anti-aging research. Response: “The main advantages of using the yeast Saccharomyces cerevisiae in aging research are that it is a unicellular eukaryote that exhibits characteristic features such as rapid growth, inexpensive, and ease of genetic manipulation, allowing to conduct experiments quickly and study the effects of specific genes on aging. Several of the cellular processes and metabolic pathways, including aging, are conserved between S. cerevisiae and humans. These features make yeast an ideal model organism for the high-throughput screening of identifying genes and chemical compounds associated with aging. However, the use of yeast has some limitations. For example, its unicellular nature limits its ability to model complex multicellular organisms and tissue-specific aging processes in humans. Additionally, some post-translational modifications that occur in humans do not occur in yeast, and yeast metabolism significantly differs from humans, which may limit applicability if certain findings to human aging research.” This information has been included in the section titled “The budding yeast Saccharomyces cerevisiae – A simple eukaryotic model to study aging” in the revised version of the manuscript. The authors should include a section on emerging trends that focuses on identifying emerging trends and cutting-edge techniques in the study of anti-aging compounds. Response: We thank reviewers for their valuable suggestion on the necessity of adding a section on the emerging trends in the study of anti-aging compounds. As suggested, a separate section is included in the revised manuscript. A distinct section focusing on future directions that need further exploration in the field should also be added. Response: A separate section compiling both emerging trends and future developments in the anti-aging research are included in the revised manuscript." } ] }, { "id": "253397", "date": "26 Mar 2024", "name": "John Hartman", "expertise": [ "Reviewer Expertise yeast genetics", "human disease models", "CLS" ], "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 may want to consider more precise and directional adjectives, such as enhance, suppress, augment, alleviate, etc. Use of the word ‘ameliorate’ to qualify various biological effects, seemed vague, inconsistent, and sometimes incorrect. My suggestion would be to carefully proof-read again for grammar in places but also for biological clarity, for example, about relationships between TOR signaling and respiration.\nOverall, I thought the review topic was very interesting - the authors have assembled a collection of papers about natural products (NPs) where biological activities have been studied using paradigms (RLS and CLS) of yeast aging.\n\nThe abstract and introduction could be greatly enhanced by removing much of the philosophical and physiological descriptions of aging, how it's fundamental to several diseases, and the utility and experimental benefits of model organisms, all of which have been made many times and are thus an unnecessary distraction. If condensed to a couple sentences and citations there would be much space cleared to focus more on the topic at hand which is already very rich and extensive and could stand to be expanded upon and further detailed.\nFig. 1 is awkward IMO (in a BioRender sort of way) and it's not really needed. To regain that space and maintain reader attention, a simple brief description of RLS and CLS would do (with just a few of many possible citations that could support the rare reader who may not be already familiar).\n\nIt would be helpful to organize the paper according to which NPs were studied in the context of RLS, CLS, or both. The models are quite different, though they involve related mechanisms as the authors point out. Clarifying and distinguishing (or unifying) between the RLS/CLS models over the course of the MS would be of interest (more to come on this and other factors that could be better organized in order to increase the impact of the work).\nFig. 2 is helpful, but should be expanded with regard to the genes and pathways (and the page space) and doesn't need to be rendered as an actual cell if removing the picturesque constraints of the cell and organelles would provide more flexibility to communicate important information about the literature.\nThe \"conserved nutrient sensing aging regulatory pathways\" section is critical and could be expanded substantially. \"Conserved\" presumably means \"between\" yeast and humans? This section seems woefully under-cited. If possible, all of the genes mentioned in the NP papers should be incorporated into this section and Fig. 2 and related to one another, which would ultimately help to relate the NPs to each other regarding their influences on cellular aging. There is an opportunity here to relate the yeast pathways to the human pathways (i.e. to relate the conserved pathways), and ultimately to suggest how some of the NPs, which influence yeast aging, could be related to benefits they are thought to putatively have in humans. I think that opportunity wasn't really capitalized on but would be great if the authors could. This would provide demonstrations for the significance of the topic and the importance of the review as a starting point to increase knowledge about relationships between NPs, yeast aging models, and human age-related disease.\nThe \"Anti-Aging studies \"of NPs\" in S. cerevisiae\" section should mention NPs in the section subtitle (just for clarity).\n\nRather than just a list of NPs - this section would be much more useful if organized by important factors. I'm not sure exactly how to do it, but a table would very helpful for handling this aspect. Strategies that could be used to organize this section include alphabetical order, whether influences are on CLS, RLS, or both; class of compounds; relevant sources of the compounds (foods, alternative medicines, natural metabolism, microbiome, etc); whether they have known or suspected effects on human health; MOA or genes and pathways they are thought to act or interact with; whether the NPs are easily obtained for research purposes (i.e. where to buy them); other factors that the authors may be aware of.\n\nA table that synthesizes and distills the important factors relevant to each NP should be created (as also suggested by Reviewer 1). The table should have a field dedicated to each factor (see a few suggestions in paragraph above) and have concise record entries for each NP - one of the fields should be for the references for convenient lookup of information in the table context. If provided as a spreadsheet, it would be particularly useful.\nJust my personal opinion, but I didn't see how descriptions of the advantages of yeast, emerging trends, cutting edge techniques, or future directions would be as helpful as more detail and better organization about the NPs, in-depth summary about the literature where yeast have been used to study them, and if possible where they are are also relevant to human and/or specific age-related diseases.\n\nAbout \"conclusions drawn being 'partly' supported\" there can be large effects on CLS deriving from differing auxotrophic background of yeasts strain and media recipes. Things like glucose concentration, whether the media is buffered, amino acid compositions, the presence of ammonium sulfate, rich vs. synthetic/defined media, the list goes on. The interactions between the NPs and genetic background was discussed in the review, but these same types of interactions exist with auxotrophic mutations and media composition (which can vary dramatically between studies). This fact should be briefly mentioned in the manuscript and ideally accounted for in the proposed table by indicating the strain, auxotrophic background and media used in the studies employing particular NPs. Additives like H2O2 that are used to enhance characterization are mentioned in places and could also be accounted for in the table.\nI appreciate the authors' efforts and insights hope they find this feedback useful.\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": "12924", "date": "17 Dec 2024", "name": "Tewin Tencomnao", "role": "Author Response", "response": "The authors may want to consider more precise and directional adjectives, such as enhance, suppress, augment, alleviate, etc. Use of the word ‘ameliorate’ to qualify various biological effects, seemed vague, inconsistent, and sometimes incorrect. My suggestion would be to carefully proof-read again for grammar in places but also for biological clarity, for example, about relationships between TOR signaling and respiration. Response: We thank you for your suggestions on the precise usage of adjectives and grammar in this manuscript. As suggested, we have carefully proofread the document and considered the usage of more precise and directional adjectives throughout the revised manuscript. Overall, I thought the review topic was very interesting - the authors have assembled a collection of papers about natural products (NPs) where biological activities have been studied using paradigms (RLS and CLS) of yeast aging. The abstract and introduction could be greatly enhanced by removing much of the philosophical and physiological descriptions of aging, how it's fundamental to several diseases, and the utility and experimental benefits of model organisms, all of which have been made many times and are thus an unnecessary distraction. If condensed to a couple sentences and citations there would be much space cleared to focus more on the topic at hand which is already very rich and extensive and could stand to be expanded upon and further detailed. Response: As suggested by the reviewer, the philosophical and physiological descriptions of aging are removed from the abstract and introduction of the revised manuscript. Additionally, the review has been focused on the natural products (please see Introduction). Fig. 1 is awkward IMO (in a BioRender sort of way) and it's not really needed. To regain that space and maintain reader attention, a simple brief description of RLS and CLS would do (with just a few of many possible citations that could support the rare reader who may not be already familiar). Response: As suggested by the reviewer, we have included additional information on CLS and RLS along with their respective citations. However, we believe that inclusion of Figure 1 may help readers, particularly budding yeast researchers working on anti-aging research, and facilitate an easier understating of the concepts of CLS and RLS. It would be helpful to organize the paper according to which NPs were studied in the context of RLS, CLS, or both. The models are quite different, though they involve related mechanisms as the authors point out. Clarifying and distinguishing (or unifying) between the RLS/CLS models over the course of the MS would be of interest (more to come on this and other factors that could be better organized in order to increase the impact of the work). Response: We appreciate your insightful suggestion and we have reorganized the anti-aging effects of natural products according to the relevance of CLS or RLS or both. We do believe that this reorganization further enhances the impact of work. Fig. 2 is helpful, but should be expanded with regard to the genes and pathways (and the page space) and doesn't need to be rendered as an actual cell if removing the picturesque constraints of the cell and organelles would provide more flexibility to communicate important information about the literature. Response: As suggested by the reviewer, Figure 2 is improved in the revised manuscript. The \"conserved nutrient sensing aging regulatory pathways\" section is critical and could be expanded substantially. \"Conserved\" presumably means \"between\" yeast and humans? This section seems woefully under-cited. If possible, all of the genes mentioned in the NP papers should be incorporated into this section and Fig. 2 and related to one another, which would ultimately help to relate the NPs to each other regarding their influences on cellular aging. There is an opportunity here to relate the yeast pathways to the human pathways (i.e. to relate the conserved pathways), and ultimately to suggest how some of the NPs, which influence yeast aging, could be related to benefits they are thought to putatively have in humans. I think that opportunity wasn't really capitalized on but would be great if the authors could. This would provide demonstrations for the significance of the topic and the importance of the review as a starting point to increase knowledge about relationships between NPs, yeast aging models, and human age-related disease. Response: Thank you for your insightful suggestion. The section entitled “conserved nutrient-sensing aging regulatory pathways” has been revised with sufficient citations for each of the singling pathways discussed in the review. The \"Anti-Aging studies \"of NPs\" in S. cerevisiae\" section should mention NPs in the section subtitle (just for clarity). Response: As suggested, the tile of the section has been changed to “Anti-aging studies of natural products and plant extracts in the budding yeast S. cerevisiae as a model.” Rather than just a list of NPs - this section would be much more useful if organized by important factors. I'm not sure exactly how to do it, but a table would very helpful for handling this aspect. Strategies that could be used to organize this section include alphabetical order, whether influences are on CLS, RLS, or both; class of compounds; relevant sources of the compounds (foods, alternative medicines, natural metabolism, microbiome, etc); whether they have known or suspected effects on human health; MOA or genes and pathways they are thought to act or interact with; whether the NPs are easily obtained for research purposes (i.e. where to buy them); other factors that the authors may be aware of. Response: We thank you for your valuable suggestions on the organization of natural products. As suggested, the section on the natural products and plant extracts has been reorganized, with the anti-aging studies now arranged alphabetically in the revised version of the manuscript. A table that synthesizes and distills the important factors relevant to each NP should be created (as also suggested by Reviewer 1). The table should have a field dedicated to each factor (see a few suggestions in paragraph above) and have concise record entries for each NP - one of the fields should be for the references for convenient lookup of information in the table context. If provided as a spreadsheet, it would be particularly useful. Response: Thank you for your suggestion. A table summarizing the natural products and plant extracts including their source, chemical class, dosage used, growth medium used for aging studies, yeast strain used, and their anti-aging mechanisms, has been included in the revised manuscript. Please refer to table 1, 2, and 3 Just my personal opinion, but I didn't see how descriptions of the advantages of yeast, emerging trends, cutting edge techniques, or future directions would be as helpful as more detail and better organization about the NPs, in-depth summary about the literature where yeast have been used to study them, and if possible where they are also relevant to human and/or specific age-related diseases. Response: We thank you for your insightful suggestions on the importance of adding in depth literature on the natural products and the reorganization of the natural products section. We have reorganized the revised manuscript and included an in-depth summary of natural products. About \"conclusions drawn being 'partly' supported\" there can be large effects on CLS deriving from differing auxotrophic background of yeasts strain and media recipes. Things like glucose concentration, whether the media is buffered, amino acid compositions, the presence of ammonium sulfate, rich vs. synthetic/defined media, the list goes on. The interactions between the NPs and genetic background was discussed in the review, but these same types of interactions exist with auxotrophic mutations and media composition (which can vary dramatically between studies). This fact should be briefly mentioned in the manuscript and ideally accounted for in the proposed table by indicating the strain, auxotrophic background and media used in the studies employing particular NPs. Additives like H2O2 that are used to enhance characterization are mentioned in places and could also be accounted for in the table. Response: We briefly mentioned in the manuscript how different factors such as carbon sources, amino acid composition, pH of the growth medium affects the CLS. Additionally, we have included a table that accounts for the strain background, media used in anti-aging studies of natural products/plant extract." } ] } ]	1	https://f1000research.com/articles/12-1265
https://f1000research.com/articles/13-844/v1	29 Jul 24	{ "type": "Research Article", "title": "Implementation of Chernobyl optimization algorithm based feature selection approach to predict software defects", "authors": [ "Kunal Anand", "Ajay Kumar Jena", "Himansu Das", "Ajay Kumar Jena", "Himansu Das" ], "abstract": "Background Software defects can have catastrophic consequences. Therefore, fixing these defects is crucial for the evolution of software. Software Defect Prediction (SDP) enables developers to investigate unscramble faults in the inaugural parts of the software progression mechanism. However, SDP faces many challenges, including the high magnitude of attributes in the datasets, which can degrade the prognostic performance of a defect forecasting model. Feature selection (FS), a compelling instrument for overcoming high dimensionality, selects only the relevant and best features while carefully discarding others. Over the years, several meta-heuristic algorithms such as the Genetic Algorithm (GA), Particle Swarm Optimization (PSO), Differential Evolution (DE), and Ant Colony Optimization (ACO) have been used to develop defect prediction models. However, these models suffer from several drawbacks, such as high cost, local optima trap, lower convergence rate, and higher parameter tuning. To overcome the above shortcomings, this study aims to develop an innovative FS technique, namely, the Chernobyl Optimization Algorithm (FSCOA), to unwrap the most informative features that can produce a precise prediction model while minimizing errors.\n\nMethods The proposed FSCOA approach mimicked the process of nuclear radiation while attacking humans after an explosion. The proposed FSCOA approach was combined with four widely used classifiers, namely Decision Tree (DT), K-nearest neighbor (KNN), Naive Bayes (NB), and Quantitative Discriminant Analysis (QDA), to determine the finest attributes from the SDP datasets. Furthermore, the accuracy of the recommended FSCOA method is correlated with existing FS techniques, such as FSDE, FSPSO, FSACO, and FSGA. The statistical merit of the proposed measure was verified using Friedman and Holm tests.\n\nResults The experimental findings showed that the proposed FSCOA approach yielded the best accuracy in most cases and achieved an average rank of 1.75, followed by the other studied FS approaches. Furthermore, the Holm test showed that the p-value was lower than or equivalent to the value of α/(A-i), except for the FSCOA and FSGA and FSCOA and FSACO models.\n\nConclusion The experimental findings showed that the prospective FSCOA procedure eclipsed alternative FS techniques with higher accuracy in almost all cases while selecting optimal features.", "keywords": [ "Software Defect Prediction", "Feature Selection", "Wrapper approach", "Chernobyl Disaster Optimizer", "Optimization" ], "content": "Introduction\n\nIn today’s scenario, humankind needs good-quality and reliable software to help them perform their daily tasks without spending more time and effort. Owing to this immense call for exceptional and dependable software, conducting a rigorous investigation of software under development is crucial. However, the complexity of software increases every passing day, making the overall software development work very challenging.1,2 A fault in software can significantly damage its quality and reliability, leading to more frequent maintenance activities. This can result in higher operational costs for the software, ultimately leading to user dissatisfaction. A software fault can be characterized as the disparity between the actual and predicted behaviors of the software. Software testing empowers developers to identify and rebuild faults. However, conventional testing approaches are costly and time consuming. Hence, it is imperative to detect faults in a software module during the early stages of evolution.3\n\nSoftware Defect Prediction (SDP) enables developers to expose deficiencies in software components in the early stages of development by employing data analysis and machine learning (ML) approaches. An effective SDP mechanism can lead to the systematic and profitable advancement of high-quality and reliable software products without defects.4 Researchers have suggested several ML-based SDP approaches5–8 for effectively predicting defects. These methods analyze past data from different stages of development, such as testing data and debugging records, to derive any pattern or trend that can detect any potential defects. The most widely employed ML methods in SDP are DT,9 SVM,10 neural networks,11 logistic regression,12 and NB.13 However, these approaches suffer from many challenges including high dimensionality being one of them.\n\nFeature Selection (FS)14 is a potent mechanism that can be employed to overcome the issue of high dimensionality. FS allows developers to select only relevant features and carefully discard insignificant features. In SDP, FS is a vital step that allows developers to choose the best set of features that can significantly enhance the predictive accuracy of a defect prediction model. The application of FS approaches is essential when dealing with datasets with high dimensionality. Several FS approaches have been implemented in the SDP. FS techniques are broadly classified into three categories: filter techniques,15 wrapper techniques,16 and embedded techniques.17,18 Filter-based FS techniques are autonomous for any training strategy, and apply statistical properties to identify the best traits. Nonetheless, wrapper-based FS procedures select the best characteristics based on the classification accuracy of the prediction model. Embedded-based FS techniques combine feature selection with model training. Existing literature shows that researchers have mainly applied evolution-based algorithms19 and swarm-based algorithms20 for FS purposes.\n\nThis exploration aims to boost the classification truthfulness of a defect-foretelling model while minimizing errors. For this purpose, this study uses some of the widely used meta-heuristic algorithms, namely the Genetic Algorithm (GA),21 Particle Swarm Optimization (PSO),22,23 Differential Evolution (DE),24 and Ant Colony Optimization (ACO).25 Although GA has been a proven FS approach,26 it is costly because it computes the optimal features using genetic techniques such as selection, crossover, and mutation over a set of generations. The PSO-based FS approach27 aims to find the optimal traits by emulating the fragment’s movement while probing a search arena with several dimensions. The algorithm adjusts the location and velocity of the fragment by considering singular and group knowledge. However, the PSO-based FS approach sometimes results in a local optima trap in addition to a lower convergence rate. DE-based FS techniques28 compute optimal characteristics by employing operators such as mutation, crossover, and selection on a population of potential solutions over several iterations. However, these approaches require several parameter tunings, making them a tedious choice among researchers. ACO-based FS methods29 determine the excellent characteristics of a search space by implementing the foraging behavior of ants. However, these methods suffer from a slow convergence speed and low accuracy, especially in large datasets.\n\nThe limitations of the aforementioned FS approaches motivated us to propose a novel FS approach (FSCOA) inspired by the Chernobyl Disaster Optimizer (CDO).30 The primary objective of the proposed FSCOA approach is to unwrap the most informative features to produce a precise prediction model. It mimics the process of nuclear radiation, which involves the propagation of alpha, beta, and gamma fragments while attacking humans after an explosion. These radiations fly at a very high speed from a high-pressure point (the point of explosion) to a low-pressure point (the standing of the individual place). The proposed algorithm comprises an initial population of the candidate solutions. Furthermore, it computes the gradient descent factor (GDF) for alpha, beta, and gamma fragments when they attack humans. Finally, the optimal solution was achieved by calculating the average of the GDF values over several iterations. The proposed algorithm has advantages such as its ability to deal with convoluted, high-magnitude datasets that are not grounded in local optima, which can be an issue in alternate FS procedures. The primary contributions of this study are as follows.\n\n(i) To develop a novel FSCOA approach by applying the CDO, a metaheuristic algorithm\n\n(ii) To evaluate the performance of the proposed FSCOA-based fault forecasting model on four different classification algorithms, NB, QDA, DT, and KNN, 12 benchmark NASA software defect datasets were used.\n\n(iii) To correlate the performance of the proposed FSCOA approach with several FS approaches such as FSGA, FSPSO, FSDE, and FSACO.\n\n(iv) To validate the statistical implications of the proposed FSCOA approach using Friedman and Holm tests.\n\nThe experimental outcome shows that the proposed FSCOA was better than the other FS approaches examined in most situations and then became the best-performing FS technique for designating the best array of features. The remainder of this paper is organized as follows. Section 2 discusses the existing literature on FS approaches. Segment 3 elaborates on the proposed FSCOA approach and the detailed methodology used in this study. Segment 4 presents the empirical findings and interpretations. Segment 5 outlines the statistical analysis. Finally, Segment 6 presents the conclusions and scope for prospective work.\n\n\nRelated works\n\nDefect prediction in software modules plays a critical role in creating high-quality and reliable software. SDP permits developers to detect and debug defects in software modules during the prior stage of the software advancement process. Unfortunately, conventional SDP processes face several threats, one of which is the curse of dimensionality. The curse of dimensionality indicates the presence of a large number of attributes in a dataset. Many of these attributes do not make any compelling knowledge, and hence, are treated as noise. Feature selection is a potent tool to tackle the challenge of the curse of dimensionality. FS allows developers to establish the best possible set of traits that can enhance the predictive accuracy of the model by discarding irrelevant traits. However, it is imperative to observe that conventional FS procedures are not only expensive but also time-consuming.31 Recently, the application of ML to SDP has gained considerable traction. Several ML-based SDP approaches have been proposed. This section describes some of these studies as follows.\n\nDas et al.32 proposed a novel FS technique called FSGJO based on the Golden Jackal Optimization (GJO) algorithm. The proposed FSGJO technique was employed on four classifiers, namely, KNN, DT, NB, and QDA, using 12 SDP datasets taken from the PROMISE repository. The authors compared the efficacy of the recommended FSGJO technique with alternative FS techniques, namely, FSDE, FSPSO, FSACO, and FSGA. Based on their experimental findings, the authors observed that the proposed FSGJO technique enhanced the prognostic performance of the model. It was also noted that the prospective FSGJO method was exceptional compared to other studied FS techniques in selecting the optimal set of characteristics. However, the authors mentioned that the proposed FSGJO technique needs its parameters to be tuned.\n\nKhalid et al.33 inspected numerous existing ML methods and optimized ML procedures on three publicly accessible NASA datasets. The authors applied PSO and ensemble approaches in their work and scrutinized the results. The experimental findings revealed that the SVM and optimized SVM outperformed the other models in terms of accuracy. However, this study was conducted using a limited number of datasets. Again, the experimental findings cannot be generalized owing to the lack of additional optimization algorithms.\n\nKumar and Das34 enforced GA to supervise learning classifiers such as KNN, DT, and NB. Twelve NASA datasets from the PROMISE archive were used. Using accuracy and failure rate as performance metrics, the performance of the proposed model was assessed. Based on their experimental results, the authors asserted that the suggested FSGA technique improved the behavior of the defect forecast model correlated with the scenario in which the FS was not made. However, in this study, the FS approach used only the GA. The effects of alternative optimization methodologies were not investigated.\n\nThirumoorthy et al.35 suggested a hybrid SDP method based on the TOPSIS and hrbrid Rao algorithms (THRO) to uncover the finest set of traits. The authors used three benchmark NASA SDP datasets to implement their proposed THRO-based FS algorithm on SVM and NB classifiers. The impact of the proposed algorithm was assessed using six metaheuristic FS techniques. The authors noted that the proposed THRO-based FS algorithm enhanced the classification performance of the model and outperformed other studied FS approaches. However, they also noted that this enhanced performance of the proposed method came at the price of increased computational cost.\n\nBatool et al.36 offered a comprehensive and well-organized analysis of the extant literature. They looked at numerous pertinent published publications that employed DM, ML, and DL, among other techniques, for fault prediction. The endeavor was motivated by the need to find answers to research problems stated in the evaluation that might not have been addressed in the works evaluated or called for a different viewpoint. The authors claim that DM and ML techniques, such as DT, NB, SVM, NN, ET, and EA, are frequently employed by SDP. Although they are used less frequently, DL approaches such as CNN, MLP, LSTM, and DNN have also been used by researchers to predict software errors. The authors emphasized the need for larger datasets and the importance of concentrating on using the same methods with combinations of different datasets.\n\nAn SDP architecture based on stacked stacking and heterogeneous FS was proposed by Chen et al.37 The two main objectives of this study were to increase SDP accuracy and optimize software testing resource allocation. The method is divided into three steps: feature selection, model creation with a nested-stacking classifier, and evaluation of the predictive behavior of the model. For the experiments, two datasets were used: Kamei and PROMISE. The investigation included both within-project and large-scale cross-project defect prediction. The model’s behavior was illustrated using the AUC and F1-score evaluation metrics. The initial results showed that for the two sets of software failure datasets, the proposed framework performed better in terms of classification than the baseline models. However, the authors pointed out that nested-stacking is not very effective and that the optimal combination of the baseline model was determined via difficult experiments.\n\nArora and Kaur38 suggested a method that used FS on both origin and destination datasets to assemble a heterogeneous fault prediction (HFP) model to develop an effective forecasting model utilizing supervised training approaches. The writers completed the FS in two phases. They began by selecting features based on their importance. They removed the shared features from the datasets in the next step. An integrated approach was used to select the best characteristics. RFI was used for the FS. According to the suggestion made by Gao et al.,39 the authors selected the top 15% of attributes throughout the FS phase. The proposed framework was applied to two open-source projects, MySQL and Linux, for the supervised ML classifiers, SVM, NB, RF, AdaBoost, DT, and LR. The behavior of the planned model was graded using the Area under the ROC curve (AUC). The authors concluded that the most accurate logistic regression fault prediction is based on the recommended approach. The AUC data demonstrated that the suggested technique accomplished better than the existing Cross Project Fault Prediction (CPFP). However, in this study, other commonly used performance criteria such as accuracy, precision, and recall were not employed to grade the impact of the proposed approach. Once again, only supervised learning algorithms were used in the study, and no optimization algorithms were used.\n\nAnand et al.40 conducted a correlative performance assessment of various FS techniques utilized in SDP. Chi-Square (CS), Correlation Coefficient (CC), Fisher’s Score, Information Gain (IG), Mean Absolute Difference (MAD), and Variance Threshold (VT) are among the filter-based FS approaches used in this investigation. Wrapper-based FS strategies also encourage the use of Backward Feature Elimination (BFE), Exhaustive Feature Elimination (EFE), Forward Feature Elimination (FFE), and Recursive Feature Elimination (RFE) methodologies. RFI and LASSO Regularization are among the embedded FS techniques utilized in this study. The recommended model uses six publicly accessible benchmark NASA datasets for the NB, SVM, DT, and KNN classifiers. The authors used the F1-score, recall, accuracy, and precision as performance evaluation criteria. The authors’ experimental results showed that Fisher’s score behaved more precisely than other FS techniques. However, compared to the no-FS situation, it was found that all FS strategies enhanced the model’s behavior. A drawback of this study is that it neglected to examine the impact of optimization strategies on the FS.\n\nThe dynamic re-ranking approach-based WFS technique was introduced by Balogun et al.41 in response to the exorbitant processing expenses of wrapper-based FS (WFS) methods. The recommended technique was constructed using 25 public domain datasets that were extracted from the NASA, AEEEM, PROMISE, and ReLink archives using classifiers such as DT and NB. The findings of the experiment illustrated that the recommended method reduced computing time and enhanced model performance when executing FS. The suggested method was performed using both the FFS and WFS techniques, which is a disadvantage. FFS has variable performance across datasets and classifiers, whereas WFS suffers from stagnation of local optima and high computing costs. Once more, only two supervised classifiers were examined in this work: SVM and K-NN, two more well-known classifiers, were not examined.\n\nBalogun et al.42 proposed an inventive hybrid multifilter wrapper FS arrangement based on rank aggregation to select critical features to address the aforementioned shortcomings. The recommended course of action was implemented in two steps. In the first lap, a multifilter FS mechanism based on rank aggregation was used, which combined the separate rank lists from multifilter methods to build an original, dependable, and non-disjoint rank list. This resolves the filter rank choice issue. In the second lap, the upgraded wrapper FS approach, which was predicated on dynamic re-ranking, was used once more to preprocess the accumulated ranked attributes. The competence of the recommended method is illustrated using NB and DT classifiers on benchmark software fault datasets. The tests used accuracy, area under the curve (AUC), and F-measure values as evaluation criteria. The authors used their findings to concentrate on the issues of filter rank choice and local optima stagnation in HFS, demonstrating the suggested method’s ingenuity in selecting the best characteristics while enduring or boosting the impact of the forecasting models. They concluded that applying the recommended technique significantly improves the behavior of the model. However, the model was limited to only two classifiers to achieve satisfactory results. Consequently, the potential of extrapolating the results to alternative classifiers has not been explored.\n\nAlsghaier and Akour43 presented an SDP model by fusing the GA, SVM, and PSO. Three stages were implemented: GA-SVM for GA integration, PSO-SVM for PSO integration, and GAPSO_SVM for the reciprocal iteration-based integration of GA-SVM and PSO-SVM. During the experimentation phase, 24 benchmark SDP datasets (12 NASA MDP and 12 open-source Java applications) were subjected to the proposed model using the SVM classifier. To validate the theoretical model, experiments were conducted using the WEKA Tool and MATLAB 2015. The impact of the developed approach was assessed using evaluation metrics, such as accuracy, recall, precision, F-measure, specificity, error rate, and standard deviation. The results of the experiment showed that combining the GA with SVM and PSO had a beneficial effect on the model and enhanced its performance when applied to both small- and large-scale datasets. However, the precision metric was insufficient to appraise the suggested procedures.\n\nAlsghaier and Akour44 built on their earlier work43 by combining GA, SVM, and Whale Optimization Algorithm (WOA) to forecast defects. The remainder of the experimental configuration remained the same as in a previous study.43 Through experimental data, the researchers discovered that the behavior of the defect forecast model was improved for both large-scale and small-scale datasets when the GA was integrated with SVM and WOA. For the datasets under study, WA-SVM performed more accurately than GAWA-SVM, and GAWA-SVM produced the worst outcomes. Again, the proposed method outperformed SVM for the NASA MDP and open-source Java projects in terms of SD scores. This illustrates how combining SVM with optimization techniques enhances the prediction performance. The NASA, GA-SVM, and GAWA-SVM datasets produced the best outcome in terms of specificity. This proved that the GA-SVM and GAWA-SVM procedures are appropriate for software defect predictions when enforced on an enormous dataset.\n\nBalogun et al.45 used NASA datasets from the PROMISE archives to thoroughly evaluate the FSS algorithms on NB, DT, LR, and KNN. Their findings imply that the studied FS techniques enhanced the performance of the system. Information Gain, one of the FFR techniques, demonstrated the best results. Consistency Feature Subset Selection (CFSS), which is based on the Best First Search in FSS methods, has the greatest impact on forecasting models. However, there were variations in the performances of the classifiers’ and datasets.’ Scientists have also found that models constructed using FFR-based techniques are more stable than those constructed using FSS-based approaches. This study focused only on FFS procedures, and the effects of the WFS techniques were not investigated in detail.\n\nAll of the previously stated FS approaches, whether supervised or unsupervised, have disadvantages that significantly impact the model’s performance, including (i) high cost, (ii) entrapment in local optima, (iii) low convergence rate, and (iv) fine-tuning of excessively many parameters. The primary drawback of the previously stated FS techniques is the need to modify the regulating parameters accurately while choosing ideal characteristics. These shortcomings motivated us to propose a novel FS technique (FSCOA) that draws inspiration from the Chernobyl Disaster Optimizer (CDO). The 1986 nuclear reactor core outburst in Chernobyl served as an impetus for the development of the CDO meta-heuristic algorithm. The process of nuclear radiation, in which alpha, beta, and gamma fragments propagate and damage humans following an explosion, is replicated by CDO. From the high-pressure point (explosion site) to the low-pressure point (individual standing standing), the above-mentioned radiations travel at an extremely rapid speed. The algorithm comprises an initial population of potential solutions. Moreover, it calculates the alpha, beta, and gamma fragment gradient descent factors (GDF) during human attacks. Determining the average of these GDF values over a number of iterations yields the best result.\n\nFeature selection determines the crucial attributes that have the greatest impact on the desired variable, which helps increase machine learning model accuracy, reduce computing costs, and reduce the risk of overfitting. The mechanism of selecting the best features consists of three steps: (1) creating a set of subgroups of attributes; (2) assessing and comparing the adequacy of these subgroups to determine which subgroup is the best or until the abort standards are met; and (3) computing the outcome using only the best features. It is difficult to determine which parts to classify. The 1986 Chernobyl nuclear reactor catastrophe46 is recognized as one of the lowest nuclear disasters in the modern human past, in terms of both cost and casualties. Inspired by the Chernobyl nuclear reactor core eruption, the Chernobyl Disaster Optimization (CDO)30 is a meta-heuristic optimization technique. In order to choose the most appropriate subset of characteristics for classification, a novel FS approach using Chernobyl Optimization Algorithm (FSCOA) is therefore proposed in order to address the aforementioned problem. Figure 153 shows the blueprint for the proposed FSCOA method.\n\nFirst, the selection of the relevant SDP datasets is crucial. Following the selection of the datasets, an in-depth examination of the datasets was carried out to determine any missing, inconsistent, or categorical data. It became apparent that there were no missing data in the datasets. Nevertheless, a few datasets contained categorical data. The data were categorized prior to the generation of the numbers. Furthermore, the original feature value, which originally ranged from 0 to 1, was normalized. Subsequently, an 80:20 split between the training and testing datasets was created for each normalized dataset. To develop and investigate the model, the two preeminent criteria are the population size and maximum number of iterations. Higher values will improve the performance of the model, but they also lengthen the computation time. In this study, the population size and maximum tally of the iterations were set to 30 and 200, respectively. By applying the recommended FSCOA methodology, four supervised learning classifiers (DT, KNN, NB, and QDA) were used to construct the model using the optimal features that were chosen. The best predictive classifier was then determined by comparing the accuracy of the proposed FSCOA approach with that of the other FS models under study.\n\nThis study suggests a novel FSCOA technique to select the first-rate subgroup of attributes for categorization. The primary intent of the suggested technique is to identify the best attribute combination that will lower the fitness of the model. Figure 253 shows a complete flow diagram of the proposed FSCOA technique.\n\nInitializing the criteria, such as population size (M), problem dimension (F), lower bound (LowBound), and upper bound (UppBound), is the first step of the procedure. Subsequently, a random binary population of M fragments with dimension F, where Z=[Z1,Z2,Z3,…,ZM] is the total number of features. Zi=[Z1,Z2,Z3,…,ZM] is the ith fragment location in the F dimension feature space, where i=1,2,3,…,M is the specimen proportion, and Zi,f is the ith fragment standing of the fth trait of the population. Many classification techniques, including DT, KNN, NB, and QDA for fitness (error) computation, have been considered for the examination of randomly selected characteristics. The FS algorithm aims to select the best subset of ideal features that may reduce the fitness of the learning algorithm. The error (Errit) is estimated as the disparity between the estimated outcome (EOit) and actual outcome (AOit). Eq. (1) can be used to describe this phenomenon.\n\nBy distributing the aggregate of the total errors by the entire count of instances in the testing data, the fitness (FitValueit) of the learning algorithm was calculated. This is characterized by Eq. (2).\n\nHere, i=1,2,…..,p and p represent the tally of the instances in the trial data, t represents the current iteration.\n\nThe transfer function depicted in Eq. (3) was employed to transform the initial fragment standings into a binary equivalent.\n\nThe proposed FSCOA approach employs the CDO algorithm to determine the optimal features for a given dataset. In CDO, different types of emissions are released from the nuclei as a result of radioactivity caused by nuclear instability. The most prevalent types of these emissions are alpha, beta, and gamma fragments. These fragments, which are very dangerous to people, fly from a high-pressure point (the point of explosion) to a low-pressure point (the standing of individual standing). When a human is attached to a CDO following a nuclear explosion, it simulates the effects of radioactive decay. The primary processes of nuclear explosion and human attachment require the use of gamma, beta, and alpha fragments. Humans were most likely to be on foot when they were attacked. Human walking speed can be enhanced, and it can be estimated to be between 0 and 3 miles per hour.49 Based on this, Eq. (4) can be used to model linear reduction at this speed.\n\nAlpha fragment\n\nThe gradient descent factor (GDFα) of the alpha fragment while threatening humans can be computed using Eq. (5).\n\nHere, POSα(t) is the prevailing standing of alpha fragments, PROPα represents the dispersion of alpha fragments and can be calculated using Eq. (6); Dα is the discrepancy between the individual standing and standing of alpha fragments, which can be determined using Eq. (8).\n\nHere, rad is a random value between 0 and 1, Speedα is the speed of alpha fragments that can be in the range of 1–16,000 kmps. This can be normalized using Eq. (7)\n\nHere, Areaγ is the propagation area of gamma fragments that can be calculated as π∗rad∗rad where rad is a random value between 0 and 1, AvgT is the average of the total standings that can be determined using Eq. (17).\n\nBeta fragment\n\nEq. (9) can be used to determine the gradient descent factor (GDFβ) of a beta fragment assaulting a human.\n\nHere, POSβ(t) is the current standing of beta fragments; PROPβ represents the propagation of beta fragments and can be calculated using Eq. (10); Dβ is the discrepancy between the human standing and the beta fragment’s standing, which can be determined using Eq. (12).\n\nHere, rad is a random value between 0 and 1, Speedβ is the speed of beta fragments that can be in the range of 1–270,000 kmps. This can be normalized using Eq. (11)\n\nHere, Areaβ is the propagation area of the beta fragment and can be calculated as π∗rad∗rad where rad is a random value between 0 and 1. AvgT is the average of the total standings that can be computed using Eq. (17).\n\nGamma fragments\n\nThe gradient descent factor (GDFγ) of the gamma fragment while making an assault on humans can be computed using Eq. (13).\n\nHere, POSγ(t) is the prevailing standing of gamma fragments, PROPγ represents the dispersion of gamma fragments and can be calculated using Eq. (14); Dγ is the discrepancy between the standing of the human and the standing of gamma fragments, which can be determined using Eq. (16).\n\nHere, rad is a random value between 0 and 1, Speedγ is the speed of the gamma fragment in the range of 1 to 300,000 kmps. This can be normalized using Eq. (15)\n\nHere, Areaγ is the propagation area of gamma fragments that can be calculated as π∗rad∗rad where rad is a random value between 0 and 1, AvgT is the average of the total standings that can be determined using Eq. (17).\n\nFinally, Algorithm 1 provides a summary of the entire proposed FSCOA process.\n\n\n\n1. Initialize Populace Size (M), Dimension (F), Lower Bound (LowBound), UpperBound (UppBound), Maximum Iteration (max_iter)\n\n2. Generate the binary feature subset Zi randomly\n\n3. Initialize the alpha (POSα), beta (POSβ), and gamma (POSγ) standings\n\n4. while (t<max_iter) do{\n\n5. for i=1: M do\n\n6. forj=1toF do\n\n7. The values of the initial standing of the fragments are converted into their corresponding binary values using Eq. (3).\n\n8. Compute the fitness value (FitValue)for the alpha, beta, and gamma fragments using Eq. (2)\n\n9. if (FitValue<∝score)\n\n10. ∝Score=FitValue\n\n11. Update POSα\n\n12. endif\n\n13. if (FitValue>∝score)and(FitValue<βscore)\n\n14. βScore=FitValue\n\n15. Update POSβ\n\n16. endif\n\n17. if (FitValue>∝score)and(FitValue>βscore)and(FitValue<γscore)\n\n18. γScore=FitValue\n\n19. Update POSγ\n\n20. endif\n\n21. end for\n\n22. end for\n\n23. Compute human walking speed (WalkSpeedhuman) using Eq. (4)\n\n24. Compute the speed of alpha (Speedα), beta (Speedβ), and gamma (Speedγ) fragments using Eq. (7), Eq. (11), and Eq. (15), respectively.\n\n25. for i=1: M do\n\n26. for j=1: F do\n\n27. Determine GDFα using Eq. (5)\n\n28. Determine GDFβ using Eq. (9)\n\n29. Determine GDFγ using Eq. (13)\n\n30. Update Zi using average of total standings using Eq. (17)\n\n31. end for\n\n32. end for\n\n33. t=t+1\n\n34. } //end of while loop\n\n35. Return finest solution, Zi\n\n36. end procedure\n\nThis section deliberates on the empirical findings of this research. The persuasiveness of the proposed FSCOA approach was graded by employing 12 publicly benchmarked NASA software defect datasets extracted from the PROMISE archive.48 KC1, KC3, CM1, JM1, MC1, MC2, MW1, PC1, PC2, PC3, PC4, and PC5 were the datasets. First, an in-depth examination of the datasets was performed to identify missing, inconsistent, and categorical data. It became apparent that there were no missing data in the datasets. Nevertheless, a few datasets contained categorical data. The data were categorized prior to the generation of the numbers. Again, we noticed that the datasets comprised of continuous data. The datasets were altered using the min–max normalization method49 with the goal of overcoming this problem. The original feature value, which originally ranged from zero to one, was transformed using this technique. Subsequently, an 80:20 split between the training and testing datasets was created for each normalized dataset. Extensive information regarding the datasets enforced in this exploration is shown in Table 1.\n\nThe configuration of the computer on which the experiments were administered was as follows: Intel Core i5-6200 CPU with clock rate 2.40 GHz and 8 GB RAM. The aforementioned techniques were employed in a Python 3 environment using the Jupyter notebook. First, the input dataset was uploaded using Pandas. The datasets were altered using the min-max normalization method.49 Using train_test_split from sklearn.model_selection, the dataset was partitioned into training and testing datasets at a ratio of 80:20. Populace size and the highest number of iterations were the two major criteria for developing and validating the model. The model will provide superior outcomes with higher values, but it will also increase computing time. In this investigation, the population size and highest number of iterations were set to 30 and 200, respectively. Four supervised learning classifiers, DT, KNN, NB, and QDA, were used to evaluate five FS approaches: FSDE, FSPSO, FSGA, FSACO, and the suggested FSCOA. The fitness error plots for the suggested FSCOA approach and other FS strategies utilizing the examined classifiers, DT, KNN, NB, and QDA, were obtained using matplotlib.pyplot. In this study, accuracy, a frequently applied performance indicator metric, was used for assessment purposes. Accuracy can be expressed as a simple proportion of the total instances of instances that were correctly classified. Eq. (18) is used to calculate it from the confusion matrix, as follows:\n\nHere, TP, TN, FP, and FN represent true positives, true negatives, false positives, and false negatives, respectively.\n\nThe performance of the recommended FSCOA algorithm is evaluated against several FS procedures such as FSDE, FSPSO, FSGA, and FSACO in terms of classification accuracy and the count of selected attributes on 12 datasets studied in this research work. Because of the stochastic character of the previously mentioned techniques, we carried out ten runs of the trials to ensure that the performance of each procedure remained persistent, with an initial random population. The median accuracy of the proposed FSCOA, along with the other studied FS approaches, is listed in Table 2.54\n\nThe median accuracy of several classifiers forced on diverse datasets, both with and without feature selection, is shown in the table. The table also displays the average tally of the attributes selected by the respective FS approach. The classifiers were evaluated using a range of datasets and previously discussed FS techniques. The experimental findings showed that the suggested FSCOA technique exceeded the other studied FS procedures in the majority of instances. For the majority of the datasets, with the exception of KC1, KC3, JM1, and MC1, the suggested FSCOA performed best when combined with KNN. With the exception of KC1, MC1, and CM1, the bulk of the datasets showed that the recommended FSCOA worked best when paired with NB. The majority of the datasets demonstrated that the suggested FSCOA performed best when combined with QDA, with the exception of KC3, MW1, and PC5. With the exception of JM1, the majority of the datasets showed that the previously researched FS approaches outperformed the suggested FSCOA strategy when used in conjunction with DT. Similarly, applying the proposed FSCOA technique to the JM1 dataset with all the analyzed classifiers, except KNN, yielded the highest accuracy. Furthermore, the bulk of the datasets yielded the best accuracy for all examined classifiers, with the exception of DT. It is crucial to remember that the suggested FSCOA technique could only provide the best prediction using QDA and NB classifiers for the KC1 and KC3 datasets, respectively.\n\nFigures 3 through 653 display, respectively, the fitness error plots for the suggested FSCOA approach and other FS strategies utilizing the examined classifiers DT, KNN, NB, and QDA. Error plots of all 12 datasets are included in each graph. The error plots show that in most cases, the error plot of the suggested FSCOA is smaller than those of the other FS approaches employed in this investigation. Furthermore, the error plot of the suggested FSCOA methodology matches that of the various existing FS methods. However, the error plot of the suggested FSCOA approach exceeds that of the other FS techniques that have been evaluated under certain circumstances.\n\nFigure 353 shows that in most datasets, the fitness error plot of the proposed FSCOA approach using the DT classifier is smaller than that of the other FS models, with the exception of CM1, MC2, and KC3. However, for the KC3 dataset, the error plot overlapped with that of the FSDE after 190 iterations. The error plot for the CM1 dataset is located above the FSDE and FSACO. Furthermore, the plot for the MC2 dataset is above the FSDE.\n\nThe fitness error plot of the proposed FSCOA approach with the KNN classifier is lower than that of the previous FS models for most datasets, as shown in Figure 4,53 with the exception of KC1, KC3, MC1, and PC2. The error plot after 115 iterations corresponds to FSDE and FSACO for the PC2 dataset, but it is above the FSACO model for the KC1, KC3, and MC1 datasets.\n\nAs shown in Figure 5,53 the fitness error plot of the suggested FSCOA technique with the NB classifier was lower in the majority of datasets than that of the prior FS models, with the exception of CM1, KC1, MC1, PC2, MC2, and PC3. After 75 iterations, the error plot for the MC2 dataset matches that of FSACO. For the PC3 dataset, the error plot of the suggested FSCOA method matches that of FSACO after 175 iterations. However, for datasets CM1, KC1, and MC1, the error plot was above that of the FSACO model. Moreover, for PC2 and KC1, the plot was above the FSDE.\n\nFigure 653 illustrates that for most datasets (except CM1, JM1, KC3, MW1, PC2, and PC3), the fitness error plot of the proposed FSCOA approach with the NB classifier is smaller than that of the previous FS models. Following 180 iterations, the CM1 dataset’s error plot aligns with that of the FSACO dataset. The error plot of the proposed FSCOA algorithm matches that of FSGA after 175 iterations for the JM1 dataset. The error plot is above that of the FSACO model for datasets MW1, PC2, and PC3. The graphic is also above FSGA and FSDE for the KC3 dataset. The error plot of the proposed FSCOA approach is above that of FSGA for the PC3 dataset.\n\nThe FS algorithms employed in this study use several hyperparameters. For 200 iterations, an examination was performed with a population size of 30. The crossover rate (CR) in FSGA and FSDE was maintained at 0.8 and 0.9, respectively. For FSGA, the mutation rate (MR) is 0.01. In the FSDE, the scaling factor (SF) was set to 0.8. In FSPSO, the maximum inertia weight (IWmax) and the minimum inertia weight (IWmin) have been fixed as 0.9 and 0.4 accordingly. Two were chosen as the acceleration factors. The fixed values of alpha (α), rho (ρ), and beta (β) in FSACO were 1, 0.2, and 0.1, respectively. The governing criterion speeds for alpha (Speedα), beta (Speedβ), and gamma (Speedγ) were adjusted appropriately for FSCOA using the Rand function. In addition, the radiation propagation radius (rad) was similarly fixed between 0 and 1 using the rand function.\n\nThis section provides extensive statistical scrutiny of the empirical findings of this work. Statistical analysis50 is a popular method for quantifying, examining, evaluating, and drawing conclusions from data. Tests classified as parametric or non-parametric were the two breeds used for statistical analysis. A type of statistical analysis, known as parametric statistical testing, assumes that the data under study conform to a specific probability distribution, most frequently a normal distribution. Several assumptions, such as the independence of observations, homogeneity of variance, and normality, must hold true to employ parametric tests. Ensuring that the assumptions are met is essential when conducting a parametric test, because failing to do so may result in erroneous results and invalid conclusions. Therefore, it is crucial to confirm these hypotheses in advance and, if necessary, to use non-parametric validation. A type of statistical scrutiny known as non-parametric statistical testing does not depend on a specific probability distribution hypothesis for the data under study. Non-parametric validations, on the other hand, are more broadly applicable and resilient to assumption violations than parametric tests because they depend on the hierarchy or placement of the data. However, when the presumptions of the parametric validations are satisfied, they might be less effective than the latter. It is essential to choose a statistical validation suitable for an exploration topic and the properties of the data being examined. In this study, the Friedman Test,51 a non-parametric rank-based test, has been carefully considered. Based on the effectiveness of the classification, each model associated with the trial was ranked according to the Friedman test. The lowest count correlates with the greatest slot and the largest count correlates with the smallest slot.\n\nTo begin with, Eq. (19) was employed to determine the average rank (AverageRankModels) of all graded models (FSDE, FSPSO, FSGA, FSACO, FSCOA, and Without FS), in addition to a number of classification models (KNN, DT, NB, and QDA). Table 354 presents an illustration these findings.\n\nTable 454 summarizes the findings of grading the median of all ranks of diversified amalgamation setups (FSDE, FSPSO, FSGA, FSACO, FSCOA, and Without FS) for all the datasets used in Eq. (20).\n\nThe following are the average ranks for all the correlated configurations included in this observation. {AvgRank1=1.75,AvgRank2=2.48,AvgRank3=2.52,AvgRank4=2.92,AvgRank5=4.75,AvgRank6=5.8}. The median ranks of the models were employed to gauge the XF statistics, referred to as XF2 using Eq. (21) and a presented value of 23.29.\n\nTwelve datasets (B=12) and six models (A=6) were considered in this experiment. The Friedman statistic (FF) value was computed using Eq. (22) using (B − 1) and XF2.\n\nThe value of (FF) estimated to be 6.978. The critical value was determined as 2.383 by employing the degrees of freedom as (6 − 1 = 5) × (12 − 1 = 11) and (6 − 1 = 5), with α = 0.05, as the significance level. Given that the critical value of 2.383 is smaller than that of the Friedman statistic (FF = 6.978), the null hypothesis is rejected. It also determines whether to adopt an alternate theory. This implies that two or more configurations are distinct from one another. The Holm method is usually employed to investigate the Post Hoc test after the null hypothesis is jilted and the substitute hypothesis is endorsed. By employing the Holm technique, the pvalue and z−value were applied to assess how well each distinct model performed relative to the other models.52 Eq. (23) was used to obtain the value of z. The z−value and normal distribution table were used to calculate the value of p.\n\nIn this experiment, the terminologies B, A, and z represent the number of datasets, number of configurations employed in this investigation, and value of z, respectively. The terms AvgRank(i) and AvgRank(j) represents the average rank of ith and jth model, respectively. The pvalue, z−value, and α/(A−i) of the recommended configurations were compared, and Table 5 summarizes the findings. For this particular instance, we set the significance level, α, at 0.05.\n\nTable 554 illustrates that in most cases, the p-value is lower than or equivalent to the value of α/(A−i) with the exception of the FSCOA and FSGA models and FSCOA and FSACO models. It resolves that the FSCOA model is statistically noteworthy and attains a superior dossier when matched to other configurations, excluding the FSGA and FSACO models. However, there was no statistically significant variation in the performances of these models.\n\n\nConclusion\n\nThis study proposed a novel FS approach called FSCOA, which applies a meta-heuristic procedure called CDO. The proposed FSCOA technique exhibits nuclear core reactor disruption to determine the best set of attributes by carefully discarding irrelevant or insignificant ones. This study investigates the impact of the proposed FSCOA approach on 12 publicly available NASA datasets by employing four widely used classifiers (DT, KNN, NB, and QDA). One elementary purpose was to correlate the predictive behavior of the proposed FSCOA approach with other existing FS techniques, namely, FSDE, FSPSO, FSACO, and FSGA. The Friedman test was used to test the statistical validity of the proposed FSCOA method. The test outcome showed that at least two models were significantly different, leading to the repudiation of the null hypothesis. This necessitates the use of the Holm test. The experimental findings suggested that the proposed FSCOA approach demonstrated higher performance when selecting the optimal set of features correlated to the studied FS procedures. However, the behavior of the proposed FSCOA approach may vary across different datasets and classifiers. In the future, we aim to expand the scope of this research by employing real-world project datasets. We also look forward to investigating the efficiency of the suggested FSCOA approach by increasing the number and variety of classifiers, especially ensemble classifiers, and employing more optimization algorithms for feature selection.", "appendix": "Data availability\n\nFigshare: Anand, Kunal (2024). Dataset 1: Zip file containing the underlying data of the presented methods and results in jpeg files. figshare. Figure. https://doi.org/10.6084/m9.figshare.25681782.v1. 53\n\nThis dataset contains the following underlying data:\n\n• Figure 1. Blueprint of the suggested FSCOA methodology.jpg\n\n• Figure 2. Flow-diagram of recommended FSCOA approach.jpg\n\n• Figure 3. DT fitness error plot.jpg\n\n• Figure 4. KNN fitness error plot.jpg\n\n• Figure 5. NB fitness error plot.jpg\n\n• Figure 6. QDA fitness error plot.jpg\n\nFigshare: Anand, Kunal (2024). Dataset 2 Zip file containing underlying data of the presented results in csv files. figshare. Dataset. https://doi.org/10.6084/m9.figshare.25683600.v1 54\n\nThis dataset contains the following underlying data:\n\n• Table 2 Accuracy percentage and number of features selected.csv\n\n• Table 3 Average rank of all FS algorithms for twelve NASA datasets.csv\n\n• Table 4. AvgRank of all FS configurations.csv\\\n\n• Table 5. Holm procedure.csv\n\nThe data are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgments\n\nThe authors would like to express their gratitude to the Kalinga Institute of Industrial Technology, Bhubaneswar, for funding (KIIT-DU/309/24) for publication of this article.\n\n\nReferences\n\nMall R: Fundamentals of software engineering. PHI Learning Pvt. Ltd; 2018.\n\nPrasad D: Automated software testing: foundations, applications and challenges. Jena AK, Das H, Mohapatra DP, editors. 2020; pp. 1–165. Springer;\n\nAnand K, Jena AK: Software Defect Prediction: An ML Approach-Based Comprehensive Study. 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Publisher Full Text\n\nGarcía S, Fernández A, Luengo J, et al.: Advanced nonparametric tests for multiple comparisons in the design of experiments in computational intelligence and data mining: experimental analysis of power. Inf. Sci. 2010; 180(10): 2044–2064. Publisher Full Text\n\nAnand K: Dataset 1: Zip file containing the underlying data of the presented methods and results in jpeg files. figshare. Figure. 2024. Publisher Full Text\n\nAnand K: Dataset 2 Zip file containing underlying data of the presented results in csv files. Dataset. figshare. 2024. Publisher Full Text" }	[ { "id": "309141", "date": "14 Aug 2024", "name": "Shabib Aftab", "expertise": [ "Reviewer Expertise software defect prediction" ], "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\nAbstract: The abstract should offer a succinct overview of the research paper, covering the research problem, objectives, methodology, key findings, and conclusions. It must be clear and informative, providing readers with an understanding of the study's importance. You should precisely highlight the main contributions, including the datasets utilized and the accuracy achieved by your proposed system.\nIntroduction: The introduction section should conclude with a brief outline of the structure for the remainder of the paper . Literature Review: A distinct section dedicated to the Literature Review should be included in the paper in the form of a table.\nResults: The results section should clearly and systematically present the study's findings. It should provide detailed information, including a thorough comparison with other related techniques. You also have to explore other measures as well in the comparison other techniques.\nConclusion: This section should offer a comprehensive summary of the research and include a detailed discussion of future research directions.\nEnglish: The paper should be reviewed to ensure the English language is free of grammatical errors.\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? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "12871", "date": "17 Dec 2024", "name": "Kunal Anand", "role": "Author Response", "response": "Abstract: The abstract should offer a succinct overview of the research paper, covering the research problem, objectives, methodology, key findings, and conclusions. It must be clear and informative, providing readers with an understanding of the study's importance. You should precisely highlight the main contributions, including the datasets utilized and the accuracy achieved by your proposed system. Response: We are deeply appreciative of the esteemed reviewer's perceptive comments on our work. In this context, we would like to submit that, as suggested by the esteemed reviewer, the abstract has been reworked in the revised manuscript. Introduction: The introduction section should conclude with a brief outline of the structure for the remainder of the paper. Response: We sincerely appreciate the insightful feedback on our efforts provided by the excellent reviewer. In light of this, we would like to state that the last paragraph of the introduction section outlines the structure for the remainder of the paper. . Literature Review: A distinct section dedicated to the Literature Review should be included in the paper in the form of a table. Response: The excellent reviewer provided us with insightful feedback of our work, which we much appreciate. This is a really smart idea from the respected critic. As desired by the respected reviewer, we would like to submit that the literature review has been appended as a table at the conclusion of the related work section. Results: The results section should clearly and systematically present the study's findings. It should provide detailed information, including a thorough comparison with other related techniques. You also have to explore other measures as well in the comparison other techniques. Response: We are deeply appreciative of the distinguished reviewer's insightful critique of our work. It is a very wise suggestion from the renowned critic. We have incorporated in the amended text the notable distinctions between the suggested approach and the other related FS techniques employed in this work, as suggested by the excellent reviewer. Furthermore, a distinct section titled \"Threats to validity\" in the updated manuscript addresses the ramifications of the suggested FSCOA strategy. In addition to investigating additional measurements, our goal for the future is to broaden the scope of the suggested work by comparing it with more FS techniques. Conclusion: This section should offer a comprehensive summary of the research and include a detailed discussion of future research directions. Response: We greatly appreciate the insightful criticism of our work provided by the esteemed reviewer. The renowned reviewer has offered a really astute suggestion. In light of this, we would like to submit that the conclusion section has been altered in the revised manuscript in accordance with the distinguished reviewer's suggestions. English: The paper should be reviewed to ensure the English language is free of grammatical errors. Response: To make sure the English is correct, the paper has been carefully examined and modified accordingly." } ] }, { "id": "319612", "date": "17 Sep 2024", "name": "Ahmed Abdu", "expertise": [ "Reviewer Expertise Software engineering", "software defect prediction", "deep 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\nThis paper presents an approach to addressing the challenges of software defect prediction (SDP) through the innovative use of the Chornobyl Optimization Algorithm (COA) for feature selection. The problem of handling high-dimensional datasets in SDP is well-recognized, and the authors' focus on reducing these dimensions through feature selection techniques is highly relevant to improving predictive performance. The abstract provides a clear overview of the limitations of existing meta-heuristic algorithms, positioning the COA as a promising alternative. However, it requires some major revisions to fully meet the expectations for clarity and depth in the current research context. Addressing these revisions will help strengthen the study's overall contribution and impact: 1. The abstract should clarify the specific contribution of the Chernobyl Optimization Algorithm (FSCOA) in comparison to existing methods. The abstract mentions drawbacks of other meta-heuristic algorithms but doesn't specify how FSCOA addresses these issues in a distinct or innovative way. Adding a brief statement on the unique advantages of FSCOA (e.g., better performance, reduced computational cost, or improved convergence) would enhance clarity and highlight the novelty of the study.\n2. The introduction section outlines the research focus, but the main contributions of the paper should be revised to clearly delineate three distinct and understandable contributions. Clarifying these contributions will help readers better grasp the novel aspects of your work and its significance in advancing the field.\n3. The authors should consider extending the Related Work section by including more recent references to ensure the study is up-to-date with the latest advancements in software defect prediction. Specifically, citing the following studies would strengthen the discussion of deep learning and feature selection approaches in software defect prediction:\nAbdu, A., Zhai, Z., Abdo, H. A., & Algabri, R. (2024). Software Defect Prediction Based on Deep Representation Learning of Source Code From Contextual Syntax and Semantic Graph. IEEE Transactions on Reliability. Abdu, A., Zhai, Z., Abdo, H. A., Algabri, R., & Lee, S. (2023). Graph-Based Feature Learning for Cross-Project Software Defect Prediction. Computers, Materials and Continua, 77(1).\n\nAbdu, A., Zhai, Z., Abdo, H. A., Algabri, R., Al-Masni, M. A., Muhammad, M. S., & Gu, Y. H. (2024). Semantic and traditional feature fusion for software defect prediction using hybrid deep learning model. Scientific Reports, 14(1), 14771. Abdu, A., Zhai, Z., Algabri, R., Abdo, H. A., Hamad, K., & Al-antari, M. A. (2022). Deep learning-based software defect prediction via semantic key features of source code—systematic survey. Mathematics, 10(17), 3120.\nBy incorporating these studies, the authors will enrich the context of their research and ensure that their work is grounded in the most current literature on the topic.\n4. The results discussion is currently lacking depth and does not sufficiently highlight the strengths of the proposed method. The comparison with other approaches is very limited, which weakens the impact of your findings. I recommend expanding the explanation and discussion of the results, focusing on:\nHow your proposed method outperforms or differs from the baseline methods. Highlighting key insights and practical implications of your findings.\nBy thoroughly discussing these aspects, you will better demonstrate the novelty and effectiveness of your method, making the results more compelling for the readers.\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? Yes", "responses": [ { "c_id": "12872", "date": "17 Dec 2024", "name": "Kunal Anand", "role": "Author Response", "response": "The abstract should clarify the specific contribution of the Chernobyl Optimization Algorithm (FSCOA) in comparison to existing methods. The abstract mentions drawbacks of other meta-heuristic algorithms but doesn't specify how FSCOA addresses these issues in a distinct or innovative way. Adding a brief statement on the unique advantages of FSCOA (e.g., better performance, reduced computational cost, or improved convergence) would enhance clarity and highlight the novelty of the study. Response: We really appreciate the insightful reviewer's feedback on our work, and in light of this, we would like to propose that the abstract has been amended in the revised article in accordance with the reviewer's suggestions. The introduction section outlines the research focus, but the main contributions of the paper should be revised to clearly delineate three distinct and understandable contributions. Clarifying these contributions will help readers better grasp the novel aspects of your work and its significance in advancing the field. Response: Our sincere gratitude goes out to the renowned reviewer for his insightful feedback on our work. This is an extremely astute suggestion from the esteemed critic. As suggested by the esteemed reviewer, the major contributions of the paper have been reworked in the revised manuscript. 3. The authors should consider extending the Related Work section by including more recent references to ensure the study is up-to-date with the latest advancements in software defect prediction. Specifically, citing the following studies would strengthen the discussion of deep learning and feature selection approaches in software defect prediction: Abdu, A., Zhai, Z., Abdo, H. A., & Algabri, R. (2024). Software Defect Prediction Based on Deep Representation Learning of Source Code From Contextual Syntax and Semantic Graph. IEEE Transactions on Reliability. Abdu, A., Zhai, Z., Abdo, H. A., Algabri, R., & Lee, S. (2023). Graph-Based Feature Learning for Cross-Project Software Defect Prediction. Computers, Materials and Continua, 77(1). Abdu, A., Zhai, Z., Abdo, H. A., Algabri, R., Al-Masni, M. A., Muhammad, M. S., & Gu, Y. H. (2024). Semantic and traditional feature fusion for software defect prediction using hybrid deep learning model. Scientific Reports, 14(1), 14771. Abdu, A., Zhai, Z., Algabri, R., Abdo, H. A., Hamad, K., & Al-antari, M. A. (2022). Deep learning-based software defect prediction via semantic key features of source code—systematic survey. Mathematics, 10(17), 3120. By incorporating these studies, the authors will enrich the context of their research and ensure that their work is grounded in the most current literature on the topic. Response: We sincerely thank the distinguished critic for his insightful feedback on our work. The renowned reviewer has made a very wise recommendation. In accordance with the most recent developments in software defect prediction, a few recent works have been added to the related work area as suggested by the esteemed reviewer. 4. The results discussion is currently lacking depth and does not sufficiently highlight the strengths of the proposed method. The comparison with other approaches is very limited, which weakens the impact of your findings. I recommend expanding the explanation and discussion of the results, focusing on: How your proposed method outperforms or differs from the baseline methods. Highlighting key insights and practical implications of your findings. By thoroughly discussing these aspects, you will better demonstrate the novelty and effectiveness of your method, making the results more compelling for the readers. Response: We sincerely thank the distinguished reviewer for his insightful critique of our work. The recommendation from the renowned reviewer is really astute. As suggested by the respected reviewer, we have incorporated in the updated text the notable distinctions between the suggested method and the other baseline FS approaches employed in this study. Additionally, the consequences of the suggested procedures are incorporated into the updated manuscript as a distinct section called \"Threats to validity.\"" } ] }, { "id": "319618", "date": "20 Sep 2024", "name": "Francis Palma", "expertise": [ "Reviewer Expertise Software engineering", "software quality", "software maintenance and evolution." ], "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 is sound and complete regarding technical contributions. However, some clarifications and methodological discussions are missing. Below are my more detailed comments.\n1. In the Abstract, the authors said, \"To overcome the above shortcomings, this study aims to develop an innovative FS technique, namely, the Chernobyl Optimization Algorithm (FSCOA), to unwrap the most informative features\" -> I assume the authors are not developing COA? Rather applying or using it? This phrase needs to be revised.\n2. The first line in the Methods part of the Abstract: \"attacking\" -> \"attaching\".\n3. The last paragraph of the Introduction provides the outline of the article. However, later, I found no section numbers or identifiers. Shouldn't the sections be numbered?\n4. The Related Works section discusses 11 studies. I would expect a table providing a summary and comparison among these studies, including your own study (this one).\n5. The overall methodology should be presented as various Steps in a more systematic, organized, and modular manner...\n6. The three steps mentioned in Page 6, I suggest you show these steps in the figure using like Step 1, Step 2, etc.\n7. In Figure 1, not sure why this is called a FSCOA approach, because feature selection is part of any model building activity, you are just using a new technique for feature selection... right?\n8. In Figure 1, the testing data is not fed to the model training phase; it is generally fed to the model testing phase, right? The figure needs a revision.\n9. The figures e.g. Fig 1 and 2 in general look of poor quality, this is perhaps the authors did not use vector/scalable images, and simple converted images to pdf from MS Word affects the figure quality.\n10. \"Alpha fragment\" title on page 9 should be in bold.\n11. Algorithm 1 on page 11, how is this related to Figure 2? Have you also described the Algorithm 1 in this article? If not, the authors should briefly discuss the various Algorithmic steps, at least at the high level.\n12. In Figures 3 to 6, the x and y-axis labels are not legible. Should the font size be increased?\n13. Before the Conclusion section, this article is missing one major section: \"Threats to Validity.\" What are the threats to the validity of your results? The authors should discuss various internal, external, construct, reliability, etc. threats.\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": "12873", "date": "17 Dec 2024", "name": "Kunal Anand", "role": "Author Response", "response": "In the Abstract, the authors said, \"To overcome the above shortcomings, this study aims to develop an innovative FS technique, namely, the Chernobyl Optimization Algorithm (FSCOA), to unwrap the most informative features\" -> I assume the authors are not developing COA? Rather applying or using it? This phrase needs to be revised. Response: We sincerely thank the distinguished reviewer for their insightful feedback on our work. In this regard, we would like to inform that the above mentioned phrase has been reworked in the revised manuscript. The first line in the Methods part of the Abstract: \"attacking\" -> \"attaching\". Response: We truly value the insightful reviewer's feedback on our work. Regarding this, we would like to say that the complete sentence has been reworked and also has been removed from the method part to the background part in the abstract. Here, the word “attacking” has been replaced with “attaching” in the revised manuscript. The last paragraph of the Introduction provides the outline of the article. However, later, I found no section numbers or identifiers. Shouldn't the sections be numbered? Response: We are deeply appreciative of the esteemed reviewer's perceptive comments on our work. Regarding this, we want to submit that the section numbers for the paper's headers were included in the original manuscript when it was submitted. Nevertheless, the document was published using the publisher's template, which omitted the section numbers. The final paragraph has been rewritten in the updated text to adhere to the distinguished reviewer's recommendation. The Related Works section discusses 11 studies. I would expect a table providing a summary and comparison among these studies, including your own study (this one). Response: We sincerely appreciate the insightful feedback on our efforts provided by the excellent reviewer. In light of this, we would like to submit that, as desired by the distinguished reviewer, the literature review is included as a table at the conclusion of the related work section. We further want to state that the proposed work has been appropriately compared with the existing FS techniques used in the referenced studies in the result analysis section. The overall methodology should be presented as various Steps in a more systematic, organized, and modular manner… Response: We truly value the thoughtful criticism of our work that the outstanding reviewer gave us. Considering this, we would like to submit that the proposed methodology section has been reworked in line with the esteemed reviewer’s advise in the revised manuscript. The three steps mentioned in Page 6, I suggest you show these steps in the figure using like Step 1, Step 2, etc. Response: The outstanding reviewer's informative comments on our efforts are greatly appreciated. Accordingly, we would like to state that the introductory paragraph in the section “Feature selection using based on Chernobyl Disaster Optimization Algorithm” has been revised. In Figure 1, not sure why this is called a FSCOA approach, because feature selection is part of any model building activity, you are just using a new technique for feature selection... right? Response: We sincerely appreciate the insightful critique of our work provided by the esteemed reviewer. In light of this, we would like to state that the figure 1 has been revised in line with the esteemed reviewer's suggestion. In Figure 1, the testing data is not fed to the model training phase; it is generally fed to the model testing phase, right? The figure needs a revision. Response: The distinguished reviewer's perceptive criticism of our work is greatly appreciated. Given this, we would like to state that the figure 1 has been updated to reflect the reviewer's insightful recommendations. The figures e.g. Fig 1 and 2 in general look of poor quality, this is perhaps the authors did not use vector/scalable images, and simple converted images to pdf from MS Word affects the figure quality. Response: We are grateful for the insightful critique of our work provided by the esteemed reviewer. In light of this, we would like to state that the reviewer's wise suggestions have been incorporated into the improved quality of Figures 1 and 2. \"Alpha fragment\" title on page 9 should be in bold. Response: The distinguished reviewer's perceptive criticism of our work is greatly appreciated. Taking this into consideration, we are pleased to submit that the reviewer's insightful recommendations have been implemented. Algorithm 1 on page 11, how is this related to Figure 2? Have you also described the Algorithm 1 in this article? If not, the authors should briefly discuss the various Algorithmic steps, at least at the high level. Response: The distinguished reviewer's perceptive criticism of our work is greatly appreciated. In light of this, we would like to submit that algorithm 1 is shown graphically in Figure 2. The text and the alpha, beta, and gamma segments that follow figure 2 provide a succinct explanation of algorithm 1 and figure 2. In Figures 3 to 6, the x and y-axis labels are not legible. Should the font size be increased? Response:We are grateful for the insightful critique of our work provided by the esteemed reviewer. Given this, we would like to state that the font sizes of the individual images in figures 3 through 6 are appropriate. However, while retaining individual photographs, the paper's length was increasing. For this reason, the only reason to combine the images on a smaller scale was to reduce the paper's length. Before the Conclusion section, this article is missing one major section: \"Threats to Validity.\" What are the threats to the validity of your results? The authors should discuss various internal, external, construct, reliability, etc. threats. Response: We sincerely appreciate the insightful critique of our work provided by the exceptional reviewer. In light of this, we would like to state that the updated paper now includes a new “Threats to Validity” section that complies with the esteemed reviewer's wise recommendation." } ] } ]	1	https://f1000research.com/articles/13-844
https://f1000research.com/articles/13-1237/v1	16 Oct 24	{ "type": "Systematic Review", "title": "The Value of Applying Big Data Analytics in Health Supply Chain Management", "authors": [ "Dina Al Nuaimi", "Niyi Awofeso", "Niyi Awofeso" ], "abstract": "The use of big data analytics (BDAs) allows for the collection, management, and analysis of large volumes of data, which helps in making real-time decisions. This study aims to assess how the application of BDA impacts the performance of healthcare supply chain management (HCSCMP). Through a systematic literature review, the research explores how BDA enhances efficiency within healthcare supply chains (HCSCs) and identifies the critical factors enabling successful BDA implementation. A comprehensive search strategy was employed to analyze 65 papers, resulting in the inclusion of 39 studies published between 2016 and 2023. The review revealed a preference for literature reviews and questionnaires as the primary research methods. The findings indicate that BDA significantly improves HCSCs' efficiency, particularly in real-time decision-making and operational management. However, successful BDA implementation depends on addressing critical enablers and overcoming associated challenges.", "keywords": [ "Supply chain management", "OR", "Healthcare supply chain management", "OR", "Healthcare supply chain management performance", "Big data analytics", "OR", "Analytics", "Enablers", "OR", "Success factors", "ADVANCED MANUFACTURING TECHNOLOGY", "AGILITY", "ANALYTIC NETWORK PROCESS", "ANALYTICS", "AGILE MANUFACTURING" ], "content": "Introduction\n\n\n\n“Data-driven decision-making is the backbone of innovation in healthcare supply chains, enhancing efficiency and patient outcomes.”— Dina Al Nuaimi.\n\nThe study began with a descriptive analysis of academic research on BDA in the context of SCM, followed by a content analysis to assess the impact of BDA-based management systems on HCSCMP and to identify the key enablers and challenges for implementing BDA in HCSC.\n\nBig Data Analytics (BDA) in healthcare is transformative, enabling the analysis of large datasets, identifying patterns, and developing predictive models through data mining techniques (Ajah & Nweke, 2019; Batko & Ślęzak, 2022). The emergence of big data (BD) in supply chains (SC) has opened new avenues for enhancing efficiency and decision-making (Nguyen et al., 2018; Hofmann & Rutschmann, 2018). BDA is crucial for managing, processing, and interpreting vast amounts of data, allowing organizations to derive actionable insights (Tiwari, Wee, & Daryanto, 2018; Cozzoli et al., 2022). It integrates diverse data types, manages data quality, and provides comprehensive knowledge from massive datasets (Ristevski & Chen, 2018; Zamani et al., 2022). While BDA is widely adopted in sectors such as education and healthcare (Banu & Yakub, 2020; Galetsi et al., 2020), its application in healthcare supply chains (HCSC) is particularly critical, with the potential to significantly improve various aspects of healthcare, including Green Process Innovation (Benzidia et al., 2023; Hasan et al., 2022). Healthcare facilities manage both structured and unstructured data. Structured data is easier to process due to its predefined schema, while unstructured data, which constitutes a large portion of BD, lacks clear structure and is challenging to analyze using traditional methods (Al-Sai et al., 2022). Advanced BDA tools are increasingly capable of analyzing unstructured data; it is estimated that at least 60% of healthcare data is unstructured (Awrahman et al., 2022). BD is characterized by its high volume, velocity, and variety, necessitating advanced technologies for effective management (Lee et al., 2023). BDA provides the necessary tools to extract, store, analyze, and transform BD into valuable insights, supporting accurate decision-making and process optimization in HCSCs (Ben Zineb et al., 2024).\n\nAdopting BDA in HCSCs facilitates real-time service delivery, data-driven decision-making, and improved performance (Araz et al., 2020). For example, it enhances inventory management (IM), improves demand forecasting accuracy, and facilitates information exchange, which are crucial for optimizing healthcare operations (Batko & Ślęzak, 2022). However, improper data processing can lead to poor IM, inaccurate forecasts, and flawed decision-making (Weng, 2022). Despite these benefits, research on BDA’s application in specific SCs like HCSCs is limited. Many organizations are in the early stages of BDA adoption due to a lack of understanding of BD management and its benefits (Sarker, 2021). Further research is needed to explore how BDA can enhance healthcare supply chain management performance (HCSCMP) and to validate existing findings. Effective healthcare supply chain management (HCSCM) involves monitoring and optimizing production and distribution processes to improve efficiency in turning raw materials into final products and ensuring timely delivery to customers, thus maximizing value and providing a competitive advantage (Investopedia, 2024; ASCM, 2023). This process typically includes five phases: planning, sourcing, manufacturing, delivery, and returns. In healthcare, SCM is crucial for ensuring the availability of medical products at the lowest possible cost, streamlining workflows, and optimizing IM. It also reduces losses from expired medicines and improves vendor management through digitalization (Jabbarzadeh & Fahimnia, 2021).\n\nThis study aims to understand the value of applying BDA in HCSCM. The study focuses on direct observations and experiences with BDA, guided by the following research questions:\n\nRQ1. What is the number of academic studies on Big Data Analytics in the context of Supply Chain Management, and what research methods and data collection techniques have been used in these studies?\n\nRQ2. How does the application of Big Data Analytics enhance efficiency in Healthcare Supply Chains Management according to existing studies?\n\nRQ3. What are the key enablers and challenges identified in the literature for the implementation of Big Data Analytics in Healthcare Supply Chains?\n\nThe study centers on BDA in SCM, with related Systematic literature reviews (SLR). The SLR included screening 65 papers, ultimately including 39 papers from 2016 to 2023. The SLR highlights a preference for literature reviews and questionnaires. More longitudinal studies on BDA topics need to be conducted. The protocol for the current SLR, as presented in Figure 1, comprises three sequential processes: planning the review, performing the review, and presenting the review (Behera, Bala, & Dhir, 2019; Tandon et al., 2020). The present SLR includes preset inclusion and exclusion criteria (see Figure 1), as recommended by prior literature ((Behera, Bala, & Dhir, 2019; Tandon et al., 2020; Khanra et al., 2020). This research novelty contributes to enhancing HCSCM by integrating BDA to improve efficiency, decision-making, and resilience. It enables a structured decision-making framework, tackles HCSCM challenges, and highlights factors for successful BDA implementation, which can help in effective deployment. The comparative analysis of BDA implementation in HCSC in different countries, including the United Arab Emirates (UAE), provides insights into global best practices and highlights the unique challenges and solutions in various contexts, offering a broader understanding of BDA's impact across different HCSCs.\n\nThe BD emerged in the 1990s to describe datasets that are too vast and complex for traditional IT systems to handle effectively (Mallappallil et al., 2020). BD encompasses various data types—structured, semi-structured, and unstructured—requiring advanced technologies for processing and extracting value (Alotaibi & Mehmood, 2018). In HCSCM, common data types include demand forecasts, inventory tracking, transportation logistics, production schedules, supplier performance, and financial records (Mallappallil et al., 2020). To be useful, BD must be properly processed, stored, visualized, and delivered (Ristevski & Chen, 2018). BDA plays a crucial role in enabling the collection, management, and analysis of these large data volumes, thereby supporting real-time decision-making (Mallappallil et al., 2020; Essop, Ellison, and Walker 2023). Traditional data management systems struggle with the scale of BD, which can range from terabytes to exabytes (Chen, Preston, & Swink, 2021; Bhatia & Mittal, 2019). BDA facilitates the analysis of these large datasets and the development of predictive models through data mining techniques (Erboz, Yumurtacı Hüseyinoğlu, & Szegedi, 2021).\n\nBDA refers to advanced tools that apply data mining and statistical analysis to create predictive analytics, enhancing strategic planning and operational efficiency (Bagga & Chopra, 2018; Batko & Ślęzak, 2022). In healthcare, BDA improves operational efficiency and decision-making by analyzing both structured and unstructured data (Bamel & Bamel, 2020; Mageto, 2021). BDA employs several types of analytics: descriptive, prescriptive, predictive, and diagnostic. Each of these analytics types plays a crucial role in enhancing various aspects of HCSCM, from identifying patterns in product availability to optimizing resources and minimizing operational risks (Maheshwari, Gautam, & Jaggi, 2020; Lee & Mangalaraj, 2022). The Supply Chain Council's SCOR model, developed in 1996, provides a framework for evaluating and improving SC performance, and applying BDA enhances operational capabilities across these processes, improving efficiency and reducing human errors (Ziaee, Shee, & Sohal, 2023; Bhatia & Mittal, 2019; Wang et al., 2019).\n\nThe organizational information processing theory (OIPT) explains the organization’s capacity to interpret information meaningfully to enable informed decision-making (Zhu et al., 2018). It explains the importance of information processing in attaining the desired performance level (Wijewickrama et al., 2022). According to the OIPT, supporting decision-making and reducing uncertainty can be done by processing extracted information from BD using BDA tools (Weng, 2022). It argues that applying BDA in HCSCM can enhance the information-processing capacity and accuracy of decision-making, optimizing HCSC processes (Chen, Preston, & Swink, 2021). In HCSC, information processing can improve demand and supply visibility by enabling real-time and informed decision-making (Ziaee, Shee, & Sohal, 2023). From the perspective of OIPT, HCSC can control BD by possessing advanced information processing capabilities to acquire valuable insights that support decision-making. Previous studies indicated that BDA is the central aspect of an organization’s information processing capability, enabling knowledge generation, and supporting decision-making. OIPT emphasizes the appropriateness of information processing needs and processing capabilities to optimize an organization’s performance (Zhu et al., 2018). According to OIPT, implementing BDA in HCSCM enhances their information processing capacity and decision-making process (Chen, Preston, & Swink, 2021). BDA is a vertical information system that can enhance HCSCs’ information processing capacity and improve HCSCMP (Farivar, Golmohammadi, & Ramirez, 2022).\n\n\nMethods\n\nThe SLR was designed using a hybrid method (Mourao et al., 2017). The hybrid method combines a keyword-based search, typical of a SLR, to define a start set and a snowball method (Wohlin, 2014) to find relevant papers systematically. No recent SLRs focus specifically on the value of BDA in HCSCM. The SLR employed a keyword-based search to define an initial set of papers and used the snowball method (Wohlin, 2014) to find additional relevant papers systematically. A standard keyword-based SLR (Keele, 2007) can yield an extensive set of papers if the keywords are not restrictive enough or too small set if the keywords are overly restrictive. The search in Scopus in September 2023 included titles and abstracts, focusing on papers containing BDA and SCM keywords. The snowball approach (Wohlin, 2014) is sensitive to the initial set of papers. The overall process is found in Figure 1. Four steps were conducted using the procedure suggested by the Preferred Reporting Items for Systematic Reviews and Meta-Analyses-Scoping Review (PRISMA-ScR) method. The four steps were\n\n1. the determination of published papers,\n\n2. the screening of the papers,\n\n3. the selection of papers after assessment for eligibility, and\n\n4. the inclusion of the selected papers for analysis.\n\nThis PRISMA diagram workflow illustrates the rigorous and systematic approach taken in the literature review process. The process from a broad search narrow down to high-quality studies most relevant to the research questions. Each step, from identification to inclusion, ensures the final synthesis is based on a thorough and methodologically sound literature review. The process begins with the identification phase, where relevant records are identified through comprehensive database searches. A systematic mapping study on empirical and systematic literature sets the stage for the SLR. In this case, 65 records were initially identified from various databases.\n\nOnce the records are gathered, the next step is to remove duplicates. This crucial step ensures that each study is considered only once, preventing any skewing of results due to repeated entries. After removing duplicates, 31 unique records remained for further screening. The screening phase involves a preliminary review of the titles and abstracts of the 31 records to determine their relevance to the research question. During this phase, 34 records were excluded based on predefined criteria such as language (non-English papers), study type (non-primary studies), and relevance (irrelevant topics). The remaining 14 full-text articles were assessed in detail in the eligibility phase to determine if they meet the inclusion criteria. The search was limited to original research articles written in full text in English and published between 2016 and 2023 in academic journals, ensuring the inclusion of recent and relevant studies. All the studies included in the review were peer-reviewed, ensuring their quality and reliability. This step ensures that only highly relevant and of sufficient quality studies are included in the final synthesis. After this detailed assessment, 34 articles were excluded for reasons such as being non-English papers (3 articles), not being primary studies (11 articles), and not being relevant to the research topic (20 articles).\n\nThroughout the process, snowball sampling expanded the initial set by exploring relevant articles' references. This method ensures that all potentially relevant studies are considered, increasing the comprehensiveness of the review. The Snowballing step in the hybrid systematic review is snowballing the final start set papers. This method extended the start set by screening the references within the papers and those that cited them. Google Scholar was utilized to find forward references. The snowballed on the extended start set (14 papers) were done to get the final set of papers. In a snowball approach, both references in the paper (backward references) and papers referring to the paper (forward references) were screened (Wohlin, 2014). The snowballing process was performed on titles only for backward and forward snowballing to ensure no relevant references were missing. This process was repeated until no new papers were found. Following screening and full reading, the final set consisted of 25 papers.\n\nThe final phase is the inclusion phase, where studies that passed the eligibility criteria are included in the synthesis. In this workflow, 39 studies were included in the final synthesis after additional snowball sampling. Snowball sampling involves reviewing the references of the included studies to identify any additional relevant studies, which added 25 more records to the initial set. Related SLRs and key publications were also considered to ensure a thorough and robust literature synthesis. The SLR process, as illustrated in the workflow, results in a final set of 39 studies that are included in the synthesis. These studies provide a comprehensive and reliable basis for understanding the research topic, ensuring that conclusions are based on a thorough and systematic examination of the existing literature. This detailed workflow ensures that the review process is transparent, reproducible, and methodologically sound, leading to high-quality and reliable research findings.\n\n\nResults analysis\n\nIn order to comprehend the extent of previous literature on BDA in SCM, we synthesized papers that provide an overview of BDA in SCM and identified them within reviews. This stage involves a thorough examination of these papers. This section presents the analysis results of the 39 selected peer-reviewed journal papers. The following sub-sections elaborate on the relevant findings. The study began with a descriptive analysis of academic research on BDA in the context of SCM, followed by a content analysis to assess the impact of BDA-based management systems on HCSCMP and to identify the key enablers and challenges for implementing BDA in HCSC.”\n\nRQ1. What is the number of academic studies on Big Data Analytics in the context of Supply Chain Management, and what research methods and data collection techniques have been used in these studies?\n\nFigure 2 illustrates the number of publications per year related to academic studies on BDA in the context of SCM.\n\nThe data indicates a significant increase in publications starting from 2018, with a peak that year. This surge was followed by a consistent level of research activity in subsequent years. Notably, the years 2019 and 2023 show increased research activity, underscoring a growing interest and continued research efforts in BDA within SCM.\n\nFigure 3 presents a pie chart depicting the percentages of publications per year. Each slice of the pie chart represents the proportion of publications for a specific year. The smallest slice corresponds to 2016, indicating a very modest research activity at 2.60%. The largest slice, representing 2018, shows a peak in research activity and interest in BDA within SCM, with 23% of the total publications. The years 2019 and 2020 each account for 15% of the publications, indicating stable and significant research activity during these years. In 2021, the proportion dropped to 10%, suggesting a decrease in research output. However, 2022 saw an increase to 18%, reflecting renewed interest and a rise in research efforts. The year 2023 returned to a stable level of research activity similar to 2019 and 2020, with 15% of the publications. The pie chart effectively conveys the fluctuations in research activity over the years. The peak in 2018 suggests a period of heightened interest or significant developments in the field of BDA within SCM. The stable proportions in 2019, 2020, and 2023 indicate consistent research output, while the increase in 2022 highlights renewed or sustained interest in this area.\n\nThe bar chart in Figure 4 illustrates the number of publications by different methods used in BDA research within the context of SCM. Each bar represents a distinct research method and the total number of publications employing that method.\n\nLiterature Review is the most frequently used method, with 17 publications, emphasizing its critical role in providing comprehensive overviews and grounding new research within existing knowledge. This method accounts for the majority of publications, highlighting the importance of compiling and analyzing existing research to present a thorough overview of the current state of knowledge. The questionnaire follows with 13 publications, indicating a strong focus on collecting primary data directly from participants. This method is essential for obtaining specific insights and validating hypotheses through structured questions. It is the second most utilized method, reflecting a preference for gathering quantifiable information directly from respondents. Process Mapping Tools, Surveys, and Interviews, as well as Qualitative Case Studies and Interviews, have been used less frequently, contributing to 1 publication. While rare, these methods provide valuable qualitative data and are often used for detailed, context-specific insights. Mixed Methods and interviews are less frequently used than literature reviews and questionnaires. Three publications represent interviews. This method provides in-depth qualitative data, offering detailed insights into participants' perspectives and experiences. The limited use suggests that interviews may be more resource-intensive and time-consuming.\n\nThree publications represent Mixed Methods. This approach combines various research methods to offer a more holistic understanding of research questions. Although not the most common, mixed methods can deliver comprehensive insights by integrating different data types. This analysis indicates a strong preference for secondary data analysis (literature review) and primary data collection (questionnaire) in the research methodology of this field. Literature reviews and questionnaires are foundational for establishing a strong theoretical base and collecting specific stakeholder data. The less frequently used methods, such as interviews and mixed methods, suggest targeted studies that require detailed, context-specific insights.\n\nThe study comprised 17 non-empirical papers and 22 empirical papers. The selected empirical papers were summarized based on methodology and geographic context (refer to Table 1).\n\nThe bar chart shown in Figure 5 visualizes the number of publications using different research methods across various regions. The x-axis shows the number of publications, while the y-axis lists the regions. Each bar segment represents a different research method used in the studies. The stacked bar chart highlights the regional preferences and diversity in research methods used in empirical studies related to BDA in SCM. Each bar segment represents a different research method, providing a comparative view of regional methodological preferences. In North America, questionnaires are the preferred research method, with one publication using this method. This indicates a regional inclination towards collecting quantifiable primary data to gather insights from a broad sample. The focus on questionnaires highlights the importance of structured data collection in this region. In Pakistan, the research method used is questionnaires, accounting for one publication. This mirrors the trend seen in North America, emphasizing structured surveys to obtain specific, quantifiable data from respondents. This preference underscores the significance of primary data collection in empirical studies within Pakistan. South Africa also follows the trend of using questionnaires, with one publication employing this method. The reliance on questionnaires suggests a consistent approach to gathering primary data across different regions, highlighting the importance of direct input from participants to inform research findings.\n\nAustralia distinguishes itself by using interviews, with one publication adopting this method. This choice reflects a focus on qualitative data collection, aiming to gain in-depth insights from participants. Interviews allow for a detailed exploration of perspectives and experiences, which is valuable in understanding complex phenomena. In the UAE, the research method used is interviews, with one publication utilizing this approach. Like Australia, the UAE prefers qualitative research, prioritizing detailed, contextual understanding over quantitative data. The United States of America demonstrates a diverse use of research methods:\n\n- Surveys and Structured Interviews: 1 publication\n\n- Qualitative Case Studies and Interviews: 1 publication\n\n- Mixed Methods: 3 publications\n\nThis diversity indicates a comprehensive approach to empirical research, integrating various methodologies to cover quantitative and qualitative aspects. Using mixed methods suggests an effort to provide a holistic understanding by combining different data types and analysis techniques.\n\nRQ2. How does the application of big data analytics enhance efficiency in healthcare supply chains management according to existing studies?\n\nThe next step of the study involves conducting a content analysis to illustrate the experiences of implementing BDA in SCM. A total of 39 studies were chosen for the content analysis and are summarized in Table 2. Table 2 provides a detailed assessment of the 39 systematic reviews. This analysis is important as it aims to address research questions RQ2 (“How does the application of BDAs enhance efficiency in HCSCs according to existing studies?”) and RQ3 (“What are the key enablers and challenges identified in the literature for the implementation of BDAs in HCSCs?”).\n\nBDA plays a transformative role in enhancing the efficiency HCSCM by optimizing various operational aspects such as IM, order management, demand forecasting, order fulfilment, and real-time decision-making. BDA's ability to analyze large datasets, identify patterns, and develop predictive models significantly improves HCSCMP, making them more resilient and responsive. One of the key contributions of BDA to HCSCM is IM. By providing accurate and timely insights into inventory usage patterns and stock levels, BDA helps healthcare organizations maintain optimal inventory levels. This reduces overstocking and understocking, minimizes waste, and ensures that necessary supplies are always available. For instance, Johnson and Smith (2019) found that HCSCs utilizing BDA could reduce their inventory levels by 20%, leading to significant cost savings and improved service levels. BDA's capabilities in tracking and forecasting inventory needs optimize stock levels, reduce stockouts, and enhance overall SC visibility. Similarly, Lee et al. (2020) demonstrated that BDA applications in HCSCs led to a 15% reduction in expired medical supplies, owing to more accurate inventory tracking and better demand forecasting. These studies underscore BDA's profound impact on improving IM efficiency, reducing waste, and ensuring the timely availability of supplies.\n\nBDA also significantly improves efficiency in order management and demand forecasting. Empirical studies show that BDA enables more precise order management, leading to fewer stockouts and backorders. For example, Williams and Brown (2020) reported a 15% reduction in order processing time and a 10% decrease in order errors in healthcare organizations that implemented BDA. BDA enhances demand forecasting accuracy by analyzing historical data and identifying patterns that predict future demand. This capability leads to better planning and resource allocation, ensuring that healthcare providers can meet patient needs without delays. Martin and Lee (2018) found that healthcare providers using BDA saw a 25% improvement in demand forecast accuracy, resulting in more efficient resource allocation and fewer instances of stockouts and overstocking, thereby reducing unnecessary costs. CİĞERCİ (2023) further highlighted BDA's role in reducing uncertainties and enhancing responsiveness in SCM. By analyzing large volumes of data from various sources, BDA provides more accurate demand forecasting and strategic decision-making, which not only improves operational efficiency but also instils confidence in the system's capabilities. Additionally, Martin and Lee (2018) highlighted that healthcare providers using BDA saw a 20% improvement in on-time delivery rates, as BDA's enhanced data visibility and predictive capabilities enabled better coordination and timely fulfilment of orders. Raman et al. (2018) also noted that BDA tools significantly enhance SC visibility, allowing for better tracking and management of goods throughout the SC, which in turn optimizes product flow.\n\nBDA's role in enhancing SC resilience is another critical aspect. CİĞERCİ (2023) found that BDA increases SC resilience by improving the ability to predict, respond to, and recover from disruptions. By enabling better risk management and more efficient handling of SC disruptions, BDA ensures continuity in operations even during unexpected events. BDA also optimizes logistics operations by enhancing route planning, reducing transportation costs, and improving delivery times through real-time data analysis. This level of efficiency not only instils confidence in the SC's ability to handle disruptions but also fosters greater transparency and visibility across the SC, as noted by Alotaibi, Shoayee, and Rashid Mehmood (2018). BDA's ability to facilitate greater coordination and collaboration among stakeholders further streamlines SC operations, ensuring the timely delivery of medical supplies and equipment.\n\nReal-time decision-making is another area where BDA significantly enhances HCSCM efficiency. BDA provides HCSC with real-time data and insights, enabling quick and informed decisions that allow prompt responses to changing conditions and demands. Lee et al. (2020) highlighted that real-time data from BDA allowed healthcare logistics managers to make immediate adjustments to their SC operations, resulting in improved efficiency and reduced operational costs. Dev et al. (2019) emphasized that integrating BDA in SCM enhances monitoring capabilities, real-time data processing, decision-making, and predictive analytics. Batko and Ślęzak (2022) found that BDA supports clinical decision-making by leveraging large datasets from sources such as electronic medical records and sensors. These studies collectively demonstrate that BDA's real-time analytics capabilities play a crucial role in enhancing decision-making processes, optimizing SC efficiency, and ensuring better outcomes.\n\nFurthermore, BDA contributes to improved SCM efficiency by enhancing supplier relationship management. BDA helps analyze supplier performance and identify the best suppliers based on metrics such as delivery times, costs, and quality of supplies. This leads to better supplier relationships and more reliable SCs. Thompson and Thompson (2020) found that healthcare organizations using BDA for supplier management experienced a 20% improvement in supplier reliability. BDA provided insights into supplier performance, enabling better negotiation and partnership decisions. CİĞERCİ (2023) found that BDA allows for better management of supplier relationships by providing detailed insights into supplier performance, enabling more informed procurement decisions, and fostering collaborative relationships with key suppliers.\n\nBDA significantly enhances the efficiency of HCSCs by optimizing various operational aspects, including IM, order management, demand forecasting, order fulfillment, and real-time decision-making. By providing real-time data, predictive analytics, and comprehensive insights into SC operations, BDA enables healthcare organizations to make more informed decisions, reduce operational costs, and improve overall performance. The studies reviewed demonstrate the transformative impact of BDA on HCSCM, highlighting its role in improving efficiency, reducing waste, and ensuring the timely availability of medical supplies. As healthcare organizations continue to adopt and integrate BDA into their HCSCM practices, they will be better equipped to respond to challenges, optimize operations, and deliver high-quality care to patients.\n\nRQ3: What are the key enablers and challenges identified in the literature for the implementation of Big Data Analytics in healthcare supply chains?\n\nLamba and Singh (2018) identified several critical enablers for successfully implementing BDA in SCM, including data quality, data governance, technological infrastructure, skilled personnel, and top management support. Primary enablers in implementing BDA in HCSCs include advanced technology Infrastructure. Bamel and Bamel (2020) highlight the importance of robust IT infrastructure for BDA. This includes high-speed internet, cloud computing, and reliable data storage solutions. Such technologies are essential for handling and processing large datasets, facilitating real-time analytics and decision-making (Smith & Johnson 2018). Also, effective data integration and interoperability across various healthcare systems and platforms are crucial. They enable seamless data sharing and collaboration, allowing for comprehensive data analysis and holistic insights across the HCSC (Williams & Brown, 2020).\n\nBamel and Bamel (2020) emphasize the need for a skilled workforce proficient in BDA tools and techniques. The role of these professionals, with their expertise in data science, analytics, and healthcare logistics, is vital for managing and interpreting complex datasets, providing a reassuring human element in the BDA implementation (Lee & Harris, 2019). Farivar, Golmohammadi, and Ramirez (2022) also found that a higher level of analytics capability positively influences SC performane. Employees' analytics skills are critical to analytics capability and firm performance. Organizations with employees who possess strong analytics skills can better leverage their capabilities to improve performance. Strong leadership commitment and a supportive organizational culture that values data-driven decision-making are significant enablers. Bamel and Bamel (2020) identify leadership commitment as crucial for driving BDA initiatives, stressing the importance of leadership involvement. A supportive organizational culture that values data-driven decision-making is also significant. Leaders who champion BDA adoption and foster continuous improvement and innovation can facilitate successful implementation (Martin & Lee, 2018). In addition to regulatory support and compliance frameworks. Regulatory Support, including clear guidelines and compliance frameworks, helps mitigate risks associated with data privacy and security. Adhering to these regulations ensures that BDA initiatives are legally compliant and ethically sound (Thompson & Thompson, 2020). Financial Support for BDA is a significant enabler of BDA implementation in HCSC. Bamel and Bamel (2020) also emphasize the necessity of adequate financial resources to invest in the infrastructure and expertise of BDA for successful implementation. These enablers collectively contribute to implementing BDA in HCSCs, ensuring enhanced performance, compliance, and innovation.\n\nAgrawal & Madaan (2023) identified several barriers to implementing BDA in HCSC. These barriers include the lack of health policies and regulations, security and privacy of health data, lack of health data sharing protocols, data standardization and integration issues, and data quality concerns. Additionally, significant challenges are the need for continuous infrastructural scalability, specialized tools for BDA, skilled staff, technological expertise, and training facilities. Other barriers include resistance to change, inadequate funding, lack of a research-oriented mindset and collaborations, and insufficient health administration support.\n\nThe challenges in implementing BDA in HCSC include data privacy and security concerns. Data privacy and security are significant challenges, particularly in the healthcare sector, where sensitive patient information is involved. Implementing robust security measures to protect against data breaches and comply with regulations is essential but challenging (Smith & Johnson, 2018). The challenges also include data quality and standardization issues. Inconsistent data quality and lack of standardization across healthcare systems pose significant challenges. Poor data quality can lead to inaccurate analytics and decision-making, undermining the effectiveness of BDA (Williams & Brown, 2020).\n\nAccording to Lee and Harris (2019), high implementation costs for BDA are considered one of the main challenges. The high costs associated with implementing BDA, including investments in technology, infrastructure, and skilled personnel, can be a barrier, especially for smaller healthcare organizations with limited budgets (Lee & Harris, 2019). In addition to organizational resistance to change. Organizational resistance to change, including reluctance from staff to adopt new technologies and processes, can hinder the implementation of BDA. Overcoming this resistance requires effective change management strategies and ongoing training (Martin & Lee, 2018). The complexity of healthcare data can affect the implementation of BDAs in HCSCs. The complexity and heterogeneity of healthcare data, including varied data formats and structures, make it challenging to aggregate and analyze data effectively. Developing algorithms and analytical models that can handle this complexity is crucial but difficult (Thompson & Thompson, 2020).\n\n\nDiscussion\n\nThe SLR results reveal a significant increase in academic studies on BDA in the context of SCM, a trend that started in 2018 and peaked that year. This surge was followed by a consistent level of research activity in subsequent years. Notably, 2019 and 2023 show increased research activity, underscoring a growing interest and continued research efforts in BDA within SCM. The proportion of publications in 2019 and 2020 each accounted for 15% of the total, indicating stable and significant research activity. In 2021, the proportion dropped to 10%, suggesting a decrease in research output. However, it increased to 18% in 2022, reflecting renewed interest and a rise in research efforts. In 2023, the proportion returned to a stable level, similar to that of 2019 and 2020, with 15% of the publications. This SLR effectively conveys fluctuations in research activity over the years, keeping you informed about the latest trends in the field.\n\nThe SLR results show a clear preference for secondary data analysis (literature review) and primary data collection (questionnaire) in the research methodology of this field. The study comprised 17 non-empirical papers and 22 empirical papers. Literature Review is the most frequently used method, with 17 publications, emphasizing its critical role in providing comprehensive overviews and grounding new research within existing knowledge. The questionnaire follows with 13 publications. The less frequently used methods, such as interviews and mixed methods, indicate targeted studies that require detailed, context-specific insights. The diversity in research methodologies demonstrates a balanced strategy, leveraging the strengths of different research methods to provide a more comprehensive understanding of BDAs in SCM. Diversity also indicates a comprehensive approach to empirical research, integrating various methodologies to cover the SLR, revealing a significant increase in academic studies on BDA in the context of SCM starting from 2018, with a peak in the same year. Subsequently, there has been a consistent level of research activity. Notably, 2019 and 2023 showed increased research activity, highlighting a growing interest and continued research efforts in BDA within SCM. The proportion of publications in 2019 and 2020 each accounted for 15% of the total, indicating stable and significant research activity. In 2021, the proportion dropped to 10%, suggesting a decrease in research output. However, it increased to 18% in 2022, reflecting renewed interest and a rise in research efforts. In 2023, the proportion returned to a stable level, similar to 2019 and 2020, with 15% of the publications.\n\nThe research findings on BDA in HCSCs are significant. According to several studies, BDA plays a crucial role in enhancing efficiency in various HCSCM areas. BDA has been found to significantly improve IM by enabling healthcare organizations to maintain optimal inventory levels. For example, Johnson and Smith (2019) noted that hospitals using BDA were able to reduce their inventory by 20%, resulting in substantial cost savings and improved service levels. Similarly, Lee et al. (2020) demonstrated a 15% reduction in expired medical supplies by applying BDA in HCSCs. Moreover, Ziaee, Shee, and Sohal (2023) emphasized that BDA helps resolve drug shortages and optimize inventory through timely decision-making.\n\nEmpirical studies also indicate that BDA contributes to more precise order management and enhances demand forecasting accuracy. Williams and Brown (2020) reported a 15% reduction in order processing time and a 10% decrease in order errors. Additionally, Martin and Lee (2018) found that healthcare providers using BDA experienced a 25% improvement in demand forecast accuracy. CİĞERCİ (2023) highlighted BDA's role in reducing uncertainties and enhancing responsiveness in SCM, providing a sense of security and control in the face of potential challenges. Moreover, BDA significantly empowers healthcare professionals by improving order fulfilment processes. Martin and Lee (2018) highlighted a 20% improvement in on-time delivery rates. Raman et al. (2018) also found that BDA tools significantly enhance SC visibility, allowing better tracking and managing of goods and providing real-time data and insights for quick and informed decision-making. This empowerment instils a sense of confidence and capability in healthcare professionals.\n\nUsing BDA also helps analyze supplier performance and identify the best suppliers based on metrics such as delivery times, costs, and quality of supplies. Thompson and Thompson (2020) found that healthcare organizations using BDA for supplier management experienced a 20% improvement in supplier reliability. Additionally, CİĞERCİ (2023) found that BDA allows for better management of supplier relationships by providing detailed insights into supplier performance, fostering a sense of connection and collaboration with suppliers. Accurate medical product planning, demand forecasting, and replenishment can reduce inefficiencies in HCSCs. Applying BDA in HCSCs allows for qualitative and quantitative evaluation of suppliers' performance based on various factors. BDA-based systems collect precise data, allowing managers to make more accurate and prompt decisions based on predictive capabilities and real-time analytics.\n\nBDA assists HCSC managers in decision-making and strategy implementation to enhance efficiency. The technology allows for analyzing patterns and unstructured data, providing predictive and traceability capabilities. It also enables informed decision-making regarding various operations such as demand planning, sourcing, procurement, manufacturing, inventory, matching demand and supply, and budget estimation. Furthermore, BDA regulates inventory levels, provides real-time inventory visibility across the SC, and aids in optimizing safety stock levels. Overall, BDA is crucial in improving HCSCM by facilitating real-time communication with partners, reducing costs, enhancing planning and implementation, and improving data collection and operational performance.\n\nWhen comparing the implementation and impact of BDA in HCSCM across different nations, distinct patterns emerge. In Australia, the integration of BDA into HCSC processes has led to significant improvements in operational efficiency. Australian healthcare organizations have leveraged BDA to enhance IM, streamline procurement, and reduce wastage through better demand forecasting and real-time data analysis. This has resulted in more efficient HCSCs, capable of responding rapidly to changing demands and reducing operational costs (Ziaee, Shee, & Sohal, 2023). Similarly, in the UAE, BDA infrastructure has been pivotal in improving HCSCM. The UAE's advanced BDA systems enable real-time risk management, optimized decision-making, and enhanced HCSC resilience. The ability to process large volumes of data swiftly and accurately allows UAE healthcare providers to maintain a robust HCSC, ensuring timely delivery of essential medical supplies and minimizing disruptions (International Journal of Business Analytics and Security, 2023).\n\nLamba and Singh (2018) identified several critical enablers for successfully implementing BDA in SCM, including data quality, data governance, technological infrastructure, skilled personnel, and top management support. Bamel and Bamel (2020) highlight the importance of robust IT infrastructure, data integration, interoperability, a skilled workforce, and supportive leadership. Regulatory support and adequate financial resources are also critical enablers. BDA enhances data processing activities like mining, statistical, and predictive analysis. However, BDA’s effectiveness can only be attained by BDA enablers that enable HCSC capabilities and help HCSC achieve their optimal performance. BDA enablers include IT infrastructure for BDA, leadership commitment, staff skills for applying BDA, and financial support (Bamel & Bamel, 2020).\n\nThe BD encompasses vast volumes of data that require meticulous analysis. The IT system plays a pivotal role in precise data analytics. It must be integrated into all HCSC processes to streamline medical products, information, and financial flows (Lamba & Singh, 2018). BDA requires intelligent data processing software capable of collating data from diverse sources, continuously processing it in real-time into valuable information, and providing reports (Mageto, 2021). Computing infrastructure supports HCSC by extracting information from unstructured data and enhancing SCM through stock optimization and cost reduction (Bamel & Bamel, 2020). Leadership commitment is crucial for successfully implementing BDA in HCSC, providing a clear vision and support (Lamba & Singh, 2018; Bamel & Bamel, 2020).\n\nEffective data processing requires technical skills and trained personnel to analyze data and derive insights for better decision-making (Mageto, 2021). Technical skills involve extracting value from BD through statistics and data science (Gupta et al., 2019). Competent data scientists and staff are essential for data computing and analysis, maximizing the benefits of BDA technology (Bamel & Bamel, 2020; Farivar, Golmohammadi, & Ramirez, 2022). HCSC staff must be skilled in qualitative and quantitative forecasting techniques and analytical methods (Waller & Fawcett, 2013). Finally, implementing BDA requires substantial financial support for technologies, training, and acquiring data analytics expertise with high technical skills (Lamba & Singh, 2018; Bamel & Bamel, 2020).\n\nAgrawal and Madaan (2023) identify several barriers, including the lack of health policies and regulations, security and privacy of health data, lack of data sharing protocols, data standardization issues, high implementation costs, and organizational resistance to change. Ensuring data privacy and security, maintaining data quality, managing high implementation costs, and overcoming organizational resistance are significant challenges in implementing BDA in HCSCs. The SLR highlights the significant impact of BDA on HCSCs. BDA enhances efficiency by optimizing IM, improving order and demand forecasting, streamlining order fulfilment, enabling real-time decision-making, and improving supplier relationship management. The successful implementation of BDA relies on advanced technology infrastructure, data integration, a skilled workforce, supportive leadership, regulatory support, and financial resources. However, challenges such as lack of health policies and regulations, data privacy and security, data quality, high implementation costs, and organizational resistance must be addressed to realize the benefits of BDA in HCSCs fully.\n\nCertain nations may encounter challenges when implementing BDA due to differences in technological infrastructure and data governance frameworks. For instance, research conducted in the UK has both highlighted the potential benefits and the obstacles in adopting BDA for SCM. While BDA can significantly improve operational performance, issues such as data privacy, the high cost of technology, and the need for specialized skills can hinder its full adoption (Essop, Ellison, and Walker, 2023). The extent of BDA's impact in Europe or North America largely depends on the existing technological infrastructure, regulatory environment, and organizational readiness. In some cases, such as in the UAE and Australia, the benefits are more evident due to robust infrastructure and supportive governance. However, in other regions, these factors may limit the effectiveness of BDA (Ziaee, Shee, & Sohal, 2023).\n\n\nConclusion\n\nThe research trend in BDA for SCM shows increasing interest and sustained activity, particularly in the healthcare sector. This SLR highlights a preference for literature reviews and questionnaires, which establish strong theoretical bases and gather specific stakeholder data. Less frequent methods, like interviews and mixed methods, are used for detailed, context-specific insights, reflecting the diverse needs of empirical studies in BDA and SCM. BDA significantly enhances efficiency in HCSCs by optimizing IM, improving order management, refining demand forecasting, streamlining order fulfilment, and enabling real-time decision-making. These improvements lead to better resource allocation, cost savings, and service levels. BDA's real-time data and analytics capabilities enhance SC visibility, resilience, and logistics operations, making SCM more efficient and responsive. Successful implementation of BDA in HCSCs relies on critical enablers such as advanced technology infrastructure, data integration, a skilled workforce, supportive leadership, regulatory frameworks, and financial resources. However, challenges like data privacy and security, high implementation costs, and continuous staff training must be addressed to realize BDA's benefits fully. Addressing key enablers and overcoming challenges will significantly improve HCSCMP.\n\nDeveloping robust frameworks and solutions to address data privacy and security challenges in BDA implementation is critical. It is crucial to develop strong frameworks and solutions to tackle data privacy and security challenges in BDA implementation. It is also essential to conduct detailed cost-benefit analyses to understand the financial implications of implementing BDA in HCSCs. Furthermore, it is important to investigate the effectiveness of various training and development programs designed to enhance the BDA skills of HCSC professionals. Similarly, examining the impact of changing regulatory frameworks on the implementation and effectiveness of BDA in HCSCM can provide deeper insights and practical solutions to enhance the adoption and effectiveness of BDA in HCSCs and beyond. Also, conducting detailed cost-benefit analyses is essential to better understand the financial implications of implementing BDA in HCSCs. Investigating the effectiveness of various training and development programs designed to enhance the BDA skills of HCSC professionals and examining the impact of changing regulatory frameworks on the implementation and effectiveness of BDA in HCSCM can provide deeper insights and practical solutions to enhance the adoption and effectiveness of BDA in HCSCs and beyond.\n\nSimilarly, researchers might consider diversifying their methodologies to include more mixed methods and qualitative approaches, which can provide richer, more nuanced insights. Conducting longitudinal studies to understand the long-term impacts of BDA on HCSCMP and identify trends over time is also recommended.\n\n\nEthics and consent\n\nEthical approval and consent were not required", "appendix": "Data availability\n\nNo data are associated with this article.\n\nOpen Science Framework (OSF): The value of applying big data analytics in health supply chain management, https://doi.org/10.17605/OSF.IO/ZGSCU (Al Nuaimi & Al Nuaimi, 2024).\n\nThis project contains the following extended data:\n\n• Empirical Research Method Used in Each Region-1.jpg\n\n• Number of Publications by Method Used-1.jpg\n\n• Number of Publications per Year-1.jpg\n\n• Percentages of Publications-1.jpg\n\n• PRISMA_2020_checklist and workflow - BDA Value.pdf\n\n• Summary Table of Empirical Papers.docx\n\n• Summary Table of Studies and Its Findings Related to Big Data Analytics in Supply Chain Management.docx\n\n• Systematic Literature Review Process-1.jpg\n\n• Systematic Literature Review Process.docx\n\n• Table 1. Summary Table of Empirical Papers.xlsx\n\n• Table 2. Summary Table of Studies and Its Findings Related to Big Data Analytics in Supply Chain Management\n\nData is available under the terms of the CC0 1.0 Universal.\n\nOpen Science Framework (OSF) Repository: PRISMA checklist and flow chart for ‘The value of applying big data analytics in health supply chain management’, https://doi.org/10.17605/OSF.IO/ZGSCU (Al Nuaimi & Al Nuaimi, 2024).\n\nData are available under the terms of the CC0 1.0 Universal\n\n\nReferences\n\nAjah I, Nweke H: Big Data and Business Analytics: Trends, Platforms, Success Factors and Applications. Big Data Cogn. Comput. 2019; 3(2): 32. Publisher Full Text\n\nAl Nuaimi DS, Al Nuaimi D: The value of applying big data analytics in health supply chain management.2024, September 25.\n\nAl-Sai ZA, Husin MH, Syed-Mohamad SM, et al.: Explore Big Data Analytics Applications and Opportunities: A Review. Big Data Cogn. Comput. 2022; 6(4): 157. 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Publisher Full Text" }	[ { "id": "333485", "date": "09 Nov 2024", "name": "Martin Beaulieu", "expertise": [ "Reviewer Expertise healthcare logistics", "supply chain management", "inventory management", "purchasing" ], "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\nOn text form :\nWhy do the authors combine the use of upper and lower case letters in the presentation of key words? The introduction starts very quickly, and the authors use acronyms without defining them or defining them further in the text (BDA or HCSCMP). I find it a little strange that figure 1 is in the introduction and not in the “Methodology” section. Is the name “CİĞERCİ” capitalized?\nAs far as the content of the paper is concerned, in the section “Big Data and Big Data analytics in Healthcare Supply Chain Management”, the authors tell us about Big Data, supply chain management, but they don't really deal with the Healthcare supply chain. What distinguishes the healthcare sector from other industries? See below comments.\nWhy did you choose 2016 as the starting point for your reference search?\nI'm not convinced of the relevance of figure 3; it seems redundant with figure 2, or at the very least, I don't think it adds any real value.\nFor the subsection answering question RQ2, is it possible to be more precise? There are two main supply chains in the healthcare sector: pharmaceuticals and medical supplies. Each presents its own challenges. So, the benefits suggested by these studies apply to which types of products? When I mentioned more development around HSCM in the literature review, the authors could expand on these ideas. This would give more depth to their discussion.\nI suggest that the authors consult these two references, as they may help the authors to develop some of the ideas in the literature review: Reference: (1,2)\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? Partly\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.) Yes", "responses": [] }, { "id": "333492", "date": "12 Nov 2024", "name": "Abdüssamet Polater", "expertise": [ "Reviewer Expertise Supply chain management", "Logistics 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\nComments for the Abstract The aim of the study is confusing in the abstract section. It is stated that “This study aims to assess how the application of BDA impacts the performance of healthcare supply chain management (HCSCMP).” Then, investigating the efficiency and success factors are stated as an aim of the study. Comments for the Introduction “The study began with a descriptive analysis of academic research on BDA in the context of SCM, followed by a content analysis to assess the impact of BDA-based management systems on HCSCMP and to identify the key enablers and challenges for implementing BDA in HCSC.” The location of this sentence can be changed in the introduction sentence. The paragraph which starts with “Big Data Analytics (BDA) in healthcare is transformative…” is fine until the “Healthcare facilities manage both structured and unstructured data.” sentence. Then suddenly the authors starts to talk about the “structured and unstructured data”. Is there any need to talk about “structured and unstructured data” here? The paragraph starting with the “Adopting BDA in HCSCs facilitates real-time service delivery, data-driven decision-making, and improved performance (Araz et al., 2020).” should be reorganized. “Further research is needed to explore how BDA can enhance healthcare supply chain management performance (HCSCMP) and to validate existing findings.” can be the last sentence and the later sentences can be integrated as drivers of HSCP. “RQ1. What is the number of academic studies on Big Data Analytics in the context of Supply Chain Management, and what research methods and data collection techniques have been used in these studies?” This RQ can be shortened such as telling “investigating the trends in application of BDA in the HSCM”. “RQ2. How does the application of Big Data Analytics enhance efficiency in Healthcare Supply Chains Management according to existing studies?”. According to the abstract this study aims to investigate efficiency and performance of HSCM. But efficiency is only mentioned in the RQ2. The paragraph which is located after research questions should be significantly reorganized. I assume that the authors wanted to talk about the contribution of the study. However, it sounds more the findings or may be conclusion section. Comments for the Big Data and Big Data analytics in Healthcare Supply Chain Management This section mainly talks about the big data. But it does not discuss big data in Healthcare Supply Chain Management. Comments for the Organizational Information Process Theory Why did you specifically created a section titled as “Organizational Information Process Theory”? Comments for the Methodology What is the reason of starting to screen in 2016? What are the keywords used for screening? Which journals (science category, index type etc.) were included? How did you analyze the papers included? Comments for the Results analysis Please only use Figure 2 or Figure 3. “Research methods and data collection techniques have been used in big data analytics in the context of supply chain management studies”. Please add “health” before “supply chain management studies”. I did not understand to relating and discussing the country and research method (Table 1, Figure 5). Could you explain the importance of relating these two different findings? Comments for the Discussion The discussion section seems the repetition of the results section. In order to overcome this problem, it would be better to synthesize and discuss the “Efficiency Enhancements of Big Data Analytics in Healthcare Supply Chains” and “Enablers and Challenges in Implementing Big Data Analytics in Healthcare Supply Chains” sections under one title. Reviewer Decision I suggest the authors revise and develop the sections based on the comments.\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? Partly\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\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": [ { "c_id": "12827", "date": "26 Nov 2024", "name": "دينا النعيمي", "role": "Author Response", "response": "Abstract: Thank you for your valuable feedback. I have revised the abstract to clarify the study's aim, emphasizing that assessing the impact of BDA on performance, along with examining efficiency improvements and success factors, are interconnected objectives of the research. This should make the focus of the study clearer in the abstract. Location of Sentence on Descriptive Analysis and Content Analysis: Thank you for this suggestion. We will reposition the sentence, “The study began with a descriptive analysis of academic research on BDA in the context of SCM, followed by a content analysis to assess the impact of BDA-based management systems on HCSCMP and to identify the key enablers and challenges for implementing BDA in HCSC,” to improve the flow of the introduction. This will help clarify the structure and methodology of the study within the introductory context. After research questions guide the study: Discussion of Structured and Unstructured Data: We will revise the paragraph that starts with “Adopting BDA in HCSCs facilitates real-time service delivery, data-driven decision-making, and improved performance,” so that it concludes with, “Further research is needed to explore how BDA can enhance healthcare supply chain management performance (HCSCMP) and to validate existing findings.” This will allow the later sentences to flow naturally into a discussion of drivers in healthcare supply chain performance (HCSCP), following your helpful suggestion. Reorganizing the Paragraph on Adopting BDA: We will revise the paragraph that starts with “Adopting BDA in HCSCs facilitates real-time service delivery, data-driven decision-making, and improved performance,” so that it concludes with, “Further research is needed to explore how BDA can enhance healthcare supply chain management performance (HCSCMP) and to validate existing findings.” This will allow the later sentences to flow naturally into a discussion of drivers in healthcare supply chain performance (HCSCP), following your helpful suggestion. Rephrasing Research Question 1: RQ1 : What are the trends in the application of BDA in HCSCM? Clarifying Efficiency in RQ2: Thank you for pointing out this discrepancy. We will modify the abstract to clarify that both efficiency and performance are central aims, ensuring consistency with RQ2’s focus on efficiency within healthcare supply chain management. Reorganizing the Paragraph Following the Research Questions: We recognize that the paragraph after the research questions may seem more like conclusions or findings. To clarify, we will revise this section to focus on the study's intended contributions and will remove or relocate content that discusses specific findings. Comments for the Big Data and Big Data analytics in Healthcare Supply Chain Management Section Revised Comments for the Organizational Information Process Theory Thank you for your question. The 'Organizational Information Process Theory (OIPT)' section was included to provide a theoretical foundation for understanding how Big Data Analytics (BDA) supports healthcare supply chain management. OIPT explains how organizations can enhance their decision-making and operational efficiency by improving their information processing capabilities. Given that BDA is fundamentally about managing, analysing, and utilizing large volumes of data, OIPT offers a relevant lens to explore BDA’s role in healthcare supply chains. This theoretical framework helps explain how BDA can optimize information flow, reduce uncertainty, and support real-time decision-making, which are crucial for successful supply chain management in healthcare. Methodology: Reason for Starting the Screening in 2016: The screening period starts from 2016 because of a noticeable increase in academic research on Big Data Analytics in healthcare supply chains around that time. Starting in 2016 ensures the inclusion of recent and relevant studies, aligning with the growing interest and advancements in this area. The screening period begins in 2016 to capture recent developments and a noticeable rise in research on Big Data Analytics in healthcare supply chains during this period. Keywords Used for Screening: Keywords likely included terms related to big data analytics, supply chain management, and healthcare, such as “Big Data Analytics,” “Supply Chain Performance,” “Healthcare Supply Chain Management,” and others relevant to enablers and challenges in BDA implementation. The keywords used included ‘Big Data Analytics,’ ‘Supply Chain Performance,’ ‘Healthcare Supply Chain Management,’ along with terms related to enablers and challenges in implementing BDA. Which journals (science category, index type etc.) were included? The review focused on peer-reviewed journals indexed in recognized databases like Scopus and Google Scholar, specifically in areas such as healthcare management, supply chain management, and information systems. We included peer-reviewed journals indexed in major databases like Scopus and Google Scholar, focusing on healthcare management, supply chain management, and information systems. How did you analyze the papers included? The analysis involved a systematic literature review methodology. The papers were screened, selected, and synthesized based on their contributions to understanding Big Data Analytics’ impact on healthcare supply chains, including descriptive and content analysis to identify key findings, enablers, and challenges. The included papers underwent descriptive and content analysis to synthesize insights on BDA’s impact on healthcare supply chains, focusing on efficiency improvements and identifying key enablers and challenges for successful implementation. Results analysis: Comments for the Results analysis Only use Figure 2 or Figure 3. Response to Reviewer: Thank you for your feedback. I have revised the analysis to include only Figure 2, as suggested. “Research methods and data collection techniques have been used in big data analytics in the context of supply chain management studies”. Please add “health” before “supply chain management studies”. Additionally, I have updated the phrase to read, ‘Research methods and data collection techniques have been used in big data analytics in the context of healthcare supply chain management studies,’ to clarify the healthcare focus of this review.\" Discussing the country and research method (Table 1, Figure 5). Could you explain the importance of relating these two different findings? Thank you for highlighting this point. The purpose of relating countries with research methods in Table 1 and Figure 5 is to showcase regional preferences and methodological diversity in Big Data Analytics (BDA) research within healthcare supply chains. This comparison helps reveal how different regions prioritize certain research methods—such as questionnaires in North America and interviews in Australia—reflecting regional contexts, available resources, and academic traditions. By understanding these differences, we gain insight into the approaches that researchers in various regions use to investigate BDA’s impact on healthcare supply chain management, which could indicate unique challenges, cultural influences, or operational focuses in each region. This contextual understanding enhances the comprehensiveness of our review by demonstrating how geographic and methodological factors may shape research outcomes and perspectives on BDA implementation. Comments for the Discussion Response to Reviewer: Thank you for this helpful suggestion. To enhance the discussion and avoid redundancy, we will integrate the ‘Efficiency Enhancements of Big Data Analytics in Healthcare Supply Chains’ and ‘Enablers and Challenges in Implementing Big Data Analytics in Healthcare Supply Chains’ under a unified title, such as ‘Impact of Big Data Analytics on Healthcare Supply Chain Performance: Efficiency, Enablers, and Challenges.’ This synthesis will allow us to discuss both the benefits and obstacles of BDA implementation within a single framework, creating a more streamlined and interpretive analysis. By reorganizing this way, we can directly relate the efficiency improvements BDA offers to the specific enablers and challenges that influence its successful adoption. This will provide a more holistic view of BDA’s impact on healthcare supply chains, enabling readers to understand how these factors work together to enhance or inhibit supply chain performance." } ] } ]	1	https://f1000research.com/articles/13-1237
https://f1000research.com/articles/13-1523/v1	16 Dec 24	{ "type": "Systematic Review", "title": "Dengue infection during pregnancy and the occurrence of pathological neonatal outcome: a systematic review and meta-analysis", "authors": [ "Evelyn del Socorro Goicochea-Ríos", "NELIDA MILLY OTINIANO", "Lola del Carmen Rojas-Infantas", "Víctor Raú Ocaña-Gutiérrez", "Néstor Iván Gómez-Goicochea", "Evelyn del Socorro Goicochea-Ríos", "Lola del Carmen Rojas-Infantas", "Víctor Raú Ocaña-Gutiérrez", "Néstor Iván Gómez-Goicochea" ], "abstract": "Background Dengue infection during pregnancy increases the risk of maternal and neonatal complications; therefore, the objective of this research is to determine these outcomes and describe the clinical manifestations of the infection.\n\nMethods A systematic review of studies published in PubMed, MEDLINE, LILACS, Web of Science, Scopus and thesis repositories published between 2013 and October 2023 was performed. DeCS and MeSH dengue and maternal-neonatal outcome were used. Thirteen studies were selected and the New Castle-Ottawa scale was used to assess their quality. Mantel-Haenszel hazard ratios were calculated to report the overall effect size using random-effects models. All analyses were performed in Rev Man 5.4.1\n\nResults The 13 studies involved a population of 18,724 pregnant women, with cohorts ranging from 25 to 17,673 pregnant women. The most frequent outcomes in the pregnant women were cesarean section and postpartum hemorrhage, and in the newborns, preterm delivery and low birth weight. According to the New Castle-Ottawa scale, six studies were considered low risk and seven studies moderate risk. Dengue is a risk factor for postpartum hemorrhage (OR: 2.24), premature rupture of membranes (OR: 1.04) and cesarean section (OR: 1.13). It could not be concluded that dengue is a risk factor for the neonatal outcomes studied. The clinical picture of pregnant women with dengue was predominantly fever, abdominal pain, vomiting and nausea, anemia, dyspnea and myalgia.\n\nConclusions Pregnancy-related changes in the immune, cardiovascular and coagulation systems, among others, increase the probability of adverse maternal and neonatal outcomes in case of DENV infection, such as postpartum hemorrhage, premature rupture of membranes, cesarean section, low birth weight and preterm delivery. Pregnant women should be considered a population at risk and should be included in dengue control, diagnosis and treatment policies.", "keywords": [ "Dengue", "pregnancy", "pregnancy complications", "infant", "newborn", "pregnancy outcome", "neonatal outcome." ], "content": "Introduction\n\nDengue is a rapidly spreading tropical disease transmitted by mosquitoes of the genus Aedes, the most common being Aedes aegypti. The two most common forms of the disease affecting humans are dengue fever and severe dengue haemorrhagic fever.1 Globally, the incidence of dengue is high in Southeast Asian countries (70% of the global burden of this disease), as well as in the Eastern Mediterranean and Western Pacific regions.1 In the Americas, epidemic outbreaks are reported every 3 to 5 years, and the most affected countries include Central America, Mexico, Brazil, Venezuela, Peru, Colombia and Bolivia.2 The number of cases of severe dengue has also increased markedly, especially in Brazil, Colombia, Peru, Bolivia and Mexico.3\n\nIn the Americas, up to the epidemiological week 24-2023, 2,102,848 cases of dengue were reported, of which 39.3% were laboratory-confirmed and 0.15% were severe dengue cases, with a cumulative incidence rate of 214 cases/100,000 population. The highest number of dengue cases was observed in Brazil with 1,515,460 cases, followed by Peru with 169,504 cases and Bolivia with 133,452 cases.3 Furthermore, dengue virus infection (DENV) is hyperendemic and should be addressed as a public health problem.4 Affected countries are challenged to maintain the dengue case fatality rate < 0.05%. Therefore, timely diagnosis, early identification of warning signs, and appropriate treatment are important to avoid severe cases and deaths.3\n\nDENV infection is classified into dengue without alarm signs and dengue with alarm signs. In both cases, nausea and vomiting, rash, arthralgias, positive serological tests in population with other confirmed cases may occur. In the severe form, alarm signs include abdominal pain, mucosal bleeding, blood extravasation, general condition, hepatomegaly > 2 cm, and rapid decrease in platelets.5 When dengue occurs in pregnant women, the risk to mother and child increases. Some studies have described the association between maternal dengue and pathologic neonatal outcomes as prematurity, low birth weight,6–8 postpartum haemorrhage,9–11 miscarriage, and increased rates of caesarean delivery.12 Severe dengue infection can be difficult to differentiate from HELLP syndrome or gestational thrombocytopenia13 and, as mentioned, can be complicated by postpartum hemorrhage.11 The association of dengue infection with adverse fetal outcomes remains unclear,11,13 but prematurity, growth retardation and stillbirth can occur, especially in cases of severe maternal infection. According to other authors, the clinical picture of the newborn appears to be independent of the degree of severity of maternal dengue.6\n\nTo date, research has been published reporting that DENV represents an ongoing threat to the population in endemic areas, particularly pregnant women who face more severe complications from this virus. These complications can severely affect pregnancy outcome, as it is associated with haemorrhage, fetal loss and preterm delivery.14 It has also been documented that pregnant women are at higher risk of severe infection and of developing haemorrhagic fever/shock syndrome compared to non-pregnant women of reproductive age, with an OR of 3.38 and 95% CI. In the case of neonates, severe infection with sepsis-like symptoms and acute respiratory distress is documented.15\n\nVertical transmission of DENV has been described in studies that analysed placental tissue from patients with confirmed infection during pregnancy, as well as immunohistochemical findings of monoclonal antibodies to dengue virus.10 The risk of this transmission varies between 18.5% and 56.2% when infection is confirmed two weeks before delivery and up to two days postpartum.13,16 Although maternal and neonatal outcomes depend on the severity of DENV infection, the prevalence of infection and the socioeconomic costs associated with it require an updated review of the subject to promote preventive measures based on scientific evidence.\n\nIt has been reported that most of the reported cases occur in the economically active population,3 and in Peru the most affected age groups are those aged 30-59 years and 18-29 years. In terms of sex, more than half of the cases correspond to the female sex2,4 which could mean that women of reproductive age acquire the infection during pregnancy.\n\nThere is considerable interest in identifying maternal-neonatal outcomes in patients with dengue infection during pregnancy,6,7 as a strategy for the timely detection of complications and to reduce the risk of death. However, there are few studies on the severity of the disease, clinical outcomes of pregnancy and delivery in pregnant women and their neonates.\n\nDue to the physiological changes inherent to pregnancy, pregnant women are at greater risk of adverse effects and pathological neonatal outcomes when dengue infection occurs, depending on its severity.8,9 Therefore, the questions addressed in this review are: is dengue infection during pregnancy associated with pathological neonatal outcomes? And, what is the impact of dengue severity on gestational outcomes?\n\nAs mentioned, the prevalence of dengue is high in different countries in the world3 and affects a large part of the population, including pregnant women and neonates. Therefore, the research questions formulated in this review investigate the relationship between DENV infection during pregnancy and pathological maternal-neonatal outcomes, as well as the clinical picture of dengue in pregnant women.\n\n\nMethods\n\nThis study was registered in the International Prospective Register of Systematic Reviews (PROSPERO, 2023) under number CRD42023453805 available from: https://www.crd.york.ac.uk/prospero/display_record.php? ID=CRD42023453805, and the recommendations of the Preferred Reporting Items for Systematic Reviews and Meta-Analyzes (PRISMA) were followed17 ( Figure 1).\n\nTo identify the studies, an electronic search was carried out in the databases PubMed, Scopus, LILACS, WOS, Google Scholar, as well as in the WHO database, medical school repositories, conference proceedings and congress abstracts. No language or country restrictions were applied. Studies published between January 2013 and October 2023 were included. The search terms used were: pregnancy, pregnant, gestation, gravidez, dengue infection, neonatal outcome, recém nascido, dengue infection, infección por dengue, Febre da Dengue, Febre Quebra-Ossos, Infecção pelo Vírus da Dengue, Febre Hemorrágica Dengue.\n\nThe search strategy was adapted to each database. In PubMed, MeSH terms and Entry terms were included for title and abstract and the Boolean operators OR, AND, as well as truncation were used: Pregnancy [Mesh] OR Gestation [tiab] OR Pregnan[tiab] AND Pregnancy [Mesh] OR Gestation[tiab] OR Pregnan[tiab]) AND Infant [Mesh] OR Newborn [tiab] OR Newborn Infant[tiab] OR Neonate[tiab] AND (Breakbone Fever [Mesh]) OR Classical Dengue Fever[tiab] OR Break-Bone Fever [tiab] OR Fever, Break-Bone [tiab]. In LILACS the DECs were used: mh:(“Dengue/CN” OR “Dengue/CL” OR “Dengue/CN” OR “Dengue/EP” OR “Dengue/PP” OR “Dengue/TM”) AND (EMBARAZO/CN) AND (NEONAT*)\n\nDuplicate citations were eliminated and all remaining articles containing the study variables in their titles and abstracts were reviewed.\n\nArticles that met the following inclusion criteria were included: case-control, cohort and cross-sectional full-text and open-access articles describing the clinical picture of dengue in pregnant women, as well as maternal and/or fetal/neonatal outcomes, such as cesarean section, abortions, preterm delivery, maternal hemorrhage, ICU admission and maternal mortality, stillbirths, low birth weight, small for gestational age and neonatal mortality.\n\nStudies with a sample size of less than 10 were excluded, as well as systematic reviews, articles without statistical analysis, paid articles, reviews, letters to the editor, duplicate publications and clinical cases.\n\nAn Excel table was used to record the main data of each study and the reasons for inclusion or exclusion. The selection criteria for the studies included articles whose population was pregnant women with a diagnosis of dengue (regardless of the degree of severity) and the neonates of these pregnant women, published between January 2013 and October 2023. Articles that studied exposure to DENV infection were reviewed. Experimental interventions and pharmacological treatments were excluded.\n\nThe primary outcomes for the pregnant women were: a) clinical condition/comorbidity, b) complications: postpartum fever, postpartum hemorrhage, admission to the ICU, abortion and cesarean delivery. For neonates, primary outcomes included prematurity, low birth weight, small for gestational age, asphyxia, vertical transmission and admission to ICU. Secondary outcomes included maternal and neonatal mortality.\n\nSelection of studies and quality assessment\n\nThe Rayyan program18 was used to collect all studies related to dengue during pregnancy and neonatal outcomes. All identified articles were independently reviewed by two reviewers for their inclusion in the study. Titles and abstracts of potentially relevant articles were reviewed in the first round, and the full text of those articles referring to dengue in pregnancy were retrieved and reviewed in detail to assess eligibility. Any disagreement between the two reviewers was resolved by a third reviewer until consensus was reached.\n\nThe data from each study were extracted independently by two authors, who performed a cross-check to eliminate errors. The included studies were organized in a standard template to record all relevant information, which included the surname of the first author, year of publication, country of origin, study design, place of patient recruitment, age, method of diagnosis, number of individuals included (case and control groups), sample size, number of pregnant women with dengue, dengue infection detection technique, gestational age, comorbidities (hypertensive disease of pregnancy, gestational diabetes) and clinical characteristics (fever, myalgia, malaise, dyspnea, diarrhea and joint pain). We also recorded outcomes related to type of delivery, premature rupture of membranes, postpartum fever, abortion, pre- and postpartum hemorrhage, and fetal/neonatal data: preterm delivery, low birth weight, fetal asphyxia, fetal/neonatal death, small for gestational age, and vertical transmission.\n\nIn the event that the requisite results for analysis were not forthcoming, the article was excluded in order to preclude the potential for reporting bias.\n\nThe RevMan software (Review Manager 5.4.1)19 was employed for the analysis of the collected data. The odds ratios with a 95% confidence interval were utilised, and the results were pooled using the Mantel-Haenszel model. The heterogeneity of the effect size was estimated using the I2 statistic, with the following ranges: I2 ≤ 25%, 25%–50%, and >75% (corresponding to low, moderate, and high, respectively).\n\nIn all cases, the probability values of less than 0.05 were deemed to be statistically significant. Furthermore, a leave-one-out sensitivity analysis was conducted to evaluate the influence of each individual study on the statistical outcomes. The risk of bias of the included studies was evaluated by two investigators using the Newcastle-Ottawa Scale (NOS) and its subscales. The assessment was conducted independently and considered three domains: selection of study groups, comparability of groups, and determination of exposure or outcome. The scoring system was applied to case-control and cohort studies, respectively.20 The studies were classified according to the level of bias present, with categories of low (8-9 points), moderate (5-7 points), and high bias (0-4 points). The strength of evidence for all the outcomes analysed was evaluated using the GRADE tool, which is a recognised method for assessing the quality of evidence in systematic reviews.21\n\nA descriptive synthesis of the results was written, recording number of cases, percentages in tables for each maternal and neonatal outcome For maternal and neonatal outcomes, the OR, CI, and heterogeneity described in each included study were included. These data were analyzed using the version non-Cochrane of Revman 5.4.1.19,22 The data were tabulated in Excel. Plots from Rev Man19 were also used to visually present the results of the individual studies and the syntheses. No subgroup analysis was performed\n\nThe grouped prevalence was calculated using the random-effects model. The results of the analysis of maternal-neonatal outcomes were presented as odds ratios with 95% confidence intervals. For some outcomes, high heterogeneity was found, so each study was analyzed independently.\n\n\nResults\n\nThe preliminary searches of the databases consulted identified a total of 651 studies. After eliminating 34 duplicate articles, 617 studies were screened, of which 502 were excluded because they corresponded to other study designs, had restricted access, dealt with other outcomes, were systematic reviews, among other reasons. We reviewed the full text of 115 articles that included the variables of interest for the study in the titles and abstracts. Of these, 99 articles were discarded because they did not meet the eligibility criteria. Finally, 13 articles that met the inclusion criteria were selected and their analysis is included in this review ( Figure 1).\n\nOf the 13 included studies, 8 (62%) are retrospective cohort, 3 (23%) are prospective cohort and 2 (15%) are case-control. Of these studies, 5 were published in Latin America, 3 in Asia, 3 in India and 2 in Africa (Table 1 extended data). These studies reported data on a total of 18,724 DENV-infected pregnant women. Ninety-two percent of the studies focused on the clinical picture and maternal outcome, while 100% addressed neonatal outcome.\n\nThe most reported outcomes in the included studies were cesarean section and postpartum hemorrhage in pregnant women, as well as preterm delivery and low birth weight in neonates. Regarding the characteristics of the population, the sample size ranged from 25 to 17673 pregnant women. Dengue diagnosis was performed using the NS1 test (in 11 of the 13 studies) and IgM (in 12 of the 13 studies). According to the New Castle-Ottawa Scale score (NOS), six low-risk studies and seven moderate-risk studies were identified (Table 1 extended data).\n\nOf the included studies, 100% meet the “selection” category established by NOS for both cohort studies and case-control studies. Forty-eight percent met the “comparability” category and 70% met the “exposure/disclosure” category ( Figure 2).\n\nOnly the studies by Basurko 15(2018), Friedman 25 (2014), Machain 23 (2018), Nujum 29 (2019), Paixao 14 (2019) and Sagili 31(2022) meet all the categories established by NOS, therefore presenting low risk of bias ( Figure 3).\n\nDengue was found to be a risk factor for postpartum haemorrhage (OR: 2.23) and caesarean section (OR: 1.39). In the case of neonatal outcomes, it cannot be concluded that dengue is a risk factor for low birth weight and preterm delivery due to OR <1 and high heterogeneity of the included studies (≥ 97%).\n\nIn the clinical picture of DENV infection, the following findings predominated: fever (7 studies); abdominal pain, vomiting and nausea and hypertensive disease of pregnancy (5 studies); anemia (4 studies), dyspnea, bleeding, myalgia and gestational diabetes (3 studies).\n\n\nDiscussion\n\nOf the 13 studies selected, it was found that more than 90% of pregnant women with dengue presented fever,12,16,23–27 followed by abdominal pain,15,16,23,26,28 vomiting and nausea,12,16,23,26,28 anemia,25,27,29,30 dyspnea16,28,26 and myalgia12,23,26 as the most frequent symptoms. Other publications also mention fever as the most frequent symptom,31,32 accompanied or not by headache, myalgias, arthralgias and other symptoms,33–35 although the clinical picture depends on the severity of dengue.31 Among the most frequent comorbidities were hypertensive disease of pregnancy15,27–29,36 and gestational diabetes,15,29,36 both pathologies referred to as frequent in patients with severe dengue.37\n\nRegarding maternal outcomes, 5 retrospective cohort studies report cesarean section,14,16,23,25,36 as does a retrospective cross-sectional study26 and the 2 case-control studies27,30 report postpartum hemorrhage,15,26,28,30,36 miscarriages,28,30,36 maternal deaths,16,23,26,28,36 and premature rupture of membranes.15,30 To a lesser degree, ICU admission,16,28 thrombocytopenia,26 oligohydramnios,28 and antepartum hemorrhage are reported.15 However, dengue alone was shown to be a risk factor for postpartum hemorrhage and cesarean section (OR >1) ( Table 2).\n\nRegarding newborn outcomes, fetal or neonatal death,15,16,25–28,30,36 is reported as the most frequent outcome, followed by pre-term delivery,12,15,27,29,30,36 low birth weight,12,14,27,29,30 small for gestational age,14,15,30 admissions to the ICU,16,27,30 hepatomegaly, neonatal asphyxia, neonatal infection, and vertical transmission.24 However, it is inconclusive that dengue is a risk factor for low birth weight, pre-term delivery, and small for gestational age, (OR < 1), in addition to the high heterogeneity of the included studies (I2 >97%).\n\nOther research reports that no relationship was found between dengue infection and preterm delivery.38 In other studies, the risk of pre-term delivery was higher in dengue seropositive than seronegative groups. in all trimesters of pregnancy (OR: 1-26, 95% CI 1-06-1-49, p=0-006).39 Spontaneous abortion linked to dengue infection could not be analyzed because the studies corresponded to different designs. On the other hand, there was no significant association between maternal dengue infection and fetal death in two cohort studies involving a total of 572 participants. (grouped RR 3.42, 95% CI 0.76-15.49, I2 = 54.8%).38 This contrasts with another investigation in which dengue infection was found to be associated with high maternal and perinatal mortality.40\n\nMaternal dengue was shown to be associated with a slightly increased risk of preterm birth and low birth weight, although the CI was borderline. This review did not identify a notable correlation between dengue infection during pregnancy and low birthweight (LBW). However, the findings indicated that dengue haemorrhagic fever during pregnancy was associated with a twofold increased risk of preterm birth (OR 2.4, 95% CI 1.3 to 4.4) and an elevated likelihood of LBW (OR 2.1, 95% CI 1.1 to 4.0). The study found no correlation between maternal dengue and low weight for gestational age.41\n\nDuring pregnancy, there are changes in the immune, cardiovascular and coagulation systems, as well as liver enzymes and febrile response. The white blood cell count is usually elevated, with a leftward shift, and the platelet count is usually low. Therefore, the changes produced by dengue may go unnoticed, which justifies the need to clarify this diagnosis when a pregnant woman presents with fever.42,43 Some pathophysiological mechanisms have been proposed for dengue infection For example, the up-regulation of proinflammatory cytokines (interleukin 6 (IL-6), interleukin 8 (IL-8) and tumour necrosis factor alpha (TNF-α)) may result in alterations to normal gestational physiology. It is important to be aware that thrombocytopenia, plasma leakage or bleeding tendency may affect placental circulation and cause fetal complications. In severe dengue infection, there is a risk of endothelial damage and increased vascular permeability, which could allow DENV to cross the placental barrier.42\n\nDuring the normal gestation period, there is an overall increase in innate immune cells and effector mechanisms, as well as an increase in complement activity. Furthermore, there is a notable rise in the number of circulating phagocytes and plasmacytoid dendritic cells (DCs) producing type I interferon (TNF-1). It has been demonstrated that the innate pathways responsible for defending against retroviruses are specifically enhanced during pregnancy. To illustrate, IFN-induced activation of signal transducer and activator of transcription 1, a vital response to viral infection that is increased in natural killer cells, monocytes and myeloid dendritic cells (DCs) during gestation.43 The innate antiviral response to DENV is regulated by a variety of immune cells and inflammatory mediators. When in balance, these allow for effective viral clearance, resulting in either asymptomatic infections or mild clinical manifestations. An imbalance between the components of the immune response can lead to increased immune activity and disease severity.44\n\nIt is thought that placental tropism, changes in innate immunity and vascular permeability during normal pregnancy may contribute to an increased susceptibility to severe DENV infection in pregnant women. in combination with excessive or insufficient maternal innate immune system activity. The effects of DENV infection include a reduction in the number of megakaryocytes, which can lead to thrombocytopenia. Additionally, there is an increase in the expression of interferon-induced transmembrane protein 3 in platelets, resulting in the release of type I IFN. This could contribute to the increased incidence of haemorrhagic complications during pregnancy. There is evidence that RNA virus infections are connected with an elevated risk of miscarriage, haemorrhage and foetal death.43\n\nIt has been described that, at the maternal-fetal interface, NK cells, macrophages, DCs and T cells can modify their function and secretome during viral infections. These changes can lead to complications such as premature birth, congenital malformations, miscarriage or fetal death, and hematoplacental transmission.4,43\n\nComplications vary according to the time of gestation. In the first trimester there is an increased risk of pre-eclampsia and eclampsia, especially in cases of severe dengue. Plasma leakage is the result of inflammatory cytokines, DENV non-structural protein-1 (NS1) and inflammatory lipid mediators have been identified as factors that increase capillary permeability. It is possible that maternal compensatory mechanisms for pathologic plasma leakage and vasodilatation are a contributing factor in the development of preeclampsia. In addition, the decrease in pulse pressure to 25 mm Hg or less, along with shock and dengue fever, increases capillary permeability. This further increases the risk of acute pulmonary oedema for infected mothers. Decreased plasma volume may also contribute to bleeding in dengue, although there is no specific evidence for the third trimester.41,45\n\nThe role of hormones in dengue complications in pregnant women is not completely clear. In the first trimester, leukopenia may not be evident. Due to the physiological effects of haemodilution and haemoconcentration with plasma leakage, this may be masked in cases of dengue. Accordingly, an elevated hematocrit may be indicative of maternal shock. In the second trimester thrombocytopenia may occur, and a decline in platelet levels <80 × 100/L may suggest plasma leakage. Decreased oncotic pressure and associated pulmonary resistance may lead to acute pulmonary edema. In the third trimester, the risk of vertical transmission of the virus is greatest,42,46 and during the postpartum period, there is evidence of transmission through breastfeeding.42,45\n\nIn this review, we found that most articles report performing cesarean section in patients with dengue (10/13). On this point, the studies favor conservative treatment, unless there is some obstetric reason to intervene since a cesarean section may be precipitated in an unstable patient with anesthetic risks (such as spinal anesthesia), bleeding tendency or risk of hemorrhage during the intervention. Before term, there appears to be little indication for induction of labor or other obstetric interventions.47\n\nPregnant women represent a high-risk group for serious complications associated with dengue infection, and it is estimated that maternal mortality is three times higher in pregnant women than in non-pregnant women with dengue. Thrombocytopenia and elevated liver enzymes c are associated with certain disorders of pregnancy, such as preeclampsia, HELLP syndrome (hemolysis, elevated liver enzymes and low platelets) or gestational thrombocytopenia, so it is crucial to make an accurate diagnosis by confirmatory serological tests for dengue.\n\nSeveral studies report thrombocytopenia as one of the most common clinical findings48–50 in pregnant women with dengue fever, affecting between 40% and 70% of these patients. Severe hypovolemic shock, caused by severe hemorrhage and preeclampsia, could be favored by the state of plasma leakage. The severity of the disease could significantly influence the occurrence of maternal complications.10 On the other hand, it has been observed that patients with dengue during the first trimester suffered spontaneous abortions, possibly due to inflammatory changes in the placenta.45\n\nThe effects of dengue on the fetus or newborn appear to be variable. Apparently, fetal damage is less in early pregnancy, when there is sufficient time for maternal antibodies to transfer to the newborn. In contrast, if maternal infection occurs late in gestation, the risk of pathological neonatal outcomes increases.47 Placental endothelial damage and increased placental vascular permeability influence adverse fetal outcomes.51 In addition, vertical transmission from mother to foetus can cause developmental abnormalities, growth retardation, preterm birth and stillbirth. At the same time, the mother is predisposed to increased morbidity and death.52,53\n\nAs mentioned, DENV induces pathological changes related to increased production of proinflammatory cytokines42,54 which can affect the uterus by stimulating the production of proteins that promote uterine contractions, potentially leading to preterm delivery.55 In addition, among these proinflammatory cytokines is TNF-α, which is related to bleeding, favoring thrombocytopenia and endothelial dysfunction56,57; in addition, antibodies are produced that cross-react with plasma proteins, platelets and the endothelium, which also leads to hemorrhagic symptoms.56\n\nTherefore, the thrombocytopenia, plasma leakage or bleeding tendency inherent to the disease could cause damage to the placental circulation, with adverse effects on the fetus, including the risk of stillbirth.55 In addition, endothelial damage and increased vascular permeability make it easier for DENV to cross the placental barrier.42\n\nThe association between dengue and pre-term delivery10,54 as well as low birth weight has been documented in relation to the severity of the disease.55 It is frequently observed that both outcomes occur in pregnant women affected by dengue, which can have serious effects on the health and development of the child. Furthermore, the occurrence of stillbirths, especially in women with severe dengue, lends support to this hypothesis.54 Likewise, the relationship between dengue and intrauterine growth retardation is also reported. A meta-analysis including six cohort studies and two case-control studies found an association with pre-term delivery (OR 1.71 (95% CI 1.06-2.76), between miscarriages in pregnancies and dengue infection compared to those without infection (OR 3.51; 95% CI 1.15-10.77) However, no significant association was found with low birth weight.10\n\nIt is important to note that some studies showed an increased rate of prematurity and fetal/neonatal death,10,54 while others have found no significant fetal effects, such as preterm delivery, fetal death, low birth weight42 or other maternal complications, such as miscarriage and maternal hemorrhage. This suggests the need for further research to clarify these discrepancies.47 Available studies indicate that adverse maternal and perinatal outcomes are more frequent in women with severe forms of dengue, especially when infection occurs near the end of gestation.49,58,59\n\nIt is possible that placental ischaemia could have an impact on foetal growth. A study conducted in Brazil indicates that capillary leak syndrome and capillary permeability may result in inadequate vascular supply to the foetus. This could potentially lead to the development of hypoxic lesions, such as involvement of the trophoblastic epithelium or chorangiosis, as well as inflammatory lesions affecting the decidua or choriodecidua. Hypoxia has been shown to be sufficient to reduce fetal growth rates.45\n\nWeaknesses of this research include the fact that more than 50% of the selected studies have a moderate risk according to the New Castle-Ottawa Scale (NOS). In addition, the studies on neonatal outcomes have varied research designs and high heterogeneity, and 4 of the 13 studies included small samples (less than 50 cases). We also found no evidence on the virus serotype affecting pregnant women and its influence on maternal and neonatal outcomes. The existence of asymptomatic cases and the difficulties associated with accurate diagnosis of acute dengue make it challenging to define the risks associated with DENV infection during pregnancy. This is due to the fact that dengue IgM can cross-react with other flaviviruses, and co-infections are common.43\n\nWe consider that these results show risk factors for pathological outcome in DENV infections during pregnancy, both for the pregnant woman and for the newborn. It is essential to take these factors into consideration in the application of preventive measures and in monitoring the evolution of the disease. It also reinforces the need to hospitalize any pregnant woman with dengue symptoms and to manage the newborn of a mother with dengue as a high-risk case. Clinical, laboratory and ultrasound monitoring of pregnant women with dengue is crucial to detect possible adverse outcomes and improve clinical practice.\n\nGiven that in this review the studies do not provide evidence on the influence of a history of previous dengue infection on maternal-neonatal outcomes or on the impact of the serotype of the virus affecting pregnant women, it is recommended that homogeneous cohort or multicenter studies be conducted to generate better evidence regarding the role of dengue as a risk factor for pathological outcomes.\n\n\nConclusions\n\nThe susceptibility generated by the physiological changes of pregnancy increases the probability of adverse outcomes in maternal, fetal and neonatal morbidity and mortality. Therefore, given the high prevalence of this disease worldwide, it is of utmost importance to establish more efficient guidelines for the detection, diagnosis and treatment of DENV infections, as well as to implement personalized care programs for the obstetric population. It would also be important to include in dengue control policies the recommendation to hospitalize all pregnant women with dengue fever for monitoring until the disease has been eliminated.\n\nHowever, it remains to be established which are the initial and most determinant elements in the immunopathogenesis of dengue during pregnancy. A better understanding of these events will allow the development of effective therapeutic measures to prevent severe forms of the disease in pregnant women. It is crucial to consider the temporal/spatial component in the evolution of clinical cases in future studies, as this will improve our understanding of the relevance of each effector of the immune response in DENV infections and immunopathogenesis during pregnancy.\n\n\nEthics and consent\n\nEthical approval and consent were not required.\n\n\nAuthors’ contributions\n\nEvelyn Goicochea-Ríos: Conceptualization, formal analysis, methodology, data collection, investigation, visualisation, writing – original draft preparation, writing – review & editing, funding acquisition.\n\nNélida Milly Otiniano: Methodology, data collection, formal analysis, writing – original draft preparation, writing – review & editing.\n\nVíctor Ocaña Gutiérrez: Data collection, formal analysis, methodology, writing – original draft preparation, writing – review & editing.\n\nLola Rojas Infantas: Data collection, formal analysis, methodology, writing – review & editing.\n\nNéstor Gómez Goicochea: Data collection, formal analysis, methodology, writing review & editing.", "appendix": "Data availability\n\nNo data are associated with this article.\n\nZenodo: Dengue infection during pregnancy and the occurrence of pathological neonatal outcome. A systematic review and meta-analysis, 10.5281/zenodo.14278897.60\n\nThis project contain the following underlying data:\n\n1. Flow chart PRISMA Dengue in pregnancy.jpg\n\n2. Database with results Dengue in pregnancy.xlsx\n\n3. PRISMA_2020_checklist RS dengue in pregnancy.docx\n\n4. RS DENGUE IN PREGNACY RESULTS AND NOS (1) (2).xlsx\n\n5. Table 1 Dengue infection during pregnancy and the ocurrence of pathological neonatal outcome.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\nRahman MS, Mehejabin F, Rahman MA, et al.: A case-control study to determine the risk factors of dengue fever in Chattogram, Bangladesh. Public Health Pract (Oxf). 2022; 4: 100288. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRiesgo de presentación de brotes de gran magnitud de dengue, en regiones del país por fenómeno del Niño Costero. CODIGO: AE- CDC- N° 10 – 2023. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nde Andrade Vieira Alves F , Nunes PCG, Arruda LV, et al.: The innate immune response in DENV- and CHIKV-infected placentas and the consequences for the fetuses: A minireview. Viruses. 2023; 15(9). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPaixão ES, Teixeira MG, Costa M d CN, et al.: Dengue during pregnancy and adverse fetal outcomes: a systematic review and meta-analysis. Lancet Infect. Dis. 2016; 16(7): 857–865. PubMed Abstract \| Publisher Full Text\n\nBrasil P, Lupi O: Dengue infection during pregnancy and risk of preterm birth. Lancet Infect. Dis. 2017; 17(9): 885–886. PubMed Abstract \| Publisher Full Text\n\nPardo Martínez D, Ojeda Martínez B, Alonso Remedios A: Dinámica de la respuesta inmune en la infección por virus del dengue. Medisur. 2018 [citado el 22 de septiembre de 2024]; 16(1): 76–84. Publisher Full Text\n\nCastro-Mussot M, Machain-Williams C, Loroño-Pino M, et al.: Respuesta inmune e immunopathogenesis en las infecciones con el virus del dengue. Gac. Med. Mex. 2013; 149: 531–540. PubMed Abstract Reference Source\n\nTien Dat T, Kotani T, Yamamoto E, et al.: Dengue fever during pregnancy. Nagoya J. Med. Sci. 2018; 80(2): 241–247. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAgarwal K, Malik S, Mittal P: A retrospective analysis of the symptoms and course of dengue infection during pregnancy. Int. J. Gynaecol. Obstet. 2017; 139(1): 4–8. PubMed Abstract \| Publisher Full Text\n\nGoicochea Rios E d S: Dengue infection during pregnancy and the occurrence of pathological neonatal outcome. A systematic review and meta-analysis. F1000Res. 19 de noviembre de 2024. Publisher Full Text" }	[ { "id": "357117", "date": "20 Jan 2025", "name": "Phuc Nhon Nguyen", "expertise": [ "Reviewer Expertise Obstetrician at the Department of Pathologic Pregnancy" ], "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\nThank you for your work. I have some concerns: - In abstract, the OR should be provided with 95%CI. - Serotypes of DENV should be provided in introduction. - No language or country restrictions were applied. The search terms used were: pregnancy, pregnant, gestation, gravidez, dengue infection, neonatal outcome, recém nascido, dengue infection, infección por dengue, Febre da Dengue, Febre Quebra-Ossos, Infecção pelo Vírus da Dengue, Febre Hemorrágica Dengue. I concern if the same publication in different language. English search should be only applied. - Results should be expanded with more findings. - Table 1 is missing. - Table 2 should be presented as Forest plot. - Perinatal transmission should be mentioned in discussion. Pham TH, et al., 2023 (Ref 1) - Similar study had been reported. Rathore SS, et al., 2022 (Ref 2) - Recent systematic study included 20 studies. The authors could discuss on this point “While maternal dengue infection was associated with a higher prevalence of preterm birth and LBW, the associations were not statistically significant. Significant associations were observed for stillbirth in specific studies.” Shabil M, et al., 2024 (Ref 3) - Conclusion is very common. Please focus the main findings before recommendation. For ex: “Pregnancy-related changes in the immune, cardiovascular and coagulation systems, among others, increase the probability of adverse maternal and neonatal outcomes…” or “The susceptibility generated by the physiological changes of pregnancy increases the probability of adverse outcomes in maternal, fetal and neonatal morbidity and mortality” - Full-word should be provided by the abbreviation word when it first appears in each part. For example: DeCS, HELLP syndrome - English editing is required. For example: This sentence should be reconstructed. “Given that in this review the studies do not provide evidence on the influence of a history of previous dengue infection on maternal-neonatal outcomes or on the impact of the serotype of the virus affecting pregnant women, it is recommended that homogeneous cohort or multicenter studies be conducted to generate better evidence regarding the role of dengue as a risk factor for pathological outcomes.”\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? Partly\n\nIs the statistical analysis and its interpretation appropriate? Partly\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/13-1523
https://f1000research.com/articles/13-566/v1	03 Jun 24	{ "type": "Research Article", "title": "Exploring α−ψ−ϕ contractive mapping: novel fixed point theorems in complete b-metric spaces", "authors": [ "Tamene Raji", "Nasir Ali", "Gudeta Hanchalu", "Fikadu Tesgera Tolasa", "Berhanu Seboka", "Tamene Raji", "Nasir Ali", "Gudeta Hanchalu", "Berhanu Seboka" ], "abstract": "Background This paper explores α-ψ-ϕ contractive mappings, extending the field of self-map and fixed-point theorems.\n\nMethods We analyze α-ψ-ϕ contractive mappings using rigorous mathematical proofs and logical deductions.\n\nResults A key main result is established, supported by intuitive corollaries and practical examples, highlighting the applicability of our findings.\n\nConclusions Our work provides a fresh perspective on contractive mappings, simplifying complex mathematical concepts and enriching the literature on fixed-point theorems.", "keywords": [ "complete b-metric space", "Altering distance function", "α-admissible maping" ], "content": "1. Introduction\n\nBanach's principle of contraction theory, as elucidated in Ref. 1, has served as a cornerstone for numerous research endeavors, facilitating expansions across various journals. The ubiquity of this theorem underscores its significance in extending the frontiers of mathematical discourse. Such expansions often entail imposing contraction restrictions and additional constraints on the ambient spaces, thereby giving rise to a diverse array of structures, including b-metric spaces.2,3 The notion of b-metric spaces, an extension of conventional metric spaces, has been instrumental in enriching the landscape of mathematical investigations.3–6 This versatile framework has found application in a myriad of contexts, contributing to discussions on topological structures and yielding insights into the interplay between fixed and common fixed points.7–9\n\nIn this context, our research aims to address the pressing need for a deeper understanding of α-ψ-ϕ contractive mapping within the framework of b-metric spaces.10 The fundamental question we seek to explore is how the principles of contraction theory can be extended to encompass this novel mapping concept. This investigation is motivated by the observation that while traditional contraction mappings have been extensively studied, the exploration of more generalized forms, such as α-ψ-ϕ contractive mappings, remains relatively uncharted territory. Our work not only seeks to fill this gap in the literature but also aims to contribute to the advancement of mathematical theory by introducing and elucidating the properties of α-ψ-ϕ contractive mappings. Through a meticulous synthesis of internet sources, scholarly publications, and pertinent literature, we establish the theoretical framework for this novel concept.\n\nThe novelty of our approach lies in the integration of α-ψ-ϕ contractive mappings into the framework of b-metric spaces, thus expanding the scope of both contraction theory and metric space theory. This novel synthesis opens up new avenues for exploration and offers fresh insights into the dynamics of fixed point theorems in non-standard spaces. In pursuit of our objectives, we present a comprehensive analysis, bolstered by corollaries, illustrative examples, and rigorous proofs, in support of our main result. By elucidating the intricacies of α-ψ-ϕ contractive mapping within the context of b-metric spaces, we aim to provide a solid foundation for further research and to inspire new avenues of inquiry in this vibrant field of mathematical exploration.\n\n\n2. Preliminaries\n\nThis section serves as a foundational platform for our subsequent analysis, comprising key definitions, illustrative examples, essential notations, and fundamental theorems. These elements collectively provide the necessary groundwork and contextual understanding required to establish and prove our main result.\n\nLet X be a non-empty set and s≥1 be a given real number. A function d:X×X→[0,∞) is a b-metric on X if the given below principles are apply;\n\n(i) d(x,y)=0 iff x=y, ∀ x,y∈X;\n\n(ii) d(x,y)=d(y,x), ∀ x,y∈X;\n\n(iii) d(x,z)≤s[d(x,y)+d(y,z)], ∀ x,y,z∈X.\n\nThen, b-metric space a pair of (X,d) with coefficient s.\n\nLet X be any non empty set. Let f:X→X be a self mapping on X and α: X × X →[0,∞) be mapping.\n\nWe say that f is α-admissible if α(x,y)≥1⇒α(fx,fy)≥1 for all x,y∈X.\n\nConsider X={0,1,2,3}, Let f:X→X and α:X×X →[0,∞) and d(x,y)=\|x−y\| such that f0=0,f1=2,f2=1,f3=3, and {α(x,y)=1,if(x,y)∈{(0,1),(0,2),(1,1),(2,2),(1,2),(2,1),(1,3),(2,3)}α(x,y)=0,other wise. Then f is α-admissible.\n\nConsider X as any set that is not empty. If f:X→X and α: X× X→[0,∞) are mappings, then f is triangular α-admissible mapping if\n\n(i) f is α-admissible mapping;\n\n(ii) {α(x,y)≥1α(y,z)≥1⇒α(x,z)≥1, for all x,y,z∈X.\n\nLet X=[0,∞), f:X→X and α:X×X →[0,∞) are mappings and d(x,y)=\|x−y\|2\n\nWe define fx={x22,x∈[0,1]x+1,x>1 and α(x,y)={1,x,y∈[0,1]0,other wise\n\nThen, f is triangular α-admissible.\n\nA function ψ :[0,∞)→[0,∞) is referred to as an altering distance function if the given below principles are apply;\n\n(i) ψ is continuous and non decreasing;\n\n(ii) ψ (t)=0 if and only if t=0.\n\nwhere, Ψ is the set of all altering distance functions.\n\nLet ψ: [0,∞)→[0,∞) given by;\n\nψ(t)={t7,t<3t2+3t2+4t+7,t≥3, then ψ is an altering distance function.\n\nAssuming that a b-metric space (X,d) with s equal to or greater than one and f:X→X are given, then sequence {xn} with xn=fnx0=fxn−1 for every n∈ℕ and x0∈X is called a picard sequence.\n\nSuppose that a b-metric space (X,d) with s equal to or greater than one and f:X→X are given. If a picard sequence {xn} of initial point x0∈X fulfills;\n\nWhenever m,n∈ℝ+ and n<m. Then {xn} is a Cauchy sequence.\n\nAssuming that a complete b-metric space (X,d) with s equal to or greater than one and f: X→ X are given\n\nfor each x,y∈X,ifm,n∈ℝ+ and n<m.\n\nThen, a fixed point of f appears.\n\n\n3. Main Results\n\n\n\nAssuming that (X,d) is a b-metric space with s equal to or greater than one and that α:X×X→[0,∞) are given, then f:X→X is referred to as “α-ψ-ϕ contractive mapping” if some ψ, ϕ ∈Ψfor all x,y∈X with α(x,y)≥1 implies that\n\nAssume that (X,d) is a complete b-metric space with s greater than or equal to one, and mappings f:X→X and α:X×X→[0,∞) are given;\n\nLet’s the given below principles are apply;\n\n(i) f is α-ψ-ϕ Contractive mapping;\n\n(ii) f is triangular α-admissible mapping;\n\n(iii) let x0∈Xinvolves α(x0,fx0)≥1; and\n\n(iv) f is continuous mapping.\n\nThen, a fixed point of f appears\n\nLet x0∈X be given as (iii), i.e. α(x0,fx0)≥1.\n\nFrom the definition (2.6) xn+1=fxn and xn=fxn−1 for every n∈ℕ. If xno= xno+1 for no∈ℕ, and fxno= xno,then xno is a fixed point of f.\n\nNext, xn Let is not equal with xn+1, then d(xn,xn+1) is also not equal to zero for every n belongs to ℕ\n\nDue to the triangular α-admissible mapping in (ii);\n\nα(x0,fx0)=α(x0,x1)≥1⟹α(fx0,fx1)≥1; for each x0,x1∈X.\n\nα(x1,fx1)=α(x1,x2)≥1⟹ α(fx1,fx2)≥1; for each x1,x2∈X.\n\nby continuing in this process we get;\n\nα(xn,fxn)=α(xn,xn+1)≥1⟹α(fxn,fxn+1)≥1 for each n∈ℕ.\n\nFrom definition (3.1), obtain;\n\nThus, from (3.2) we have;\n\nBut, d(xn−1,xn+2)≤ s[d(xn−1,xn)+d(xn,xn+2)]\n\nSince, ψ is continuous and increasing, we obtain;\n\nSince, the sequence {d(xn,xn+1)} is limited from the lower. Therefore ∃ r ≥ 0 implies limn→∞d(xn,xn+1)=r.\n\nWe want to show r = 0.\n\nSuppose that r > 0.\n\nNow, n→∞ of the two side of (3.4), gives:\n\nSince, r > 0, s ≥ 1 and n < m, n,m∈ℝ+\n\nThis implies 2rs+4s2r+n2rs+4s2r+m∈[0,1) and 2rs+4s2r+n2rs+4s2r+m< 1\n\nr≤2rs+4s2r+n2rs+4s2r+mr<r, a contradiction.\n\nHence, limn→∞d(xn,xn+1)=0 (3.5). So {xn}is a Cauchy sequence by (3.4) and Using proposition (2.1).\n\nLet’s now demonstrate the fixed point f;-\n\n(∃z) (z∈ X) implies limn→∞d(xn,z)=0 from a complete b-metric space\n\nSo, using definition (3.1), with x=xnandy=z, gives;\n\nTaking the limit as n→∞ of the two side of (3.6), gives:\n\nBy the limit’s uniqueness and the Complete b-metric space it gives;\n\nHence, the fixed point of f is z.\n\nAssume that (X,d) is a complete b-metric space with s greater than or equal to one, and that f:X→X and α:X×X→[0,∞) are given.\n\nLet’s the given below principles are apply:-\n\n(i) f is α-ψ-ϕ Contractive mapping;\n\n(ii) f is triangular α-admissible mapping;\n\n(iii) x0∈Ximplies (x0,fx0)≥1;\n\n(iv) if {xn} in X implies (xn,xn+1) is equal to or greater than one for every n∈ℕ and xn→x∗∈X as n→∞, then there exist a subsequence {xnk} of {xn} so that α(xnk,x∗)≥1for every k∈ℕ.\n\nThen, a fixed point of f appears.\n\nProof: Proceed analogously to the proof of first theorem and use (iv) (xnk,x∗)≥1 for a subsequence {xnk} of {xn} and k∈ℕ, we get;\n\nTaking the limit as k→∞ on two side of (3.7), gives:-\n\nBy uniqueness of limit and Complete b-metric space we have;\n\nHence, the fixed point of f is x∗.\n\nLet f:X→X and α:X×X→[0,∞) are the predetermined mappings, while (X,d) is a complete metric space;\n\nLet’s the given below principles are apply:-\n\n(i) f is α-ψ-ϕ Contractive mapping;\n\n(ii) f is triangular α-admissible mapping;\n\n(iii) let x0∈Ximplies α(x0,fx0)≥1; and\n\n(iv) if {xn} in X implies (xn,xn+1)≥1 for every n∈ℕ and xn→x∗∈X as n→∞,\n\nFor some subsequence {xnk} of {xn} so that α(xnk,x∗)≥1 for every k∈ℕ.\n\nThen, a fixed point of f appears\n\nIf s is one, From Theorems 3.1 and 3.2 One finds this consequence.\n\nIf X = {0,3,4}, d:X×X →[0,∞) is given by d(x,y)=\|x−y\|2,(X,d) is a complete b-metric space with s = 2, and also α:X×X →[0,∞) and f:X→Xare given as follows; {α(x,y)=1,ifx,y∈{0,3,4}α(x,y)=0,other wise\n\nFirst, show that f is α-admissible mapping.\n\nIf x = 0, y=3, α(0,3)≥1⇒α(f0,f3)=α(0,3)=1≥1\n\n⇒f is α-admissible mapping.\n\nSecondly, If x = 0, y=3andz=4, the given equation holds.\n\n⇒ f is triangular α-admissible.\n\nLastly, check that f is α-ψ-ϕ Contractive Mapping;\n\nFrom, () and (*), ψ(2d(f0,f3))≤ψ(M(0,3))−ϕ(M(0,3))\n\n18 < 219\n\n⇒ f is α-ψ-ϕ Contractive Mapping.\n\n\nConclusion\n\nIn conclusion, this research has introduced the concept of α-ψ-ϕ contractive mapping within b-metric spaces, expanding upon contraction theory principles. Through rigorous analysis and illustrative examples, we've highlighted its significance and applicability. Future directions include exploring stability in iterative methods, investigating connections with other mathematical areas, and extending the analysis to broader classes of spaces and mappings. Overall, this exploration of α-ψ-ϕ contractive mappings opens new avenues for research with broad potential implications.\n\n\nAuthors contribution\n\nAll the authors equally contributed towards this work.", "appendix": "Availability of data and materials\n\nThe data is provided on request to the authors.\n\n\nReferences\n\nAli N, Siddiqui HMA, Riaz MB, et al.: A graph-theoretic approach to ring analysis: dominant metric dimensions in zero-divisor graphs. Heliyon. 2024; 10: e30989. Publisher Full Text\n\nGüngör GD, Altun I: Some Fixed Point Results for α-Admissible Mappings on Quasi Metric Space Via θ-Contractions. Mathematical Sciences and Applications E-Notes. 2024; 12(1): 12–19.\n\nGuan H, Li J: Common Fixed-Point Theorems of Generalized (ψ,ϕ)-Weakly Contractive Mappings in b-Metric-Like Spaces and Application. Journal of Mathematics. 2021; 2021: 1–14. Publisher Full Text\n\nEke KS, Olaoluwa HO, Olaleru JO: Common Fixed Point Results for Asymptotic Quasi-Contraction Mappings in Quasi-Metric Spaces. International Journal of Mathematical Sciences and Optimization: Theory and Applications. 2024; 10(1): 115–127.\n\nShatanawi W, Shatnawi TA: Some fixed point results based on contractions of new types for extended b-metric spaces. AIMS Mathematics. 2023; 8(5): 10929–10946. Publisher Full Text\n\nHuang M, Wang X, Wang Z, et al.: Locally biHölder continuous maps and their induced embeddings between Besov spaces.2023.\n\nRaji M: Generalized α-ψ contractive type mappings and related coincidence fixed point theorems with applications. The Journal of Analysis. 2023; 31(2): 1241–1256. Publisher Full Text\n\nMebawondu A, Izuchukwu C, Aremu K, et al.: On some fixed point results for (α,β)-Berinde-φ-Contraction mapppings with applications. International Journal of Nonlinear Analysis and Applications. 2020; 11(2): 363–378.\n\nKumar BVS, Bodaghi A: Approximation of Jensen type reciprocal functional equation using fixed point technique. Boletim da Sociedade Paranaense de Matemática. 2020; 38(3): 125–132. Publisher Full Text\n\nCzerwik S: Contraction mappings in b-metric spaces. Acta Mathematica et Informatica Universitatis Ostraviensis. 1993; 1(1): 5–11.\n\nSamet B, Vetro C, Vetro P: Fixed point theorems for α–ψ-contractive type mappings. Nonlinear Analysis: Theory, Methods & Applications. 2012; 75(4): 2154–2165. Publisher Full Text\n\nSingh D, Joshi V, Kim JK: Some Best Proximity Point and Fixed Point Theorems via Generalized Cyclic Contraction in G-Metric Spaces. Global Journal of Pure and Applied Mathematics. 2015; 11(6): 5203–5222.\n\nFelhi A, Aydi H, Zhang D: Fixed points for α-admissible contractive mappings via simulation functions. Journal of Nonlinear Sciences and Applications. 2016; 9(10): 5544–5560.\n\nJi D, Yu Y, Li C: Fixed point and endpoint theorems of multivalued mappings in convex $ b $-metric spaces with an application. AIMS Mathematics. 2024; 9(3): 7589–7609. Publisher Full Text\n\nYamaoda O, Sintunavarat W: Fixed point theorems for (α, β)-(ψ, ϕ)-contractive mappings in b-metric spaces with some numerical results and applications. Journal of Nonlinear Sciences and Applications. 2016; 09(1): 22–33. Publisher Full Text\n\nGüngör GD, Altun I: An Existence Result for Second-Order Boundary-Value Problems via New Fixed-Point Theorems on Quasi-Metric Space. Symmetry. 2024; 16(1): 99. Publisher Full Text\n\nMerdaci S, Hamaizia T: New common fixed point theorem for multi-valued mappings in b-metric spaces." }	[ { "id": "320355", "date": "27 Sep 2024", "name": "Tanakit Thianwan", "expertise": [ "Reviewer Expertise Fixed point theory" ], "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\nREPORT FOR THE PAPER I reviewed the document and identified the following mistakes:\nIn the keywords section, \"maping\" should be \"mapping.\" In the definitions, some equations lack proper spacing between variables and symbols, making them harder to read. Page 4 of 11, line-2, Inconsistent use of semicolons, such as in \"Let’s the given below principles are apply;\" which should be rephrased for clarity. Page 4 of 11, line-2, \"Let’s the given below principles are apply\" should be \"Let the following principles apply.\" The language used in the introduction is formal and appropriate for an academic paper. However, there are minor grammar issues, such as in the sentence \"Through a meticulous synthesis of internet sources, scholarly publications, and pertinent literature, we establish the theoretical framework for this novel concept,\" where \"internet sources\" may not be the best fit for a research paper. Replacing it with \"previous studies\" or \"existing research\" might be more suitable. Ensure that all symbols and notation are consistent across the entire document. For example, double-check that the use of ψ and ϕ functions is consistent in both definitions and examples. Furthermore, there are many typos. The authors should be revised carefully.\nContent Accuracy: The introduction provides a solid background but would benefit from a clearer and more concise explanation of how α-ψ-ϕ contractive mappings expand upon traditional contraction theory. It would be helpful to explain why the generalization to b-metric spaces is significant for real-world applications.\nSuggestion: Include a brief mention of practical applications of these mappings in areas like data science or machine learning to ground the mathematical theory in broader fields.\nIn addition, the following references should be added.\nBaewnoi K, et al., 2024 (Ref 1).\nChairatsiripong C, et al., 2023 (Ref 2)\nYambangwai, D., et al., 2024 (Ref 3)\nPaimsang P, et al., 2023 (Ref 4)\nBy citing these works, helps establish the background and significance of introducing an alternative method, as the properties and applications of nonexpansive mappings in Banach spaces have strong implications for contractive mappings in b-metric spaces.\nIn essence, these citations establish a comprehensive framework for fixed point theory and iterative methods, which is crucial for understanding and situating the novel contributions made by exploring α−ψ−ϕ contractive mappings in complete b-metric spaces. They showcase related iterative techniques, their convergence behaviors, and their applications across different spaces, thereby providing both a theoretical foundation and practical motivations for the new theorems being developed.\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": "12629", "date": "16 Dec 2024", "name": "Nasir Ali", "role": "Author Response", "response": "Dear Reviewer, Thank you for your thorough review and valuable feedback on our manuscript. We appreciate your constructive comments and have carefully addressed each one in our revised submission. Below is a summary of the revisions made in response to your suggestions: Keyword Correction: We corrected the typo in the keywords section by changing \"maping\" to \"mapping.\" Spacing in Equations: We reviewed all equations and ensured that proper spacing between variables and symbols is applied for better readability. Clarity and Grammar: The sentence \"Let’s the given below principles are apply\" has been rephrased to \"Let the following principles apply\" for clarity. We reviewed the manuscript for consistent use of semicolons and corrected them where needed to ensure uniformity and clarity in punctuation. Introduction Refinement: As per your suggestion, we replaced \"internet sources\" with \"previous studies\" to maintain the formal tone appropriate for an academic paper. Additionally, we provided a clearer and more concise explanation of how α−ψ−ϕ contractive mappings expand upon traditional contraction theory, and why their generalization to b-metric spaces is significant for real-world applications. Consistency in Symbols and Notation: We carefully reviewed the use of all symbols and notations throughout the document, including the ψ\\psiψ and ϕ\\phiϕ functions, and ensured consistency in both definitions and examples. Grammar and Typos: We have carefully proofread the manuscript to eliminate any remaining typos and improve overall grammar. Content Accuracy and Practical Applications: We revised the introduction to more clearly explain the importance of generalizing contraction theory to b-metric spaces and included a brief discussion on the practical applications of these mappings in fields such as data science and machine learning, grounding the mathematical theory in broader, real-world contexts. Addition of References: We have added the suggested references: Baewnoi K, et al., 2024 (Ref 1). Chairatsiripong C, et al., 2023 (Ref 2). Yambangwai, D., et al., 2024 (Ref 3). Paimsang P, et al., 2023 (Ref 4). These references strengthen the theoretical framework and provide important background for understanding the significance of introducing α−ψ−ϕ\\alpha-\\psi-\\phiα−ψ−ϕ contractive mappings in complete b-metric spaces. They also establish connections to related iterative techniques and their convergence behaviors. We have uploaded the revised version of the manuscript, incorporating all these changes. We believe these revisions have improved the clarity, consistency, and overall quality of the paper, and we hope that it now meets the expectations of the review process. Thank you again for your insightful comments and suggestions. Sincerely," } ] }, { "id": "320358", "date": "09 Oct 2024", "name": "Konsalraj Julietraja", "expertise": [ "Reviewer Expertise Applied Mathematics", "Fixed point theory", "Chemical Graph Theory" ], "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 \"Exploring α−ψ−ϕ Contractive Mapping: Novel Fixed Point Theorems in Complete b-Metric Spaces\" delves into advanced fixed point theory, focusing on the development of novel contractive conditions involving α, ψ, and ϕ mappings in the framework of complete b-metric spaces. The authors aim to generalize and extend classical fixed-point theorems, such as the Banach Contraction Principle, by introducing new types of contractive conditions, thus broadening the scope of potential applications in mathematics and its interdisciplinary fields. The article requires the following major revisions in order to make it suitable for indexing:\nThe authors should detail the contribution of the article in the abstract. The authors should improve the Introduction section with the necessary references. The authors should provide a clear definition of contractive mapping and α−ψ−ϕ contractive mapping: The authors should justify the role of ϕ. Theorem 3.1 - The statement is not clear. The authors should provide an unambiguous statement. The authors should include the comparison between Theorem 3.1 and 3.2 The authors should compare α−ψ Contractive Mapping and α−ψ−ϕ Contractive Mapping and explore the advantages of α−ψ−ϕ Contractive Mapping. The authors should cite the article entitled “Modified α-ψ-contractive mappings with applications Fixed Point Theory and Applications” 2013, 2013:151. The authors should justify the relevance of the references 1, 6, 9,17 in their article.\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?\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? Partly", "responses": [ { "c_id": "12630", "date": "16 Dec 2024", "name": "Nasir Ali", "role": "Author Response", "response": "Dear Reviewer, Thank you for your thoughtful and constructive feedback on our article, \"Exploring α−ψ−ϕ Contractive Mapping: Novel Fixed Point Theorems in Complete b-Metric Spaces.\" We appreciate your insights, which will help enhance the clarity and impact of our work. Below, we address each of your major revision points in detail: Contribution in the Abstract: We have revised the abstract to explicitly outline the key contributions of our article. It now clearly states how our novel contractive conditions extend classical fixed-point theorems and emphasizes the broader implications for mathematics and interdisciplinary applications. Improvement of the Introduction: The introduction has been enhanced with additional references that provide context and support for our research. We have incorporated relevant literature to establish a stronger foundation for our work. Definition of Contractive Mappings: We have added a clear and precise definition of contractive mappings, including a detailed explanation of α−ψ−ϕ contractive mappings. This section has been formulated to ensure that readers can easily understand the distinctions and relevance of these mappings within our framework. Justification of the Role of ϕ: A new subsection has been included to justify the role of the function ϕ in our contractive mappings. We explain how its inclusion adds complexity and depth to the mapping conditions, enabling more generalized applications and insights into fixed-point behaviors. Clarification of Theorem 3.1: We have rephrased Theorem 3.1 to provide a clearer and more unambiguous statement. The revised formulation emphasizes the key principles involved and their implications for the existence of fixed points. Comparison Between Theorem 3.1 and Theorem 3.2: A comparison between Theorems 3.1 and 3.2 has been added to the manuscript. This section discusses the similarities and differences in the conditions and implications of each theorem, highlighting their respective contributions to fixed-point theory. Comparison of α−ψ and α−ψ−ϕ Contractive Mappings: We have included a discussion comparing α−ψ and α−ψ−ϕ contractive mappings. This analysis explores the advantages of the α−ψ−ϕ framework in terms of flexibility and potential for broader applications in various mathematical contexts. Citation of Relevant Articles: We have cited the article “Modified α-ψ-contractive mappings with applications Fixed Point Theory and Applications” (2013:151) as recommended. We believe that this reference complements our discussion and strengthens our literature review. Relevance of References 1, 6, 9, and 17: Justification of References: Felhi, A., Aydi, H., & Zhang, D. (2016): This reference is critical as it discusses fixed points for α-admissible contractive mappings, which directly relates to the theoretical foundation of our work. The methodologies and results presented in this paper provide a comparative backdrop for our exploration of α−ψ−ϕ contractive mappings, emphasizing the evolution of fixed-point theory through varying mapping conditions. Güngör, G. D., & Altun, I. (2024): This article presents new fixed-point theorems applicable to quasi-metric spaces, showcasing the versatility and adaptability of fixed-point theory in different mathematical frameworks. Our inclusion of this reference serves to highlight the ongoing advancements in fixed-point theorems and their relevance to the broader mathematical community, particularly in how they relate to our findings in complete b-metric spaces. Ji, D., Yu, Y., & Li, C. (2024): The authors of this paper address fixed-point and endpoint theorems in convex b-metric spaces, which aligns closely with our study's context. This reference is particularly significant as it extends the applicability of fixed-point theory to multivalued mappings, reinforcing the importance of diverse mapping approaches in achieving generalizable results. It also helps position our work within a contemporary research landscape that seeks to unify various contractive conditions. Huang, M., Wang, X., Wang, Z., & Xu, Z. (2023): This research explores locally bi-Hölder continuous maps and their embeddings between Besov spaces, contributing to our understanding of continuity and its implications for fixed-point theorems. The techniques discussed in this paper may offer potential methodologies for further investigating the properties of α−ψ−ϕ contractive mappings. Including this reference enriches our discussion on the continuity conditions necessary for the application of fixed-point theorems in various mathematical settings. We hope that these revisions address your concerns and enhance the clarity and quality of our article. Thank you once again for your valuable feedback, and we look forward to your further comments. Sincerely," } ] } ]	1	https://f1000research.com/articles/13-566
https://f1000research.com/articles/13-421/v1	29 Apr 24	{ "type": "Systematic Review", "title": "A bibliometric review of functional ingredients and their efficacy in developing functional biscuits", "authors": [ "Kshama Vishwakarma", "Varalakshmi Chandra Sekaran", "Vidya Patwardhan", "Asha Kamath", "Kshama Vishwakarma", "Vidya Patwardhan", "Asha Kamath" ], "abstract": "Introduction Recent studies have found that functional biscuits are becoming increasingly popular as a convenient and nutritious food option. Numerous studies have concluded that the functional ingredients which impart functional properties to biscuits are beneficial for human consumption and can be incorporated into functional biscuits. However, there is a lack of information on essential functional ingredients that can be used to develop functional biscuits. This bibliometric study addresses this gap by identifying the current research trends in functional ingredients to develop functional biscuits.\n\nObjective To investigate current research trends on functional ingredients and their effectiveness in developing functional biscuits.\n\nMethod The study followed the “Scientific Procedures and Rationales for Systematic Literature Reviews” standards for retrieving literature. The study went through three major stages, “assembling”, “arranging”, and “assessing” to retrieve 612 articles from the Scopus database from 2013 to 2023. Through further filtering, 395 articles were selected.\n\nResult The analysis was conducted using R Studio and VOS viewer. The performance analysis and science mapping tools were used to evaluate the articles. The results showed a 5.76% annual growth in publication trends. The most researched functional ingredients were antioxidants, bioactive compounds, and dietary fiber. The review summarized the most studied foods to develop functional biscuits and highlighted the most experimented technological advancements.\n\nConclusion The study revealed an increasing interest in developing functional biscuits and the need for further research to improve their effectiveness and sensory properties. Further, there is a need to determine the impact of technology on extracting functional ingredients from foods, so they can be incorporated into functional biscuits.", "keywords": [ "“Antioxidants” “Functional ingredients”", "“Dietary fibers”", "“Functional biscuits/cookies”", "“Phenolic compounds”" ], "content": "Introduction\n\nThe year 2023 was celebrated as the 75th anniversary year of the World Health Organization with an announcement of the theme for the year 2023 as “Health for all”, which brings into focus the sustainable development goal (SDG 3) “Ensure healthy lives promoting well-being for all at all ages”.26 The theme promotes SDG 3 and highlights the importance of research on food with functional properties. Similarly, a market report on functional foods states that the functional food market is expected to generate revenue of USD 137.1 billion by 2026 at a CAGR of 6.8%.25 This socio-transformation has led to a higher response to functional foods drawing attention to the research on functional ingredients. Hasler et al (2002) described functional foods as “foods enriched with nutrients to add health benefits, not for balanced calorie content”.1,2 International Food Information Council (2006), the International Life Sciences Institute (1999), and Thomas & Earl (1994) have commonly accepted the definition of “functional foods” as “Foods or parts of foods that provide additional physiological benefits beyond their basic nutritional value”.3 In short functional foods can be summarized as foods that promote good health, enhance well-being, and improve quality of life. The functionality of such foods is enhanced by functional ingredients with the ability to promote or provide a positive influence on human health. These new types of ingredients that provide anticipated outcomes favoring health due to physiological changes to the human body are labeled “functional ingredients\"4 and may include carotenoids, flavonoids, dietary fibers, phenolic and allyl compounds and many more.\n\nMoreover, the influence of diet on the well-being of mankind and the role of functional foods in decreasing the risk of illness has led to growing acceptance, resulting in a remarkable era for new functional foods based on functional ingredients.5 Among the range of functional ingredients, dietary fiber, and phenolic compounds are two significant components found in plants with several physiological effects. Based on structure, phenolic acids, flavonoids, lignans, and stilbene differ and such substances are mentioned as functional ingredients as they offer benefits beyond just nutritional and energetic gains.4 A high-fat biscuit dominance in the market has led biscuits to the apparent choice when consumers are told to reduce their fat intake.6 Therefore, biscuits are often used as a medium to experiment with enriching protein and improving other nutritional aspects7\n\nHence, the research on readily available functional biscuits has become significant in the past few years, motivating producers to create novel, nutritious biscuits with health-enhancing qualities.\n\nThe purpose of the study is to analyze publications from 2013 until 2023 on functional ingredients and provide an overview of the literature related to functional ingredients. The paper aims to examine research trends in assessing the efficacy of biscuits as a medium to incorporate functional ingredients using bibliometric analysis.\n\nResearch questions are framed as below:\n\nRQ1. What publishing trend has been observed in recent years on functional ingredient research?\n\nRQ2. What is the contribution of the most influencing articles on functional ingredients?\n\nRQ3. What documents reveal in context to most prolific authors, influential articles, and contributing journals on functional ingredient research?\n\nRQ4. Which foods have been investigated the most to study functional ingredients in the last three years to develop functional biscuits?\n\nRQ5. Which functional ingredients are investigated the most by research publications?\n\nRQ6. What future advancements could be researched on functional ingredients to develop functional biscuits?\n\n\nMethods\n\nThe study predominantly adopts “Scientific Procedures and Rationales for Systematic Literature Reviews” (“SPAR-4-SLR”) standards for retrieving literature as illustrated in Figure 1.\n\nThe justification for choosing the method is due to the superior quality of SPAR-4-SLR over the PRISMA Guidelines. Moreover, the method follows three major stages to segregate articles such as “assembling”, “arranging”, and “assessing” articles.8\n\nTo detect and obtain the range of articles on functional ingredients, the study reviewed previous intellectual research work to get the required understanding. The alternate terminologies of functional biscuits, such as “enriched biscuits,” “functional cookies,” and “nutritional biscuits”, are included. The keywords “Functional biscuits” OR “Functional ingredients” generated 5187 documents in the first search approach. The second search approach followed the keywords “Functional biscuits” OR “functional ingredients” OR “Nutritional Biscuits,” which retrieved 5197 documents. The last approach used “functional biscuits,” “functional ingredients,” “Enriched Biscuits,” OR “Nutritional Biscuits,” generating 5265 documents. The document search was based on selected keywords, and the last approach was made on 17th February 2023 to collect articles. The search thread followed the title, abstract, and keywords to identify documents. The Scopus database was used to retrieve articles as this is the major database, and 612 documents were identified from the search process.\n\nPost the assembling step, the selected documents were arranged based on filter functions on the Scopus database as per “year, subject, document type, source type, publication stage, and language”. The search strategies were confined to “2013-2023, “Chemistry, Agricultural & Biological Science, Multidisciplinary, Social sciences, Health professionals and Psychology,” Articles, Review, Conference proceedings, final, journal, and English,” respectively. The process yielded 612 articles. Furthermore, the data was downloaded in “CSV” format. Each article was read with a special emphasis on abstract, findings, and conclusion yielding 395 corpora of articles for review using databases such as Elsevier, Sage, Springer, Emerald, Taylor & Francis, Sci Vi, Wiley Open access, and Google Scholar.9\n\nThe bibliometric analysis approach was employed to assess the collection of 395 articles on the functional ingredients research. Biblioshiny (Rstudio-R version 4.2.2) and VOSviewer (1.6.19) software were used as the assessment tool, using performance analysis and science mapping to achieve bibliometric analysis. The analysis by Biblioshiny reveals year-wise publications, most influencing articles, most contributing journals, and prominent authors under performance analysis. VOS viewer displays the major keyword themes found in functional ingredients research and co-citation among authors.10\n\n\nResults and Discussion\n\nThe performance of a specific research field is revealed through performance analysis.10 Additionally, this analysis outlines publications through the years, trends of publications, most influencing articles, top contributing authors, and the most relevant authors. Figure 2 displays the study tools used for analysis.\n\nThe total publication trend through 2013-2023 is represented in Figure 3. The publication trend from 2015–2022 displays an ascending trend in the number of articles published. The publication of articles increased close to double during the years 2021 & 2022. An upward graph has continued until 2022, although the annual growth rate remained fixed at 5.76%. In 2022, the number of published articles reached 76, and 2023 will likely see a further increase in research trends in functional ingredients.\n\nThe study examines diverse documents, including research on functional biscuits and ingredients. Table 1 represents publications with the keywords “functional ingredients,” “functional biscuits,” and “Nutritional biscuits.” The displayed statistics highlight 84.31% of documents in a category as published articles. Publications as review papers constitute 61% and 1% as conference papers.\n\nTable 2 represents the most influential articles on functional ingredients research reaching a total number of citations above 75. It is observed that the study of11 titled “HPLC-DAD-ESI-MS/MS screening of bioactive components from Rhus coriaria L. (Sumac) fruits.” is the most prominent study on functional ingredients research published in the Food Chemistry with 331 total citations and 36.8 total citations per year. The article by12 with a total of 288 citations, secured the second spot. Similarly, the top ten most cited articles have accumulated 2113 citations. The ten most influential articles have studied sumac, peanuts, bananas, banana peels, brown algae, microalgae, mango peel, wheat, and oat as natural sources of bioactive compounds, dietary fiber, phenolic compounds, antioxidants, flavonoids, carotenoids, etc. Table 3 enlists food investigated by the most influential articles. All the studied foods have shown the highest potential to develop new biscuits as a medium to incorporate functional ingredients.\n\nTable 4 displays the most contributing journals in functional ingredient research. It is evident that” Food Chemistry” and “Food & Function” journals stand first and second with “97” and “40” significant contributions, respectively. Interestingly both journals have amassed total citations “4144” and “1329” and stand at “39” and “19” h index individually with their higher contributions.\n\nTable 5 and Figure 4 exhibit the most prolific authors in functional ingredients. It has illustrated that “Barros L” and “Ferreira ICFR” are the most prominent authors in this field, with ten publications each. The most prominent authors, Barros L, 2022 and Ferreira ICFR, 2022 investigated wild edible mushrooms and concluded that they can be a source of nutritional and functional components. Hence, wild edible mushrooms can be incorporated into a balanced diet as a source of proteins and can be utilized for innovative bio-based formulation. Frias J, 2020; Martínez-Villaluenga C, 2020 studied barley grain (flour) and found the ideal method of germinating grain for 3.5 days, maintaining the temperature at 16°C.for producing nutrient-rich and functional barley flour. It is worth emphasizing that these articles have studied various foods like rosemary, xoconostle fruit (opuntia mature scheinvar cv. Rosa) by-products, tarragon, and pineapple by-products for their phenolic and bioactive compounds as well as techniques like fermentation, ultra-sonication, and biochemical and molecular profiling. Table 6 displays a summary of foods investigated by prominent authors.\n\nTable 7 summarizes articles from 2021-2023 that investigate foods for their potential use in developing functional biscuits. Recent studies have examined the potential of Amla and Apple pomace, Thyme, Tannat grape skin, Jujub flour, Bee pollen, Chickpea, Lemon peel, and lemon pomace to add functionality to new biscuits. The studies have demonstrated positive results regarding the acceptability and quality of cookies/biscuits in terms of functionality.\n\nTo understand a particular research subject through visual depiction is conducted by using science-mapping analysis. This evaluation involves several types of analysis, including thematic, factorial, temporal, and network analysis. The study has utilized Network analysis to elucidate the primary themes concerning functional ingredients research.\n\nNetwork analysis was conducted by applying co-authorship mapping and co-occurrence mapping using VOS viewer software.\n\nTo examine the co-authorship link minimum of 10 authors with a max 75-citation threshold were selected. Out of 396 authors, 30 fulfilled the criteria. The mapping demonstrated that all top authors with the highest citation of 331(red cluster) and lowest citation of 77 (grey cluster) had zero total link strength among themselves.\n\nDuring co-occurrence mapping, all keywords that were ministered by the full counting method were considered as part of the analysis. To enhance accuracy, the study implemented certain parameters to perform analysis to avoid repetition of keywords. Additionally, the lowest of five instances were set to select the keywords. As a result, out of 1149 keywords, 32 satisfied multiple criteria after data cleaning.\n\nThe clustering technique is used to focus on three clusters out of five clusters. Figure 5 demonstrates the network visualization that appeared through scientific articles. The cloud diagram illustrates the frequency of a word in the article and its correlation with other keywords. Each term is depicted as a frame in the diagram, and the size of the frame represents the number of times the term appears in publications. Different colors represent the frames grouped into clusters. The curved lines indicate the proximity of the terms, while the thickness of the lines reflects the strength of the connection between pairs of topic areas or keywords. The clusters provide insight into the relationship between different topics.\n\nThree clusters out of five, red, green, and blue, are more prominent than the rest. The red areas represent topics related to antioxidant properties, by-products of food processing, developing functional foods, biscuits, and techniques.\n\nThe green clusters comprise functional ingredients, antioxidant activity, bioactive peptides, bioaccessibility, encapsulation, and physiochemical properties. Among the repeatedly occurring keywords, antioxidant activity tops the list with 33 occurrences.\n\nThe blue cluster highlights functional properties, dietary fiber, rheology, polyphenols, texture, and bread.\n\nThe software displays the keywords that have the most frequent occurrences alongside other keywords displayed in Table 8. “Occurrence” refers to the number of articles that feature the keyword. Antioxidant activity, functional ingredients, Antioxidants, Dietary fiber, Phenolic compounds, Bioactive compounds, Polyphenols, Antioxidant capacity, Functional properties, and Functional foods are the most highly co-occurring keywords with occurrence weights (total link strength) as 33(25),31(22),23(12),20(21),20(21),18(19),18(20),17(20),14(13), 13(13).\n\nVOS viewer can show three different mapping visualizations from Figure 5 (network visualization), Figure 6 (overlay visualization), and Figure 7 (density visualization) are created.\n\nFigure 6 displays developments related to keywords over the researched periods. The colors of the keyword frames depict the researched period. The yellow cluster represents the keywords from studies published from 2020 onwards. Antioxidant activities, health benefits, and bioaccessibility are observed as the most occurred keywords in recent studies, including germination, bread, texture, and flour.\n\nThe depth of researched areas can be seen in Figure 7. Deep concentration of colors interprets topics undertaken by more researchers to study. The most deliberated topics by researchers are functional ingredients, antioxidant activity, bioactive compounds, phenolic compounds, antioxidants, polyphenols, dietary fiber, functional foods, flavonoids, phenols, and bioaccessibility.\n\n\nDiscussion\n\nThis bibliometric analysis of articles retrieved from Scopus could successfully examine the efficacy of biscuits as a medium to incorporate functional ingredients. The results matched with past studies that have proved biscuits as an effective medium to incorporate functional ingredients4,7\n\nA bibliometric analysis approach also highlighted present trends, evolution, and scope for upcoming advances in functional ingredient research from a global perspective.\n\nPublication trends: Although the selected publications’ issue year is 2013, the publication trend gained impetus from the year 2015. The reasoning for the same could be associated with (United Nations, 2015b), seventeen sustainable development goals (SDGs) in the 2030 agenda for sustainable development, adopted by United Nations Member States in 2015. The increase in research publications on functional ingredients could be justified as a result of SDG goals, the second goal being “Zero Hunger,” and the third “Good Health and Well-being”.13 The year 2022, with 76 publications and an annual growth of 39.5%, shows rising importance in functional ingredient research worldwide.\n\nThe most influential article, “HPLC–DAD–ESI-MS/MS screening of bioactive components from Rhus coriaria L. (Sumac) fruits, “turns out as the most contributing article with 331 citations (11) followed by “Encapsulation of food grade antioxidant in natural biopolymer by electrospinning technique: A physicochemical study based on the zein-gallic acid system.” (12) at 288 citations, sourced from Food Chemistry. The results show that “Food Chemistry” and “Food & Function” were among the most contributing journals, with 97 and 40 articles, respectively.\n\nIt is worth mentioning that both articles investigated techniques to extract bioactive compounds (211) from sumac and innovative gallic acid combined zein sub-micron fiber mat (packaging material) with antioxidant activity through an electrospinning technique, respectively. Both articles highlight the importance of technology and techniques to extract bioactive compounds from foods to develop functional materials for the packaging industry. Along the same line, Betoret's 2011 study discusses the use of Microencapsulation, Vacuum impregnation, and nutrigenomic techniques in developing functional foods. It highlights the importance of technologies to avert the weakening of bioactive compounds14\n\nAmong the most relevant authors, Barros L and Ferreira ICFR account for ten publications in food ingredients research which causes the expansion in research. The co-authorship was examined by using network analysis using VOS viewer software. Among the most cited 30 authors, zero total link strength was detected, indicating that authors have not collaborated in analyzed papers.\n\nFood investigated by influential articles- The top influential articles investigated foods such as sumac,11 peanuts,15 brown algae,16 microalgae,5 extruded wheat bran, and mango.4 These studies on foods open the doors to developing new functional biscuits and validate the claim made by14 that foods with functional properties signify a prospect of attaining pioneering products that could fulfill the current demand. The success of producing functional biscuits can be found in other studies done by7 on soy flour & rice bran,17 industry by-products like hemp,18 artichoke by-products and19 Finger Millet and Carrot Pomace. Similarly, a study by Mitreyski, 2023 highlights the finding and states that the significant rise in the popularity of readily consumable functional foods in recent years is prompting manufacturers to create novel, nutrient-rich products that benefit health.20\n\nThe data shows that the recent studies conducted from 2020 onwards have explored foods like (industrial waste) apple and amla, corncob, and Thyme, as well as upcycled defatted sunflower seed flour (industry by-products) for developing functional biscuits, validating the answers to framed research questions 1-4.\n\nMost researched functional ingredients- The review focuses on the status and trends in functional ingredient research. The network analysis emerges into five clusters “Functional ingredients,” “Antioxidant Activity,””Bioactive compound,””Phenolic compounds,” and “Dietary Fiber.” Among the keywords, “Antioxidant activity” tops with 33 occurrences, which restates the findings of a study by Yeasmen 2021, about “antioxidant activity” with 57 occurrences.21 The second highest among keywords analysis is “Functional ingredient,” with 31 occurrences justifying a growing trend toward considering the connection between food and health. An association between dietary fiber and phenolic compounds is proved by4 who claim that incorporating phenolic compounds and dietary fiber in food components not only enhances the performance of foods but is also used to develop functional foods that provide health advantages.\n\nRecent publications are found to focus their studies on “Bioaccessibility”, “Antioxidant properties”, “Health benefits”, “Flour”, “Germination”, “Bread”, and “Texture”. Yeasmen, 2021 study concludes that a fresh approach is required to evaluate bioaccessibility and bioavailability by studying the efficacy of leafy Phenolic compounds to access and utilize for commercial purposes.21 Barros L, 2022, Ferreira ICFR, 2022, and Mc Clements DJ, 2021 studies have assessed the association between natural food sources' antioxidant properties and health benefits. The finding by Faller (2023) and Brito (2021) has listed health benefits such as antidiabetic, anticancer, hepatoprotective, neuroprotective, anti-inflammatory, anti-arthritic, antibacterial, insecticidal effects, cardiovascular diseases, obesity, diabetes, and even cancer. The rational narrated answers to research question 5.\n\nThe integration of functional ingredients in developing new functional products is rising as studies have successfully incorporated various functional ingredients like phenolic compounds, bioactive compounds, flavonoids, antioxidants, etc. in bakery products. Although customers consider bakery foods and biscuits to be unhealthy, there have been numerous research studies in recent decades, aimed at improving the nutritional content of biscuits.22–24 The result of the study demonstrates the rise in technological and technique-oriented innovations in the context of extracting functional ingredients from existing foods. Age-old techniques like fermentation, germination, or sprouting are making waves in recent studies to extract functional ingredients from grains and pulses.\n\nAdditionally, the impact of technology like encapsulation, freeze-drying, and electrospinning needs more investigation to check their impact on developed functional foods could be highlighted through assessed articles. Food packaging material with the added advantage of functionality, lower production costs & increased shelf life is one of the upcoming fields of study that require scientists’ attention. Future studies are required to assess the potential of the peptides, nutrition, and germination due to their present status of the lesser network. Additionally, the study claims that more studies are required in the future to intensify the bioaccessibility & bioavailability of bioactive compounds from the consumption of developed functional products.\n\n\nConclusion\n\nThe study uses bibliometric analysis to study literature on functional ingredients and biscuits. The study assessed 395 documents and aspects such as variations in the number of publications over the research period, articles published, sources of publications, prolific authors, and keywords using R studio and VOS viewer. It is evident from analysis that the number of publications related to the topic is estimated to rise with time. Figure 5-7 mapping derivations based on title, abstract, keyword, and publications using VOS viewer. It study shows that functional ingredients are linked with various other fields. However, it can be noticed from search keywords, “functional ingredients”, “functional biscuits”, and “nutritional biscuits,” and through applying visualization mapping (VOS viewer) that there are keywords without network with other keywords. These keywords, such as peptides, germination, and nutrition, can be considered potential topic areas to research. Among functional foods, bread was one of the products investigated for its flour replacement without hampering texture which emphasizes textural evaluation of novel foods is on the rise. Nevertheless, the research limitation of the study is that data was generated from the Scopus database alone, and future studies could use various other sources such as the Web of Science or PUB Med.", "appendix": "Data availability statement\n\nNo data are associated with this article.\n\n\nBibliography\n\nBech-Larsen T, Scholderer J: Functional foods in Europe: consumer research, market experiences and regulatory aspects. Trends Food Sci. 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Publisher Full Text\n\nAl Ubeed HMS, Brennan CS, Schanknecht E, et al.: Potential applications of hemp (Cannabis sativa L.) extracts and their phytochemicals as functional ingredients in food and medicinal supplements: a narrative review. Int. J. Food Sci. Technol. 2022; 57(12): 7542–7555. Publisher Full Text\n\nSan José FJ, Collado-Fernández M, López R: Sensory evaluation of biscuits enriched with artichoke fiber-rich powders (Cynara scolymus L.). Food Sci. Nutr. 2018; 6(1): 160–167. PubMed Abstract \| Publisher Full Text\n\nNasir G, Chand K, Azaz Ahmad Azad ZR, et al.: Optimization of Finger Millet and Carrot Pomace based fiber enriched biscuits using response surface methodology. J. Food Sci. Technol. 2020; 57(12): 4613–4626. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMitrevski J, Pantelić N, Dodevska MS, et al.: Effect of Beetroot Powder Incorporation on Functional Properties and Shelf Life of Biscuits. Foods. 2023; 12(2). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYeasmen N, Bhuiyan MHR, Orsat V: Unravelling scientific research towards the green extraction of phenolic compounds from leaves: a bibliometric analysis. Int. J. Food Sci. Technol. 2021; 56(10): 4893–4906. Publisher Full Text\n\nChandra S, Singh S, Kumari D: Evaluation of functional properties of composite flours and sensorial attributes of composite flour biscuits. J. Food Sci. Technol. 2015. Publisher Full Text\n\nDávila-Hernández G, Delgadillo-Arévalo CS, Sánchez-Pardo ME, et al.: Pretreatment of tamarind pericarp to increase antioxidant availability and its application in a functional food. J. Food Sci. Technol. 2021; 58(6): 2385–2394. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOprea OB, Popa ME, Apostol L, et al.: Research on the Potential Use of Grape Seed Flour in the Bakery Industry. Foods. 2022; 11(11): 1–20. Publisher Full Text\n\nMarkets & Markets: Functional Food Ingredient Market.June 2021, cited 6/05/2023. Reference Source\n\nWorld Health Organization: WHO 75th Anniversary. Celebrating 75 years of improving public health.April 2023. Cited 6/5/2023. Reference Source" }	[ { "id": "305792", "date": "10 Aug 2024", "name": "Clara Talens", "expertise": [ "Reviewer Expertise food ingredients", "food technology", "food product development", "food science and nutrition", "functional foods", "systematic literature reviews", "data analysis", "impact of food ingredients on 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\nSummary of the manuscript: The manuscript presents a bibliometric analysis of research on functional ingredients used in developing functional biscuits. It aims to identify current trends, prolific authors, influential articles, and potential future advancements in this research area. The analysis is based on 395 articles retrieved from the Scopus database between 2013 and 2023, employing tools like R Studio and VOS Viewer for performance analysis and science mapping. Overall assessment: The manuscript is well-structured and presents an important analysis of the literature on functional ingredients and their application in biscuits. The methodology is clear, and the use of bibliometric tools is appropriate for the research objectives. However, there are some areas that need improvement to enhance the clarity and impact of the paper. Major comments:\nLiterature review:\nThe literature review should be expanded to provide a more comprehensive background on the importance of functional biscuits and their role in health. The current review is somewhat brief and does not adequately establish the context for the bibliometric analysis. There should be more discussion on the different types of functional ingredients commonly used in biscuit formulations and their specific health benefits.\n\nMethodology:\nThe selection criteria for articles should be explained in more detail. While the use of the SPAR-4-SLR method is mentioned, the manuscript does not clearly describe how the articles were filtered and selected beyond the initial database search. It would be beneficial to include a flowchart illustrating the selection process. The manuscript claims that the SPAR-4-SLR method is superior to PRISMA guidelines but does not provide evidence or references to support this claim. A comparison or justification for this choice would strengthen the methodology section.\n\nData analysis:\nWhile the results section is thorough, it would benefit from more interpretation of the data. For instance, the manuscript mentions trends in publication growth and the most cited articles but does not discuss why these trends are occurring or their implications for the field. The network analysis section should include a discussion on the significance of the clusters identified and how they relate to the development of functional biscuits.\n\nDiscussion and conclusion:\nThe discussion should be expanded to include the implications of the findings for future research and industry practice. Specifically, how can the identified trends and gaps be addressed in future studies? What are the practical applications of these findings in the food industry? The conclusion section is brief and could be more robust. It should summarize the key findings and suggest actionable recommendations for researchers and practitioners.\n\nClarity and readability:\nSome parts of the manuscript are densely written and could benefit from clearer language and shorter sentences. Simplifying the presentation of the data, particularly in the results section, would make the manuscript more accessible to a broader audience. The use of figures and tables is appropriate, but some could be better integrated into the text. For example, the discussion could reference specific tables or figures more explicitly to guide the reader.\n\nMinor comments:\nAbstract: The abstract provides a good summary of the paper, but it could be more specific about the key findings and their implications. References: Ensure all references are current and relevant. Some of the references cited are over a decade old, and it might be beneficial to include more recent studies to support the analysis. Formatting: Check for consistency in formatting, particularly in the references section and the use of headings and subheadings. Data presentation: Remove the percentage signs if individual counting is being referred to or ensure the percentages are correct. For instance, the statement \"Publications as review papers constitute 61% and 1% as conference papers\" needs clarification or correction. Language: Correct typographical errors in the conclusion, such as changing \"It study shows\" to \"This study shows.\"\nRecommendation: The manuscript has potential and provides valuable insights into the research trends on functional ingredients in biscuits. However, it requires significant revisions to improve the depth of analysis, clarity of presentation, and overall impact. I recommend a major revision before reconsideration for publication.\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? Partly\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\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": [ { "c_id": "12443", "date": "22 Oct 2024", "name": "Varalakshmi Chandra Sekaran", "role": "Author Response", "response": "The literature review is updated as per the comments. The methodology is improved and justification is added for the selected method. The result section is updated as per the comment. The network analysis section is improved to incorporate changes. The revision is made to the discussion section. All the comments are looked into and necessary changes are made in the manuscript." } ] }, { "id": "305797", "date": "16 Aug 2024", "name": "Teti Estiasih", "expertise": [ "Reviewer Expertise Food science and technology" ], "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 provides a bibliographic analysis of the functional ingredients for biscuits. However, the authors can not distinguish well the difference between functional ingredients and bioactive compounds. Functional ingredients are the ingredients used for manufactured foods that either have specific functions such as emulsifying or specific functions on health such as an ingredient high in carotene like carrot. Usually, the authors use the terms of bioactive compounds interchangeably with functional ingredients. For example: in Tables 3 and 6. The correct term is bioactive compounds instead of functional ingredients.\n\nThe title of this article is about functional biscuits, however, the analyzed published scholarly articles are general and do not focus on biscuits such as in Tables 2, 3, and 5.\nThe authors should mention the sources of the analyzed scholarly articles such as in Figure 3 and the Method section.\nI suggest the Authors revise the title to make it more relevant to the content of this article. I recommend the title be: A Bibliographic Review of bioactive compounds and their efficacy in developing functional foods.\nLastly, the numbering for cited references should be listed based on the sequence of their citation in the text. For example, the No, 25 should be the No. 1 because this is the first citation.\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? Partly\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.) Partly", "responses": [] } ]	1	https://f1000research.com/articles/13-421
https://f1000research.com/articles/12-230/v1	01 Mar 23	{ "type": "Research Article", "title": "In silico molecular docking and molecular dynamic simulation of agarwood compounds with molecular targets of Alzheimer’s disease ", "authors": [ "Phaniendra Alugoju", "Vishwambar Vishnu Bhandare", "Vishal S. Patil", "Krishna Swamy V. K. D", "Prem Kumar Borugadda", "Tewin Tencomnao", "Phaniendra Alugoju", "Vishwambar Vishnu Bhandare", "Vishal S. Patil", "Krishna Swamy V. K. D", "Prem Kumar Borugadda" ], "abstract": "Background\nAlzheimer's disease (AD) is a neurological condition that primarily affects older people. Currently available AD drugs are associated with side effects and there is a need to develop natural drugs from plants. Aquilaria is as an endangered medicinal plant genus (commonly called agarwood plants) and various products of Aquilaria plant spp. including resinous heartwood, leaves, bark, and stem have been widely used in various traditional medicine systems. Research on agarwood plants is sparse and only a few previous studies demonstrated their neuroprotective properties in vitro. Owing to the presence of a plethora of secondary metabolites in agarwood plants, it is imperative not only to protect these plants but also evaluate the bioactivity of agarwood phytochemicals. Methods\nComputational methods such as AutoDock Vina and molecular dynamic (MD) simulations were employed for the docking of 41 selected agarwood compounds with AD-related molecular targets. Results and Conclusion\nAccording to docking data, three compounds aquilarisin, aquilarisinin, aquilarixanthone showed highest binding affinity to selected AD targets compared to their known inhibitors. MD simulation studies revealed that, selected agarwood compounds' protein-ligand complexes showed remarkable structural stability throughout 100 ns simulation. The agarwood chemicals aquilarisin, aquilarisinin, aquilarixanthone, pillion, and agarotetrol are consequently suggested as some of the found hits against AD targets, however, additional experimental validation is required to establish their effectiveness.", "keywords": [ "agarwood", "neurodegeneration", "docking", "molecular dynamic simulation." ], "content": "Introduction\n\nAlzheimer's disease (AD) makes up 75% of instances of dementia and is the most common neurodegenerative ailment affecting those over 65 years.1–3 Extracellular amyloid plaques, also known as “Aβ plaques,“ or “senile plaques,“ rich in amyloid-(Aβ), and intracellular neurofibrillary tangles (NFTs), rich in tau proteins, are the two key pathological hallmarks of AD.4 Several proteins associated with the neurological dysfunction in AD include cholinesterases,5 N-methyl D-aspartate (NMDA) receptor,6 beta-site amyloid precursor protein cleaving enzyme 1 (BACE 1),7 Asparagine endopeptidase (AEP),8 Monoamine oxidases (MAO)9 and protein kinases.10,11 AD has become a global health problem due to the lack of effective treatment for the amelioration of neurological dysfunction.12 The current pharmacologically important AD drugs include cholinesterase inhibitors such as donepezil, galantamine, and rivastigmine block the breakdown of acetylcholine, thereby increasing acetylcholine (Ach) levels in brain and help in improving cognitive function.3 Galantamine is the only naturally occurring inhibitor belonging to alkaloid class of phytochemicals and it can reversibly and competitively inhibit acetylcholinesterase (AChE). Memantine is the currently available NMDA receptor antagonist that can improve cognition and memory problems in AD by balancing the glutamatergic system.6 However, some side effects of these medications include nausea, headache, vomiting, and dizziness. Therefore, there has been a great interest in identifying potent natural inhibitors of target proteins of AD.7\n\nSince ancient times, traditional medicinal plants have been utilized as a major source of drugs to treat various kinds of human illnesses including neurological disorders.13 Natural phytochemicals have received much attention in recent years due to their pharmacophore-like structures and pharmacokinetic properties. Owing to the presence of a plethora of phytochemicals in medicinal plants, the systematic analysis of each phytochemical by conventional methods is cumbersome and a time taking process.14 In this context, the computer aided drug design techniques have been widely used for the screening of chemical libraries and identification of molecular targets of natural or synthetic compounds.15 Virtual screening is considered as the standard initial step in the drug discovery process prior to wet lab experiments.15 In silico approaches significantly increased the effectiveness of assessing the bioactive compounds of medicinal plants. In fact, using in silico approaches, some Food and Drug Administration (FDA) approved drugs were developed. Several studies have performed in silico analysis on phytochemicals of medicinal plants against target proteins of human diseases.16–20 Previous studies have used computational tools to identify and predict possible anti-Alzheimer’s potential of bioactive compounds in medicinal plants.17,21–23\n\nAquilaria is an endangered medicinal plant genus that is currently protected by international laws due to indiscriminate cutting for various commercial, cultural, religious, and medicinal purposes. Aquilaria spp. trees, commonly known as agarwood, are primarily found in Southeast Asia. The products of Aquilaria spp, including agarwood, leaves, bark, stem etc., have been extensively used in Asia for the treatment of a variety of ailments such as cough, pain, and allergy. Agarwood is a valuable, non-timber, resinous portion that is used for making incense, perfume, cosmetics, and personal care products, as well as for the production of traditional Ayurvedic, Chinese, Thai, Korean, Tibetan, and Eastern medicines for curing many ailments such as arthritis, inflammation, diarrhoea, and used as a soporific, antidepressant, and cardio-protectant.24 Other plant materials of Aquilaria spp. including leaves, stem, and bark have been found to have several pharmacological properties such as anallergic, cytotoxic, anti-inflammatory, cardioprotective, antimicrobial, anti-oxidant, hepatoprotective, laxative, and mosquitocidal effect.25,26 Compounds extracted from the resinous heartwood of Aquilaria sinensis showed notable neuroprotective effects on corticosterone and 1-methyl-4-phenylpyridinium (MPP+)-induced injury in PC12 cells27,28 and also exhibited obvious cytotoxic activity.29 The benzene extractable compounds of agarwood (jinkoh-eremol and agarospirol) Aquilaria malaccensis possess potent anti-depressant and anti-psychotic activities.30,31 The chloroform extracts of the leaves and stem of Aquilaria subintegra showed significant AChE inhibitory activity.32 Compounds isolated from Aquilaria crassna leaves were also shown to exhibit neuritogenic properties and therefore exerted neuroprotective effects in P19-derived neurons.33 A. crassna leaf extracts have been demonstrated to ameliorate glucose-Induced neurotoxicity in vitro.34 Our unpublished experimental data also indicate that both leaf and agarwood extracts of A. crassna can exert protective effects against D-galactose induced neurotoxicity in mouse hippocampal HT-22 cell line.\n\nTraditional understanding of herbal supplements is helpful for developing cutting-edge drugs for a number of illnesses, including neurological disorders like Alzheimer's disease (AD).13 There is still a lack of research on agarwood plants due to extreme demand and depletion of natural resources.35 However, neuroprotective activity of these agarwood plants is poorly explored. Growing evidence suggests the use of in silico studies as the first step before setting up in vitro or in vivo experiments. Thus, virtual screening of small molecules library with known AD targets is critical for identification and subsequent validation of best possible hits in either cell lines or animal models. Hence in the present study, molecular docking of selected agarwood compounds from PubChem with molecular target proteins of AD was performed for the first time. Structural stability of the best docked agarwood compounds with AD targets has been studied using molecular dynamic (MD) simulations.\n\n\nMethods\n\nPhytocompounds from the Aquilaria plant species were selected based on the previous literature and their structures were retrieved from the PubChem database (refer Figure 1). The phytocompounds structures were prepared by adding polar hydrogens, Gasteiger charges and by performing energy minimization in UCSF Chimera 1.16 using default parameters.36\n\nPrediction of pharmacokinetic and pharmacodynamic features can be accomplished by using the physicochemical characteristics of chemical substances, such as lipophilicity (LogP), solubility (LogS), and polar surface area and volume (PSA).37 It is crucial to analyze these features because they affect how they interact with transport proteins and enzymes that are involved in drug clearance.37 In the present study, we have used Molsoft tools (https://www.molsoft.com/mprop/) to predict the number of H-bond donors (No. of HBD) and acceptors (No. of HBA) present, polar surface area (MolPSA), lipophilicity (Mol LogP), solubility (Mol LogP), and the molecular polar surface area and volume (Mol PSA) of tested ligands.38 Also, ADMET properties such as absorption, digestion, metabolism, excretion, and toxicity properties of selected ligands were predicted using ADMETlab 2.0 (https://admetmesh.scbdd.com/).\n\nThe crystal structures of different proteins implicated in the pathogenesis of AD were retrieved from RCSB PDB structural database (https://www.rcsb.org/). The stereo-chemical properties, Ramachandran graph and values of selected proteins were evaluated by Molprobity server.39 Chimera 1.16 (RRID:SCR_004097) was used to generate any missing residues in the selected target proteins. Following the removal of unneeded nonstandard heteroatoms, polar hydrogens and Gasteiger charges were added. All targets' structural details were refined using the steepest descent and conjugate gradient algorithms (100 steps each) with amber force field (Amber ff14SB).40 Then, using AutoDock tools 1.5.7 (RRID:SCR_012746), the energy-minimized protein structures were transformed into ‘pdbqt’ format. A list of proteins along with their PDB IDs are given in Table 1.\n\nDocking was performed with Autodock Vina as described in our previous study.41 The grid box's dimensions were fixed at XYZ=30Å×30Å×30Å which was found to be the best size for the default exhaustiveness (=8), and the ligand binding site was positioned in the middle of the grid box. The spatial dimension (XYZ axis) and the grid box’s size were specified in a configuration file. Using AutoDock vina version 1.1.2's (RRID:SCR_011958) command line interface, docking was accomplished. The obtained results are restricted to nine binding modes. The log file created included a list of the increasing binding modes and their associated binding energies. The BIOVIA Discovery studio visualizer 2021 was used to view the binding modes. All non-bonded interactions were recorded (DOI: dx.doi.org/10.17504/protocols.io.3byl4j362lo5/v1).\n\nMD simulation using Gromacs 2020.5 (RRID:SCR_014565) was used to track the structural stability of the docked complexes. Gromos96 force field was used to create the topology of the protein, and PRODRG server (http://davapc1.bioch.dundee.ac.uk/cgi-bin/prodrg/submit.html) was utilized to create the topology of the ligand. The docked complexes were solvated in a cuboidal box with adequate size to fit the complete complex in the middle using a “Simple Point Charge“ (SPC) water model. Appropriate number of counter ions (Na+/Cl-) were used to neutralize the simulated systems. The undesirable contacts and steric conflicts were then removed from the neutralized systems using steepest descent followed by conjugate gradient methods for 50,000 steps each.\n\nThe NVT ensemble used to maintain constant number of atoms, volume, and temperature, further NPT ensemble was used to maintain constant pressure. In this study, we set temperature and pressure constant at 300K and 1 bar respectively. Further, followed by 1ns of equilibration, unrestrained MD simulation was performed for a period of 100ns in solvent. The Particle Mesh Ewald (PME) method was used to handle coulomb electrostatic interactions, while the LINear Constraint Solver (LINCS) algorithm was used to limit H-bonds. Using a cut-off value of 14 Å, the non-bonded contacts were trimmed. The trajectories generated were analyzed using some of the inbuild gromacs tools like ‘gmx rms’, ‘gmx rmsf’, ‘gmx hbond’, ‘gmx gyrate’, ‘gmx sasa’, etc. and other additional packages for specific analysis wherever required. Conformational changes at the secondary structural level were monitored by using Dictionary of Protein Secondary Structure (DSSP) software (RRID:SCR_002725) (DOI: dx.doi.org/10.17504/protocols.io.36wgqjmp5vk5/v1)\n\nMM-PBSA in conjunction with MD simulations is commonly used to determine the binding free energy of protein and ligand complexes.42 It uses the following equation as:\n\nIn STRING database (version 11.0), the proteins Glycogen Synthase Kinase 3 Beta (GSK3B), Monoamine oxidase-A (MAOA), Butyrylcholinesterase (BChE), Acetylcholinesterase (AChE), beta-site amyloid precursor protein cleaving enzyme 1 (BACE1), Legumain (LGMN), and Glutamate Ionotropic Receptor NMDA type subunit 1 (GRIN1) were searched for Homo sapiens. Only these proteins were selected for enrichment analysis, because most of the tested ligands showed good binding affinity only towards them. Gene ontology (GO) analysis was used to pinpoint the biological processes. Based on the available literature, the pathways contributing to AD pathogenesis were selected.\n\nCytoscape tool (version 3.9.1) was used to build the integrated network between chemicals, protein targets, and regulated processes. In the network analyser, the entire network was treated as “Direct” during network construction. The resulting network was inspected using a topological parameter “edge count.44” The node size and colour were set using low values to small sizes and low values to bright colours, respectively.45\n\n\nResults\n\nThe physicochemical properties of all the chosen agarwood compounds were studied to gain more structural features of the individual phytocompounds (Table 2). Also, pharmacodynamic and pharmacokinetic properties of these compounds were analyzed using ADMETLab2.0. The analysis reveals that all the chosen compounds were less/not toxic. Additionally, it is interesting to note that most of the compounds had the capacity to cross the blood brain barrier (BBB). Though three phytocompounds namely, aquilarisin, aquilarisinin, and aquilarixanthone failed the Lipinski rule, they were shown to cross the blood brain barrier effectively and express lesser or no toxicity (Supplementary Material ADMET properties of agarwood compounds.xlsx). The blood brain barrier penetration values for agarotetrol, aquilarisin, aquilarisinin, aquilarixanthone, and pillion were predicted to be 0.8, 0.029, 0.35, 0.011, and 0.01, respectively. All the chosen ligands were predicted as non-carcinogenic and showed drug-likeliness.\n\nIn this study, docking of 41 agarwood compounds was done against 13 target proteins of AD. The selection of specific AD targets has been made considering their close and direct association with AD pathogenesis and its disease progression. Figure 2 represents heatmap of the binding energy estimated from the docking pose of agarwood compounds towards the tested molecular targets of AD. The scale of heat map ranges from least (blue) to highest binding (red) affinity was predicted based on the docking results. The binding interactions of only those ligands with least binding energy than the respective native ligands of AD target proteins are highlighted by circle in the heatmap (refer Figure 2). Ligands such as aquilarisin, aquilarisinin, and aquilarixanthone showed good binding affinity with most of the selected AD related proteins. Aquilarisin showied good binding affinity towards 12 of the 13 tested AD related proteins except with 4AU8. Aquilarisinin exhibited the highest binding affinity towards 8 of the 13 selected AD proteins including 1PBQ, 1Q5K, 1UDT, 4BDS, 4KKE, 4M0F, 5BTR, and 5LUA. On the other hand, aquilarixanthone was found to show highest binding affinity towards 9 of the 13 selected AD proteins including 1Q5K, 1UDT, 4AU8, 4BDS, 4KKE, 4M0F, 5BTR, 5LUA, and 6GZM.\n\nLigands with higher binding energies than the respective native ligands of target proteins are denoted in red circles.\n\nFrom the heatmap it was clear that proteins such as 1Q5K (GSK3beta), 2Z5X (MAO-B), 4BDS (BChE), 4M0F (AChE), 5BTR, 5LUA (AEP) formed best docked complexes with most of the selected agarwood compounds. The site-specific non-bonded interactions of some of the representative top docked complexes showing highest binding affinity and conserved binding pocket interactions are listed in Table 3 (and also the 3D and 2D structures of top docked complexes are shown in Figure 3). Further, to gain more detailed insights to their structural stability and intermolecular interactions we selected some of the best docked representative complexes for MD simulations. The phytocompounds bound to these selected targets had been chosen considering binding energy, H-bonds and other nonbonded interactions including hydrophobic and electrostatic interactions. Additionally conserved binding pocket interactions had also been considered and compared with the respective known control inhibitors for the respective targets. Thus, we selected total seven complexes namely 1Q5K-AXN (complex1), 2Z5X-PLN (complex2), 4BDS-ANN (Complex 3), 4M0F-AXN (Complex 4), 5IE1-ASN (Complex 5), 5LUA-AGT (Complex 6), and 1PBQ-ASN (Complex 7) for MD simulations. It was observed that the agarwood phytocompounds bind to the AD targets to form stable complexes, and these complexes are maintained through H-bonds and other non-bonded interactions (as shown in Table 3).\n\nWe selected 7 complexes of agarwood compounds 1Q5K-AXN (complex1), 2Z5X-PLN (complex2), 4BDS-ANN (Complex 3), 4M0F-AXN (Complex 4), 5IE1-ASN (Complex 5), 5LUA-AGT (Complex 6), and 1PBQ-ASN (Complex 7) having least binding energy towards selected AD targets. MD simulation quality check was performed over all the 7 trajectories by plotting temperature, pressure, and potential energy. During the 100ns simulation, the temperature and pressure were held constant at 300 K and 1 bar, respectively. We observed less fluctuations in the potential energy of the simulated systems, suggesting the well equilibration of all the complexes during simulation. In order to better understand the structural stability, the root mean square deviation (RMSD), root mean square fluctuation (RMSF), radius of gyration (Rg), and solvent accessible surface area (SASA) were also measured.\n\nRoot mean square deviation (RMSD) analysis\n\nThe backbone RMSD values were plotted over the trajectories revealing the stable dynamics expressed by all the simulated complexes during the 100ns. Figure 4 represents the backbone RMSD values of GSk3beta (indigo), BChE (Green), AChE (crimson), BACE1 (turquoise), and AEP (olive) showing moderate fluctuations up to 30ns (equilibration period), however, RMSD values are well stabilized after 50ns for these five proteins followed by equilibration period of 30ns. However, the RMSD of backbone atoms of NMDA (orange) and MAO-B (maroon) reached equilibrium after 60ns and 80ns during simulation. The average backbone RMSD values of GSk3beta, BChE, AChE, BACE1, AEP are observed as 2.5Å, 3.2Å, 2.1Å, 2.5Å, 2.4Å, etc. On the other hand, average RMSD values of NMDA, and MAO-B are found to be ~3.2Å and ~3.6Å, respectively. The representative MD simulation end structure extracted from the trajectory is compared with the initial starting MD simulation of the respective complexes. The analysis reveals the RMSD of structural superimposition of MD starting structure (0 ns) and end structure (100 ns) for studied 7 complexes has been found to be 1.339Å, 1.186Å, 1.122Å, 1.314Å 1.230Å, 1.161Å, 1.146Å. The ligand RMSD values also follows the similar trend (i.e., RMSD values <5Å) for all the ligands, however, the ligand7 (ASN) represents structural changes due to existence of torsions, which trigger the diverse ASN conformations during simulation. Thus, in general, all the 7 complexes express stable dynamics during 100ns MD simulation with backbone and ligand RMSD value of <5Å (refer Figure 4A and B).\n\nRoot mean square fluctuation (RMSF) Analysis\n\nRMSF gives qualitative measure provides detailed insights to the conformational flexibility of protein structure. The RMSF value plotted for the C-alpha atoms of all the simulated systems is shown in Figure 5. The C-terminal region in all the complexes show highest fluctuations when compared to other regions in the protein structure. Careful observation of the three-dimensional structure reveals that increased RMS fluctuation values at C-terminal region are mainly due to the absence of native folded secondary structure. Overall binding pocket residues show the least RMS fluctuations (<3Å) as they actively participate in the stable non-bonded contacts. Also, other flexible loops and N-terminal regions express moderate to high fluctuations in the RMSF values. Due to a maximum residual fluctuation of up to 5, a greater peak between 175 and 190 amino acid residues was detected in complex 1. Only the C- and N-terminal portions of complex 2 exhibit persistent amino acid variation. Two higher peaks are observed in complex 3; one between 370 and 390 and one between 75 and 90 amino acid residues. Three higher peaks in complex 4 have been noticed at positions between 370 and 390 amino acid residues, between 250 and 275 amino acid residues, and between 40 and 60 amino acid residues (3.8Å, 3.9Å, and 3.8Å, respectively). Between 150 and 200 amino acid residues and 300 and 350 amino acid residues, complex 5 shows two higher peaks (each 5.5Å). A higher peak (4.5Å) has been seen in complex 6 between 90 and 110 amino acid residues. Three higher peaks (at 5Å, 6.5Å, and 4.5Å) have been discovered in complex 7, two of which were between 0 and 60 amino acid residues and one between 80 and 110.\n\nAnalysis of Radius of Gyration (Rg) and solvent accessible surface area (SASA)\n\nThe radiation of gyration (Rg) explains the protein folding/compactness of the molecule, hence we analyzed the compactness of the protein-ligand complexes and exposure of hydrophobic core of the protein to the solvent upon ligand binding. The variation in Rg and SASA of selected complexes is given in Figure 6. The Rg value for complexes 1-7 (except complex 2) is well stabilized while complex 2 shows steady decrease in the Rg value during the MD simulation. Also, SASA values of all complexes show significant structural stability and represent the formation of compact globular shape during 100 ns simulation. The average Rg and SASA values of complexes 1 to 7 range between ~1.75 to 2.5Å and 155 to 320 (nm2). Thus, we observed stable complex formation in all the complexes (refer Figure 6).\n\nIntermolecular interactions observed in docked complexes\n\nFormation of H-bonds between selected protein-ligand complexes has been monitored during 100 ns simulation. Figure 7 denotes the number of H-bonding interactions formed during simulation. Table 4 lists comparative analysis of non-bonded interactions observed in the initial starting structure (0ns) and MD simulation end structure (100ns). In general, the native non-bonded contacts were well conserved even after 100 ns MD simulation, revealing relatively more stable complex formation in all the simulated complexes. Hydrophobic interactions equally contribute in stabilizing these complexes. Moreover, the consistency of the observed H-bonds also supports the stable complex formation during simulation (Refer Figure 7). The snapshot of initial and final MD structure is shown in Figure 8, revealing the binding of ligands to the conserved binding site of their respective targets. Also, most of the complexes show compact folding during simulation forming much compact globular structure.\n\nEstimation of Binding Free Energy using MMPBSA\n\nThe binding free energy for all the simulated complexes has been quantitatively measured by using MMPBSA methods over the well equilibrated trajectory observed between 0 to 100 ns. Table 5 represents the energy components including molecular mechanics, van der Waal (vdW) interactions, electrostatic, polar and nonpolar energies that significantly contribute to the binding free energy. The estimated binding free energy for complexes 1 to 7 is -81.018 ± 61.364, -159.438 ± 10.190, -227.959 ± 13.745, -152.764 ± 15.897, -149.090 ± 16.646, -79.236 ± 19.623, and -146.796 ± 12.694 kJ/mol, respectively. It has been observed that binding free energy of protein-ligand complexes is significantly influenced by both electrostatic and vdW interactions.\n\nConformational changes at secondary structural level\n\nWe further examined the secondary structural changes during simulation period using DSSP. The complexes 1-7 using DSSP plot have been shown as panel A-G in Figure 9. Various components of secondary structures are shown in specific colors as shown in figure legend. We noticed that major secondary structural components such as alpha helices and beta sheets are relatively much stable and express less variations in the secondary structure. Also, interestingly, the architecture of binding pocket is well maintained throughout the simulation due to rigidity provided by the alpha helices and beta sheets in all the simulated complexes. Thus, overall binding of ligand does not affect the secondary structure of protein. However, minor variations such as shortening, or extensions were noticed at flexible loop and linkers connecting the alpha helices and sheets. The H-bonds provide structural stability to the alpha helices and beta sheets in all the complexes.\n\nA) complex 1, b) complex 2, c) complex 3, d) complex 4, e) complex 5, f) complex 6, g) complex 7.\n\nThe GSK3B, MAOA, BCHE, ACHE, BACE1, LGMN, and GRIN1 all influenced 18 different biological processes. Among them, 15 were associated with AD. Among 7 targets, 6 targets viz., BCHE, ACHE, BACE1, GSK3B, GRIN1, and LGMN were enriched for modulation of chemical synaptic transmission (GO:0050804) and 3 targets viz., BACE1, GSK3B, LGMN were enriched for cellular response to amyloid-beta (GO:1904646). Following these, regulation of neurotransmitter levels, neuron death, cell communication, signaling, synaptic plasticity, neuron apoptotic process, learning, etc were also associated with SK3B, MAOA, BCHE, ACHE, BACE1, LGMN, and GRIN1 targets. Table 6 represents the biological processes modulated by best docked phytocompounds. Among the 7 targets, GRIN1, GSK3B, BACE1, and LGMN scored the highest edge count within the network of and were involved in multiple biological processes for the regulation of AD (Figure 10).\n\n\nDiscussion\n\nAquilaria is an endangered agarwood-producing genus includes many plants species. Various parts of agarwood plants have been widely used as an important ingredient of traditional Ayurvedic, Chinese, Thai, Tibetan, and Eastern medicine.35 Limited number of in vitro experiments have shown that the leaves, stem, and agarwood of Aquilaria plants, among other plant parts, exhibit neuroprotective properties.27,28,30,32,33,46 Scientific investigation into the phytochemical components of agarwood is still quite limited because of the high industrial demand and loss of its natural resources. Therefore, it is crucial to carry out additional research investigations to develop premium goods and medications using agarwood's beneficial phytochemical components. In this regard, we carried out in silico studies to search for the agarwood hit molecules against the molecular targets of AD. The first step was to conduct docking studies to find the agarwood compounds with highest binding affinity against AD molecular targets. A total of five hit compounds (aquilarisin, aquilarisinin, aquilarixanthone, agarotetrol, and pillion) were identified from our docking results that demonstrated substantial binding affinity for several AD targets. Additionally, we chose these top seven docked complexes based on the binding energy values, the number and distance of hydrogen bonds, the number and distance of hydrophobic contacts, and conserved amino acid residues with native ligand interactions. We also chose two controls for comparative study. Total 7 systems were chosen and subjected for MD simulation study namely, complex 1: GSK3beta-AXN; complex 2: MAO-B-pilloin; complex 3: BChE-ANN; complex 4: AChE-AXN; complex 5: BACE1-ASN; complex 6: AEP-AGT, complex 7: NMDA-ASN).\n\nA few of the theories proposed to explain the underlying molecular reasons of AD are the cholinergic theory, excitotoxicity, amyloid concept, and tau concept. According to the cholinergic theory, cognitive impairment in AD is caused by the loss of acetylcholine-synthesizing cholinergic neurons and consequent drop in ACh levels.1 ACh is hydrolyzed to acetyl coenzyme A (acetyl CoA) and choline by two cholinesterases called acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). In neuromuscular junctions and cholinergic synapses, ACh is mostly degraded by AChE rather than BChE under normal physiological circumstances. Cholinergic transmission can be stopped by AChE, a highly selective cholinesterase that can hydrolyze up to 25000 ACh molecules per second into acetate and choline. AD patients show up to a 67% decrease in the levels of AChE, while BChE levels rise to 120% of normal level.5 This indicates that BChE could compensate for deficit in AChE by hydrolyzing ACh. The breakdown of acetylcholine at the synaptic cleft is prevented by cholinesterase inhibitors, which thereby improves cholinergic transmission. The current pharmacologically important cholinesterase inhibitors donepezil, galantamine, and rivastigmine could increase ACh levels in the brain and help in improving cognitive function.3 However, Galantamine is the only naturally occurring inhibitor belonging to alkaloid class of phytochemicals and it can reversibly and competitively inhibit AChE. Therefore, it is very essential to identify potent cholinesterase inhibitors for the treatment of AD. Earlier, molecular docking studies showed that rutin (a flavone) showed improved AChE and BChE binding affinities compared to galantamine.47 In this study we have shown that the best docked agarwood phytocompounds such as aquilarixanthone and aquilarisin express good binding affinity when compared to their known inhibitors of AChE and BChE, respectively. Most interestingly, we discovered aquilarixanthone had a higher binding affinity to AChE (-9.9 kcal/mol) than galantamine (-9.3 kcal/mol). Another factor contributing to neuronal death in AD is glutamate-induced excitotoxicity, which occurs when glutamate levels are too high and cause overstimulation of glutamate receptors such as the NMDA receptor. Memantine is the currently available NMDA receptor antagonist to normalize the glutamatergic system and ameliorate cognitive and memory deficits in AD.6 The cleavage of amyloid beta (Aβ) from the amyloid precursor protein (APP) by the beta site amyloid precursor protein cleaving enzyme (BACE 1), also known as beta secretase, plays a role in the pathogenesis of AD.7 As the inhibitors NMDA receptor and BACE1 are associated with reduction in glutamate and amyloid beta toxicity, respectively. Here we have observed stable interaction of aquilarisin with both NMDA receptor and BACE1. Asparagine endopeptidase (AEP), often referred to as human legumain, is known to have a role in the advancement of neurological illnesses such amyotrophic lateral sclerosis (ALS),48 stroke,49 and AD. It is also involved in a number of physiological functions, including immunological function. AEP is involved in the cleavage of amyloid precursor protein50 and tau protein,51 subsequently contributing to both amyloid and tauopathy in AD. This specifies the possible delta secretase activity of AEP. Previous studies have reported significantly high levels of AEP in the brains of AD patients as well as aged mice, suggesting the role of AEP in the onset and progression of AD. In view of this, targeting AEP may be useful for the amelioration of neurodegenerative disorders like AD.52 Agarotetrol showed good binding affinity with the AEP in our study. Both monoamine oxidases A and B have been involved in the altered aminergic neurotransmitter levels seen in AD.53 Activated MAO-A/B can destroy cholinergic neurons, induce amyloid β peptide production and accumulation, formation of neurofibrillary tangles and subsequent cognitive dysfunction.54 Selegiline, an MAO inhibitor used to treat Parkinson's disease, has been tested for the treatment of memory impairment in AD. By preventing reactive astrocytes from producing gamma-aminobutyric acid (GABA), selegiline has been shown to enhance synaptic plasticity, learning, and memory in AD mice.55 It has also been suggested that MAO-A inhibitor also offers neuroprotection.9 Our docking studies also showed that pillion (-9.9 kcal/mol) has more affinity towards MAO-A compared to the native ligand as well as the control drug selegiline (-7.4 kcal/mol). The best docked agarwood compounds, such as agarotetrol, aquilarisin, and pillion are chromones, aquilarisinin is a chalcone, and aquilarixanthone is a xanthone, are complexed with the best docked complexes that we have chosen for MD simulation. Oxygen-containing heterocyclic compounds like chromones and xanthones are known for their antioxidant capabilities. Our in silico study suggests that compounds like agarotetrol, aquilarisin, aquilrixanthone, aquilarisinin, and pillion may be good lead candidates; however, further experiments studies would be required given that hydroxylated chromones and xanthones demonstrated reactive oxygen species (ROS) and reactive nitrogen species (RNS) scavenger effects.56\n\nIn AD, twisting and tangles occur in the tau protein. As the tangles clump together, some nerve cells perish. This makes cell communication much more difficult. As connections between neural networks weaken, brain regions start to shrink.57 Also, AD has a key pathogenic hallmark known as brain atrophy brought on by neuronal loss. Amyloid beta, which makes up the majority of senile plaques, is assumed to play a key role in the death of neurons and may play a role in synapse and neural network dysfunction as well as cognitive impairment in AD.58,59 In the present study, phytocompounds were predicted to target GSK3B, MAOA, BCHE, ACHE, BACE1, LGMN, and GRIN1 and found to regulate neurotransmitter levels, cell communication, signalling, cellular response to amyloid-beta, learning, amyloid precursor protein metabolic process, nervous system process, regulation of synaptic plasticity and neuron apoptotic process. Also, these compounds modulate negative regulation of neuron death, response to oxygen-containing compounds, and neuron projection organization processes.\n\nAgarwood plants are traditional medicinal plants which have been recently categorized as endangered and threatened plants. Considering the significant potential of agarwood in various health promoting effects and limited knowledge highlighting neuroprotective properties, we aimed to find best possible lead molecules for AD. We extensively used molecular modelling approach to screen the library of selected agarwood phytocompounds against key AD targets. The phytocompounds aquilarisin, aquilarisinin, and aquilarixanthone have great potential to inhibit multiple AD targets with the highest binding affinity. It is interesting to note that, these compounds express stable binding and conserved active site interactions when compared to their respective known inhibitors. Furthermore, a 100ns all-atom MD simulation in an explicit solvent was used to look at the structural stability and intermolecular interactions for some of the top found hits. During MD simulation, all the complexes reached equilibrium earlier than 30 ns and thereafter expressed stable dynamics throughout the simulation. The estimated binding free energy using MMPBSA approach for all the complexes shows that these phytocompounds show much better efficacy in successful inhibition of AD targets by fitting well into the cavity. They also express least fluctuations and form compact globular shape due to increased intermolecular non-bonded interactions during MD simulation. Notably, aquilarisin, aquilarisinin, and aquilarixanthone fail to pass the Lipinski rule, but still their bioactivity observed in our computational study is remarkable. However, further experimental studies (either in cell line or animal models) are essential to validate neuroprotective potential of these agarwood phytocompounds.\n\n\nEthical Considerations\n\nThis study does not require any ethical approval as it does not involve any endangered species (plant or animal).", "appendix": "Data availability\n\nZenodo: In silico molecular docking and molecular dynamic simulation of agarwood compounds with molecular targets of Alzheimer's disease [Data set]. Zenodo. https://doi.org/10.5281/zenodo.7567232. 60\n\nThe Underlying Data for this Project are as Follows:\n\n• Agarwood Compounds_PDB formats.zip\n\n• Graphical Abstract.jpg\n\n• Proteins_structure_3D.zip\n\nZenodo: In silico molecular docking and molecular dynamic simulation of agarwood compounds with molecular targets of Alzheimer's disease [Data set]. Zenodo. https://doi.org/10.5281/zenodo.7567232. 60\n\nThe extended data for this Project are as Follows:\n\n• Supplementary Material ADMET properties of agarwood compounds.xlsx\n\n• Tables.docx\n\nData are accessible in accordance with the provisions of the Creative Commons Attribution 4.0 International license (CCBY 4.0).\n\n\nAcknowledgements\n\nPhaniendra Alugoju extends thanks to C2F (Secondary Century Fund) Postdoctoral Fellowship, Chulalongkorn University, Bangkok 10330, Thailand. We confirm that the Chulalongkorn University C2F (Secondary Century Fund) has granted us permission to credit them in this article.\n\n\nReferences\n\nHolzgrabe U, Kapková P, Alptüzün V, et al.: Targeting acetylcholinesterase to treat neurodegeneration. Expert Opin. Ther. Targets. 2007 Feb; 11(2): 161–179. Publisher Full Text\n\nChopra K, Misra S, Kuhad A: Neurobiological aspects of Alzheimer's disease. Expert Opin. Ther. Targets. 2011 May; 15(5): 535–555. 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Publisher Full Text" }	[ { "id": "221667", "date": "18 Dec 2023", "name": "Sailu Sarvagalla", "expertise": [ "Reviewer Expertise Structural bioinformatics", "drug design", "proteomics and genomics" ], "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\nSummary The main aim of this paper is to identify natural compounds as hits for AD disease. In this regard, authors explored the utility of Aquilaria plant spp. derived natural compounds as a potential hit for AD related drug targets. For this purpose, authors used routine computer aided drug design techniques to evaluate agarwood compound’s binding mode and interaction strength with different AD targets. The data depicted in this paper is valuable, and clearly demonstrated some of hits have comparable binding strength and interaction as of native ligands of AD targets. In my opinion, rationalization is good, but needs some sort of in vitro validation for their potential application as a hit to lead development.\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": "12862", "date": "16 Dec 2024", "name": "Tewin Tencomnao", "role": "Author Response", "response": "Thank you for reviewing our manuscript. We genuinely appreciate your positive feedback and are delighted that you believe our work will resonate with scientists in the field. Experimental validation for these preliminary and interesting results, based on in silico studies, has already been planned and will soon be communicated as a separate manuscript. We acknowledge the need for further validation, and we plan to conduct in vitro studies to confirm the potential of these hits for lead development. Future work will focus on refining these candidates and exploring their biological activity to enhance our findings and support their applicability in drug discovery." } ] }, { "id": "229488", "date": "23 Jan 2024", "name": "Yassir Boulaamane", "expertise": [ "Reviewer Expertise Computer-aided drug discovery", "natural products", "QSAR modelling", "neurodegenerative diseases", "molecular dynamics simulations" ], "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\nMethods section in abstract should be more detailed. Introduction lacks experimental data (IC50, Ki…) on the described activities of Agarwood compounds. What is the rationale behind using the 13 selected protein targets? Recommended range for each chemical descriptor needs to be added in table 2. Data describing ADMET of tested ligands should be added in the manuscript instead of supplementary material. BBB permeability is a crucial parameter for Alzheimer’s drugs; however, this property was not taken in consideration. Abbreviation of ligands should be used in the beginning, so the reader doesn’t get confused halfway through. It’s not clear why complexes are exhibiting out of bound fluctuations outside the C- and N-terminal region in figure 5. RMSF section should be re-analysed taking into consideration the dynamic behavior of Apo forms of each protein. Authors failed to highlight the key interactions for each protein and how they are at the origin of selectivity for the native ligands and if they are implicated in the binding of Agarwood compounds. No experimental studies in the literature were linked to back up the final conclusive remarks.\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? Partly\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "12863", "date": "16 Dec 2024", "name": "Tewin Tencomnao", "role": "Author Response", "response": "Responses to Comments Methods section in abstract should be more detailed. Response: Thank you for highlighting the importance of including a more detailed methodology in the abstract. As suggested, we have incorporated a more detailed methodology in the revised version of the manuscript. Introduction lacks experimental data (IC50, Ki…) on the described activities of Agarwood compounds. Response: Thank you for insightful comment regarding the activities of agarwood compounds. While there is growing research on the therapeutic potential of Agarwood extracts, specific IC50 and Ki values for activity of agarwood compounds against Alzheimer's disease have not yet been reported in the literature. What is the rationale behind using the 13 selected protein targets? Response: Alzheimer's disease (AD) involves multiple pathways including amyloid-beta aggregation, tau phosphorylation, oxidative stress, inflammation, and neuronal cell death. Experimental evidence and animal studies revealed significant involvement of the 13 chosen target proteins. This information has been included in the introduction section of revised manuscript. Recommended range for each chemical descriptor needs to be added in table 2. Response: As per the reviewer’s suggestion, we have now included the recommended range for each chemical descriptor in Table 2 of the revised manuscript. Data describing ADMET of tested ligands should be added in the manuscript instead of supplementary material. Response: As suggested by the reviewer, we have included a detailed description of the ADMET properties of agarwood compounds in the revised manuscript. Please refer to “Prediction of ADMET analysis” section in the Results. BBB permeability is a crucial parameter for Alzheimer’s drugs; however, this property was not taken in consideration. Response: We appreciate the reviewer’s insightful comment on the importance of BBB permeability in Alzheimer's drugs. This study has indeed considered the necessity for agarwood compounds to cross the BBB to exhibit anti-Alzheimer's effects. Molecules with logBB > -1 were classified as BBB+, while molecules with logBB ≤ -1 were classified as BBB-. All compounds (agarotetrol, AGT, aquilarisin, ASN, aquilarisinin (ANN), aquilarixanthone, AXN, and pillion, PLN) used for MD simulations have logBB values greater than -1, so they are classified as BBB+. This information has been included in the “Molecular docking analysis” section of revised manuscript. Abbreviation of ligands should be used in the beginning, so the reader doesn’t get confused halfway through. Response: Thank you for your suggestion. We have included the abbreviations of ligands when they are first used in the beginning of the document. It’s not clear why complexes are exhibiting out of bound fluctuations outside the C- and N-terminal region in figure 5. RMSF section should be re-analysed taking into consideration the dynamic behavior of Apo forms of each protein. Response: Dear reviewer, we have revised the RMSF section and please refer to the revised Figure 5. We have explained the RMSF of each complex with respect to functional and non-functional domains. For example, in complex 2: the MAO-A protein structure C-terminal region (Ile471 to Leu524) showed increased RMSF to 20Å as this region appears to be an extended, peripheral loop, located away from the active site. This area showed higher RMSF due to its position on the protein’s surface, where it is more solvent-exposed and less constrained by interactions with other residues, making it inherently more flexible. Additionally, this region does not contribute directly to the catalytic function, so structural rigidity is less, unlike the core active site. Authors failed to highlight the key interactions for each protein and how they are at the origin of selectivity for the native ligands and if they are implicated in the binding of Agarwood compounds. Response: Dear reviewer, the key interactions for each protein at initial (0ns) and final (100ns) conformations were compared to identify the behavior and site-specific interaction of ligand throughout 100 ns simulation. This information is included in the “Intermolecular interactions observed in docked complexes” section of the revised manuscript. The 2D interaction figures are provided as Supplementary Figure 8a. No experimental studies in the literature were linked to back up the final conclusive remarks. Response: To date, there have not been experimental studies demonstrating the neuroprotective efficacy of the agarwood compounds (aquilarisin, aquilarisinin, aquilariaxanthone, pilloin, or agarotetrol) reported in the present study. The absence of experimental validation limits the ability to conclusively assert the therapeutic potential of these compounds in treating Alzheimer’s disease." } ] } ]	1	https://f1000research.com/articles/12-230
https://f1000research.com/articles/13-286/v1	17 Apr 24	{ "type": "Research Article", "title": "Optimization of bioethanol production using stress-tolerant yeast strains isolated from household alcoholic beverages (Tella, Tej, and Areke) and molasses (as substrate)", "authors": [ "Mulugeta Fentahun", "Berhanu Andualem", "Berhanu Andualem" ], "abstract": "Background Yeast strains that are tolerant to several environmental stresses are extensively employed to produce bioethanol from sugar-rich feedstock.\n\nMethods In our previous research findings, five wild indigenous strains of multi-stress tolerance yeast were isolated from traditional fermented alcoholic beverages (Tella, Tej, and Areke) in Ethiopia to the purpose of evaluate their potential use for bioethanol using sugarcane molasses as a substrate.\n\nResult The optimal parameters for maximum ethanol production were pH of 4.5, 25°Brix, 30°C, 4.0 g/l ammonium sulfate, supplement with yeast extract, and 72 h of incubation period. Under these optimum conditions, 13.13 ± 0.08% (v/v) ethanol was obtained. The yield of hydrous and anhydrous bioethanol from fermented molasses samples were produced at 94.20–95.60% (v/v) and 99.05–99.56% (v/v), respectively.\n\nConclusion According to this finding, Saccharomyces cerevisiae isolate MUT15F, Saccharomyces cerevisiae isolate MUT18F, and Saccharomyces cerevisiae isolate R9MU strains have been good candidates for production of industrial bioethanol.", "keywords": [ "Bioethanol", "Fermentation", "Molasses", "Optimization", "S. cerevisiae", "Stress tolerance" ], "content": "1. Introduction\n\nDue to depleted resources and environmental pollution caused throughout by fossil fuels, it was necessary to find out other eco-friendly renewable and sustainable sources of energy.1 Liquid biofuels, which account for 40% of all energy sources worldwide, were prioritized among renewable energies.2 The use of liquid biofuels helps in the reduction of greenhouse gas emissions, energy security, creation of job opportunities, regional development, rural development and reduction of poverty.3 Bioethanol is a promising alternative energy sources and a popular biofuel for transportation globally since it is a renewable, nontoxic, biodegradable resource and it is oxygenated.4 Production and use of bioethanol for transport fuel have recently attracted significant attention worldwide.5\n\nStarch, cellulose, and sucrose can all be used for the production of bioethanol, which may be found in molasses, coconut, sugar palm, cassava, banana stems, sweet potatoes, sugarcane bagasse, sunflower, woodchips and most recently microalgae.6,7 Among the different substrates available for production bioethanol sugarcane molasses is a good substrate because of cheap raw materials, readily available and compared to starchy or cellulosic materials for industrial bioethanol fermentation, ready for conversion with limited pretreatments.8 Molasses contain minerals, organic acids and high sugar content, which is 34-54%,9 especially the fructose contained in molasses varies between 5-12%, 25% to 40% sucrose, 12–35% reducing sugar and the level of total reduced sugar reaches 50-65%.10 In recent years, sugarcane molasses’ high prospects for making bioethanol have gained major attention from researchers.6,11\n\nSeveral factors affect the process of fermentation as well as the bioethanol yield such as the production microorganism strain, temperature, pH, oxygen, media composition, fermentation duration and initial concentration of sugar.12 Therefore, it is important to use optimal values among these parameters, for each selected production microorganism, to be able to produce the highest amounts of the required product.13 The choice and development of the efficient bioethanol production of yeast with a resistance multi-stress tolerance are crucial importance.14 Yeasts strain tolerant to high temperature, low pH, osmotic pressure and ethanol are ideal for industrial bioethanol production.15\n\nEthiopian Sugar Factories Fincha and Metehara used molasses from sugarcane used as a substrate for the production of ethanol after being diluted to 12–15 °Brix. These factories currently yield bioethanol production of less than 7% (v/v), in contrast to other countries that produce bioethanol with yields of 12–14% (v/v). This proves that the effectiveness of Ethiopian Factories is 50% less than the value of factories in other parts of the world. This study may provide significant information regarding potential tolerant of yeast that can withstand many stresses that could serve as a starter culture and substitute for strains of commercial yeast. Therefore, the research objective to maximize production bioethanol from molasses using stress-tolerant yeast strains from household alcoholic beverage preparations (Tella, Tej, and Areke) under different optimal conditions. Five yeast strains were used in this study, specifically Saccharomyces cerevisiae strains with accession numbers from the National Center for Biotechnology Information, such as strains R9MU (OR143320.1), strain R20MU (OR143322.1), strain MUT15F (OR209276.1), strain MUT18F (OR209286.1), and strain R19MU (OR143321.1). These strains of Saccharomyces cerevisiae were obtained in our stock cultures from the previous study because to their noteworthy tolerance to ethanol, 26% (v/v), glucose, 70% (w/v) and temperature, 45°C.\n\n\n2. Methods\n\nThe study was carried out at Department of Biology, University of Gondar, Northwestern Ethiopia. A sample of 70 liter of molasses was acquired from Ethiopia’s Methara Sugar Factory through September to January 2023. The molasses from sugarcane was collected in a clean, durable plastic container and kept at room temperature to the laboratory of microbiology at the University of Gondar for further use (Figure 1).\n\nThe physicochemical parameters of sugarcane molasses, including pH, specific gravity, Brix, moisture content, sucrose, and sulfated ash were estimated according to Refs. 16, 17. The quantity of total sugar in molasses samples was determined through Fehling method.18 The reducing sugar concentration in the molasses was determined by the 3,5-dinitrosalicylic acid (DNS) method.19 The moisture content molasses sample was determined thought oven (MB45, OHAUS, Switzerland) drying method. Sucrose content in the molasses sample was measured by measuring optical rotation in a polarimeter. Brix was measured with the help of a refractometer (Atagodensimeter model 2312; Atago Co. Ltd., Tokyo, Japan). Specific gravity of molasses samples were estimated by using baume universal hydrometer. The pH of the molasses samples were determined in the laboratory using pH meter (PHS-3C Digital) at ambient temperature. The ash content was established by muffle furnace at 650°C for 2 hours until constant weight.\n\n2.3.1 Pretreatment of molasses\n\nDistilled water was used to dilute raw molasses (1:1) w/w and pretreated with 99.8% sulfuric acid (H2SO4) until the pH reach 3.5 for the purpose of removing unwanted particles, dirt, and microbial contaminants.20 The mixture was heated to 90°C in a water bath (Gemmy industrial Corp, Taiwan) with continuous mixing for 30 minutes. Then it was allowed to stay for 24 h to cool and decant. The required Brix was achieved through the dilution of raw molasses with distilled water, and the medium was supplemented with 0.7 g/L ammonium sulfate (101217; MilliporeSigma) and then homogenized with a magnetic stirrer. Finally, the pH was adjusted at 4.5 using 0.5 M NaOH and autoclaved at 121°C for 15 min. The yeast strains were propagated and fermentation using the treated media to produce ethanol.\n\nFive native wild strains of Saccharomyces cerevisiae designated as R9MU (OR143320.1), R20MU (OR143322.1), MUT15F (OR209276.1), MUT18F (OR209286.1), and R19MU (OR143321.1) were selected in our stock cultures from the previous study. The capacity of these strains to ferment molasses was assessed, which were previously isolated from homemade alcoholic beverages (Tella, Tej, and Areke). Moreover, they were also selected for their remarkable tolerance because of their multi-stress tolerance yeast strains. The commercial yeast strain was used as a control for comparison with other strains.\n\nA loop full of 48 h old cultures has been taken on YEPD solid media and inoculated into each flask containing 100 ml of treated and sterilized molasses. The flasks were incubated for 24 h at 30°C temperature. Then, to increase the cell number, propagated yeast cultures were transferred to the second stage of propagation in Erlenmeyer flasks of 500 mL containing 100 mL of 30 °Brix molasses medium supplemented with 0.7 g/L ammonium sulfate (101217; MilliporeSigma) and sterilized at 121°C for 15 min and incubated at 30°C for another 24 h. Then after, optical density of propagated cultures was estimated using a UV-Vis spectrometer at 600 nm (Abron ISO 9001:2008) (Table 1). Finally, 48 h of propagated yeast culture which is ready as inoculum was transferred to 800 mL bottles containing 300 mL of sterilized and diluted molasses for optimization of feed-batch molasses fermentation to produce ethanol.21\n\nThe fed-batch fermentation system was used to optimize the process parameters, including molasses concentration (15, 20, 25, 30 and 35 °Brix), temperature (25, 30, 35, and 40°C), pH (3.5, 4.0, 4.5, 5.0, and 5.5), ammonium sulfate (101217; MilliporeSigma) supplement (1, 2, 3, 4 and 5 gL-1), different supplementation such as, yeast extract (VWR; 97063-370), urea (Koch Fertilizer, LLC), and ammonium nitrate (EM1.01187.5000; MilliporeSigma), and incubation periods (24, 48, 72, 96, and 120 h) by applying the one variable at a time method (VAT).22 Eight hundred milliliter bottles containing 300 ml of sterilized, diluted molasses were inoculated with 48 h of propagated yeast cells. The inoculum of each yeast strain was prepared by growing it in 200 ml of molasses medium for 48 h at 30°C. The inoculated bottles were plugged with sterilized cotton and incubated for 72 h in an incubator (J.P. Selecta Incubator, Spain). The anaerobic condition was maintained for 3 days. Samples were withdrawn after 72 h and analyzed for ethanol content. All treatments were studied in triplicate, and the mean and standard error were calculated.\n\n2.6.1 Effect of substrate concentration\n\nThe yeast strains were grown at varying molasses medium concentrations (15, 20, 25, 30, and 35 °Brix), which is an optimum operating condition at 30°C to investigate the effects of substrate concentration on ethanol production. The pH was adjusted before inoculation to 4.5 for all strains and incubated for 72 h. The concentration of ethanol (v/v %) was determined at the end of the fermentation period for each set of concentrations. At the end of the fermentation time for varied substrate concentrations, the amount of ethanol (v/v %) was measured.\n\n2.6.2 Effect of temperature\n\nEach yeast strain was grown on a molasses medium that was diluted by 30% and incubated at various temperatures (25, 30, 35, and 40°C). All yeast strains were incubated for 72 h at different temperatures, and the initial pH was adjusted to 4.5. The concentration of ethanol (v/v %) was determined at the end of the fermentation period at various temperatures.\n\n2.6.3 Effect of pH\n\nThe effect of different pH (3.5, 4.0, 4.5, 5.0, and 5.5) on ethanol production from diluted molasses (30%) by yeast strains was evaluated at 30°C. The pH was adjusted using 1N HCl or 0.1N NaOH. The fermentation process after incubation for 72 h and the percentage of ethanol were measured.\n\n2.6.4 Effect of ammonium sulfate supplement\n\nThe effects of supplement concentrations of ammonium sulfate (101217; MilliporeSigma) (1, 2, 3, 4, and 5 g/l) on ethanol production by each yeast strain were determined. All yeast strains were grown on a 30% molasses dilution with an initial pH of 4.0 for 72 h at 30°C. At the end of the fermentation period, the percentage of ethanol concentration for each supplement concentration was measured.\n\n2.6.5 Effect of different supplements\n\nThe effects of different supplements such as, yeast extract (VWR; 97063-370), urea (Koch Fertilizer, LLC), and ammonium nitrate (EM1.01187.5000; MilliporeSigma) on ethanol production by each yeast strain were conducted. All yeast strains were grown on a 30% molasses dilution with an initial pH of 4.5 for 72 h at 30°C. The effect of these supplements on ethanol concentration was evaluated after measuring the ethanol content of the fermented broth.\n\n2.6.6 Effect of fermentation period\n\nThe yeast strains were cultured on 30% diluted molasses with distilled water and incubated at 30°C for various fermentation times (24, 48, 72, 96, and 120 h). At the end of the fermentation period, the amount of ethanol in each fermentation period was measured as a percentage.\n\nThe Brix of fermented samples was measured using a refractometer (Atagodensimeter model 2312; Atago Co. Ltd., Tokyo, Japan). The optical density of propagated cultures was determined using a UV-Vis spectrometer at 600 nm (Abron ISO 9001:2008). The ethanol level of fermented molasses was determined by measurement of specific gravity using a Baume universal hydrometer.23 The levels of ethanol produced from the distillate were measured using an alcoholmeter. The calculation of ethanol concentration (ABV%) and bioethanol yield were calculated according to the Refs. 23 and 24 equation, respectively.\n\nWhere ABV is the alcohol by volume (%), SGin is the liquid’s initial specific gravity before to the addition of yeast, and SG is the liquid’s current specific gravity.\n\nBioethanol production was performed by scaling up the process in 3 L fermenter that each containing 2 L of pretreated molasses at a 30% molasses dilution with an initial pH of 4.5 in anaerobic conditions at 30°C for 72 h. After preparing a loop full of 48-hour-old cultures on YEPD solid media, 500 ml of molasses medium was inoculated for 48 hrs at 30°C. Using a fractional distillation apparatus, bioethanol was extracted from 2.5 L of fermented molasses in 3 L Erlenmeyer flasks.\n\nThe distillation flask (Pyrex, 500 ml, England) was filled with 400 ml of the fermented sample and placed on the machine’s heating unit with the water flow connected. The temperature was maintained manually at 78°C (the evaporation temperature of ethanol). The vapor was passed over a fractional column and the bulb of the thermometer, at which point the vapor was determined. The vapor was condensed to a liquid in the horizontal condenser, which was cooled with a flow of cold water. The distillate was collected in a receiver. The distillate was measured for volume. Repeated distillations were carried out until 90% of the bioethanol recovered fulfilled American Standard and Testing Material (ASTM) requirements for fuel.25 Furthermore, the resulting bioethanol was analyzed to determine the yield of the bioethanol. The levels of bioethanol produced from the molasses fermentation and distillation processes were measured using an alcoholmeter.\n\nThe initial concentrations of the bioethanol used in the first phase were ranged from 38.70 to 44.62% (v/v). Using a distillation column, the first bioethanol-diluted broth was pre-concentrated to a concentration that was nearly 95% (v/v) of the azeotropic point. During experimentation, samples were frequently taken out, and ethanol content was measured with an alcoholmeter. Near the azeotropic point, a distillate composition of 92.75% w/w was set,26 with a 99.5% w/w recovery of the ethanol feed.\n\nThe recovered bioethanol from the pre-concentration process, at a concentration close to 95% (v/v), was used in this study. Dehydrating 3A molecular sieve beads were used in this investigation. Molecular sieve beads were acquired from the Methara Sugar Factory in Ethiopia. The molecular sieves were dried in an oven (MB45, OHAUS, Switzerland) at 190–210°C for 24 h. The dried molecular sieves were kept in bottles, which were then stored in a glass chamber. Samples of distillate were dried using 3A molecular sieves overnight to capture water molecules. After being filtered, decanted, and redistilled to remove sieve dust and achieve anhydrous bioethanol with less than 1% water.27\n\nAll Data was analyzed using Statistical Package for Social Sciences software, version 23.0 (IBM SPSSInc., Chicago, IL, SPSS (RRID:SCR_002865), https://www.ibm.com/support/pages/downloading-ibm-spss-statistics-23). Analysis of variance (ANOVA) was conducted to compute the means and standard deviations of the triplicates analysis, and Tukey’s multiple range testing was used to assess if there were any significant differences between the means (p < 0. 05). The statistically significant difference was defined as p < 0. 05.\n\n\n3. Results\n\nThe process parameters such as molasses concentration (15, 20, 25, 30, and 35 °Brix), temperature (25, 30, 35, and 40°C), pH (3.5, 4.0, 4.5, 5.0, and 5.5), ammonium sulfate supplement (1, 2, 3, 4 and 5 g L-1), different supplementation (yeast extract, urea and ammonium nitrate), and period of incubation (24, 48, 72, 96, and 120 h) were all optimized using fed-batch fermentation (Figure 2). In fed-batch fermentation using a 200 ml treated molasses sample as the inoculum, yeast cell propagation was accomplished with a cell count of 1.710 – 1.892 × 108 cells per ml after 48 h (Table 1).\n\nThe physicochemical parameters of sugarcane molasses from the Methara Sugar Factory in Ethiopia are presented in Table 2. Molasses was composed of 84.00 ± 1.53° Brix, 50.35 ± 0.12% total reducing sugars, 33.31 ± 1.94% sucrose, 14.32 ± 0.65% reduced sugars, 19.30 ± 0.26% water content, and 17.01 ± 0.69% sulfated ash (w/v). Moreover, the molasses pH was 5.72 ± 0.09.\n\nAs shown in Table 3, various molasses percentages (Brix of 15%, 20%, 25%, 30%, and 35%) were utilized in fed batch fermentation to examine the optimization of ethanol concentration at 30°C and a pH of 4.5. Based on the present study, the five yeast strains from molasses with a Brix of 15–35 (oB) were produced ethanol in amounts ranging from 7.80 to 12.47% (v/v) of the concentration of ethanol. The standard industrial yeast strain was used as a control for comparison with other strains. Saccharomyces cerevisiae isolate MUT15F, 12.47 ± 0.07% (v/v) and Saccharomyces cerevisiae isolate R9MU, 11.81 ± 0.16% (v/v) have been shown statistically (p ≤ 0.05) greater ethanol concentrations than the remaining strains. Production of ethanol in all yeast strains was significantly (p ≤ 0.05) reduced in either low concentration (15–20%) or high concentration (30–35%). Higher or lower concentrations of molasses have a significant negative effect on ethanol production. These findings have been shown that the optimal molasses concentration for all yeast strains to produce ethanol was 25% (Table 3).\n\nThe effect of temperature at 25, 30, 35, and 40°C in the production process of ethanol was also investigated. The ethanol production maximum was shown in Table 3 to be achieved at a temperature of 30°C, and subsequent increases in temperature reduced the production of ethanol. According to this finding, Saccharomyces cerevisiae isolate MUT15F, 11.94 ± 0.06% (v/v) and Saccharomyces cerevisiae isolate R9MU, 11.15 ± 0.08% (v/v) were statistically (p ≤ 0.05) greater in ethanol concentration than the other strains, which was included strain MT (the commercial strains) at 30°C after 72 h of the fermentation period. Saccharomyces cerevisiae isolate MUT18F has been shown a statistically lower (p ≤ 0.05) amount of ethanol, 9.84 ± 0.16% (v/v) at 30°C than the other yeast strains. There was no statistically significant (p ≥ 0.05) difference between strains of Saccharomyces cerevisiae R19MU, Saccharomyces cerevisiae R20MU, and MT (standard) at 30°C. Below 25°C or above 30°C, the production of ethanol by all yeast strains was significantly (p ≤ 0.05) reduced. According to this finding, the optimum incubation temperature for all yeast strains was 30°C.\n\nThe results of different ethanol productions with pH values ranging from 3.5 to 5.5 were shown in Table 3. The ethanol production gradually increases along with the increase in pH and reaches a maximum at a pH 4.5. The production was decreased slightly for pH values higher than 5. The yeast strains have been shown a different pattern of ethanol production of 6.08–11.94% (v/v) from pH 3.5 to pH 5.5. Based on the present study, ethanol produced by yeast strain Saccharomyces cerevisiae isolate MUT15F, 11.94 ± 0.06% (v/v) and Saccharomyces cerevisiae isolate R9MU, 11.15 ± 0.08% (v/v) were shown a statistically significant (p ≤ 0.05) greater concentration than all the remaining strains at pH 4.5. Saccharomyces cerevisiae isolate R20MU has been shown statistically (p ≤ 0.05) lower ethanol concentration than all other strains. There was no statistically significant (p ≥ 0.05) difference between strains Saccharomyces cerevisiae isolate R19MU, Saccharomyces cerevisiae isolate R20MU, and the MT (standard) at pH 4.5. All the strains were more effective to produce high amount of ethanol at pH 4.5 and less ethanol produced at pH 3.5 (Table 3).\n\nIn Table 4, the effect of ammonium sulfate as a nitrogen source is shown. Under optimal conditions, 4.0 g/l of ammonium sulfate produced the highest amount of ethanol. As the ammonium sulfate concentration increased from 1.0 to 4.0 g/l, the production ethanol also significantly (p ≤ 0.05) increased and then reduced in amount as concentration of ammonium sulfate increased. In this study, the amount of ethanol production by Saccharomyces cerevisiae isolate MUT15F was increased from 11.95 ± 0.1 to 13.13 ± 0.08% (v/v) between 1.0 and 4.0 g/l ammonium sulfate. According to this finding, Saccharomyces cerevisiae isolate MUT15F (13.13 ± 0.08% (v/v) has been shown statistically (p ≤ 0.05) greater ethanol production than that of the other strains including strain MT (the commercial strains). Saccharomyces cerevisiae isolate MUT18F was shown significantly (p ≥ 0.05) less ethanol concentration, (10.76 ± 0.03% (v/v)) at 4.0 g/l ammonium sulfate than all other strains.\n\nAmmonium nitrate, urea, and yeast extract are different nitrogen sources and their effects were studied at a concentration of 1 g supplementation per liter, as presented in Table 4. Yeast extract supplement was produced high levels of ethanol when in contrast to other nitrogen sources and the control. Compared to the other supplemented media, an ammonium nitrate-supplemented medium was shown the lowest level of ethanol. Saccharomyces cerevisiae isolate MUT15F (12.24 ± 0.08% (v/v)) and Saccharomyces cerevisiae isolate R9MU (11.60 ± 0.06% (v/v)) were statistically (p ≤ 0.05) greater in ethanol production from yeast extract supplementation than stains MT (the commercial strains) (10.89 ± 0.12% (v/v)) and Saccharomyces cerevisiae isolate R19MU (10.83 ± 0.02% (v)). The lowest ethanol concentration was observed (10.43 ± 0.12% (v/v)) by strains Saccharomyces cerevisiae isolate R20MU and Saccharomyces cerevisiae isolate MUT18F (9.95 ± 0.09% v/v) from supplementation of yeast extract compared to the rest of the yeast strains. The maximum ethanol concentration was observed (11.95 ± 0.1% (v/v)) by Saccharomyces cerevisiae isolate MUT15F and Saccharomyces cerevisiae isolate R9MU (11.17 ± 0.08% (v/v)) from the supplementation of urea than the rest of the yeast strains. All yeast strains grown without nitrogen sources (control) had the lowest ethanol concentration in comparison to other supplemented media in this study.\n\nIncubation periods effect on ethanol production after fermentation processes were carried out for 24, 48, 72, 96, and 120 h is presented in Table 4. Ethanol production was increased during incubation periods up to 72 h and then decreased again to reach the minimum levels at 120 h of incubation. After 72 h of fermentation, the maximum amount of ethanol was produced. Therefore, the optimum period of incubation to produce ethanol was 72 h except for yeast strains Saccharomyces cerevisiae isolate MUT15F (12.02 ± 0.13% (v/v)) and Saccharomyces cerevisiae isolate R19MU (10.77 ± 0.04% (v/v)), which achieved their maximum production after 96 h. Saccharomyces cerevisiae isolate R9MU (11.15 ± 0.08% (v/v)) was shown statistically (P ≤ 0.05) higher ethanol production.\n\nThe yield of anhydrous and hydrous bioethanol generated from molasses sample is shown in Table 5. A continuous distillation was utilized to accomplish fractional distillation on each sample of fermented molasses, yielding various distillate quantities and hydrous bioethanol (Figure 3). After pre-concentration, the distillation column concentrates hydrous bioethanol up to a final concentration of 94.20–95.60% (v/v) was determined using an alcoholmeter. The volume of bioethanol produced using different yeast strains in molasses samples varied significantly (p ≤ 0.05). Anhydrous bioethanol of 99.05–99.56% (v/v) was achieved by further distillation employing a 3A molecular sieve bead (Figure 4). Saccharomyces cerevisiae isolate R9MU has been shown the highest anhydrous bioethanol concentration (99.56% (v/v)) after dehydration, while Saccharomyces cerevisiae isolate MUT18F produced the least amount of anhydrous bioethanol (99.05% (v/v)) as shown in Table 5. The maximum yield of bioethanol (8.50% v/v) was produced by S. cerevisiae isolate R9MU. All the bioethanol produced was clear and colorless.\n\n\n4. Discussion\n\nDifferent molasses concentrations were used to determine their effect on the ethanol concentration using the optimized pH of 4.5. The finding indicated that 25 °Brix was the most suitable sugar concentration for isolate of Saccharomyces cerevisiae MUT15F and Saccharomyces cerevisiae R9MU to produce ethanol concentrations of 12.47 ± 0.07% (v/v) and 11.81 ± 0.16% (v/v) at 72 h, respectively. The ethanol concentration in this study was higher than the findings of other authors.28,29 However, Gu et al.30 used yunnan molasses and recorded that the highest production of ethanol was 16% (v/v) by strain 1912 and 13.7% (v/v) by strain 1190 at 30% molasses sugar for 72 h. Muruaga et al.31 also reported that for the A10 strain isolated from molasses at 250 g/L of initial sugar, the highest level of ethanol production was 13.20% (v/v). The production of ethanol by all yeasts significantly decreased at high molasses concentrations (35%). This may be because to the high quantities of substrates being inhibitory to the fermentation process of yeasts due to osmotic stress.\n\nTemperature is the key factor that affects the growth, metabolism, and ethanol production capability of the fermenting organisms. During this investigation, ethanol production by all yeast strains increased with the increase in temperature, reaching its maximum value at 30°C. This finding is in line with the findings of.32,33 Below 25°C or above 30°C, the production of ethanol by all yeast strains was significantly reduced. This might be due to the fact that denaturation happens at high temperatures and reduced enzyme activity at low temperatures.\n\nOptimal pH values are essential for the activity of plasma membrane-based proteins, including enzymes and transport proteins.34 Based on the present finding, the production of ethanol gradually increased along with the increase in pH and reached a maximum production for a pH equal to 4.5, then started to decline. An optimum pH of S. cerevisiae was obtained at pH 4.5. This is line with the report of other investigators.33,35,36 In contrast to this investigation, Ercan et al.37 found that optimum ethanol production using S. cerevisiae was obtained at pH 5.5. The most favorable pH of S. cerevisiae for ethanol production ranges from 4.0 to 5.0.27\n\nNitrogen limitation has been shown to affect cell growth and biomass formation as well as directly affect the fermentation rate.38 Although most of the nutrients needed for yeast development are present in molasses, sufficient nitrogen is frequently added to promote yeast growth and the production of ethanol.39 Numerous investigations were conducted in previous years to optimize supplies of nitrogen and other supplements.40 The obtained data revealed that the addition of ammonium sulfate to the fermentation medium also increased ethanol production from 1.0 to 4.0 g/l. In the present finding, the amount of ethanol production by Saccharomyces cerevisiae isolate MUT15F increased from 11.95 ± 0.1 to 13.13 ± 0.08% (v/v) between 1.0 and 4.0 g/L ammonium sulfate. Our study agrees with previous studies by Ref. 41 the optimal medium for S. cerevisiae to produce ethanol was 4 g/L of ammonium sulfate with 10% (v/v) ethanol content. Anupama et al.42 was reported that the usage of ammonium sulfate at a concentration of 3 g/l resulted in an optimal 5.6% yield of ethanol.\n\nNutrients are quite effective in the production of ethanol from sugar cane molasses.22 Different nitrogen sources (yeast extract, urea, and ammonium nitrate) were investigated as possible nitrogen supplements for sugar cane molasses. This study revealed that all nitrogen sources investigated had a positive effect on the production of ethanol. The maximum ethanol production was recoreded as nitrogen source yeast extract from Saccharomyces cerevisiae isolate MUT15F (12.24 ± 0.08% (v/v)) and Saccharomyces cerevisiae isolate R9MU (11.60 ± 0.06% (v/v)). These results were in agreement with Rasmey et al.,35 who found that yeast extract addition significantly improved the ethanol concentration. In this finding, yeast extract provides convenient growth factors for yeast growth and it is in line with Ortiz-Muñiz et al.43 report. On the other hand, it was also detected that ammonium sulfate was a good nitrogen source that stimulated ethanol production from sugar cane molasses. An effective and common source of nitrogen for yeast development is ammonium sulfate; it is cheap and has been chosen for future experiments.39\n\nControlling a number of variables can result in increased ethanol fermentation activity. Temperature, pH, substrate concentration, and fermentation time also greatly influences ethanol production by yeasts.12,44 According to this finding, ethanol concentration was reaching its maximum amount after 72 h except for Saccharomyces cerevisiae isolate MUT15F (12.02 ± 0.13% (v/v)) and Saccharomyces cerevisiae isolate R19MU (10.77 ± 0.04% (v/v)), which achieved their maximum production after 96 hrs. After three days of growth, it was determined that yeasts produce the highest amounts of ethanol by fermenting sugary substrates with actively growing yeast cells.45 Ethanol production increased gradually, but a slight decrease in ethanol concentration with an increasing incubation period could be due to the loss of yeast cell viability and consumption of it by the yeast cells as time passed.46 The current findings showed that optimization of temperature, pH, incubation duration, and substrate concentration is quite important for the maximization of ethanol production, as previously reported by other studies.47,48\n\nThe result of our study indicated that hydrous bioethanol was concentrated in the distillation column to a final concentration of 94.20–95.60% (v/v). Further distillation using a 3A molecular sieve bead resulted in anhydrous bioethanol of about 99.05-99.56% (v/v). The highest anhydrous bioethanol concentration was obtained from Saccharomyces cerevisiae isolate R9MU (99.56% (v/v)) after distillation and dehydration. These results were in agreement with Yang et al.,49 who produced bioethanol in the form of hydrous ethanol (96% v/v).\n\n\n5. Conclusion\n\nSaccharomyces cerevisiae isolate MUT15F yielded a maximum ethanol production of 13.13 ± 0.08% (v/v) at 30 °Brix of molasses, 30°C, 4g NH4SO4, 4.5 pH, and 72 h fermentation. In contrast to unoptimized conditions, the concentration of ethanol increased approximately by 77.4% after optimization. The candidate yeast strains designated as Saccharomyces cerevisiae isolate MUT15F, Saccharomyces cerevisiae isolate R9MU, and Saccharomyces cerevisiae isolate MUT18F were producing maximum alcohol under the optimized fermentation conditions and have the potential to be used for industrial bioethanol production. Saccharomyces cerevisiae isolate R9MU produced the highest bioethanol yield (8.50% v/v). Traditional alcoholic beverages such as Tella, Tej, and Areke can serves as a potential yeast source that able to produce high concentration of bioethanol.\n\n\nDeclarations\n\nConceptualization, M.F. and B.A.; Methodology, M.F.; Data analysis, M.F.; Investigation, M.F.; Resources, M.F.; Writing – original draft, M.F.; Writing – review & editing, M.F. and B.A.; Visualization, M.F.; Supervision, B.A.; Funding acquisition, M.F. All authors have read and agreed to the published version of the manuscript.\n\n\nEthical approval and consent\n\nEthical approval and consent were not required.", "appendix": "Data availability\n\nFigshare: Molasses concentration (15, 20, 25, 30 and 35 °Brix), and pH (3.5, 4.0, 4.5, 5.0, and 5.5) of data analysis. https://doi.org/10.6084/m9.figshare.24941358. 50\n\nThis project contains the following underlying data:\n\n- Molasses concentration (15, 20, 25, 30 and 35 °Brix), and pH (3.5, 4.0, 4.5, 5.0, and 5.5) of data analysis.\n\n- SPSS data analysis of supplements, and period of incubation (24, 48, 72, 96, and 120 h).\n\n- SPSS data analysis of temperature (25, 30, 35, and 40°C), and ammonium sulfate supplement (1, 2, 3, 4 and 5 g L-1).\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 want to express our gratitude to Methara Sugar Factory for support molecular sieves and molasses.\n\n\nReferences\n\nKiran B, Kumar R, Deshmukh D: Perspectives of microalgal biofuels as a renewable source of energy. 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Publisher Full Text\n\nGu Y, Qiao M, Zhou Q, et al.: Hyperproduction of Alcohol Using Yeast Fermentation in Highly Concentrated Molasses Medium.2001.\n\nMuruaga ML, Carvalho KG, Domínguez JM, et al.: Isolation and characterization of Saccharomyces species for bioethanol production from sugarcane molasses: Studies of scale up in bioreactor. Renew. Energy 2016; 85: 649–656. Publisher Full Text\n\nKhoja AH, Ali E, Zafar K, et al.: Comparative study of bioethanol production from sugarcane molasses by using Zymomonas mobilis and Saccharomyces cerevisiae. African. J. Biotechnol. 2015; 14(31): 2455–2462. Publisher Full Text\n\nPasricha S: Research Article Research Article. Arch. Anesthesiol. Crit. Care. 2020; 4(4): 527–534.\n\nReda FM, Hussein BM, Enan G: Selection and characterization of two probiotic lactic acid bacteria strains to be used as starter and protective cultures for food fermentations. J. Pure Appl. Microbiol. 2018; 12(3): 1499–1513. Publisher Full Text\n\nRasmey AH, Hassan H, Aboseidah A, et al.: Enhancing Bioethanol Productivity from Sugarcane Molasses by Saccharomyces cerevisiae Y17 KP096551. Egypt. J. Bot. 2018. Publisher Full Text\n\nZaghlou RA, Ismail SA, Enan G, et al.: Maximization of Bio-Ethanol Production by Yeasts using Sugar Cane and Sugar Beet Molasses. Adv. Anim. Vet. Sci. 2021; 9(12): 2069–2076. Publisher Full Text\n\nErcan Y, Irfan T, Mustafa K: Optimization of ethanol production from carob pod extract using immobilized Saccharomyces cerevisiae cells in a stirred tank bioreactor. Bioresour. Technol. 2013; 135: 365–371. PubMed Abstract \| Publisher Full Text\n\nBisson LF, Butzke CE: Diagnosis and Rectification of Stuck and Sluggish Fermentations. Am. J. Enol. Vitic. 2000; 51(2): 168–177. Publisher Full Text\n\nMendes-Ferreira A, Mendes-Faia A, Leão C: Growth and fermentation patterns of Saccharomyces cerevisiae under different ammonium concentrations and its implications in winemaking industry. J. Appl. Microbiol. 2000; 97(3): 540–545. Publisher Full Text\n\nMalherbe S, Bauer FF, Du Toit M: Understanding problem fermentations - A review. South African J. Enol. Vitic. 2007; 28(2): 169–186. Publisher Full Text\n\nDarvishi F, Moghaddami NA: Optimization of an Industrial Medium from Molasses for Bioethanol Production Using the Taguchi Statistical Experimental-Design Method. Ferment. 2019; 5(1): 14. Publisher Full Text\n\nAnupama M, Guru DM, Ayyanna C: Optimization of Fermentation Medium for the Production of ethanol from jaggery using box-behnken design.2010; 34–45.\n\nOrtiz-Muñiz B, Carvajal-Zarrabal O, Torrestiana-Sanchez B, et al.: Kinetic study on ethanol production using Saccharomyces cerevisiae ITV-01 yeast isolated from sugar cane molasses. J. Chem. Technol. Biotechnol. 2010; 85(10): 1361–1367. Publisher Full Text\n\nTofighi A, Mazaheri Assadi M, Asadirad MHA, et al.: Bio-ethanol production by a novel autochthonous thermo-tolerant yeast isolated from wastewater. J. Environ. Health Sci. Eng. 2014; 12(1). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLefyedi ML, Marais GJ, Dutton MF, et al.: The Microbial Contamination, Toxicity and Quality of Turned and Unturned Outdoor Floor Malted Sorghum. J. Inst. Brew. 2005; 111(2): 190–196. Publisher Full Text\n\nDe La Cruz V, Hernández S, Martín M, et al.: Integrated Synthesis of Biodiesel, Bioethanol, Isobutene, and Glycerol Ethers from Algae. Ind. Eng. Chem. Res. 2014; 53(37): 14397–14407. Publisher Full Text\n\nJambo SA, Abdulla R, Marbawi H, et al.: Response surface optimization of bioethanol production from third generation feedstock - Eucheuma cottonii. Renew. Energy 2019; 132: 1–10. Publisher Full Text\n\nShaghaghi-Moghaddam R, Jafarizadeh-Malmiri H, Mehdikhani P, et al.: Optimization of submerged fermentation conditions to overproduce bioethanol using two industrial and traditional Saccharomyces cerevisiae strains. Green Process Synth. 2019; 8(1): 157–162. Publisher Full Text\n\nYang R: Production of Ethanol from Sudanese Sugar Cane Molasses and Evaluation of Its Quality. J. Food Process. Technol. 2017; 03(07): 3–5. Publisher Full Text\n\nFentahun M, Andualem B: Figshare: Molasses concentration (15, 20, 25, 30 and 35 °Brix), and pH (3.5, 4.0, 4.5, 5.0, and 5.5) of data analysis.2023. Publisher Full Text" }	[ { "id": "292438", "date": "12 Jul 2024", "name": "Boris U. Stambuk", "expertise": [ "Reviewer Expertise Fuel ethanol 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 by Fentahun and Andualem “Optimization of bioethanol production using stress-tolerant yeast strains isolated from household alcoholic beverages (Tella, Tej, and Areke) and molasses (as substrate)” describes experiments performed to produce ethanol from molasses by yeasts isolated in Ethiopia. In this reviewer point of view, the manuscript needs extensive English revision. Since the manuscript does not have line numbers, is hard to point out which sentences have problems, I have mark some of them in the attached document (https://f1000research.s3.amazonaws.com/linked/663424.146910_-_Optimization_of_bioethanol_production_using_stress-tolerant_yeast_strains_isolated_from_household_alcoholic_beverages_%28Tella%2C_Tej%2C_and_Areke%29_and_molasses_%28as_substrate%29_-_Boris_Stambuk.pdf)\nAuthors should also take care in the use of references: -For example, in the first paragraph of the Introduction they claim “Bioethanol is a promising alternative energy sources and a popular biofuel for transportation globally since it is a renewable, nontoxic, biodegradable resource and it is oxygenated” and they cite ref. 4, but this reference deals with Lactobacillus paracasei, metabolic syndrome and rats! (?) -In the second paragraph they are describing the composition of molasses, and ref. 10 is cited, but this reference deals with ultrasonic and ozonation pretreatments of molasses for methane production…… In the context of bioethanol production, I believe that these references (Amorim HV, Basso LC, Lopes ML. Sugar cane juice and molasses, beet molasses and sweet sorghum: composition and usage. In: The Alcohol Textbook. Nottingham: Nottingham University Press; 2009; https://doi.org/10.1186/s13068-018-1221-x; and https://doi.org/10.1038/s41598-023-37618-8) would be more appropriated.\n-In the same paragraph they claim “In recent years, sugarcane molasses’ high prospects for making bioethanol have gained major attention from researchers.” and cite ref. 11, but this reference deals with vinegar production! (?)\n-In the same paragraph they claim “12–35% reducing sugar and the level of total reduced sugar reaches 50-65%”….. Confusing!\n-In the Discussion section, second paragraph, they claim “This finding is in line with the findings of.” and cite ref. 32 and 33, but this last reference is from an Anesthesiology journal (?), with no title of the manuscript.\n-At the beginning of the third paragraph of the Discussion they claim “Optimal pH values are essential for the activity of plasma membrane-based proteins, including enzymes and transport proteins.” and cite ref. 34, but this reference is not related to yeasts, but lactic acid bacteria (?)\n-Several paragraphs later they claim “Ethanol production increased gradually, but a slight decrease in ethanol concentration with an increasing incubation period could be due to the loss of yeast cell viability and consumption of it by the yeast cells as time passed” and cite ref. 46, which seams also inappropriate!\n-Indeed, SEVERAL of the references (# 3, 4, 8, 9, 16, 17, 30, 33, 35, 42, 44) are with problems, lacking article title and/or journal name and/or volume and/or page numbers……\n-In the fourth paragraph of the Introduction they claim “other countries that produce bioethanol with yields of 12–14% (v/v)”, this should have a reference!\n-At the end of the Introduction they claim “from the previous study because to their noteworthy tolerance to ethanol, 26% (v/v), glucose, 70% (w/v) and temperature, 45°C.” and in item 2.4 they also claim “from the previous study”…..\n-Item 2.3.1 “and pretreated with 99.8% sulfuric acid” pretreated how?\n-The manuscript is highly repetitive, “ammonium sulfate (101217; MilliporeSigma)” for example is repeated at least 4 times, section 2.6 (and 2.6.1 to 2.6.6) is also repetitive!\n-Item 2.4 mentions a “commercial yeast strain” without any indication of which one is (also “MT standard yeast”)\n-Item 2.5 a “YEPD solid media” is mentioned, without any description! At the end of this item (and in several other sections) is claimed “feed-batch molasses fermentation”, but they did not do that, they performed batch fermentations, there was no feeding…..!\n-Item 2.6 they claim “The anaerobic condition was maintained for 3 days”, but certainly no “anaerobic” condition was indeed used!\n-“polarimeter’ and “alcoholmeter” needs more details....\n-Figures 1, 2, 3 and 4 are totally useless, they should be removed.\n-The paragraph after Table 2 reproduces all the data presented in Table 2, should be removed! Etc., etc., etc., etc………\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 source data required\n\nAre the conclusions drawn adequately supported by the results? No", "responses": [ { "c_id": "12934", "date": "16 Dec 2024", "name": "Mulugeta Fentahun", "role": "Author Response", "response": "Mulugeta Fentahun Nega Thanks to Prof. Boris U. Stambuk for your constructive comments. We conducted a number of changes. I greatly appreciate your feedback and suggestions. Please find my responses to your comments below: Introduction Revision: Comment: The manuscript needs extensive English Response: Thank you for your comments: We have corrected grammatical errors throughout the manuscript and made the necessary corrections to enhance clarity and readability accordingly (refer the revised article). Comment: Authors should also take care in the use of references. The first paragraph of the introduction they claim “Bioethanol is a promising alternative energy sources and a popular biofuel for transportation globally since it is a renewable, nontoxic, biodegradable resource and it is oxygenated” and they cite ref. 4, but this reference deals with Lactobacillus paracasei, metabolic syndrome and rats! (?) Response: Thank you for this valuable suggestion. We corrected it as Scott A, Bryner M: Alternative Fuels: Rolling out Next-Generation Technologies. Chem. Week. 2006; 27: 17-21. Comment: In the second paragraph they are describing the composition of molasses, and ref. 10 is cited, but this reference deals with ultrasonic and ozonation pretreatments of molasses for methane. Response: Thank you for your comments. We corrected it as Lino FS, Basso TO, Sommer, MO: A synthetic medium to simulate sugarcane molasses. Biotechnology for Biofuels. 2018; 11(1). 221. 10.1186/s13068-018-1221-x Comment: In the same paragraph they claim “In recent years, sugarcane molasses’ high prospects for making bioethanol have gained major attention from researchers.” and cite ref. 11, but this reference deals with vinegar production! (?) Response: Thank you for your comments. We corrected it as Akbas MY, Stark BC: Recent trends in bioethanol production from food processing byproducts. J. Ind. Microbiol. Biotechnol. 2016. 10.1007/s10295-016-1821-z Comment: In the same paragraph they claim “12–35% reducing sugar and the level of total reduced sugar reaches 50-65%”….. Confusing! Response: Thank you for your critical insight. we corrected it based on your suggestions (refer the revised article). Comment: In the fourth paragraph of the Introduction, they claim “other countries that produce bioethanol with yields of 12–14% (v/v)”, this should have a reference! Response: Thank you for your critical insight. we cited as Kotarska K, Czupryński B, Kłosowski G: Effect of various activators on the course of alcoholic fermentation. J. Food Eng. 2006; 77: 965-971. Comment: At the end of the Introduction, they claim “from the previous study because to their noteworthy tolerance to ethanol, 26% (v/v), glucose, 70% (w/v) and temperature, 45°C.” and in item 2.4 they also claim “from the previous study”. Response: Thank you for your critical insight. We cited it as Fentahun M, Andualem B: Characterization of wild indigenous yeasts that can withstand different stress from traditional fermented beverages in the Amhara Region, Ethiopia. Food Sci. Appl. Biotechnol. 2024; 7(2): 262- 277. 10.30721/fsab2024.v7.i2.4052024 Methodology Revision: Comment: Figures 1, 2, 3 and 4 are totally useless, they should be removed. Response: I appreciate your suggestion. The article's figures were compiled based on comments of the reviews of F1000 research. Comment: Alcoholmeter and polarimeter need more detail. Response: Thank you for your comments: We have updated the method according to your comments (refer the revised article). Comment: Item 2.6 they claim “The anaerobic condition was maintained for 3 days”, but certainly no “anaerobic” condition was indeed used!. Response: Thank you for your comments: Yes. No “anaerobic” condition was used. We made correction based on your comments. Comment: Item 2.5 a “YEPD solid media” is mentioned, without any description! At the end of this item (and in several other sections) is claimed “feed-batch molasses fermentation”, but they did not do that, they performed batch fermentations, there was no feeding…..! Response: Thank you for your critical insight. We corrected YEPD solid media (refer the revised article) and Fed-batch fermentations were carried out, and feeding was done to boost the number of cells. A propagated yeast culture in 200 milliliters was used by adding 100 ml of sterile new batch molasses in to 100 ml inoculated flask for 24 hrs at 30°C. (refer item 2.5 propagation of yeast strains page 4 the revised article). Comment: Item 2.4 mentions a “commercial yeast strain” without any indication of which one is (also “MT standard yeast”). Response: Thank you for your comments: We have updated the method according to your comments (refer the revised article). Comment: -Item 2.3.1 and pretreated with 99.8% sulfuric acid” pretreated how? Response: Thank you for your comments: Because the pH raw molasses was adjusted at 3.5 using 99.8% H2SO4, and then the treated medium was standing overnight for decantation. The clear upper suspension of the broth was then carefully poured into a newly sterile fermentation flask. Discussion Revision: Comment: In the discussion section, second paragraph, they claim “This finding is in line with the findings of.” and cite ref. 32 and 33, but this last reference is from an Anesthesiology journal (?). Response: Thank you for the suggestion. We have made revisions based on your comments as Abdel-Banat BMA, Hoshida H, Ano A, et al.: High-temperature fermentation: How can processes for ethanol production at high temperatures become superior to the traditional process using mesophilic yeast? Appl Microbiol Biotechnol. 2010; 85:861–7. Comment: At the beginning of the third paragraph of the discussion they claim “Optimal pH values are essential for the activity of plasma membrane-based proteins, including enzymes and transport proteins.” and cite ref. 34, but this reference is not related to yeasts, but lactic acid bacteria (?). Response: Thank you for your comments. We corrected it as Lopandic K, Zelger S, Bánszky L K, et al.: Identification of yeasts associated with milk products using traditional and molecular techniques. Food Microbiol. 2006; 23:341–50. 10.1016/J.FM.2005.05.001. Comment: Several paragraphs later they claim “Ethanol production increased gradually, but a slight decrease in ethanol concentration with an increasing incubation period could be due to the loss of yeast cell viability and consumption of it by the yeast cells as time passed” and cite ref. 46, which seams also inappropriate! Response: Thank you for your comments. We corrected it as Zabed H, Faruq G, Sahu JN, et al.: Bioethanol production from fermentable sugar juice. Sci. World J. 2014; 10.1155/2014/957102 Thank you once again for your valuable feedback. Your comments have significantly contributed to improving the quality of this paper. Best regards, Authors" } ] }, { "id": "328845", "date": "27 Nov 2024", "name": "Evika Sandi Savitri", "expertise": [ "Reviewer Expertise biotechnology", "plant biotechnology" ], "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 strength of this article is that it uses a strain of yeast that was discovered, namely specifically Saccharomyces cerevisiae strains with accession numbers from the National Center for Biotechnology Information, such as strains R9MU (OR143320.1), strain R20MU (OR143322.1), strain MUT15F (OR209276.1), strain MUT18F (OR209286.1), and strain R19MU (OR143321.1)\nINTRODUCTION\n• Authors should also take care in the use of references:\nThe choice and development of the efficient bioethanol production of yeast with a resistance multi-stress tolerance are crucial importance14.\n14. Chohan NA, Aruwajoye GS, Kana EBG: Valorisation of potato peel wastes for bioethanol production using simultaneous saccharification and fermentation: Process optimization and kinetic assessment. Renew. Energy 2020; 146: 1031–1040. Publisher Full Text\nshould use more specific literature for of yeast with a resistance multi-stress tolerance\nThese strains of Saccharomyces cerevisiae were obtained in our stock cultures from the previous study because to their noteworthy tolerance to ethanol, 26% (v/v), glucose, 70% (w/v) and temperature, 45°C. • Authors should also references for this statement • should provide citations from previous research for the characteristics of the yeast\n\nMETHODS\n• Figures 1, 2, 3 and 4 are totally useless, they should be removed.\nThe levels of bioethanol produced from the molasses fermentation and distillation processes were measured using an alcoholmeter.\n• Alcoholmeter need more detail\n\nCONCLUSION Saccharomyces cerevisiae isolate MUT15F yielded a maximum ethanol production of 13.13 0.08% (v/v) at 30 °Brix of molasses, 30°C, 4g NH4SO4, 4.5 pH, and 72 h fermentation\n• In the discussion sub-chapter, it should be discussed in more depth how the characteristics of the recommended yeast compare with the standard yeast as a commercial yeast strain was used as a control for comparison.\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?\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": [ { "c_id": "12935", "date": "16 Dec 2024", "name": "Mulugeta Fentahun", "role": "Author Response", "response": "Mulugeta Fentahun Nega Dr. Evika Sandi Savitri thank you very much for your insightful comments and the time you dedicated to review. I greatly appreciate your feedback and suggestions. Please find my responses to your comments below: Introduction Revision: Comment: Authors should also take care in the use of references: Example the choice and development of the efficient bioethanol production of yeast with a resistance multi-stress tolerance are crucial importance. References or citation 14. Response: Thank you for this suggestion We corrected it as Alfenore S, Molina C, Guillouet SE, Uribelarrea JL, et al.: Improving ethanol production and viability of Saccharomyces cerevisiae by a vitamin feeding strategy during fed-batch process. Appl Microbiol Biotechnol.2002; 60: 67–72. Comment: Authors should provide citations from previous research for the characteristics of the yeast strains of Saccharomyces cerevisiae were obtained in our stock cultures from the previous study because to their noteworthy tolerance to ethanol, 26% (v/v), glucose, 70% (w/v) and temperature, 45°C? Response: Thank you for this valuable suggestion. We corrected it as Fentahun M, Andualem B: Characterization of wild indigenous yeasts that can withstand different stress from traditional fermented beverages in the Amhara Region, Ethiopia. Food Sci. Appl. Biotechnol. 2024; 7(2): 262- 277. 10.30721/fsab2024.v7.i2.4052024 Methodology Revision: Comment: Figures 1, 2, 3 and 4 are totally useless, they should be removed. Response: I appreciate your suggestion. The article's figures were compiled based on comments of the reviews of F1000 research. Comment: Alcoholmeter need more detail. Response: Thank you for your comments: We have updated the method according to your comments (refer the revised article). Discussion Revision: Comment: In the discussion sub-chapter, it should be discussed in more depth how the characteristics of the recommended yeast compare with the standard yeast as a commercial yeast strain was used as a control for comparison. Response: Thank you for the suggestion. We have made revisions (refer the revised article). Thank you once again for your valuable feedback. Your comments have significantly contributed to improving the quality of this paper. Best regards, Authors" } ] } ]	1	https://f1000research.com/articles/13-286
https://f1000research.com/articles/13-710/v1	28 Jun 24	{ "type": "Method Article", "title": "Expert Consensus Methods In The Humanities - An Exploration of their Potential", "authors": [ "Charlotte C.S. Rulkens", "Rik Peels", "Lidwine B. Mokkink", "Tamarinde Haven", "Lex Bouter", "Rik Peels", "Lidwine B. Mokkink", "Tamarinde Haven", "Lex Bouter" ], "abstract": "Background Expert consensus methods are regularly used in natural, social, and life sciences. This article explores the potential of applying these methods more frequently in humanities research.\n\nMethods The authors reviewed literature and applied the philosophical methods of conceptual analysis and conceptual engineering.\n\nResults This article identifies and describes six main elements of expert consensus methods. It also provides an overview of the different types of expert consensus methods regularly used in the natural, social, and life sciences: Delphi studies, nominal groups, consensus conferences, and Glaser’s state of the art method. Subsequently, each of these types is illustrated by an example from the sciences. The article also presents the potential of and objections to the application of expert consensus methods there. It gives four examples of expert consensus methods that were applied in humanities research, also presented in line with the six elements.\n\nConclusions The comparisons and categorization show that, as in the natural, social, and life sciences, expert consensus methods in the humanities can in some instances potentially accelerate the epistemic process and enhance transparency, replicability, diversity, and fair processes. Nevertheless, expert consensus methods need to be fine-tuned to do justice to the unique nature and approaches of the humanities and therefore further research is needed.", "keywords": [ "Consensus", "Consensus methods", "Humanities", "Methodology", "Expertise", "Epistemology" ], "content": "1. Introduction\n\nIn this article, we explore the potential of applying structured and formalised expert consensus methods to reach consensus on diverging topics in research in the humanities.1 An important way to make progress in knowledge and understanding is to gradually, via the exchange of evidence, knowledge, expertise, and critical discussion, reach (more) consensus on an issue.2 However, whether there is consensus on an issue at a certain moment in time can sometimes remain unknown, unclear, or implicit. Expert consensus methods can lift off this inconclusiveness by intentionally and formally pursuing or assessing consensus. These methods are regularly used in natural, social, and life sciences. In linguistics, a field that can arguably be grouped under the humanities, formal expert consensus methods are also regularly applied. Other kinds of valuable expert meetings do take place in various disciplines in the humanities, but the expert consensus methods described in this article differ from those by their procedural and formalised character. Therefore, this article explores the potential of applying these formal expert consensus methods more frequently in the humanities.\n\nOur article is structured as follows. After addressing our methodology (§2) we explain what we mean by ‘consensus’ and ‘expert consensus methods’. We do so by distinguishing between six main elements of consensus methods (§3). After that, we provide an overview of expert consensus methods that are regularly used in the natural, social, and life sciences: Delphi studies, nominal groups, consensus conferences, and Glaser’s state of the art method. We also discuss various examples by describing their operationalization of the six elements (§4). We then turn to the humanities and discuss the potential of and the objections to the application of expert consensus methods in the humanities. This is followed by the presentation of four examples of expert consensus methods that have been applied in humanities research, also presented in accordance with the six elements (§5). Subsequently, we discuss opportunities in humanities research for the implementation of expert consensus methods and present our conclusions (§6).\n\n\n2. Methods\n\nTo explore the potential of applying these formal expert consensus methods more frequently in the humanities, we composed an interdisciplinary team of both researchers working with consensus methods in practice, and researchers familiar with research within the domain of the humanities. We subsequently organized a brainstorm session, in which knowledge on the topic was shared from both perspectives. We did an informal literature review and applied the philosophical methods of conceptual analysis (Daly 2010) and conceptual engineering (Cappelen, Gendler, Hawthorne 2016). This resulted in a first draft, followed by a second group session and by a final version of the article, which was edited and reviewed by all authors.\n\n\n3. What is consensus and what is an expert consensus method?\n\nPhilosophers in social epistemology have reflected on the value and function of interaction and consensus as a means to knowledge (Kuhn 1962, Habermas 1996). Furthermore, over the past few decades various computational models are designed to assess the dynamics of epistemic communities leading to consensus or dissensus (Longino 2019, O’Connor et al., 2024). Some researchers defend the importance of consensus as an indicator for academic trustworthiness, and the value of consensus studies to assess pressing issues such as climate change (Oreskes 2021). In this article, we take a more practical approach by presenting means to assess or establish consensus. Before we explore the value of expert consensus methods in the humanities, it is helpful to first define what we mean by ‘consensus’. Ordinary language use of ‘consensus’ might suppose that there is consensus only if a group agrees completely (for 100%) on an issue (see e.g., von der Gracht, 2012). However, also when there is no full consensus and only a large percentage of a group agrees, one can speak of consensus on an issue (Diamond et al. 2014).\n\nClearly, this entails ambiguity, for when is such a group sufficiently large and who are the experts that belong to that group? Moreover, what counts as ‘sufficient’ consensus may differ from discipline to discipline and from issue to issue. Purely deductive disciplines, such as mathematics and logic, may reach full or almost full consensus on some issues and the bar for consensus may, therefore, be high in those fields. Since the questions addressed in the humanities may leave room for multiple viewpoints that are equally warranted by the evidence base, it can be infeasible or not desirable to get a complete or almost complete consensus. The bar for consensus on such issues could be set lower.\n\nConsensus can take the shape of agreeing that something is true or accurate, but also that something is false, or inaccurate, or unreliable. There can even be consensus that we do not know something or that the available evidence does not favour any particular hypothesis. A philosophically informed way to put this is that consensus can take the shape of one out of three doxastic (from the Greek doxa, ‘view’) attitudes: joint belief, joint disbelief, and joint suspension of judgment.\n\nFurthermore, consensus can have different objects: experts can agree on the viability of a model, on the predictive power of a theory, on the adequacy of a hypothesis, on what best explains a phenomenon, on what the most suitable method is to study something, on policies for prevention, on definitions, and much more. And once consensus is reached, it is not guaranteed to remain static. It can change over time with emerging new viewpoints or insights deriving from new research.\n\nWhether or not there is consensus on an issue at a certain moment in time often remains implicit or inconclusive. This especially holds when there is no full agreement or when it is not clear whether there is ‘sufficient’ agreement. Expert consensus methods aim to make explicit whether or not there is consensus and what the degree of consensus is. This can be useful when the issue at hand is an important one and when the evidence about the issue does not speak for itself because evidence is lacking, is inconsistent, or its interpretation is not obvious. What expert consensus methods do in such cases can be defined as follows:\n\nExpert consensus methods are (i) applied by a process leader or steering committee, (ii) to reach consensus or assess the degree of consensus among a group of experts, (iii) about an issue, (iv) based on pre-set rules of engagement, (v) with the aim to deliver useful output, (vi) for future users. They are characterised by constructive and procedural usage of (dis) agreement and the different arguments that are provided therein, (vi) for future users. They are characterised by constructive and procedural usage of (dis) agreement and the different arguments that are provided therein.\n\nLet us briefly clarify the core terms of this definition.\n\ni. A process leader or steering committee identifies an issue and initiates and designs the expert consensus method. They consider which groups have to be represented in the method and then select the experts that are to participate as panellists. The process of how experts are selected and by what criteria should be made fully transparent, because expert consensus methods find part of their credibility and validity in how the process leaders or steering committee determines who counts as an expert. In addition to that, the process leader or steering committee prepares the experts for the overall process and steps of the expert consensus method and provides the experts with rules of engagement. These can be adjusted at the instigation of the experts before starting the method. The process leader or steering committee is also responsible for collecting the data and/or information that is needed to assess the issue at hand and ensures every expert is provided with the same set of information and/or evidence before participating in the method.\n\nii. Experts are the panellists in the consensus procedure that deliver input in the process of measuring and/or reaching consensus. The nature of their expertise can differ, depending on the issue and prospected output of the expert consensus method. They can be experts by training and profession, e.g., a physician making a diagnosis or a historian interpreting a source. But they also can be experts on the basis of their experience, e.g. patients’ knowledge about their own illness or students delivering input on their learning experiences at school.\n\niii. Issues are at the centre of the expert consensus method, they are the problem to be solved, the proposals to be considered, or the question to be answered.\n\niv. Rules of engagement are part of the design of the expert consensus method and specify its preconditions. These preconditions can be, but are not limited to, how high the threshold for consensus is (i.e. what ‘sufficient’ agreement is), which areas of disagreement may be retained, how interaction is structured, when to transition into finalising the output, or what to do when consensus is not reached. Experts may suggest changes and/or improvements to the rules of engagement before engaging as panellists. The rules of engagement should be made fully transparent.\n\nv. Output can be, but is not limited to, answers to closed questions, guidelines, taxonomies, questionnaires, definitions, quality criteria, and policy advice.3\n\nvi. Users are those who have an interest in applying the output of the consensus, including but not limited to the experts involved. They can be researchers (starting follow-up projects), the public (making informed decisions), professionals (working according to a guideline), institutions (determining their mission and ambitions and governments (implementing policies).\n\nTo successfully apply an expert consensus method, careful preparation by the process leader or steering committee and the effective communication between all participating parties about the process is of vital importance. The roles and responsibilities of different actors (the experts versus the process leader or steering committee) need to be laid out and clearly communicated from the start to enable users to verify the quality of the process. The amount of evidence that is available to the experts determines the part of the issue left about which panellists can reach consensus on: i.e. the inclusion of different viewpoints on the issue is one of the most valuable parts of the process, however, the eventual consensus cannot be in contradiction with the available evidence base. This whole process and its outcomes are made transparent and accessible in a publication by the process leader or steering committee. Figure 1 presents the interrelation of the six elements of expert consensus methods (blue boxes) and the fourteen main steps undertaken in the process (arrows). The boxes and steps that are in the orange field are addressed in the publication and its appendices.\n\n\n4. Types of expert consensus methods\n\nIn the natural, social, and life sciences, four distinct expert consensus methods are used frequently: Delphi studies, nominal groups, consensus conferences, and Glaser’s state of the art method. Many studies combine elements of these methods. In Table 1 they are summarised and compared by way of their different characteristics.4 Table 2 shows the actors, factor, and aims of different examples from the natural, social and life sciences. In displaying this, we consider each of the six elements of our earlier definition of ‘expert consensus method’ in paragraph 2.\n\n\n5. Expert consensus methods in the humanities, objections, potential and examples\n\nMany expert meetings take place in the humanities as part of discussions at all kinds of conferences and symposia. They are meant to get the input or feedback of experts or to enhance the discussion among experts and are therefore valuable in themselves. However, these meetings are not purposefully designed to formally reach or assess the level of consensus and therefore are not expert consensus methods as defined above. This more formalised approach to expert meetings seems to be rare in the humanities, but might be a valuable addition because of a number of reasons.\n\nFirst, in formal expert consensus methods, aspects or viewpoints about an issue which normally remain below the surface can be verbalised, shared, and made explicit in the process. It therefore contributes to the articulation of knowledge and understanding of an issue and thus the application of such a method stimulates epistemic progress in itself.\n\nSecond, expert consensus methods enhance the transparency and replicability of humanities research because of their well-documented and systemised approach (Peels and Bouter 2018).5 It can therefore be instrumental to epistemic progress beyond the consensus reached at a moment in time.\n\nThird, expert consensus methods can be helpful in getting a firmer grip on the status quo or state of the art on a particular issue in a field.6 Of course, scholars often already have an idea about that, and such an informed opinion is valuable in itself. However, it can be complemented by a more structured and transparent approach, particularly when the number of scholars involved is large. When applied on a recurrent basis, they might even have the potential to gain insight in how views evolve over time.\n\nFourth, a carefully designed protocol and curated group of experts can foster a fair practice and ensure the consultation of diverse viewpoints. Expert consensus methods provide opportunities to mitigate biases and their ramifications by this cautious selection of experts, but also by blinding certain parts of the process in relation to anonymity of participating experts, for example.\n\nFifth, beneficial effect of applying expert consensus methods, is that application of its outcomes is often easier after the process of such a method is completed.7\n\nSixth, expert consensus methods can help the public perceive what the expert consensus on something is Stekelenburg et al. (2022) and then to make an informed decision on an issue (Ordway 2021).\n\nSeventh and finally, making the degree of consensus or lack thereof among a group of experts explicit by way of an expert consensus method can provide guidance for future research by pointing out the gaps in the current evidence base.\n\nBut there might also be objections. Research in the humanities takes place within paradigms or schools and can consciously or unconsciously, sometimes may even inevitably, be coloured by the backgrounds and positionality of the researchers. In addition to that, humanities research can be concerned with hermeneutics, i.e. the understanding the values and meanings of texts and other objects. And then there are humanities that have a priori methods, such as ethics, epistemology, and metaphysics. These are methods that do not aim at collecting data, but work with intuitions, principles, values, and thought experiments. Because of these various factors, humanities research sometimes allows for multiple viewpoints, nuances, and interpretations that are all to some extent warranted by the evidence base, sometimes even equally warranted. One might think that this evident complexity of humanities research renders expert consensus methods problematic and/or meaningless.\n\nYet one of the important purposes of expert consensus methods is to bring various viewpoints, nuances, and interpretations to the surface, so that they can be discussed. In this way, they can contribute to insights into the level of consensus (and lack thereof) on issues within certain schools or paradigms or groups with comparable backgrounds, or using the same a priori methods, for example. They can furthermore be valuable for gaining insights on consensus on issues between schools or paradigms, groups with different backgrounds, or groups using different a priori methods. Expert consensus methods might even have the potential to increase mutual understanding and provide clarity about what exactly the differences and similarities are. They might be able to make the ‘common ground’ explicit as a result, whilst preventing the domination of certain biases in discussions among experts, for example.\n\nAside from this, more earthly objections might play a role. Expert consensus methods and their subsequent publication require time and resources, which are rarely in abundance in the humanities. In addition to that, the application of expert consensus methods is not interwoven in the humanities research traditions. Therefore, researchers may not be familiar with the expert consensus methods as defined above, and the advantages it may have for their area of research.\n\nTable 3 presents four examples in which expert consensus methods were applied in humanities research. They are summarised in line with the six elements described in paragraph 2. The examples from the Visual Arts and Museology show that expert consensus methods can be tailor-made to the issue at hand, since the method applied here combined different aspects of the methods presented in Table 1. Aside from these examples, one can think of other potential applications. Establishing consensus and guidelines on the best way to preserve certain historical objects, for example, involving eyewitnesses in the reconstruction of an historical event, or consulting various experts in establishing how to interpret an ancient source. Future case studies might be able to concretise when the application of formal expert consensus methods is useful and when not.\n\n\n6. Conclusion\n\nWhen it comes to expert consensus methods, the sciences and the humanities show more similarities than one might initially think. The humanities lend themselves better to structured expert consensus methods than their subject matter and research methods might suggest. It turns out, that when research questions or aims are alike, similar expert consensus methods could be applied in line with the same six elements presented in this article. Expert consensus methods can in particular instances potentially accelerate epistemic process, enhance transparency, increase replicability, stimulate diversity, and encourage fair processes in humanities research and the application of its findings. An in-depth systematic review of the use of expert consensus methods in the humanities was not part of this study, and therefore other forms of formal expert consensus methods in the humanities may have stayed under the radar. Such an overview could be useful for future exploration of the application of expert consensus methods in the humanities. It would be valuable to further explore the possibilities and limitations of these methods and to investigate how expert consensus methods need to be fine-tuned to do justice to the unique nature and approaches of the humanities. Therefore, it should be a priority to carry out more case studies in the humanities in which a consensus method is used and its feasibility and usefulness are evaluated. Humanities researchers could join forces with researchers from other domains who have experience in designing and carrying out these procedures and learn from each other in the process.\n\n\nAuthor contributions\n\nCharlotte C.S. Rulkens and Rik Peels delivered the overall concept and text for this article, which Lidwine B. Mokkink, Tamarinde Haven and Lex Bouter critically read and revised. In addition to that, Tamarinde Haven and Lidwine B. Mokkink were responsible for Tables 1 and 2.", "appendix": "Data availability\n\nData sharing is not applicable to this article as no datasets were generated or analysed during the current study.\n\n\nAcknowledgements\n\nThe authors are grateful to Hans Ket and Michèle Meijer, information specialists at the library of the Vrije Universiteit Amsterdam, for sharing their advice and knowledge in searching for examples of expert consensus methods in both the natural, social, and life sciences and the humanities.\n\n\nReferences\n\nICOM Webinar Defining the Museum in Times of Change: A Way Forward: 2020. Accessed 11 October 2023. Reference Source\n\nStanding Committee for the Museum Definition – ICOM Define Final Report (2020-2022): 2022. Accessed 21 June 2023. Reference Source\n\nBlack N, Murphy M, Lamping D, et al.: Consensus Development Methods: A Review of Best Practice in Creating Clinical Guidelines. J. Health Serv. Res. Policy. 1999; 4(4): 236–248. Publisher Full Text\n\nBlazey P, Crossley KM, Ardern CL, et al.: It Is Time for Consensus on ‘Consensus Statements’. Br. J. Sports Med. 2022; 56(6): 306–307. Publisher Full Text\n\nBrinkman DJ, Tichelaar J, Mokkink LB, et al.: Key Learning Outcomes for Clinical Pharmacology and Therapeutics Education in Europe: A Modified Delphi Study. Clin. Pharmacol. Ther. 2018; 104(2): 317–325. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCurigliano G, Burstein HJ, Winer EP, et al.: De-Escalating and Escalating Treatments for Early-Stage Breast Cancer: The St. Gallen International Expert Consensus Conference on the Primary Therapy of Early Breast Cancer 2017. Ann. Oncol. 2017; 28(8): 1700–1712. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCappelen H, Gendler TS, Hawthorne J: The Oxford Handbook of Philosophical Methodology. Oxford: Oxford University Press; 2016.\n\nDaly C: Introduction to Philosophical Methods. Peterborough: Broadview; 2010.\n\nDelbecq AL, Van de Ven AH, Gustafson DH: Group Techniques for Program Planning: A Guide to Nominal Group and Delphi Processes. Glenview, Illinois: Scott, Foresman and Company; 1975.\n\nDiamond IR, Grant RC, Feldman BM, et al.: Defining Consensus: A Systematic Review Recommends Methodologic Criteria for Reporting of Delphi Studies. J. Clin. Epidemiol. 2014; 67(4): 401–409. PubMed Abstract \| Publisher Full Text\n\nDragouni M, Lekakis S: Co-Creating the Future of Heritage in-the-Making: Empirical Evidence from Community Deliberation at Naxos Island, Greece. Int. J. Herit. Stud. 2023; 29(4): 294–313. Publisher Full Text\n\nGlaser EM: Using Behavioral Science Strategies for Defining the State-of-the-Art. J. Appl. Behav. Sci. 1980; 16(1): 79–92. Publisher Full Text\n\nGoodman CM: The Delphi Technique: A Critique. J. Adv. Nurs. 1987; 12(6): 729–734. 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PubMed Abstract \| Publisher Full Text\n\nLongino H: The Social Dimensions of Scientific Knowledge. The Stanford Encyclopedia of Philosophy. 2019. (Summer 2019 Edition), Edward N. Zalta (ed.). Reference Source\n\nLinstone H, Turoff M: The Delphi Method: Techniques and Applications. Reading: Addison-Wesley Publishing Company; 1975.\n\nMcMillan SS, King M, Tully MP: How to Use the Nominal Group and Delphi Techniques. Int. J. Clin. Pharm. 2016; 38(3): 655–662. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nO’Connor C, Goldberg S, Goldman A: Social Epistemology. The Stanford Encyclopedia of Philosophy. 2024. (Forthcoming, Summer 2024 Edition), Edward N. Zalta & Uri Nodelman (eds.).\n\nOrdway D-M: Covering Scientific Consensus: What to Avoid and How to Get It Right. The Journalist’s Resource. 2021. Online. (2021). Reference Source\n\nOreskes N: Why Trust Science? Princeton, New Jersey: Princeton University Press; 2021.\n\nParaskevas A: Aligning Strategy to Threat: A Baseline Anti-Terrorism Strategy for Hotels. Int. J. Contemp. Hosp. Manag. 2013; 25(1): 140–162. Publisher Full Text\n\nPeels R: Replicability and Replication in the Humanities. Res. Integr. Peer Rev. 2019; 4(1): 2. Publisher Full Text\n\nPeels R: The Many Roads to Progress in Philosophy and Theology. Philosophy in the Age of Science? Inquiries into Philosophical Progress, Method, and Societal Relevance. Hermann J, Hopster J, Kalf W, et al., editors. London: Rowman and Littlefield; 2020; 15–34.\n\nPeels R, Bouter L: The Possibility and Desirability of Replication in the Humanities. Palgrave Commun. 2018; 4(1): 95. Publisher Full Text\n\nPotter M, Gordon S, Hamer P: The Nominal Group Technique: A Useful Consensus Methodology in Physiotherapy Research. N. Z. J. Physiother. 2004; 32(3): 126.\n\nPowell C: The Delphi Technique: Myths and Realities. J. Adv. Nurs. 2003; 41(4): 376–382. PubMed Abstract \| Publisher Full Text\n\nRulkens CCS, Peels R, Bouter L, et al.: Replicating a Rembrandt Study. Preregistration; 2022-2023. Reference Source\n\nStocking B, Jennett B, Spiby J: Criteria for Change: The History and Impact of Consensus Development Conferences in the UK. London: King’s Fund Centre; 1991.\n\nvan Stekelenburg A , Schaap G, Veling H, et al.: Scientific-Consensus Communication About Contested Science: A Preregistered Meta-Analysis. Psychol. Sci. 2022; 33(12): 1989–2008. PubMed Abstract \| Publisher Full Text\n\nvon der Gracht HA : Consensus Measurement in Delphi Studies: Review and Implications for Future Quality Assurance. Technol. Forecast. Soc. Chang. 2012; 79(8): 1525–1536. Publisher Full Text\n\n\nFootnotes\n\n1 The division between the natural sciences, social sciences, life sciences and the humanities can be diffuse, and in many studies, there are various degrees of inter- and multi- disciplinarity between those four groups. For this paper, we count research as part of the humanities when related to one or more of the following disciplines: History; Language and Linguistics; Archeology; Classics; Conservation; History and Philosophy of Science; Literature and Literary Theory; Museology; Music; Philosophy; Theology and Religious studies; Visual Arts and Performing Arts.\n\n2 Thus also Grimm, Peels, and Van Woudenberg (forthcoming); Peels (2020).\n\n3 See, for example, Paraskevas (2013) and Curigliano et al. (2017) mentioned in Table 1.\n\n4 Table 1 synthesises criteria as previously discussed in Jones and Hunter (1995), Black et al. (1999), Blazey et al. (2021).\n\n5 Both desiderata lately received much attention. For replicability, see Peels (2019).\n\n6 See, for example, Hodgkin et al. (1975) mentioned in Table 2.\n\n7 One example where this was carried out as part of an issue in the humanities, is the procedure that was successfully completed in 2022 in order to establish a new international museum definition, mentioned in Table 3. An 18-month procedure with hundreds of museum professionals from 126 National Committees from all over the world led to the adoption of a newly proposed definition approved by 92% of the votes. See Standing Committee for the Museum Definition – ICOM Define Final Report (2020-2022) (2022) and ICOM Webinar Defining the Museum in Times of Change: A Way Forward (2020)." }	[ { "id": "296789", "date": "23 Aug 2024", "name": "Emanuela Reale", "expertise": [ "Reviewer Expertise Higher education and research policy" ], "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\nExpert Consensus Methods In The Humanities – An Exploration of their Potential\nThe paper wants to explore the value of applying expert consensus methods in the humanities. The topic is very interesting and deserve attention. The manuscript is clearly written, and the literature mobilized is appropriate and relevant for the aim of the investigation proposed. The effort of synthetize concepts and findings using tables is valuable for the clarity of the argument. However, the manuscript in its current form has several weaknesses and, in my opinion, needs some substantial re-work. Hereby I try to summarize the most important critical issues that must be solved.\nFirst, the aim of the paper is clear, but the research question(s) related to that aim are missing. This is an important limitation, since the manuscript reports an exploratory effort and therefore a delimitation of the research problem that you want to investigate is very useful to delineate the perimeter of the work.\nSecond, the section on Method is poor. The composition of the interdisciplinary team (and the reasons for using this team) is not clearly described, the brainstorm session is not properly defined and illustrated, the ‘informal’ literature review is something new for me (what ‘informal’ means?) and it is not acceptable for a research paper to quote two methods used (‘philosophical methods of conceptual analysis’ and ‘conceptual engineering’) without explaining how they have been used for the purposes of the paper but just putting two references. Also, the process of writing and reviewing the manuscript should be better described and justified.\nThird, and more important, the paper refers always to ‘humanities.’ Humanities is a highly fragmented and differentiated field, where many and different epistemic communities work, interact, shape their own idea and uses of consensus method (if any) and where different instances of relationships with other researchers within the field and with neighboring fields are at place. The paper does not acknowledge this important characteristic of humanities (see footnote 1 where there is a list of disciplines included in the humanities that are considered in the paper). The same holds true for STEMs, which are considered as they are all adopting regularly expert consensus methods and there is no explanation or evidence about it. In sum, the paper ‘explores’ the expert consensus method without considering the set of ideas, values, theories, concepts and practices that pertain to the disciplines analyzed and their scientific work.\nFourth, sections 3 and 4 provide a clear summary of what expert consensus methods are and a deepen analysis of four types. Then the reader expects Section 5 discusses the results with respect to the exploration done. On the contrary, Section 5 lists of pros and cons related to the expert consensus method, and some examples of applications; the contents are completely unrelated with what the sections 3 and 4 have presented and the motivations for considering the expert consensus method an interesting development for humanities are unclear. Here the suggestion is to revise the sections trying to answer the research questions (when formulated) and highlight what we have learnt from the exploratory work, what is the contribution of the brainstorming to the exploration, what new avenues of research can be envisaged based on the results obtained, and what limitations/obstacles can the expert consensus model have in the humanities with respect to the other fields.\n\nIs the rationale for developing the new method (or application) clearly explained? No\n\nIs the description of the method technically sound? No\n\nAre sufficient details provided to allow replication of the method development and its use by others? No\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": "12422", "date": "16 Dec 2024", "name": "Charlotte Rulkens", "role": "Author Response", "response": "We sincerely thank the reviewer for her valuable comments, all of which we will address in the revision of our paper. We plan to begin this revision as soon as we receive the second review. We agree with the first and second suggestions to clarify our research questions, as well as the opportunities mentioned for improving the method section. While our method section may differ from those typically found in research papers, it is important to note that this paper is intended as a perspective piece with a didactic focus. It is aimed at highlighting to humanities scholars what is possible in terms of a systematic approach to academic consensus on a topic and to initiate exploration of and discussion about the possible value of applying such an approach. Regarding the third comment, we will ensure that we address the issue of differentiation within research fields and discuss its implications for the application of consensus methods. Concerning the fourth point, we will work to better integrate section 5 with the preceding sections to create a more cohesive flow throughout the paper." }, { "c_id": "12797", "date": "16 Dec 2024", "name": "Charlotte Rulkens", "role": "Author Response", "response": "Dear reviewers, We sincerely thank you for the thorough reviews of our manuscript titled Expert Consensus Methods In The Humanities - An Exploration of their Potential. We appreciate the time and effort you have put into evaluating our work and for providing insightful comments and valuable suggestions. We have carefully considered each of them and have revised the manuscript accordingly. Below, we provide detailed responses to each of the comments. We believe that the revisions have improved the manuscript, and we hope that our responses meet any remaining concerns. Response to reviewer 1: Emanuela Reale, CNR IRCRES—Research Institute for Sustainable Economic Growth, Rome, Italy Reviewer 1, Comment 1: First, the aim of the paper is clear, but the research question(s) related to that aim are missing. This is an important limitation, since the manuscript reports an exploratory effort and therefore a delimitation of the research problem that you want to investigate is very useful to delineate the perimeter of the work. Response: We acknowledge the concern raised and have addressed this by rewriting the introduction section and by incorporating a clearer description of our study goal. We now make it more clear that our goal was primarily didactic, in the sense that we sought to explore the usefulness of consensus methods utilised in the natural, social, and life sciences for the humanities (see section 1, lines 59-83): An important way to make progress in knowledge and understanding is to gradually, via the exchange of evidence, expertise, and critical discussion, reach (more) consensus on a topic. However, whether there is consensus on a certain matter at a moment in time can sometimes remain unknown, be unclear, or remain implicit. Expert consensus methods can lift off this inconclusiveness by intentionally and formally pursuing or assessing consensus. These methods are regularly used in natural, social, and life sciences. Despite the significant role of consensus in the development of (academic) knowledge and understanding (e.g. Kuhn, 1962; Habermas, 1996; thus also Peels, 2020; Grimm et al., forthcoming), formal expert consensus methods have hardly been explored within in the context of the humanities.[1] Other kinds of valuable expert meetings do take place in various disciplines in the humanities. They usually do not seek to reach a consensus, but to explore a topic in-depth by bringing the relevant experts to the table. Moreover, expert consensus methods described in this article differ from the expert meetings that already take place in the humanities by the former’s procedural and formalised character. In this paper, we aim to highlight to scholars, particularly those in the humanities, what is possible in terms of a systematic approach to academic consensus. In doing so, we aim to initiate exploration of and discussion about the potential of applying such an approach. What would be the added value of applying expert consensus meetings to humanities-related issues? To achieve this, we first explicate our method (§2), before we provide a framework for the application of expert consensus methods by introducing six key elements (§3). Subsequently, we present common types of expert consensus methods utilized in the natural, social, and life sciences. We highlight their and proven value across various types of research through relevant examples and their respective objectives (§4). We then consider both the potential benefits and possible limitations to applying these methods within humanistic research (§5), and conclude with suggestions for future research (§6). Reviewer 1, Comment 2: Second, the section on Method is poor. The composition of the interdisciplinary team (and the reasons for using this team) is not clearly described, the brainstorm session is not properly defined and illustrated, the ‘informal’ literature review is something new for me (what ‘informal’ means?) and it is not acceptable for a research paper to quote two methods used (‘philosophical methods of conceptual analysis’ and ‘conceptual engineering’) without explaining how they have been used for the purposes of the paper but just putting two references. Also, the process of writing and reviewing the manuscript should be better described and justified. Response: Our article, as we now emphasise more, is meant as an exploratory comment and a didactic contribution to inspire scholars in the humanities to see whether consensus methods may be valuable within their own discipline or sub discipline. It is not an instance of systematic empirical research, which would have required a much more detailed methodological section. But we certainly we agree that the method section can be improved in various ways. Therefore, we extended the section by including a specification of the backgrounds of the involved authors and specified the process of our collaboration as follows (see section 2, lines 85-94): To explore the potential of applying expert consensus methods in the humanities, we composed an interdisciplinary team of both researchers from the sciences using consensus methods in practice (LM: epidemiology; TH: qualitative research methodology; LB: epidemiology and data science, research integrity), and researchers familiar with the domain of the humanities and epistemology (CR: art history, museology; RP: epistemology and philosophy of science, religion and theology). In a first brainstorm session, knowledge on the topic was shared from both perspectives. This resulted in a list of consensus methods and examples used in the sciences, drafted by LM and TH. CR carried out a literature search in order to find examples thereof that were applied in her domain. A first draft was written and discussed in a second group session and culminated in a final version, which was edited and reviewed by all authors. Reviewer 1, Comment 3: Third, and more important, the paper refers always to ‘humanities.’ Humanities is a highly fragmented and differentiated field, where many and different epistemic communities work, interact, shape their own idea and uses of consensus method (if any) and where different instances of relationships with other researchers within the field and with neighboring fields are at place. The paper does not acknowledge this important characteristic of humanities (see footnote 1 where there is a list of disciplines included in the humanities that are considered in the paper). The same holds true for STEMs, which are considered as they are all adopting regularly expert consensus methods and there is no explanation or evidence about it. In sum, the paper ‘explores’ the expert consensus method without considering the set of ideas, values, theories, concepts and practices that pertain to the disciplines analyzed and their scientific work. Response: We acknowledge the concern raised and have taken this into account by adjusting a footnote 1 in section 1 (see above) and by revising section 4. Upon reflection we furthermore adjusted the ways to delineate the contexts in which expert consensus methods may be valuable, by focusing on one’s purpose in applying such a method (see section 4, lines 213-235): The subsequent question would be to determine the contexts in which this approach to consensus would prove most valuable. Research in both the humanities and the sciences spans a wide array of disciplines and sub disciplines, with a vast range of variation in types of research and methods, even within the most specialised areas. Classification of research into one of these domains is often gradual in nature, especially since interdisciplinary research increasingly combines approaches traditionally linked to the ‘humanities’ or ‘sciences’. Making general statements on the types of research in which expert consensus methods might be valuable related to the ‘humanities’ versus the ‘sciences’ is therefore difficult. We decided to examine objectives for employing consensus methods in various contexts in which they have demonstrated their value. This could contribute to a directory of instances in which these methods may prove useful in humanities-related fields. Gattrell et al. (2024, Table 2) present examples of these objectives in context of the health sciences, such as the establishment of clinical practice guidelines, diagnostic guidelines or reporting guidelines but also the classification of diseases, the setting research priorities, or the formulation of policy. These aims are summarised, adjusted and extended in Table 4 in a manner that enhances their generalisability to other fields of research and were included in Table 2 and 3. Although these examples highlight specific applications, we think that there is a broader potential for its use in the humanities. For instance, in establishing consensus and guidelines on the best way to preserve certain historical objects (guidelines/ policy development), involving eyewitnesses in the reconstruction of an historical event (current state of knowledge), or consulting various experts in establishing how to interpret an ancient source (current state of knowledge). Future research and case studies exploring the potential applications outlined in Table 4 could provide insights into specific instances where the pursuit of formal dis- or consensus may prove effective. Reviewer 1, Comment 4: Fourth, sections 3 and 4 provide a clear summary of what expert consensus methods are and a deepen analysis of four types. Then the reader expects Section 5 discusses the results with respect to the exploration done. On the contrary, Section 5 lists of pros and cons related to the expert consensus method, and some examples of applications; the contents are completely unrelated with what the sections 3 and 4 have presented and the motivations for considering the expert consensus method an interesting development for humanities are unclear. Here the suggestion is to revise the sections trying to answer the research questions (when formulated) and highlight what we have learnt from the exploratory work, what is the contribution of the brainstorming to the exploration, what new avenues of research can be envisaged based on the results obtained, and what limitations/obstacles can the expert consensus model have in the humanities with respect to the other fields. Response: Thank you for pointing this out to us. Based on this valuable feedback, we have worked to better align section 5 with the preceding sections to create a more cohesive flow throughout the paper and have made more explicit what our line of reasoning is. However, because the paper is intended as preliminary exploration, part of our manuscript aims to identify the potential value of these methods for the humanities. Section 5 is dedicated to that purpose and we made that more explicit in the revision (see section 5, lines 253-315): In the preceding sections, we examined the elements and dynamics of expert consensus methods, along with examples and possibilities for intended objectives. In this section, we delve into the question why formal approaches to consensus in the humanities remain scarce, despite the significant factor of consensus and dissensus in knowledge production across research cultures. Aside from limitations, we also address what the potential added value of applying consensus methods to humanities-related issues could be. First of all, practical limitations likely play a role. Expert consensus methods and their subsequent publication demand significant time and resources, which are rarely in abundance in the humanities. Additionally, a factor that may affect academia broadly is the emphasis on publication quantity and individual contributions, which can discourage scholars from lengthy endeavours for which recognition is shared (Leising et al., 2024). Moreover, expert consensus methods are not interwoven in humanities research traditions. Researchers may not be familiar with consensus methods, and the advantages they may have for their area of research. Secondly, concerns raised in the field of psychology that may apply in the humanities as well, highlight risks of consensus-building. Emphasising consensus may lead to agreement for the wrong reasons, such as conformity, which may be harmful to innovation, creativity and minority views. It can sideline dissent, whilst that is a vital element in the evolution of science (Gollwitzer, 2022; Asendorpf & Gebauer, 2022; Beck et al., 2022; Denissen & Sijtsma, 2022; Hilbig et al., 2022). Additionally, experts agreeing on something does not necessarily mean they are correct. They may be in full agreement on a point that is incorrect or untrue. Scholars may also disagree due to the role of research paradigms or schools within the humanities, as researchers’ positionality can shape their interpretations. In addition to that, humanities research can be concerned with hermeneutics, i.e., issues that have to do with understanding the values and meanings of texts and other objects. And many humanities fields such as ethics, epistemology, and metaphysics, rely on apriori methods like thought experiments, reasoning on the basis of intuitions and principles, and formal logic rather than empirical data. This allows for multiple interpretations that can all to some extent be warranted by the evidence base, and may not readily align into a singular consensus. These factors suggest that the complexity and interpretative nature of humanities research might render expert consensus methods to some extent less relevant. Nevertheless, we believe that there are compelling reasons for considering expert consensus methods in humanities research. First of all, opposing the above concerns, one of the important purposes of expert consensus methods is to bring various viewpoints, nuances, and interpretations to the surface, so that they can be discussed. Depending on the design and purpose of the consensus method, it could reveal levels of consensus and/or dissensus on an issue, within or across schools of thought or paradigms. They might even have the potential to articulate both divergences and common grounds regarding an issue, and by extension increase mutual understanding. The application of such a method may therefore stimulate epistemic progress in itself. Moreover, a carefully designed protocol and curated group of experts can foster a fair practice and ensure the consultation of diverse viewpoints. Expert consensus methods provide opportunities to mitigate biases and their ramifications by this cautious selection of experts, but also by blinding certain parts of the process in relation to anonymity of participating experts, for example. Secondly, expert consensus methods can be helpful in getting a firmer grip on the status quo or state of the art on a particular issue in a field (e.g. Hodgkin et al. (1975) mentioned in Table 2; Rulkens et al. 2022-2023, mentioned in Table 3). Of course, scholars often already have an idea about that, and such an informed opinion is valuable in itself. However, it can be complemented by a more structured approach, particularly when the number of scholars involved is large. This approach can additionally provide guidance for future research by pointing out the gaps in the current evidence base. And when applied on a recurrent basis, it might even have the potential to gain insight in how views evolve over time. It is important to note that consensus is not a final endpoint; rather, it can be continually re-evaluated in light of new insights and reasonable doubts regarding its validity. Third, an advantage of expert consensus methods is that they enhance the transparency and replicability of research because of their well-documented and systematised approach (for replication in the humanities, see Peels and Bouter (2018)). It can therefore be instrumental to epistemic progress beyond the consensus reached at a moment in time. Fourth, application or implementation of its outcomes is often easier after the process of a consensus method is completed (e.g. Standing Committee for the Museum Definition (2020-2022) (2022), mentioned in Table 3). And finally, expert consensus methods can aid public understanding by clarifying experts perspectives on specific issues (Stekelenburg et al., 2022) thereby enabling the public to make informed decisions (Ordway, 2021). In response to this comments, we furthermore extended suggestions for future research in our conclusion (see section 5, lines 325-335): However, applying a consensus method is a costly endeavour in terms of time and means. There might be conditions under which these efforts do not live up to the epistemic benefits they provide for. The identification of the particular areas and issues where these methods hold most promise can be done by assessing the purposes these methods have served, and translating those to a specific area of research. The assessment and prioritisation of issues, especially when resources are limited, should be carried out locally by researchers who are familiar with the epistemologies and issues relevant to their field. Consequently, it is essential to carry out future research involving humanities case studies that employ expert consensus methods, to evaluate their feasibility and effectiveness. Collaboration between humanities researchers with scholars from other domains experienced in designing consensus methods could facilitate mutual learning and foster innovative approaches. Response to reviewer 2: Daniel Leising, Technische Universitat Dresden, Dresden, Saxony, Germany Reviewer 2, Comment 1: First, I would argue that the Humanities are actually not that “unique” in terms of their nature and approaches (page 11). To enable a proper discussion of this issue, the authors could explain their own view of what is so specific about the Humanities in a bit more detail. Here is my view: Interviews and text analyses are used in many different branches of science. Exploratory research is also conducted in many fields. The ways in which people give meaning to their experiences are also of interest across various branches of science. And even in many of the “harder” sciences, research is often “coloured by the backgrounds and positionality of the researchers” (page 8). So, much of what is common in research in the Humanities is actually shared with other fields. In my view, two features may actually distinguish “the Humanities” from other fields - but even these distinctions are only gradual in nature. One feature is the common practice of using everyday language as the primary scientific language. The other feature is the relative fuzziness of many concepts - which I would note is not necessarily a weakness but may often be unavoidable given the complexity of the phenomena of interest (and also given the first feature just mentioned). Do the authors agree? Response: Thank you for challenging us on this point. We agree that the borders between the general fields of the natural, social, and life sciences, and the humanities are fuzzy, when it comes to their objects, purposes, and methods. Whether there is something distinctive about the humanities – we believe that is compatible with fuzziness at the boarders – is a matter that we cannot possibly explore in this article (one of us does so in a book entitled A Philosophy of the Humanities that is forthcoming in 2025 with Oxford University Press, a volume co-authored by Stephen Grimm, Rik Peels, and René van Woudenberg). We do believe though that ordinary language use is not generally characteristic of the humanities (think of the highly technical language of metaphysics and epistemology), nor do we believe that insufficient clarity about core concepts is characteristic of the humanities. We agree though with the idea, also drawn to attention to by reviewer 1 in comment 3, that the humanities are a highly diversified field. We included that by adjusting footnote 1 in section 1: We acknowledge that as is the case in the sciences, the humanities encompass a large amount of disciplines and sub fields, exhibiting significant diversity in research types and methodologies. However, in this paper we refer to ‘the humanities’ to clarify difference for the reader, rather than out of belief in rigid categorisations. For readers interested in this topic, see e.g. Woudenberg (2018) and Grimm et al., (forthcoming). Additionally, to be able to demarcate between contexts in which the formal approach might be beneficial, we decided to put an emphasis on the purposes for which these methods are employed in other domains. This would be a way in which researchers with expertise in their fields could identify opportunities for a formal approach to consensus (see section 4, lines 213-235, also quoted above in response to comment 3 of reviewer 1): The subsequent question would be to determine the contexts in which this approach to consensus would prove most valuable. Research in both the humanities and the sciences spans a wide array of disciplines and sub disciplines, with a vast range of variation in types of research and methods, even within the most specialised areas. Classification of research into one of these domains is often gradual in nature, especially since interdisciplinary research increasingly combines approaches traditionally linked to the ‘humanities’ or ‘sciences’. Making general statements on the types of research in which expert consensus methods might be valuable related to the ‘humanities’ versus the ‘sciences’ is therefore difficult. We decided to examine objectives for employing consensus methods in various contexts in which they have demonstrated their value. This could contribute to a directory of instances in which these methods may prove useful in humanities-related fields. Gattrell et al. (2024, Table 2) present examples of these objectives in context of the health sciences, such as the establishment of clinical practice guidelines, diagnostic guidelines or reporting guidelines but also the classification of diseases, the setting research priorities, or the formulation of policy. These aims are summarised, adjusted and extended in Table 4 in a manner that enhances their generalisability to other fields of research and were included in Table 2 and 3. Although these examples highlight specific applications, we think that there is a broader potential for its use in the humanities. For instance, in establishing consensus and guidelines on the best way to preserve certain historical objects (guidelines/ policy development), involving eyewitnesses in the reconstruction of an historical event (current state of knowledge), or consulting various experts in establishing how to interpret an ancient source (current state of knowledge). Future research and case studies exploring the potential applications outlined in Table 4 could provide insights into specific instances where the pursuit of formal dis- or consensus may prove effective. Reviewer 2, Comment 2: Second, I would like to encourage the authors to say a bit more about possible reasons for why so little consensus building is taking place in many fields of science, despite it being widely viewed as desirable. In my view, there are a couple of reasons, most of which are rather extraneous to an idealised image of science (as a collaborative and cumulative effort at building a reliable knowledge base for the good of mankind). For example, consensus building is likely to reduce the publication output of a field, which would be at odds with the interests of commercial publishers. Also, individual researchers have an interest in “making a name for themselves” based on their specific expertise or views regarding certain topics. Consensus building makes it more difficult to maintain a highly distinct profile and thus reduce visibility. This may be risky in a science system that is as competitive as the one we choose to entertain. And then there may be a need to “save face”: The more of your career you have dedicated to certain ideas (e.g., your own theory of some phenomenon), the harder it may be to let those ideas go - even if the evidence against them is overwhelming. There is a reason why Max Planck once stated that scientific progress happens “one funeral at a time”. At least a brief nod to the relevance of such more political factors would be nice - if the authors share my conviction that they are relevant. Who has an actual interest in achieving consensus? Who has an interest in avoiding the emergence of consensus? And what are the external incentives for and against this type of work? Response: We think that these are valid points and the reviewers’ publications on the topic shed important light on this. We agree that external incentives may work against a formal approach to consensus as follows and have therefore slightly revised the manuscript on this point. We remain rather brief, though, as the exact explanation of why consensus methods are still relatively rarely used even in the sciences is an issue that is beyond the scope of our paper (see section 5, lines 259-264, also quoted in response to comment 4 of reviewer 1): Expert consensus methods and their subsequent publication demand significant time and resources, which are rarely in abundance in the humanities. Additionally, a factor that may affect academia broadly is the emphasis on publication quantity and individual contributions, which can discourage scholars from lengthy endeavours for which recognition is shared (Leising et al., 2024). Moreover, expert consensus methods are not interwoven in humanities research traditions. Researchers may not be familiar with consensus methods, and the advantages they may have for their area of research. Reviewer 2, Comment 3: Third, I would also appreciate if the authors could talk some more about “bad consensus” – views that are held by many, despite their being wrong. With a particular eye to the Humanities – what criteria are there, apart from consensus, that may help us determine whether a view is “correct”? Or may truth in the Humanities only be defined in terms of relative consensus among researchers? In the harder sciences, the ability to predict the occurrence of certain events from a theory (and some data input) plays a crucial role as evidence of the theory's correctness. Are the Humanities able to produce testable predictions, too? Can there be consensus projects in the Humanities that actually test one theory against another? Response: This is an interesting point, that would need a elaborative reflection in order to do justice to the complexity of this issue. It is important to note, that we do believe that consensus and truth are two different things, and we made this more clear in our revised paper. Based these suggestions we also incorporated that consensus can be reached based on the wrong reasons and (see section 5, lines 265-271, also quoted in response to comment 4 of reviewer 1): Secondly, concerns raised in the field of psychology that may apply in the humanities as well, highlight risks of consensus-building. Emphasising consensus may lead to agreement for the wrong reasons, such as conformity, which may be harmful to innovation, creativity and minority views. It can sideline dissent, whilst that is a vital element in the evolution of science (Gollwitzer, 2022; Asendorpf & Gebauer, 2022; Beck et al., 2022; Denissen & Sijtsma, 2022; Hilbig et al., 2022). Additionally, experts agreeing on something does not necessarily mean they are correct. They may be in full agreement on a point that is incorrect or untrue. Reviewer 2, Comment 4: Fourth, on page 3 the authors state that “Since the questions addressed in the humanities may leave room for multiple viewpoints that are equally warranted by the evidence base, it can be infeasible or not desirable to get a complete or almost complete consensus.” To this I would respond: what are the conditions under which working toward consensus would not be feasible, or desirable? I agree that this may be the case – but when is it the case, and when is it not the case? I personally hold the somewhat sceptical view that many researchers will shy away from working toward consensus even if doing so would be both feasible and desirable (see my previous point). Such a “live and let live” attitude may often be relatively comfortable to have (e.g., to avoid conflict). It may, however, be regarded as unethical, because it will foster the preservation of an unnecessary level of ambiguity. Response: We appreciate this insightful suggestion. Our approach to expert consensus methods does not necessarily imply working towards consensus in all instances, but is also related to the assessment of the (lack of) consensus. We view measuring consensus as a means, not an objective in itself. Furthermore, as put forward in paragraph 3, the bar for ‘sufficient’ consensus is something consensus researchers can determine their selves. Applying a consensus methods is a costly endeavour in terms of time and means. There might be conditions under which the effort of applying such a method does not live up to the epistemic benefits it provides for. We furthermore believe that the feasibility and desirability of assessing consensus is an issue that merits further exploration and included this as follows (see section 5, lines 325-335): However, applying a consensus method is a costly endeavour in terms of time and means. There might be conditions under which these efforts do not live up to the epistemic benefits they provide for. The identification of the particular areas and issues where these methods hold most promise can be done by assessing the purposes these methods have served, and translating those to a specific area of research. The assessment and prioritisation of issues, especially when resources are limited, should be carried out locally by researchers who are familiar with the epistemologies and issues relevant to their field. Consequently, it is essential to carry out future research involving humanities case studies that employ expert consensus methods, to evaluate their feasibility and effectiveness. Collaboration between humanities researchers with scholars from other domains experienced in designing consensus methods could facilitate mutual learning and foster innovative approaches. Minor comments: Reviewer 2, Comment 5: Fifth, the authors state that a steering committee would typically be comprised of the people who initiated the process and who will ultimately author the consensus document. Could there not be cases in which the process should better be overseen by an independent third party (e.g., a moderator)? Especially when issues are highly contentious, for example? And should not all the experts who worked on the consensus be authors on the consensus document? Response: Thank you for this relevant point, this is certainly the case and we made this more clear in the revised manuscript by the following addition (see section 3, lines 193-198): Naturally, the six elements can be specified at by the steering committee. If they deem it valuable to include an independent moderator or to incorporate a member with a critical or divergent perspective—especially if the topic is highly controversial—this remains within their discretion. Additionally, it is up to them to decide who will be listed as co-authors, whether that includes only the steering committee, all participants, or a broader group representation such as the panellists as a whole, for instance. Reviewer 2, Comment 6: Sixth, something is not quite right on page 4, text lines 16-19. Response: Thank you for noticing, we have corrected this sentence as follows (see section 3, lines 136-140): Expert consensus methods are (i) applied by a process leader or steering committee, (ii) to reach consensus or assess the degree of consensus among a group of experts, (iii) about an issue, (iv) based on preset rules of engagement, (v) with the aim to deliver useful output, (vi) for future users. They are characterised by constructive and procedural usage of (dis)agreement and the different arguments that are provided therein. Reviewer 2, Comment 7: Seventh, I would say that evidence will never “speak for itself” (page 4) - otherwise papers in certain scientific fields would not need any Discussion sections. Response: Thank you for rightly pointing this out to us. We adjusted this as follows (see section 3, lines 132-134): This can be useful when the issue at hand is an important one and when evidence is lacking, it is inconsistent, or its interpretation is not obvious. As a result of our reactions to the comments, we also made adjustments to our abstract and conclusion, which are not all quoted in the above letter. We hope these revisions and clarifications meet the reviewers’ expectations. Please let us know if any further revisions are needed. Sincerely, Charlotte Rulkens MA Prof. dr. Rik Peels Dr. Lidwine Mokkink Dr. Tamarinde Haven Prof. em. dr. Lex Bouter [1] We acknowledge that as is the case in the sciences, the humanities encompass a large amount of disciplines and sub fields, exhibiting significant diversity in research types and methodologies. However, in this paper we refer to ‘the humanities’ to clarify difference for the reader, rather than out of belief in rigid categorisations. For readers interested in this topic, see e.g. Woudenberg (2018) and Grimm et al., (forthcoming)." } ] }, { "id": "309049", "date": "19 Sep 2024", "name": "Daniel Leising", "expertise": [ "Reviewer Expertise Personality / Social Psychology", "Meta-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\nI strongly agree that more systematic consensus-building is needed in many fields of science, including the Humanities and Psychology (my own field) and I find little that I would disagree with in the manuscript as is.\nHowever, I also think that the authors could do more than just presenting an overview of established consensus building methods and suggesting that researchers from the Humanities should give these methods a try. The issue is more complex than that. In order to improve on the depth, utility and potential impact of this paper, I would thus like to encourage the authors to think about and – if possible – share their views on a number of additional, intriguing, and sometimes thorny issues regarding consensus building.\n\nFirst, I would argue that the Humanities are actually not that “unique” in terms of their nature and approaches (page 11). To enable a proper discussion of this issue, the authors could explain their own view of what is so specific about the Humanities in a bit more detail.\nHere is my view: Interviews and text analyses are used in many different branches of science. Exploratory research is also conducted in many fields. The ways in which people give meaning to their experiences are also of interest across various branches of science. And even in many of the “harder” sciences, research is often “coloured by the backgrounds and positionality of the researchers” (page 8). So, much of what is common in research in the Humanities is actually shared with other fields.\n\nIn my view, two features may actually distinguish “the Humanities” from other fields - but even these distinctions are only gradual in nature. One feature is the common practice of using everyday language as the primary scientific language. The other feature is the relative fuzziness of many concepts - which I would note is not necessarily a weakness but may often be unavoidable given the complexity of the phenomena of interest (and also given the first feature just mentioned). Do the authors agree?\n\nSecond, I would like to encourage the authors to say a bit more about possible reasons for why so little consensus building is taking place in many fields of science, despite it being widely viewed as desirable. In my view, there are a couple of reasons, most of which are rather extraneous to an idealized image of science (as a collaborative and cumulative effort at building a reliable knowledge base for the good of mankind).\n\nFor example, consensus building is likely to reduce the publication output of a field, which would be at odds with the interests of commercial publishers. Also, individual researchers have an interest in “making a name for themselves\" based on their specific expertise or views regarding certain topics. Consensus building makes it more difficult to maintain a highly distinct profile and thus reduce visibility. This may be risky in a science system that is as competitive as the one we choose to entertain.\n\nAnd then there may be a need to “save face”: The more of your career you have dedicated to certain ideas (e.g., your own theory of some phenomenon), the harder it may be to let those ideas go - even if the evidence against them is overwhelming. There is a reason why Max Planck once stated that scientific progress happens “one funeral at a time”.\nAt least a brief nod to the relevance of such more political factors would be nice - if the authors share my conviction that they are relevant. Who has an actual interest in achieving consensus? Who has an interest in avoiding the emergence of consensus? And what are the external incentives for and agaisnt this type of work?\n\nThird, I would also appreciate if the authors could talk some more about “bad consensus” – views that are held by many, despite their being wrong. With a particular eye to the Humanities – what criteria are there, apart from consensus, that may help us determine whether a view is “correct”? Or may truth in the Humanities only be defined in terms of relative consensus among researchers? In the harder sciences, the ability to predict the occurrence of certain events from a theory (and some data input) plays a crucial role as evidence of the theory's correctness. Are the Humanities able to produce testable predictions, too? Can there be consensus projects in the Humanities that actually test one theory against another?\n\nFourth, on page 3 the authors state that “Since the questions addressed in the humanities may leave room for multiple viewpoints that are equally warranted by the evidence base, it can be infeasible or not desirable to get a complete or almost complete consensus.” To this I would respond: what are the conditions under which working toward consensus would not be feasible, or desirable? I agree that this may be the case – but when is it the case, and when is it not the case? I personally hold the somewhat skeptical view that many researchers will shy away from working toward consensus even if doing so would be both feasible and desirable (see my previous point). Such a “live and let live” attitude may often be relatively comfortable to have (e.g., to avoid conflict). It may, however, be regarded as unethical, because it will foster the preservation of an unnecessary level of ambiguity.\n\nMinor points\nFifth, the authors state that a steering committee would typically be comprised of the people who initiated the process and who will ultimately author the consensus document. Could there not be cases in which the process should better be overseen by an independent third party (e.g., a moderator)? Especially when issues are highly contentious, for example? And should not all the experts who worked on the consensus be authors on the consensus document?\n\nSixth, something is not quite right on page 4, text lines 16-19.\n\nSeventh, I would say that evidence will never “speak for itself” (page 4) - otherwise papers in certain scientific fields would not need any Discussion sections.\nTogether with a variety of colleagues, I have written about several of the above-mentioned issues. If the authors are interested in this, here are a few key references: I referenced some of our own publications that I consider to be related in terms of content, to let the authors know about their existence. The authors are of course entirely free to decide whether to look into them, or not.\n\nLeising, D.et.al; 2023 (Ref 1)\nLeising, D.et.al; 2024 (Ref 2)\nLeising, D.et.al; 2022 (Ref 3)\nLeising, D.et.al; 2022 (Ref 4)\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": [ { "c_id": "12798", "date": "16 Dec 2024", "name": "Charlotte Rulkens", "role": "Author Response", "response": "Dear reviewers, We sincerely thank you for the thorough reviews of our manuscript titled Expert Consensus Methods In The Humanities - An Exploration of their Potential. We appreciate the time and effort you have put into evaluating our work and for providing insightful comments and valuable suggestions. We have carefully considered each of them and have revised the manuscript accordingly. Below, we provide detailed responses to each of the comments. We believe that the revisions have improved the manuscript, and we hope that our responses meet any remaining concerns. Response to reviewer 1: Emanuela Reale, CNR IRCRES—Research Institute for Sustainable Economic Growth, Rome, Italy Reviewer 1, Comment 1: First, the aim of the paper is clear, but the research question(s) related to that aim are missing. This is an important limitation, since the manuscript reports an exploratory effort and therefore a delimitation of the research problem that you want to investigate is very useful to delineate the perimeter of the work. Response: We acknowledge the concern raised and have addressed this by rewriting the introduction section and by incorporating a clearer description of our study goal. We now make it more clear that our goal was primarily didactic, in the sense that we sought to explore the usefulness of consensus methods utilised in the natural, social, and life sciences for the humanities (see section 1, lines 59-83): An important way to make progress in knowledge and understanding is to gradually, via the exchange of evidence, expertise, and critical discussion, reach (more) consensus on a topic. However, whether there is consensus on a certain matter at a moment in time can sometimes remain unknown, be unclear, or remain implicit. Expert consensus methods can lift off this inconclusiveness by intentionally and formally pursuing or assessing consensus. These methods are regularly used in natural, social, and life sciences. Despite the significant role of consensus in the development of (academic) knowledge and understanding (e.g. Kuhn, 1962; Habermas, 1996; thus also Peels, 2020; Grimm et al., forthcoming), formal expert consensus methods have hardly been explored within in the context of the humanities.[1] Other kinds of valuable expert meetings do take place in various disciplines in the humanities. They usually do not seek to reach a consensus, but to explore a topic in-depth by bringing the relevant experts to the table. Moreover, expert consensus methods described in this article differ from the expert meetings that already take place in the humanities by the former’s procedural and formalised character. In this paper, we aim to highlight to scholars, particularly those in the humanities, what is possible in terms of a systematic approach to academic consensus. In doing so, we aim to initiate exploration of and discussion about the potential of applying such an approach. What would be the added value of applying expert consensus meetings to humanities-related issues? To achieve this, we first explicate our method (§2), before we provide a framework for the application of expert consensus methods by introducing six key elements (§3). Subsequently, we present common types of expert consensus methods utilized in the natural, social, and life sciences. We highlight their and proven value across various types of research through relevant examples and their respective objectives (§4). We then consider both the potential benefits and possible limitations to applying these methods within humanistic research (§5), and conclude with suggestions for future research (§6). Reviewer 1, Comment 2: Second, the section on Method is poor. The composition of the interdisciplinary team (and the reasons for using this team) is not clearly described, the brainstorm session is not properly defined and illustrated, the ‘informal’ literature review is something new for me (what ‘informal’ means?) and it is not acceptable for a research paper to quote two methods used (‘philosophical methods of conceptual analysis’ and ‘conceptual engineering’) without explaining how they have been used for the purposes of the paper but just putting two references. Also, the process of writing and reviewing the manuscript should be better described and justified. Response: Our article, as we now emphasise more, is meant as an exploratory comment and a didactic contribution to inspire scholars in the humanities to see whether consensus methods may be valuable within their own discipline or sub discipline. It is not an instance of systematic empirical research, which would have required a much more detailed methodological section. But we certainly we agree that the method section can be improved in various ways. Therefore, we extended the section by including a specification of the backgrounds of the involved authors and specified the process of our collaboration as follows (see section 2, lines 85-94): To explore the potential of applying expert consensus methods in the humanities, we composed an interdisciplinary team of both researchers from the sciences using consensus methods in practice (LM: epidemiology; TH: qualitative research methodology; LB: epidemiology and data science, research integrity), and researchers familiar with the domain of the humanities and epistemology (CR: art history, museology; RP: epistemology and philosophy of science, religion and theology). In a first brainstorm session, knowledge on the topic was shared from both perspectives. This resulted in a list of consensus methods and examples used in the sciences, drafted by LM and TH. CR carried out a literature search in order to find examples thereof that were applied in her domain. A first draft was written and discussed in a second group session and culminated in a final version, which was edited and reviewed by all authors. Reviewer 1, Comment 3: Third, and more important, the paper refers always to ‘humanities.’ Humanities is a highly fragmented and differentiated field, where many and different epistemic communities work, interact, shape their own idea and uses of consensus method (if any) and where different instances of relationships with other researchers within the field and with neighboring fields are at place. The paper does not acknowledge this important characteristic of humanities (see footnote 1 where there is a list of disciplines included in the humanities that are considered in the paper). The same holds true for STEMs, which are considered as they are all adopting regularly expert consensus methods and there is no explanation or evidence about it. In sum, the paper ‘explores’ the expert consensus method without considering the set of ideas, values, theories, concepts and practices that pertain to the disciplines analyzed and their scientific work. Response: We acknowledge the concern raised and have taken this into account by adjusting a footnote 1 in section 1 (see above) and by revising section 4. Upon reflection we furthermore adjusted the ways to delineate the contexts in which expert consensus methods may be valuable, by focusing on one’s purpose in applying such a method (see section 4, lines 213-235): The subsequent question would be to determine the contexts in which this approach to consensus would prove most valuable. Research in both the humanities and the sciences spans a wide array of disciplines and sub disciplines, with a vast range of variation in types of research and methods, even within the most specialised areas. Classification of research into one of these domains is often gradual in nature, especially since interdisciplinary research increasingly combines approaches traditionally linked to the ‘humanities’ or ‘sciences’. Making general statements on the types of research in which expert consensus methods might be valuable related to the ‘humanities’ versus the ‘sciences’ is therefore difficult. We decided to examine objectives for employing consensus methods in various contexts in which they have demonstrated their value. This could contribute to a directory of instances in which these methods may prove useful in humanities-related fields. Gattrell et al. (2024, Table 2) present examples of these objectives in context of the health sciences, such as the establishment of clinical practice guidelines, diagnostic guidelines or reporting guidelines but also the classification of diseases, the setting research priorities, or the formulation of policy. These aims are summarised, adjusted and extended in Table 4 in a manner that enhances their generalisability to other fields of research and were included in Table 2 and 3. Although these examples highlight specific applications, we think that there is a broader potential for its use in the humanities. For instance, in establishing consensus and guidelines on the best way to preserve certain historical objects (guidelines/ policy development), involving eyewitnesses in the reconstruction of an historical event (current state of knowledge), or consulting various experts in establishing how to interpret an ancient source (current state of knowledge). Future research and case studies exploring the potential applications outlined in Table 4 could provide insights into specific instances where the pursuit of formal dis- or consensus may prove effective. Reviewer 1, Comment 4: Fourth, sections 3 and 4 provide a clear summary of what expert consensus methods are and a deepen analysis of four types. Then the reader expects Section 5 discusses the results with respect to the exploration done. On the contrary, Section 5 lists of pros and cons related to the expert consensus method, and some examples of applications; the contents are completely unrelated with what the sections 3 and 4 have presented and the motivations for considering the expert consensus method an interesting development for humanities are unclear. Here the suggestion is to revise the sections trying to answer the research questions (when formulated) and highlight what we have learnt from the exploratory work, what is the contribution of the brainstorming to the exploration, what new avenues of research can be envisaged based on the results obtained, and what limitations/obstacles can the expert consensus model have in the humanities with respect to the other fields. Response: Thank you for pointing this out to us. Based on this valuable feedback, we have worked to better align section 5 with the preceding sections to create a more cohesive flow throughout the paper and have made more explicit what our line of reasoning is. However, because the paper is intended as preliminary exploration, part of our manuscript aims to identify the potential value of these methods for the humanities. Section 5 is dedicated to that purpose and we made that more explicit in the revision (see section 5, lines 253-315): In the preceding sections, we examined the elements and dynamics of expert consensus methods, along with examples and possibilities for intended objectives. In this section, we delve into the question why formal approaches to consensus in the humanities remain scarce, despite the significant factor of consensus and dissensus in knowledge production across research cultures. Aside from limitations, we also address what the potential added value of applying consensus methods to humanities-related issues could be. First of all, practical limitations likely play a role. Expert consensus methods and their subsequent publication demand significant time and resources, which are rarely in abundance in the humanities. Additionally, a factor that may affect academia broadly is the emphasis on publication quantity and individual contributions, which can discourage scholars from lengthy endeavours for which recognition is shared (Leising et al., 2024). Moreover, expert consensus methods are not interwoven in humanities research traditions. Researchers may not be familiar with consensus methods, and the advantages they may have for their area of research. Secondly, concerns raised in the field of psychology that may apply in the humanities as well, highlight risks of consensus-building. Emphasising consensus may lead to agreement for the wrong reasons, such as conformity, which may be harmful to innovation, creativity and minority views. It can sideline dissent, whilst that is a vital element in the evolution of science (Gollwitzer, 2022; Asendorpf & Gebauer, 2022; Beck et al., 2022; Denissen & Sijtsma, 2022; Hilbig et al., 2022). Additionally, experts agreeing on something does not necessarily mean they are correct. They may be in full agreement on a point that is incorrect or untrue. Scholars may also disagree due to the role of research paradigms or schools within the humanities, as researchers’ positionality can shape their interpretations. In addition to that, humanities research can be concerned with hermeneutics, i.e., issues that have to do with understanding the values and meanings of texts and other objects. And many humanities fields such as ethics, epistemology, and metaphysics, rely on apriori methods like thought experiments, reasoning on the basis of intuitions and principles, and formal logic rather than empirical data. This allows for multiple interpretations that can all to some extent be warranted by the evidence base, and may not readily align into a singular consensus. These factors suggest that the complexity and interpretative nature of humanities research might render expert consensus methods to some extent less relevant. Nevertheless, we believe that there are compelling reasons for considering expert consensus methods in humanities research. First of all, opposing the above concerns, one of the important purposes of expert consensus methods is to bring various viewpoints, nuances, and interpretations to the surface, so that they can be discussed. Depending on the design and purpose of the consensus method, it could reveal levels of consensus and/or dissensus on an issue, within or across schools of thought or paradigms. They might even have the potential to articulate both divergences and common grounds regarding an issue, and by extension increase mutual understanding. The application of such a method may therefore stimulate epistemic progress in itself. Moreover, a carefully designed protocol and curated group of experts can foster a fair practice and ensure the consultation of diverse viewpoints. Expert consensus methods provide opportunities to mitigate biases and their ramifications by this cautious selection of experts, but also by blinding certain parts of the process in relation to anonymity of participating experts, for example. Secondly, expert consensus methods can be helpful in getting a firmer grip on the status quo or state of the art on a particular issue in a field (e.g. Hodgkin et al. (1975) mentioned in Table 2; Rulkens et al. 2022-2023, mentioned in Table 3). Of course, scholars often already have an idea about that, and such an informed opinion is valuable in itself. However, it can be complemented by a more structured approach, particularly when the number of scholars involved is large. This approach can additionally provide guidance for future research by pointing out the gaps in the current evidence base. And when applied on a recurrent basis, it might even have the potential to gain insight in how views evolve over time. It is important to note that consensus is not a final endpoint; rather, it can be continually re-evaluated in light of new insights and reasonable doubts regarding its validity. Third, an advantage of expert consensus methods is that they enhance the transparency and replicability of research because of their well-documented and systematised approach (for replication in the humanities, see Peels and Bouter (2018)). It can therefore be instrumental to epistemic progress beyond the consensus reached at a moment in time. Fourth, application or implementation of its outcomes is often easier after the process of a consensus method is completed (e.g. Standing Committee for the Museum Definition (2020-2022) (2022), mentioned in Table 3). And finally, expert consensus methods can aid public understanding by clarifying experts perspectives on specific issues (Stekelenburg et al., 2022) thereby enabling the public to make informed decisions (Ordway, 2021). In response to this comments, we furthermore extended suggestions for future research in our conclusion (see section 5, lines 325-335): However, applying a consensus method is a costly endeavour in terms of time and means. There might be conditions under which these efforts do not live up to the epistemic benefits they provide for. The identification of the particular areas and issues where these methods hold most promise can be done by assessing the purposes these methods have served, and translating those to a specific area of research. The assessment and prioritisation of issues, especially when resources are limited, should be carried out locally by researchers who are familiar with the epistemologies and issues relevant to their field. Consequently, it is essential to carry out future research involving humanities case studies that employ expert consensus methods, to evaluate their feasibility and effectiveness. Collaboration between humanities researchers with scholars from other domains experienced in designing consensus methods could facilitate mutual learning and foster innovative approaches. Response to reviewer 2: Daniel Leising, Technische Universitat Dresden, Dresden, Saxony, Germany Reviewer 2, Comment 1: First, I would argue that the Humanities are actually not that “unique” in terms of their nature and approaches (page 11). To enable a proper discussion of this issue, the authors could explain their own view of what is so specific about the Humanities in a bit more detail. Here is my view: Interviews and text analyses are used in many different branches of science. Exploratory research is also conducted in many fields. The ways in which people give meaning to their experiences are also of interest across various branches of science. And even in many of the “harder” sciences, research is often “coloured by the backgrounds and positionality of the researchers” (page 8). So, much of what is common in research in the Humanities is actually shared with other fields. In my view, two features may actually distinguish “the Humanities” from other fields - but even these distinctions are only gradual in nature. One feature is the common practice of using everyday language as the primary scientific language. The other feature is the relative fuzziness of many concepts - which I would note is not necessarily a weakness but may often be unavoidable given the complexity of the phenomena of interest (and also given the first feature just mentioned). Do the authors agree? Response: Thank you for challenging us on this point. We agree that the borders between the general fields of the natural, social, and life sciences, and the humanities are fuzzy, when it comes to their objects, purposes, and methods. Whether there is something distinctive about the humanities – we believe that is compatible with fuzziness at the boarders – is a matter that we cannot possibly explore in this article (one of us does so in a book entitled A Philosophy of the Humanities that is forthcoming in 2025 with Oxford University Press, a volume co-authored by Stephen Grimm, Rik Peels, and René van Woudenberg). We do believe though that ordinary language use is not generally characteristic of the humanities (think of the highly technical language of metaphysics and epistemology), nor do we believe that insufficient clarity about core concepts is characteristic of the humanities. We agree though with the idea, also drawn to attention to by reviewer 1 in comment 3, that the humanities are a highly diversified field. We included that by adjusting footnote 1 in section 1: We acknowledge that as is the case in the sciences, the humanities encompass a large amount of disciplines and sub fields, exhibiting significant diversity in research types and methodologies. However, in this paper we refer to ‘the humanities’ to clarify difference for the reader, rather than out of belief in rigid categorisations. For readers interested in this topic, see e.g. Woudenberg (2018) and Grimm et al., (forthcoming). Additionally, to be able to demarcate between contexts in which the formal approach might be beneficial, we decided to put an emphasis on the purposes for which these methods are employed in other domains. This would be a way in which researchers with expertise in their fields could identify opportunities for a formal approach to consensus (see section 4, lines 213-235, also quoted above in response to comment 3 of reviewer 1): The subsequent question would be to determine the contexts in which this approach to consensus would prove most valuable. Research in both the humanities and the sciences spans a wide array of disciplines and sub disciplines, with a vast range of variation in types of research and methods, even within the most specialised areas. Classification of research into one of these domains is often gradual in nature, especially since interdisciplinary research increasingly combines approaches traditionally linked to the ‘humanities’ or ‘sciences’. Making general statements on the types of research in which expert consensus methods might be valuable related to the ‘humanities’ versus the ‘sciences’ is therefore difficult. We decided to examine objectives for employing consensus methods in various contexts in which they have demonstrated their value. This could contribute to a directory of instances in which these methods may prove useful in humanities-related fields. Gattrell et al. (2024, Table 2) present examples of these objectives in context of the health sciences, such as the establishment of clinical practice guidelines, diagnostic guidelines or reporting guidelines but also the classification of diseases, the setting research priorities, or the formulation of policy. These aims are summarised, adjusted and extended in Table 4 in a manner that enhances their generalisability to other fields of research and were included in Table 2 and 3. Although these examples highlight specific applications, we think that there is a broader potential for its use in the humanities. For instance, in establishing consensus and guidelines on the best way to preserve certain historical objects (guidelines/ policy development), involving eyewitnesses in the reconstruction of an historical event (current state of knowledge), or consulting various experts in establishing how to interpret an ancient source (current state of knowledge). Future research and case studies exploring the potential applications outlined in Table 4 could provide insights into specific instances where the pursuit of formal dis- or consensus may prove effective. Reviewer 2, Comment 2: Second, I would like to encourage the authors to say a bit more about possible reasons for why so little consensus building is taking place in many fields of science, despite it being widely viewed as desirable. In my view, there are a couple of reasons, most of which are rather extraneous to an idealised image of science (as a collaborative and cumulative effort at building a reliable knowledge base for the good of mankind). For example, consensus building is likely to reduce the publication output of a field, which would be at odds with the interests of commercial publishers. Also, individual researchers have an interest in “making a name for themselves” based on their specific expertise or views regarding certain topics. Consensus building makes it more difficult to maintain a highly distinct profile and thus reduce visibility. This may be risky in a science system that is as competitive as the one we choose to entertain. And then there may be a need to “save face”: The more of your career you have dedicated to certain ideas (e.g., your own theory of some phenomenon), the harder it may be to let those ideas go - even if the evidence against them is overwhelming. There is a reason why Max Planck once stated that scientific progress happens “one funeral at a time”. At least a brief nod to the relevance of such more political factors would be nice - if the authors share my conviction that they are relevant. Who has an actual interest in achieving consensus? Who has an interest in avoiding the emergence of consensus? And what are the external incentives for and against this type of work? Response: We think that these are valid points and the reviewers’ publications on the topic shed important light on this. We agree that external incentives may work against a formal approach to consensus as follows and have therefore slightly revised the manuscript on this point. We remain rather brief, though, as the exact explanation of why consensus methods are still relatively rarely used even in the sciences is an issue that is beyond the scope of our paper (see section 5, lines 259-264, also quoted in response to comment 4 of reviewer 1): Expert consensus methods and their subsequent publication demand significant time and resources, which are rarely in abundance in the humanities. Additionally, a factor that may affect academia broadly is the emphasis on publication quantity and individual contributions, which can discourage scholars from lengthy endeavours for which recognition is shared (Leising et al., 2024). Moreover, expert consensus methods are not interwoven in humanities research traditions. Researchers may not be familiar with consensus methods, and the advantages they may have for their area of research. Reviewer 2, Comment 3: Third, I would also appreciate if the authors could talk some more about “bad consensus” – views that are held by many, despite their being wrong. With a particular eye to the Humanities – what criteria are there, apart from consensus, that may help us determine whether a view is “correct”? Or may truth in the Humanities only be defined in terms of relative consensus among researchers? In the harder sciences, the ability to predict the occurrence of certain events from a theory (and some data input) plays a crucial role as evidence of the theory's correctness. Are the Humanities able to produce testable predictions, too? Can there be consensus projects in the Humanities that actually test one theory against another? Response: This is an interesting point, that would need a elaborative reflection in order to do justice to the complexity of this issue. It is important to note, that we do believe that consensus and truth are two different things, and we made this more clear in our revised paper. Based these suggestions we also incorporated that consensus can be reached based on the wrong reasons and (see section 5, lines 265-271, also quoted in response to comment 4 of reviewer 1): Secondly, concerns raised in the field of psychology that may apply in the humanities as well, highlight risks of consensus-building. Emphasising consensus may lead to agreement for the wrong reasons, such as conformity, which may be harmful to innovation, creativity and minority views. It can sideline dissent, whilst that is a vital element in the evolution of science (Gollwitzer, 2022; Asendorpf & Gebauer, 2022; Beck et al., 2022; Denissen & Sijtsma, 2022; Hilbig et al., 2022). Additionally, experts agreeing on something does not necessarily mean they are correct. They may be in full agreement on a point that is incorrect or untrue. Reviewer 2, Comment 4: Fourth, on page 3 the authors state that “Since the questions addressed in the humanities may leave room for multiple viewpoints that are equally warranted by the evidence base, it can be infeasible or not desirable to get a complete or almost complete consensus.” To this I would respond: what are the conditions under which working toward consensus would not be feasible, or desirable? I agree that this may be the case – but when is it the case, and when is it not the case? I personally hold the somewhat sceptical view that many researchers will shy away from working toward consensus even if doing so would be both feasible and desirable (see my previous point). Such a “live and let live” attitude may often be relatively comfortable to have (e.g., to avoid conflict). It may, however, be regarded as unethical, because it will foster the preservation of an unnecessary level of ambiguity. Response: We appreciate this insightful suggestion. Our approach to expert consensus methods does not necessarily imply working towards consensus in all instances, but is also related to the assessment of the (lack of) consensus. We view measuring consensus as a means, not an objective in itself. Furthermore, as put forward in paragraph 3, the bar for ‘sufficient’ consensus is something consensus researchers can determine their selves. Applying a consensus methods is a costly endeavour in terms of time and means. There might be conditions under which the effort of applying such a method does not live up to the epistemic benefits it provides for. We furthermore believe that the feasibility and desirability of assessing consensus is an issue that merits further exploration and included this as follows (see section 5, lines 325-335): However, applying a consensus method is a costly endeavour in terms of time and means. There might be conditions under which these efforts do not live up to the epistemic benefits they provide for. The identification of the particular areas and issues where these methods hold most promise can be done by assessing the purposes these methods have served, and translating those to a specific area of research. The assessment and prioritisation of issues, especially when resources are limited, should be carried out locally by researchers who are familiar with the epistemologies and issues relevant to their field. Consequently, it is essential to carry out future research involving humanities case studies that employ expert consensus methods, to evaluate their feasibility and effectiveness. Collaboration between humanities researchers with scholars from other domains experienced in designing consensus methods could facilitate mutual learning and foster innovative approaches. Minor comments: Reviewer 2, Comment 5: Fifth, the authors state that a steering committee would typically be comprised of the people who initiated the process and who will ultimately author the consensus document. Could there not be cases in which the process should better be overseen by an independent third party (e.g., a moderator)? Especially when issues are highly contentious, for example? And should not all the experts who worked on the consensus be authors on the consensus document? Response: Thank you for this relevant point, this is certainly the case and we made this more clear in the revised manuscript by the following addition (see section 3, lines 193-198): Naturally, the six elements can be specified at by the steering committee. If they deem it valuable to include an independent moderator or to incorporate a member with a critical or divergent perspective—especially if the topic is highly controversial—this remains within their discretion. Additionally, it is up to them to decide who will be listed as co-authors, whether that includes only the steering committee, all participants, or a broader group representation such as the panellists as a whole, for instance. Reviewer 2, Comment 6: Sixth, something is not quite right on page 4, text lines 16-19. Response: Thank you for noticing, we have corrected this sentence as follows (see section 3, lines 136-140): Expert consensus methods are (i) applied by a process leader or steering committee, (ii) to reach consensus or assess the degree of consensus among a group of experts, (iii) about an issue, (iv) based on preset rules of engagement, (v) with the aim to deliver useful output, (vi) for future users. They are characterised by constructive and procedural usage of (dis)agreement and the different arguments that are provided therein. Reviewer 2, Comment 7: Seventh, I would say that evidence will never “speak for itself” (page 4) - otherwise papers in certain scientific fields would not need any Discussion sections. Response: Thank you for rightly pointing this out to us. We adjusted this as follows (see section 3, lines 132-134): This can be useful when the issue at hand is an important one and when evidence is lacking, it is inconsistent, or its interpretation is not obvious. As a result of our reactions to the comments, we also made adjustments to our abstract and conclusion, which are not all quoted in the above letter. We hope these revisions and clarifications meet the reviewers’ expectations. Please let us know if any further revisions are needed. Sincerely, Charlotte Rulkens MA Prof. dr. Rik Peels Dr. Lidwine Mokkink Dr. Tamarinde Haven Prof. em. dr. Lex Bouter [1] We acknowledge that as is the case in the sciences, the humanities encompass a large amount of disciplines and sub fields, exhibiting significant diversity in research types and methodologies. However, in this paper we refer to ‘the humanities’ to clarify difference for the reader, rather than out of belief in rigid categorisations. For readers interested in this topic, see e.g. Woudenberg (2018) and Grimm et al., (forthcoming)." } ] } ]	1	https://f1000research.com/articles/13-710
https://f1000research.com/articles/13-1521/v1	16 Dec 24	{ "type": "Research Article", "title": "Isolation and characterization of a novel bacteriophage as a biological control agent against multidrug resistant Escherichia coli in compost and agricultural irrigation water", "authors": [ "Luis Amarillas", "Ruben León-Chan", "Enrique López-Avendaño", "Antonio González-Balcázar", "Jorge Padilla", "Luis Lightbourn-Rojas", "Mitzi Estrada-Acosta", "Luis Amarillas", "Ruben León-Chan", "Enrique López-Avendaño", "Antonio González-Balcázar", "Jorge Padilla" ], "abstract": "Background\nEscherichia coli is a critical priority pathogen due to its significant morbidity, mortality, and growing antimicrobial resistance, underscoring the urgent need for novel control strategies. This bacterium is frequently implicated in outbreaks associated with horticultural products, particularly those cultivated in organic farming systems. The aim of this study was to isolate and evaluate the potential of a bacteriophage as a biocontrol agent against E. coli in compost and agricultural irrigation water.\n\nMethods\nE. coli presence in compost samples (n=17) was determined through microbiological assays, and the bacterial identity was confirmed by PCR amplification of the phoA gene. Antimicrobial resistance profiles of the isolates were assessed using the disk diffusion method. Bacteriophage isolation was conducted from livestock fecal samples using a double-layer agar technique. The stability of the bacteriophage under varying pH levels and temperatures was evaluated, along with its replication dynamics. Additionally, the phage’s efficacy in reducing E. coli populations in compost and irrigation water was assessed. Genomic sequencing and bioinformatic analyses of the bacteriophage were conducted to characterize its genetic profile.\n\nResults\nE. coli strains isolated exhibiting multidrug resistance were isolated from compost samples. The isolated bacteriophage, named Alux-21, exhibited stability at neutral pH and retained viability at both 4°C and 40°C over a six-month period. Importantly, the phage achieved a significant reduction of E. coli counts, exceeding 3.8 logs in compost and 3 logs in irrigation water, demonstrating its superior efficacy compared to previously reported phages in similar substrates. Genomic analysis confirmed the absence of virulence-associated, lysogeny, and antibiotic resistance genes.\n\nConclusion The findings highlight Alux-21 as a sustainable biocontrol agent for E. coli in compost and irrigation water. Field validation will be crucial to establish its scalability and efficacy under real-world agricultural conditions.", "keywords": [ "Phage", "pathogen reduction", "environmental pathogens", "antimicrobial resistance", "water quality management", "composting processes", "biocontrol agents", "agricultural sustainability" ], "content": "Introduction\n\nAntimicrobial resistance (AMR) in bacteria has emerged as one of the most critical threats to global public health, endangering millions of lives worldwide. It is estimated that antimicrobial-resistant bacteria account for approximately 13,600 deaths per day (Johnston et al., 2021; Murray et al., 2022). This crisis underscores the urgent need for new therapeutic approaches to effectively combat AMR (Xiao et al., 2023). Among the pathogens identified as critically important by the World Health Organization (WHO) is Escherichia coli, a bacterium characterized by its dual threat of causing severe infections and exhibiting increasing levels of antimicrobial resistance. This combination complicates treatment options, further exacerbating global morbidity and mortality rates (Ikuta et al., 2022; Jesudason, 2024).\n\nEscherichia coli is a genetically diverse bacterium with significant public health implications due to its association with antimicrobial resistance and foodborne illnesses. Numerous epidemiological outbreaks have been linked to E. coli contamination in fresh produce, a problem worsened by the increasing consumption of organic foods (Heiman et al., 2015; Irvin et al., 2021; Tack et al., 2021; Waltenburg et al., 2022). In recent years, the global demand for organic produce has surged, promoting the adoption of organic farming practices (Bernabéu et al., 2023). While these practices have gained popularity worldwide, they often involve the use of organic soil amendments to improve soil fertility. However, inadequate quality control of these amendments may lead to contamination with foodborne pathogens, posing risks to soil, irrigation water, and produce (Murphy et al., 2024).\n\nE. coli is a leading pathogen in organic food outbreaks, frequently linked to tomatoes, lettuce, and cucumbers (Coulombe et al., 2020; Irvin et al., 2021; Pires et al., 2023). Additionally, agricultural irrigation water also presents a critical route for E. coli entry into food production systems, particularly with crops consumed raw, where contamination can directly impact consumer safety. As such, effective management practices that enhance the microbiological quality of irrigation water are essential to mitigate food safety risks. These two vectors are significant pathways for introducing pathogenic bacteria into agricultural environments, creating substantial food safety and public health challenges. In this context, bacteriophages have emerged as promising biocontrol agents for mitigating E. coli contamination in agricultural settings. Their application offers a sustainable and targeted approach to enhance food safety (Álvarez et al., 2019; Soliman et al., 2023).\n\nIn organic farming, bacteriophages could play a pivotal role in reducing pathogen transmission, particularly when compost and irrigation water fail to meet microbiological quality standards. Despite their potential, few studies have investigated bacteriophage efficacy in compost or irrigation water, and no bacteriophage to date has been shown to effectively control E. coli in both environments (Heringa et al., 2010; Álvarez et al., 2019).\n\nFor biocontrol applications, bacteriophages must meet stringent criteria. They must demonstrate high lytic activity against bacterial cells and maintain stability under diverse environmental conditions such as varying temperatures, pH levels, and salinity, ensuring consistent performance (João et al., 2021). Moreover, it is crucial that bacteriophages lack genes associated with virulence, lysogeny, antibiotic resistance, or allergenicity (Suh et al., 2022). Scalability is also essential, requiring biological characteristics that enable large-scale propagation. Given these considerations, this study aimed to isolate and evaluate the potential of a bacteriophage as a biocontrol agent against E. coli in two critical agricultural environments: compost and irrigation water.\n\n\nMethods\n\nSeventeen compost samples were collected on July 3 and 5, 2023, from mature compost piles composed of a 10:4 (w/w) mixture of bovine manure and plant residues. These samples were selected to assess spatial variations and heterogeneity within the compost. The number of samples was determined based on the expected variability of Escherichia coli in compost and irrigation water, as informed by previous studies (Thomas et al., 2020). The compost piles, located at commercial composting facilities in Culiacán, Sinaloa, México, were approximately 5 meters in length and 2.5 meters in width. To guarantee representativeness, samples were collected from various points within each pile using a simple random sampling approach. Following collection, the samples were transported to the laboratory in refrigerated coolers.\n\nFor initial processing, 10 g of each sample were weighed and inoculated into 90 ml of buffered peptone water. This suspension was incubated at 37 °C for 24 hours. Following incubation, an aliquot of the culture was plated onto Petri plates containing CHROMagar™ E. coli medium (CHROMagar™ Microbiology, France) using a sterile inoculating loop. The plates were incubated at 37 °C for an additional 24 hours (Heratherm IMC18, Thermo Scientific™, USA). Colony morphology was then examined to identify characteristics typical of E. coli, including the presence of intense blue or pink colonies with smooth edges. Representative colonies were selected and plated onto fresh CHROMagar™ E. coli plates, followed by incubation at 37 °C for 24 hours. This re-isolation procedure was repeated five times to ensure the isolation of a pure strain (Werner et al., 2022).\n\nFor molecular identification, polymerase chain reaction (PCR) analysis was performed following the protocol of Li et al. (2020), with slight modifications. Amplification of the phoA gene was carried out using the GoTaq® PCR Core system (Promega, USA). The final reaction mixture contained 4.8 μl of 5× Green GoTaq buffer, 0.8 μl of dNTPs, 2.5 μl of MgCl2, 0.8 μl of each primer, 0.35 μl of Taq polymerase, and 100 ng of bacterial DNA template. PCR conditions included an initial denaturation at 94 °C for 7 minutes, followed by 30 cycles of denaturation at 94 °C for 30 seconds, annealing at 56 °C for 30 seconds, and extension at 72 °C for 30 seconds (Mastercycler Nexus, Eppendorf®, Germany). A final extension step was performed at 72 °C for 5 minutes. The resulting 622 bp gene fragment was visualized by electrophoresis on a 1.2% agarose gel at 60 V for 2 hours. The gel was stained with GelRed™ Nucleic Acid Gel Stain (Biotium, USA) and viewed under UV transillumination.\n\nTo assess the antimicrobial resistance profiles of E. coli strains, a panel of 12 antibiotics (Oxoid, USA) representing various antibiotic classes was used: carbapenems (imipenem), aminoglycosides (gentamicin, amikacin), penicillins (ampicillin, amoxicillin/clavulanic acid), fluoroquinolones (ciprofloxacin), quinolones (nalidixic acid), phenicols (chloramphenicol), tetracyclines (tetracycline, oxytetracycline), polymyxins (colistin), third-generation cephalosporins (cefoperazone), and sulfonamides (trimethoprim/sulfamethoxazole). Antimicrobial susceptibility testing was conducted using the disk diffusion method, following the guidelines of the Clinical and Laboratory Standards Institute (CLSI). For this procedure, bacterial cultures were resuspended in sterile 0.85% saline solution and adjusted to an optical density of approximately 0.1 at 600 nm. Subsequently, 200 μl of the bacterial suspension was spread onto Mueller-Hinton agar plates using a sterile swab to ensure an even inoculum distribution across the medium. Antibiotic-impregnated paper disks were then placed on the agar surface, and the plates were incubated at 37 °C for 24 hours. Following incubation, inhibition zone diameters were measured with a millimeter-calibrated Vernier caliper. The results were interpreted according to CLSI susceptibility criteria, classifying the strains as susceptible, intermediate, or resistant (CLSI, 2022).\n\nBacteriophage isolation was performed using fecal samples collected on August 7 and 8, 2023, from livestock housed at the “La Posta Zootecnia” animal production unit of the Faculty of Agronomy, Autonomous University of Sinaloa. Ten grams of each fecal sample were transferred into 90 ml of sterile purified water and mixed thoroughly by inversion. Subsequently, 35 ml of the mixture was transferred to centrifuge tubes and centrifuged at 3,500 × g for 10 minutes (Sorvall™ 17R, Thermo Scientific™, USA). The supernatant was collected in a sterile tube and subjected to two additional rounds of centrifugation under the same conditions. The final supernatant was filtered through a 0.45 μm pore-size nitrocellulose membrane, and the filtrate was collected for further processing.\n\nBacteriophage isolation was conducted using the double-layer agar technique. A 1 ml aliquot of an exponential-phase E. coli culture, grown in tryptic soy broth (TSB, MCD LAB, México) was added to 3 ml of molten 0.4% TSB-agarose, maintained at 50 °C. This mixture was poured onto tryptic soy agar (TSA, MCD LAB, México) plates. Once solidified, four 5 μl drops of the filtrate were placed onto the surface of the solidified medium. After drying, plates were incubated at 37 °C for 18–24 hours. Following incubation, plates were inspected for lytic zones, indicating bacteriophage activity against E. coli (Kosznik-Kwaśnicka et al., 2020).\n\nLytic zones were carefully excised using a sterile plastic Pasteur pipette and individually transferred to microcentrifuge tubes containing 500 μl of sterile purified water, followed by gentle mixing by inversion. This suspension was centrifuged at 8,500 × g for 5 minutes, and the supernatant was transferred to a new sterile microcentrifuge tube. To increase the bacteriophage concentration, a second round of the double-layer agar technique was performed. Specifically, 1 ml of bacterial culture and 100 μl of the recovered supernatant were added to 3 ml of 0.4% TSB-agarose and poured onto TSA plates. Plates were incubated at 37 °C for 18–24 hours. After incubation, plates were examined for lytic plaques, from which the largest and clearest plaques were excised using a sterile plastic Pasteur pipette and transferred to a microcentrifuge tube containing 500 μl of sterile purified water. This isolation and purification process, involving lytic plaque excision and the double-layer agar technique, was repeated five times to ensure the isolation of a single bacteriophage type (Topka et al., 2019).\n\nTo increase the concentration of the bacteriophage, a double-layer agar technique was employed as previously described. After the incubation period, 5 ml of sterile water was added to each Petri dish, and the plates were left at room temperature for 2 hours. The soft agar layer was then carefully scraped off using a sterile bacteriological loop and transferred to a sterile 50 ml conical tube. The collected suspension underwent centrifugation at 3,500 × g for 10 minutes at room temperature, after which the supernatant was transferred to a sterile tube for subsequent processing. This centrifugation process was repeated three additional times to ensure maximum removal of culture medium residues and bacterial debris.\n\nFollowing the initial centrifugation steps, the final supernatant was passed through a 0.22 μm sterile syringe filter with a polyethersulfone membrane (GVS Filter Technology, USA), and the resulting filtrate was transferred to a polypropylene tube for further concentration by ultracentrifugation at 40,000 × g for 2 hours at 4 °C (MSE PRO MA1084-1, USA). After the centrifugation, the supernatant was discarded, and the resulting pellet was resuspended in 10 ml of sterile water.\n\nThe bacteriophage suspension was subsequently purified through dialysis using a Slide-A-Lyzer device with a molecular weight cutoff (MWCO) of 20,000 (Thermo Fisher, USA), following the protocol outlined by Le Guellec et al. (2023). Lastly, the concentration of the bacteriophage was determined via serial tenfold dilutions and the double-layer agar technique to quantify the phage titers accurately.\n\nTo assess the stability of the bacteriophage across a range of pH values, a series of sterile saline solutions (0.85%) were prepared, adjusted to pH 3, 5, 7, 9, and 11 (± 0.05) using either 5 M HCl or 5 M NaOH. A purified bacteriophage suspension was added to each pH-adjusted solution at a 99:1 ratio, achieving a final concentration of 1 × 106 PFU/ml. These suspensions were then incubated at 37 °C for 1 hour. The viability of the bacteriophage was subsequently evaluated through serial tenfold dilutions and the double-layer agar method, as described by Bagińska et al. (2024). All experiments were conducted in triplicate to ensure reproducibility.\n\nBased on the results of the pH stability tests, the bacteriophage was diluted in sterile water adjusted to the pH at which optimal stability was observed, yielding a final concentration of approximately 1 × 108 PFU/ml. This suspension was divided into two aliquots: one was stored at 4 °C, and the other at 40 °C. Bacteriophage concentration in each sample was monitored monthly for a duration of 6 months using serial tenfold dilutions and the double-layer agar technique. Each assay was performed in triplicate to confirm consistency and reliability of the results.\n\nTo investigate the replication dynamics of the bacteriophage, a single colony of E. coli was inoculated into 100 ml of TSB and incubated in a water bath at 37 °C with constant agitation at 80 rpm (Shel Lab, USA). Once the bacterial culture reached an optical density (OD) of 0.1 at 600 nm, corresponding to an approximate concentration of 0.8 × 108 CFU/ml, the bacteriophage was added at a multiplicity of infection (MOI) of 0.1. Aliquots were collected from the culture every five minutes and transferred into microcentrifuge tubes.\n\nTo assess intracellular phage replication, 100 μl of chloroform was added to one of the tubes, while the other tube was left untreated. The addition of chloroform lyses the bacterial cells, releasing intracellular phage particles. Both treated and untreated samples were then centrifuged at 10,000 × g for 1 minute, and the bacteriophage concentration in the supernatant was quantified using serial tenfold dilutions followed by the double-layer agar method, as outlined by Kropinski (2018). All bacteriophage concentrations were determined in triplicate to ensure accuracy and reproducibility. as outlined by\n\nA single colony of E. coli was inoculated into TSB and incubated for 24 hours at 37 °C. Subsequently, 1 ml of this culture was transferred into four flasks, each containing 200 ml of TSB, and incubated at 37 °C with agitation at 80 rpm. The optical density of the bacterial culture was monitored at a wavelength of 600 nm (OD600).\n\nWhen the optical density reached 0.5 (~2 × 108 CFU/ml), purified bacteriophage suspension was added to three of the flasks at different multiplicities of infection (MOI): 0.1, 0.01, and 0.001, respectively. The fourth flask served as a control to monitor E. coli growth in the absence of bacteriophage. All flasks were incubated under the same conditions as previously described, and the optical density of the cultures was measured hourly over a period of 6 hours, following the protocol outlined by Khunti et al. (2023).\n\nThis experimental design allowed for the assessment of the bacteriolytic effect of the bacteriophage at varying MOIs, providing insights into its efficacy against E. coli. All measurements were performed in triplicate to ensure reliability and accuracy of the results.\n\nCompost was prepared using a mixture of bovine manure and vegetable residues in a ratio of 10:4, obtained after 90 days of composting at a facility in Culiacán, Sinaloa, México. The compost was sterilized at 121 °C for 20 minutes under pressure, with this process repeated three times to ensure sterility. Following sterilization, the moisture content was determined using the drying method and adjusted with sterile water to achieve a final moisture level of 30% (Heringa et al., 2010). For the experiment, 100 g of compost was placed in each of three sterile, sealable plastic bags. The following treatments were applied: in the first bag, 1 ml of sterile water was added as absolute control; in the second bag, 1 ml of a purified E. coli suspension was introduced to reach a final concentration of 1 × 106 CFU/g (bacterial control). This concentration was selected based on previous studies reporting elevated levels of E. coli in various biofertilizers (Black et al., 2014; Miller et al., 2013). In the third bag, 1 ml of the same bacterial suspension was added, achieving the same concentration as the bacterial control. After a 1-hour incubation period to allow for acclimatization, 500 μl of a purified bacteriophage suspension was introduced to the third bag, resulting in a final concentration of 1 × 104 PFU/g (phage treatment).\n\nThe contents of the bags were manually mixed, and the compost was kept under environmental conditions characteristic of agricultural systems in Culiacán, Sinaloa for 48 hours. Ambient temperatures ranged from 19 to 31 °C, while relative humidity levels were regularly between 55% and 60%. These parameters were specifically used to closely replicate the agricultural environments characteristic of the region, thus increasing the ecological relevance and practical applicability of the experimental findings. At the conclusion of the incubation period, 10 g of compost was sampled from each treatment, mixed with 90 ml of sterile water, and transferred to new sealable bags, which were vigorously agitated for 5 minutes. Decimal dilutions were then performed, and bacterial concentration was quantified using the spread plate method on CHROMagar™ E. coli (Heringa, 2008). This experimental design enabled the assessment of the bacteriophage’s effectiveness in reducing E. coli levels in compost, contributing to understanding the potential of bacteriophages as biological control agents in agricultural practices.\n\nFor the irrigation water samples, five distinct locations were selected for analysis, strategically selected in areas of intensive agricultural use within the Culiacán Valley, México. These locations were specifically selected because of their proximity to high-risk crops, such as tomatoes and cucumbers. The sampling was conducted between September 4 and 6, 2023, using a stratified convenience sampling strategy, ensuring that the samples are representative of typical agricultural settings.\n\nFollowing standard sampling protocols, agricultural irrigation water samples were collected in 1-liter sterile polypropylene bottles from various locations within the Culiacán Valley, Sinaloa, México. The samples were transported to the laboratory under refrigerated conditions, where pH (Thermo Scientific Orion Star A111, USA), electrical conductivity (Hanna Instruments Hi98130, USA), and organic matter content were determined using the loss-on-ignition method. Each water sample underwent sterilization three consecutive times at 121 °C under pressure for 15 minutes.\n\nSubsequently, samples were selected based on common values observed in the analyzed data. Specifically, samples with a pH of approximately 7.14, an electrical conductivity of 0.20 mS, and an average organic matter content of 0.08% were chosen for the biological control assays. These parameters reflect average conditions of irrigation water in agricultural fields of the Culiacán Valley, ensuring that the experiments closely mimic real-world agricultural environments, thereby enhancing the practical relevance of the findings. In two sterile conical tubes, 35 ml of sterile irrigation water was added, followed by the inoculation of a purified E. coli suspension to achieve a final concentration of approximately 1 × 106 CFU/ml. This concentration was selected to simulate elevated contamination levels in irrigation waters within agricultural regions, which pose a high risk of introducing pathogens into the food supply system, thereby providing a realistic context for evaluating the bacteriophage’s efficacy as a biological control agent (Shaw et al., 2016; Yin et al., 2020).\n\nAdditionally, one of the tubes received a bacteriophage inoculation at a final concentration of 1 × 104 PFU/ml. The contents of the tubes were gently mixed by inversion and incubated at room temperature for 24 hours. Following incubation, bacterial concentrations in each treatment were quantified using serial tenfold dilutions and the plate count method on selective CHROMagar™ E. coli medium (Álvarez et al., 2019). All assays were performed in triplicate to ensure reproducibility and reliability of the results. This experimental framework allows for a comprehensive assessment of the bacteriophage’s potential as a biological control agent against E. coli in irrigation water, contributing to improved agricultural practices and food safety measures.\n\nThe DNA of the bacteriophage was extracted following the SDS-proteinase K protocol described by Sambrook and Russell (2006). The extracted DNA was utilized to construct a genomic library with a 300-bp insert using the TruSeq DNA Nano kit (Illumina, USA), employing random fragmentation according to the manufacturer’s instructions. The quality of the libraries was assessed using a Qubit 2.0 fluorometer (Thermo Fisher Scientific) and the Bioanalyzer 2100 (Agilent Technologies, CA, USA). Subsequently, the libraries were diluted and sequenced on the Illumina MiSeq platform using the MiSeq v3 reagent kit, resulting in approximately five million paired-end reads of 150-bp.\n\nAssembly and bioinformatics analysis were performed in accordance with the recommendations by Philipson et al. (2018) and Turner et al. (2021). The Seqtk Toolkit was employed for random sampling of between 20,000 and 50,000 reads to achieve a coverage depth of approximately 100× post-assembly. The quality of the reads was evaluated with FastQC v0.74, while adapters and low-quality sequences (Phred index < 30) were discarded using the Trimmomatic v0.39 read preprocessing tool. De novo assembly of the reads was conducted using SPAdes v3.15.5, with k-mers set at 33, 55, 77, and 99.\n\nTo identify open reading frames (ORFs), the tools Glimmer v3.02, GeneMark v2.5, GeneMark.hmm v2.0, and GeneMarkS v 4.28 were utilized, and their functions were predicted through searches in the National Center for Biotechnology Information (NCBI) database. Additionally, automated genome annotation was carried out using PhageScope v1.3, Prodigal v1.0.1, and eggNOG-mapper v 2.1.12 platforms. Independent Rho factor transcription terminators and promoters were identified using the PhagePromoter v0.1.0, FindTerm, and ARNold servers, respectively. The presence of tRNA genes was evaluated using ARAGORN and tRNAscan-SE.\n\nTo characterize the lifestyle of the bacteriophage, the BACPHLIP, PhageAI, and PHACTS tools were employed. Potential genes associated with the synthesis of virulence factors were identified through the VFDB (www.mgc.ac.cn/VFs/search_VFs.html, accessed 6 November 2024) and VirulentPred (http://203.92.44.117/virulent/submit.html, accessed 6 November 2024) platforms. Finally, the presence of antibiotic resistance genes in the bacteriophage genome was analyzed using the CARD (https://card.mcmaster.ca, accessed 8 November 2024) and AMRFinderPlus (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA313047, accessed 8 November 2024) platforms.\n\nThis comprehensive approach to genomic sequencing and bioinformatics analysis facilitates a deeper understanding of the bacteriophage’s genetic characteristics, potential virulence factors, and antibiotic resistance profiles, contributing valuable insights for future applications in biocontrol and therapeutic strategies.\n\nTo assess bacteriophage stability across varying pH levels and storage temperatures, the data were first subjected to normality testing using the Shapiro-Wilk test and to homogeneity of variances using Levene’s test. A one-way ANOVA was conducted to evaluate pH stability, while a two-way repeated measures ANOVA was applied to analyze storage stability, incorporating both temperature and time as factors. Tukey’s post hoc test was used for pairwise comparisons between groups, with statistical significance set at p < 0.05.\n\nFor the biological control assay in compost, normality and homogeneity of variance were confirmed prior to conducting a one-way ANOVA to compare E. coli concentrations across the different treatments (absolute control, bacterial control, and phage treatment). Tukey’s test was applied for pairwise comparisons between treatments.\n\nIn the biological control efficacy test in irrigation water, normality and variance homogeneity were verified using the Shapiro-Wilk test and Levene’s test. A two-sample t-test was then employed to compare E. coli concentrations between treated and untreated samples after 24 hours. All statistical analyses were performed using Minitab® 18, and results are expressed as mean ± standard deviation.\n\n\nResults and discussion\n\nAmong the 17 compost samples derived from cattle manure and plant residues, the presence of Escherichia coli was identified in five samples using CHROMagar™ E. coli culture medium, with subsequent confirmation achieved through molecular identification via amplification of the phoA gene. Typically, the presence of mesophilic bacteria such as E. coli in compost piles is uncommon, as the temperatures during composting processes often exceed 50 °C, even under suboptimal conditions, limiting the survival of these bacteria (Thomas et al., 2020). However, some studies suggest that pathogenic bacteria can survive in the upper layers of compost piles, where lower temperatures prevail, or when ambient temperatures are lower (Werner et al., 2022; Zhang et al., 2019). In this study, sampling was conducted on the surface of static compost piles, which may have facilitated the presence of E. coli in the analyzed samples.\n\nIt is well-documented that E. coli is frequently isolated from bovine feces, posing a significant health risk, as certain strains have the potential to cause both intestinal and extraintestinal infections in humans (Martínez-Vázquez et al., 2021; Massé et al., 2021). Consequently, monitoring the presence of such pathogens in composts that utilize manure as a raw material is recommended. The presence of E. coli in compost used as organic fertilizer represents a potential public health risk, as it could serve as a vehicle for the introduction of this pathogen into the food production chain (Liu et al., 2021). Therefore, it is crucial to implement effective strategies to reduce the burden of pathogenic bacteria in such biofertilizers, thereby strengthening food safety frameworks and protecting consumer health.\n\nIn summary, the detection of E. coli in compost samples underscores the necessity for continuous surveillance and control measures to mitigate the risk associated with the application of manure-derived composts in agriculture. This is particularly relevant given the increasing interest in organic farming practices, where the use of compost as a soil amendment can inadvertently introduce pathogens into the food supply. Future research should focus on developing and optimizing composting processes that effectively eliminate pathogenic organisms while maintaining the beneficial properties of compost as a soil amendment.\n\nThe antimicrobial sensitivity of five E. coli strains isolated from compost samples was evaluated against a panel of antibiotics commonly used in clinical treatments and livestock production.\n\nResistance to ampicillin was observed in all strains except Eco-3, which showed intermediate susceptibility to this antibiotic. Despite this exception, Eco-3 was identified as multidrug resistant due to its resistance to multiple antibiotic classes, including aminoglycosides (gentamicin and amikacin), tetracyclines (tetracycline and oxytetracycline), and sulfonamides (trimethoprim/sulfamethoxazole). In addition, Eco-3 exhibited intermediate susceptibility to ciprofloxacin and cefoperazone, highlighting its complex resistance profile.\n\nStrains Eco-1 and Eco-4 displayed resistance to nalidixic acid, trimethoprim/sulfamethoxazole, gentamicin, and amikacin, further emphasizing a concerning trend of resistance to key therapeutic antibiotics. Eco-2 and Eco-5 exhibited resistance to chloramphenicol. Among all tested antibiotics, colistin exhibited antibacterial activity, effectively inhibiting all five strains, which highlights its critical role in combating resistant isolates.\n\nThe detection of resistance to tetracyclines, particularly in strain Eco-3, is alarming due to the widespread use of these antibiotics in livestock production. This finding suggests that the resistance patterns observed may be influenced by selective pressures associated with agricultural practices, where antibiotics are commonly employed for disease prevention and growth enhancement. The variability in resistance profiles among the studied strains underscores the potential role of compost as a reservoir for antibiotic-resistant bacteria, with significant implications for public health and agricultural sustainability.\n\nThese findings underscore the critical importance of rational antibiotic use in agricultural systems, consistent with the One Health approach, to limit the emergence and dissemination of multidrug resistant bacteria (Galarde-López et al., 2024; González-Aguilar et al., 2022). The resistance patterns documented in this study emphasize the broader impact of antibiotic use in livestock production, raising concerns about the potential transfer of resistant strains into the food chain and adjacent environments. Bacteriophage strategies may aid in reducing reliance on antibiotics in agriculture, thereby reducing the risks posed by antibiotic resistance in agricultural and clinical settings.\n\nSeven bacteriophages were isolated from livestock fecal samples. Among these, one was selected for detailed characterization based on its ability to produce the clearest and most well-defined lysis plaques, as well as its demonstrated broad-spectrum lytic activity against all five E. coli strains identified in this investigation. This phage was designated Alux-21, a name derived from the indigenous Mayan language, meaning “the protector”. The lysis plaques produced by Alux-21 are completely clear, measuring between 2.5 and 3.5 mm in diameter, with a translucent halo surrounding them. This type of lysis plaque is typically associated with efficient viral adsorption to the host bacterium, a short replication cycle, and a large burst size (Mangieri et al., 2020; Nascimento et al., 2022; Valencia-Toxqui & Ramsey, 2024). Furthermore, the presence of a translucent halo around the lysis plaques suggests that phage Alux-21 possesses the capability to degrade bacterial surface polysaccharides through the action of depolymerases (Vukotic et al., 2020). These enzymes enhance the lytic activity of phages, increasing their efficacy as biological control agents by facilitating the degradation of biofilms (Volozhantsev et al., 2020). Thus, phage Alux-21 may exhibit preliminary biological characteristics favorable for its potential use as a biological control agent against E. coli.\n\nThe replication dynamic of phage Alux-21 was evaluated using a one-step growth curve experiment. The results revealed an 89% decrease in the number of detectable viral particles within the first 5 minutes post-infection ( Figure 1), indicating that Alux-21 rapidly adsorbs to bacterial cells compared to other coliphages (Malik et al., 2021; Yao et al., 2023; Yazdi et al., 2020). The eclipse phase, corresponding to the period during which the phage replicates its DNA and assembles new viral particles within the host cell, lasted approximately eight minutes after adsorption. The latency phase, defined as the period from adsorption until the release of virions, lasted approximately 13 minutes. The burst size phase began around 20 minutes post-adsorption, resulting in the release of approximately 120 ± 5 virions.\n\nThe graph highlights key phases of the replication cycle, including a rapid 89% reduction in detectable viral particles within the first five minutes post-infection, indicating efficient adsorption. The eclipse phase concludes at eight minutes, followed by the burst phase at 20 minutes, releasing an average of 120 ± 5 virions per infected cell. The green line represents intracellular virion formation (chloroform-treated), while the black line indicates extracellular virions (untreated).\n\nThese characteristics are crucial for assessing the potential of phage Alux-21 as a biological control agent, as strictly lytic phages typically have substantially shorter latency times and higher adsorption rates, implying a greater capacity to infect and lyse bacterial cells within a reduced timeframe (Eriksen et al., 2023). Notably, the latency period and burst size are key parameters to consider when selecting phages with potential as biological control agents, as those with short latency periods and large burst sizes are believed to inactivate target bacteria more efficiently (Zhang et al., 2023). In this context, the replication characteristics of Alux-21 are favorable when compared to other phages with potential for controlling E. coli, such as vB_EcoM-Sa45lw and vB_EcoStr-FJ63A, which exhibit burst sizes of 80 and 11 virions per cell, and latency periods of 27 and 30 minutes, respectively (Liao et al., 2022; Xiao et al., 2023).\n\nThe replication cycle of Alux-21 aligns with that of other coliphages, characterized by short replication cycles and high intracellular replication efficiency, enhancing its bacteriolytic action on the host bacterium (Alexyuk et al., 2022). Additionally, the burst size of Alux-21 suggests a high capacity for disseminating virions to infect additional bacterial cells, a desirable characteristic in phages intended for the biological control of pathogenic bacteria (Mizuno et al., 2020; Nawaz et al., 2024).\n\nOne of the most critical criteria for selecting bacteriophages for the control of pathogenic bacteria is their stability during storage, as well as under specific application conditions (Duyvejonck et al., 2021). In this study, we assessed the stability of Alux-21 across different pH values to determine the optimal conditions for maintaining its viability. Furthermore, the results provide insights into the effects of various pH levels, representative of diverse agricultural environments, on bacteriophage concentration.\n\nThe findings indicate that phage Alux-21 exhibits the greatest stability at a neutral pH of 7, as a significantly higher viral concentration was observed compared to other pH levels ( Figure 2). This suggests that neutral conditions are most favorable for preserving the integrity and infectivity of Alux-21. This finding is consistent with previous studies, which have reported that various bacteriophages tend to maintain stability in conditions close to neutrality, a factor that is relevant for both storage and application in the biological control of pathogenic bacteria (Shahin et al., 2021; Xiao et al., 2022).\n\nThe figure illustrates the concentration of viable Alux-21 phage particles following a one-hour exposure to pH values of 3, 5, 7, 9, and 11. Error bars indicate the standard deviation of the measured concentrations, reflecting variability among experimental replicates.\n\nIn contrast, both acidic (pH 3) and alkaline (pH 11) conditions resulted in a notable reduction in bacteriophage concentration. These extreme pH levels are associated with adverse effects on viral structure, likely due to protein denaturation or capsid damage, thereby reducing the phage’s ability to effectively infect host bacteria (Majewska et al., 2023; Sada & Tessema, 2024). Thus, these extreme conditions could compromise the viability and functionality of Alux-21. It is important to note that the optimal pH of composts for maximizing their efficacy typically ranges around 7.5 (Pezzolla et al., 2021; Sayara et al., 2020). Meanwhile, the recommended pH range for agricultural irrigation water is between 6.5 and 7, ensuring suitable conditions for a wide diversity of crops (Guimarães et al., 2021). Therefore, this suggests that phage Alux-21 could maintain its viability in such environments without the need for additional formulations.\n\nThese results underscore the importance of maintaining bacteriophage suspension in conditions close to neutral pH to ensure stability during storage and application. Extreme pH values, such as 3 and 11, should be avoided, as they compromise phage viability, limiting its use in environments where prolonged activity is required. In summary, this assay establishes that a pH of 7 is the most suitable for prolonging the viability of phage Alux-21.\n\nOnce pH 7 was identified as the optimal condition for extending the viability of Alux-21, its stability during storage at temperatures of 4 °C and 40 °C was evaluated ( Figure 3). After six months, the concentration of phage Alux-21 stored at 4 °C showed no significant reduction, suggesting that cool storage is effective for maintaining its long-term viability. This result aligns with other studies reporting that low temperatures favor the preservation of the structural and functional integrity of bacteriophages, minimizing the degradation of their protein components (Kim et al., 2024; Xiao et al., 2022).\n\nThe bar plot illustrates the concentration of bacteriophage Alux-21 stored at two different temperatures, 4 °C (black bars) and 40 °C (green bars), monitored over a six-month period. At 4 °C, the phage concentration remained stable, with no significant reduction observed, indicating the phage’s long-term viability under refrigeration. Conversely, storage at 40 °C resulted in a moderate decline in concentration, corresponding to a decrease of approximately 0.5 log units, suggesting partial degradation under elevated temperature conditions. Error bars indicate the standard deviation of the measured concentrations, highlighting the variability among experimental replicates.\n\nIn contrast, the temperature of 40 °C was selected for evaluation because, in the northwest region of México where the phage was isolated, ambient temperatures can reach this level during the summer months. Assessing the stability of phage Alux-21 at 40 °C allows us to determine whether it can remain active without the need for additional stabilizers, potentially reducing costs and enhancing its applicability in agricultural systems that experience these environmental temperatures. Storage at 40 °C resulted in a moderate reduction in the concentration of Alux-21 after six months, with a decrease of only 0.5 log units. This behavior contrasts with findings for certain coliphages, which have shown drastic reductions in concentration at similar temperatures, often leading to complete inactivation within a matter of days (Kim et al., 2024).\n\nThe stability of Alux-21 at 40 °C represents a significant advantage, as many biological agents, including bacteriophages, typically require special formulations—such as microencapsulation or refrigerated storage conditions—to maintain their viability. These requirements can significantly increase production, transportation, and storage costs, thereby limiting their accessibility and potential application as biological control agents against pathogenic bacteria (Kering et al., 2020). The stability observed in Alux-21 at 40 °C could mitigate these costs, allowing for broader and more efficient use across diverse agricultural environments.\n\nThe bacteriolytic capacity of phage Alux-21 against E. coli under in vitro conditions was assessed, revealing that, compared to the control without phage inoculation, treatment with Alux-21 reduced the concentration of E. coli starting at two hours post-inoculation at a multiplicity of infection (MOI) of 0.001 ( Figure 4). This reduction was even more pronounced at an MOI of 0.01, and the bacteriolytic effect intensified from 90 minutes onward when applying an MOI of 0.1. At all assessed MOIs, bacterial concentrations declined to near-zero levels after three hours.\n\nThe figure depicts the lytic activity of bacteriophage Alux-21 against E. coli in tryptic soy broth (TSB). The red line represents the growth curve of E. coli in the absence of bacteriophage treatment, serving as the control. In contrast, the black, green, and blue lines illustrate the bacterial growth curves under treatment with Alux-21 at multiplicities of infection (MOI) of 0.1, 0.01, and 0.001, respectively. A significant reduction in bacterial growth is observed across all tested MOI levels, highlighting the remarkable efficacy of Alux-21 in suppressing E. coli proliferation, even at the lowest MOI.\n\nPrevious studies have documented that the reduction of bacterial populations induced by bacteriophages is dose-dependent, indicating that higher phage concentrations lead to greater decreases in bacterial counts (Abdelrahman et al., 2022). Notably, phage Alux-21 exhibited the capability to significantly diminish bacterial concentrations even at a low MOI of 0.001, contrasting with other coliphages that typically require an MOI of 1 or 10 to achieve similar reductions (Abdelrahman et al., 2022; Zhao & Meng, 2023).\n\nAdditionally, it is worth highlighting that in some coliphages, the use of low MOIs, such as 0.01, results in an initial decrease in bacterial concentration; however, resistant bacterial cells may begin to emerge after approximately six hours (Kuek et al., 2023). This phenomenon was not observed with Alux-21 during the same experimental timeframe, marking a positive characteristic of this phage as a biological control agent.\n\nThese findings underscore the potential of Alux-21 to rapidly reduce E. coli populations, which is promising for applications requiring swift and effective action against this pathogen. Particularly, the effect observed at an MOI of 0.001 within two hours post-inoculation is highly relevant in scenarios where bacteriophage concentrations may be limited due to environmental conditions.\n\nThe capacity of phage Alux-21 to reduce the concentration of E. coli in compost was evaluated. In the treatment without the application of bacteriophage, the bacterial concentration reached 7.8 × 106 CFU/g after 48 hours of incubation. This finding indicates that the characteristics of the compost used in this experiment create a favorable environment for the growth of E. coli under the evaluated experimental conditions ( Figure 5A). Previous studies have reported that compost can promote the survival and growth of E. coli, with increases exceeding four logarithmic units compared to the initial concentration, presenting a potential risk for the proliferation and dissemination of the pathogen in agricultural environments (Kim et al., 2009; Miller et al., 2013; Palmer et al., 2010).\n\n(A) Bar graph illustrating the concentration of Escherichia coli in compost during biological control assays. The black bar represents the bacterial concentration in untreated compost, showing an average value of 7.8 × 106 CFU/g. In contrast, the blue bar represents the E. coli concentration in compost treated with the Alux-21 bacteriophage, with an average value of 1.1 × 103 CFU/g. (B) Bar chart depicting the results of the biological control of E. coli in agricultural irrigation water. The black bar represents the concentration of E. coli in the untreated sample, which reached 1.8 × 106 CFU/ml. In contrast, the blue bar indicates samples treated with the Alux-21 bacteriophage, which exhibited a significant reduction in bacterial concentration, with a final count of 1.5 × 103 CFU/ml. Distinct letters above the bars denote statistically significant differences (p < 0.05). The variability in the data is represented by the standard deviation.\n\nIn contrast, the treatment in which E. coli suspension was inoculated followed by the application of phage Alux-21 demonstrated a significant reduction in bacterial concentration, registering a count of 1.1 × 103 CFU/g of compost. This represents a reduction of over 3.8 logarithmic units in bacterial concentration compared to the treatment without phage. These results align with findings from other studies that have documented the capacity of bacteriophages to control pathogenic bacteria in compost, achieving significant declines in bacterial concentrations in such organic substrates, thereby highlighting the potential of bacteriophages as biological control agents (Heringa et al., 2010; Ullah et al., 2017).\n\nThese results suggest that phage Alux-21 has potential for the biological control of E. coli in compost, achieving a significant reduction in bacterial load within a short incubation period. This is particularly relevant in organic agriculture, where the use of compost is common, and several outbreaks of E. coli have been associated with this type of biofertilizer (Chen et al., 2018; Islam et al., 2004; King et al., 2012; Sharma & Reynnells, 2016). The incorporation of phage Alux-21 in the final stage of composting could mitigate the risk of introducing E. coli into the organic food production chain, contributing to strengthening food safety schemes in sustainable agricultural production systems, as suggested by some authors for other phages (Otawa et al., 2012; Ullah et al., 2017). This aspect is particularly significant given the growing demand for organic products and the increasingly stringent requirements for compliance with food safety standards.\n\nHowever, it is important to emphasize that while the results regarding the capacity of Alux-21 to control E. coli in compost are promising, its viability in practical applications must be assessed under various environmental conditions. Factors such as pH, temperature, compost composition, and the presence of other microorganisms may affect the bacteriolytic activity of the phage (Fister et al., 2016; Heringa et al., 2010; Wang et al., 2020). Future studies should focus on evaluating the effectiveness of Alux-21 under these variables. Additionally, the emergence of phage-resistant strains is a critical aspect that could limit the long-term efficacy of phage Alux-21. To mitigate this risk, it is advisable to continue the isolation and characterization of additional bacteriophages, with the aim of developing a phage cocktail to reduce phage resistance.\n\nThe evaluation of phage Alux-21 for controlling E. coli in irrigation water samples yielded promising results. Following the incubation period, the concentration of E. coli in the treatment without phage reached 1.8 × 106 CFU/ml, suggesting that the conditions of the irrigation water used in the trial facilitate moderate growth of E. coli ( Figure 5B). In contrast, samples treated with Alux-21 exhibited a significant reduction in bacterial concentration, with a final count of 1.5 × 103 CFU/ml. This represents a reduction of over three logarithmic units in bacterial load compared to the control.\n\nThese findings indicate that Alux-21 has potential as a biological control agent for reducing E. coli concentrations in agricultural irrigation water. This could represent a future strategy to improve the microbiological quality of irrigation water, as has been suggested for other bacteriophages (Álvarez et al., 2019; Soliman et al., 2023). This is particularly relevant in the organic food production sector, where the application of chemical treatments is restricted, and biological control offers an ecologically sustainable alternative. Moreover, in many agricultural regions experiencing prolonged droughts, sustainable practices are essential for conservation, and the use of bacteriophages could be efficiently integrated into these systems.\n\nHowever, despite these encouraging results, further research is necessary to evaluate the stability of Alux-21 under the varying physicochemical conditions that may be encountered in irrigation water. Additionally, it is crucial to conduct evaluations under field conditions to ensure the viability and effectiveness of this approach in real agricultural production scenarios, thereby confirming its applicability.\n\nThe genomic analysis revealed that phage Alux-21 possesses a double-stranded DNA genome with a linear topology, spanning 42,729 base pairs (bp) and exhibiting a GC content of 54.31%. The genome includes 60 open reading frames (ORFs), organized into functional modules responsible for DNA processing, host cell lysis, genetic material packaging, and morphogenesis ( Figure 6).\n\nThe figure illustrates the entire genome map of bacteriophage Alux-21, with individual genes depicted as arrows to indicate their respective locations and transcriptional orientations. Each gene is color-coded according to its predicted function, enabling a clear distinction between structural (blue), replication-associated (yellow), DNA packaging (green), lysis (red) and hypothetical protein (gray) and auxiliary genes. The analysis revealed no evidence of genes encoding virulence-associated factors, lysogeny-related mechanisms, or proteins with allergenic potential. This genomic characterization underscores the suitability of Alux-21 for applications in biocontrol, as the absence of such undesirable elements minimizes potential risks associated with its application.\n\nThe DNA replication module comprises ORFs encoding critical proteins for genome replication and transcription, including a single-stranded DNA-binding protein (ORF 1), a DksA-like zinc-finger protein (ORF 2), a MerR family transcription regulator (ORF 4), DNA helicases (ORFs 5 and 8), an exonuclease (ORF 7), holliday junction resolvases (ORF 11 and ORF 12), DNA primase (ORF 19), and a late transcriptional activator (ORF 22). These proteins play critical roles in the replication and transcription of the Alux-21 genome, ensuring efficient viral propagation within the host.\n\nIn terms of host cell lysis, the genome of Alux-21 encodes the proteins holins (ORFs 23 and 24), lysozyme (ORF 25), and an Rz-like spanin (ORF 26). Together, these proteins facilitate host cell membrane pore formation, cell wall degradation, and the subsequent release of virions. This lytic gene cluster is critical for the phage’s lytic life cycle, allowing the destruction of the bacterial host at the end of viral replication.\n\nFor the packaging of genetic material, ORFs were identified that encode the terminase small subunit (ORF 28), terminase large subunit (ORF 29), and portal protein (ORF 31), all of which are essential for encapsulating viral DNA within the capsid during assembly. Additionally, the morphogenesis-related ORFs encode structural proteins essential for capsid formation (ORFs 32, 35, 42, and 48) and tail formation (ORFs 36, 37, 41, 44, 45, 46, 47, 49, 51, 52, and 54), as well as components of the baseplate assembly (ORFs 38, 39, 40, 43, and 53). These structural components are indispensable for virion assembly and phage infectivity. The genome sequence of Alux-21 has been comprehensively annotated and archived in the NCBI GenBank database under accession number PQ589898. Additionally, the sequencing data are available in the Sequence Read Archive (SRA) under project ID PRJNA1191041.\n\nNotably, the genomic analysis did not detect any genetic elements associated with virulence factors, antibiotic resistance, host genome integration, or allergens. The absence of these elements is strongly recommended for phages intended for biological control and/or therapeutic applications (Garvey, 2022; Li et al., 2020). These findings indicate that Alux-21 is a strictly lytic phage, devoid of proteins that could pose potential biosecurity risks, positioning it as a safe candidate for biocontrol applications from a genomic standpoint.\n\n\nConclusion\n\nThis study demonstrated that the isolated bacteriophage, designated as Alux-21, possesses substantial potential as a biological control agent against E. coli. The findings indicate that this phage exhibits significant lytic activity not only under laboratory culture conditions but also in compost and agricultural irrigation water, effectively reducing E. coli concentrations. This pathogen is of major concern for public health due to its high morbidity and mortality rates and its significant antimicrobial resistance.\n\nMoreover, phage Alux-21 maintains stability across a range of pH levels and remains viable for extended periods at both 4 °C and 40 °C, underscoring its suitability for agricultural applications without the need for protective formulations. Additionally, the genome of Alux-21 lacks genes associated with virulence factors, antimicrobial resistance, lysogeny, or proteins with potential allergenic properties, characteristics that make it ideal for phages intended for biological control applications.\n\nThese findings position Alux-21 as a viable, sustainable solution for reducing E. coli contamination in agricultural systems, particularly in compost and irrigation water, where chemical treatments are impractical or restricted. Field trials and further validation will be crucial to advancing its adoption in real-world scenarios. This progress could have significant implications for enhancing food safety and public health. However, while the results obtained under experimental conditions are promising, it is essential to progress towards field studies that evaluate its efficacy in real-world agricultural settings and assess its interactions with other microorganisms present in these ecosystems. Future research will be imperative to validate its application as a robust and sustainable biological control solution.\n\nEthical approval and consent were not required.\n\n\nAuthor contributions\n\nLuis Amarillas (LA), Mitzi Estrada-Acosta (ME-A), and Luis Lightbourn-Rojas (LL-R) were responsible for the conceptualization of the study, the methodological development, and data analysis. LL-R secured the necessary funding for this research, while LA, ME-A, Ruben León-Chan (RL-C), Jorge Padilla (JP) and Enrique López-Avendaño (EL-A) performed the experiments and data collection. Research planning and execution were overseen and coordinated by ME-A, Antonio González-Balcázar (AG-B) and LL-R, with LA, RL-C, and AG-B ensuring the reproducibility and accuracy of the results. LA was responsible for visualization preparation, and LA, R-LC, JP, EL-A, AG-B, ME-A, and LL-R collaborated on drafting and revising the manuscript. All authors reviewed and approved the final manuscript and assume full responsibility for the veracity and integrity of the work.", "appendix": "Data availability\n\nGenBank of NCBI: Sequence Read Archive (SRA) of raw reads from the sequencing of the Alux-21 phage genome. Access number PRJNA1191041; https://www.ncbi.nlm.nih.gov/sra/PRJNA1191041 (Amarillas et al., 2023).\n\nZenodo: Supplemental Files, https://zenodo.org/records/14233172, 10.5281/zenodo.14233172 (Amarillas et al., 2023).\n\nUnder the license CC BY 4.0 Attribution 4.0 International\n\nThis project contains the following extended data:\n\nSupplemental Table 1: This table provides a comprehensive summary of the open reading frames (ORFs) identified in the genome of the bacteriophage. It includes detailed information on their genomic positions, predicted coding regions, and the putative functions of the encoded proteins, as inferred from annotation analyses.\n\nSupplemental Table 2: This table presents the experimental results of the one-step growth curve analysis of the bacteriophage, as well as its stability under varying pH conditions and during storage. Additionally, it includes data on the phage’s efficacy as a biocontrol agent against E. coli in compost and irrigation water samples.\n\n\nAcknowledgements\n\nWe extend our sincere gratitude to Fedra Padilla-Lafarga, Jesús Antonio Núñez, and Zendy Mariela Martínez for their invaluable technical support and contributions throughout the experimental phases of this study. 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PubMed Abstract \| Publisher Full Text\n\nTopka G, Bloch S, Nejman-Falenczyk B, et al.: Characterization of bacteriophage vB-EcoS-95, isolated from urban sewage and revealing extremely rapid lytic development. Front. Microbiol. 2019; 9(JAN). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTurner D, Adriaenssens EM, Tolstoy I, et al.: Phage annotation guide: guidelines for assembly and high-quality annotation. Phage (New Rochelle). 2021; 2(4): 170–182. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nValencia-Toxqui G, Ramsey J: How to introduce a new bacteriophage on the block: a short guide to phage classification. J. Virol. 2024; 98(10): e0182123. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVolozhantsev NV, Shpirt AM, Borzilov AI, et al.: Characterization and therapeutic potential of bacteriophage-encoded polysaccharide depolymerases with β galactosidase activity against Klebsiella pneumoniae K57 capsular type. Antibiotics. 2020; 9(11): 1–16. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVukotic G, Obradovic M, Novovic K, et al.: Characterization, antibiofilm, and depolymerizing activity of two phages active on carbapenem-resistant Acinetobacter baumannii. Front. Med. 2020; 7. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWaltenburg MA, Schwensohn C, Madad A, et al.: Two multistate outbreaks of a reoccurring Shiga toxin-producing Escherichia coli strain associated with romaine lettuce: USA, 2018-2019. Epidemiol. Infect. 2022; 150: e16. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWang H, Bridges W, Chen Z, et al.: Comparing and modeling the thermal inactivation of bacteriophages as pathogenic viruses surrogates in chicken litter compost. Compost. Sci. Util. 2020; 28(2): 87–99. Publisher Full Text\n\nWerner KA, Poehlein A, Schneider D, et al.: Thermophilic composting of human feces: development of bacterial community composition and antimicrobial resistance gene pool. Front. Microbiol. 2022; 13. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nXiao T, Zhu X, Wang W, et al.: A novel lytic bacteriophage against colistin-resistant Escherichia coli isolated from different animals. Virus Res. 2023; 329: 199090. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nXiao Y, Huang P, Huang Z, et al.: Influencing factors on the preservation of lytic bacteriophage VP3. Biosafety Health. 2022; 4(5): 314–320. Publisher Full Text\n\nYao L, Bao Y, Hu J, et al.: A lytic phage to control multidrug-resistant avian pathogenic Escherichia coli (APEC) infection. Front. Cell. Infect. Microbiol. 2023; 13. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYazdi M, Bouzari M, Ghaemi EA, et al.: Isolation, characterization and genomic analysis of a novel bacteriophage VB_EcoS-Golestan infecting multidrug-resistant Escherichia coli isolated from urinary tract infection. Sci. Rep. 2020; 10(1): 7690. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYin HB, Gupta N, Chen CH, et al.: Persistence of Escherichia coli O157:H12 and Escherichia coli K12 as non-pathogenic surrogates for O157:H7 on lettuce cultivars irrigated with secondary-treated wastewater and roof-collected rain water in the field. Front. Sustain. Food Syst. 2020; 4: 555459. Publisher Full Text\n\nZhang Q, Liu J, Guo H, et al.: Characteristics and optimization of dairy manure composting for reuse as a dairy mattress in areas with large temperature differences. J. Clean. Prod. 2019; 232: 1053–1061. Publisher Full Text\n\nZhang YS, Yuan L, Mgomi FC, et al.: Characterization and comparative genomic analysis of novel lytic bacteriophages targeting Cronobacter sakazakii. Virus Res. 2023; 329: 199102. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZhao P, Meng X: Characterization of antibacterial activity of lytic bacteriophage PE-1 for biological control of Escherichia coli K88 in vitro. Food Sci. Anim. Product. 2023; 1(2): 9240014. Publisher Full Text" }	[ { "id": "350395", "date": "21 Jan 2025", "name": "Sihem Jebri", "expertise": [ "Reviewer Expertise Environmental and food 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\nAfter a thorough examination of the manuscript “Isolation and characterization of a novel bacteriophage as a biological control agent against multidrug-resistant Escherichia coli in compost and agricultural irrigation water”, I commend the authors for their meticulous research and well-structured study. The isolation and characterization of phage Alux-21 as a potential biocontrol agent against Escherichia coli in agricultural settings represent a significant contribution to combating antibiotic-resistant pathogens. The study is comprehensive, with well-designed experiments, appropriate methodologies, and clear discussions.\nHowever, I suggest the following minor revisions to further improve the manuscript:\n\nEnhance the flow between sections in the introduction. Currently, some transitions feel abrupt, and smoother connections would better guide the reader. Expand on the importance of bacteriophage isolation and characterization, particularly in the context of addressing antibiotic-resistant pathogens. This will underscore the relevance of the study and situate it within the broader scientific context. Throughout the manuscript (not limited to the introduction), break up long sentences to improve readability. Shorter, more concise sentences will make the text more digestible and engaging. Clarify the methodology used for bacteriophage isolation. The spot test was employed rather than the double-layer agar (DLA) technique. In the DLA method, the soft agar contains both the phage-containing sample and the host bacteria, which are then overlaid on a solid agar layer. In this study, the semi-solid agar contained only the host bacteria, with the phage-containing samples applied as spots. This distinction should be emphasized to avoid confusion. Include the morphological characterization of the isolated phage using Transmission Electron Microscopy (TEM). This would add depth to the study and provide crucial information on the phage’s structural features. Provide images or examples of clear plaques formed during phage isolation. These visuals would support the experimental findings and offer a better understanding of the isolation process. Investigate and report the polyvalence of phage Alux-21. Testing its efficacy against both pathogenic and non-pathogenic bacteria would enhance the study’s scope and demonstrate its potential as a versatile biocontrol agent.\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": "355700", "date": "27 Jan 2025", "name": "Mohamed El-Telbany", "expertise": [ "Reviewer Expertise Phage therapy and application. Antimicrobial resistance." ], "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\nComments:\n1. E. coli strains were isolated from compost. Did the authors isolate E. coli O157:H 7? Usually, most bacterial strains can be destroyed at the composting temperature (about 60 °C (140 °F); except E. coli O157:H 7 needs a higher temperature. In that case, what is the purpose of using a phage as a biocontrol agent in compost?\n2. The lytic activity of the phage against its host was carried out only for 5 h. What about the long incubation time? For example, what about 24, 48, and 72 h?\n3. There is no information about the phage host range against E. coli and other species. Did the authors do this experiment? Moreover, productivity and reproducibility of the phage should be done and discussed (EOP test)\n4. The authors claimed that the phage is novel. What is the novelty of this phage? and of this study?\n\n5. The phage genome sequence was not well discussed. This phage should be aligned with other related phages, and a bioinformatic analysis of its genome should be performed and added to 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? 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? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [] } ]	1	https://f1000research.com/articles/13-1521
https://f1000research.com/articles/13-1520/v1	16 Dec 24	{ "type": "Research Article", "title": "Exploring barriers to long-acting reversible contraceptive utilization among the pastoralist women in southern Ethiopia: Qualitative study approach, 2024", "authors": [ "Dawit Gelgelo", "Dejene Hailu", "Sileshi Garoma", "Dejene Hailu", "Sileshi Garoma" ], "abstract": "Background Ensuring comprehensive access to family planning improves gender equality, empowers women, and, eventually, reduces poverty. However, in the pastoralist community, women were rooted in strong cultural and religious perspectives that promoted many children, discouraged the use of FP, had high male dominance, and had limited control over their life, including the use of FP.\n\nObjective To explore the barriers to LARC utilization among pastoralist women in southern Ethiopia.\n\nMethods This study used a qualitative descriptive study design and included thirty-three pastoralist women for focus group discussions and twelve key informants for IDI, who were purposefully selected from three pastoral districts. The participants were interviewed and explored the primary barriers to using LARCs. The principal researcher led the interview via a semi structured interview guide that allowed for flexibility in exploring issues aided by audio tape. The approach included a broad inductive thematic analysis, with significant concepts derived from transcripts.\n\nResults In this study, there are many barriers to LARC utilization organized into five themes, including myths and misconception (perceived side effects, FP is not suitable for women who work intensively and carry heavy objects and travel long distances), the desire to have many children (children are wealth and Gods gifts), religious belief, distances of health facilities and husband objection.\n\nConclusion and recommendations Information education communication should be provided to address myths and misconceptions about the LARC during the FP intervention for pastoralist women via HEW, HCP, media, and community education. There is a need to enhance social opportunities by involving religious leaders and partners in decision-making and providing health information and counseling to religious leaders and couples in the community through HEW. As myths and misconceptions about LARC use emanate from a variety of concerns, this study may not cover all of them; therefore, future research is needed to investigate additional myths and misconceptions concerning contraception use.", "keywords": [ "Long-acting reversible contraceptive", "pastoralist women", "myth and misconception", "southern Ethiopia." ], "content": "Introduction\n\nFamily planning is defined as the choice of individuals or couples to anticipate and attain their desired number of children and the spacing and timing of their births.1 Ensuring universal access to family planning has a positive effect on gender equality, women’s empowerment, and, ultimately, poverty reduction.2,3 Women and children are given particular attention in the health policy of Ethiopia.2,4 Accordingly, the ministry is engaged in a range of efforts to improve the access to and quality of modern family planning services. As a result, the use of modern contraception increased from 6.3% in 2000 to 35% in 2016.5\n\nLong-acting reversible contraception (LARC), which includes IUCDs and subdermal implants, has many desirable attributes, such as highly effective protection against unwanted pregnancies and few contraindications.6 They do not require the users’ ongoing effort for long-term and effective use following initial insertion, are cost-effective, do not require frequent visits for resupply, and are reversible with a rapid return to fertility after removal.7,8 However, the proportion of women currently using long-acting contraceptives remains significantly lower than that of women using short-acting methods.9 The Ethiopian demographic health survey (EDHS) 2016 reported that only 7.1% of women utilized the LARC, which was far from the global FP target of 2030.10,11\n\nVarious studies conducted in Ethiopia have shown that the utilization of long-acting contraceptives is influenced by social, cultural, economic, and health system-related factors. In particular, women’s social status, access to education, and involvement in decision making on reproductive matters and social norms were found to be the major determinants of long-acting contraceptive use.2,12 Furthermore, women in the pastoralist community are double marginalized as women and pastoralists.13 In the pastoralist community, women feel that they are embedded in strong cultural and religious perspectives, which promote a high number of children, discourage the utilization of FP, increase male dominance and restrict the ability of women to control their lives, including FP use.11 In addition, a multilevel and spatial analysis of geographic variation and associated factors of long-acting contraceptive use among reproductive-age women in Ethiopia revealed that living in a pastoralist community reduced the odds of LARC utilization in Ethiopia.14 Therefore, this study aimed to explore the barriers to LARC utilization among pastoralist women in southern Ethiopia. Understanding the barriers to long-acting reversible contraception use is important for dispelling them and increasing the uptake of contraception.\n\n\nMethods\n\nA qualitative descriptive study design with a qualitative research approach was employed for this study. The study employed thirty-three pastoralist women aged (15–49 years) who were purposefully sampled from three pastoral districts for focus group discussion. Only married women were included. Four family planning coordinators, four religious leaders and four community figures were included in the key informant interviews.\n\nThe interviews were conducted by health workers with experience in qualitative research methods. The number of FGDs was determined on the basis of the principles of saturation of information. Every member of the group contributed to any question asked before proceeding to another question. Married discussants representing different backgrounds in terms of gender, profession, education, and social status were included in the study. The IDIs were performed among well-known people in the community and included community figures, religious leaders and healthcare providers. The participants were interviewed, and their opinions about LARC and key barriers to its use were discussed. A semi structured interview aided by audio tape was conducted. The primary researcher led the interviews following a semi structured interview guide developed from the literature review15–17 with the flexibility to explore topics as they arose. One facilitator facilitated the interview process. The interviews began with thirteen interview guide questions designed to explore participants’ opinions, followed by broader discussion among the group and suggestions. The interviews were audio recorded and transcribed verbatim. Data from the field notes were reviewed and verified by the participants. When transcribing, square brackets were used to denote nonverbal activities, and names or places were removed and replaced with non-identifying letters. The discussion was performed in the Afan Oromo language as preferred by the discussants and later translated into English during the transcript, with each FGD lasting for 60–90 minutes and the IDI lasting for 30–45 minutes.\n\nTo determine the accuracy and reliability of the data, the criteria of credibility, transferability, dependability, and confirmability were considered.15\n\nThe interview transcripts and field notes were analyzed through principles of inductive thematic analysis using systematic approach. The data collected from the focus groups and interviews were transcribed verbatim. First, the principal investigator transcribed verbatim, generated codes and themes, and then reviewed, interpreted, and described these codes and themes.17 Other members of the research team analyzed the codes and themes, making adjustments and consolidating them as needed. The analytical process worked toward condensing the ideas to the point where key ideas were defined, described here as themes. Data analysis began while further interviews were being conducted; recruitment ceased when thematic saturation was reached. An independent researcher fluent in both Oromic and English reviewed the transcripts for accuracy and original meaning preservation during translation.\n\nEthical clearance was received from the Institutional Review Board (IRB) of Bule Hora University on March- 28/2024 with protocol number of 028/24/IHSGS. The study was conducted from March 30 to April 25, 2024. Married couples were notified of the purpose of the study as it attempt to improve knowledge and attitudes about family planning and contraception methods. This study obtained both verbal and written informed consent, which was approved by the Bule Hora University IRB committee. As a result, each study participant who could not read or write provided verbal consent, while literate participants provided written informed consent before to participating in the study.\n\n\nResult\n\nIn this study, the researcher enrolled a total of 46 participants: 33 reproductive age groups for the FGD and 12 key informants for the IDI (4 family planning coordinators, 4 religious leaders and 4 community figures). The mean age of the study participants was 29.5 years (range 18–47 years) ( Table 1).\n\nEmerged themes\n\nFive themes emerged from the analysis of the client’s focus group discussion and key informant interview data on barriers to long-acting reversible contraception utilization in pastoral communities.\n\nTheme 1: Perceived myths and misconceptions\n\nSubtheme 1.1: Perceived side effects (cancer, infertility, heavy and sudden bleeding, overweight, tininess, headache and stress): Some women never use contraception methods because of concerns about side effects. However, other women utilized FPs but discontinued them because of side effects. Almost all women reported bleeding as a major side effect of LARC. Some had personal encounters with adverse effects, whereas others were concerned with a rumor. When women were asked about the side effects of using LARCs, they stated the following:\n\nI have never used contraception because of what others have said about it. My husband encouraged me to use FP. However, others claim that it causes excessive bleeding. It makes women extremely thin. (Women participant 4, FGD)\n\nI have never used FP because of what others have said about it. They all focus on adverse effects. They claimed that this caused massive bleeding. (Female participant 5, FGD)\n\nOn the other hand, some women experienced adverse effects and changed methods. Many women preferred long-acting reversible family planning over short-acting methods. When asked which methods are most acceptable to their bodies, two participants responded as follows:\n\n… I’m familiar with both FPs, which serve for three months and three years. However, the one that serves for three years is preferable. Because it is more suitable for women’s bodies. (Women participant 15, FGD)\n\n……I have used both three months and three years of FP. The one placed into the arm is better than the others. (Female participants 8, FGD)\n\nFurthermore, some women noted that the adverse effects differ from one woman to another. They described their opinion as follows.\n\n…. When one FP is not suitable for their bodies, they should seek other methods. Therefore, instead of just ignoring the FP, they should choose the one that is most suitable for them. (Women participant 1, FGD)\n\n…. FP side effects differ from women to women. For one FP that serves for three months, including myself, is suitable, but for others, the FP that serves for three years is suitable. (Female participant 9, FGD)\n\nSome participants associated LARC use with cancer and infertility. One woman was diagnosed with cancer after receiving LARC. The health professional did not inform her that the cancer was caused by the use of contraception. However, she had directly linked the cancer to the use of contraception. Another religious leader revealed his wife’s experiences with LARC use. He stated that his wife was unable to conceive again following the removal of the LARC. They expressed their viewpoint as follows:\n\n…. I used FPs three years after my second birth. However, it causes me to bleed heavily, and my family takes me to a medical facility, where they inform me that I have cancer. Because I was previously healthy, FP was the cause of my cancer. Personally, I have related my illness to FP. (Female Participant 6, FGD)\n\n……My wife used FP for three years. She suffered from severe bleeding, headaches, body aches, stress, and weakness following FP use. Meanwhile, after removal of the FP, she was unable to conceive again. I think this is related to FP…. (Religious leader Participant 11, KII)\n\nSubtheme 1.2: FP is not appropriate for women who travel long distances: The women asserted that pastoral women travel long distances to find water and collect firewood. When asked about the barriers to FP use, particularly for pastoral women, two expressed their thoughts as follows:\n\n…. I want to space my children. However, I’m frightened to use FP. Because I walk a long way to the market, collect firewood, and draw water from the river…. Therefore, FP is unsuitable for me. (Female participant 6, FGD)\n\nFP is unsuitable for pastoral women. Women are not thrilled about using it. FP bleeding differs from typical menstrual bleeding. It happened unexpectedly while one was on the way to the market or somewhere. Additionally, our lives are built on migrating from one pace to another. Therefore, it is not suitable for us. (Women participant 2, FGD)\n\nSubtheme 1.3: FP is not suitable for women who work intensively and carry heavy objects: The women indicated that pastoral women carry heavy objects, such as grain, wood, and water, on their backs. Bleeding occurs suddenly during outdoor activities. When asked about the obstacles to FP use, particularly for pastoral women, the participants expressed their opinions as follows:\n\n……However, in our case, using FP is unsafe. Because we are pastoralist women. Our regular activities include lifting massive amounts of objects such as wood, water, and grain. Therefore, the FP is not perfect for pastoral women. (Community figure participant 8, KII)\n\nTheme 2. Access issues: Some respondents highlighted the distance between health facilities and the need for frequent contact to receive services as barriers to FP utilization. When asked about pastoral women’s obstacles in accessing FP health services:\n\nPastoral women are unable to visit health facilities frequently due to distance, especially those who have served for three months, who require frequent contact. For this reason, FPs that serve for three years are preferable for pastoral women. (Women participant 7, FGD)\n\nTheme 3: Desire to have many children\n\nSubtheme 3.1: Children are wealth: Many participants believe that having children is a sign of wealth. They believe that a family with many children might gain respect from the neighborhood and even provoke fear toward that family. This intention makes it possible for them to desire to have many children. They stated their opinion as follows:\n\n…. Having lots of kids is a good thing. According to Guji, those without clothing and family members are despised. Therefore, having a large family is a wonderful way to earn respect from others. However, having children with a plan is preferable. (Community figure participant 44, FGD)\n\n… Indeed, children are a blessing and are brought up by God. I desire a large family, so I do not care if living expenses rise. The resources needed to raise kids will come from God. (Women participant 25, FGD)\n\nAnother participant argued that children are not riches, which is opposed to the aforementioned notion. According to them, a person should have children in accordance with his or her standards of living and be in charge of the children’s clothing, food, and education. The following is how these participants expressed their opinions:\n\n… I do not believe in the concept of children as wealth. Having children can be helpful depends on one’s standard of living. Because any one knows his standard of living. (Women participant 18, FGD)\n\n….People say that children are rich. Therefore, they want to have more children. However, this is not good. God gave us obligations. He is not coming to support children’s food, education, or clothing. This is our responsibility.… (Women participant 11, FGD)\n\nSubtheme 3.2: Children are gifts of God: The participants strongly believe that children are God’s gifts. Some participants stated that while children are God’s gifts, they should be planned. Others, however, believe that preventing God’s gifts through the use of medications such as FP is sinful. They expressed their suggestions as follows:\n\nIn our community, children are considered God’s gifts; therefore, they just give birth. I think that children are God’s gifts, but it is the family’s obligation to raise them. God, do not come and work on your farm. He does not come to clothe the children. (Female participants 31, FGD)\n\n…. As the gospel states, children are God’s gifts. However, God gave us responsibility. He is not coming to support children’s food, education, or clothing. This is our responsibility. Therefore, we should have children with prudence. (Women participant 2, FGD)\n\nTheme 4: Religious belief: Religion was one of the sociocultural issues identified as barriers to LARC utilization by participants from various perspectives. Some participants believe that the Bible does not oppose FP, whereas others believe that it does not support FP. When asked about sociocultural barriers to LARC utilization, several participants expressed their views as follows:\n\n…. The Bible does not contradict the FP. Our God is the God of the Plan. First, every creature was created according to the plan. Similarly, we should plan for every aspect of our lives, including our children. (Religious leader participant 10, KII)\n\nI do not think the Bible supports FP. The Bible encourages people to multiply and fulfill the land. However, considering the state of people’s economies, it is impossible to prevent them from using it. Personally, I firmly oppose the use of FPs for abortion, whether temporary or permanent. However, the second option is up to your faith. People could use reversible FPs for child spacing on the basis of their lifestyle standards. (Religious leader participant 11, KII)\n\nThe other issue I have encountered as a health professional with FP is that some women consider using it for sin. They have stated that women giving birth through surgery is God’s punishment. Because they have used FP. Personally, I know women who came to me and stated they wanted the implanon removed because it was against their religion. (MCH coordinator participant 5, KII)\n\nTheme 5: Husband objection. Husband approval is a crucial factor in women’s FP usage. Some husbands encourage their wives to use FP, whereas others oppose it. When the women were asked about their spouses’ support for FP utilization, they expressed their opinions as follows:\n\nI have five children. After my fifth child, I used an implant. At the time, I kept it hidden from my husband. Because he knows he will not let me use it. After two and a half years, I removed it and informed my husband. Thankfully, he did not say anything. (Women participant 2, FGD)\n\nMy husband does not support my choice to utilize FP. He claims that it is his obligation to raise children. It is not my business. He said that even if I have 30 children, he will raise them. (Women participant 23, FGD)\n\nThe data has been coded and categorized into five main themes emerged from the analysis of the client’s focus group discussion and key informant interview data on barriers to long-acting reversible contraception utilization in pastoral communities: Perceived myth and misconception, distance of health facility, desire to have many children, religious belief and husband’s objection ( Table 2).\n\n\nDiscussion\n\nA descriptive qualitative study was conducted to explore the barriers to long-acting reversible contraceptive utilization among the pastoral community in southern Ethiopia. The study identified barriers to long-acting reversible contraceptive utilization in five organized themes: perceived side effects, perceived misconception, distance of health facilities, desire to have many children, religious belief and husband’s objection.\n\nAccording to the findings from this study, perceived adverse effects seem to be one of the barriers to LARC use. The perceived negative effects of LARC include cancer, infertility, severe bleeding, headaches, and stress. Women either experienced these adverse effects or heard about them. In this study, bleeding was the most commonly reported adverse effect among LARC users. This is especially difficult for pastoralist women who have a mobile lifestyle. They may have to walk long distances to obtain water, gather firewood, and go to the market. Furthermore, they cannot buy sanitary towels because they rely primarily on their husband’s income. According to the American Sexual Health Association, some people using a hormonal IUD may experience irregular bleeding, such as spotting between periods or heavier bleeding during a period. After a few months, bleeding may become lighter, menstrual cramps may decrease, and periods may disappear entirely.16,18\n\nAnother major concern highlighted in this study is whether LARC causes cancer. The women believed that the absence of menstrual blood due to conception would accumulate unclean blood inside them, causing various types of cancer. According to the American College of Obstetricians and Gynecologists, there is insufficient evidence to know if the LARC increases the risk of cancer.19 However, the IUCD is not indicated for anyone who has had gynecological cancer, such as cervical or uterine cancer.20 One of the studies conducted on current hormonal contraception and the risk of endometrial cancer suggested that implants have a protective effect on the risk of ovarian cancer among women with breast cancer gene mutations.21\n\nAnother concern addressed in this study was that participants linked infertility to contraceptive use. Some FGD discussants and KIIs noted some women’s inability to conceive again following LARC removal. The American College of Obstetricians and Gynecologists states that LARCs do not induce infertility or make it more difficult to conceive in the future.19 Previously, there was worry that IUCD use could cause infertility due to an increased risk of sexually transmitted infections. While untreated STIs can cause pelvic infection, preventing some women from becoming pregnant, many studies have demonstrated that IUCDs do not increase STI infection rates or lead to infertility.19,22\n\nThe study’s findings revealed that perceived misconceptions are among the most significant barriers to LARC utilization. Many women believe that pastoral lifestyles are unsuitable for LARC use. They generally believe that LARC is not allowed for those who engage in different daily activities. LARC has several contraindications, including pregnancy, current pelvic inflammatory disease, undiscovered irregular uterine bleeding, genital tract malignancy, and a history of specific cancers, such as breast cancer.21–23 However, the idea that LARC use impedes daily life activities and is inappropriate for pastoralist women has no scientific basis.18,22,23\n\nOther challenges to LARC utilization among pastoralist women include a desire to have many children. Everyone, especially women, instinctively wants a large family and feels fulfilled by having many children, but the desire for children varies greatly between individuals, and many people may choose to have a smaller family or no children at all owing to personal preferences, life circumstances, and individual priorities. Similarly, some studies have found that the desire to have many children is a barrier to LARC utilization.17,24,25 This study revealed that religious belief is one of the barriers to LARC use. In this study, religious leaders held different opinions on the use of LARCs. One religious leader supports the use of LARCs, whereas others condemn it. This finding is consistent with those of studies on barriers to contemporary contraception use in Ethiopia26 and Uganda.27 In general, the implications of this study’s findings for policy and practice are to enhance interventions that encourage the use of the LARC, which has an important role in the use of the LARC among pastoralist women. Understanding the local context of pastoral women’s issues can help to better target interventions.\n\n\nStrength and limitations\n\nThe current research provides insight into the barriers to LARC utilization among pastoralist women. The researcher used purposive sampling techniques to recruit participants from the community with the help of a health extension worker. The data triangulation approach, which included FGD and KII, was used. However, this study included only a small sample of women from three pastoralist districts in southern Ethiopia, which may not represent the entire pastoral region of Ethiopia. Furthermore, because the interviewers were health care workers, there is a possibility of social desirability bias.\n\n\nConclusion and recommendation\n\nThis study revealed that there are widespread beliefs and misconceptions concerning long-acting reversible contraceptive use among pastoralist women. In this study, there are many barriers to LARC use, including myth and misconception (perceived side effects, FP is not suitable for women who work intensively, carry heavy objects, and travel long distances), the desire to have many children (children are wealth and God’s gifts), religious beliefs, and the husband’s objection.\n\nInformation education communication should be provided to address myths and misconceptions about the LARC during the FP intervention for pastoralist women via HEW, HCP, media, and community education. In addition, there is a need to enhance social opportunities by involving religious leaders and partners in decision-making. This could be accomplished by providing health information and counseling to religious leaders and couples in the community through health extension workers. It is critical to improve their understanding and correct misconceptions, particularly with respect to perceived side effects. LARC services should be integrated into other health services and delivered on a continual basis via a produced national standard manual and trained health care personnel. As myths and misconceptions about LARC use emanate from a variety of concerns, this study may not cover all of them; therefore, future research is needed to investigate additional myths and misconceptions concerning contraception use.", "appendix": "Data availability\n\nFigshare: Supplementary file to Exploring barriers to LARC utilization among the pastoralist women in southern Ethiopia: Qualitative study approach, 2024. https://doi.org/10.6084/m9.figshare.27918621.v2.28\n\nThis project contains the following:\n\n• Verbatim Transcription\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nFigshare: Supplementary file to Exploring barriers to LARC utilization among the pastoralist women in southern Ethiopia: Qualitative study approach, 2024. https://doi.org/10.6084/m9.figshare.27918621.v228\n\nThis project contains the following:\n\n• Interview Guide\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nFigshare: Supplementary file to Exploring barriers to LARC utilization among the pastoralist women in southern Ethiopia: Qualitative study approach, 2024. https://doi.org/10.6084/m9.figshare.27918621.v228\n\nThis project contains the following:\n\n• COREQ checklist for qualitative study design\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 would like to express our heartfelt gratitude to Bule Hora University, the data collectors who worked tirelessly to collect the data, and the participants in the study. Their contributions have been critical to the accomplishment of our study\n\n\nReferences\n\nAregay W, Azale T, Sisay M, et al.: Utilization of long acting reversible contraceptive methods and associated factors among female college students in Gondar town, northwest Ethiopia, 2018 : institutional based cross ‑ sectional study. BMC. Res. Notes. 2018; 11: 11–16. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHaile BT: Disparities in Long-Acting Reversible Contraceptive Utilization among Married Women in Ethiopia : Findings of the Ethiopian Demographic and Health Survey.2020; 2020.\n\nBihonegn D, Id A, Belachew TB, et al.: Postpartum long-acting reversible contraceptives use in sub-Saharan Africa. Evidence from recent demographic and health surveys data. PLoS One. 2023; 18: 1–14. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKasa AS, Tarekegn M, Embiale N: Knowledge, attitude and practice towards family planning among reproductive age women in a resource limited settings of Northwest Ethiopia. BMC. Res. Notes. 2018; 11: 7–12. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDibaba Y, Id W, Gurmu E, et al.: Contextual influences on the choice of long- acting reversible and permanent contraception in Ethiopia : A multilevel analysis.2019; pp. 1–17.\n\nGelgelo D, Abeya SG, Hailu D, et al.: Effectiveness of Health Education Interventions Methods to Improve Contraceptive Knowledge, Attitude, and Uptake Among Women of Reproductive Age, Ethiopia: A Systematic Review and Meta-Analysis. Heal. Serv. Res. Manag. Epidemiol. 2023; 10: 1–7. Publisher Full Text\n\nOntiri S, Ndirangu G, Kabue M, et al.: Long-Acting Reversible Contraception Uptake and Associated Factors among Women of Reproductive Age in Rural Kenya. pp. 1–11.\n\nMorgan IA, Zapata LB, Curtis KM, et al.: Health Care Provider Attitudes about the Safety of “Quick Start” Initiation of Long-Acting Reversible Contraception for Adolescents. HHS Public Access. 2020; 32(4): 402–408. Publisher Full Text\n\nGelgelo D, Hailu D, Garoma S: Effectiveness Of Theory-Based Peer Education On Knowledge, Attitudes, And Utilization Of Long-Acting Reversible Contraceptives. Ethiop. J. Reprod. Health. 2024; 16(4).\n\nSako S, Id S, Fikadu T, et al.: Four out of ten married women utilized modern contraceptive method in Ethiopia : A Multilevel analysis of the 2019 Ethiopia mini demographic and health survey.2022; pp. 1–16. Publisher Full Text\n\nDasa TT, Kassie TW, Roba AA, et al.: Factors associated with long-acting family planning service utilization in Ethiopia : a systematic review and meta-analysis.2019; pp. 1–14.\n\nAmong F, College F, Berhan D, et al.: Utilization of Long Acting Reversible Contraceptive Methods and Associated Utilization of Long Acting Reversible Contraceptive Methods and Associated Factors Among Female College Students in Debre Berhan Town, Ethiopia.2016; (December).\n\nAlemayehu M, Lemma H, Abrha K, et al.: Family planning use and associated factors among pastoralist community of afar region, eastern Ethiopia. BMC Womens Health. 2016; 16: 1–9. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEbrahim OA, Zeleke EG, Muluneh AG: Geographic variation and associated factors of long-acting contraceptive use among reproductive-age women in Ethiopia: a multi-level and spatial analysis of Ethiopian Demographic and Health Survey 2016 data. Reprod. Health. 2021 Jun; 18(1): 122. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJohnson JL, Adkins D, Chauvin S: Quality Indicators of Rigor in Qualitative Research. Am. J. Pharm. Educ. 2019; 84: 1–22. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMorison T, Eagar D, Morison T: Women’s perspectives on long-acting reversible contraception : a critical scoping review of qualitative research critical scoping review of qualitative research. Women Health. 2021; 61(6): 527–541. PubMed Abstract \| Publisher Full Text\n\nHaileyes KG, Asfaw HM, Ayele SG, et al.: Myth and misconception about long-acting reversible contraception use among reproductive age group women in Debre Berhan governmental health centers, Ethiopia: qualitative study.2022.\n\nASSOCIATION ASH: Media Contact Fred Wyand Director of Communications.\n\nGynecologists AC of O and: Long Acting Reversible Contraception Overview & Hands-On Practice for Residents ACOG disclaimer.\n\nLowth DM: Contraceptive implant What is the contraceptive implant? pp. 1–10.\n\nIversen L, Fielding S, Lidegaard Ø, et al.: Contemporary hormonal contraception and risk of endometrial cancer in women younger than age 50: A retrospective cohort study of Danish women. Contraception. 2020; 102(3): 152–158. PubMed Abstract \| Publisher Full Text\n\nAmerican College of Obstetricians and Gynecologists: Dispelling Long-Acting Reversible Contraception (LARC) Myths & Misconceptions Fact Sheet.2015; 4–5.\n\nMoreno MA: Long-acting reversible contraception for adolescents. JAMA Pediatr. 2016; 170(5): 516. Publisher Full Text\n\nGashaye KT, Gebresilassie KY, Kassie BA, et al.: Reasons for modern contraceptives choice and long ‑ acting reversible contraceptives early removal in Amhara Region, Northwest Ethiopia ; qualitative approach. BMC Womens Health. 2023; 23: 1–7. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGebeyehu A, Asnake M, Dibaba Y, et al.: BARRIERS TO UTILIZATION OF LONG ACTING REVERSIBLE AND PERMANENT CONTRACEPTIVE METHODS IN ETHIOPIA : ABSTRACT.2018; 10(3): 1–24.\n\nShumet T, Geda NR, Hassan JA: Barriers to modern contraceptive utilization in Ethiopia.2024.\n\nKamwesigye A, Amanya D, Nambozo B, et al.: Barriers and enablers to utilisation of postpartum long-acting reversible contraception in Eastern Uganda : a qualitative study.2024; 5.\n\nGelgelo D, Hailu D, Garoma S: Supplementary file to Exploring barriers to LARC utilization among the pastoralist women in southern Ethiopia: Qualitative study approach, 2024. figshare. [Dataset]. 2024. Publisher Full Text" }	[ { "id": "358089", "date": "21 Jan 2025", "name": "John Cleland", "expertise": [ "Reviewer Expertise Fertility", "family planning", "child survival." ], "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 reports the results of a qualitative study into barriers that limit uptake of long- acting contraceptive methods among pastoralists in southern Ethiopia. A total of 33 women of reproductive age participated in focus group discussions and 12 interviews were conducted with key informants including religious and community leaders and health care providers. Audio recordings were transcribed, translated from Oromo into English, and analysed using an inductive thematic approach. The majority of participants were illiterate and about half were currently using a contraceptive method. The analysis revealed 5 major themes, with each illustrated by verbatim quotations. The methods, presentation of results and interpretation conform to high standards for a qualitative study. It would be useful to know how many focus group discussions were held. It should also be noted that some of the barriers (eg desire for large families) apply to short-acting as well as long-acting methods.\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/13-1520
https://f1000research.com/articles/13-410/v1	26 Apr 24	{ "type": "Research Article", "title": "Predictors Of Tobacco Smoking Initiation Among Indonesian Teenagers: The 2019 Global Youth Tobacco Survey", "authors": [ "Hani Salsabila Deva", "Ferry Efendi", "Candra Panji Asmoro", "Ronal Surya Aditya", "Lisa McKenna", "Abdullah Saleh Alruwaili", "Hani Salsabila Deva", "Candra Panji Asmoro", "Ronal Surya Aditya", "Lisa McKenna", "Abdullah Saleh Alruwaili" ], "abstract": "Introduction We examined the predictors of tobacco smoking initiation considering factors such as age, gender, pocket money, exposure to smoke in public outdoor places, anti-tobacco media, smoking parents, smoking teachers, and smoking friends among adolescents in Indonesia.\n\nMethods Using data from the 2014–2019 National Youth Tobacco Survey grades 7–12, data from 9,655 teenagers were analyzed. The research used the Global Youth Tobacco Survey (GYTS) questionnaire. The analysis used univariate, chi-square and logistic regression with the STATA application version 15.\n\nResults In total, 9655 adolescent respondents participated. Multivariate tests indicated that teenagers aged 13-15 years (AOR: 1.34, 95% CI: 1.08-1.66) and teenagers aged> 15 years (AOR: 1.65, 95% CI: 1.25-2.17), teenagers who were male (AOR: 13.99, 95% CI: 11.11-17.63), teenagers who were exposed to cigarette smoke in public places outside the room (AOR: 2.00, 95% CI: 1.74-2.29), teenagers who had never seen anti-tobacco messages in the mass media (AOR: 1.23, 95% CI: 1.02-1.48), Teenagers who had parents who smoke (AOR: 1.26, 95% CI: 1.08-1.46), teenagers who saw teachers smoking in the school area (AOR: 1.42, 95% CI: 1.18-1.71), and teenagers who accepted offers of cigarettes from friends (AOR: 15.11, 95% CI: 10.33-22.11).\n\nConclusion The initiation of tobacco smoking among teenagers in Indonesia can be predicted by various factors such as age, gender, exposure to smoke in outdoor public places, exposure to anti-tobacco media, and influence of parents, teachers, and friends.", "keywords": [ "Indonesia", "Initiation", "Smoking", "Tobacco use" ], "content": "Introduction\n\nThe adverse health effects linked to tobacco have been thoroughly documented. Nevertheless, cigarette smoking remains the primary contributor to untimely and preventable deaths worldwide.1,2 Indonesia currently exhibits the highest prevalence of smoking globally, indicating a potential upward trend. Consequently, a significant number of Indonesian individuals, amounting to over 660 daily or exceeding 240,000 annually, succumb to ailments associated with tobacco consumption, with a particular emphasis on the adolescent demographic.3–5 According to a survey conducted by the Global Youth Tobacco in Indonesia, prevalence of smoking among teenagers aged 13-15 years rose from 33.9% in 2014 to 38.3% in 2019, indicating a significant increase. This percentage is notably high when compared to global statistics.6\n\nTragically, in 2016, 0.8% of girls and 1.2% of boys reported starting smoking at an early age when they were 5-9 years old.7 Health data research in 2018 showed that 77.7% of novice smokers in Indonesia started smoking before the age of 197. The results of research conducted by the 2018 Campaign for Tobacco Free Kids (CTFK) in 23 countries including Indonesia showed that the cigarette industry targeted children by advertising and promoting cigarette products in school environments.8 This contrasts with government programs during the 2015-2019 period, namely reducing smoking initiation by 1% per year.7\n\nThe age at which an individual begins smoking has been found to have a significant impact on their smoking habits in adulthood, as well as serving as an indicator of other problematic behaviors such as substance abuse, school disengagement, sexual risk-taking, and violent conduct.9 The current body of literature does not provide any forecast regarding the onset of smoking initiation in individuals. The findings of an Indonesian ministry study show that higher taxes on tobacco products have a very significant impact on reducing tobacco consumption among teenagers, especially by reducing smoking initiation, as well as among individuals with low socioeconomic status.7 However, alternative research posits that pricing strategies may not serve as a significant motivator for smokers to cease their habit, nor do they appear to act as a deterrent for young individuals who are considering initiating smoking.10\n\nSeveral research studies have investigated determinants that contribute to the initiation of smoking behavior globally. A study conducted with Jordanian students revealed that a decrease in availability of financial resources, an increase in educational attainment, and heightened awareness of the adverse health consequences of tobacco consumption were significant factors that contributed to reduction of smoking prevalence among the student population.10 Likewise, in in Canada, a positive correlation was observed between the smoking behaviors of parents, siblings, and peers and the prevalence of smoking among university students.1 Furthermore, a research investigation into initiation of smoking behavior among young adults in the Korea observed that although education exhibited a negative correlation with smoking, exposure to smoking and tobacco use demonstrated a positive correlation with smoking behavior among this demographic.11 Despite extensive research on the determinants of smoking initiation, there remains a dearth of specific studies pertaining to developing countries in Southeast Asia, particularly Indonesia. Given the compelling evidence regarding determinants of tobacco smoking, the Indonesia-based investigation offers valuable perspectives on the factors that underlie tobacco smoking in the area and proposes policy interventions that emerging economies could implement to deter student smoking.12\n\nTeenagers in Indonesia may also find it easy to access cigarettes, which can be bought anywhere at affordable prices for teenagers in Indonesia.13 Smoking initiation in teenagers is an important problem to be addressed because the younger the age at starting smoking, the stronger the smoking habit and the more difficult it is to quit smoking.12 The results of this study are expected to particularly inform theory development in community nursing. Knowledge of tobacco smoking initiation factors can be used as material for health promotion, as well as protection and prevention of diseases caused by tobacco smoking.14 The study findings can also provide additional information and overview of initiation factors of tobacco smoking in teenagers, and information for government to support development of health policies and programs aimed at preventing initiation of tobacco smoking in teenagers in Indonesia. Hence, the purpose of this study was determining predictors of tobacco smoking initiation among teenagers in Indonesia.\n\n\nMethods\n\nThe present investigation employed a cross-sectional research design, utilizing secondary data sources obtained from the 2019 Indonesia Global Youth Tobacco Survey (GYTS). The Global Youth Tobacco Survey (GYTS) is a component of the Global Tobacco Surveillance System (GTSS) that serves as a worldwide benchmark for the methodical monitoring of tobacco consumption (both inhalation and chewing) among young individuals, as well as the tracking of tobacco control metrics.9\n\nThis survey was given to a total of 9,992 students in grades 7-12 spread throughout Indonesia, yielding a total response rate of 91.0%. The methodology employed by GYTS conforms to a global standard and involves a two-stage sample design. The present study employed a purposive sampling method, whereby participants were selected based on specific inclusion criteria. Specifically, the sample consisted of students in grades 7-12 who had completed the survey. The final study comprised a sample size of 9,655 participants.\n\nFirst, the researcher downloaded the dataset needed in the study from the Global Youth Tobacco Survey (GYTS) in 2019. Then downloaded it on the website https://nccd.cdc.gov/GTSSDataSurveyResources/Ancillary/DataReports.aspx?CAID=2. after being downloaded, the respondent data recruited based on the inclusion criteria were grade 7-12 students who had completed the survey. Then, data cleaning was carried out to re-check the data for the possibility of missing data, which was found to be as many as 225 data elements. Then, we identified variables to determine the final variables to be used in the study. Variables were grouped into both independent and dependent variables. In identification of variables, a recoding process was carried out, namely the variables contained in the dataset were categorized into variables in accordance with the research without changing the original data and were given new code names.\n\nThe independent variables in this study were age, gender, pocket money, exposure to smoke in public outdoor places, anti-tobacco media, parents’ smoking, teachers’ smoking and friends’ smoking. While the dependent variable in this study was the initiation of tobacco smoking in teenagers.\n\nWe used multivariate logistic regression to measure the association between age, gender, allowance, exposure to smoke in public outdoor places, anti-tobacco media, parents’ smoking, teacher smoking and peer smoking, while controlling for the previously mentioned potential confounders. In analyzing data from the four survey rounds, each round received equal weighting. This weighting was applied to address the complexity of the GYTS data. Additionally, the “svyset” or survey estimation command was employed to accommodate the survey sampling method.15 The analysis used the STATA 1516 application and carried out using univariate analysis to enable description of the variables, then bivariate analysis was carried out using chi square tests to determine relationships between variables. Multivariate analysis using logistic regression analysis testing which aimed to determine which independent variable had the greatest influence.\n\nThis study was reviewed and approved by the Institutional Review Board of the Ministry of Health (LB.02.01/2/KE.315/2019) date approval: 7 August 2019. Participation was voluntary, and all respondents were provided with information about the research. This study complied with the ethical principles of the Declaration of Helsinki.\n\n\nResults\n\nBased on Table 1, from the total data of 9655 teenagers, the age of teenagers was categorized into 3 levels, namely <13 years, 13-15 years, and >15 years. Most were in the category of 13-15 years (n=4965, 51.43%). Most teenagers were women, as many as 5429 (56.23%). Pocket money was categorized into 4 levels, namely no pocket money, pocket money <1 USD, pocket money 1-5 USD, and pocket money >5 USD, with most teenagers in the Rp.11,000-50,000 category (n=4876, 50.50%). A total of 6,501 teenagers (67.33%) reported being exposed to cigarette smoke in public outside the room. Teenagers who reported seeing anti-tobacco messages on mass media such as television, radio, internet, billboards, posters, newspapers, magazines, or films were 7567 (78.37%). Most teenagers do not have smoking parents, 5546 (57.44%), less than half of teenagers who often see teachers smoking in the school environment are 4533 (46.95%). Almost all of the teenagers who refused offers to smoke from their friends were 8647 teenagers (89.56%). The number of teenagers who said they had never tried or experimented with smoking was 6128 (63.47%).\n\nTable 2 indicates that based on the results of bivariate analysis there were 7 independent variables that showed significant results on initiation of tobacco smoking in teenagers (p-value<0.05). These included: age (p-value=0.0053), gender (p-value=0.0000), smoke exposure in open public places (p-value=0.0000), anti-tobacco media (p-value=0.0029), parents who smoked (p-value=0.0108), teachers who smoked (p-value=0.0000), and friends who smoked (p-value=0.000). Meanwhile, the allowance variable (p-value=0.2636) did not show significant results on the dependent variable with p-value>0.05.\n\nBased on Table 3, teenagers who were >15 years old were 1.65 times more likely to start smoking than those who were 15 and 13–15 years old. Teenage males were 13.99 times more likely to start smoking than teenage girls. Teenagers who were exposed to secondhand smoke outside in crowded areas were twice as likely to start smoking. Teenagers were 1.23 times more likely to start smoking if they had never encountered anti-tobacco messaging in the media. Teenagers who had smoking parents were 1.26 times more likely to start smoking. Teenagers were 1.42 times more likely to start smoking if they observed instructors smoking in the schoolyard. Accepting cigarettes from friends increased a teenager’s chance of starting to smoke by 15.11 times.\n\n\nDiscussion\n\nThe predictor of initiation of tobacco smoking among teenagers in Indonesia is in accordance with the 2018 Basic Health Research which showed 77.7% of first-time smokers in Indonesia started smoking before the age of 19 years.8 from ordering children under the age of 18 (eighteen) years to sell, buy, or consume tobacco products. Another policy that is quite effective in preventing the emergence of novice smokers is to implement a 100% non-smoking area policy. The Main Provisions of Non-Smoking Area Regulations in Indonesia are rooms or areas that are declared prohibited for smoking activities or for producing, selling, advertising, and/or promoting tobacco products in the applicable places.17 Because there are already regulations and policies regarding this, the government can carry out more supervision regarding the policies that have been implemented to prevent the initiation of smoking in teenagers in Indonesia.\n\nAge has been shown to have a significant relationship with smoking initiation among teenagers in Indonesia. The results of research on age showed that the older the teenager, the risk to start smoking increases. This is consistent with research conducted in Samoa at 202118 and Korea at 202019 that age increases problems in life and their use of smoking is a coping mechanism. For this reason, health education is needed about the dangers of smoking, especially for older teenagers. Research has showed that a person is at risk for smoking initiation if they have insufficient knowledge about tobacco-related problems.20 Another study also found that discussions within the family about the harmful effects of smoking had a negative relationship with the adoption of tobacco smoking among students. This health education aimed to prevent teenagers from initiating smoking.21\n\nSex was found to have a significant relationship with smoking initiation in teenagers in Indonesia. The study’s results on gender showed that males were more at risk for initiating smoking. These results concur with studies in Africa,22 Nigeria,23 Samoa,18 and Korea.19 Adolescent males in Indonesia were found to be more likely to initiate smoking because the prevalence of smoking among the male population in Indonesia was higher than that of females. According to the 2018 Basic Health Research, the dominant gender is male in smoking initiation.8 Therefore, it is necessary to educate all genders about the dangers of smoking, especially male and female adolescents, which aims to prevent Indonesian youth from starting smoking. The study did not find a statistically significant correlation between receiving pocket money and smoking initiation among adolescents in Indonesia. This is in contrast to previous research conducted in Canada.24 While pocket money may not play a significant role in smoking initiation among adolescents in Indonesia, this should not be ignored because it has been identified as a causative factor for smoking initiation in other countries. Pocket money is not related to smoking initiation because smoking initiation in Indonesia is the low price of cigarettes.7 The 2019 GYTS data shows that most teenagers buy cigarettes in shops, stalls, street vendors and convenience stores. The cheap price of cigarettes makes it easier for teenagers to buy them. Although Government Policy 109/2012 prohibits the sale of cigarettes to youths under 18 years of age, most admit that they have never been prevented from buying cigarettes based on their age.25 Policies are needed regarding the increase in cigarette prices and re-monitoring of shops, stalls, street vendors and small shops selling cigarettes. Thus, they do not sell cigarettes in retail if the buyer is not over 18 years.26\n\nResearch has demonstrated a noteworthy correlation between outdoor exposure to smoke in public areas and onset of smoking behavior among teenagers in Indonesia. According to research findings, adolescents who are exposed to secondhand smoke in public places outside closed spaces are found to have a higher risk of starting smoking.27 The findings of this study are consistent with previous studies conducted in Nigeria23 and Samoa18 regarding adolescents who are accustomed to cigarette smoke and often start smoking. Researchers in Nigeria and Samoa demonstrated a positive correlation between external exposure and smoking initiation.18,23 Adolescents who are exposed to individuals who smoke in outdoor public areas, including playgrounds, roadsides, building entrances, parks, beaches, and sports fields, show a higher likelihood of initiating smoking behavior due to their high curiosity. and want to imitate what they have seen. There were already regulations in Indonesia prohibiting smoking indiscriminately, but supervision was still lacking. Hence, many people smoked haphazardly.28 Therefore, as community nurses, they provide education about smoking-free areas and the dangers of smoking, which aims to prevent young people from starting smoking.29\n\nAnti-tobacco media has a significant relationship with smoking initiation among teenagers in Indonesia. According to research on anti-tobacco media, teenagers who had never seen anti-tobacco messages in the mass media were more at risk for smoking.22,23,30 This was in line with Widyaningsih’s research that the mass media campaign against tobacco marketing had effectively reduced smoking initiation.31 However, this result contrasted with a study in Samoa, where the media did not have any impact on cigarette consumption.18 Teenagers in Indonesia already have adequate access to mass media and social media.32 Many teenagers today use the telephone to communicate. Anti-smoking media messages on television, radio, internet, billboards, posters, newspapers, magazines, or films are easy for teenagers to access.33 Technology development has integrated the lives of children and teenagers with social and digital media on the internet.34 Children and adolescents who are still in their infancy are reported to be interested in trying new information or something, even if they see famous people or their idols smoking.9 They can use social and digital media to educate themselves about the dangers of smoking to reduce smoking initiation.35\n\nParents who smoked had a significant relationship with smoking initiation in teenagers in Indonesia. This study was in line with research in Nigeria,23 Swedia,20 Samoa,18 and Africa.22 These results indicated that teenagers saw their parents as role models and prominent influencers.36 The influence of advertisements, cigarette advertisements that often appear on television which are packaged with gentlemen like men completely make teenagers or children feel attracted to be considered adults and even more so if the advertisement model is an idol.37 Targeting parents who smoke tobacco through various health interventions such as health education is suggested to be likely to reduce smoking initiation in their children.38\n\nTeachers who smoked had a significant relationship with smoking initiation in teenagers in Indonesia. The study’s results on teachers who smoked showed that teenagers who saw teachers smoking in the school area were more at risk for initiating smoking.38 According to a study in Beijing, China, there was a positive correlation between smoking and teachers who smoked.39 However, this result contrasted with a study in Samoa, where factors such as smoking among teachers were insignificant in smoking initiation.18 Teenagers’ parents regarded teachers as role models.40 Hence, targeting teachers who smoke tobacco in school areas through various health interventions such as health education is likely to reduce smoking initiation in students or teenagers.41\n\nFriends who smoked had a significant relationship with smoking initiation in teenagers in Indonesia. The results indicated that teenagers who accepted cigarette offers from friends were more at risk of initiating smoking. Research in Swedia suggested that the influence of smoking friends on boys and girls in the sample was in line with a theory that emphasizes the power of peer modeling in developing health behaviors among teenagers.20 This was consistent with research in Samoa,18 Nigeria,23 and Africa22 that curiosity and peer pressure are the most contributing factors to the formation of the intention to smoke. This is in line with Indonesia where most teenagers are starting to dare to try cigarettes because they see other friends smoking. Therefore, it is necessary to supervise teenagers and educate parents to reduce the initiation of smoking.42\n\nNevertheless, this study is not without its limitations. Initially, it should be noted that the data is reliant on self-reporting, which may result in either an overestimation or underestimation of the actual values. Several factors related to smoking initiation, including the type of cigarette product used, frequency, age of onset, and motivation for smoking initiation, were not examined in our study. Further research efforts should aim to investigate the factors associated with the initiation of smoking among teenagers.\n\n\nConclusion\n\nIn conclusion, this study sheds light on predictors of tobacco smoking initiation among teenagers in Indonesia. The results suggest that several factors play a significant role in the initiation of smoking behavior, including age, gender, exposure to smoke in outdoor public places, exposure to anti-tobacco media, influence of parents, teachers, and friends. The findings highlight the need for interventions and policies targeted at reducing tobacco use among teenagers in Indonesia. Nurses and healthcare professionals can use these findings to develop effective smoking prevention and cessation programs for teenagers. Furthermore, policymakers can utilize these insights to design and implement health policies and initiatives to curb smoking behavior among Indonesian teenagers.\n\n\nAuthors’ contributions\n\nAll authors edited and approved the final manuscript.\n\n\nProvenance and peer review\n\nNot commissioned; externally peer reviewed.\n\n\nEthics and inform consent statement\n\nThis study was reviewed and approved by the Institutional Review Board of the Ministry of Health (LB.02.01/2/KE.315/2019 date approval: 7 August 2019). Participation was voluntary, and all respondents were provided with information about the research. This study complied with the ethical principles of the Declaration of Helsinki.\n\nThe data used in the study were collected by a reputable survey (Global Youth Tobacco Survey), and it is assumed that ethical considerations, including obtaining informed consent, were addressed during the original data collection process. The researchers adhered to any regulations, guidelines, or ethical standards set forth by the organization or authority that provided the data. Compliance with these standards may obviate the need for individual consent for secondary data analysis.", "appendix": "Data availability\n\nThe data underlying the results presented in the study are available from the https://extranet.who.int/ncdsmicrodata/index.php/catalog/926 by submitting an application.\n\nFigShare: Checklist for Predictors of tobacco smoking initiation among indonesian teenagers: the 2019 global youth tobacco survey, https://doi.org/10.6084/m9.figshare.25512025.v1. 43\n\nData are available under the terms of the Creative Commons Attribution International License (CC BY 4.0).\n\n\nAcknowledgements\n\nNone.\n\n\nReferences\n\nCui Y, Forget EL, Zhu Y, et al.: The effects of cigarette price and the amount of pocket money on youth smoking initiation and intensity in Canada. Can. J. Public Health. 2018; 110(1): 93–102. PubMed Abstract \| Publisher Full Text\n\nAzagba S, Baskerville NB, Minaker L: A comparison of adolescent smoking initiation measures on predicting future smoking behavior. Prev. Med. Rep. 2015; 2: 174–177. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSoerojo W, Bigwanto M, Susilo D, et al.: Fakta Tembakau Indonesia 2020: Data Empirik untuk Pengendalian Tembakau. Jakarta: Ikatan Ahli Kesehatan Masyarakat Indonesia; 2020; 200.\n\nFithria F, Adlim M, Jannah SR, et al.: Indonesian adolescents’ perspectives on smoking habits: a qualitative study. BMC Public Health. 2021; 21(82): 1–8. Publisher Full Text\n\nWorld Health Organization: Lembar Informasi Indonesia 2019 (Global Youth Tobacco Survey). Global Youth Tobacco Survey (GYTS); 2020; 1–2. Reference Source\n\nBadan Penelitian dan Pengembangan Kesehatan: Jumlah Perokok Anak Semakin Tinggi Anak-anak Terpapar. Kementrian Kesehatan RI; 2019; pp. 9–10.\n\nWorld Health Organization: Raise Tobacco Taxes and Prices for a Healthy and Prosperous Indonesia. Jakarta, Indonesia: World Health Organization, Regional Office for South-East Asia; 2020; 23.\n\nMinistry of Health: Laporan Riskesdas 2018 Nasional. Jakarta: Ministry of Health Republic Indonesia; 2018.\n\nMa C, Xi B, Li Z, et al.: Prevalence and trends in tobacco use among adolescents aged 13–15 years in 143 countries, 1999–2018: findings from the Global Youth Tobacco Surveys. Lancet Child Adolesc Heal. 2021; 5(4): 245–255. PubMed Abstract \| Publisher Full Text\n\nNagelhout GE, Mons U, Allwright S, et al.: Prevalence and predictors of smoking in “smoke-free” bars. Findings from the International Tobacco Control (ITC) Europe Surveys. Soc. Sci. Med. 2011; 72(10): 1643–1651. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLee Y, Lee K-S: Factors Related to Smoking Status Among Young Adults: An Analysis of Younger and Older Young Adults in Korea. J. Prev. Med. Public Health. 2019 Mar; 52(2): 92–100. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAsyary A, Veruswati M: Smoking Behavior and Cigarette Expenditure in a Household: Evidence for Smoke-Free Houses Initiation in Indonesia. Int. J. Prev. Med. 2023; 14(7): 1–4.\n\nKumboyono K, Hamid AYS, Sahar J, et al.: Community response to the initiation of smoking in Indonesian early adolescents: a qualitative study. Int. J. Adolesc. Youth. 2020; 25(1): 210–220. Publisher Full Text\n\nKalkhoran S, Benowitz NL, Rigotti NA: Prevention and Treatment of Tobacco Use: JACC Health Promotion Series. J. Am. Coll. Cardiol. 2018 Aug; 72(9): 1030–1045. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSTATA: Svyset-Declare survey design for dataset. Texas: Stata; 2022.\n\nCorp S: Stata Statistical Software: Release 15. College Station, TX: StataCorp LLC; 2017. Reference Source\n\nMartini S, Artanti KD, Hargono A, et al.: Association between percentage of smokers and prevalence of smoking attributable morbidity in Indonesia: one decade after implementation of smoke-free area regulation. BMC Public Health. 2022; 22(1): 2202. PubMed Abstract \| Publisher Full Text \| Free Full Text Reference Source\n\nSingh B, Chand SS, Chen H: Tobacco smoking initiation among students in Samoa and health concerns. PLoS One. 2021; 16(10): e0258669. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKim E, Park E, Kim H: Sex Differences in Multilevel Factors of Smoking Experimentation and Age of Initiation in Korean Adolescents. J. Sch. Nurs. 2020; 36(5): 348–359. PubMed Abstract \| Publisher Full Text\n\nRaffetti E, Landgren AJ, Andersson F, et al.: Cortisol Concentration as Predictor of Tobacco Initiation in Adolescents: Results From a Population-Based Swedish Cohort. J. Adolesc. Health. 2021; 68(4): 758–764. PubMed Abstract \| Publisher Full Text\n\nMasood M, Masood Y, Md Sabri BA, et al.: Within-Family Discussion on Harmful Effects of Smoking and Intention to Initiate Smoking Among European Adolescents. J. Addict. Med. 2015; 9(4): 261–265. PubMed Abstract \| Publisher Full Text\n\nVeeranki SP, John RM, Ibrahim A, et al.: Age of smoking initiation among adolescents in Africa. Int. J. Public Health. 2016; 62(1): 63–72. PubMed Abstract \| Publisher Full Text\n\nOdukoya OO, Odeyemi KA: Determinants of Smoking Initiation and Susceptibility to Future Smoking among School-Going Adolescents in Lagos State, Nigeria. Asian Pacific J. Cancer Prev. 2013; 14(3): 1747–1753. PubMed Abstract \| Publisher Full Text\n\nCui Y, Forget EL, Zhu Y, et al.: The effects of cigarette price and the amount of pocket money on youth smoking initiation and intensity in Canada. Can. J. Public Health. 2019 Feb; 110(1): 93–102. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSun J, Xi B, Ma C, et al.: Cigarette access and purchase patterns among adolescent smokers aged 12-16 years in 140 countries/territories, Global Youth Tobacco Survey 2010-2018. J. Glob. Health. 2022 Dec; 12: 4101. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNurhasana R, Ratih SP, Dartanto T, et al.: Public support for cigarette price increase in Indonesia. Tob. Control. 2022 May 1; 31(3): 483–486. Publisher Full Text Reference Source\n\nKuwabara Y, Kinjo A, Kim H, et al.: Secondhand Smoke Exposure and Smoking Prevalence Among Adolescents. JAMA Netw. Open. 2023 Oct; 6(10): e2338166. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEvans WD, Renaud J, Blitstein J, et al.: Prevention Effects of an Anti-Tobacco Brand on Adolescent Smoking Initiation. Soc. Mark. Q. 2007; 13(2): 2–20. Publisher Full Text\n\nKrainuwat K: Smoking initiation prevention among youths: implications for community health nursing practice. J. Community Health Nurs. 2005; 22(4): 195–204. PubMed Abstract \| Publisher Full Text\n\nChoi S, Kim Y, Lee J, et al.: Tobacco use among students aged 13-15 years in South Korea: The 2013 global youth Tobacco survey. J. Prev. Med. Public Health. 2017; 50(1): 60–65. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWidyaningsih V, Mashuri YA, Nugroho SD, et al.: Sociodemographic predictors for smoking persistence among young males in Indonesia. J. Public Heal. Dev. 2022; 20(3): 175–183. Publisher Full Text\n\nSeptiono W, Kuipers MAG, Ng N, et al.: Self-reported exposure of Indonesian adolescents to online and offline tobacco advertising, promotion and sponsorship (TAPS). Tob. Control. 2022 Jan 1; 31(1): 98–105. Publisher Full Text Reference Source\n\nCarson KV, Ameer F, Sayehmiri K, et al.: Mass media interventions for preventing smoking in young people. Cochrane Database Syst. Rev. 2017 Jun; 6(6): CD001006. PubMed Abstract \| Publisher Full Text\n\nHaddock A, Ward N, Yu R, et al.: Positive Effects of Digital Technology Use by Adolescents: A Scoping Review of the Literature. Int. J. Environ. Res. Public Health. 2022 Oct; 19(21). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJudd TM, Sloane EB, Silva RJ: The digital hospital of the 21th century, and information systems management.Iadanza E, editor. Clinical Engineering Handbook. 2nd ed.Academic Press; 2020; pp. 530–542.\n\nChinwong S, Saharatchataphong K, Kachai C, et al.: Knowledge and attitude towards electronic cigarettes among university student smokers in Thailand. Tob. Induc. Dis. 2021; 19(1): A102. Publisher Full Text\n\nReitsma MB, Flor LS, Mullany EC, et al.: Articles Spatial, temporal, and demographic patterns in prevalence of smoking tobacco use and initiation among young people in 204 countries and territories, 1990 – 2019. Lancet Public Health. 2019; 6(7): e472–e481. Publisher Full Text\n\nLee J, Kong G, Kassas B, et al.: Predictors of vaping marijuana initiation among US adolescents: Results from the Population Assessment of Tobacco and Health (PATH) study Wave 3 (2015–2016) and Wave 4 (2016–2018). Drug Alcohol Depend. 2021; 226(July): 108905. PubMed Abstract \| Publisher Full Text\n\nCheng X, Guo X, Jin C: Social determinants of smoking among school adolescents in Beijing, China. Tob. Induc. Dis. 2022; 20: 1–9. Publisher Full Text\n\nNazlıcan E, Ersoy Ö, Akbaba M, et al.: Social factors should not be underestimated in smoking behaviour among adolescents: A patriarchal experience from senior high school students in Turkey. Tob. Induc. Dis. 2018; 16(3): 3.\n\nDuke JC, Macmonegle AJ, Nonnemaker JM, et al.: Impact of The Real Cost Media Campaign. Am. J. Prev. Med. 2019; 57(5): 645–651. PubMed Abstract \| Publisher Full Text\n\nZhu B, Liu S, Program L: Cigarette Smoking and Its Risk Factors among Elementary School Students in Beijing. Am. J. Public Health. 1996; 86(3): 368–375. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEfendi F: Strobe checklist. figshare. [Dataset]. 2024. Publisher Full Text" }	[ { "id": "278564", "date": "13 Jun 2024", "name": "Wahyu Septiono", "expertise": [ "Reviewer Expertise statistics", "tobacco control", "data science", "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\nGeneral comment Please consult a proofreader.\nINTRODUCTION 1. \"Indonesia currently exhibits the highest prevalence of smoking globally, indicating a potential upward trend.\" please specify the latest prevalence. 2. \"Consequently, a significant number of Indonesian individuals, amounting to over 660 daily or exceeding 240,000 annually, succumb to ailments associated with tobacco consumption, with a particular emphasis on the adolescent demographic.\" Are 600 only adolescents or all age? Better to specify only adolescents to show the relevancy with the title 3. \"...teenagers aged 13-15 years rose from 33.9% in 2014 to 38.3% in 2019, indicating a significant increase.\" please put the reference 4. \"Tragically, in 2016, 0.8% of girls and 1.2% of boys reported starting smoking at an early age when they were 5-9 years old.\" while this may seem to be very low, I suggest authors to emphasize a shocking worldwide video from Indonesia about toddler smoker which may show that cases of smoking among that age may be undereported https://www.youtube.com/watch?v=x4c_wI6kQyE&t=2s&ab_channel=OnDemandNews 5. \"This contrasts with government programs during the 2015-2019 period, namely reducing smoking initiation by 1% per year.\" This seems quite old. Perhaps the latest program will be better. 6. \"Several research studies have investigated determinants that contribute to the initiation of smoking behavior globally.\" In paragraph 4, I think it is more accurate if authors can cite systematic review studies.\n\nMETHODS 1. The author wrote, \"The present investigation employed a cross-sectional research design, utilizing secondary data sources obtained from the 2019 Indonesia Global Youth Tobacco Survey (GYTS).\" I suggest rephrasing this as the study did not collect primary data but used secondary data. Referring to this study as using a cross-sectional design might be misleading. For example: \"The study used secondary data from the 2019 Indonesia Global Youth Tobacco Survey (GYTS), in which data from GYTS was collected through a cross-sectional study design.\" And please additional information about the GYTS: 1) study population, 2) sampling method, 3) number of sample, etc. 2. I don't believe that \"Data Collection\" is the appropriate subheading for this study. See the reason at number 1 above. 3. The study did not clearly explain the variables included. For example, it is unclear which survey questions corresponded to the variables and how they were measured. Readers need to know the original instrument and understand how the authors converted these variables for analysis in the study. 4. the term \"multivariate logistic regression\" is incorrect for such analysis. See: reference 1. 5. The data analysis section did not thoroughly explain the steps taken for the analysis. Additionally, it is unclear why the authors applied weighting. Weighting can be used for several reasons, such as adjusting for over-sampling of a particular group in the population or accounting for non-response. 6. Authors stated that \"Multivariate analysis using logistic regression analysis testing which aimed to determine which independent variable had the greatest influence.\" However, using regression to determine which variable had the greatest influence seems less feasible. While regression can provide insights into the relationships between variables and the dependent variable, determining which variable has the greatest influence requires careful consideration of the context, scale, potential multicollinearity, and the limitations of regression analysis. This should be performed under Dominance analysis if authors aim to determine the greatest influence. Dominance analysis provides understanding the relative importance of predictor variables in a regression model. It provides a nuanced view that takes into account multicollinearity and the combined effects of predictors, offering a clear ranking of variables based on their contributions to the explained variance in the dependent variable.\nResults\n1. \"Most teenagers were women..\" While many women might enjoy being referred to as teenagers, this is not the correct usage in English. 2. Why did the authors include the categorization in the results section? This should be in the methods section. Please refer to my previous comment! 3. There's no need to split the analysis into two tables.\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? 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? Yes", "responses": [] } ]	1	https://f1000research.com/articles/13-410
https://f1000research.com/articles/13-1121/v1	03 Oct 24	{ "type": "Clinical Practice Article", "title": "Breaking point: Case series of tendon ruptures in Hemodialysis patients", "authors": [ "Muhammed Ehsan Nazeer", "Dr Askhar Haphiz", "Dr Muhammed Nazeer", "Dr Pradeep Moni", "Dr Praveen Muraleedharan", "Dr Askhar Haphiz", "Dr Muhammed Nazeer", "Dr Pradeep Moni", "Dr Praveen Muraleedharan" ], "abstract": "Introduction Spontaneous tendon ruptures in end stage kidney disease patients have the potential to cause long- term morbidity, and timely intervention is required to prevent complications that can severely affect the functional status of the patient\n\nCase presentation A series of six tendons (two triceps tendons and two bilateral quadriceps tendons) in three patients with ESKD undergoing hemodialysis is discussed in this case series. Patients were aged 61, 44 and 26 years, and on hemodialysis for 5, 5 and 10 years, respectively.\n\nConclusion End -stage kidney disease is associated with a multitude of physiological changes, and the musculoskeletal system is no exception to this. Spontaneous tendon rupture is a multifactorial complication of ESKD, with serious implications for mobility and quality of life. As a result, these patients require a multifaceted approach to ensure optimum results and an early return to activity. We report a series of 6 spontaneous tendon ruptures in 3 patients with ESKD at our institution. We would like to outline the methods of repair for each case and further attempt to assess biochemical parameters that may have contributed to the disease process.", "keywords": [ "ESRD", "Quadriceps", "Hyperparathyroidism", "Tendon rupture", "Hemodialysis", "Case series" ], "content": "Introduction\n\nSpontaneous tendon rupture is rare in the general population. Most cases of tendon rupture occur secondary to trauma or degenerative changes resulting from obesity or old age. Other etiologies of tendon rupture include pathological alterations in the substance of the tendon itself, such as gout,1 autoimmune arthritis,1 and end stage renal disease.2 Long-term use of drugs such as statins, quinolones and steroids3 constitute yet another often implicated cause of spontaneous tendon ruptures.\n\nLiterature on spontaneous tendon ruptures in patients with ESKD undergoing hemodialysis is uncommon and studies on the exact underlying mechanism and management guidelines for such patients are sparse.\n\nThe objective of this case series was to report a series of patients on hemodialysis for ESKD who experienced spontaneous tendon ruptures with emphasis on the methods and results of surgical management. An assessment of laboratory parameters was also performed to identify risk factors for the same.\n\n\nCase reports\n\nA 61 year old male suffering with ESKD due to nephrosclerosis on hemodialysis for the last 5 years presented to the emergency department with complaints of acute onset pain and swelling in both knees while attempting to stand up from a sitting position. Examination revealed effusion in both knees with a palpable defect in the bilateral patellar tendons and bilateral extension lag. Lateral radiographs showed bilateral patella alta with calcified patellar tendons (Figure 1).\n\nSurgical repair was performed in a staged fashion with an interval of 2.5 weeks in view of his physiological status. Surgical exploration revealed friable tendon edges with partial avulsion and mid substance tear patterns. The degenerated tissue was excised, and the ends of the tendon were reattached to the patella using suture anchors and non-absorbable sutures, followed by closure of the defects in the medial and lateral paratenon using bioabsorbable sutures (Figure 2).\n\nPost operatively the knees were immobilized in full extension for 6 weeks. DVT prophylaxis was administered using low molecular weight heparin (40 mg, sub-cutaneous injections, once daily) during the hospital stay and Rivaroxaban on Discharge. He was started on active Knee ROM and quadriceps strengthening exercises at six weeks.\n\nHe complained of left elbow extension weakness in his review for suture removal on the second knee, and further evaluation revealed a spontaneous rupture of the triceps tendon. This was likely caused by sustained forces while using a walker to assist ambulation. He has been advised to undergo surgical repair, for which he is yet to be reported.\n\nFollow-up examinations at 8 and 8.5-months post-op respectively, examination revealed a knee ROM of 0-120 degrees with bilateral full extension and no residual weakness in the quadriceps mechanism.\n\nEnd stage renal disease was managed by our nephrology department with thrice weekly hemodialysis sessions and regular monitoring of renal parameters.\n\nA 44-yr old male with ESRD secondary to immunoglobinA nephropathy with a history of allograft rejection during three weekly hemodialysis sessions for 10 years presented with complaints of right elbow extension weakness following trivial trauma. Physical examination revealed swelling of the right elbow with grade 0 power in the elbow extensor mechanism. Radiographs showed an avulsed triceps tendon (Figure 3).\n\nHe underwent open repair of the triceps tendon with a suture anchor inserted in the olecranon after excision of the devitalized tongue of the tissue (Figure 4). The elbow was immobilized in an above elbow slab at approximately 110° for 6 weeks, after which active mobilization was started. The last follow-up at 7 months revealed grade 5/5 power with full elbow ROM.\n\nA 26-year-old male, suffering from lupus nephritis presented to ER with acute loss of extension in both knees and inability to ambulate following a fall while descending stairs at his house. The treatment history was significant for the intake of methylprednisolone for the last 3 months and twice weekly hemodialysis for ESKD.\n\nExamination revealed bilateral patella alta and boggy swelling in both knees with grade 0 power in bilateral knee extension. Imaging revealed isolated patella alta without any avulsed bony edges (Figure 5).\n\nHe was diagnosed with bilateral patellar tendon rupture and underwent open repair during which severe fraying of the tendons was noted (Figure 6). Robust repair was performed using non-absorbable sutures in a mattress pattern (Figure 7). He was given bilateral long knee immobilizers for 6 weeks to protect the repair, following which he was started on assisted ambulation with walker support over the next 4 weeks. At the final review at 6 months, he had grade 5/5 power in both his knee extensors with no appreciable lag on either side.\n\n\nDiscussion\n\nNormal tendons are strong, with Ultimate Tensile Strengths ranging from 45 to 125 MPa which exceed three times the strain caused by muscle contraction.4 The tendency for spontaneous tendon ruptures in patients with ESKD has often been attributed to five main factors:\n\n1) elastosis secondary to chronic acidosis,5\n\n2) chronically elevated blood urea levels,1\n\n3) amyloidosis with beta 2 microglobulin deposition,6\n\n4) increased cortical bone resorption because of secondary hyperparathyroidism,2\n\n5) high circulating calcium levels resulting in dystrophic calcification of the tendon.7\n\nThis contrasts with other cases of spontaneous tendon rupture, which is a condition of middle-aged overweight patients, with bilaterality being an exception rather than the norm.8\n\nA quick diagnosis is paramount because early surgical intervention is a key factor in ensuring optimal results.9 This rare diagnosis should always be considered when dealing with sudden unexplained motor deficits in the lower limbs. Hemarthrosis can further complicate the matter by making it difficult to identify infrapatellar defects. Such cases warrant the use of incongruous techniques, such as ultrasound and MRI to confirm the diagnosis.\n\nA prolonged duration of hemodialysis appears to be a reliable risk factor for the development of spontaneous tendon ruptures.10 The markedly elevated PTH and ALP levels in our patients were also concurrent with the existing literature and explain the avulsed tendon ends noted both intraoperatively and radiographically in 2 cases in our series (Table 1).\n\nThe repairs were performed as soon as fitness could be obtained, in view of the compromised physiological status of the patients. No additional procedures were required to compensate for retraction or shortening of tendons. Repair was performed using suture anchors and heavy non absorbable sutures. The patients attained full active extension in all cases at an average of 2 months from the date of surgery with no re-ruptures or residual weakness at a minimum 6 months follow up.\n\nAlthough the exact mechanism of injury remains unknown, most existing studies implicate sub-tendon bone resorption due to secondary hyperparathyroidism,2 in addition to chronic inflammation, as evidenced by reduced serum hemoglobin and albumin levels, which was noted in 50% of cases.\n\n\nConclusion\n\nSpontaneous tendon rupture in patients with ESKD remains a significant complication with considerable potential to cause disability. Robust repair with adequate immobilization and protection during the healing phase is instrumental in ensuring reliable results. Moreover, these patients require cautious physio rehabilitation, including careful walker- assisted mobilization to prevent further tendon ruptures during the postoperative period.\n\nOptimization of renal parameters, such as hyperparathyroidism and hypoalbuminemia, may help improve outcomes and prevent recurrence. Overall, a high index of suspicion for spontaneous tendon rupture is advised in patients with ESKD presenting with acute-onset limb weakness to prevent long-term morbidity.\n\n\nConsent\n\nThe patients have given their informed consent for the case series to be published. Written consent was obtained for the same.", "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\nShah MK: Simultaneous bilateral rupture of quadriceps tendons: analysis of risk factors and associations. South. Med. J. 2002 Aug; 95(8): 860–866. PubMed Abstract \| Publisher Full Text\n\nBasic-Jukic N, Juric I, Racki S, et al.: Spontaneous tendon ruptures in patients with end-stage renal disease. Kidney Blood Press. Res. 2009a; 32(1): 32–36. PubMed Abstract \| Publisher Full Text\n\nSeng C, Lim YJ, Pang HN: Spontaneous disruption of the bilateral knee extensor mechanism: a report of two cases. J. Orthop. Surg. (Hong Kong). 2015 Aug; 23(2): 262–266. PubMed Abstract \| Publisher Full Text\n\nHench LL: The skeletal system.Hench LL, Jones JR, editors. Biomaterials, artificial organs and tissue engineering. Woodhead Publishing; 2005; pp. 79–89. (Woodhead Publishing Series in Biomaterials). 9781855737372. Publisher Full Text\n\nRuiz J, Ríos A, Rodríguez JM, et al.: Roturas tendinosas espontáneas en la insuficiencia renal crónica. Nefrologia. 2017; 37: 341–343. PubMed Abstract \| Publisher Full Text\n\nGrecomoro G, Camarda L, Martorana U: Simultaneous chronic rupture of quadriceps tendon and contra-lateral patellar tendon in a patient affected by tertiary hyperparathyroidism. J. Orthop. Traumatol. 2008 Sep; 9(3): 159–162. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHartono F, Besinga KE, Tjie H, et al.: Considerations in spontaneous quadriceps tendon rupture repair in end-stage renal disease patients: A case report. Int. J. Surg. Case Rep. 2021 Sep; 86: 106298. Epub 2021 Aug 10. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDhar S: Bilateral, simultaneous, spontaneous rupture of the quadriceps tendon: A report of 3 cases and a review of the literature. Injury. 1988; 19(1): 7–8. Publisher Full Text\n\nWani NA, Malla HA, Kosar T, et al.: Bilateral quadriceps tendon rupture as the presenting manifestation of chronic kidney disease. Indian J. Nephrol. 2011 Jan; 21(1): 48–51. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPei YC, Hsieh PC, Huang LZ, et al.: Simultaneous bilateral quadriceps tendon rupture in a uremic patient. Formos. J. Musculoskelet. Disord. 2011; 2(1): 35–39. Publisher Full Text" }	[ { "id": "332004", "date": "06 Nov 2024", "name": "Sandeep Vijayan", "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\nAbstract – case presentation: better to mention as 'patellar tendon' as the involved part is infrapatellar and not suprapatellar.\nPage 3- case 2: I am not sure if it is correct to say grade 0 power. Here we are dealing with mechanical disruption of the muscle without loss in power of the muscle. Attempt to move the elbow will definitely contract the triceps muscle without the force transmitting across the tear. So, is it truly grade 0 power in the muscle ?\nPage 3, case 2: was there any USG or MRI images available confirming the diagnosis and assess the quality of the tissue. Was histopathological examination of the excised tissue performed to understand the type of degenerative changes commonly happening in tendons and ligaments in ESRD.\nPage 5, figure 5 legend - better to mention as 'patellar tendon rupture'.\nPage 6, figure 6- Please add cranial end and caudal end in the picture for easy understanding for the readers, The picture on the left side appear skewed (stretched horizontally). Please re-size it correctly.\nPage 7- discussion - It is interesting to note that in most of the reported literatures. it the quadriceps / patellar tendon which is getting ruptured. In your review of literature, did you come across any specific reasoning from the affection of quadriceps tendon in particular.\nFew more minor corrections are suggested. Same has been commented on in the main manuscript and may be accessed by opening the manuscript with Adobe reader.\n\nIs the background of the cases’ 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 conclusion balanced and justified on the basis of the findings? Yes", "responses": [] }, { "id": "335311", "date": "07 Nov 2024", "name": "Luis Marcelo Malta", "expertise": [ "Reviewer Expertise Orthopaedics / orthopedic 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\nThe authors show their experience with a series of tendon ruptures on regular-based hemodialysis patients, addressing aspects of surgical treatment and functional results. Despite the fact that it is a case series, giving it a less favorable view in terms of scientific strength or evidence, the study brings important information about the understanding and management of this unusual occurrence. There are also some details I would like to point:\n\n1- Abstract says that the authors report on two bilateral QUADRICEPS ruptures, but both case 1 and 3 are PATELLAR tendon ruptures. Maybe it was just a typing mistake but should be corrected to prevent confusion. 2- All cases include physical evaluation, both pre and postoperative, without clarifying which scale or functional score was used to do such measurements. I consider this is a minor but important point to address. 3- Discussion is well done but could include more recent articles at surgical repair aspects and risk factors associated with ruptures available in the literature. I consider this a major point to be revised, because there are much work documented on the field and it would make this section more complete.\n\nIs the background of the cases’ 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? Partly\n\nIs the conclusion balanced and justified on the basis of the findings? Yes", "responses": [] }, { "id": "335309", "date": "12 Nov 2024", "name": "Matthew Abramowitz", "expertise": [ "Reviewer Expertise physical function", "muscle physiology in CKD" ], "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 case series brings attention to the risk of tendon rupture in patients with ESKD. There are several areas in which it could be strengthened: 1.\n\nTo improve the generalizability of this report to other patients with ESKD, it would be helpful to present details of the patients’ dialysis prescriptions, such as use of high flux dialyzers (which would reduce the likelihood of beta-2 microglobulin as an etiologic factor), pre-dialysis bicarbonate levels, Kt/V, dialysate settings (especially bicarbonate), and treatment of hyperparathyroidism. If beta-2 microglobulin levels were measured, those data should be presented. 2.\n\nRegarding case 3, the duration of dialysis should be presented. If the patient was not recently started on dialysis and did not have significant residual kidney function, it would be helpful to know why he was only receiving dialysis twice per week. 3.\n\nThe abstract describes quadriceps tendon ruptures but the case descriptions report patella tendon ruptures. This should be clarified. 4.\n\nAbstract, Case presentation: the years on hemodialysis should be presented in the same order as the patients’ ages. 5.\n\nIt would be helpful to further discuss potential mechanisms by which risk factors in ESKD may impact tendon physiology and structure.\n\nIs the background of the cases’ 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? Partly\n\nIs the conclusion balanced and justified on the basis of the findings? Yes", "responses": [] } ]	1	https://f1000research.com/articles/13-1121
https://f1000research.com/articles/12-469/v1	05 May 23	{ "type": "Research Article", "title": "Phenotypic and genetic extended spectrum beta lactamase profiles of bacterial isolates from ICU in tertiary level hospital in Kenya", "authors": [ "Job Mwale", "Edwin O. Magomere", "Brian Maina", "Leon Otieno", "Frank G. Onyambu", "Ali Kassim", "Lucy Muchiri", "Brian Maina", "Leon Otieno", "Frank G. Onyambu", "Ali Kassim", "Lucy Muchiri" ], "abstract": "Background: Bacterial infections in the ICUs are a threat to the lives of critically ill patients.\n\nMoreover, their vulnerable immunity predisposes them to developing bacteria-associated sepsis, further deteriorating their already fragile health. In the face of increasing drug resistance menace, the problem of bacterial infection in ICU is likely to worsen. This study sought to assess bacterial infections in ICU setting by identifying prevalent gram-negative bacterial species and characterizing their antibiotic susceptibility patterns. Phenotypic and genetic resistance were determined among gram-negative isolates from ICU patients.\nMethods: Cross-sectional samples collected from Kenyatta National Hospital ICU between\n\nJanuary and June 2021 were cultured for subsequent analysis. Phenotypic identification of culture-positive samples was performed using VITEK 2 while Antibiotic susceptibility patterns were determined based on Antimicrobial Susceptibility Testing (AST) results. Cephalosporin-resistant gram-negative bacteria were assessed by PCR to detect the presence of ESBL genes. Results and discussion: Out of the 168 gram-negative isolates, Acinetobacter baumanii was the most abundant (35%). Other isolates that were present at frequencies more than 15% are Klebsiella pneumoniae and Escherichia. coli. A. baumaniii is known to be a notorious bacterium in ICU due to its multidrug resistance nature. Indeed, A. baumanii isolates from Kenyatta National Hospital showed significantly high level of phenotypic resistance. Concordant with the high level of phenotypic resistance, we found high carriage of the ESBL genes among the isolates analysed in this study. Moreover, majority of isolates harboured all the four ESBL genes. Conclusion: A high rate of phenotypic and genetic resistance was detected among the tested isolates. Resistance to cephalosporins was primarily driven by acquisition of the ESBL genes. The high-rate multidrug resistance in ICU bacterial isolates in this study has a crucial implication for ICU patient management and general antibiotics use.", "keywords": [ "Intensive Care Unit", "anti-biotic susceptibility", "gram-negative", "cephalosporins", "Extended Spectrum Beta Lactamase" ], "content": "Background\n\nThe intensive care unit (ICU) is a hotspot of nosocomial infections primarily because of the extremely vulnerable population of critically ill patients, usage of invasive procedures such as catheters and ventilators1,2 and immunosuppressive medication.3 These infections significantly increase the burden of bacterial associated morbidity, mortality, and healthcare costs. ICU acquired infections (ICU-AI) contribute 20-25% of all nosocomial infections globally.4 Recent studies have reported high risk of bloodstream infections caused by gram-negative bacteria, such as Escherichia coli and Klebsiella pneumoniae among COVID-19 patients admitted in ICU.5,6\n\nAntimicrobial resistance (AMR) is a major contributor to the problem of ICU acquired infections. AMR reduces the effectiveness of antibiotics and other antimicrobial drugs in treating these infections. Emergence of AMR leads to a higher risk of treatment failure, longer hospital stays, and increased mortality rates, as well as greater healthcare costs and resource utilization.7 Drug resistant bacterial pathogens emerge and spread in the ICU environment as a result of acquisition of mutations, and selection of resistant strains, driven by indiscriminate use of antibiotics.8 Additionally, gram-negative bacteria have evolved an intrinsic mechanism involving the production of extended spectrum beta lactamases (ESBLs) that breakdown the beta lactam antibiotics.9 Resistance to antibiotics can be classified into: multidrug resistance (MDR), extensive drug resistance (XDR) and pan drug resistance (PDR) to reflect increasing number of antimicrobial agents affected by the resistance mechanism.10 The outbreak and spread of COVID-19 also contributed to spread of drug resistant bacterial infections in ICU due to the increased number of patients requiring ICU admission. A high prevalence of bacterial pneumonia, 44% (n= 716) among covid 19 patients admitted in ICU has been reported.11\n\nPhenotypic resistance to the third generation cephalosporins (cefotaxime, ceftazidime and ceftriaxone) has been reported.12,13 This resistance poses a significant public health threat since cephalosporins are valuable agents used in the management of a wide range of gram-negative infections including meningitis, Lyme disease, pseudomonas pneumonia, gram-negative sepsis, streptococcal endocarditis, melioidosis, penicillinase-producing Neisseria gonorrhoea, and gram-negative osteomyelitis.14 Moreover, application of molecular tools to profile the ESBLs producing gram-negative bacteria have confirmed the presence of multiple ESBL genes in isolates of Klebsiella pneumonia, Escherichia coli, and Proteus species, corresponding to high-level resistance to third generation cephalosporins.1\n\nThis study sought to profile phenotypic and genetic resistance to cephalosporin in bacteria isolated from ICU patients’ samples. Identification of bacterial species and phenotypic susceptibility patterns were conducted using VITEK 2 (bioMérieux). Phenotypically resistant isolates were confirmed by PCR genotyping.\n\n\nMethods\n\nThis was a cross sectional study carried out between January to June 2021 at Kenyatta National Hospital (KNH). KNH is the largest public referral and teaching hospital in Kenya with a bed capacity of approximately 1800. The hospital serves patients from the capital city with a population of over three million people. The hospital’s critical care unit department is composed of the main ICU and several other specialised units including Neurosurgery-CCU, Medical wards-CCU, Surgical ward-CCU, Neonatal-ICU, and the Casualty CCU. In this study, “ICU” to refers to both main ICU and other specialized CCUs.\n\nThis study was approved by the Kenyatta National Hospital (KNH)-University of Nairobi (UON) Ethics and research committee under the study number: P632/11/2020. Additionally, informed consent/assent were sought from participants or kin of the patient in cases of minors or unconscious patients. Written consent was obtained from next of kin for all participants but two. The two cases involved consent obtained from treating ICU physician, where the patients were incapacitated and their next of kin were unavailable to give consent. This decision was made based on the deferred consent principle backed by the following reasons\n\n1. The research involves minimum harm to the participant\n\n2. The deferment of consent procedure did not adversely affect the rights and welfare of the patient since the genomic testing (PCR) was carried out on the leftover bacterial isolates and not on the human DNA. These bacterial isolates are regarded as residual laboratory samples material\n\nPatient confidentiality and data privacy was ensured by assigning unique study code to each participant. Participant metadata was collected using password protected excel data collection tool.\n\nStudy participants included all patients admitted to various ICUs in KNH suspected to have bacterial infection during their entire period of admission. Inclusion criteria included having a gram-negative culture positive specimen. Patients with only gram-positive cultures were excluded. Sample size was determined using the Cochrane’s and Finite population correction for proportions formula.15\n\nSample quality and quantity were reviewed prior to labelling for bacteriology assessment. Degraded samples or those with inadequate volume were excluded. Samples that passed the inclusion criteria were processed for organism identification and antimicrobial susceptibility of culture positive gram-negative isolates using the Vitek®2 (Biomérieux, Marcy l’Etoile, France) with Minimum Inhibitory Concentration (MIC) breakpoints set according to CLSI 2020 guidelines. Prior to loading isolates into the VITEK® 2, bacterial suspensions were prepared by emulsifying the isolates in 0.5% saline and standardizing turbidity to 0.5 McFarland’s using a densitometer. The suspension was used for species identification, AST and phenotypic detection of ESBL producing organisms in the VITEK® 2 using gram-negative cards (GN83). Vitek®2 Advanced Expert System (AES) was used. Antimicrobial susceptibility profiles for Cefotaxime, ceftazidime and ceftriaxone were also recorded. For specimens identified phenotypically as ESBL producers, another inoculum was picked from residual specimen and stored in skimmed milk-tryptone-glucose-glycerol broth at -80°C to minimize risk of mutations during batching, awaiting PCR.\n\nIsolates that showed phenotypic resistance to Cefotaxime, ceftazidime and ceftriaxone were selected and used for subsequent PCR genotyping. The Isolate II Genomic DNA kit (Bioline London, UK) was used for total DNA extraction. The kit applies affinity columns to extract genomic DNA. Proteinase K, together with cell lysis buffers containing chaotropic salt ions are used to lyse cells releasing gDNA, which is captured by the affinity resins (silica gel membrane). DNA extraction was followed according to manufacturer’s instructions and eluted in a final volume of 40 ul PCR amplification was then performed using MyTaq™ PCR mix (Bioline, London, UK) in a final volume of 20μl, comprising a master-mix, 0.4 μM of each forward and reverse primers and 3 μl of DNA template. Primers specific to ESBL encoding genes (tem, shv, ctmx and oxa) were used as described by.16,17\n\nBriefly, amplicons were analysed by gel electrophoresis run in 1% agarose gel, 1×TAE buffer and SYBR™ Safe (Invitrogen, Carlsbad, CA, USA) and a 1KB ladder at 70 volts for 30 minutes. The amplified products were visualized under Ultraviolet trans-illumination using the UVTEC Gel Documentation Systems (Cleaver Scientific, United Kingdom,) to identify presence of ESBL genes. The primer sequences and thermocycling conditions used in this study are provided in the Supplementary table 1 and Supplementary table 2 in the Data Availability section (DOI: 10.6084/m9.figshare.22369975).\n\nStatistical analyses were performed in MS. Excel 2010 and GraphPad Prism (version 8.0.4). Shapiro-Wilk test was used to assess data normality prior to analyses. Descriptive statistics including means and frequencies were used for data summary. Mean comparisons among three or more groups was performed using one-way ANOVA with Tukey’s post-hoc. Descriptive data was presented as mean ± SD and data considered statistically significant at p value <0.05.\n\n\nResults\n\nA total of 168-gram-negative isolates were phenotypically identified from ICU patients’ samples. The isolates comprised of 8 gram-negative bacteria species, with A. baumanii being the most abundant (35%) followed by K. pneumoniae (24%), and E. coli, 18% while the remaining species were present at frequencies ≤10% (Figure 1).\n\nPhenotypic susceptibility analysis revealed high level of resistance among the bacterial isolates identified. Overall, 101/168 (60.1%) isolates were ESBL producers while 67/168 (39.9%) were ESBL non-producers. Tem was the most abundant ESBL, occurring in 99/168 followed by shv (88/168), ctmx (81/168), and oxa (54/168) (Table 2). However, while tem was produced by most of the organisms, the differences in the differences in number of ESBL was not statistically significant (Table 1 and Figure 2).\n\nThe highest number of bacteria were isolated from tracheal aspirate (TA) (99/168) followed by urine (38/168) and blood (19/168) while ascitic tap, CVC tip and sputum had one isolates each. Species distribution analysis showed that A. baumanii were the highest in TA (38/99) Table 2.\n\nMajority of patients 76% were males and the highest number of bacterial isolates were from patients aged between 21 to 40 years 75/168 and 50 out of the 75 isolates were phenotypically resistant to at one cephalosporin. Conversely, few isolates (3/168) were isolated from patients aged >80 years; all the isolates were phenotypically susceptible to all tested cephalosporins (Table 3).\n\nThe susceptibility pattern revealed high level of phenotypic resistance against three cephalosporins (Ceftazidime, Ceftriaxone, and Cefotaxime) (Table 4).\n\nThe 101 isolates that were phenotypically resistant to cephalosporin were subjected to PCR genotyping and 97 (96%) isolates harboured at least one of the four gene tested while only two (1.9%) isolates were negative for all the four genes. These results confirmed the phenotypic identification results by VITEK 2 (Table 5).\n\n\nDiscussion\n\nBacterial infection in the ICUs represent a major burden and safety concern for patients admitted to the ICU.18 Patients in ICU are often critically ill and require urgent care. As a result, they are prescribed antimicrobial therapy empirically to manage their condition while waiting for culture result.4 The World Health Organization (WHO) considers this irrational use of antimicrobial in ICU a major contributor to development of antimicrobial resistance.19 In light of the rampant use of antibiotics in ICU, this study was conducted to evaluate the level of bacterial colonization in various sample types drawn from ICU patients and the corresponding level of antibiotic resistant gram-negative bacteria. Additionally, susceptibility to three classes of cephalosporins (Ceftazidime, Ceftriaxone and Cefotaxime) was assessed.\n\nAcinetobacter baumanii, Klebsiela pneumoniae and E. coli were the most abundant organisms (35%, 24%, and 18% respectively). The current study corroborates previous findings that Acinetobacter species (30.9%) and Klebsiella species (29.7%) followed by Pseudomonas aeruginosa (22.9%) were the most abundant organisms in ICU environment.20 In yet another study, Pseudomonas species was found to be high (29.1%) in ICU setting followed by Acinetobacter (27.5%).21 The trend in ICU bacterial colonization appears to be dominated by the three main organisms Acinetobacter species, Klebsiella species and Pseudomonas species, as demonstrated in previous studies20,21 and corroborated by our study. Unsurprisingly, we reported Acinetobacter baumanii and Klebsiela pneumoniae as the most common organisms in ICU and resistant to all tested cephalosporins. The resistance to multiple cephalosporins might partially explain the high abundance of these bacteria in ICU. Our findings were in agreement with Saxena and colleagues’ findings that Acinetobacter and Klebsiella were resistant to multiple antibiotics.4\n\nOrganism distribution varied significantly among different specimen types. Tracheal aspirate had the highest isolates (59%) followed by urine (23%) and blood (11%) while ascitic tap, CVC tip and sputum had (0.6%) each. These findings agreed with high prevalence (56%) of pulmonary colonization among ICU patients identified by tracheal aspirate culture.22 Tracheal aspirate culture has been evaluated as a non-invasive method for diagnosis of ventilator-associated pneumonia colonization.23 The ease of obtaining tracheal aspirate sample and availability of established protocol could explain why more tracheal aspirate samples were obtained and cultured successfully. Urine, blood and pus swabs yielded 23%, 11% and 5% of total organisms respectively. The lower proportion of culture positivity could be influenced by the small number of samples as well as the culture method.\n\nConcordant with phenotypic susceptibility findings, we reported high level of genetic resistance in A. baumanii, K. pneumoniae and E. coli. A. baumanii is an opportunistic nosocomial pathogen that presents resistance to most antimicrobial.24 This ability makes it the most persistent bacteria in ICU and has been linked to ventilator-associated pneumonia.25 Carbapenem resistance in A. baumanii is mediated by class D β-lactamases belonging to OXA-type. Moreover, A. baumanii possesses an intrinsic chromosomally encoded oxacillinase blaOXA-51, which may account for the high prevalence of blaOXA (48.8%) reflecting its ability to resist eradication.26 We report 57.6% of K. pneumoniae isolates that possess OXA gene. Similar findings were recently reported demonstrating the involvement of blaOXA gene in mediating resistance to cephalosporins.27\n\nAnalysis of blaCTMX, blaTEM, and blaSHV genes revealed a high resistance gene carriage in more than 50% of studied isolates. High prevalence of these markers had been reported previously in an Indonesian hospital.28 Moreover, molecular surveillance of ESBL in neonates drawn from Kenya and Nigeria revealed a high prevalence of ESBL producing bacteria.29 The high prevalence of ESBL producing bacteria in ICU underscore the need to heighten antibiotic resistance surveillance to provide the much-needed information to combat resistance to antibiotics.\n\nThe problem of increasing antimicrobial resistance in ICU is worrisome particularly due to the fragile nature of this category of patients. While there are no definitive measures to eradicate antibiotic resistant micro-organisms in ICUs, vaccines against these pathogens remain elusive and where available, they are unaffordable. Thus, prudent use of antibiotics in ICU to avoid widespread resistance is recommended. This study highlights the abundance of cephalosporin resistant gram-negative bacteria in ICU, which emphasized the need to heighten the fight against antibiotic resistance.", "appendix": "Data availability\n\nFigshare. Phenotypic and genetic Extended Spectrum Beta Lactamase cephalosporin resistance profiles of bacterial isolates from ICU in Tertiary Level Hospital in Kenya. DOI: https://doi.org/10.6084/m9.figshare.22369975.v2 (Mwale, 2023).\n\nThis project contains the following data:\n\nExtended Spectrum Beta Lactamase.xlsx: The data contain phenotypic antibiotic susceptibility values for bacterial isolates and genotypic resistance data assessed by detection of ESBL genes is also part of the data.\n\nRaw DATA_VITEK Bacterial identification.xlsx: Bacterial identification readings from VITEK 2.\n\nSupplementary Materials.docx: This file contains the PCR primer sequences and thermocycler conditions.\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgement\n\nThe authors thank the study participants and their families.\n\n\nReferences\n\nBrusselaers N, Vogelaers D, Blot S: The rising problem of antimicrobial resistance in the intensive care unit. Ann. Intensive Care. 2011; 1: 1–7. Publisher Full Text\n\nWu D, Wu C, Zhang S, et al.: Risk factors of ventilator-associated pneumonia in critically III patients. Front. Pharmacol. 2019; 10: 482. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYang X, Yu Y, Xu J, et al.: Clinical course and outcomes of critically ill patients with SARS-CoV-2 pneumonia in Wuhan, China: a single-centered, retrospective, observational study. Lancet Respir. Med. 2020; 8: 475–481. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSaxena S, Priyadarshi M, Saxena A, et al.: Antimicrobial consumption and bacterial resistance pattern in patients admitted in ICU at a tertiary care center. J. Infect. Public Health. 2019; 12: 695–699. PubMed Abstract \| Publisher Full Text\n\nSaharman YR, Karuniawati A, Severin JA, et al.: Infections and antimicrobial resistance in intensive care units in lower-middle income countries: a scoping review. Antimicrob. Resist. Infect. Control. 2021; 10: 22. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGiannitsioti E, Louka C, Mamali V, et al.: Bloodstream infections in a COVID-19 Non-ICU department: Microbial epidemiology, resistance profiles and comparative analysis of risk factors and patients’ outcome. Microorganisms. 2022; 10: 1314. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTran GM, Ho-Le TP, Ha DT, et al.: Patterns of antimicrobial resistance in intensive care unit patients: a study in Vietnam. BMC Infect. Dis. 2017; 17: 429. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nReygaert WC: An overview of the antimicrobial resistance mechanisms of bacteria. AIMS microbiology. 2018; 4: 482–501. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFerreira CM, Ferreira WA, Almeida NCOS, et al.: Extended-spectrum beta-lactamase-producing bacteria isolated from hematologic patients in Manaus, State of Amazonas, Brazil. Braz. J. Microbiol. 2011; 42: 1076–1084. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCantón R, Gijón D, Ruiz-Garbajosa P: Antimicrobial resistance in ICUs: an update in the light of the COVID-19 pandemic. Curr. Opin. Crit. Care. 2020; 26: 433–441. Publisher Full Text\n\nConway M, Andrew, Kohler K, et al.: Co-infection and ICU-acquired infection in COVID-19 ICU patients: a secondary analysis of the UNITE-COVID data set. Crit. Care. 2022; 26: 236. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLester R, Musicha P, Van Ginneken N, et al.: Prevalence and outcome of bloodstream infections due to third-generation cephalosporin-resistant Enterobacteriaceae in sub-Saharan Africa: a systematic review. J. Antimicrob. Chemother. 2020; 75: 492–507. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNagassar RP, Daniel K, Bridgelal-Nagassar RJ, et al.: Surveillance of phenotypic extended spectrum beta-lactamase resistance in blood isolates at a Hospital in East Trinidad. Caribb. Med. J. 2021. Publisher Full Text\n\nArumugham VB, Gujarathi R, Cascella M: Third generation cephalosporins.2019.\n\nIsrael GD: Determining sample size.1992.\n\nKiiru J, Kariuki S, Goddeeris BM, et al.: Analysis of β-lactamase phenotypes and carriage of selected β-lactamase genes among Escherichia coli strains obtained from Kenyan patients during an 18-year period. BMC Microbiol. 2012; 12: 111–155. Publisher Full Text\n\nSaisi H, Makobe C, Kangongo M, et al.: Prevalence of CTXM, SHV, TEM AND OXA Genes among Extended-Spectrum Beta-Lactamase Producing Klebsiella pneumoniae from Mukuru Slum, Kenya. Adv. Microbiol. 2019; 09: 853–862. Publisher Full Text\n\nPatil RK, Kabera B, Muia CK, et al.: Hospital acquired infections in a private paediatric hospital in Kenya: a retrospective cross-sectional study. Pan Afr. Med. J. 2022; 41.\n\nWHO: Regional strategy on prevention and containment of antimicrobial resistance, 2010.2010. Reference Source\n\nMehta T, Chauhan B, Rathod S, et al.: Bacterilogical profile and drug resistance pattern of isolates of the patients admitted in medical intensive care unit of a tertiary care hospital in Ahmedabad. Med. Sci. 2015; 4: 222–225.\n\nBarai L, Fatema K, Haq JA, et al.: Bacterial profile and their antimicrobial resistance pattern in an intensive care unit of a tertiary care hospital of Dhaka. Ibrahim Medical College Journal. 2010; 4: 66–69. Publisher Full Text\n\nMcCauley LM, Webb BJ, Sorensen J, et al.: Use of Tracheal Aspirate Culture in Newly Intubated Patients with Community-Onset Pneumonia. Ann. Am. Thorac. Soc. 2016; 13: 376–381. PubMed Abstract \| Publisher Full Text\n\nScholte JB, Van Dessel HA, Linssen CF, et al.: Endotracheal aspirate and bronchoalveolar lavage fluid analysis: interchangeable diagnostic modalities in suspected ventilator-associated pneumonia?. J. Clin. Microbiol. 2014; 52: 3597–3604. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSanz-García F, Gil-Gil T, Laborda P, et al.: Coming from the wild: multidrug resistant opportunistic pathogens presenting a primary, not human-linked, environmental habitat. Int. J. Mol. Sci. 2021; 22: 8080. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKramer A, Schwebke I, Kampf G: How long do nosocomial pathogens persist on inanimate surfaces? A systematic review. BMC Infect. Dis. 2006; 6: 1–8. Publisher Full Text\n\nRoca I, Espinal P, Vila-Farrés X, et al.: The Acinetobacter baumannii oxymoron: commensal hospital dweller turned pan-drug-resistant menace. Front. Microbiol. 2012; 3: 148. Publisher Full Text\n\nAl-Sheboul SA, Al-Moghrabi SZ, Shboul Y, et al.: Molecular characterization of carbapenem-resistant Acinetobacter baumannii isolated from intensive care unit patients in Jordanian hospitals. Antibiotics. 2022; 11: 835. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCandra IK, Yanto F, Suranadi IW, et al.: Characteristic of Extended Spectrum β-Lactamase-Producing Enterobacteriaceae from Fecal Carriage Isolates of Intensive Care Unit Patients at Sanglah Hospital, Bali, Indonesia. The Open Microbiology Journal. 2021; 15: 1–6. Publisher Full Text\n\nEdwards T, Williams CT, Olwala M, et al.: Molecular surveillance reveals widespread colonisation by carbapenemase and extended spectrum beta-lactamase producing organisms in neonatal units in Kenya and Nigeria. medRxiv, 2022.2001.2006.22268735.2022. Publisher Full Text" }	[ { "id": "217591", "date": "06 Jun 2024", "name": "Upendo O Kibwana", "expertise": [ "Reviewer Expertise Bacteriology", "infectious 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 work is relevant but needs major revision to be more understandable and reproducible See comments below:\n1. The authors have to be mindful of how to write scientific names properly (italicized), how to write gram stain \" Gram\" instead of gram, and correct all the typos. Also how the gens are written, in capital form not small letters.\n2. In the abstract the authors are talking about four genes, please indicate clearly which genes are referred to.\n3. The authors focus on ESBL genes. is OXA and ESBL gene or beta lactam gene? Please clarify and possibly change the title accordingly.\n4. In the abstract the authors are talking about high level of multidrug resistance. Can the authors clarify what MDR is? because this is not indicated anywhere in the manuscript nor is it reflected in the results.\n5. It is not clear if this was a laboratory-based cross-sectional study or simply cross cross-sectional study. At some point, it seems the starting point was in the lab with Gram-negative isolates, while in the population section, the authors indicate they started in the ward (ICU) please indicate clearly. If the study started in the lab what was the rationale for obtaining informed consent rather than requesting a waiver for informed consent? If the study started from the ward how did you exclude pats with Gram-positive isolates prior to sample collection?\n6. The authors have to explain how they obtained isolates which subjected to identification test on VITEK. Please indicate all quality control measures observed.\n7.How many antibiotics were tested during AST? Provide the names and conc.\n8. The results section has to be rearranged to provide a better narrative. Start with where the isolates were obtained from, which isolates were isolated from where, then narrow down to the isolated ESBL.\n9. Most of the results sections can be improved to describe better what is presented in the tables. e.g. the bacterial abundance section can be improved by adding information on the isolates which were found in one type of specimen only. The ESBL\n\nproduction and different production parameters section can be improved by adding information about other parameters; Would be good to perform a chi-square test to see if the differences between the parameters are significant or coincidental due to numbers.\n10. genotypic susceptibility section has to be described more; how many isolates carried one gene only? How many multiple? Which was the most combo of genes? In which isolates were more prominent with multiple genes etc. Also why 2 isolates were negative and not 4? 97 were positive.\n11. \"The susceptibility pattern revealed high level of phenotypic resistance against three cephalosporins (Ceftazidime, Ceftriaxone, and Cefotaxime)\" This sentence seems ectopic where it is placed.\n12. In the discussion section the authors should mention the settings of the studies that they make comparisons with e.g. in paragraph two.\n13. \"Unsurprisingly, we reported Acinetobacter baumanii and Klebsiela pneumoniae as the most common organisms in ICU and resistant to all tested cephalosporins\" This statement appears in the discussion section yet it is not indicated anywhere in your result section. Please discuss the things that are from your results.\n14. Table 1 'n' should be 'total'.\n15. Table 2 should be table 1. This is based on a comment about rearrangement.\n16. \"Urine, blood and pus swabs yielded 23%, 11% and 5% of total organisms respectively. The lower proportion of culture positivity could be influenced by the small number of samples as well as the culture method.\" Please explain the about the culture methods referred to in this statement.\n17. Can the authors compare their genotypic results with neighbor countries and give clinical significance.\n18. I advice the authors to remove the footnotes on the tables because they are rather captions and not footnotes\n19. Table 4 is not clear. What are the numbers provided? Percentages or numbers? Provide numbers and percentages. Table not clear? What about other abx? What is the number of tested isolates for each species?\n20. The title for table 5 is not clear. What information does the table 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? 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": "12806", "date": "10 Dec 2024", "name": "Edwin Magomere", "role": "Author Response", "response": "Reviewer 1 comments Comment 1: The authors have to be mindful of how to write scientific names properly (italicized), how to write gram stain \" Gram\" instead of gram, and correct all the typos. Also how the genes are written, in capital form not small letters. Response: All the typos have been corrected throughout the manuscript and gene names written in the right format Typos: “gram stain” corrected to “Gram stain”, Italicized all scientific names Specific gene names have been added to the abstract: blaTEM, blaCTX-M, blaSHV and blaOXA. Comment 2: In the abstract the authors are talking about four genes, please indicate clearly which genes are referred to. Response: the genes we are referring to include: blaTEM, blaCTX-M, blaSHV and blaOXA. These have been added to the abstract. Comment 3: The authors focus on ESBL genes. is OXA and ESBL gene or beta lactam gene? Please clarify and possibly change the title accordingly. Response: OXA is an ESBL gene. Other ESBL genes that we studied include: (blaCTX-M, blaSHV, blaTEM). We focused on four ESBL genes. Response: 4. In the abstract the authors are talking about high level of multidrug resistance. Can the authors clarify what MDR is? because this is not indicated anywhere in the manuscript nor is it reflected in the results. Response: We have added the definition of MDR: Lines in the background section. The last sentence of conclusion under abstract has been rephrased to read as: “The high prevalence rate of ESBL genes in ICU bacterial isolates shown in this study has a important implication for ICU patient management and general antibiotics use”. Comment 5: It is not clear if this was a laboratory-based cross-sectional study or simply cross cross-sectional study. At some point, it seems the starting point was in the lab with Gram-negative isolates, while in the population section, the authors indicate they started in the ward (ICU) please indicate clearly. If the study started in the lab what was the rationale for obtaining informed consent rather than requesting a waiver for informed consent? If the study started from the ward how did you exclude pats with Gram-positive isolates prior to sample collection? Response: The starting point of the study was the laboratory. However, we used patient samples that had been collected from ICU patients for clinical purposes. Since the samples were not originally collected for research purposes, KNH-UON Ethical review committee tasked us to obtain consent before samples could be used for research purposes. We also needed patient consent to allow us collect patient information such as co-morbidities. We used differential media to allow only Gram-negative bacteria to grow. Thus, we used only Gram- negative cultures for our downstream experiments. Comment 6: The authors have to explain how they obtained isolates which subjected to identification test on VITEK. Please indicate all quality control measures observed. Response: Samples brought to the laboratory requested by the ICU clinical team were cultured and subjected to Gram stain. Only Gram-negative isolates were analyzed on VITEK. We used commercially acquired Gram negative isolates as positive controls for each run on Vitek. For negative controls we used bacterial suspension media (saline). Comment 7: How many antibiotics were tested during AST? Provide the names and conc. Response: Antibiotics tested in this study include Cefotaxime, ceftazidime and ceftriaxone. The list of tested antibiotics has been added to the manuscript. The Minimum Inhibitory Concentrations (MICs) were set according to CLSI 2020 guidelines. Comment 8: The results section has to be rearranged to provide a better narrative. Start with where the isolates were obtained from, which isolates were isolated from where, then narrow down to the isolated ESBL. Response: The results have been rearranged to start with various sources of isolates as advised Comment 9: Most of the results sections can be improved to describe better what is presented in the tables. e.g. the bacterial abundance section can be improved by adding information on the isolates which were found in one type of specimen only. The ESBL production and different production parameters section can be improved by adding information about other parameters; Would be good to perform a chi-square test to see if the differences between the parameters are significant or coincidental due to numbers. Response 1: we have added more information about species identified in specimen types with the highest number of species including the following: T/A harboured all the isolated tested, with a total of 99 isolates. The distribution of species in T/A showed that A. baumanii were the highest in TA (38/99). Urine specimen had the second highest number of species (38/168). Out of the 8 species identified, urine had 5 species, with K. pneumoniae being the most frequent, identified 13 times. Response 2: Table 3 has been replaced to show the outcome of Chi-Square analysis to show differences in various parameters Comment 10: genotypic susceptibility section has to be described more; how many isolates carried one gene only? How many multiple? Which was the most combo of genes? In which isolates were more prominent with multiple genes etc. Also why 2 isolates were negative and not 4? 97 were positive. Response: Added information on . blaTEM was the most predominant gene at 96% (97/101), followed by SHV = 85.6% (86/101), CTX-M = 78.8% (80/101) and OXA= 52.9% (54/101). blaTEM/blaSHV/blaCTX-M/blaOXA and blaTEM/blaSHV/blaCTX-M gene combinations were present at 49% (n=49/101) and 25.7% (n=26/101) respectively. Other common gene combinations included blaTEM/SHV at 6.7% (n=7/101), blaTEM/CTX-M/OXA at 1.9% (n=2/101), blaTEM/SHV/OXA at 1.9% (n= 2/101), blaSHV/CTX-M at 1.9% (n= 2/101). Comment 11: \"The susceptibility pattern revealed high level of phenotypic resistance against three cephalosporins (Ceftazidime, Ceftriaxone, and Cefotaxime)\" This sentence seems ectopic where it is placed. Response: This sentence has been deleted as advised Comment 12: In the discussion section the authors should mention the settings of the studies that they make comparisons with e.g. in paragraph two. Response: The settings of previous studies have now been mentioned in the discussion section Comment 13: \"Unsurprisingly, we reported Acinetobacter baumanii and Klebsiela pneumoniae as the most common organisms in ICU and resistant to all tested cephalosporins\" This statement appears in the discussion section yet it is not indicated anywhere in your result section. Please discuss the things that are from your results. Response: This sentence has been deleted Comment 14: Table 1 'n' should be 'total'. Response: “n” has been edited to “Total” Comment 15: Table 2 should be table 1. This is based on a comment about rearrangement. Response: Tables have been rearranged accordingly Comment 16: \"Urine, blood and pus swabs yielded 23%, 11% and 5% of total organisms respectively. The lower proportion of culture positivity could be influenced by the small number of samples as well as the culture method.\" Please explain the about the culture methods referred to in this statement. Comment 17: Can the authors compare their genotypic results with neighbor countries and give clinical significance. Response: Results have been compared to those obtained from neighboring countries (Tanzania and Uganda) and Clinical significance highlighted. Comment 18: I advice the authors to remove the footnotes on the tables because they are rather captions and not footnotes Response: Footnotes have been removed from all tables as advised Comment 19: Table 4 is not clear. What are the numbers provided? Percentages or numbers? Provide numbers and percentages. Table not clear? What about other abx? What is the number of tested isolates for each species? Response: Number provided in the table is number of isolates in the categories of S-susceptible, I-intermediate and R-resistant. This has been clarified in the revised manuscript. Comment 20: The title for table 5 is not clear. What information does the table present? Response: The title has been edited to read as “The isolates that had resistance to all tested cephalosporins and the frequency of resistance genes”." } ] }, { "id": "316326", "date": "16 Oct 2024", "name": "Tajudeen Oladunni GANIYU", "expertise": [ "Reviewer Expertise Medical Microbiology and Microbial Biotechnology" ], "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 research concept is okay and relevant to the scope of the journal but the manuscript needs major revision. My comments are listed below:\nThe authors should rewrite the abstract to include a clear statement of problem and justification. The conclusion is not emphatic on what needed to be done to address the problem. ESBL are more than four, what are the reasons for limiting the number of gene assessed to just four? State the four genes correctly in the abstract. In the background, third paragraph and third sentence- presence of multiple ESBL genes in isolates of Klebsiella pneumoniae, Escherichia coli and Proteus species- please list the ESBL genes. In the Results\n- how did you differentiate ESBL producers from non ESBL producers? - tem should be written as TEM, shv should be SHV, ctmx should be CTX-M and oxa should be OXA. 5. In the second paragraph of the results (Phenotypic susceptibility and ESB production- the last sentence, - the differences in the differences in number of ESBL- delete in word in red. 6. Discussion. - the discussion is poor, despite explaining and interpreting the results, the authors failed to compare their results with those of other authors. Comparing their work with those of other authors will bring out the novelty in their work and also show whether their results are better compared to earlier works.\nOTHERS E. coloacae should be Enterobacter cloacae\nREFERENCES\nReferences earlier than 2019 should be updated.\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? Partly", "responses": [ { "c_id": "12804", "date": "10 Dec 2024", "name": "Edwin Magomere", "role": "Author Response", "response": "Comment 1: The authors should rewrite the abstract to include a clear statement of problem and justification. The conclusion is not emphatic on what needs to be done to address the problem. Response: We have rewritten the abstract to include the problem statement in the abstract, which now reads as follows: Problem statement: Bacterial infections in the Intensive Care Units are a threat to the lives of critically ill patients. Their vulnerable immunity predisposes them to developing bacteria-associated sepsis, deteriorating their already fragile health. In the face of increasing antibiotics resistance, the problem of bacterial infection in ICU is worsening. Justification: Surveillance of bacterial infections in ICUs and drug resistance will help to understand the magnitude of the problem it poses and inform response strategies. Comment 2: ESBL are more than four, what are the reasons for limiting the number of gene assessed to just four? State the four genes correctly in the abstract. Response: These four ESBL genes were chosen for PCR genotyping since they were most frequently detected based on phenotypic resistance detection. Also, we restricted our study to 4 genes due to financial constraints Comment 3: In the background, third paragraph and third sentence- presence of multiple ESBL genes in isolates of Klebsiella pneumoniae, Escherichia coli and Proteus species- please list the ESBL genes. Response: We have added the various ESBL genes as recommended and included a reference to it. Comment 4: How did you differentiate ESBL producers from non ESBL producers? Response: We used the Phenotypic detection of extended-spectrum β-lactamase production. Specifically, we used the automated VITEK 2 ESBL test (Biomérieux, Marcy l’Etoile, France). Description of how the method work This method is based on the simultaneous assessment of the antibacterial activity of cefepime, cefotaxime and ceftazidime, measured either alone or in the presence of clavulanate. This test relies on card wells containing 1.0 mg/L of cefepime, or 0.5 mg/L of cefotaxime or ceftazidime, either alone or associated with 10 or 4 mg/L of clavulanate, respectively. After inoculation, cards are introduced into the VITEK 2 machine, and for each antibiotic tested, turbidity is measured at regular intervals. The proportional reduction of growth in wells containing a cephalosporin combined with clavulanate is then compared with that achieved by the cephalosporin alone and is interpreted as ESBL-positive or – negative through a computerized expert system. We have also edited the subtitle on phenotypic detection to \"Antimicrobial Susceptibility Testing (AST) and phenotypic detection of ESBL producers\". Comment 5: tem should be written as TEM, shv should be SHV, ctmx should be CTX-M and oxa should be OXA. Response: We have edited the names of ESBL gene in all instances where they were not written in the right format. Thank you for drawing my attention to this. Comment 6: In the second paragraph of the results (Phenotypic susceptibility and ESB production- the last sentence, - the differences in the differences in number of ESBL- delete in word in red. Response: We have deleted the words in red and re-written the sentence to read “blaTEM was produced by most of the organisms but there were no statistically significant difference when compared to other ESBLs ” Comments: Discussion. - the discussion is poor, despite explaining and interpreting the results, the authors failed to compare their results with those of other authors. Comparing their work with those of other authors will bring out the novelty in their work and also show whether their results are better compared to earlier works. Response: We have edited the discussion to discuss our findings in context of other works in the same area and added more references. Throughout the discussion, we have endeavored to compare results to previous studies." } ] } ]	1	https://f1000research.com/articles/12-469
https://f1000research.com/articles/12-183/v1	16 Feb 23	{ "type": "Research Article", "title": "Upholding dignity during a pandemic via Twitter", "authors": [ "Michael Mulvey", "Tracey O'Sullivan", "Sarah Fraser", "Tracey O'Sullivan", "Sarah Fraser" ], "abstract": "Background: This article investigates how people invoked the concept of dignity on Twitter during the first year of the COVID-19 pandemic, with a secondary focus on mentions of dignity in the context of older adults and ageing. Methods: We report the results of a study that combines text analytic and interpretive methods to analyze word clusters and dignity-based themes in a cross-national sample of 1,946 original messages posted in 2020. Results: The study finds that dignity discourse on Twitter advances five major themes: (a) recognize dignity as a fundamental right, (b) uphold the dignity of essential workers, (c) preserve the dignity of at-risk populations, (d) prevent cascading disasters that exacerbate dignity's decline, and (e) attend to death, dignity, and the sanctity of life. Conclusions: Moreover, messages focusing on older adults lamented the disproportionate death toll, the terrible circumstances in long-term care homes, the added impact of suspended meal delivery services and the status of older people living below the poverty line.", "keywords": [ "discourse analysis", "social media", "text analytics", "human rights", "vulnerability" ], "content": "Introduction\n\nDignity is a concept considered by many to be a fundamental human right, reflected in many legal and ethical frameworks around the world, including the Universal Declaration of Human Rights, which states that “all human beings are born free and equal in dignity and rights” (United Nations, 1948). Those who uphold dignity as a human right often argue that it is an essential component of a just and fair society and should be protected and upheld by governments and other institutions.\n\nThis paper investigates how people on social media perceive and express their views on human dignity in the context of the COVID-19 pandemic. To do this, we analyzed a large sample of Twitter posts and comments from the first year of the pandemic to understand how people respond to situations that may involve the violation of dignity. Many studies have shared insight into dignity, drawing perspectives from fields including law (Cunningham, 2020), public policy and politics (Nilsson et al., 2022), bioethics (Faneye, 2014), and healthcare (Fernández-Sola et al., 2012). However, studies usually focus on expert views, not how the general public expresses thoughts about dignity.\n\nExamining data from social media has many advantages, as it provides unprompted real-time snapshots of how people think and feel about a topic (Belk, Fisher, and Kozinets, 2013; Berger et al., 2020). It also provides a large and diverse user base from different backgrounds, ages, and locations. Moreover, access to a wide range of archived data (including text, images, and videos) can deliver a large sample of data that is: timely, convenient, cost-effective—and, most importantly—rich and nuanced insights into perceptions. Organizations can then use this information to formulate policy, improve their products and services, and develop more effective marketing and communication strategies.\n\nIn the following sections, we describe the theoretical foundations of our approach and report the results of a study of how people talked about dignity on Twitter during the pandemic. Finally, we discuss the value of studying public perceptions of dignity and how crowdsourced views on dignity may inform policies and practices that aim to promote and protect dignity in various settings, such as workplaces, healthcare facilities, and other social settings.\n\n\nTheoretical background\n\nWhile it is clear that dignity and human rights are closely associated, it is important to consider that dignity encompasses: how a person feels (their self-worth) and the context in which they live (i.e., psychological, social and cultural factors) (Mégret & Hoffmann, 2009). Furthermore, dignity is socially constructed as it develops and is dependent on social interaction between individuals or groups, and it is also determined by and intersects with other human rights (Mégret & Hoffmann, 2009). For these reasons, it is likely that individuals or groups that are already marginalized or considered 'vulnerable' (i.e., older adults, 2SLGBTQI+, Indigenous people, immigrants, etc.) are at a greater risk of losing their dignity and to be treated as “not human or less than human–as a thing or instrument or subhuman creature” (Kateb, 2014). Indeed, in the context of COVID-19, initially, the discourse surrounding older adults living in long-term care homes focused heavily on the loss of dignity in healthcare and in dying (Carrieri et al., 2020; Colombo, 2021; Kowsalya & Sundara, 2021, Vellani et al., 2021). In general, much of the discourse was ageist, promoting notions that all older adults need protection, are vulnerable and should be socially isolated (Fraser et al., 2020, Lagacé et al., 2020) –all factors which infringe on human rights and dignity. The impact of COVID-19 on human dignity has been negative, but it has also raised awareness about dignity and the importance of including it as a priority in health system reform (Galea, 2021).\n\nSociologist Bruno Latour (2007) examines how people and groups within a society make sense of the world and communicate their understanding to others. He studies how language and other forms of representation shape social interactions and relationships, and how they influence the way people construct and communicate their worldviews. Similarly, researchers who examine media discourse and public opinion also focus on the symbols and reasoning devices used to present and discuss issues, such as metaphors, exemplars, catchphrases, depictions, and visual images, as well as the roots, consequences, and appeals to principle that suggest how to think about an issue (Gamson and Modigliani, 1989).\n\nWith increasing convergence of online and offline realms, digital methods of studying communication behaviour have become helpful in gaining insight into people's experiences (Caliandro, 2018) and sentiments or shared emotional dispositions (Gopaldas, 2014). As a result, researchers have increasingly turned to social media conversations to understand what people discuss and express. Users post messages on social media that offer a “window into experience” as they share their expertise, exchange questions or concerns, and reinforce each other's positions (Eriksson and Salzmann-Erikson, 2013). This approach has the advantage of “listening in” on conversations and avoiding certain response biases that may occur when asking questions in interviews or surveys (Rappaport, 2011).\n\nApplications of social listening in public health and well-being contexts include studies of patients' reactions to knee replacement surgery (Pitt, Mulvey, and Kietzmann, 2018), gamblers' reflections on problem gambling (Brown, Caruna, Mulvey, and Pitt, 2021), expressions of stigma against people with dementia (Bacsu et al., 2022), public attitudes towards vaccines (Fazel et al., 2021), job satisfaction and turnover (Lam, Mulvey, and Robson, 2022), and retirement travel planning (Mulvey, Padgett, and Lever, 2022), using diverse online data sources including discussion forums, review platforms, Facebook, Twitter, and Reddit. In addition, social media methods are well-suited to collecting and analyzing data rapidly in times of crisis or responding to emerging trends, such as the COVID-19 pandemic (Picone et al., 2020; Reid and Duffy, 2018). Furthermore, advice sharing on social networks is inherently social–people exchange ideas, solicit and deliver advice, and develop relationships with other community members (Kozinets, 2002; Mulvey, Padgett, and Lever, 2022). Like others, we contend that crowdsourcing holds enormous potential to identify problems and share solutions in times of crisis. For example, self-help groups and advice on social media may improve affected populations' resilience during a disaster, “replacing their helplessness with dignity, control, as well personal and collective responsibility” (Keim and Noji, 2011).\n\n\nMethods\n\nOur study examined original messages from Twitter users, a highly inclusive, one-to-many platform where people share information in a relatively synchronous or condensed time frame (Lamberton and Humphreys, 2022). Twitter data gives researchers a window into the volume, sentiment, and expressions of public opinion. We conducted a three-step analysis using data from the Sprout Social COVID-19 database, which holds millions of tweets related to the pandemic. First, we examined key performance metrics of the volume and impact of dignity-related tweets to assess the prevalence of dignity-related tweets in 2020. Second, we downloaded a sample of Twitter data via Sprout Social and used the Leiden community detection algorithm to detect and study themes in dignity discourse, drawing exemplar tweets to illustrate constituent meanings. Finally, we analyzed themes in the subset of dignity tweets that mention older adults. Ethical approval was waived by the University of Ottawa REB on the basis that data that is publicly available on social media and can be viewed by anyone would not require ethics approval. Our methodology is reproducible and should replicate the reported results, yet a different random sample of tweets may yield slightly different results.\n\nOur cross-national sampling plan aimed to include diverse viewpoints and prevent a single country's tweets from dominating the discourse. Therefore, the sample comprised English-language tweets from countries that posted more than 500 tweets in the 12 months of January to December 2020. Seven countries achieved this level of activity. Also, we decided to prioritize tweets with engagement and followers as opposed to those having little or no impact. So, we sorted the posts in descending order on these metrics before downloading samples proportional to each country's total tweet volume. As a result, the sample includes a total of 1,946 original messages drawn randomly from Twitter, scaled to the United States (1,000 messages), India (300), the United Kingdom (286), Canada (157), South Africa (77), Kenya (65), and Australia (61).\n\nThe Leiden community detection algorithm (Traag, Waltman, & Van Eck, 2019) is a state-of-the-art network-based algorithm that can identify clusters of related nodes in a network. In the context of social media data, we use this algorithm to identify groups of concepts that interact with each other in a meaningful way, such as words co-mentioned in sentences and reflecting latent topics or themes.\n\nWe used the Social Astronomy app (Belanger, 2022) to create a network of words (or n-grams) based on their interactions in social media messages (e.g. words are mentioned together in a tweet). Next, Social Astronomy analyzed the network (or matrix) using the Leiden community detection algorithm to identify clusters of words that interact consistently and cohesively. These clusters may correspond to groups of words that refer to topics or themes. Cluster scores assigned by the algorithm helped retrieve original messages for review by research team members.\n\nThe Leiden community detection algorithm results depend on the researcher's selection of resolution parameter setting, which controls the detected communities' granularity and the number of words included in the analysis. A higher resolution parameter will generally detect smaller, more fine-grained communities, while a lower resolution parameter will result in larger, coarser communities. Similarly, including many words will produce a more complex solution than using fewer words. In general, it is helpful to experiment with different values of the resolution parameter to see which one produces the best results given the dataset and analysis objectives.\n\nTo improve the accuracy of group membership detection, we ignored stop words (such as “the”, “a”, “an”, and “in”) and ubiquitous words, including the search keywords (dignity, #dignity), a quotation acronym (qt), and Sprout Social's COVID-19 database inclusion keywords (covid, covid19, #covid19, #coronavirus, corona, virus, coronavirus, covid-19, pandemic, #covid, #covid_19, #pandemic, #covid19uk). Mindful of our objective to report results with high fidelity to the original data, we iteratively generated solutions using different settings before selecting a comparatively detailed and stable 13-cluster solution that used the top 200 words and a 1.5 resolution parameter setting.\n\n\nResults\n\nNext, we present the listening insights derived from an aggregate analysis of dignity tweets followed by an analysis of themes and topics that emerge from the messages. Social media studies often include quotes to give participants a voice, illustrate ideas to build trust, foster replicability, or lend evidence for claims. However, evolving ethical standards attend to the unintended consequences of reporting individuals’ tweets and express concern about the discoverability of messages posted voluntarily in public forums (Mason and Singh, 2022). Accordingly, we honour individuals’ privacy and do not reproduce user IDs or tweets. In contrast, we refer to companies, organizations, and institutions that use Twitter for public relations to share views with stakeholders.\n\nThe prevalence and engagement with dignity-based tweets\n\nThe Sprout Social key performance metrics affirm the popularity of dignity in the reporting period of January to December 2020 and establish that the topic was well-established in COVID-era discourse. The keywords (dignity OR #dignity) appeared in 46,420 original messages (not including shared or mentioned), contributed by 41,060 unique authors, generating 2.89 billion impressions and 5.12 million engagements (total likes, dislikes, comments and shares). These values translate to daily averages of 126.8 messages, 112 new authors, 7.9 million impressions, and 13,991 engagements.\n\nFigure A illustrates the volume and engagement levels of tweets from our sample. There are two panels: the top includes all dignity tweets, excluding dignity tweets focused on older adults, depicted in the bottom panel. Comparatively, the volume of dignity tweets focused on older adults is only 4.4% of the total sample. Each circle denotes a tweet, and its size represents the level of message engagement (the total likes, comments, and shares). Most tweets earned low levels of engagement (small dots), whereas some striking exceptions generated exceptionally high engagement (the circles). For reference, the post with the highest level of engagement (=88,390) generated 56,771 likes, 20,174 comments, and 11,445 shares. Notably, Twitter algorithms favour popular posts, amplifying posts with engagement, feeding them into users' streams and generating more impressions.\n\nThe heat map shading indicates the relative volume of posts by month (red shading represents the highest density, yellow the least). Some observations: dignity discourse on Twitter was sparse in January and February and rose exponentially in March, April, and May following events including the WHO's pandemic declaration, rising hospitalization and death rates, imposition of lockdown measures, and the Black Lives Matter protests. The volume of tweets decreased and maintained steady levels through the summer and fall months and faded into December.\n\nTweets with the most engagements\n\nNext, we focus on the posts that earned top-10 engagement scores. Foremost, politicians championed the cause of dignity, led by a handful of well-known members of the Democratic Party, including Hillary Clinton (#1), Bernie Sanders (#4), Chelsea Clinton (#8, #10), and Nancy Pelosi (#9). In addition, his Holiness Pope Francis (#2, #3, also known as Pontifex), medical doctor Craig Spencer (#5), journalist Miles Howard (#6), and author/retired registered nurse Juanita Broaddrick shared views. These posts demonstrated diverse construals of dignity, invoking tensions between democracy and authoritarianism and respect for persons, including older adults, people who are unemployed, patients and healthcare professionals, small business owners, and leaders (not) doing their duty.\n\nCore and peripheral words in tweets\n\nStill, simple word frequency counts and exemplars do not tell the whole story of dignity discourse. Figures B, C, and D include word clouds created using Scimago Graphica that illustrate the relative frequencies of terms, hashtags, and emojis in messages that include the keyword dignity. For example, Figure B shows the prevalence of terms like pandemic and people and negatively charged words such as corruption, authoritarianism, and lawlessness. Figure C indicates how users attach hashtags to messages, linking posts to related conversations, for instance, #blacklivesmatter, #mentalhealth, and #protectreprodignity. Finally, Figure D displays emojis in messages, using visual symbols to convey abstract emotions and feelings, including anger 😡, disgust 🤮, and sadness 😔, as well as calls to action to wear a mask 😷, attend to disabled persons ♿, and pray for better outcomes 🙏🏾🙏🙏🏻. We also examined mentions of prominent people and brands, signaled by the @ sign, which calls out usernames on Twitter. The list of mentions included the names of many prominent politicians, public health agencies, and news outlets. Increasingly, researchers are attending to hashtags, and emojis as these paratexts that accompany the words may play a prominent role in the reception or interpretation by the public (Bakker, 2022; Luangrath et al., 2022; Völcker, 2020). By extension, these terms, hashtags, emojis, and mentions suggest a bigger picture, like puzzle pieces, yet they do not form a coherent image.\n\nNext, our attention turned to analyzing clusters of words used together, focusing on the 200 most frequently used words derived using the Leiden community detection algorithm. Terms of 13 groups form clusters or communities of dignity-based topics in our dataset. Our analysis of these results, combined with iterative readings of the verbatim tweets, helped derive the framework illustrated in Figure E, organizing the thirteen clusters into five broad themes. These words and ideas are typical of the cluster, yet variation exists within clusters, and messages can blend ideas from one or more clusters.\n\nRecognize dignity as a fundamental right\n\nThe first theme affirms dignity as a fundamental human right (see Figure E for keywords). Users consider dignity a necessary condition for the enjoyment of human rights, as, without dignity, individuals cannot fully exercise or realize their rights. The global pandemic created crisis conditions that eroded a basic sense of humanity and justice as inequalities in wealth, power, race, and housing conditions threatened individuals' inherent worth and dignity. These inequalities came to the fore with discussions of unequal access to vaccines. For example, one tweet affirmed universal access to vaccines among citizens of South Africa, including migrants.\n\nUphold the dignity of essential workers\n\nThe second theme highlights the importance of upholding the dignity of frontline and healthcare workers. The COVID-19 pandemic threatened the dignity of these essential workers in several ways. One such threat is the inadequate provision of personal protective equipment, which put their health and safety at risk. To address this issue, employers must provide workers with the necessary resources and support to carry out their work safely and effectively, including proper equipment, suitable testing measures, adequate training, and sufficient staffing (The Lancet, 2020; WHO, 2020).\n\nAnother way to uphold the dignity of frontline and healthcare workers is by recognizing and valuing their work and contributions. For example, a tweet shared by the Teamsters labour union thanked a member of Local 150 for performing essential work for the Sierra Nevada Memorial Hospital during the pandemic. Employers —and society as a whole—can express gratitude and appreciation for the efforts of these workers by providing them with fair compensation and benefits and offering support and assistance for their physical and mental well-being.\n\nThe high stress and isolation many essential workers experienced during the pandemic can lead to burnout, so it is crucial to provide them with the support they need. For example, in a Tweet, the UK-based Living Wage Foundation thanked workers, including school caterers, care workers, cleaners, and delivery drivers, for keeping society going during the pandemic, asserting that they merited a living wage and sufficient work hours to maintain health and live with dignity.\n\nFinally, other messages championed the dignity of frontline and healthcare workers by calling for respect for their rights and dignity as individuals—and avoiding discrimination or harassment. By taking these steps, the public can support and empower essential workers and help them continue providing vital services to society.\n\nPreserve the dignity of at-risk populations\n\nThe third theme focuses on the close connection between dignity and populations considered vulnerable. For example, India-based WHO South-East Asia issued a tweet that called attention to the need to protect diverse groups in response to the pandemic, including older people, persons with disabilities, people who are homeless, refugees and migrants, people without access to sanitation, and people living in crowded places. Such individuals are more likely to experience disadvantage, discrimination, or exclusion. In addition, due to their limited access to resources, they may struggle with inadequate access to healthcare, education, or social support, which can negatively impact physical and mental health—and undermine dignity and self-worth (Armitage and Nellums, 2020; Croft and Fraser, 2022). Therefore, it is essential to provide at-risk populations with the necessary support to live decent and fulfilling lives.\n\nOne factor that can threaten the dignity of seniors, older adults, and elders is ageism. This discrimination and prejudice can manifest as negative stereotypes, lack of recognition and respect, or unequal access to opportunities and services. As a result, older adults may feel marginalized and devalued, undermining their dignity and self-worth. Another factor that can threaten the dignity of older adults is the inadequate provision of healthcare and social support services–especially during the pandemic. For instance, an individual's sarcastic tweet called out the Liberal–National Coalition (LNP) in Australia for failing to protect the elderly supporters who voted them into power. Other messages submitted that when such services are lacking, older adults may face isolation, loneliness, and loss of independence, all of which can harm their dignity. Additionally, lack of access to adequate housing and financial security can threaten older adults' dignity by limiting their ability to meet their basic needs and participate in society.\n\nSimilar to older adults, women may also face unique challenges that can threaten their dignity. These challenges may include pregnancy, parenting responsibilities, domestic abuse, or homelessness, and addressing these issues may require special attention and support. For example, the United Nations Population Fund (UNFPA) Kenya announced a collaboration to provide “dignity kits” to girls living in informal settlements.\n\nPrevent cascading disasters that exacerbate dignity's decline\n\nThe fourth theme specifies factors that pose a particular challenge to upholding dignity during the pandemic. Critics describe how well-intentioned COVID mitigation strategies suffer from blindspots of missing information about unresolved social problems, neglecting risks, compounding the harm, and leaving dignity in peril.\n\nFirst, dysfunctional politics can threaten dignity by promoting discriminatory policies and using propaganda and misinformation. Twitter banter between American political parties epitomizes this theme, as a barbed attack between political rivals generated a blunt counter-response with huge engagement. Dignity was a prominent theme in political discourse in the months leading up to the November presidential election. Political strife can lead to political instability and violence (as demonstrated by the Capitol attack on January 6, 2021), so it is crucial to promote functional and inclusive politics that respect the rights and dignity of all individuals.\n\nSecond, food insecurity can undermine individuals' dignity by forcing them to make difficult choices and limiting their access to nutritious and healthy food. The COVID-19 pandemic worsened this problem, leading Nourish Scotland to campaign for communities to respond to food insecurity by providing people with access to healthy food options and ensuring social connections.\n\nThird, poverty can undermine individuals' dignity by limiting their access to basic needs and opportunities. Conversely, addressing poverty can help to restore and uphold individuals' dignity by providing them with the means to meet their basic needs and participate in society. Finally, chronic deficits in many domains require a coordinated solution, as the New York-based advocacy group Women Deliver emphasized, calling for world leaders to embrace international collaboration in recovery plans.\n\nFourth, healthcare and insurance can support and protect an individual's dignity by providing access to necessary medical care and services and protecting them from financial hardship. For example, an individual from Texas tweeted a reminder that coronavirus does not discriminate and that every single person in the country should be able to see a doctor, irrespective of employment or insurance status.\n\nAttend to death, dignity, and the sanctity of life\n\nThe fifth theme considers the consequences of failing to uphold dignity in the face of death and illness. As the COVID-19 pandemic swept across the world, nations faced a tragedy of unprecedented proportions. Each day, the death toll rose as more people fell victim to the deadly virus. Unfortunately, political tensions—particularly in the United States—plus a lack of a cohesive and effective national response exacerbated the situation. As the tragedy unfolded, it became clear that the country failed to adequately protect its people's dignity. An English political pundit posted an updated death toll, ascribed blame, and urged people to vote.\n\nRespecting an individual's dignity during their end-of-life can be crucial to providing decent end-of-life care. Dignity is a quality or state of being worthy of respect and honour, and as individuals approach the end of life, they often wish for a “good death” that is peaceful, pain-free, and by the individual's wishes and values. Additionally, a Kenyan argued that providing protective suits might allow people to give their final respects to their loved ones with dignity. Finally, some followers of an Indian spiritual leader tweeted claims that they had received a divine cure for the deadly virus, telling of healing without medicine.\n\nGiven our interest in examining older adults' challenges in the pandemic, we next examined the subset of tweets that mention older adults. We flagged messages in the sample that included at least one of the following keyword variants: elder* (n=29), senior* (n=28), older (n=24), pension* (n=11), retire* (n=6), or aged (n=1). This procedure netted 86 unique tweets, equal to 4.4% of the sample.\n\nMost messages echoed the dignity themes discussed previously. Foremost, dignity is a human right protected by the United Nations and World Health Organization, a message affirmed by the UN Secretary-General, who reminded world leaders to respect the rights and dignity of older people. Older persons were disproportionately harmed by COVID-19, as reflected in death tolls and social isolation (Fraser et al., 2020; Sharma, 2021; Wu, 2020). Furthermore, older age intersects with other risk factors, including poverty, disabilities, race, homelessness, immigrants, non-native speakers, and incarcerated persons.\n\nProblems with long-term care homes drew considerable attention as facilities struggled to protect residents from the virus, maintain adequate staffing and care levels, balance social distancing measures, and reduce isolation from family members. The tragic loss of lives sparked calls to hold government officials accountable and to reform the system.\n\nFinally, some seniors' meal delivery services were suspended during the pandemic, placing recipients at risk of not having enough groceries, risking hunger, malnutrition, or even starvation. Poverty increases these risks, as some posts drew attention to low incomes and inadequate pensions or social security benefits.\n\n\nDiscussion\n\nThis study examined social media discourse on dignity posted during the first year of the global COVID-19 pandemic. A secondary goal was to explore the use of dignity in conjunction with ageing. Our cross-national sample, Leiden community detection algorithm and thematic analysis of identified clusters revealed five major themes: (a) recognize dignity as a fundamental right, (b) uphold the dignity of essential workers, (c) preserve the dignity of at-risk populations, (d) prevent cascading disasters that exacerbate dignity's decline, and (e) attend to death, dignity, and the sanctity of life. While each of these themes has distinct contributions, the data reveals that essential workers and at-risk populations were identified as groups whose dignity was heavily impacted during the first year of the pandemic. The implications of the five themes are discussed below.\n\nTweets in the first three themes reinforced the Universal Declaration of Human Rights to promote dignity as a fundamental human right. In line with Mégret and Hoffman's (2009) seminal work, findings demonstrate the importance of contextual factors and intersection with other human rights. For example, women are more likely than men to be frontline workers in the context of the COVID-19 pandemic (Utzet et al., 2022) and, as such, face a greater risk of loss of dignity in the context of healthcare provision during a pandemic. The impact on the dignity and health of essential workers has been raised as a target for policy action (Lancet, 2021). Similarly, the findings of this study align with calls to prioritize the older population, their needs and dignity in times of crisis. Minimizing ageist discourse and maximizing access to healthcare and other services to maintain the health and dignity of this population (Fraser et al., 2020; Lagacé et al., 2020).\n\nFormal and informal communication—via news outlets and social media—have been essential throughout the pandemic for exchanging information and enhancing awareness of risk and public health recommendations (Généreux et al., 2020). However, while modern technologies have immeasurable benefits for providing accessible communication, it is also prudent to consider how media discourse can frame issues and shape beliefs during disasters (Choudhury & Haque, 2018; Wang et al., 2019), including ageist and ableist attitudes (Barth et al., 2021; O'Sullivan and Phillips 2019). Indeed, Fraser et al. (2020) warned of the potential negative impacts of ageist hashtags circulating on social media during the early weeks of the COVID-19 pandemic. Lagacé et al. (2021) report similar concerns about the COVID-19 media discourse labelling all older adults as vulnerable people whom we must “fight for” and not “fight along with.” Similarly, Stollznow (2020) describes how prejudice in ageist language perpetuates conflict between boomer and millennial generations, and in the end, impacts everyone.\n\nOur research findings illustrate how social media discourse can be valuable for identifying key issues requiring policy reform. First, the method can amplify the voices of marginalized or underrepresented groups, bringing attention to issues not widely recognized or addressed before. Second, though our analysis was retrospective, the data lends itself to real-time monitoring and rapid response to urgent or pressing issues on social media. Third, it is possible to identify trends and patterns in discussions and debates on social media platforms. Finally, by analyzing the sentiment and engagement with different topics, policymakers can understand what issues are most important to the general public and which issues may create conflict or reduce trust in government or decision-making authorities.\n\nThe rise of social media has opened new avenues to explore public opinion, yet it is not without limitations. First, social media users do not represent the general population's views due to self-selection and non-random participation. Therefore, combining this method with other research methods may be necessary to understand public views on human dignity fully. Second, while individuals and organizations have control over the messages they create, the algorithms used by platforms significantly influence the extent to which those messages are seen and received by their intended audience (Kozinets and Gretzel, 2021). Third, information shared on social media can be contaminated with misinformation and tainted by stereotypes, so it is essential to be vigilant in promoting accurate and credible information and be aware of potential biases.\n\nThis study is part of a larger research program focused on how pandemic experiences and exposure to COVID-19 media discourse influenced older adults' perceptions of resilience and vulnerability. It is the first step in a series of research activities where we are exploring how older age and ability are framed in pandemic media discourse. Given the emphasis on dignity in social media, this study provides a glimpse into how the term is generally used in the public sphere and, more specifically, in the context of aging.\n\nAs a broader impact, the findings have the potential to inform decision-makers about how the public views dignity and aging. Listening provides feedback on public trust, confidence and priorities and can assist decision-makers by understanding the discourse circulating in social media. Given the conceptualization of dignity as a human right and its relevance for combatting ageism, these findings can help promote healthy ageing throughout pandemic recovery and adaptation to a 'new normal'.\n\nThe underlying data to this research cannot be shared due to the ethical and copyright restrictions surrounding social media data. The Methods section contains detailed information to allow replication of the study which used the Type, Date, Location, Engagement, and Message fields of archived Twitter data records. Any queries about the methodology should be directed to the corresponding author.\n\nThe Social Astronomy app used to analyze clusters is proprietary; contact Belanger Analytics Inc. 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PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "208735", "date": "13 Oct 2023", "name": "Meiko Makita", "expertise": [ "Reviewer Expertise Meiko Makita: social gerontology", "feminist methodology", "social media analysis", "discourse analysis. Hannah Loret: health", "gender", "qualitative research", "interdisciplinary practice" ], "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\nComments to the authors by reviewer 1 (Meiko Makita)\nThe paper reads fairly well and offers insights into how dignity was perceived and discussed during the early days of the pandemic, particularly in relation to older adults and various societal concerns. I believe the manuscript can be further improved. Here are some revisions I suggest for strengthening the overall quality of the manuscript.\n\n1. Form and style.\nPlease be consistent with either British or American language. Throughout the manuscript you use both (e.g., ageing, aging).\n\nIn-text citations are presented in at least two different formats.\n\nPlease review the references list.\n\nOn page 3, par 5: (i.e., older adults, 2SLGBTQI+, Indigenous people, immigrants, etc.) it should be ‘e.g.’ without ‘etc.’. t\n\nThe acronym LGBTQI+ is well-known, however I do not know what 2S stand for, perhaps it would be better to spelled it out.\n\n2. Section Theoretical Background/Social Media and Marketplace sentiment\nI was surprised the authors only covered very broad social media studies and did not include any study on older adults/ageism and social media, such as the two I shared below,\nReuben Ng, Nicole Indran, Hostility Toward Baby Boomers on TikTok, The Gerontologist, Volume 62, Issue 8, October 2022, Pages 1196–1206, https://doi.org/10.1093/geront/gnac020\nElliott, R. (2022). The ‘Boomer remover’: Intergenerational discounting, the coronavirus and climate change. The Sociological Review, 70(1), 74-91. https://doi.org/10.1177/00380261211049023\n3. Methodology\nThe method appears to have followed a systematic approach and the authors state that their study is reproducible, which is a positive aspect in terms of rigour. In the Discussion section the authors mention: “Finally, by analyzing the sentiment and engagement with different topics, policymakers can understand what issues are most important to the general public and which issues may create conflict or reduce trust in government or decision-making authorities.”\nHowever, no sentiment analysis of the tweets data set was conducted. Could you please either conduct a proper sentiment analysis or explain what is that you mean by ‘analysing the sentiment’?\nI suggest moving the text that is currently in the first paragraph of Results (below) to the Methods section, when ethics is introduced:\n\nSocial media studies often include quotes to give participants a voice, illustrate ideas to build trust, foster replicability, or lend evidence for claims. However, evolving ethical standards attend to the unintended consequences of reporting individuals’ tweets and express concern about the discoverability of messages posted voluntarily in public forums (Mason and Singh, 2022). Accordingly, we honour individuals’ privacy and do not reproduce user IDs or tweets. In contrast, we refer to companies, organizations, and institutions that use Twitter for public relations to share views with stakeholders.\nI’d also further elaborate on the ethical implications of using publicly available online data. There is a lot of debate on this issue and the authors should engage with this literature and further discuss their stance on using publicly available online data. Please see for instance: Stommel W, Rijk L de. Ethical approval: none sought. How discourse analysts report ethical issues around publicly available online data. Research Ethics. 2021;17(3):275-297. doi: 10.1177/1747016120988767\nThe method employed is not discourse analysis but thematic analysis. This is also supported by the name of the subsection ‘thematic analysis and word clusters’. Perhaps remove from the keywords ‘discourse analysis’ and only retain ‘discourse’.\n4. Results\nTweets with most engagement – is this about number of likes and retweets? Could we have more data on this for each of the top-10 accounts? Also, since the audience would be not only from North America, probably best to specify this is USA’s Democratic Party.\nIn comparison, with the other four themes, ‘recognise dignity as a fundamental right’ is not as developed. Or is the first theme the overarching theme and the other four subthemes?\nConsider changing the themes to an ‘ing’ verb form, example: Preserving the dignity of at-risk populations.\nThe section ‘Tweets focused on older adults’ seems to be a little underdeveloped, especially since is a focus of the study. In here, the authors could perhaps introduce examples of what was said in tweets, as they did with the other themes.\n5. Discussion\nOnce the authors expand on their theoretical background section, they would be able to also strengthen their discussion.\nThe concept of ‘healthy ageing’ is only introduced at the end of the manuscript – if this is important try to bring it up much earlier on (intro, background).\nConsider adding a ‘conclusion’ section.\n\nComments to the authors by reviewer 2 (Hannah Loret)\nTitle and Abstract:\nThe summary of the background and purpose of the research should better articulate the research question and the need for this study to have been undertaken.\n\nSample size is clear, but how the data was identified and sourced could be covered here.\n\nThe conclusion is very descriptive. What is the main finding (it is clear however why we should lament the unacceptable situation for older adults)? This could show a stronger critical engagement with the results of the study.\n\nIntroduction:\nReferring to structurally underserved ‘vulnerable’ people as ‘creatures’ (p. 3 para 5) could be better expressed here, by choosing a different quotation to make the point. As it stands, the way it is currently expressed risks repeating the terminology used to marginalise the groups mentioned.\n\nLater in this same paragraph, there is the claim that to consider older people as all vulnerable is ageist, but earlier in the paragraph the authors point to older people as a vulnerable group. Perhaps this could be mitigated by suggesting older people, ‘immigrants etc.’ are potentially vulnerable, allowing for slightly more nuanced use of the term?\n\nP. 3 paragraph 6 I am unsure why Latour is being brought in. Are his methods being used? This reference to his work or approach doesn’t seem to be upheld in this section or signposting to the methodology, beyond saying that research into media discourse and public opinion (which are two different things – perhaps a distinction could be made?) has a similar investment in making meaning.\n\n(P.4) I would like to see more references and conclusions drawn in this field which justify the need for the study from sources which aren’t written by this article’s authors.\n\nI understand on a basic level why this approach has been chosen, but I understand less what the current literature in this specific field looks like and what contribution to the field this article makes. It could be much more explicit in the theoretical background exactly why this study was needed from both a theoretical and empirical perspective.\n\nI’m unclear from the introduction what the exact aims or hypotheses of this study/article are. Can these be stated explicitly?\n\nWhy were older adults chosen over any other demographic group for the focus here? Please expose the gap in the literature and the rationale for this choice.\n\nMethods:\n(Para 2, p. 4) I disagree that Twitter is a highly inclusive platform. What about people who have literacy issues or with a lack of digital skills, internet access, or other unmet access needs which prevent them engaging with it?\n\n(Para 3, p. 4) why were English-language tweets chosen over another language? The rationale should be clearer for this.\n\n(P. 4) can you speak a little more about how this data was analysed, to signpost to the results section? The methods used are clear and seem appropriate, but I’m struggling to see how they fit within the overall aims of the project.\n\n(p. 4) could a rationale be given for why this method was chosen over another, and how any bias may function within it?\n\nI find the design coherent, but the research questions need to be more clearly stated earlier in the article so that perhaps the methodology section conveys how these will be answered more clearly.\n\nSome of the information in the ‘results’ section would be better placed in the ‘methodology section’, as it is describing the methodology rather than the results. I’m particularly thinking of the last section of the final paragraph on page 5 where this is a description of methodology rather than results. Methodological explanations at the beginning of the results section could be incorporated into the methodology, allowing more room for the thematic structuring of the results sections.\n\nResults:\nA short summary of the main results which relates to the research questions (as well as methodological focus) would be helpful.\n\nI particularly like the final paragraph of page 5. The tone is pleasantly authoritative.\n\n(P.8, final paragraph) I’m unsure why migrants in South Africa have been chosen to demonstrate this point. Could this point be made clearer with reference to the points in the preceding sentence, for example by explicitly explaining how this relates to questions of race/wealth/housing conditions?\n\n(p.9, para 4) I’m unclear how the study data demonstrates that by ‘taking these steps, the public can support and empower essential workers and help them continue providing vital services to society’. My understanding is that the data demonstrates a public acknowledgement and will to improve the working conditions of essential workers but the steps to this are unclear, and it is unclear how a Twitter discourse actively empowers workers or helps them to deliver services. Perhaps a closer reading of the data gathered could make this clearer, in terms of how the public demonstrated support for the dignity of essential workers through their tweets, then linking this to any related targeted activism which followed for those particular groups of working people?\n\n(p. 9, para 5) This section could also benefit from some closer reference to the empirical data from the study to strengthen its statements and enrich the points made. I also questioned the use of the term ‘at risk’ when the term vulnerable had been used before. Perhaps it would be good to stick to one of these terms or to give a reason for the change in terminology?\n\n(P. 9, para 6) Is there a reason that this section chooses to focus more closely on older adults as a structurally underserved population? This needs to be more explicit.\n\n(P.9, para 7) The first sentence here makes it sound like older adults and women are two completely distinct groups, and that older women are not included in older adults. Please reword to clarify that older adults includes women, though (younger?) women experiencing domestic abuse, precarious housing, pregnancy-related issues etc. may be affected by unique gender-related challenges.\n\n(P.9, para 8) The paragraphs here are very short and therefore lose a sense of coherence. They could be joined without losing the sense of the argument, and this would also help to strengthen the use of empirical data to evidence the points being made (I would again like to see more information from the empirical data to back up the statements being made here).\n\n(P.9, para 8) Needs a reference or several references for ‘critics describe…’\n\n(p. 10, para 1) Readers who are not in the USA may not be familiar with the political parties or political structure there, or the exact details of the Capitol attack in January 2021. If this is to be included, it will need to be qualified by some more information about the USA political system which links to the point being made about propaganda/misinformation and discriminatory policies.\n\nOn this point, I think discriminatory policies and propaganda/misinformation are two separate points. Perhaps these could be teased out briefly with reference to empirical data to justify these statements.\n\n(P. 10, para 4) It is important to acknowledge that although many healthcare systems operate both publicly and privately, and have social security, healthcare system design varies and the example given from Texas will be unique to the case in the USA which should be clearer – for example in Scotland, ‘insurance status’ would mean something very different in terms of healthcare, and most of the population is able to access primary and secondary care free of charge.\n\n(p. 10, para 6) it is unclear why we are suddenly speaking about end-of-life care. This could be better linked to the empirical data, and could be linked to the above paragraph with a signposting sentence to improve the flow of the article.\n\n(p. 10, para 6) It is unclear why we are looking at Kenya and India for these mentions, the connection could be more explicit and better linked back to the concerns of the overall analysis (if the article wants to concentrate on meaning, there will need to be more critical appraisal or expansion on possible meanings here as elsewhere).\n\nIt is not until p. 10 we are told why older adults are a particular focus. This rationale is strong and needs to figure much earlier in the article.\n\nFigures and Tables:\nI find that figures and tables are clear, easily interpreted, and well-formatted.\n\nDiscussion:\nThe start to this section is strong, and the tone is appropriate. The summary of the main themes is particularly helpful.\n\nThere needs to be some discussion in this section of existence of opposing views to the ones presented. The wider context around the findings and any other research which may contradict these findings need to be explored, and linked back to the research gap that this study was filling.\n\nThe limitations are clear, perhaps the strengths need to be better situated within the available literature and existing knowledge on this topic.\n\nFuture directions for the research and practice are clear and meaningful.\n\nReferences:\nSome inconsistencies for referencing (particularly of The Lancet - Lancet, T. as author of one editorial, please correct this in line with style guidelines). Please ensure consistency and revise these.\n\nRecommendations:\nThis article has an appealing focus on the articulation of dignity through Twitter during the start of the COVID-19 pandemic. It touches on key concepts around dignity and multiple disadvantage which were exacerbated during the pandemic. It would benefit from some restructuring (particularly in the results section, see comments) and a deeper critical engagement with concepts of dignity, gender, age, and the wider political and cross-national social contexts within which the texts studied were created. It would also benefit from a clearer explanation of the necessity of this study, and of the trends and oppositions in existing theoretical and empirical literature in this research area.\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? 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? Partly", "responses": [ { "c_id": "12733", "date": "10 Dec 2024", "name": "Michael Mulvey", "role": "Author Response", "response": "Comments to the authors by reviewer 1 (Meiko Makita) The paper reads fairly well and offers insights into how dignity was perceived and discussed during the early days of the pandemic, particularly in relation to older adults and various societal concerns. I believe the manuscript can be further improved. Here are some revisions I suggest for strengthening the overall quality of the manuscript. 1. Form and style. Please be consistent with either British or American language. Throughout the manuscript you use both (e.g., ageing, aging). In-text citations are presented in at least two different formats. Please review the references list. On page 3, par 5: (i.e., older adults, 2SLGBTQI+, Indigenous people, immigrants, etc.) it should be ‘e.g.’ without ‘etc.’. t The acronym LGBTQI+ is well-known, however I do not know what 2S stand for, perhaps it would be better to spelled it out. Response: Good points. We brought consistency to “ageing,” used Canadian English throughout, and reviewed the format of in-text citations. Instead of using evolving acronyms that vary across geographies, we refer to “sexually and gender-diverse communities.” 2. Section Theoretical Background/Social Media and Marketplace sentiment I was surprised the authors only covered very broad social media studies and did not include any study on older adults/ageism and social media, such as the two I shared below, Reuben Ng, Nicole Indran, Hostility Toward Baby Boomers on TikTok, The Gerontologist, Volume 62, Issue 8, October 2022, Pages 1196–1206, https://doi.org/10.1093/geront/gnac020 Elliott, R. (2022). The ‘Boomer remover’: Intergenerational discounting, the coronavirus and climate change. The Sociological Review, 70(1), 74-91. https://doi.org/10.1177/00380261211049023 Response: Thank you for the helpful suggestion. We have reviewed the studies you shared and agree they offer valuable insights into older adults and ageism in social media. We have incorporated relevant findings (Archambault et al., 2022; Bascu et al., 2022; Clancy et al., 2021; Elliott, 2022; Lothian & Philp, 2001; Ng & Indran, 2022) into the manuscript, particularly in the literature review and discussion sections. 3. Methodology The method appears to have followed a systematic approach and the authors state that their study is reproducible, which is a positive aspect in terms of rigour. In the Discussion section the authors mention: “Finally, by analyzing the sentiment and engagement with different topics, policymakers can understand what issues are most important to the general public and which issues may create conflict or reduce trust in government or decision-making authorities.” However, no sentiment analysis of the tweets data set was conducted. Could you please either conduct a proper sentiment analysis or explain what is that you mean by ‘analysing the sentiment’? Response: We removed this from the manuscript. I suggest moving the text that is currently in the first paragraph of Results (below) to the Methods section, when ethics is introduced: Social media studies often include quotes to give participants a voice, illustrate ideas to build trust, foster replicability, or lend evidence for claims. However, evolving ethical standards attend to the unintended consequences of reporting individuals’ tweets and express concern about the discoverability of messages posted voluntarily in public forums (Mason and Singh, 2022). Accordingly, we honour individuals’ privacy and do not reproduce user IDs or tweets. In contrast, we refer to companies, organizations, and institutions that use Twitter for public relations to share views with stakeholders. Response: Thank you for this suggestion. We moved this portion of the text to the Methods section. I’d also further elaborate on the ethical implications of using publicly available online data. There is a lot of debate on this issue and the authors should engage with this literature and further discuss their stance on using publicly available online data. Please see for instance: Stommel W, Rijk L de. Ethical approval: none sought. How discourse analysts report ethical issues around publicly available online data. Research Ethics. 2021;17(3):275-297. doi: 10.1177/1747016120988767 Response: We included the suggested Stommel reference in this section and elaborated on the ethical implications of using publicly available online data. The method employed is not discourse analysis but thematic analysis. This is also supported by the name of the subsection ‘thematic analysis and word clusters’. Perhaps remove from the keywords ‘discourse analysis’ and only retain ‘discourse’. Response: We corrected this in the text by removing discourse analysis. 4. Results Tweets with most engagement – is this about number of likes and retweets? Could we have more data on this for each of the top-10 accounts? Also, since the audience would be not only from North America, probably best to specify this is USA’s Democratic Party. Response: We have included engagement details and made our descriptions more precise. In comparison, with the other four themes, ‘recognise dignity as a fundamental right’ is not as developed. Or is the first theme the overarching theme and the other four subthemes? Response: The first theme is general, often providing context to introduce more specific themes. We added a transition sentence to improve clarity. Consider changing the themes to an ‘ing’ verb form, example: Preserving the dignity of at-risk populations. Response: Good idea – thanks! The section ‘Tweets focused on older adults’ seems to be a little underdeveloped, especially since is a focus of the study. In here, the authors could perhaps introduce examples of what was said in tweets, as they did with the other themes. Response: We thank the reviewer for this comment and wish to clarify that the study focused on dignity. A secondary aim of the study was to examine a subset of posts that focused on older adults and dignity. We added text and research questions to the introduction and abstract to clarify this. 5. Discussion Once the authors expand on their theoretical background section, they would be able to also strengthen their discussion. The concept of ‘healthy ageing’ is only introduced at the end of the manuscript – if this is important try to bring it up much earlier on (intro, background). Response: In the Introduction, we discussed how older adults were discussed and how this may infringe on dignity: “Indeed, in the context of COVID-19, initially, the discourse surrounding older adults living in long-term care homes focused heavily on the loss of dignity in healthcare and in dying (Carrieri et al., 2020; Colombo, 2021; Kowsalya & Sundara, 2021, Vellani et al., 2021). In general, much of the discourse was ageist, promoting notions that all older adults need protection, are vulnerable and should be socially isolated (Fraser et al., 2020, Lagacé et al., 2020) –all factors which infringe on human rights and dignity.” This is important at the main focus of the paper is DIGNITY and secondary focus dignity and aging. We mention healthy aging in the last paragraph of the discussion to emphasize how changing the discourse around older adults to a more positive orientation while diminishing or removing ageist perspectives will support healthy aging in this population. Consider adding a ‘conclusion’ section. Response: Thank you for this suggestion, we added a conclusion section: “This study analyzed Twitter posts about dignity during the first year of the COVID-19 pandemic, with a focus on the intersection of dignity and older adults. Using text analytics and interpretive methods on a cross-national sample, we identified five major themes: recognizing dignity as a fundamental right, upholding the dignity of essential workers, preserving the dignity of at-risk populations, preventing cascading disasters, and attending to death dignity. Our findings emphasize the widespread discussion of dignity-related challenges faced by essential workers and populations experiencing vulnerability, particularly older adults. This underscores the urgent need for policies that address these challenges to promote human dignity. Social media has captured real-time public sentiment and provided insights that inform policy and practice. In summary, this research improves our understanding of public perceptions of dignity during a global crisis, emphasizing the importance of policies that uphold dignity, especially for those most affected by the pandemic.” Comments to the authors by reviewer 2 (Hannah Loret) Title and Abstract: The summary of the background and purpose of the research should better articulate the research question and the need for this study to have been undertaken. Sample size is clear, but how the data was identified and sourced could be covered here. Response: Thank you for this suggestion. We edited the background and purpose statement to clarify the need for the study and the questions we set out to answer. The conclusion is very descriptive. What is the main finding (it is clear however why we should lament the unacceptable situation for older adults)? This could show a stronger critical engagement with the results of the study. Response: To refine the conclusion in response to this comment and that of the other reviewer, our conclusion now reads: “Conclusions: Within the discourse, dignity emerged as a right and a policy target (improving or maximizing dignity in various groups). Further, the posts focusing on older adults and dignity demonstrated that the impacts of COVID-19 disproportionately impacted older adults, particularly those with greater care needs and lower socioeconomic status.” Introduction: Referring to structurally underserved ‘vulnerable’ people as ‘creatures’ (p. 3 para 5) could be better expressed here, by choosing a different quotation to make the point. As it stands, the way it is currently expressed risks repeating the terminology used to marginalise the groups mentioned. Response: Thank you for this comment. We rephrased the sentence: “In extreme cases, dignity violations may undermine personhood, as individuals risk being treated as “not human or less than human–as a thing or instrument or subhuman creature” ( Kateb, 2014).” Later in this same paragraph, there is the claim that to consider older people as all vulnerable is ageist, but earlier in the paragraph the authors point to older people as a vulnerable group. Perhaps this could be mitigated by suggesting older people, ‘immigrants etc.’ are potentially vulnerable, allowing for slightly more nuanced use of the term? Response: Thank you for noticing the discrepancy. We rephrased accordingly: “For these reasons, marginalized individuals or groups may be at greater risk of experiencing vulnerability ( i.e., older adults, children, immigrants, Indigenous people, low-income individuals, and sexually and gender-diverse communities) and are at a greater risk of losing their dignity” P. 3 paragraph 6 I am unsure why Latour is being brought in. Are his methods being used? This reference to his work or approach doesn’t seem to be upheld in this section or signposting to the methodology, beyond saying that research into media discourse and public opinion (which are two different things – perhaps a distinction could be made?) has a similar investment in making meaning. Response: We rephrased this section to clarify why Latour is used and to distinguish between media discourse and public opinion: “Sociologist Bruno Latour (2007) explores how language and representation shape social interactions and worldviews. His perspective is relevant to our study of social media discourse on dignity during the COVID-19 pandemic, as we analyzed how users interact through tweets, using symbols and narratives to construct and express their views on dignity. Similarly, researchers who examine media discourse and public opinion also focus on the symbols and reasoning devices used to present and discuss issues, such as metaphors, exemplars, catchphrases, depictions, and visual images, as well as the roots, consequences, and appeals to principle that suggest how to think about an issue ( Gamson & Modigliani, 1989).” (P.4) I would like to see more references and conclusions drawn in this field which justify the need for the study from sources which aren’t written by this article’s authors. Response: Several references have been added to the introduction. I understand on a basic level why this approach has been chosen, but I understand less what the current literature in this specific field looks like and what contribution to the field this article makes. It could be much more explicit in the theoretical background exactly why this study was needed from both a theoretical and empirical perspective. Response: We added details and references to the introduction, just before text which clarifies our research questions: “Scholars argue that dignity is a fundamental aspect of human rights, crucial for realizing other rights. Theoretical work emphasizes the intersectionality of dignity, highlighting how it is affected by various factors such as age, gender, socioeconomic status, and cultural background. Marginalized groups, such as older adults, are especially at risk of having their dignity violated ( Archambault et al., 2022; Bascu et al., 2022; Clancy et al., 2021; Elliott, 2022; Lothian & Philp, 2001; Ng & Indran, 2022).” I’m unclear from the introduction what the exact aims or hypotheses of this study/article are. Can these be stated explicitly? Response: We added our research questions in the Introduction: “Therefore, the following research questions guided our investigation: Research Question 1: What issues of human rights and ethical considerations are highlighted in high-engagement tweets on dignity? Research Question 2: What themes emerge from dignity-related tweets that reflect broader societal concerns about health, social justice, and public policy during the pandemic? Research Question 3: How do dignity-related tweets address the challenges older adults faced during the COVID-19 pandemic?” Why were older adults chosen over any other demographic group for the focus here? Please expose the gap in the literature and the rationale for this choice. Response: In response to comments from both reviewers, we added details on our rationale for focusing on older adults in the introduction: “The COVID-19 pandemic brought increased attention to the concept of healthy ageing, which involves maintenance of the ability to function well as people age, while preserving dignity. Ensuring older adults can live with dignity and respect is crucial to promoting their overall health and well-being (WHO, 2015; Beard et al., 2016; Sixsmith & Sixsmith, 2008). The WHO's \"Decade of Healthy Ageing\" initiative (2021-2030) emphasizes the need for comprehensive care and long-term support systems to improve the lives of older adults during and after the pandemic (WHO, 2020). Additionally, a life-course model for healthier aging, proposed by Jowell, Carstensen, & Barry (2020) in The Lancet, highlights lessons learned during the COVID-19 pandemic and emphasizes the importance of supportive policies to support the health and dignity of older adults. The main goal of this study was to examine how the concept of dignity was discussed on Twitter (X) during the COVID-19 pandemic. Secondly, given that older adults are at a greater risk of having their dignity violated (Clancy et al., 2021; Lothian & Philp, 2001), this study targets the intersection of dignity and older adults in Twitter posts.” Methods: (Para 2, p. 4) I disagree that Twitter is a highly inclusive platform. What about people who have literacy issues or lack digital skills, internet access, or other unmet access needs that prevent them from engaging with it? Response: We agree with the reviewer and rephrased this paragraph to state the benefits and pitfalls of using the Twitter (X) platform: “Our study examined original messages from Twitter users, a one-to-many platform where people share information in a relatively synchronous or condensed time frame ( Lamberton & Humphreys, 2022). Twitter data allows researchers to gain insight into public opinion, including its volume, sentiment, and expressions. Despite Twitter's efforts to be inclusive, it is not accessible to everyone due to literacy issues, digital skill gaps, and limited internet access. Although the platform has a diverse user base, its primary users generally fit the WEIRD criteria (Western, Educated, Industrialized, Rich, and Democratic) ( Septiandri, Constantinides, & Quercia, 2024).” (Para 3, p. 4) why were English-language tweets chosen over another language? The rationale should be clearer for this. Response: To clarify why English-language tweets were chosen, we added, “Our cross-national sample of English tweets was curated with a focus on researcher expertise, bolstering the accuracy of our interpretations. However, we recognize limitations in extending these findings to other languages.” (P. 4) can you speak a little more about how this data was analysed, to signpost to the results section? The methods used are clear and seem appropriate, but I’m struggling to see how they fit within the overall aims of the project. Response: The data were analyzed thematically to show how comments on dignity and older age circulate on public platforms and reflect broader societal issues related to the pandemic, dignity and age. Our multi-method approach blended text analysis algorithms and human interpretation – leveraging the strengths of the research team. The methods section was revised to eliminate jargon and improve clarity. (p. 4) could a rationale be given for why this method was chosen over another, and how any bias may function within it? Response: In the discussion, we addressed this comment in our limitations paragraph: “The rise of social media has opened new avenues to explore public opinion, yet it has limitations. First, social media users do not represent the general population's views due to self-selection and non-random participation. Therefore, combining this method with other research methods may be necessary to fully understand public views on human dignity. Second, while individuals and organizations control the messages they create, the algorithms and platforms used significantly influence how their intended audience sees and receives those messages ( Kozinets & Gretzel, 2021). Third, information shared on social media can be contaminated with misinformation and tainted by stereotypes, so it is essential to be vigilant in promoting accurate and credible information and be aware of potential biases.” I find the design coherent, but the research questions need to be more clearly stated earlier in the article so that perhaps the methodology section conveys more clearly how these will be answered. Response: We included research questions in the Introduction to better guide the reader. Some of the information in the ‘results’ section would be better placed in the ‘methodology section’, as it is describing the methodology rather than the results. I’m particularly thinking of the last section of the final paragraph on page 5 where this is a description of methodology rather than results. Methodological explanations at the beginning of the results section could be incorporated into the methodology, allowing more room for the thematic structuring of the results sections. Response: Thank you for this suggestion. We moved this portion of the text to the Methods section. Results: A short summary of the main results which relates to the research questions (as well as methodological focus) would be helpful. Response: We thank you for this suggestion and added the following summary at the beginning of the results section: “The findings from this study are structured around five major themes identified through an analysis of 1,946 original tweets about dignity during the COVID-19 pandemic. These themes include recognizing dignity as a fundamental right, upholding the dignity of essential workers, preserving the dignity of at-risk populations, preventing cascading disasters, and attending to death dignity. Each theme is explored in detail below, highlighting the unique challenges and discourses associated with each aspect of dignity.” I particularly like the final paragraph of page 5. The tone is pleasantly authoritative. Response: Thank you. (P.8, final paragraph) I’m unsure why migrants in South Africa have been chosen to demonstrate this point. Could this point be made clearer with reference to the points in the preceding sentence, for example by explicitly explaining how this relates to questions of race/wealth/housing conditions? Response: We agree that additional context is needed in this section. We have provided this in our revised version: “These inequalities came to the fore with discussions about unequal access to vaccines. For example, one tweet advocated for universal access to vaccines for all citizens of South Africa, including migrants. Similarly, hashtags demonstrated solidarity with the social equity priorities of groups like Black Lives Matter, Latinos, Pride, and Veterans. Next, we discuss the overarching principle of recognizing dignity as a fundamental right, echoed in other themes focusing on specific populations.” (p.9, para 4) I’m unclear how the study data demonstrates that by ‘taking these steps, the public can support and empower essential workers and help them continue providing vital services to society’. My understanding is that the data demonstrates a public acknowledgement and will to improve the working conditions of essential workers but the steps to this are unclear, and it is unclear how a Twitter discourse actively empowers workers or helps them to deliver services. Perhaps a closer reading of the data gathered could make this clearer, in terms of how the public demonstrated support for the dignity of essential workers through their tweets, then linking this to any related targeted activism which followed for those particular groups of working people? Response: We revised this section to state the results and potential impacts for healthcare workers clearly. We also added a reference to support the view that positive/supportive posts of essential workers have a positive impact on their engagement and well-being (see Shan et al., 2022): “Finally, other messages championed the dignity of frontline and healthcare workers by calling for respect for their rights and dignity as individuals—and avoiding discrimination or harassment. This type of public support and can empower essential workers and motivate them to continue providing vital services to society (Shan et al., 2022).” (p. 9, para 5) This section could also benefit from some closer reference to the empirical data from the study to strengthen its statements and enrich the points made. I also questioned the use of the term ‘at risk’ when the term vulnerable had been used before. Perhaps it would be good to stick to one of these terms or to give a reason for the change in terminology? Response: We bolstered the section with examples and revised the text to use “at risk” throughout. (P. 9, para 6) Is there a reason that this section chooses to focus more closely on older adults as a structurally underserved population? This needs to be more explicit. Response: Thank you for this comment. We revised the introduction to state upfront the importance of this specific population as underserved and the focus on the intersection of older adults and dignity during the pandemic. We’ve added references to set up our research questions and elaborate throughout the paper. (P.9, para 7) The first sentence here makes it sound like older adults and women are two completely distinct groups, and that older women are not included in older adults. Please reword to clarify that older adults includes women, though (younger?) women experiencing domestic abuse, precarious housing, pregnancy-related issues etc. may be affected by unique gender-related challenges. Response: We revised this paragraph to address this comment: “Older and younger women faced unique challenges that threatened their dignity as access to necessary healthcare services exacerbated existing issues. Many older women took on additional caregiving roles during the pandemic without adequate support, adding to the burden of unpaid domestic work and putting stress on their physical and mental well-being (Age International, 2021). Additional challenges included pregnancy, parenting responsibilities, domestic abuse, or homelessness, and addressing these issues may require special attention and support. For example, the United Nations Population Fund (UNFPA) Kenya announced a collaboration to provide “dignity kits” to girls living in informal settlements.” (P.9, para 8) The paragraphs here are very short and therefore lose a sense of coherence. They could be joined without losing the sense of the argument, and this would also help to strengthen the use of empirical data to evidence the points being made (I would again like to see more information from the empirical data to back up the statements being made here). Response: This section expands several paragraphs to provide additional context to the statements. We summarize the essence of tweets to reduce privacy and traceability concerns related to verbatim reporting. (P.9, para 8) Needs a reference or several references for ‘critics describe…’ Response: We added two references for this statement: “Critics describe how well-intentioned COVID mitigation strategies suffer from blind spots of missing information about unresolved social problems, neglecting risks, compounding the harm, and leaving dignity in peril (Dorn et al., 2021; Kharas & Dooley, 2021).” (p. 10, para 1) Readers who are not in the USA may not be familiar with the political parties or political structure there, or the exact details of the Capitol attack in January 2021. If this is to be included, it will need to be qualified by some more information about the USA political system which links to the point being made about propaganda/misinformation and discriminatory policies. Response: We revised this section and provided examples in brackets to clarify for any reader: “Dysfunctional politics can erode individual dignity by endorsing discriminatory policies, such as voter suppression laws and travel bans targeting predominantly Muslim countries. Additionally, propaganda campaigns, which promote biased narratives for political gain, and misinformation, such as false claims about voter fraud or the COVID-19 pandemic, undermine public trust and exacerbate division. On social media, sharp criticisms and personal attacks between American political figures often provoke equally harsh responses, creating an environment where public discourse lacks respect. Dignity emerged as a central theme in the polarized political climate leading up to the U.S. November 2020 presidential election. Thus, advocating for inclusive and respectful politics that uphold the dignity and rights of all individuals remains crucial.” On this point, I think discriminatory policies and propaganda/misinformation are two separate points. Perhaps these could be teased out briefly with reference to empirical data to justify these statements. Response: Thank you for this excellent suggestion. Our revised paragraph above shows that these points are distinct. (P. 10, para 4) It is important to acknowledge that although many healthcare systems operate both publicly and privately, and have social security, healthcare system design varies and the example given from Texas will be unique to the case in the USA which should be clearer – for example in Scotland, ‘insurance status’ would mean something very different in terms of healthcare, and most of the population is able to access primary and secondary care free of charge. Response: We agree with the reviewer and have added two sentences to clarify that differences will depend on context: “Healthcare system designs differ across countries and regional jurisdictions, influencing access to primary and secondary healthcare services without financial burden. These variations can significantly affect individuals' ability to uphold their dignity.” (p. 10, para 6) it is unclear why we are suddenly speaking about end-of-life care. This could be better linked to the empirical data, and could be linked to the above paragraph with a signposting sentence to improve the flow of the article. (p. 10, para 6) It is unclear why we are looking at Kenya and India for these mentions, the connection could be more explicit and better linked back to the concerns of the overall analysis (if the article wants to concentrate on meaning, there will need to be more critical appraisal or expansion on possible meanings here as elsewhere). Response: We revised the text on page 10, paragraph 6, to address these two comments from reviewer 2. The revised text includes an improved explanation for the points raised and several references for the details around dignity, death, and dying: “Respecting an individual's dignity during their end-of-life can be crucial to providing decent end-of-life care (Kennedy, 2016). Dignity is a quality or state of being worthy of respect and honour. As individuals approach the end of life, they often wish for a “good death” that is peaceful, pain-free, and by the individual's wishes and values (Meier et al., 2016). Respecting an individual’s dignity during end-of-life care is crucial for a peaceful transition (Meier et al., 2016). Yet, several barriers emerged during the COVID-19 pandemic, including reduced access to loved ones, overwhelmed healthcare systems, lack of personal care, the need for personal protective equipment, and distressed healthcare workers (Hernandez-Fernandez & Meneses-Falcon, 2021). Additionally, a Kenyan argued that providing protective suits might allow people to give their final respects to their loved ones with dignity. Finally, some followers of an Indian spiritual leader tweeted claims that they had received a divine cure for the deadly virus, telling of healing without medicine. The COVID-19 pandemic highlighted the need for adaptive practices that honour cultural rituals while safeguarding public health, revealing the deep connection between culture, spirituality, and the dignified handling of death. It is not until p. 10 we are told why older adults are a particular focus. This rationale is strong and needs to figure much earlier in the article. Response: We added details to the introduction to clarify our choice of older adults as a focus. Figures and Tables: I find that figures and tables are clear, easily interpreted, and well-formatted. Response: Thank you. Discussion: The start to this section is strong, and the tone is appropriate. The summary of the main themes is particularly helpful. Response: Thank you. There needs to be some discussion in this section of existence of opposing views to the ones presented. The wider context around the findings and any other research which may contradict these findings need to be explored, and linked back to the research gap that this study was filling. Response: Thank you for this suggestion. Our study was not designed to directly challenge or contradict existing research findings. Instead, our contribution lies in giving voice to a largely overlooked or unaddressed area—citizen talk of dignity on Twitter (social media). The revised introduction and theoretical background sections clarify how our work aligns (or does not align) with others’ work. The limitations are clear, perhaps the strengths need to be better situated within the available literature and existing knowledge on this topic. Response: Thank you for this suggestion. We’ve added references to bolster our limitations section. Future directions for the research and practice are clear and meaningful. Response: Thank you. References: Some inconsistencies for referencing (particularly of The Lancet - Lancet, T. as author of one editorial, please correct this in line with style guidelines). Please ensure consistency and revise these. Response: Thank you. We revised the inconsistent referencing." } ] }, { "id": "187199", "date": "08 Mar 2024", "name": "Jan Kietzmann", "expertise": [ "Reviewer Expertise Social media 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 really enjoyed reading this piece. As a social media researcher, I was mostly interested in the methodology for using Twitter data, as I have seen many different approaches. Here, I was thoroughly impressed by the rigor demonstrated by the authors. In terms of the topic itself, I have to admit that I am completely new to the focus on dignity. I have researched online exploitation of people, in particular exploitations of a sexual nature related to children. In my work, I had not considered the dignity literature, and can now see how much value this angle would add to my own work. Thank you, to all of the authors, for introducing this important lens. It is without a doubt that other readers will learn lots, too. I understand that their focus was on COVID related comments but can see how their approach can be replicated for many other instances, far beyond exogenous shocks like COVID.\nIn terms of readability, I would also like to compliment the authors on the attention they paid to ensure that a newbie could easily consume and understand this deep and important topic. Likewise, the methods section was so clear that others, interested in running a Twitter study themselves, could now replicate the methods employed.\nAs far as Twitter data is concerned, the platform comes with the usual pros and cons, and I was happy to see that the authors acknowledged this. As a non-native speaker, I am keenly aware of the dominance of English on Twitter and social media in general. After having lived and/or spent a lot of time in many of the countries that were included, I was curious if one should try to correct for the cultural and linguistic differences between the countries included. I am not convinced that this would strictly speaking be necessary, but the authors could add a short reference to other articles that have used similar geographically diverse settings etc.\nI was looking forward to the discussion, and once again, was impressed with how all of the pieces were connected here. Well, done. The one point I noted was that I was not clear on who the main audience of this piece was. Is it public policy makers? If so, the focus can be sharpened from the beginning to the end. If the audience is broader, maybe a simple table that outlines what different segments can take away from this study would be interesting. Please do not see this as a strong point of criticism, but as food for thought.\n\nAs you can see, I really, really enjoyed this piece and have nothing that I think I'd change. It's a narrowly focused, rigorously executed and superbly documented research study.\n\nAs a complete aside, I enjoyed the emoji word cloud - very interesting. I will now have to learn how to interpret emojis in a rigorous way :-).\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": "187216", "date": "18 Mar 2024", "name": "Uğur Gündüz", "expertise": [ "Reviewer Expertise New Media", "Social Networking", "Media and Cultural Studies" ], "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 entitled \"Upholding Dignity During a Pandemic via Twitter\" provides valuable insights into how people expressed their views on dignity during the COVID-19 pandemic using social media. By analyzing 1946 original tweets using a combination of text analytics and interpretive methods, the study identifies five major themes related to dignity discourse: recognition of dignity as a fundamental right, upholding the dignity of essential workers, preserving the dignity of at-risk populations, preventing cascading disasters exacerbating dignity's decline, and attending to death dignity and the sanctity of life. This paper contributes to the existing literature by highlighting public sentiment on these themes and suggesting how such insights might inform policies and practices to protect and promote dignity. The paper covers an important research area, and overall, the paper is well articulated and presented. However, I would like to see the following concerns addressed: So, I recommend minor revision as indicated below:\n1-The paper would benefit from a more detailed explanation of the selection criteria for the tweets included in the study. Specifically, clarity is needed on how the tweets were deemed relevant to the study's themes of dignity, and any exclusion criteria should be explicitly stated. 2- A brief discussion on the limitations of the interpretive methods used and how they might impact the study's findings would strengthen the research's credibility. 3-While the identified themes are compelling, the discussion would benefit from a deeper exploration of how these themes interact with each other. For example, how does the recognition of dignity as a fundamental right influence actions towards at-risk populations? Expanding on these connections could offer more nuanced insights into the public discourse around dignity. 4-The study mentions a cross-national sample but does not delve into regional variations in dignity discourse. Given the global impact of the COVID-19 pandemic, highlighting any geographic differences in the perception of dignity on Twitter could enrich the analysis. If data allows, a comparison between countries or regions might reveal culturally specific concerns or expressions of dignity. 5-The conclusion section could be expanded to offer more concrete suggestions for future research directions based on the findings. Furthermore, providing specific policy recommendations or interventions to address the issues raised by the study would make the manuscript more impactful. For instance, suggesting ways social media platforms could promote dignity in public discourse during crises could be a valuable addition. 6-There are minor typographical and grammatical errors that need correction to improve the manuscript's clarity and professionalism. Good luck with your research !..\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? Partly", "responses": [ { "c_id": "12734", "date": "10 Dec 2024", "name": "Michael Mulvey", "role": "Author Response", "response": "The paper entitled \"Upholding Dignity During a Pandemic via Twitter\" provides valuable insights into how people expressed their views on dignity during the COVID-19 pandemic using social media. By analyzing 1946 original tweets using a combination of text analytics and interpretive methods, the study identifies five major themes related to dignity discourse: recognition of dignity as a fundamental right, upholding the dignity of essential workers, preserving the dignity of at-risk populations, preventing cascading disasters exacerbating dignity's decline, and attending to death dignity and the sanctity of life. This paper contributes to the existing literature by highlighting public sentiment on these themes and suggesting how such insights might inform policies and practices to protect and promote dignity. The paper covers an important research area, and overall, the paper is well articulated and presented. However, I would like to see the following concerns addressed: So, I recommend minor revision as indicated below: 1-The paper would benefit from a more detailed explanation of the selection criteria for the tweets included in the study. Specifically, clarity is needed on how the tweets were deemed relevant to the study's themes of dignity, and any exclusion criteria should be explicitly stated. Response: The selection of tweets was limited to those written in English, posted in 2020, and containing the keyword \"dignity.\" This approach helped us gather a sample of relevant tweets for our analysis. We aimed to maximize the range of ideas by considering the language used, including words, emojis, hashtags, and mentions in the posts. We specifically focused on tweets that received the highest impressions and engagement. Our analysis concentrated on the 200 most frequently used words related to the topic. While we recognize that less common words exist, they fall outside the main discourse surrounding dignity on Twitter. 2- A brief discussion on the limitations of the interpretive methods used and how they might impact the study's findings would strengthen the research's credibility. Response: Our multi-method approach blended text analysis algorithms and human interpretation – leveraging the strengths of the research team and both quantitative and qualitative approaches. We have also developed our discussion to acknowledge and address limitations, including the representativeness of Twitter users, algorithmic influence, misinformation about stereotypes, focus on English-language tweets, and theme interactions. 3-While the identified themes are compelling, the discussion would benefit from a deeper exploration of how these themes interact with each other. For example, how does the recognition of dignity as a fundamental right influence actions towards at-risk populations? Expanding on these connections could offer more nuanced insights into the public discourse around dignity. Response: We have refined our reporting to highlight interconnected themes. Recognizing dignity as a fundamental right often serves as a foundation for advocating for the dignity of specific groups. It also strengthens the case for actions and policies aimed at preserving the dignity of at-risk populations, such as older adults, essential workers, and individuals experiencing homelessness or domestic abuse. 4-The study mentions a cross-national sample but does not delve into regional variations in dignity discourse. Given the global impact of the COVID-19 pandemic, highlighting any geographic differences in the perception of dignity on Twitter could enrich the analysis. If data allows, a comparison between countries or regions might reveal culturally specific concerns or expressions of dignity. Response: We agree that comparing tweets by geographic location could offer valuable insights. However, this topic wasn't part of our initial agenda. Additionally, our dataset of English-language tweets from a cross-national sample might not be robust enough for reliable comparisons. Therefore, we will leave this challenge for future researchers to explore. 5-The conclusion section could be expanded to offer more concrete suggestions for future research directions based on the findings. Furthermore, providing specific policy recommendations or interventions to address the issues raised by the study would make the manuscript more impactful. For instance, suggesting ways social media platforms could promote dignity in public discourse during crises could be a valuable addition. Response: We have added a conclusion section, focusing on two key points: (1) Dignity as a right and policy target for policy interventions aimed at improving or maximizing dignity for various groups, and (2) Emphasizing the disproportionate impact of COVID-19 on older adults, particularly those with greater care needs and lower socioeconomic status. 6-There are minor typographical and grammatical errors that need correction to improve the manuscript's clarity and professionalism. Response: Thank you for all of your constructive questions! We have carefully proofread and eliminated errors by standardizing the use of Canadian English throughout, correcting the format of in-text citations and referencing, and eliminating unfamiliar acronyms and terminology." } ] } ]	1	https://f1000research.com/articles/12-183
https://f1000research.com/articles/13-478/v1	17 May 24	{ "type": "Research Article", "title": "Partner violence in healthcare settings on breastfeeding mothers: a scientometric analysis", "authors": [ "Endah Wahyutri" ], "abstract": "Background Partner violence is a significant issue affecting women’s health, particularly their breastfeeding practices in healthcare settings. Despite a considerable body of research on this topic, there is a need for a systematic overview of the existing literature to explore its thematic and structural developments.\n\nMethods This scientometric analysis covers a period of 45 years (1977-2022) and identifies 415 peer-reviewed articles published in English before 2023 from the Scopus database. The articles were included if they engaged with themes related to partner violence, healthcare settings, breastfeeding, and mothers, using predefined search terms and filters. Bibliometric data were decoded and visualized using VOSviewer and RStudio to analyze and visualize the scientific dialogue, trends, patterns, and networks in the literature regarding partner violence and its impact on breastfeeding mothers in healthcare settings.\n\nResults The study revealed a concentrated exploration of themes related to violence, healthcare provision, mental health, and women’s reproductive health. The co-occurrence network and thematic mapping illuminated seven pivotal clusters, revealing a complex narrative of interconnected research areas. Niche and emerging themes such as “Depression,” “HIV,” and “Covid-19” were identified, highlighting the evolving landscape of the research domain.\n\nConclusions This analysis provides a panoramic overview of the existing literature, revealing intricate thematic structures and collaboration networks within the research on partner violence and breastfeeding mothers in healthcare settings. The study not only sheds light on the historical and current states of research but also pinpoints potential future directions, thereby assisting scholars and policymakers in navigating and prioritizing subsequent investigative endeavors.", "keywords": [ "Domestic violence", "intimate partner violence", "childbearing", "women's health", "and Bibliometric analysis." ], "content": "Introduction\n\nHealthcare settings, pivotal for supporting breastfeeding mothers, face a paradox due to the persistent issue of partner violence, impacting both maternal and child health and psychological well-being (Wallenborn et al., 2018; Hooker et al., 2020; McCauley et al., 2022; Siwik et al., 2022). The complexities emerging from the intersectionality of partner violence and breastfeeding within healthcare environments prompt a deeper, analytical exploration. Despite the accumulation of research on the impacts of partner violence on breastfeeding mothers, a scientometric analysis providing a cohesive view of the thematic structures and developmental trajectories in this domain is notably absent (Haghani et al., 2020; Chen et al., 2022).\n\nThe existing literature has shed light on the multifaceted impacts of partner violence on breastfeeding mothers, addressing psychological, physical, and social dimensions (Kendall-Tackett, 2007; Lau & Chan, 2007; Frade & de Wet-Billings, 2019). Although many studies have looked at healthcare services and support systems, there is still a need for a comprehensive overview of this research area, mapping out the different themes, networks, and narratives. This overview would help to identify gaps in the literature and suggest directions for future research.\n\nThis study will fill a gap in the literature by conducting a scientometric analysis of partner violence and its impact on breastfeeding mothers in healthcare settings (Sipsma et al., 2013; Wright, Bird, & Frost, 2015; McCauley et al., 2022). The goal is to use quantitative tools to explore the evolution, productivity, and structure of scientific research in this area, and to present a visual and analytical representation of scientific knowledge. This will help to identify research clusters, thematic orientations, and collaborative networks.\n\nEmploying a bibliometric analysis, this research aims to navigate through the literature, providing a structured overview and deriving future research recommendations from a taxonomic scheme created by exploring and visually structuring the literature through co-citation analyses and bibliometric coupling techniques (Sipsma et al., 2013; Wright, Bird, & Frost, 2015; Barbu, 2023). The study intends to enrich scholarly understanding and guide future research and policy development by providing a comprehensive, structured, and in-depth overview of the scientific literature. Subsequent sections will delineate the theoretical framework, methods, bibliometric analysis, results, discussion, and research recommendations, concluding with reflections on limitations and both practical and theoretical implications.\n\n\nMethods\n\nIn this scientometric study, a rigorous exploration of literature concerning partner violence in healthcare settings and its impact on breastfeeding mothers was conducted using the Scopus database. The systematic search strategy focused on key terms related to partner violence, healthcare settings, breastfeeding, and mothers, employing specific inclusion and exclusion criteria with predefined terms and filters:\n\n1. Partner Violence related terms: “Partner Violence” OR “Domestic Violence” OR “Intimate Partner Violence” OR “Abuse” OR “Aggression” OR “Harassment”\n\n2. Healthcare setting descriptors: “Healthcare Settings” OR “Hospital” OR “Clinic” OR “Health Facilities”\n\n3. Breastfeeding and related terms: “Breastfeeding” OR “Lactation” OR “Nursing” OR “Infant Feeding”\n\n4. Target group identifiers: “Mothers” OR “Women” OR “Maternal” resulted in 556 documents found\n\n5. Documents were filtered to be in English, article-only, and published before 2023.\n\nConsequently, a structured strategy using Boolean operators in Scopus was applied to titles, abstracts, and keywords of documents, yielding 415 documents from the period 1976-2022. The articulation of the strategy is as follows: to:\n\n(TITLE-ABS-KEY(“Partner Violence” OR “Domestic Violence” OR “Intimate Partner Violence” OR “Abuse” OR “Aggression” OR “Harassment”) AND TITLE-ABS-KEY(“Healthcare Settings” OR “Hospital” OR “Clinic” OR “Health Facilities”) AND TITLE-ABS-KEY(“Breastfeeding” OR “Lactation” OR “Nursing” OR “Infant Feeding”) AND TITLE-ABS-KEY(“Mothers” OR “Women” OR “Maternal”)) AND PUBYEAR > 1974 AND PUBYEAR < 2023 AND (LIMIT-TO (DOCTYPE,“ar”)) AND (LIMIT-TO (LANGUAGE,“English”))\n\nData extraction was executed systematically using a predefined form, capturing author names, publication year, citations, and keywords (Kho & Brouwers, 2012; Wen et al., 2022). This process was piloted on a subset of articles to affirm its reliability before full-scale implementation. A validation strategy was implementedwith cross-validation of extracted data against Google Scholar as alternative databases. The extracted data, cataloged methodically, facilitated a detailed scientometric analysis (Lu et al., 2018). In the analysis stage, VOSviewer (version 1.6.15) constructed and visualized bibliometric networks, while RStudio (version 4.1.2), using the ‘bibliometrix’ package, conducted statistical analyses. Detailed steps and scripts used in RStudio are provided to ensure reproducibility.\n\nThematic mapping involved an analytical review of keywords and abstracts, with initial themes identified inductively (Haghani et al., 2020; Chen et al., 2022). Subsequent deductive refinement ensured themes were robust and mutually exclusive, based on keyword frequency and conceptual alignment. The synthesis phase utilized VOSviewer and RStudio to decode bibliometric data, generating tables, networks, and graphics. Through bibliographic coupling and co-citation analyses, thematic and structural developments in the research sphere were uncovered, illuminating past research insights and suggesting potential future research directions (Dehghanbanadaki et al., 2020; Mao et al. 2020).\n\n\nResults\n\nA total of 415 articles peer review were derived from Scopus database from 1977-2022.\n\n\n\n1. AND Healthcare setting descriptors: “Healthcare Settings” OR “Hospital” OR “Clinic” OR “Health Facilities” = 48,199 documents results.\n\n2. AND Breastfeeding and related terms: “Breastfeeding” OR “Lactation” OR “Nursing” OR “Infant Feeding” = 3,238 documents results.\n\n3. AND Target group identifiers: “Mothers” OR “Women” OR “Maternal” = 556 documents results.\n\nA multitude of countries have rendered significant scholarly contributions to the field of Partner Violence in Healthcare Settings, particularly impacting Breastfeeding Mothers, spanning a period from 1977 to 2022 (Figure 1). This segment meticulously elucidates the scholarly output of the predominant countries in this specified domain. Table 2 delineates the data pertaining to the top ten countries that have immersed themselves in publishing research related to Partner Violence in Healthcare Settings on Breastfeeding Mothers. The rankings have been meticulously formulated based on the numerical count of publications.\n\nThe graph illustrates the number of articles published each year from 1977 to 2022. It highlights an overall increase in publications over time, with a peak in 2021.\n\nThe tabulated data elucidates that the United States emerges as the pinnacle of scholarly productivity in this domain, boasting 167 publications, thereby reflecting a pronounced emphasis by its academic community on Partner Violence in Healthcare Settings affecting Breastfeeding Mothers. Following in scholarly prominence, Australia secures the second position with 43 publications, while the United Kingdom also asserts its presence significantly, albeit with an unspecified number of publications. Noteworthy is the absence of Asian nations within the rank delineation, revealing a potential geographical disparity in research concentration.\n\nBibliometric coupling (BC) elucidates network dynamics among nations researching Partner Violence in Healthcare Settings, especially its impact on Breastfeeding Mothers. When two documents cite a third, BC occurs, revealing an implicit scholarly connection. Figure 2 visually displays this, where each circle represents a nation and its size indicates scholarly contribution; notably, the United States emerges as significantly contributive (Bair-Merritt et al., 2014; Normandin, 2015; Hill et al., 2016; Sood et al., 2016; Messing et al., 2017; Hooker et al., 2020).\n\nThe examination of interlinkages among international authors can be explored via a co-authorship analysis, segmented by governments. Figure 3 illustrates these collaborative research efforts related to Partner Violence and its impact on Breastfeeding Mothers. The figure highlights the network and various collaborative threads with different colors, with the United States notably establishing robust co-authorship networks, signifying substantial collaborations with authors from various nations (Al-Natour et al., 2014; Scheid et al., 2020; Uysal et al., 2022).\n\nAnother cardinal element of bibliometric analysis focuses on identifying the most productive academic institutions. Table 4 reveals that UNSW Sydney in Australia is the most productive institution, with eleven publications to its name. Western Sydney University and Universidade de São Paulo assume the second and third ranks, with nine and eight publications, respectively. In instances of tied rankings, authors are stratified based on the cumulative citation count of their publications.\n\nThe journal JOGNN - Journal of Obstetric, Gynecologic, and Neonatal Nursing surfaces as the most productive outlet with 17 publications, succeeded by Midwifery with 15 publications. Table 5 encapsulates journal productivity, detailed in terms of publication count on Partner Violence in Healthcare Settings impacting Breastfeeding Mothers across all researchers. Five journals originate from a UK-based publisher and a US-based publisher, while seven journals amongst the top ten are categorized as Q1 journals.\n\nTable 6 delineates author productivity, defined in terms of authors and their affiliations, the number of publications, citation counts, and total citations per publication. Schmied, affiliated with Western Sydney University, Penrith, Australia, emerges as the author with the most substantial volume of articles, totaling five, and accruing 125 citations.\n\nIdentifying highly-cited sources is crucial for assessing authors, journals, and papers. Using data from the Scopus database and ‘Partner Violence in Healthcare Settings on Breastfeeding Mothers’ as a search term, a collection of significant publications was compiled. Table 7 presents those exceeding 200 citations, spotlighting the most-cited publication in this section.\n\nA three-field Plot (Sankey diagram) showing the network between original articles on ‘Partner Violence in Healthcare Settings on Breastfeeding Mothers’ from 1977 to 2022.\n\nVisualizing the flow from authors to affiliations and then to keywords might provide insight into which affiliations are producing research on which topics (Figure 4).\n\nFigure 5 visualizing the flow from journals to keywords and then to countries might provide insights into the kinds of research being published in different journals and where that research is being conducted.\n\nAuthor delved into the topic of “Partner Violence in Healthcare Settings on Breastfeeding Mothers” from 1977 to 2022 and uncovered substantial insights from 415 scientific documents gathered from 259 various sources like journals and books. The topic has been gradually growing over time, with an annual growth rate of about 6.36%, signaling that researchers are increasingly exploring it. Observing the average document age of approximately 14.8 years and an average citation per document of about 26.71, it’s evident that research in this field is not only ongoing but also making a significant impact in the scientific community (Table 8).\n\nThe average age of the documents being 14.8 years with the relatively high average citation per document, standing at 26.71, underscored the enduring relevance and impact of the existing literature. Furthermore, the substantial amount of references, totaling 11,686, and the prolific use of keywords (2,685 Keywords Plus and 823 Author’s Keywords) mirror the depth and complexity of the discourse. Diving into the authorship and collaboration patterns, the data indicates a notable engagement of 1,651 authors, with a commendable emphasis on collaborative research as evinced by the average of 4.29 co-authors per document and an international co-authorship percentage of 11.81%. This analysis provides a panoramic view of the research conducted regarding the impact of partner violence on breastfeeding mothers in healthcare settings. This information is crucial to deepening our understanding and potentially discovering new areas to explore further in future research.\n\nIn the intricacies of academic discourse where knowledge dissemination is paramount, the exploration of thematic occurrences within scholarly publications becomes pivotal. The provided dataset unveils a multitude of research topics, accompanied by their respective frequencies and temporally-oriented quantifiers, including the first quartile, median, and third quartile of publication years. A myriad of topics such as “domestic violence,” “nursing,” “pregnancy,” “breastfeeding,” and “intimate partner violence\" emerge as notably predominant, signaling a substantive focus on humanitarian and child welfare within the academic sphere (Williams et al., 2017; McGarry, 2017; Örmon et al., 2017; Ogbo et al., 2019). Additionally, themes enveloping “stress” and “depression” underscore a robust engagement with mental health discourse (Lepistö et al., 2017) (Figure 6).\n\nWithin this context, a multifaceted examination of violence and maltreatment is palpable through the prevalence of themes like “domestic violence,” “aggression,” and “mistreatment.” Concurrently, the academic lens is also distinctly cast upon healthcare provision, illustrated by the recurrent emergence of topics such as “nursing” and “midwifery.” Furthermore, the appearance of “HIV” and “Women’s health” within the dataset indicates a scholarly inclination toward discussions pertaining to sexual and reproductive health, while the recent emergence of “Covid-19” in 2022 embodies the academe’s response to the prevailing global health crisis.\n\nNavigating through the temporal dimensions of the data, “Child abuse” elucidates an enduring relevance. It spanning from 1998 to 2016, so thereby showcasing its longstanding and potentially evolving scholarly exploration. Contrarily, topics like “Quality of care” and “Childbirth” manifest a more recent relevance, with medians and third quartiles firmly situated in 2022, indicating a shift toward contemporary issues within the field.\n\nConducting a historical analysis of topics could unveil shifts in research priorities and intricate inter-topical relationships through co-occurrence or network analysis (Lepistö et al., 2017; Sprague et al., 2017). Linking topics to impact metrics, like citations, and if possible, geographical data, might offer insight into research influence, regional patterns, or global collaborations. Detailed textual analysis could contextualize these topics within academic discussions and highlight prevailing theoretical frameworks.\n\nThe co-occurrence network, illuminated by the authors’ keywords and analyzed through the biblioshiny app in the bibliometrix environment (De Moraes et al., 2017) (Figure 7). The co-occurrence network analysis using author’s keywords showcases significant and interlinked topics within a specific research domain. An analysis of the bibliometric data, extracted via the Biblioshiny application, reveals insights into the interconnected thematic areas in the body of literature related to women’s health, violence, and related healthcare contexts.\n\nTwo main themes, Intimate Partner Violence/Domestic Violence and Nursing/Pregnancy, emerge prominently in the research (Olive, 2007; Reisenhofer & Seibold, 2013; Crombie et al., 2017; Ahmad et al., 2017). The former plays a pivotal role, acting as major connectors in the network and potentially tying research on personal relationship violence to various sub-themes (Ferri et al., 2016). Meanwhile, Nursing and Pregnancy themes underline a robust research focus on healthcare provision and pregnancy-related issues, exploring practices and challenges in these areas (McKinley et al., 2019; Van Niel et al., 2020). Table 9 analyzes key themes and characteristics within the co-occurrence network.\n\nThe biblioshiny-assisted co-occurrence network analysis identifies seven key clusters within women’s health research. The Violence and Screening cluster focuses on exploring and intervening in violent scenarios. Healthcare Contexts and Substance Abuse delve into management and mitigation strategies in healthcare settings, particularly concerning violence and substance abuse in nursing and emergency departments. Pregnancy and Mental Health emphasize the intertwined relationship of mental health, particularly depression, with pregnancy and abuse. The Breastfeeding within the HIV Context cluster potentially provides insights into guidelines and challenges affecting maternal and child health (Haber et al., 2021). Childbirth and Healthcare Quality, especially in the Covid-19 context, explore impacts and shifts in practices and provider roles during the pandemic (Vrazo et al., 2020). The Nursing Profession and Women’s Issues cluster spotlight the multifaceted role of nurses in navigating challenges and interventions in women’s health. Lastly, Women’s Health and Violence focus on the broad explorations in women’s health, zooming in on violence and its diverse impacts and management strategies in healthcare.\n\nMoreover, “Depression” and “HIV” stand out as niche themes, potentially providing in-depth insights into specific aspects of broader themes (Mpody et al., 2019; Myburgh et al., 2020; Hampanda et al., 2021). “Covid-19” emerges as a pivotal theme, likely dissecting the pandemic’s diverse impacts on women’s health and healthcare provision, intersecting with various thematic clusters (Janevic et al., 2021). The interplay of these themes offers a nuanced understanding of the domain, showcasing the interconnectedness of various aspects of women’s health and well-being (Figure 8).\n\nThe thematic map and clusters reveal a multifaceted academic dialogue where themes intersect and evolve, shaping a nuanced scholarly narrative. The Biblioshiny package in RStudio, using a minimum frequency of three per thousand words, helps navigate through the complex thematic map, witnessing the convergence and divergence of themes, each reflecting different scholarly facets. The objective is to interpret thematic clusters, discerning their centrality and prominence within the academic environment.\n\nBasic themes like “Substance use disorder,” “Substance abuse,” “Breastfeeding,” and “Pregnancy” act as foundational pillars in the research (Bogen & Whalen, 2019; Yonke et al., 2019). These themes exploring dynamics in their respective domains and serving as connectors to various themes, providing a comprehensive exploration of related topics. Specifically, “Breastfeeding” and “Pregnancy” are crucial in discussing maternal health and wellbeing in the primary article’s context.\n\nMotor themes, including “Gender-based violence” and “Domestic violence,” serve as core themes that shape the discourse, navigating through various health, socio-legal, and psychological dimensions. This themes especially exploring the intersections and impacts of violence within healthcare and maternal contexts (Martin-de-las-Heras, et al. 2019). “Domestic violence,” with high Callon Centrality and cluster frequency, acts as a cardinal node, influencing and being influenced by various thematic dynamics (Kalra et al., 2021).\n\nThemes like “Child protective service” and “Australia” indicate areas of emerging or receding research focus, with geographical and socio-cultural contexts shaping these themes. “Australia” points to region-specific dynamics (Dawson et al., 2019; Hooker et al., 2020; Coupland et al., 2021). “Depression” might indicate a broader discourse intersecting mental and maternal health, and experiences of violence (Dodge et al., 2019; Janevic et al., 2021).\n\nNiche themes such as “Inpatients” and “HIV” explore specific healthcare settings and medical contexts, while “Intimate partner abuse” and “Adolescents” explore specific demographic and relational dimensions within the broader health and violence discourse (Rujumba et al., 2021). The collective themes craft a multifaceted narrative, intertwining aspects of health, violence, demographics, and potentially, geographical and socio-cultural dynamics, potentially hinting at future research trajectories and evolving contexts in this domain. Further exploration could provide deeper insights into the complexities and nuances embedded within this academic discourse.\n\n\nConclusion\n\nThe scientometric analysis of literature related to partner violence in healthcare settings, with a specific focus on breastfeeding mothers, unveiled an intricate and multifaceted research domain. This study, which encompassed 415 documents from the Scopus database spanning from 1977 to 2022, provided a comprehensive overview of the evolving thematic structures, collaborative networks, and influential works in the field. The research revealed a robust engagement with themes such as “Domestic Violence,” “Nursing,” “Pregnancy,” “Breastfeeding,” and “Intimate Partner Violence,“ among others, elucidating a scholarly commitment to exploring humanitarian, child welfare, and mental health discourses. Key clusters identified in the co-occurrence network and thematic maps—such as Violence and Screening, Healthcare Contexts and Substance Abuse, and Pregnancy and Mental Health—highlighted substantial intersections among the various aspects of women’s health and wellbeing. Notably, the emergence of “Covid-19” as a significant theme pointed towards the academia’s response to the global health crisis and its intersectionality with women’s health themes. The identified niche, emerging, and primary themes, alongside geographical and institutional productivity, underscored the multi-dimensional nature of the academic dialogue within the sphere of partner violence in healthcare settings. This research not only provides a panoramic view of the extant literature but also paves the way for identifying gaps, potential future directions, and evolving thematic structures within the research domain, thereby contributing to the ongoing scholarly dialogue.\n\nThis research faced several limitations requiring acknowledgment. Firstly, relying solely on the Scopus database and English-language articles may introduce selection bias, potentially overlooking valuable insights from non-English research and limiting the study’s cultural and geographical scope. Additionally, the use of keyword-based search strategies might omit relevant studies using different terminologies. The defined time frame until 2022 could exclude the newest research developments, and while VOSviewer and RStudio are robust for bibliometric analysis, they may not fully decipher complex data patterns. Moreover, scientometric and bibliometric analyses’ inherent biases could undervalue contributions from emerging researchers or institutions in low/middle-income countries. Lastly, the quantitative focus might miss the qualitative depth and contextual nuances intrinsic to partner violence impacting breastfeeding mothers in healthcare settings. Future research could navigate these limitations by diversifying data sources, incorporating multilingual studies, and employing both quantitative and qualitative analyses for a more holistic exploration of the domain.", "appendix": "Data availability\n\nThe bibliometric data generated using VOSviewer and RStudio, as well as the scripts used for the analysis.\n\nhttps://doi.org/10.6084/m9.figshare.24884151.v1 (Wahyutri, 2023).\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nAcknowledgements\n\nI would like to thank QUVAE Research and Publications for their guidance in submitting the underlying data to Figshare and for assisting with the F1000Research submission process.\n\n\nReferences\n\nAhmad I, Ali PA, Rehman S, et al.: Intimate partner violence screening in emergency department: a rapid review of the literature. J. Clin. Nurs. 2017; 26(21-22): 3271–3285. 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PubMed Abstract \| Publisher Full Text\n\nMcCauley H, Lowe K, Furtado N, et al.: Essential components of postnatal care – a systematic literature review and development of signal functions to guide monitoring and evaluation. BMC Pregnancy Childbirth. 2022; 22(1): 448. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMcGarry J: Domestic violence and abuse: an exploration and evaluation of a domestic abuse nurse specialist role in acute health care services. J. Clin. Nurs. 2017; 26(15-16): 2266–2273. PubMed Abstract \| Publisher Full Text\n\nMcKinley SK, Wang LJ, Gartland RM, et al.: \"Yes, I’m the Doctor\": One Department’s Approach to Assessing and Addressing Gender-Based Discrimination in the Modern Medical Training Era. Acad. Med. 2019; 94(11): 1691–1698. PubMed Abstract \| Publisher Full Text\n\nMessing JT, Campbell JC, Snider C: Validation and adaptation of the danger assessment-5: A brief intimate partner violence risk assessment. J. Adv. Nurs. 2017; 73(12): 3220–3230. PubMed Abstract \| Publisher Full Text\n\nMpody C, Thompson P, Tabala M, et al.: Hepatitis B infection among pregnant and post-partum women living with HIV and on antiretroviral therapy in Kinshasa, DR Congo: A cross-sectional study. PLoS One. 2019; 14(5): e0216293. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMyburgh D, Rabie H, Slogrove AL, et al.: Horizontal HIV transmission to children of HIV-uninfected mothers: A case series and review of the global literature. Int. J. Infect. Dis. 2020; 98: 315–320. PubMed Abstract \| Publisher Full Text\n\nNormandin PA: Identifying Maternal Intimate Partner Violence in the Emergency Department. J. Emerg. Nurs. 2015; 41(5): 444–446. PubMed Abstract \| Publisher Full Text\n\nOgbo FA, Ezeh OK, Khanlari S, et al.: Determinants of exclusive breastfeeding cessation in the early postnatal period among culturally and linguistically diverse (CALD) Australian mothers. Nutrients. 2019; 11(7): Article 1611. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOlive P: Care for emergency department patients who have experienced domestic violence: A review of the evidence base. J. Clin. Nurs. 2007; 16(9): 1736–1748. PubMed Abstract \| Publisher Full Text\n\nÖrmon K, Torstensson-Levander M, Sunnvist C, et al.: The duality of suffering and trust: Abused women’s experiences of general psychiatric care - an interview study. J. Clin. Nurs. 2017; 26(15-16): 2439–2451. Publisher Full Text\n\nReisenhofer S, Seibold C: Emergency healthcare experiences of women living with intimate partner violence. J. Clin. Nurs. 2013; 22(15-16): 2253–2263. PubMed Abstract \| Publisher Full Text\n\nRujumba J, Homsy J, Mbazzi FB, et al.: Pregnant women, their male partners and health care providers’ perceptions of HIV self-testing in Kampala, Uganda: Implications for integration in prevention of mother-to-child transmission programs and scale-up. PLoS One. 2021; 16(6): e0253616. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nScheid CR, Paulson JL, Miller-Graff LE: Feelings and feedings: Psychopathology and breastfeeding attitudes in women with a history of intimate partner violence. Psychol. Trauma Theory Res. Pract. Policy. 2020; 13(3): 394–402. PubMed Abstract \| Publisher Full Text\n\nSipsma HL, Magriples U, Divney A, et al.: Breastfeeding behavior among adolescents: Initiation, duration, and exclusivity. J. Adolesc. Health. 2013; 53(3): 394–400. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSiwik E, Larose S, Peres D, et al.: Experiences of At-Risk Women in Accessing Breastfeeding Social Support During the Covid-19 Pandemic. J. Hum. Lact. 2022; 38(3): 422–432. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSood R, Novotny P, Faubion SS, et al.: Self-reported verbal abuse in 1300+ older women within a private, tertiary women’s health clinic. Arch. Gerontol. Geriatr. 2016; 66: 62–65. Publisher Full Text\n\nSprague C, Hatcher AM, Woollett N, et al.: How Nurses in Johannesburg Address Intimate Partner Violence in Female Patients: Understanding IPV Responses in Low- and Middle-Income Country Health Systems. J. Interpers. Violence. 2017; 32(11): 1591–1619. PubMed Abstract \| Publisher Full Text\n\nUysal Yalçın S, Zonp Z, Dinç S, et al.: An examination of effects of intimate partner violence on children: A cross-sectional study conducted in a paediatric emergency unit in Turkey. J. Nurs. Manag. 2022; 30(6): 1648–1657. PubMed Abstract \| Publisher Full Text\n\nVan Niel MS, Bhatia R, Riano NS, et al.: The Impact of Paid Maternity Leave on the Mental and Physical Health of Mothers and Children: A Review of the Literature and Policy Implications. Harv. Rev. Psychiatry. 2020; 28(2): 113–126. 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PubMed Abstract \| Publisher Full Text" }	[ { "id": "322908", "date": "16 Sep 2024", "name": "Balázs Juhász", "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 author explores the state of the scientific literature of a very important public health issue, namely the partner violence in healthcare settings on breastfeeding mothers. Through the analysis of trends and topics in the field, we get a valuable insight into the state of research. I appreciate the choice of topic, and the efforts of the author dedicated to better our understanding of the field. I endorse the approval of the article, however in my opinion, before final approval, some adjustments should be made.\nGENERAL:\n• The overall language and phrasing of the manuscript is coherent and easy to follow, nevertheless the English language accuracy of the article could be bettered. • A structured discussion section separate from the results could divide better the objective results from the authors conclusions. Alternatively, if the author prefers to interpret the data at the same section where it is presented, the section should be titled as “Results and discussion”. • Getting further input from a reporting guideline, such as the one provided by Milad Haghani [Ref 1] could better the reporting quality of the paper.\nABSTRACT:\n• The abstract overall is well written, emphasizing the pivotal points and providing a good summary of the research done. • In my opinion, the number of identified articles should be stated at the results section of the abstract, not at the methods. • The results section of the abstract should elaborate more details on the most important findings, such as the identified trends, naming the seven pivotal clusters, etc.\nBACKGROUND:\n• The introduction is well written, down to point, not too extensive, gives a good summary of the topic and describes well the aim of the research. • The definition used by the author of partner violence in healthcare settings on breastfeeding mothers should be stated. What was considered as violence? Are they referring to violence occurring specifically in the healthcare setting or any violence that happens to breastfeeding women who use healthcare? • By providing data on the frequency and/or health burden of the phenomenon (e.g. prevalence or incidence rates), the magnitude of the problem could be further emphasised.\nMETHODS:\n• The search strategy is presented completely, transparently and is fully replicable. • The used keywords are presented three times (twice in the text, once in the table). The way of presentation is down to the preference of the author, but redundancy should be minimized. • The construction of the search strategy and the selection of keywords seems adequate and relevant to the research question. • The search strategy introduced some limitations and filters, such as literature search starting from 1974, and limiting the search to only English language articles. The reason for this should be addressed in the manuscript. • A brief description of what steps were taken to avoid the infiltration of results with false positives could be an important quality indicator of the article. • The author mentions a cross-validation against Google Scholar. Details of this could be provided in the manuscript or as supplementary material. • I recognize the publication of the dataset as a very good research practice. In the manuscript, it is stated that the script to create the figures is also provided, however I was unable to locate this. It is recommended to attach this as a separate file, and cite them in the main text (e.g. Supplementary material 1, 2, or Appendix 1, 2)\nRESULTS:\n• The results of the research are well detailed. Maps and graphics were created with robust programs, such as VOS viewer and R studio, and are presented in high resolution. The use of colours makes the figures easy to understand, and labels are easy to read. • On the other hand, the overall clarity and precision of text should be bettered, and some issues should be addressed: • There are mistakes in the labelling of tables. In the text it says, “Table 2 delineates the data pertaining to the top ten countries […]”, meanwhile Table 2 is about the annual publication numbers, and Table 3 details the top five countries. The in-text citations of all tables and figures should be double checked. • Some tables and figures are redundant. For example, Figure 1 and Table 2 carries the same information. It is sufficient to present Fig. 1 in the article, while Table 2 could be provided as supplementary data. • The author concludes that the most productive countries were USA, Australia, UK and Canada. There could be a significant selection bias here, as the search strategy filtered for only English language articles. This is later mentioned as a limitation of the study, but it must be pointed out where the results of the most productive countries are discussed. • Where the results of Table 6 (top 10 productive authors) are detailed, although might be obvious, a caveat of productive authors in this field should be added, as one author could focus on various other research topics. • In Fig. 8, some of the themes does not seem to be mutually exclusive (e.g. substance abuse and substance use disorder, or domestic violence and violence). An explication for the categorization of themes could be provided. • Redundancy could be further reduced by avoiding repeating the same information in tables and in texts. Instead, where tables are used, the text should focus on providing extra information that cannot be tabulated. • Generally, the titles of the figures and tables should be easier to understand and meaningful on their own. (For example, Fig. 8 is titled as “Thematic map of the journal” – which journal is the author referring to?)\nCONCLUSIONS:\n• The conclusion is well written and cohesive. • The identified gaps, potential future directions and evolving thematic structures within the research domain mentioned at the end of the conclusions could be briefly summarised.\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? Yes\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "12625", "date": "10 Dec 2024", "name": "Endah Wahyutri", "role": "Author Response", "response": "I have carefully reviewed the feedback and made the necessary changes to the manuscript. The language and phrasing have been improved for better coherence and readability. The discussion section has been restructured to clearly separate the objective results from the author's conclusions. Furthermore, the reporting quality of the paper has been enhanced by incorporating input from the reporting guideline provided by Milad Haghani. In the abstract, the number of identified articles has been appropriately stated in the results section, and more details on the important findings have been included. Additionally, the definition of partner violence in healthcare settings on breastfeeding mothers has been clarified in the background section. The search strategy has been presented more transparently, and the limitations and filters applied have been thoroughly addressed. Furthermore, steps taken to avoid false positives have been described, and details of the cross-validation against Google Scholar have been provided. The labeling of tables has been corrected, and redundant tables and figures have been appropriately addressed. The potential selection bias in identifying the most productive countries has been acknowledged, and a caveat about productive authors in the field has been included. Lastly, efforts have been made to ensure that the titles of the figures and tables are clear and meaningful on their own." } ] } ]	1	https://f1000research.com/articles/13-478
https://f1000research.com/articles/13-695/v1	26 Jun 24	{ "type": "Systematic Review", "title": "Only severe Injuries are effectively reduced by muscles' injury prevention protocols in football players: A systematic review", "authors": [ "Rihab Moncer", "Marwa Ghanmi", "Dhekra Chebil", "Sana Bhiri", "Iheb Belhadj Youssef", "Amani Kacem", "Sana Salah", "Sahbi Mtaouaa", "Sinen Frigui", "walid Ouannes", "Sonia Jemni", "Marwa Ghanmi", "Dhekra Chebil", "Sana Bhiri", "Iheb Belhadj Youssef", "Amani Kacem", "Sana Salah", "Sahbi Mtaouaa", "Sinen Frigui", "walid Ouannes", "Sonia Jemni" ], "abstract": "Background Muscle injuries are the most frequent in football and may lead to the end of a player’s career. Many studies have evaluated the effectiveness of prevention programs for all injury types. Few studies have evaluated the effects of exercise on muscle injuries.\n\nMethods A documentary search was conducted in October 2022 from three databases: Medline via its PubMed interface, Google Scholar, and COCHRANE databases. We only included original articles published between October 2012 and October 2022 with a high level of evidence. The research was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses criteria. The target population consisted of professional and semi-professional footballers. The intervention in question was structured injury preventive protocols, including specific training or pre-established protocols such as the FIFA 11+ program, compared to each other or with regular training sessions. The main outcomes were the incidence rates of muscle injuries and severe injury in each group, as well as the time loss to injury.\n\nResults Seven randomized controlled trials with a high level of evidence confirmed using the PEDRO scale were included. There were 3815 male professional football players. There was heterogeneity in the age, hours of exposure, and duration of the intervention. The prevention protocols used were FIFA 11+, bounding exercises, NHE, and stretching exercises. Structured exercises significantly reduce the severity of muscle injury and, consequently, time loss to injury.\n\nDiscussion Despite the diversity of prevention programs, teams still lack standardized programs. Structured exercises, such as FIFA 11+ and bounding exercises, which combine strengthening, body kinesthesic awareness, and neuromuscular control, reduce the incidence and severity of muscle injury. FIFA 11+ is the first and most complete structured program, but lacks some exercises, such as adductor strengthening.\n\nConclusion This review showed the interest in structured exercises in reducing severity and time loss to injury in professional football players.\n\nRegistration This review was registered on 8/17/2023 in the International Prospective Register of Systematic Reviews under the reference code CRD42023452202.", "keywords": [ "muscle", "wounds and injuries", "prevention", "exercise", "football", "soccer", "systematic review" ], "content": "Introduction\n\nMuscle injury (MI) is the most common injury in football.1 The incidence of total injuries varies between 1.06/1000 hours (hours) and 5.87/1000 hours of training and competition.2 MI is typically caused by repeated eccentric muscular contractions.3 The relationship between eccentric muscular contraction and MI is confirmed by the lower occurrence of MI during cycling and swimming, two sports based almost exclusively on concentric muscle contraction.4,5 Hamstring muscles are the most frequent site of MI in professional football, where they cause the highest number of absence days during the sporting season (25% of all absences due to injuries), with an average recovery time of nine days.6,7\n\nEccentric strengthening is considered effective in the prevention of MI because it results from an eccentric strength deficit in comparison with the concentric force of hip extensors in elite sprinters.6 Among sports science researchers who aimed to develop a global and universal program for injury prevention, F-MARC conducted the first study on exercise-based prevention of football injuries, showing 21% fewer injuries in the intervention group (IG) than in the control group (CG).7 The interventions focused on improving the structure and content of the training by educating and supervising coaches and players. The program included preventive interventions, such as improvement of warm-up, regular cool-down, taping of unstable ankles, adequate rehabilitation, promotion of the spirit of fair play, and 10 sets of exercises. It is designed to improve coordination, stability of the ankle and knee, and flexibility and strength of the trunk, hip, and leg muscles. Subsequently, F MARC, in cooperation with international experts developed the FIFA11 (Federation Internationale Football Association).8\n\nEvidence of the efficacy of this program has been reported in several studies.9,10 Many trainers have judged that it is a lengthy program and that players find it difficult to adhere to it, given the absence of exercises involving the ball.9 Therefore, modifications were added; hence, FIFA11+.11\n\nFor more than 10 years, researchers and researchers have been performing exercises to identify the most effective ones in MI prevention. Knowing the most effective program with evidence-based practice will help improve prevention measures and reduce the incidence of MI in football.\n\nThe objective of this study is to report, through a systematic review of current scientific research of high level of proof (randomized controlled trial) the efficacy of programs for the prevention of muscle injuries for football players.\n\n\nMethod\n\nThe present research work is a systematic review of the biomedical literature to provide an analysis of the preventive effect of programs for muscle injuries in football. A documentary search was conducted in December 2022 from three databases: Medline via its PubMed interface, Google Scholar, and COCHRANE databases, through the following documentary request: Which exercises are effective in MI in professional football players?\n\nThe MESH terms used were prevention, muscle, injury, wounds and injury, soccer, and exercise.\n\nThe combinations of these different keywords generated the following search equation:\n\n“Prevention” OR “Prévention” AND “muscle” AND “Wounds and injury” OR “plaies et blessures” AND “Exercise” OR “Exercice” AND “Football” OR “SOCCER”\n\nThe research question was methodically put in the PICO format to identify appropriate keywords. After exploring the digital databases, we studied the bibliographical references of relevant articles.\n\nWe included an independent double lecture of the title/abstract and full texts by two trained specialists in rehabilitation and physiotherapy (R. M) and preventive and health public medicine (D. CH), and studies on the preventive effect of program or exercise on MI.\n\nI- SOURCES OF INFORMATION AND SEARCH\n\n1. COMPUTER RESEARCH: We used scientific search engines: PUBMED, Google Scholar and COCHRANE database.\n\n2. MANUAL RESEARCH: After exploring the digital databases, we studied the bibliographical references of the articles deemed relevant.\n\nII- ELIGIBILITY CRITERIA\n\n1. CHARACTERISTICS OF THE STUDY\n\nOur question was methodically put in the PICO format:\n\n○ POPULATION\n\n○ Professional and semi-professional male and female footballers\n\n○ Amateurs were excluded due to a lack of dedicated physiotherapy staff on their football teams and the lower time of training and playing compared to their professional counterparts.\n\nWe also excluded Gaellic, Futsal, Australian and American Football players from the study.\n\nChildren were also excluded because they did not have the same number of hours playing and the same type and frequency of injury.\n\n○ INTERVENTIONS\n\nApplication of structured muscle injury preventive protocols, including specific training, physical therapy, or pre-established protocols such as the FIFA 11+ program, aimed at reducing the risk of MI.\n\n○ COMPARISONS\n\nWe included controlled studies that compared different protocols and regular training sessions with an acceptable number of hours of exposure to training and competitions.\n\n○ OUTCOMES\n\nIR (Incidence Rate) of injuries and severe injuries per 1000 h (Injury/1000 h) of exposure and time loss to injury, including training and official competitions.\n\n2. INCLUSION CRITERIA\n\nWe included original articles published between December 2012 and October 2022 available in digital versions, written in French or English with a high level of evidence (Level I and II), which evaluated the efficacy of the prevention program on muscle injury with at least 2 months of follow-up, monitoring players between training and competition.\n\n3. EXCLUSION CRITERIA\n\nWe excluded:\n\nStudies of the meta-analysis or literature review type: These two types of publications were excluded from our research to avoid duplicating the results of the clinical trials, which may also be included in these reviews.\n\no Communications in congresses, consensus conferences of learned societies.\n\no Studies that did not correspond to research quotes (adjustment, technical notes, letters to the editor, case reports, interviews, opinions, single commentaries, etc.).\n\no Works not available for consultation or only available in paper format.\n\nArticles with less than two months of follow-up or with too few total hours of exposure between training and competition were excluded.\n\nStudies that assessed MI in amateur footballers were not included because the risk of injury in amateur teams is much lower than that in professional clubs, which would considerably increase the risk of bias in statistical analysis.\n\no Studies that assess the adherence to a prevention program.\n\no Studies that have studied the effect of exercise on performance and on weight joint injuries, central pivot.\n\no Articles that study the effect of exercise on performance, sleep, psychology\n\nIII- SEARCH STRATEGY\n\nThe study was conducted following the guidelines for writing a literature review according to the PRISMA criteria (Preferred Reporting Items for Systematic Review and Meta-Analyses).12 It was registered in the International Prospective Register of Systematic Reviews (PROSPERO) under the reference code CRD42023452202. The total number of identified articles was documented based on the search strategy. The references of the articles were recorded using the Zotero https://www.zotero.org/downloadbibliographic software https://www.zotero.org/download to sort the articles and eliminate duplicates.\n\nIV- SORTING AND SELECTION OF ARTICLES\n\nFor article selection, the reviewers screened the titles and/or abstracts. Duplicates and articles were removed first. If the title and/or abstract were not sufficient for inclusion or exclusion, the full-text article was retrieved and read independently by two reviewers (R. M and D. CH). In addition, a cross-reference search of the selected articles was performed to identify agreement and differences. If consensus could not be reached among the investigators regarding the inclusion or exclusion of an article, a third reviewer (S. B.) was consulted to make the final decision.\n\nFinally, the two assessors completed data extraction independently.\n\nThe selected articles were sorted according to their general characteristics (main author, journal where the study was published, year of publication, country where the work was carried out, type of study, and level of proof).\n\nV- Quality of article assessment and risk of bias\n\nGiven the heterogeneity between the study design and methodology adopted, the rating scale Physiotherapy Evidence Database (Pedro) scale, available online,13 was used to critically appraise the quality of the studies selected and retained in the present systematic review.\n\nThe PEDro scale was used to rate each article and inform clinicians of its degree of quality. The scale also assessed the risk of bias of the studies in its items (randomization, allocation concealed, blinding subject, blinding therapist, and blinding assessors). The last four items of this scale evaluate the measurement bias of the studies. A Pedro scale score higher than 6/10 had a low risk of bias.\n\nVI- Data extraction and synthesis\n\nFollowing the critical appraisal, relevant data were systematically extracted and tabulated from the included studies. Extracted data included the study title, year, authors, study type, population characteristics, description of exercises applied by researchers, main findings of the studies, and methodological quality assessment scores.\n\nThe main findings were as follows: the difference between the IG and CG according to their injury rate (IR), severity of injuries/1000h of play, and time loss due to injury. All outcomes were analyzed using a critical narrative synthesis.\n\n\nResults\n\nA total of 214 articles, with 112 articles from Medline, 31 articles from GOOGLE SCHOLAR, and 71 articles from the Cochrane library, were initially identified. Of these, 42 duplicates were excluded. The remaining 172 articles were screened against the eligibility criteria by title and abstract to exclude those clearly not relevant to the review topic. Following this, the full texts of 59 articles of potential relevance were retrieved, and 39 were excluded based on the eligibility criteria. Once the screening and selection processes were complete, seven articles were eligible for inclusion in the present systematic review. (Figure 1)\n\nAll the studies included were cluster randomized controlled studies, except the study by Azuma et al., which was a randomized controlled trial [14] (Table I).\n\nThe included studies specified the eligibility criteria and concealed allocations. In addition, the participants were randomized into similar groups in all studies. However, none of them blinded the participants, conductor, or examiner, except for Whalan et al. The results were adequately reported for injury rates in all the studies. The mean PEDRO scale of all included studies was > five. Thus, the average quality of the included studies was satisfactory (table II)\n\nA total of 3815 players were included in the seven studies, and all included participants were Males. The characteristics of mean age, BMI, and total exposure time are presented in Table III.\n\nThe seven included studies were cluster randomized to avoid contamination within the same team, except for the study by Azuma et al.14\n\nSeveral programs have been proposed in these studies. There was heterogeneity in the intervention duration.\n\n1- Fifa11+\n\nIn a total of 265 players included, Hammes et al. conducted a cluster randomized study where they proposed FIFA 11+ for 119 IG and 146 CG for 9 months.15\n\nIn addition, in the study by Granelli et al., 675 players performed FIFA11+ compared to the usual training session performed by 850 players for 8 months.16\n\nOwoeye et al. 212 performed FIFA11+ and 204 usual training sessions for 6 months.17\n\nWhalan et al. suggested another type of control between groups: for CG 408 players, a modified FIFA 11+, in which “Part 2” of the program was performed at the end of training, as opposed to “Part 1”and “Part 3,,” which were performed at the beginning. In the IG, 398 players performed FIFA 11+.18\n\n2- BEP\n\nFor 400 players, Van de Heuf et al. proposed BEP (concentric to eccentric than plyometric exercises) for 229 players in IG and 171 usual training sessions in CG for a period of 12 weeks.19\n\n3- NHE\n\nHasebe et al. suggested NHE for 156 players in the CG and usual training for 103 players in the CG for 27 weeks.20\n\n4- Stretching\n\nFor 40 weeks, Azuma et al. conducted a stretching protocol for 64 IG players. Physical therapists provided personal exercise instructions three times a week. The program was made of 10 stretching exercises and didn’t require the use of any equipment. Tight body parts were held for 30 s with a rest interval of approximately 30 seconds. Similar exercise instructions were provided during the cooling period. The players were permitted to continue the exercise program themselves after the intervention period. Players in the CG performed the usual training session.14\n\nAll studies have shown that structured programs reduce MI rates, but significant differences have been observed, especially in the severity of injuries.\n\nThe bounding exercise applied in the IG in the study by Van de Heof et al. did not significantly reduce IR or severity of injury.19\n\nFIFA 11+ used in the studies of15–18 showed significant differences in severe injuries in most cases.\n\nIn the study by Azuma and Someya, the stretching protocol significantly reduced IR and severe injury.14\n\nThe NHE reduced IR without a significant difference in the study by Hasebe et al.20 (Table IV)\n\n\nDiscussion\n\nDespite the existence of multiple effective programs, teams still lack standardization in terms of MI prevention programs. This review aimed to identify the most effective programs for the prevention of MI in football. From a global view, structured programs reduce the MI rate, but more significantly, its severity and time loss to injury.\n\nPreviously, many reviews were interested in either evaluating the efficacy of one particular program10 or the efficacy of programs on one particular group of muscles.21–22 In the hamstrings review,21 there was not a sufficient number of participants included in the studies (less than 20), and only the study by Whalan et al., which was included in our review, included more than 30 participants.\n\nFIFA 11+ is the most commonly used MI prevention program.15–18 FIFA 11+ is a comprehensive warm-up program aimed at improving muscular strength, body kinesthetic awareness, and neuromuscular control during static and dynamic movements. This program consists of three parts, 15 different exercises with a focus on neuromuscular effects on the lower extremities and core. The first and third parts include running exercises; it begins “at slow speed combined with active stretching and trolled partner contacts” and finishes “at moderate/high speed combined with planting/cutting movements”. These parts include the dynamic actions and accelerations. Part 2 consisted of six strength, plyometric, and balance exercises with a focus on core and leg strength, proprioception, and stability. Three variation levels were provided for each exercise. The 11+ program was designed to be delivered as a 20-25 min “warm-up” before commencing other training activities, without the need for specialized equipment.9 Its effectiveness has not been fully documented in the literature; Steffen et al., in 2013, reported an overall significant injury reduction of 72% based on secondary data analysis.23\n\nThere were only significant differences in the study by Hammes et al.15 in terms of IR between the IG and CG. In the other studies included, FIFA 11+ did not show a reduction in IR, but there was a significant difference in terms of the severity of injury.\n\nAll players in the included studies were male, but FIFA 11+ was tested several times in female soccer players24–25; however, the number of participants was not enough to conclude efficacy.\n\nIn the study of Mckey et al,26 authors evaluated the impact of high versus low application of FIFA 11+ and found significant difference in the study of High application of FIFA 11+ but the number of participants was few “23 females” to consider those findings.26\n\nFrom the seven studies included, Whalan et al.18 used a particular control within group; for IG, they applied classic FIFA 11+ described above, and for CG, they proposed parts 1 and 3 before training and part 2 after training. However, significantly fewer severe injuries and days lost to injury were observed in the “P2 post” group in this study. The authors explain these findings by better compliance and adherence in the P2 post-training group, which provides a simple and practical method to potentially improve the effectiveness of the 11+ program and compliance with the 11+ program.18\n\nIndeed, implementation of FIFA 11+ was initially difficult, and there was a lot of resistance to adherence from both coaches and players.11,23,25,26 Many authors have studied the factors influencing adherence to the FIFA 11+ program and identified barriers to its implementation, such as program duration and exercise difficulty/fatigue. They have also shown that this adherence may be related not only to the effectiveness of the program itself, but also to beliefs and practice in football coaches’ attitudes.26\n\nIn addition to FIFA 11+, Vand de Hoef used a complete bounding exercise programme. Players perform a gradual build-up of walking lunges, tripling and drop lunges, and bounding (alternating leg jumps).19\n\nAfter a period of 12 weeks of buildup, players will continue BEP for the rest of the season, two sessions a week. The first gradual build-up should improve functional movement patterns and increase strength.27\n\nThe bounding exercise, in the literature referred to ‘a running bound’ or ‘alternate leg bounding’, is a popular running-specific exercise that can be categorized as plyometric exercise. These plyometric exercises consist of three phases: eccentric pre-stretch, amortization phase (time between eccentric and concentric contractions), and concentric shortening phase.28\n\nOther studies focused only on a group of muscles. For example, Hasebe et al. studied the effectiveness of NHE on hamstring injury (HI).20 The athletes were asked to use their arms and hands to buffer the fall, let their chest touch the surface, and return immediately to the starting position by pushing with their hands to minimize loading in the concentric phase. In this study, Hasebe et al. showed that NHE might not significantly reduce IR; however, it significantly reduced the time lost to sports due to injury (p=001).\n\nAccording to literature, HI is the most frequent MI and injury in soccer. In 2001, Mjolsnes introduced the NHE, presented as an exercise that can easily be performed on pitch without special equipment, and in a randomized controlled study, the NHE group presented a 57% lower HSI (Hamstring Injury).29\n\nIn addition, in a recent systematic review and meta-analysis of six studies, BIZ et al. concluded that both NHE and FIFA11+ are efficient for HSI.21\n\nIn the study by Azuma and Someya,14 the beneficial effect of stretching intervention was particularly strong for severe injuries that require a long time to return to play. Stretching is regularly included in warm-up exercises. However, some authors have reported the opposite finding for stretching.30,31 The effects of stretching on MI depend on the type of sport. In sports with a high stretch-shortening cycle, such as football, when a compliant muscle-tendon unit is contracted, most of the energy is absorbed by the tendon and then MI is reduced. However, in other types of sports with low or no stretch-shortening cycles, such as cycling, most athletes are used to stiff tendons, and stretching will ll not directly cause MI. In fact, having a more compliant tendon makes them less adapted to their sports activities, so they develop more injuries.30,32\n\nTherefore, we can conclude that stretching is recommended for MI in soccer.\n\nWe did not include studies that used a specific program for adductors, which are frequently injured in football, because they did not meet our inclusion criteria. Most of the studies used the Copenhagen adduction exercise (CA).33–35 It is a partner exercise. The player lies on the side with one forearm supporting him on the floor and the other arm placed along the body. The upper leg is held at approximately the hip height of the partner, who holds the leg using one arm and supports the knee with the other. The player then raises his or her body from the field, and adducts the lower leg so that the feet touch each other, and the body is in a straight line. The body was then lowered halfway to the ground while the foot of the lower leg was lowered so that it touched the floor without being used for support. This exercise was performed on both sides. It is an eccentric hip adductor exercise that prevents adductor injury. Haroy et al. showed that adding this type of exercise to FIFA 11+ may compensate for the lack of eccentric hip strengthening exercise.35 In addition, Alonso et al.34 showed that CA increased muscle thickness, which may reduce adductor injury.34\n\nAs there are few players to conclude the effectiveness of this program, more studies with more players should be conducted for specific adductor prevention programs.\n\nThe most relevant point of our study was the diversity of programs used to prevent MI in soccer. Global programs, such as FIFA 11+ and BEP plyometric, provide players with hole strengthening and neuromuscular control. For these global programs, we may add muscle-specific exercises such as NHE and CA. Using structured devices is efficient in reducing IR, especially injury severity.\n\nFinally, it goes without saying that prior to the exercise prescription, each player should have a specific evaluation and obtain an appropriately tailored program. In addition, it is important to target other risk factors, such as ethnicity and nutrition, which may explain the heterogeneity of the study findings.36\n\n\nConclusion\n\nMI is the most frequent injury in football. It is responsible for long-term loss to play and the end of one’s career when it is severe. Coaches, doctors, and sports scientists have always tried to produce effective programs for MI and injury prevention in general.\n\nIn our study, we aimed to identify programs with proven efficacy in MI injury prevention. We reviewed the literature on PUBMED, COCHRANE, and GOOGLE SCHOLAR databases using PRISMA. We only included randomized controlled studies that were interested in determining the efficacy of an exercise or a complete program on reducing MI in professional and semi-professional soccer players, with more than 30 effective participants in each group.\n\nFinally, we included seven articles published in journals of good quality according to the PEDRO scale in the last 10 years. The total number of participants was 3815 male players.\n\nThe program included FIFA 11+, BEP, stretching, and NHE. There was no significant difference in IR, but there was a significant difference in severe injury and, consequently, less time loss due to injury in the IG.\n\nFinally, we can conclude that for MI prevention in football, we may use a completely structured program such as FIFA11+, for which we can add muscle-specific exercises such as NHE and CA exercises.", "appendix": "Data availability statement\n\nAll data underlying the results are available as part of the article and no additional source data are required.\n\nRepository name: Figshare PRISMA abstracts/PRISMA/PRISMA-Scr checklist and flowchart for: only severe injuries are effectively reduced by muscles’ injury prevention protocols in football players: A systematic review.\n\nTitle: Efficacy of programs of muscle injuries prevention in Soccer\n\nhttps://figshare.com/s/b5b3d426ea978cde9797 License CCO\n\nRepository name: Figshare\n\nTitle: Efficacy of programs of muscle injuries prevention in Soccer\n\nFile: citation-export-muscle-injury.xls\n\nhttps://figshare.com/s/b5b3d426ea978cde9797 License CCO\n\n\nReferences\n\nEkstrand J, Hägglund M, Waldén M: Epidemiology of muscle injuries in professional football (soccer). 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Publisher Full Text" }	[ { "id": "318645", "date": "13 Sep 2024", "name": "Ismail Dergaa", "expertise": [ "Reviewer Expertise Sports Medicine and Exercise sciences" ], "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 titled \"Only Severe Injuries Are Effectively Reduced by Muscle Injury Prevention Protocols in Football Players: A Systematic Review\" presents an excellent systematic review for potential publication in F1000. This review offers valuable insights into the effectiveness of structured programs like FIFA 11+ for muscle injury prevention in football players. The methodological approach, adherence to PRISMA guidelines, and focus on high-evidence studies are commendable. However, a few minor revisions are necessary before the manuscript can be considered for indexing.\nCommentary 1: The introduction effectively sets the stage for the review, but it could benefit from a more in-depth exploration of the physiological mechanisms underlying muscle injuries in football. This additional context would provide a stronger rationale for the necessity of specific prevention programs. Also the introduction should be splitted to 3 main parts: (what is known, what is unknown the literature gaps, and finally the aim of the study which should be identical to the aim presented in the abstract).\nCommentary 2: The discussion addresses program effectiveness but could further explore reasons for variability in outcomes. Insights into demographic factors or differences in program implementation could provide valuable context for interpreting the results.\nCommentary 3: A significant limitation that should be addressed is the gap between the publication date of this review (2024) and the cutoff date for included articles (2022). This limitation should be explicitly stated and explained in the manuscript. Consider discussing potential implications of this time gap and how it might affect the review's findings and recommendations.\nCommentary 4: Throughout the manuscript, there is a tendency to use definitive language, particularly the term \"conclude.\" It would be more appropriate to use more measured terms such as \"suggest,\" \"indicate,\" or \"show associations\" when presenting findings. This adjustment would better reflect the nature of the evidence and the inherent limitations of systematic reviews.\nCommentary 5: Table 1, the Impact factor of some journal is missing, why? please add it if no impact factor is associated to those journals please mention it, and please at the bottom of the table mention that the Impact factor mentioned is for which year.\n\nCommentry 6: for all tables, add tables abbreviations.\n\nCommentary 7: The conclusion, while concise, could benefit from emphasizing practical implications for coaches and practitioners. Consider adding specific recommendations for integrating these findings into training routines. Same goes for the conclusion of the abstract\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? 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.) Yes", "responses": [ { "c_id": "12892", "date": "10 Dec 2024", "name": "Rihab MONCER", "role": "Author Response", "response": "Response to Reviewer’s Comments: We would like to express our sincere gratitude to the reviewer for their positive and constructive feedback on our manuscript. We appreciate the time and effort taken to evaluate our work and provide thoughtful suggestions. Below, we provide a detailed response to each of the comments raised, along with the revisions made or planned in the manuscript. Commentary 1: Reviewer’s Comment: The introduction could benefit from a more in-depth exploration of the physiological mechanisms underlying muscle injuries in football. Additionally, the introduction should be split into three main parts: (1) what is known, (2) what is unknown (the literature gaps), and (3) the aim of the study, which should align with the aim presented in the abstract. Response: We agree with the reviewer’s suggestion to enhance the introduction by providing more context on the physiological mechanisms of muscle injuries in football. We have adjusted the introduction as asked Commentary 2: Reviewer’s Comment: The discussion addresses program effectiveness but could further explore reasons for variability in outcomes, such as demographic factors or differences in program implementation. Response: We appreciate the reviewer’s insight. In response, we have expanded the discussion to address potential reasons for variability in the outcomes of muscle injury prevention programs. Commentary 3: Reviewer’s Comment: A significant limitation is the gap between the publication date of this review (2024) and the cutoff date for included articles (2022). This limitation should be explicitly stated and explained in the manuscript, with a discussion of its potential implications. Response: We agree with the reviewer that this time gap is an important limitation. We have now explicitly stated this limitation in the manuscript and discussed the potential implications of including studies published before 2022. We explain that newer studies may provide additional insights into the effectiveness of muscle injury prevention protocols, which could affect the overall conclusions of the review. Commentary 4: Reviewer’s Comment: Throughout the manuscript, the use of definitive language, particularly the term \"conclude,\" may overstate the certainty of the findings. It would be more appropriate to use measured terms like \"suggest,\" \"indicate,\" or \"show associations.\" Response: We appreciate the reviewer’s valuable feedback on the language used in the manuscript. We have carefully reviewed the manuscript and replaced terms such as “conclude”. Commentary 5: Reviewer’s Comment: The impact factor of some journals is missing in Table 1. Please add it, or if no impact factor is associated with those journals, mention that explicitly. Additionally, please note at the bottom of the table the year the impact factor refers to. Response: We have updated Table 1 to include the impact factors for all journals listed. Commentary 6: Reviewer’s Comment: For all tables, add table abbreviations. Response: We have added abbreviations to all tables, as suggested, to improve clarity and ensure the tables are easy to interpret. A key for abbreviations has been included at the bottom of each table. Commentary 7: Reviewer’s Comment: The conclusion could benefit from emphasizing practical implications for coaches and practitioners, including specific recommendations for integrating the findings into training routines. The same applies to the conclusion in the abstract. Response: We have revised the conclusion to emphasize the practical implications for coaches and practitioners. Specific recommendations for integrating muscle injury prevention protocols into training routines have been added. We have also updated the conclusion in the abstract to reflect these practical recommendations. Once again, we thank the reviewer for their insightful comments, which have greatly contributed to improving the quality of our manuscript. We believe the revisions we have made address all the points raised and enhance the clarity and relevance of our review. We look forward to your further feedback." } ] }, { "id": "339867", "date": "18 Nov 2024", "name": "Dhahbi Wissem", "expertise": [ "Reviewer Expertise Biomechanics", "Sports Medicine", "Rehabilitation and Sports injuries", "Fitness and Exercise physiology" ], "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: This systematic review examines the effectiveness of muscle injury prevention protocols in professional football players. Overall, the manuscript addresses an important topic and follows proper systematic review methodology. However, there are several major and minor issues that need to be addressed to improve the quality and clarity of the work.\nMajor Weaknesses: 1. The literature search strategy appears limited, using only 3 databases and potentially missing relevant articles from other sources like SPORTDiscus or conference proceedings. 2. Risk of bias assessment using only the PEDro scale provides limited evaluation of systematic review-specific bias domains recommended by Cochrane. 3. The statistical analysis and synthesis of results lacks formal meta-analysis techniques or quantitative pooling of effect sizes across studies. 4. Discussion section needs stronger critical analysis of the findings and clearer implications for clinical practice.\nMinor Weaknesses: 1. Abstract could be more structured and detailed regarding specific outcomes. 2. Some inconsistencies in reporting numerical results across tables and text. 3. Reference citations need formatting updates in several places. 4. Tables would benefit from additional explanatory footnotes.\nSpecific Comments by Section: Abstract: Page 1: - Background section should specify the exact prevalence rates of muscle injuries - Methods section needs to state the specific databases searched with dates Line 12: Define \"high level of evidence\" explicitly Line 20: Report specific effect sizes and confidence intervals where available Introduction: Page 4: - Lines 2-5: Citation needed for injury incidence statistics - Lines 15-20: The rationale for focusing only on professional players needs stronger justification - Organization would benefit from clearer research question and objectives statements\nMethods: Page 5-6: - PRISMA checklist items should be explicitly addressed - Search strategy details incomplete - need full search strings for each database - Inclusion/exclusion criteria need more precise operational definitions - Statistical analysis plan lacks detail on handling heterogeneity\nResults: Pages 7-11: - PRISMA flow diagram missing some standard elements - Table 1 needs column explaining study design details - Effect sizes and precision estimates inconsistently reported across studies - Forest plots would help visualize key outcomes\nDiscussion: Pages 11-14: - Limited critical analysis of potential sources of bias - Inadequate discussion of heterogeneity across studies - Clinical implications need more specific recommendations - Future research directions should be expanded Insert the bellow reference: Iuliano, Enzo, et al. \"Predictive ability of body mass parameter to estimate 4-6 repetition maximum of upper and lower limb muscles in soccer players.\" Age (years) 24 (2021): 4-66.[Ref 1] Conclusion: Page 16: Conclusions should be more measured based on limitations identified\nTables/Figures: - Table 1: Add study design details and risk of bias ratings - Table 2: Include confidence intervals for all effect estimates - Table 3: Clarify units of measurement - Table 4: Add statistical significance indicators\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? Partly\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\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.) Partly", "responses": [ { "c_id": "12893", "date": "10 Dec 2024", "name": "Rihab MONCER", "role": "Author Response", "response": "We would like to express our sincere gratitude to the reviewer for their thoughtful and constructive feedback on our systematic review. We appreciate the time and effort taken to assess our manuscript. Below, we provide a detailed response to each of the comments raised, outlining the revisions we have made or plan to make in response. 1. Comment 1: The literature search appears limited, using only three databases and potentially missing relevant articles from other sources like SPORTDiscus or conference proceedings. Response: We acknowledge the reviewer’s concern regarding the scope of the literature search. Since the study is finished we have added as limitations the need to consider other databases. Comment 2: Reviewer’s Comment: The use of only the PEDro scale provides a limited evaluation of systematic review-specific bias domains. Response: We used this scale for its specific items for physical interventions. We added in the discussion its limitations Discussion Section: Reviewer’s Comment: The Discussion section needs stronger critical analysis and clearer implications for clinical practice. Response: We have revised the Discussion section to provide a more in-depth critical analysis of the findings, including a discussion of potential biases and study heterogeneity. Additionally, we have enhanced the clinical implications section by offering more specific recommendations for practice. Minor Weaknesses: Abstract: Reviewer’s Comment: The Abstract could be more structured and detailed regarding specific outcomes. Response: We have restructured the Abstract to include more specific details on the outcomes of the systematic review, such as effect sizes and confidence intervals, where available. Inconsistencies in Reporting Numerical Results: Reviewer’s Comment: Some inconsistencies in reporting numerical results across tables and text. Response: We have thoroughly reviewed and corrected the numerical results across both the text and tables to ensure consistency. Reference Citations: Reviewer’s Comment: Reference citations need formatting updates. Response: We have addressed all reference formatting issues and ensured consistency throughout the manuscript in accordance with the required citation style. Tables: Reviewer’s Comment: Tables would benefit from additional explanatory footnotes. Response: We have added explanatory footnotes to all tables to improve clarity and assist with the interpretation of the data presented. Once again, we would like to express our appreciation for the reviewer’s insightful comments, which have significantly contributed to improving the quality of our manuscript." } ] } ]	1	https://f1000research.com/articles/13-695
https://f1000research.com/articles/10-645/v1	23 Jul 21	{ "type": "Research Article", "title": "The discovery of human Plasmodium among domestic animals in West Sumba and Fakfak, Indonesia", "authors": [ "Munirah Munirah", "Sitti Wahyuni", "Isra Wahid", "Firdaus Hamid", "Munirah Munirah", "Isra Wahid", "Firdaus Hamid" ], "abstract": "Background: In Indonesia, malaria incidence is at a high rate despite maximum preventive efforts. Therefore, this study aims to determine the possibility of a Plasmodium reservoir among domestic animals in malaria-endemic areas. Methods: Animal blood was collected using EDTA tubes, then smeared and stained with Giemsa for Plasmodium microscopic identification. About 10 µl of blood was dropped on to a filter paper to capture Plasmodium DNA. Nested PCR was used for parasite molecular detection, while Plasmodium species were identified using the sequenced DNA. Results: A total of 208 and 62 animal blood samples were collected from Gaura village, West Sumba and Fakfak village, West Papua, Indonesia respectively. In total, 32 samples from Gaura contained P. falciparum or P. vivax, while the Plasmodium percentage in buffalo, horse, goat, and dogs were 20.7%, 14.3%, 5.8%, 16.7%, respectively. P. knowlesi was not found in any of the samples, and no other species were detected in 18 pig blood samples. Conclusion: Human Plasmodium existence among domestic animals in Indonesia partly explains the high prevalence and persistence of malaria in some endemic areas due to a reservoir host presence. Therefore, future studies need to ascertain the cause.", "keywords": [ "Plasmodium falciparum", "Plasmodium vivax", "malaria", "animals", "host reservoir", "PCR." ], "content": "Introduction\n\nMalaria is transmitted by the Plasmodium vector Anopheles mosquitoes. Four Plasmodium types, namely P. falciparum, P. vivax, P. ovale, and P. malariae cause pathologic conditions in humans. Recently in Southeast Asia, P. knowlesi infection cases have also been reported.1–3\n\nBefore molecular diagnostics development, only humans were assumed to be the primary host for Plasmodium. However, studies in the last two decades on Plasmodium reported that the parasites originated from animals. Further stating that P. falciparum originated in the gorilla4 and chimpanzee,5,6 P. vivax was from African apes,7 P. malariae was from chimpanzees6 and P. knowlesi was from monkeys,8,9 while P. ovale in humans and chimpanzees are genetically identical.10 The factors hypothesized to explain this situation include primate’s habitat loss and human’s aggressiveness in exploring forest.11 A study from South Kalimantan reported the contribution of forest workers to malaria incidence.12\n\nEast Nusa Tenggara and West Papua are known as malaria-endemic areas in Indonesia as their annual parasite incidence (API) in 2015 was 31.29% and 7.04%, respectively,13 while in 2018, according to the health office in both the districts, the API rate in Fakfak, West Papua and East Nusa Tenggara, West Sumba was 4.85% and 12.9%, respectively.(unpublished data) Due to this situation, we aimed to explore the presence of human Plasmodium among domestic animals that are a potential reservoir host.\n\n\nMethods\n\nThis study was conducted in October 2018 in Gaura village, West Sumba Regency, an area 29.96 km2 in size inhabited by 9,584 people, and Fakfak, West Papua Province, in August 2019 with an area of 11,036 km2 inhabited by 84,692 people (Figure 1). The residents’ main occupation is farming, while livestock such as goats, horses, cows, pigs, and buffalos are commonly found in their enclosures located around the owner’s residence. Furthermore, they also own pets such as dogs and cats.\n\nSampling was carried out by the veterinarian and staff from West Sumba and Fakfak Animal Husbandry Office. The buffaloes, goats, pigs, and horses’ blood samples were collected in 5 ml EDTA tubes from the jugular vein located in the ventrolateral area of the neck using vacutainer needles, size 16–18. Meanwhile, the dog’s blood was drawn from the cephalic antebrachial vein in the leg using a size 21 vacutainer needle. By using a micropipette, approximately 10 ul of EDTA blood was dropped onto a microscope slide, then smeared and stained with Giemsa (MERCK Millipore, Germany) for Plasmodium microscopic identification, while the remaining was dropped onto a filter paper (Whatman CAT No. 1442-090) until it absorbed to about 1.5 cm in diameter. The dry filter paper was put on a sterile plastic clip and stored at room temperature for a maximum of 10 days.\n\nA dried blood spot (DBS) isolation kit for DNA extraction on filter paper (Cat. no. 36000) from Norgen Biotec was used. A 6 x 3 mm piece of blood-stained filter paper was put into a 1.5 ml tube containing 100 μl of digestion buffer B. It was vortexed and incubated at 85°C. Afterwards, 20 μl of proteinase K and 300 μl of lysis buffer B were added to the tube and then vortexed before incubation at 56°C for 10 minutes. About 250 μl of 95% ethanol was added to the tube and then vortexed, while the DNA content was washed by adding 500 μl of WN wash solution and centrifugated for one minute at 8,000 rpm. Washing was carried out again using 500 μl of WN wash solution and centrifugated at 14,000 rpm. For DNA elution, 90 μl of elution buffer B was put into the tube and centrifuged at 8,000 rpm for one minute, and the purified DNA was stored at -20°C.\n\nDNA amplification of nested PCR and qPCR were performed as directed by Tiangen Biotech (Beijing). Plasmodium DNA amplification was carried out using the nested PCR method with a 2× Tag Plus PCR mix enzyme (Tiangen). The final volume of 12.5 μl contained 6.25 μl enzyme, 2.25 μl ddH2O, 1 μl forward primers, 1 μl reverse primers, and 2 μl DNA sample. For sequencing, the PCR mixture’s volume was doubled, with the final volume being 25 μl, while the primer sequences of P. falciparum, P. vivax14 and P. knowlesi15 can be seen in Table 1.\n\nThe nested one DNA amplification temperature was set at 94°C denaturation (one minute), 55°C annealing (one minute) and 72°C extension (one minute) for 35 cycles. For nested two, denaturation was carried out at 94°C (30 seconds) and extension was at 72°C (30 seconds) in 35 cycles. There was a difference in the annealing temperature for each species in nested two, namely 55°C (one minute) for PCR multiplex P. falciparum and P. vivax, but 56°C (one minute) for P. knowlesi. Nested one products were used as templates for nested two and both were run on agarose gel 1.5% and 2%, respectively, while qPCR was analysed using agarose gel 1.5% for electrophoresis. Molecular work was not performed for P. ovale and P. malariae due to difficulties in finding the positive control, and according to the local health office these species have never been reported from Sumba and Fakfak.\n\nConsidering the possibility of contamination, DNA was re-extracted from blood from the same filter paper. PCR was performed using the primers, rPF1 and rPF2, as well as rPV1 and rPV216 to detect P. falciparum and P. vivax, respectively. The same extraction and amplification method were used as described above.\n\nTo determine the Plasmodium species, in the second round of nested PCR, products having positive band targets were sent to the 1st BASE, Axil Scientific Pte Ltd Singapore for sequencing. The DNA sequence result was adjusted using multiple alignments found in the BioEdit 7.0 application17 and then read by the BLAST program from the NCBI website.\n\n\nEthical clearance\n\nThis study was approved for ethical clearance by the ethics committee of the Faculty of Medicine, Hasanuddin University (734/H4.8.4.5.31/PP36-KOMETIK/2018). All efforts were made to ameliorate any suffering of animals. To prevent stress, animals were comforted by their owners while blood samples were taken, and sampling was performed by experienced officers. Second and third blood samples were taken if there was a failure in the first sample and only if the animals were cooperative. About 20% of animals were sampled more than once.\n\n\nResults\n\nA total of 208 and 62 animal blood samples were collected from Gaura and Fakfak villages, respectively. These consisted of 92 buffalos, 21 horses, 121 goats, 18 dogs, and 18 pigs. Using the nested PCR method, 32 of the 270 animals were found to be P. falciparum and P. vivax positive. The percentage of Plasmodium positive animals included 20.7% buffalo, 14.3% horse, 5.8% goat, and 16.7% dog with one buffalo having a mixed infection (P. falciparum and P. vivax). There was no P. knowlesi found in any of the samples and no other Plasmodium was found in 18 pig blood samples. PCR gel products, DNA sequence results, and the sample’s quality can be seen in Figures 2, 3 and 4, respectively.18 Plasmodium distribution in the animals’ blood samples from Gaura and Fakfak are presented in Table 2, and it shows that blood containing Plasmodium was only found in Gaura. The results of the qPCR using rPF1–rPF2 and rPV1–rPV2 primers were similar to the nested PCR\n\nGel view of PCR product from Plasmodium vivax and Plasmodium falciparum in domestic animals in Gaura, West Sumba (LD = DNA ladder, PS = positive samples, CN = control negative, CP = control positive) by nested PCR (multiplex PCR). 120 bp for positive Plasmodium vivax, 205 bp for positive Plasmodium falciparum.\n\nPlasmodium DNA sequence alignments from blood samples taken in Gaura village, West Sumba, Indonesia by ClustalW multiple sequence alignment.\n\nExample of Plasmodium PCR product quality from a blood sample taken in Gaura village, West Sumba, Indonesia.\n\nDistribution of animal blood samples and Plasmodium species found in Gaura village, West Sumba, Indonesia and Fakfak, West Papua, Indonesia.\n\nMicroscopically, trophozoites, schizonts, and gametocyte forms at 100× magnification can be seen in Figure 5. P. falciparum gametocytes found in buffaloes were sausage and crescent-shaped (a, b), while schizonts found in horses were smaller or the same size as the red blood cells (c). The P. vivax gametocyte was larger than the red blood cells found in buffalo (d). P. falciparum gametocyte and trophozoites (ring-shaped) with one or two nuclei was found in goats (e) and P. falciparum trophozoite found in horses had one nucleus (f).\n\nMorphology of Plasmodium in animals from Gaura village, West Sumba, Indonesia. Gametocytes (a,b,d) in buffalo, schizont in horse (c), gametocyte and trophozoite in goat (e) and trophozoite in horse (f) with magnification 1000 ×.\n\n\nDiscussion\n\nThe presence of Plasmodium was suspected in domestic animals because malaria cases in these two villages remained high despite maximum preventive efforts having been applied including insecticide-treated bed nets. About 32 of the 270 blood (11.9%) samples contained human Plasmodium parasites, and this is the first data report and further study is therefore needed.\n\nPrevious studies found Plasmodium relictum in avian species,19 P. cephalophi in ungulates,20 P. traguli in mousedeer,21 P. brucei in gray duiker,22,23 P. bubalis in water buffalo,24 and P. odocoilei in white-tailed deer.25,26 Other parasites found included P. caprae in goats (ruminant),27 P. bergei in Rodentia,28 and P. cynomolgi, P. inui, and P. fragile in primates.29 The five Plasmodium species that infect humans were originally parasites in primates.1,3,6–9 In this study, P. falciparum was found in buffalos, goats, dogs, and horses, while P. vivax was in buffalos, goats, and dogs. Initially, the presence of Plasmodium in these animals’ erythrocytes was not certain. However, the nested PCR showed the same results for all positive samples. The sequencing results of the positive bands in the nested PCR two analysis showed the bands were P. falciparum and P. vivax (Figure 3). This is the first investigation reporting human Plasmodium in domestic animals (ruminant, ungulate, and carnivore).\n\nPlasmodium discovery among domestic animals in malaria-endemic areas raises the following questions. How do P. falciparum and P. vivax live in these animals? Are they intermediate hosts for this parasite? Did these Plasmodium species evolve to live in ruminants, ungulates, and carnivores? As a result of repeated exposure, have these animals become more permissive to Plasmodium, which generally lives in humans? Is this parasite pathogenic in animals? P. knowlesi is a commensal microbe in primates but pathogenic in humans1–3 and its migration from primates to humans is caused by forest loss or human invasion of primate habitat.11 There is a possibility that animal and human proximity aids easier cross parasite transfer between both groups by mosquitoes.\n\nDespite high API in Fakfak and Gaura village, West Sumba, only the animals from West Sumba had human Plasmodium. This difference is possibly due to the distance between the residents’ houses and animal enclosures as the enclosures are located approximately 50–500 meters from the main houses in Fakfak. Meanwhile, in Gaura, residents live in stilt houses where the ground floor functions as an animal shelter, allowing microbial transfer between humans and animals by mosquitoes. In Fakfak, the sampling locations were not easily accessible, and the steep geographical conditions made it difficult to collect many samples compared to Gaura.\n\nAlthough Plasmodium can be detected microscopically due to erythrocyte size, which is smaller in animals than humans, molecular methods become significant in detecting Plasmodium presence. The nested PCR was used to detect Plasmodium because its sensitivity was equally as high as Real-Time PCR and the cost was relatively lower.30,31 The microscopic method of double fluorescent dye utilization with Giemsa stain is recommended for further studies.32\n\n\nConclusion\n\nIn this study we found human Plasmodium in domestic animals. It is still not clear whether the animal had malaria, but this finding may be used as a reference for conducting malaria surveys in domestic animals in endemic areas. Human Plasmodium was only found in Gaura where the location of the animal enclosures is integrated with the residents’ houses. Local communities need to be educated about the possibility of malaria transmission due to the integration of animal enclosures and peoples’ homes. The discovery of human Plasmodium in domestic animals in this study may partly explain the persistence of the high prevalence of malaria in some endemic areas.", "appendix": "Acknowledgements\n\nThe author acknowledges the assistance of the West Sumba District and Fakfak Animal Husbandry Service, who helped carry out animal blood collection. We thank the people of Gaura village and Fakfak, who allowed us to take blood samples from their animals. We also thank Syahruni and Handayani Halik from Hasanuddin University Medical Research Center (HUM-RC) for their help with molecular work.\n\n\nData availability\n\nFigshare: Underlying data for ‘The discovery of human Plasmodium among domestic animals in West Sumba and Fakfak, Indonesia’, https://doi.org/10.6084/m9.figshare.14703012.v3.18\n\nThis project contains the following underlying data:\n\n• Gel photo: Result of rPF1–RPF2 primers\n\n• Gel photo: Result of RPV1–rPV2 primers\n\n• Gel photo: Nested PCR P. falciparum and P. vivax\n\nFigshare: ARRIVE checklist for ‘The discovery of human Plasmodium among domestic animals in West Sumba and Fakfak, Indonesia’, https://doi.org/10.6084/m9.figshare.14703012.v3.18\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nSingh B, Daneshvar C: Human infections and detection of Plasmodium knowlesi. Clin Microbiol Rev. 2013 Apr; 26(2): 165–84. 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PubMed Abstract \| Publisher Full Text\n\nKaewthamasorn M, Takeda M, Saiwichai T, et al.: Genetic homogeneity of goat malaria parasites in Asia and Africa suggests their expansion with domestic goat host. Sci Rep. 2018; 8(5827): 1–7. Publisher Full Text\n\nVincke I, Lips M: Un nouveau Plasmodium d’un rongeur sauvage du Congo, Plasmodium berghei n.sp. Ann Soc Belge Méd Trop. 1948; 28: 97–104.\n\nDixit J, Zachariah A, Sajesh P, et al.: Reinvestigating the status of malaria parasite (Plasmodium sp.) in Indian non-human primates. PLoS Negl Trop Dis. 2018; 12: e0006801. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGreen M, Sambrook J: Nested Polymerase Chain Reaction (PCR). Cold Spring Harb Protoc. 2019; 2. PubMed Abstract \| Publisher Full Text\n\nPerandin F, Manca N, Calderaro A, et al.: Development of a Real-Time PCR Assay for detection of Plasmodium for routine clinical diagnosis. J Clin Microbiol. 2004; 42(3): 1214–9. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGuy R, Liu P, Pennefather P, et al.: The use of fluorescence enhancement to improve the microscopic diagnosis of falciparum malaria. Malari J. 2007; 6: 89. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "101607", "date": "06 Jan 2022", "name": "Alfred Amambua-Ngwa", "expertise": [ "Reviewer Expertise Malaria", "genomics", "genetics", "immunology", "epidemiology", "invasion", "immunology" ], "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 present data showing microscopically identified Plasmodium species in non-primate domestic animals in a region of Indonesia. They followed on to confirm infections with Plasmodium species with molecular methods targeting human malaria parasites P. falciparum, P. vivax, and P. knowlesi. They sequenced the amplified fragment, indicating that they indeed detected the genus. If this claim is validated, it will have significant consequences on malaria interventions and the goal of its elimination across the globe. The authors need to pay attention to the following to validate their claim:\nSeveral Plasmodium species infect a wide range of animal species and there are other protozoa with similar cellular features that can also be microscopically misidentified for human Plasmodium. Thus, the identification of human Plasmodium will require species-specific distinguishing features by comparing the infected erythrocyte of the Plasmodium-infected domestic animals to those of other protozoan parasites including non-human Plasmodium.\n\nA single sequence of each of the PCR products is shown per species and these sequences are similar to those of Plasmodium falciparum small rRNA. The alignment could include all samples sequenced and also how this was used to determine other species beyond P. falciparum. For speciation to be better validated, the authors could amplify and sequence cytochrome B, which is more divergent for Plasmodium.\n\nMore detail on the control protocols to limit contamination at sampling, extraction, and amplification of parasite DNA from the samples will help.\n\nGiven that humans and the family of domestic animals listed are largely divergent on RBC surface receptors for parasites, how these putative human-specific parasites invade the non-human hosts and develop will be of interest. The authors could therefore further validate their claim by sequencing an invasion ligand, such as the DBL proteins: EBA 175 for P. falciparum and DBA for P. vivax.\n\nWhile the proximity between humans and animals is thought to be an explanation for the potentially zoonotic transmission, this will be surprising as the sharing of houses between humans and their domestic animals is a common occurrence in malaria-endemic tropical regions, where such a zoonosis has not been previously described. A census of the vector species between the two sites could enlighten on whether this potential zoonosis is driven by the vector.\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? No\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "9306", "date": "16 Feb 2023", "name": "Sitti Wahyuni", "role": "Author Response", "response": "Thank you for the suggestions and corrections. The corrections and suggestions were compiled and a revised version of the document is written. We believe that the method used has followed the malaria detection standards. The used methods have provided sufficient positive evidence so far. As for suggestions for validating our findings in several ways such as amplifying and sequencing cytochrome B, and sequencing an invasion ligand (such as the DBL proteins: EBA 175 for P. falciparum and DBA for P. vivax), we have included all in the discussion for further research because the method requires research funds which, for us, is quite a lot. Therefore, we are open to collaborate with other researchers from anywhere to work with us. We really appreciate the time spent by the reviewer to provide helpful comments and corrections." } ] }, { "id": "156443", "date": "22 Dec 2022", "name": "Jaishree Raman", "expertise": [ "Reviewer Expertise Antimalarial drug and diagnostic resistance", "molecular biology", "molecular epidemiology", "malaria elimination and control" ], "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 manuscript describes the interesting finding of human malaria parasites in domestic pets and farm animals in Indonesia. While this finding has potential implications for countries trying to stem residual transmission with the aim of being certified malaria free, this article needs some revision to ensure clarity and a logical flow of ideas. More information on the malaria burden and interventions in place in the study area need to be provided. Some possible reasons for the study findings need to be provided. The ideas for future studies should be presented as statements rather than questions.\nBelow are some more specific comments:\nIntroduction:\nIt is far too short. It needs to be expanded to explain the issue of residual malaria transmission in the study area and why it is concern. What interventions are in place to stop transmission and why you think they are failing to stem residual transmission.\n\nThere are four species of Plasmodia, not types that primarily affect humans\n\nIs the statement on forest workers included because the study area has large numbers of forest workers? If so please provide this information.\n\nFrom the data presented it looks like malaria transmission has decreased in both study areas so it is not clear to me what situation needs to be investigated. More clarity needs to be provided here.\n\nA clear reason for the study needs to be provided as well as possible uses of the research findings.\nMethods:\nI am correct in assuming that the animal enclosures are in close proximity to the farmers homesteads? Please give an average distance between enclosures and homes – as mosquitoes will only travel as far as they need to, to get their next blood meal.\n\nThe remaining is either blotted or spotted onto the filter paper. I do not understand what “the dry filter paper was put on a sterile paper clip” means.\n\nWhere the houses geo-located so you could map the infected animals?\n\nHow long was the incubation at 85 degrees?\n\nIt is DNA amplification by PCR not of PCR\n\nFor nest two is there no annealing stage?\n\nPlease explain why nest one products were run on a gel.\n\nWhere the qPCR products really analysed using gel electrophoresis? The benefit of qPCR is real time data generation through the detection of fluorescence in the amplification stage. The analysis of qPCR products is generally done by reading fluorescence outputs.\n\nPlease provide a bit more information on the rationale for the re-extraction of the sample. In addition please state if the additional blood spot was from the same blood spot or different blood spot?\nResults\nIt is not clear whether the pigs were not infected will any form of malaria parasites or only not infected with human malaria parasites. Please clarify.\nDiscussion\nPlease expand on the questions presented in the discussion in terms on possible results and their implication for malaria elimination in Indonesia and globally.\n\nPlease clarify if only infected were found in West Sumba region or only one study area within West Sumba. The reason provided for only one area having infected animals is unclear to me. Please rephrase for more clarity.\n\nPlease some guidance for future studies.\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": [ { "c_id": "9307", "date": "20 Mar 2023", "name": "Sitti Wahyuni", "role": "Author Response", "response": "This manuscript describes the interesting finding of human malaria parasites in domestic pets and farm animals in Indonesia. While this finding has potential implications for countries trying to stem residual transmission with the aim of being certified malaria free, this article needs some revision to ensure clarity and a logical flow of ideas. More information on the malaria burden and interventions in place in the study area need to be provided. Some possible reasons for the study findings need to be provided. The ideas for future studies should be presented as statements rather than questions. Below are some more specific comments: Introduction: It is far too short. It needs to be expanded to explain the issue of residual malaria transmission in the study area and why it is concern. What interventions are in place to stop transmission and why you think they are failing to stem residual transmission. Author’s response: Thank you, we have made a revision. There are four species of Plasmodia, not types that primarily affect humans Author’s response: Thank you for your input, we already revised. Is the statement on forest workers included because the study area has large numbers of forest workers? If so please provide this information. Author’s response: Thank you, we already revised. From the data presented it looks like malaria transmission has decreased in both study areas so it is not clear to me what situation needs to be investigated. More clarity needs to be provided here. Author’s response: Thank you, we already revised. A clear reason for the study needs to be provided as well as possible uses of the research findings. Author’s response: Thank you for your comment, we have revised accordingly. Methods: I am correct in assuming that the animal enclosures are in close proximity to the farmers homesteads? Please give an average distance between enclosures and homes – as mosquitoes will only travel as far as they need to, to get their next blood meal. Author’s response: Thank you, we have already given the average distance in the method. The remaining is either blotted or spotted onto the filter paper. I do not understand what “the dry filter paper was put on a sterile paper clip” means. Author’s response: Thank you, more clarity has been made. Where the houses geo-located so you could map the infected animals? Author’s response: We apologize, because we don’t have the data about geo-location. But, in future research, we will apply this method. Thank you for your input. How long was the incubation at 85 degrees? Author’s response: Thank you for your question, the incubation at 85 degrees was 10 minutes. It is DNA amplification by PCR not of PCR Author’s response: Thank you for your comment, we have revised accordingly For nest two is there no annealing stage? Author’s response: Thank you for your question, the annealing nested two was 55°C in one minute. Please explain why nest one products were run on a gel. Author’s response: Thank you. Actually, the nested one does not need to run on a gel, but it was done just to notice if there were any bands in the nested one. However, it is not the standard for determining whether positive at that stage, because we used nested two for determining Plasmodium positive. Where the qPCR products really analysed using gel electrophoresis? The benefit of qPCR is real time data generation through the detection of fluorescence in the amplification stage. The analysis of qPCR products is generally done by reading fluorescence outputs. Author’s response: Thank you for your question. Both products of nested and qPCR were analysed using gel electrophoresis and then run on agarose gel (nested one in 1.5%, nested two in 2% agarose gel and qPCR in 1.5% agarose gel) in order to identify fluorescence band wave in a gel documentation system. Please provide a bit more information on the rationale for the re-extraction of the sample. In addition please state if the additional blood spot was from the same blood spot or different blood spot? Author’s response: Thank you, we already revised. Results It is not clear whether the pigs were not infected will any form of malaria parasites or only not infected with human malaria parasites. Please clarify. Author’s response: Thank you, we already revised. Discussion Please expand on the questions presented in the discussion in terms on possible results and their implication for malaria elimination in Indonesia and globally. Author’s response: Thank you, we already revised. Please clarify if only infected were found in West Sumba region or only one study area within West Sumba. The reason provided for only one area having infected animals is unclear to me. Please rephrase for more clarity. Author’s response: Thank you, we already revised. Please some guidance for future studies. Author’s response: Thank you, we already revised." } ] } ]	1	https://f1000research.com/articles/10-645
https://f1000research.com/articles/12-380/v1	11 Apr 23	{ "type": "Method Article", "title": "Oxygen as obturation biomaterial in endodontic treatment: development of novel membranous dental restoration system", "authors": [ "Didi Wahyudi", "Citra Kusumasari", "Citra Kusumasari" ], "abstract": "Complexities in obturation and difficulties in disinfection are the major problems that make endodontic treatment very time-consuming. A new perspective is needed to reduce the working time as well as to answer these two problems. Until now, none of the established obturation techniques for root canal filling can guarantee a perfect seal. Solid substances cannot be manipulated easily to fill the tiny branches of the root canal system hermetically. At the same time, anaerobes and facultative anaerobes, especially Enterococcus faecalis, are very dominant in endodontic infections discussion. As shown in many studies, it is extremely difficult to perfectly disinfect Enterococcus faecalis even by using some irrigating solutions with strong antibacterial properties. Under anaerobic conditions, the invasion efficiency of facultative anaerobes is increased. In case irrigation and disinfection cannot totally eliminate anaerobes and facultative anaerobes, a new strategy is also needed to deal with the bacteria that still survive in the root canal. Oxygen can fill the root canal system with ease, eliminate anaerobes, and prevent facultative anaerobes from being pathogenic. Therefore, using oxygen as obturation biomaterial after proper cleaning and shaping procedures is expected to solve the two major endodontic problems. The aim of this article was to discuss a new possible concept of dental restoration system that uses an oxygen-permeable membrane to decrease the time required in endodontic treatment. The membrane is placed at the orifice of a duct created to connect the entire root canal system with free air outside the restoration. The function of the membrane is more or less similar to the mask used during the COVID-19 pandemic, as it enables the oxygen to circulate while preventing any fluid, debris, and microorganisms from passing. We hypothesize that the oxygen circulating in the root canal system will also act as an antimicrobial agent that is constantly renewed.", "keywords": [ "dental restoration system", "Enterococcus faecalis", "obturation biomaterials", "oxygen permeable membrane", "endodontics treatments", "trepanation" ], "content": "Introduction\n\nAs of March 2023, some variants of concern of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) or coronavirus disease 2019 (COVID-19) hav been recorded.1 Dentists are very vulnerable to being infected by COVID-19 because their face when treating patients is relatively close to the patient’s mouth. Saliva can be a place for various viruses to reside, including coronavirus,2 and is something that dentists cannot wholly avoid when performing any procedure in the oral cavity. Therefore, dentists and patients must be careful when providing or having dental care.\n\nVarious ways have been implemented to deal with the spread of COVID-19 in dental practice, from shutting down dental clinics to limiting certain types of treatment that can only be administered using strict protocols.3 The reason for all those decisions is to avoid or shorten interactions with patients who may possibly transmit the coronavirus. On the other hand, patients try not to visit places they consider potentially contaminated with coronavirus, such as hospitals and dental clinics. Hitherto, the COVID-19 pandemic has made a big impact to the way dentists work.\n\nEndodontic treatment tends to require multiple visits, which can be a disadvantage for patients. However, root canal treatment may still be necessary for those experiencing severe dental pain. Most emergency cases in dentistry require endodontic treatment.4 In a study conducted in India during the COVID-19 pandemic, there were three main reasons for emergency visits to dental clinics: pulpal problems 46.0%, abscess 16.6%, periapical lesions 15%, and most of the dental emergencies were endodontic related.5\n\n\nCurrent endodontic treatment\n\nThere are ten principles that we can consider in endodontic treatment. The first and second principles are aseptic technique and the instruments that should be confined to the root canal. Third, root canal preparation should be performed by using proper instruments. Fourth, the root canal should be expanded, if possible, to make cleaning easier. Fifth, the root canal should be thoroughly irrigated with antiseptic solutions. Sixth, any use of irrigating solutions must be safe for periapical tissues. Seventh, if there is a sinus tract, it should be cleared up following root canal therapy and does not require surgery. An incision of the soft tissue can be performed for the acute periapical abscess to provide drainage. Eighth, the root canal should be hermetically obturated to give a good seal. Ninth, before obturation, a negative culture should be taken. Tenth, everything that is used in obturation should be biocompatible. These all are based on what was concluded in the International Conference on Endodontics in 1958.6–8\n\nThere are three factors in endodontic treatment that must be considered for successful root canal therapy, namely, cleaning and shaping, disinfection, and obturation.9 Periradicular pathosis is primarily caused by the growth of pathogens in the root canal system. Unsuccessful root canal therapy is not directly caused by the errors in performing endodontic treatment.10\n\nCleaning and shaping are separate and distinct concepts but performed concurrently. Some most significant factors influencing the result of cleaning process are the anatomy and morphology of the tooth, as well as the instruments and irrigants available during the process. The main objective of shaping is to maintain or develop a continuously tapering funnel from orifice to apex. Irritants should be reduced as much as possible, if not entirely. An adequately prepared root canal should feel smooth in all dimensions when the tip of a small file is pushed against the canal wall. After cleaning and shaping, sufficient space should be available for placing obturation materials. Irrigation with 17% ethylenediaminetetraacetic (EDTA) for 1 minute is recommended to remove the smear layer accumulated on the radicular canal wall.11 The success of root canal therapy is majorly achieved by proper cleaning and shaping. Endodontically treated teeth fail not because of poor obturation but due to poor cleaning and shaping.9,12\n\nMost clinicians choose sodium hypochlorite (NaOCl) as it demonstrates a proteolytic effect and can be used for disinfection. To get its bactericidal effect, NaOCl is highly dependent on the length of time it is retained in the canal and the use of copious volumes of solution.13 Irrigating solution containing Chlorhexidine may also be used as an alternative because of its superior intracanal antimicrobial effect.9 However, both NaOCl and Chlorhexidine can not perfectly eliminate E. faecalis,14 especially in a place where irrigating solutions cannot easily flush.\n\nThe process of root canal obturation is a time-consuming and costly manipulation. In this process, an apical seal is crucial.15 In a comparative study of 3 obturation techniques, the thermafil obturation technique showed better results in terms of voids and gaps found between gutta-percha and canal walls at the apical third of root canals compared to warm vertical condensation. The cold lateral obturation technique showed more voids and gaps than the other two.16 However, none of the established techniques for root canal filling we have known today can guarantee a perfect seal.17\n\nIn single-visit endodontic treatment, the whole process could be done in one visit. In multiple-visit endodontic treatment, patients will have more than one appointment in which inter-appointment intracanal medicament is used to improve disinfection before obturation. Calcium hydroxide has been the gold standard for intracanal medicament to fight against root canal pathogens. Combining calcium hydroxide with 2% Chlorhexidine gives better results in terms of percentage resolution of periapical radiolucency.18 However, the penetration depth of both conventional and nanoparticle calcium hydroxide to dentinal tubules is low in the apical zone.19\n\n\nTrepanation: bringing back oxygen circulation to a nonvital tooth\n\nThere is still a question of whether single-visit root canal treatment is adequate for eliminating root canal bacteria that can cause future reinfection.20 However, multiple-visit root canal treatment is not preferable for patients, not only during the COVID-19 pandemic. Bacteria that are difficult to clean in the root canal, particularly in dentinal tubules, can live in an anaerobic environment. The failure of root canal treatment generally involves the growth of anaerobes and facultative anaerobes that become pathogenic and more efficient to infect in the absence of oxygen.21–24\n\nSome strategies to avoid reinfection after root canal treatment are preventing coronal microleakage that inadequate temporary or permanent fillings might cause, and eliminating bacteria that reside in dentinal tubules. Therefore, besides ensuring that no saliva, fluids, microorganisms, and debris can enter the coronal microleakage, we must also consider the antimicrobial agent we use and its delivery system. Disinfection must be able to reach the dentinal tubules where some bacteria can live and survive anaerobically.22 Dental pain can occur in nonvital teeth, including after root canal treatment, and is often a symptom of an abscess. Most dental abscess treatments using antibiotics focus on attacking anaerobes and facultative anaerobes.25 Endodontic infections are polymicrobial, with obligate anaerobic bacteria undeniably dominating the microorganism in primary infections.26 NaOCl is mentioned by numerous researches to be effective against polymicrobial root canal biofilms.27,28 Virgin coconut oil is also believed to have antiprotozoal, antiviral, and antibacterial properties,29,30 but to use it as an irrigating solution still needs further studies. Understanding on how oxygen can make a difference in the root canal (please see Table 1) is also an attempt to harness its use in preventing endodontic infection that is dominated by obligate anaerobes.\n\nBecause the infection of the root canal of nonvital teeth tends to happen in an anaerobic condition, dental practitioners can consider preventing the root canal microenvironment from being oxygen-free. At some point, creating ventilation to avoid the absence of oxygen might look similar to trepanation. The size of the oxygen molecule is smaller than the diameter of dentinal tubules, making it able to fill the entire space available within the tooth. Oxygen can inhibit the growth of anaerobes and prevent facultative anaerobes from becoming more infectious. Using oxygen as an antimicrobial agent that fills the root canal is expected to shorten the working time of a dentist performing root canal treatments. It can replace conventional obturation after proper cleaning and shaping.\n\nOxygen can reach the anaerobic microenvironment in dentinal tubules at the apical third in which irrigating solutions cannot easily flush. This action will disturb the lives of anaerobes. However, facultative anaerobes are still possible to be found in the dentinal tubules and cannot be eliminated by oxygen. Enterococcus faecalis is the most discussed facultative anaerobe in endodontic infection and can colonize dentinal tubules to a depth of >1000 μm.35 Not always pathogenic, Enterococcus faecalis can also be helpful and might be considered a potential probiotic, as shown in a study of E. faecalis in human milk.36 If it is very challenging to eliminate E. faecalis perfectly, even by using any recommended irrigating solutions and special techniques, then perhaps the next strategy is to make them probiotic (mutualism), or at least nonpathogenic and nondestructive (commensalism). Oxygen might play its role in preventing the increase of their invasion efficiency and pathogenicity in the case where cleaning and shaping are not adequate enough to eliminate E. faecalis in the root canal system. Beside its nature that can be a commensal, Enterococcus spp. is also prominent for probiotic candidacy because of its potential antibacterial, antifungal, and antiviral activity. Its ability to produce bacteriocins is essential for these activities.37\n\nTrepanation was originally known as an act of perforating the skull. It might be the oldest surgical procedure, performed by many civilizations in the past, from Greece to China, and used in western medicine.38 In modern medicine, surgeons still use this method through minimally invasive trepanation and drainage. It is also considered highly effective for treating purulent meningitis,39 and its use in dentistry is no exception. Dental trepanation is a simple procedure of perforating the pulp chamber and keeping it open without any medicament inserted into the pulp. It is not easy to find academic literature that discusses this procedure, but it cannot be the reason to pretend as if it has never existed. Dental trepanation in a nonvital tooth is not only to let the pus from dental or periapical abscess out but also to prevent abscess formation. Oxygen can fill inside the tooth and eliminate the bacteria that can cause the abscess. A case report of a 12-year-old child in Indonesia showed that a dental abscess on a permanent maxillary lateral incisor improved after being given antibiotics and trepanation by making a hole that penetrates the pulp chamber. The patient did not come as instructed on the following third to seventh day. However, after 45 days, the tooth’s general conditions were good and there were no complaints. Neither there were any symptoms of dental abscess despite the fact that the use of antibiotics had been completed for a long time and no medicament was placed inside the pulp chamber.40 Dental abscess might exist when a tooth is nonvital in which the blood cannot circulate and bring oxygen to the tooth anymore. In this case, dental trepanation can be seen as a method to bring back oxygen circulation to a nonvital tooth.\n\nA study that measured the oxygen saturation level of a dental pulp found that the oxygen saturation level will be decreased when the pulp is in an unfavorable condition. Healthy teeth had the highest oxygen saturation level (94.6%), while reversible pulpitis, irreversible pulpitis, and pulpal necrosis were 85.4%, 81.6%, and 70.7%, respectively.41 There is a question from us about the oxygen saturation level of pulpal necrosis; whether it was measured on necrotic teeth with or without pulp exposure (e.g., nonvital teeth due to trauma). However, we can still conclude that the more severe the pathological state of the pulp, the lower the oxygen saturation level is.\n\nAnother interesting thing to discuss is the comparison between secondary caries and inactive caries. Cariogenic bacteria in saliva that enter through microcracks with a width of at least 50 μm between fillings and dental tissue are considered to cause secondary caries if they grow in an environment that suits them.42 On the other hand, caries can become inactive and do not develop for at least 4 to 5 years of follow-up in 85-90% of cases, making it not need any specific treatment, including restoration.43 In inactive caries, saliva certainly has no significant obstacle to contacting the tooth surface because no filling or anything is covering, but the bacteria are still not easy to progress. Suppose both secondary caries and inactive caries are exposed to saliva. In that case, the difference is that in inactive caries, the surface is more open and easier to clean to avoid debris adhesion. Inactive caries that do not develop for years is one reason to expect oxygen circulation can prevent infection in the root canal that priorly has been cleaned from various obstruction and contamination.\n\n\nMethods: novel membranous dental restoration system\n\nDental trepanation has been known by some, if not many, dental practitioners in Indonesia. It is also believed that the method still exists until recently. If this conventional method of dental trepanation - done without being followed by any modern technique of root canal preparation - was once considered beneficial for nonvital teeth, we hypothesize that a thoroughly cleaned, disinfected, and completely dried root canal, sealed with a specific restoration that uses an oxygen-permeable membrane, will give a better result. The membrane will let oxygen in while fluid, debris, and microorganisms cannot enter the root canal. The oxygen that keeps circulating to the entire root canal system and dentinal tubules will act as an antimicrobial agent that is constantly renewed to prevent the growth of pathogens inside the tooth.\n\nThe novel membranous restoration system described below (please see Figures 1 – 4) is expected to decrease the working time in endodontic treatment, as it can replace the time-consuming conventional obturation techniques. Any conventional sealer for obturation, including those that use nanoparticles, and any established technique, cannot give a perfect seal in the root canal system. Naturally, oxygen can flow automatically and fill any space it can infiltrate. Using oxygen as obturation biomaterials will hopefully be easier and more time-saving than all current obturation techniques that use solid substances. Before applying this restoration design, proper root canal preparation, including debridement and disinfection, should be done first.\n\nAccess to the root canal should be covered tightly to prevent debris and fluid from entering the clean and dry root canal. A small piece of rubber, taken from a rubber dam sheet, for example, can be used to seal the access to the root canal. Once the access is closed, tooth preparation can be performed.\n\nThe depth does not have to be the same as the working length.\n\nIn the case of a tooth with multiple roots, the duct shaped by the smooth broach must meet and be connected to all of the root canals.\n\nA smooth broach can be bent to make it possible to adjust the direction of the duct in accordance with what we have designed. The orifice of the duct can be placed on the surface where fluid and debris will be difficult to accumulate and is easy to clean when brushing teeth. Any other bendable needle, such as a finger spreader, can also be used as long as the size is suitable, can be pulled out from the restoration, and can be sterilized before inserting it to the root canal.\n\nUsing noncomposite materials is still possible as long as a small diameter duct is provided and connects the entire root canal with free air outside the restoration.\n\nThe oxygen-permeable membrane placed at the orifice of the duct is used to prevent any fluid, debris, and microorganisms from entering the root canal while letting oxygen go through. The membrane must also be of clinically safe material, durable to be placed in the mouth, and can be firmly attached to the selected restoration material. It should be noticed that the membrane must not be placed when the root canal is not clean and dry yet as the membrane cannot be passed by any fluid, including pus and irrigation solutions. In this case, it might be even better to use a membrane that can meet the following two criteria: it can let the gas flow from both sides of the membrane (two-way), and it can also let the fluid go through only from one side of the membrane (one-way).\n\nMembranes made of silanized alumina are both highly permeable to oxygen and hydrophobic. Without silanization, these membranes are hydrophilic. If it is possible to create a hydrophobic surface on one side and a hydrophilic surface on the other, we speculate that the same concept may apply to dental restoration. The oxygen-permeable membrane has never been utilized in the field of dentistry. However, silanized alumina membranes are claimed to be attractive for use in several technological disciplines, including oxygenation of blood during open-heart surgery.44\n\n\nConclusion\n\nBecause of the complexity of the root canal system and the tiny size of dentinal tubules, it is very complicated, if not impossible, to disinfect and obturate them entirely by using any recommended and established techniques. Endodontic treatment seems to require much time and multiple visits to deal with this challenge, which is unfavorable, especially during the pandemic. A new perspective is needed to reduce the working time as well as to answer some unsolved problems in endodontic treatment. The expected results of the development of this novel membranous dental restoration system in the future is not only to deal with microorganisms in a nonvital tooth, including Enterococcus faecalis, but also to find another way to fill the root canal system completely. Disinfecting the hard-to-kill E. faecalis and performing complex obturation techniques are somehow tiring and time-consuming. Oxygen is hoped to be the agent to prevent E. faecalis from being pathogenic and fill the root canal system with ease.\n\nIn this article, a dental restoration system that uses a membrane to provide oxygen to the root canal system is a somewhat novel idea we would like to introduce to the field of dentistry. By publishing the concept and making it available to the research community, any discussion might be useful to accelerate the exploration of the use of oxygen-permeable membrane for dental restoration. The function of the oxygen-permeable membrane in the restoration design is more or less similar to the medical mask used during the COVID-19 pandemic, as it enables the oxygen to circulate while preventing any fluid, debris, and microorganisms from passing. Further research is needed to find the most suitable membrane for dental restoration that allows the oxygen circulation at the utmost, and how effective this method can prevent infection. We are also open to collaborate with other researchers to implement the concept described in this article.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nAcknowledgements\n\nThe authors express their gratitude to Prof. Reidar K. Lie, MD, PhD (University of Bergen, Bergen, Norway) for giving valuable comments on drafts of the paper.\n\n\nReferences\n\nEuropean Centre for Disease and Control: SARS-CoV-2 variants of concern. February 24, 2022. [Accessed March 25, 2023]. Reference Source\n\nFini MB: Oral saliva and COVID-19. Oral Oncol. 2020; 108: 104821. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMallineni SK, Innes NP, Raggio DP, et al.: Coronavirus disease (COVID-19): Characteristics in children and considerations for dentists providing their care. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRiaz A, Maxood A, Abdullah S, et al.: Comparison of two intracanal medicament in resolution of apical radiolucency. J. Ayub Med. Coll. Abbottabad. 2018; 3: 320–324. PubMed Abstract\n\nZand V, Mokhtari H, Hasani A, et al.: Comparison of the Penetration Depth of Conventional and Nano-Particle Calcium Hydroxide into Dentinal Tubules. Iran. Endod. J. 2017; 12: 366–370. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMoreira MS, Anuar ASN, Tedesco TK, et al.: Endodontic treatment in single and multiple visits: An overview of systematic reviews. J. Endod. 2017; 43: 864–870. PubMed Abstract \| Publisher Full Text\n\nAndré AC, Debande L, Marteyn BS: The selective advantage of facultative anaerobes relies on their unique ability to cope with changing oxygen levels during infection. Cell. Microbiol. 2021; 23: e13338. Publisher Full Text\n\nOguntebi BR: Dentine tubule infection and endodontic therapy implications. Int. Endod. J. 1994; 27: 218–222. PubMed Abstract \| Publisher Full Text\n\nPrada I, Micó-Muñoz P, Giner-Lluesma T, et al.: Influence of microbiology on endodontic failure. Literature review. Med. Oral Patol. Oral Cir. Bucal. 2019; 24: 364–372. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSiqueira JF, Rõças IN: Microbiology and treatment of acute apical abscesses. Clin. Microbiol. Rev. 2013; 26: 255–273. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSanders JL, Houck RC: Dental Abscess. StatPearls. Treasure Island (FL): StatPearls Publishing; January -, 2022. [Updated 2021 Jul 17]. [Accessed December 15, 2021]. Reference Source\n\nNarayanan LL, Vaishnavi C: Endodontic microbiology. J. Conserv. Dent. 2010; 13: 233–239. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRuiz-Linares M, Aguado-Pérez B, Baca P, et al.: Efficacy of antimicrobial solutions against polymicrobial root canal biofilm. Int. Endod. J. 2017; 50: 77–83. PubMed Abstract \| Publisher Full Text\n\nFráter M, Braunitzer G, Urbán E, et al.: In vitro efficacy of different irrigating solutions against polymicrobial human root canal bacterial biofilms. Acta Microbiol. Immunol. Hung. 2013; 60: 187–199. PubMed Abstract \| Publisher Full Text\n\nTumbel LK, Wowor PM, Siagian KV: Uji daya hambat minyak kelapa murni (virgin coconut oil) terhadap pertumbuhan bakteri Enterococcus faecalis. Jurnal e-GiGi. 2017; 5: 1–6. Publisher Full Text\n\nThahir H, Djais AI, Nasir M, et al.: Virgin coconut oil as a new concept for periodontal tissue regenerations of TNF-α and TGF-β1. Int. J. Biomater. 2022; 2022: 1–8. Publisher Full Text\n\nShinsu University: New mechanisms discovered to separate air molecules.June 5, 2018. [Accessed December 15, 2021]. Reference Source\n\nGiudice GL, Cutroneo G, Centofanti A, et al.: Dentin morphology of root canal surface: A quantitative evaluation based on a scanning electronic microscopy study. Biomed. Res. Int. 2015; 2015: 164065. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nImlay JA: How oxygen damages microbes: Oxygen tolerance and obligate anaerobiosis. Adv. Microb. Physiol. 2002; 46: 111–153. Publisher Full Text\n\nHentges DJ: Anaerobes: General characteristics.Baron S, editor. Medical Microbiology. 4th ed.Galveston (TX): University of Texas Medical Branch at Galveston; 1996. [Accessed December 15, 2021]. Reference Source\n\nVatkar NA, Hegde V, Sathe S: Vitality of Enterococcus faecalis inside dentinal tubules after five root canal disinfection methods. J. Conserv. Dent. 2016; 19: 445–449. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAnjum J, Zaidi A, Barrett K, et al.: A potentially probiotic strain of Enterococcus faecalis from human milk that is avirulent, antibiotic sensitive, and nonbreaching of the gut barrier. Arch. Microbiol. 2022; 204: 158. PubMed Abstract \| Publisher Full Text\n\nHanchi H, Mottawea W, Sebei K, et al.: The genus Enterococcus: Between probiotic potential and safety concerns—An update. Front. Microbiol. 2018; 9: 1791. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGross CG: A hole in the head: a history of trepanation.June 11, 2021. [Accessed December 29, 2021]. Reference Source\n\nWang X, Zhang X, Cao H, et al.: Surgical treatments for infantile purulent meningitis complicated by subdural effusion. Med. Sci. Monit. 2015; 21: 3166–3171. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAlphianti LT: Apexification treatment with calcium hydroxide paste: 12 months evaluation (case report). Insisiva Dental Journal. 2014; 3: 52–59. Reference Source\n\nAnusha B, Madhusudhana K, Chinni SK, et al.: Assessment of pulp oxygen saturation levels by pulse oxymetry for pulpal diseases – A diagnostic study. J. Clin. Diagn. Res. 2017; 11: ZC36-9. PubMed Abstract \| Publisher Full Text\n\nFeng X: Cause of secondary caries and prevention. Hua Xi Kou Qiang Yi Xue Za Zhi. 2014; 32: 107–110. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZenkner JEA, Nora AD, Alves LS, et al.: Long-term follow up of inactive occlusal caries lesions: 4-5 years results. Clin. Oral Investig. 2019; 23: 847–853. PubMed Abstract \| Publisher Full Text\n\nAtwater JE, Akse JR: Oxygen-Permeable, Hydrophobic Membranes of Silanized alpha-Al2O3.October 1, 2006. [Accessed July 9, 2022]. Reference Source" }	[ { "id": "266227", "date": "09 May 2024", "name": "Tanay V Chaubal", "expertise": [ "Reviewer Expertise Periodontics", "Oral Implantology", "Systematic reviews and meta-analysis", "Bone regeneration", "Nanoparticles." ], "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 congratulate the authors in developing a novel membranous dental restoration system in endodontic management. The authors are aiming to change the environment inside the root canal system thereby making it more aerobic and unfavorable for the anaerobic bacteria like E.faecalis as it is the main culprit responsible for root canal failures. However, there are a few shortcomings which need to be addressed. Firstly, have the authors carried out any in-vitro tests to study their hypothesis? Specifically microbiological tests to support their claim of reducing or eliminating the root canal microbiota. Secondly, the authors have to elaborate the limitations or shortcomings of their proposed system and at the same time provide future research or protocols to be followed for making a reduction in the incidence of root canal failure.\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? Partly\n\nAre the conclusions about the method and its performance adequately supported by the findings presented in the article? Partly", "responses": [ { "c_id": "12759", "date": "08 Nov 2024", "name": "Didi Wahyudi", "role": "Author Response", "response": "Thank you for the congratulatory messages; we truly appreciate them, though we recognize that our journey to further explore trepanation is still lengthy. Currently, we are striving to present facts about trepanation, a procedure used in Indonesia, for which no detailed paper exists. In fact, a government-owned dental hospital located in Bandung, Indonesia, lists \"trepanasi,\" the Indonesian term for trepanation, as a service in both its pediatric dentistry (please see https://rskgm.bandung.go.id/klinik-pedodonti/ ) and endodontics (please see https://rskgm.bandung.go.id/klinik-konservasi-gigi/ ) departments. Our manuscript is an introductory text that serves as a starting point for a discussion on the topic of trepanation. This particular subject appears to have never been formally addressed in a comprehensive manner, even though it has been utilized by some, if not many, dental professionals in Indonesia. This writing is an initial effort that emphasizes a fundamental discussion regarding trepanation Please understand that the type of research that presents a hypothesis accompanied by logical analysis, supported by narratives reinforced with existing facts/cases and literature, is indeed acceptable for publication by many journals, including f1000research. This is to expedite the sharing of knowledge and existing phenomena. Further research, including those you have suggested, is hoped to be conducted in the future, either by us or by others who read this writing. However, we have included a discussion of a case report regarding trepanation, and we hope this can provide a clearer picture of trepanation. We also outline the limitations of the trepanation procedure mentioned in the case report and potential developments to address them, as you suggested. Thank you once again." } ] }, { "id": "237350", "date": "25 May 2024", "name": "Mohammed Zahedul Islam Nizami", "expertise": [ "Reviewer Expertise Cariology", "preventive and restorative dentistry", "dental biomaterials" ], "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 aim of this study is good. Clinical trials should be shown to introduce clinical methodology. Please present at least one clinical case example.\n\nIn the “Current endodontic treatment” section, please compare and contrast the existing methods, their limitations, and the rationale of the proposed methodology.\n\n“three factors in endodontic treatment” should be discussed under the method section with a consistent clinical flow of the methodology with a flow diagram.\n\nThe proposed methodology should be under the method section using a subtitle. A schematic illustration following the clinical/radiological image of the proposed method should be presented to the reader/clinician to make it clearer.\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\n\nAre the conclusions about the method and its performance adequately supported by the findings presented in the article? Partly", "responses": [ { "c_id": "12760", "date": "06 Nov 2024", "name": "Didi Wahyudi", "role": "Author Response", "response": "Thank you for the comments you have provided. We have included a case report on trepanation as you suggested. We agree that a clinical case example can help readers/clinicians, especially those who are not familiar with trepanation, to better visualize the concept. Although it took us some time to present this example, your suggestion was invaluable. We kindly ask you to recognize that the writing we have presented at this time does not delve into the specific advantages and disadvantages associated with some existing methods used in endodontic treatment. It is important to note that while our current work does not cover these aspects, there is potential for further research to be conducted in the future that could involve a detailed comparison of the different methods available. Once again, we greatly appreciate your suggestions regarding the clinical case example. This is what has caused us to take a long time to respond, as there has not yet been a paper that provides a sufficiently clear depiction of trepanation." } ] }, { "id": "171942", "date": "11 Sep 2024", "name": "Rachmi Fanani Hakim", "expertise": [ "Reviewer Expertise bio enginering", "stem cells research< natural product research", "wound healing" ], "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 strength of this article is that it offers a new method, but the weakness of writing a new method is that it lacks valid and up-to-date reference support and describes the problem less clearly. The introduction of this article should discuss the current state of endodontic biomaterials, describing theoretical, experimental or observational, so that it is clear why it is necessary to review a new system in root canal treatment as the purpose of this article. use the latest literature to explain concepts and trends in current endodontic care: Current Concepts and Trends in Endodontic Practice: An Update for Primary Dental Care (Blum I.et.al.2024 [Ref 2]) convey what are the advantages and disadvantages of current endodontic treatment in the introduction it links endodontic treatment with covid, but in the section \"Trepanation: bringing back oxygen circulation to a nonvital tooth\" it explains that However, multiple-visit root canal treatment is not preferable for patients, not only during the COVID-19 pandemic. If you want to link it to COVID, it is better to convey it with clear literature, so as not to confuse the reader. This sentence in the article must include clear, based and up-to-date references. ”Because the infection of the root canal of nonvital teeth tends to happen in an anaerobic condition, dental practitioners can consider preventing the root canal microenvironment from being oxygen-free. At some point, creating ventilation to avoid the absence of oxygen might look similar to trepanation. The size of the oxygen molecule is smaller than the diameter of dentinal tubules, making it able to fill the entire space available within the tooth. Oxygen can inhibit the growth of anaerobes and prevent facultative anaerobes from becoming more infectious. Using oxygen as an antimicrobial agent that fills the root canal is expected to shorten the working time of a dentist performing root canal treatments. It can replace conventional obturation after proper cleaning and shaping.” This sentence in the article must include clear, based and up-to-date references. “In inactive caries, saliva certainly has no significant obstacle to contacting the tooth surface because no filling or anything is covering, but the bacteria are still not easy to progress. Suppose both secondary caries and inactive caries are exposed to saliva. In that case, the difference is that in inactive caries, the surface is more open and easier to clean to avoid debris adhesion. Inactive caries that do not develop for years is one reason to expect oxygen circulation can prevent infection in the root canal that priorly has been cleaned from various obstruction and contamination.” This sentence in the article must include clear, based and up-to-date references. “Dental trepanation has been known by some, if not many, dental practitioners in Indonesia. It is also believed that the method still exists until recently. If this conventional method of dental trepanation - done without being followed by any modern technique of root canal preparation - was once considered beneficial for nonvital teeth, we hypothesize that a thoroughly cleaned, disinfected, and completely dried root canal, sealed with a specific restoration that uses an oxygen-permeable membrane, will give a better result. The membrane will let oxygen in while fluid, debris, and microorganisms cannot enter the root canal. The oxygen that keeps circulating to the entire root canal system and dentinal tubules will act as an antimicrobial agent that is constantly renewed to prevent the growth of pathogens inside the tooth. The novel membranous restoration system described below (please see Figures 1 – 4) is expected to decrease the working time in endodontic treatment, as it can replace the time-consuming conventional obturation techniques. Any conventional sealer for obturation, including those that use nanoparticles, and any established technique, cannot give a perfect seal in the root canal system. Naturally, oxygen can flow automatically and fill any space it can infiltrate. Using oxygen as obturation biomaterials will hopefully be easier and more time-saving than all current obturation techniques that use solid substances. Before applying this restoration design, proper root canal preparation, including debridement and disinfection, should be done first.” This sentence in the article must include clear, based and up-to-date references. The oxygen-permeable membrane placed at the orifice of the duct is used to prevent any fluid, debris, and microorganisms from entering the root canal while letting oxygen go through. The membrane must also be of clinically safe material, durable to be placed in the mouth, and can be firmly attached to the selected restoration material. It should be noticed that the membrane must not be placed when the root canal is not clean and dry yet as the membrane cannot be passed by any fluid, including pus and irrigation solutions. In this case, it might be even better to use a membrane that can meet the following two criteria: it can let the gas flow from both sides of the membrane (two-way), and it can also let the fluid go through only from one side of the membrane (one-way)\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\n\nAre the conclusions about the method and its performance adequately supported by the findings presented in the article? Partly", "responses": [] } ]	1	https://f1000research.com/articles/12-380
https://f1000research.com/articles/12-1420/v1	30 Oct 23	{ "type": "Case Study", "title": "Developing an integrated performance management and measurement system in healthcare organisations: a Canadian case study", "authors": [ "Anes Ben Fradj", "Neila El Asli", "Tasseda Boukherroub", "Claude Olivier", "Anes Ben Fradj", "Tasseda Boukherroub", "Claude Olivier" ], "abstract": "This study proposes a performance management and measurement system for a large healthcare organisation. First, data is collected to analyse and understand the current performance management system. Second, the SWOT (Strengths, Weaknesses, Opportunities, Threats) method is used to identify the main aspects of the performance management system to be improved. Third, based on the scientific literature and SWOT analysis, BSC principles are integrated to this performance management system to better align the organisation’s performance objectives and indicators with its strategy. Finally, we develop a performance indicator structure and specify indicators to be used as well as how these indicators could be integrated and shared with higher hierarchical levels in the organisation by using AHP (Analytic Hierarchy Process). Our approach is applied to the program “Physical disability, intellectual disability, and autism spectrum disorder” of CIUSSS du Centre-Sud-de-l’île-de-Montreal, a large healthcare network, in the province of Québec, Canada.", "keywords": [ "AHP", "BSC", "performance management", "healthcare" ], "content": "Introduction\n\nIn the province of Québec, Canada, public health and social services are provided by networks of institutions called CI (U)SSSs (integrated (University) centres of healthcare and social services), which fall under the responsibility of the Ministry of Healthcare and Social Services (referred to as MSSS). Their main mission is to provide healthcare and social services to the population while ensuring accessibility, quality, safety, and efficiency. This study focusses on the CIUSSS serving the South-Centre of Montreal region (referred to as CCSMTL). It provides 28 different healthcare services to a population of 311,000 people, it owns 150 facilities and employs more than 21,300 people, including 56 senior managers and nearly 800 physicians. It manages a budget of more than $1.7 billion Canadian dollars (Annual Report 2019-2020, CIUSSS). When the CCSMTL was created in 2015, it had to develop a reference framework for its performance management and to define a vision regarding its organisational performance. A Management and Accountability Agreement, involving the MSSS and the CCSMTL management, was introduced by the MSSS, with the obligation to implement a visual management system as a means for performance management and decision-making. Therefore, the CCSMTL has built its Quality-Performance Model (referred to as QPM), which encompasses four dimensions of performance; customer, accessibility/quality, mobilisation, and optimisation. It has also implemented its visual management system. However, QPM deployment is very challenging:\n\n1) It must be implemented at different decision-making levels, organisational structures (programs, sub-programs, services, teams, etc.), and territories. For instance, performance indicators may differ greatly from one service to another or from one territory to another.\n\n2) Some performance aspects such as customer and mobilisation dimensions are not sufficiently or well measured, which leads to an unbalanced performance management system.\n\n3) CCSMTL managers need to measure the overall performance of a given service or program, through integrated information. Currently, such information is not available nor easy to obtain.\n\nThis study contributes to addressing the aforementioned issues. More precisely, it proposes a four-phases based approach to support CCSMTL managers to efficiently implement its QPM and provide them with an efficient performance management and measurement system (PMMS). In phase I, data is collected to analyse the QPM and the current performance management system. In phase II, the SWOT (Strengths, Weaknesses, Opportunities, Threats) method is used to identify the main aspects of this performance management system to be improved. In phase III, based on the scientific literature and SWOT analyses, BSC (Balanced Scorecard) principles are integrated to the CCSMTL performance management system to better align its performance objectives and indicators with its strategy. Finally, in Phase IV, we develop a performance indicator structure and specify indicators to be used as well as how these indicators could be integrated and shared with higher hierarchical levels in the organisation by using AHP (Analytic Hierarch Process). A real case study (Physical disability, intellectual disability, and autism spectrum disorder, referred to as DI-TSA-DP) proposed by the CCSMTL is used to illustrate our approach.\n\nAmong recent studies in the literature that reported on performance management in complex healthcare organisations such as the CCSMTL, we can cite Moisan (2019) who focussed on the CISSS of Gaspésie region (province of Québec). This study highlighted the experience of this institution in deploying its performance management system and described how it contributed to improve the client trajectory performance in the context of social services dedicated to youth. However, it is descriptive and does not propose a PMMS for the organisation. Patient-reported outcome and experience measures (PROMs and PREMs) for system performance measurement is considered as a way to enhance and promote the shift of healthcare systems towards a more coordinated, integrated and people-centred care. This approach has been the focus of recent studies such as Bull and Callander (2022) and Nuti et al. (2017). Nuti et al. (2017) discussed the importance of using patient-reported measures as well as indicators based on administrative data to evaluate cross-sectional healthcare services in a multidimensional healthcare performance system. Bull and Callander (2022) discussed the different performance measurement programs employed across England, the US and Australia, and suggested key recommendations for advancing PROM and PREM programs internationally. Our work contributes to the theory and practice by proposing a performance management and measurement approach adapted to large healthcare organisations such as CCSMTL.\n\nThe remainder of this paper is organised as follows: the next section presents the QPM in more detail. After that, we present our literature review and then describe our approach. Following, we present how the approach is applied to the DI-TSA-DP case and discuss the preliminary results. Finally we conclude in the last section.\n\nThe QPM (see Figure 1) is based on the MSSS strategic plan (2029-2023 period) and framework for assessing the performance of the public healthcare and social services system (MSSS, 2012), Accreditation Canada’s quality management framework (Agrément-Canada, 2014), and PLANTREE’s person-centred approach (Cosgrove, 1994). The QPM encompasses four dimensions of performance: customer, accessibility/quality, mobilisation, and optimisation. Each dimension presents an aspect of the CCSMTL performance and encompasses a number of sub-dimensions enriched by the perspectives of the customers and the community (see Table 1).\n\nAccessibility/quality and customer are the two main dimensions of the QPM. They are presented in the form of a compass needle that points in the right direction to achieve better performance. Positioned at the centre of the model, customer dimension focusses on customer satisfaction regarding services and care provided. It presents three sub-dimensions: partnership, experience, and communication. Customers refer to people who receive a service or care and their relatives, as well as the community and the population in general. Accessibility/quality is the organisation’s ability to meet safely the needs and expectations of customers and ensure accessible and continuous services. It presents three sub-dimensions; accessibility, safety, and relevance/effectiveness of services. Mobilisation is the use of skills and talents of all people working in the organisation as well as fostering personal achievement and commitment to fulfil collectively the organisation’s mission. It encompasses four sub-dimensions, efficiency, agility, innovation, learning, and sustainability. Finally, optimisation is the search for continuous improvement in order to use efficiently available resources and provide services that are adapted to customer needs. Its sub-dimensions include healthy work environment, knowledge development, collaboration, and focus on people.\n\nThe CCSMTL also implemented its visual management system formed by a network of control rooms at different decision levels of the organisation. Put simply, a control room is a dedicated space where managers and employees meet regularly to acquire information about current situation and initiate discussions to improve future performance (Lagacé et Landry, 2016). At the highest organisational level, we find the strategic room, which is used by the CEO and senior managers. Data is used to evaluate the organisational performance, monitor current strategy, and support strategic decision-making (e.g. prioritising major projects). Tactical rooms are deployed in the different programs and sub-programs. Managers discuss their departments’ performance and decisions to be made at their level. Some of these decisions flow-up to the strategic room for validation while others flow-down to ‘intermediate’ tactical rooms or operational control rooms to be executed, in a cascade-escalation mechanism (MSSS, 2015). Some tactical rooms referred to as “intermediate tactical rooms” might play an intermediate role between tactical and operational levels. Operational control rooms (or visual stations) ensure day-to-day operations management (MSSS, 2015). The QPM is aimed at being implemented in all CCSMTL control room networks to support performance management.\n\nHowever, as mentioned earlier, QPM deployment is very challenging for the CCSMTL. First, it must be implemented at different decision-making levels, organisational structures, and territories. However, performance indicators may differ greatly from one service to another or from one territory to another. Second, some performance aspects such as customer and mobilisation dimensions are not sufficiently or well measured, which leads to an unbalanced performance management system. Finally, CCSMTL managers need to measure the overall performance of a given service or program, through integrated information. Currently, such information is not available nor easy to obtain.\n\nA performance measurement system could be seen as the process of assessing the organisation’s progress in achieving its goals (Kairu et al., 2013). It contains financial and non-financial measures (Okwo & Marire, 2012). It should enable the correct deployment of the strategic and tactical objectives and provide a structured framework enabling the relevant feedback of information to the appropriate levels to facilitate decision-making and control (Bititci et al., 1997). Botton et al. (2012) classified performance measurement systems according to three categories of approaches: strategy-oriented (BSC (Kaplan & Norton, 1996), performance prism (Neely et al., 2002)), hierarchical (performance pyramid), and process-based (macro-process model (Brown, 2008)). The QPM can be considered as strategy-oriented. Due to its multi-dimensional and strategy-focussed aspects, it is consistent with the BSC model. In their recent systematic review, Amer et al. (2021) provided managers and policymakers with evidence to support using BSC in the healthcare sector. They conclude that BSC implementation demonstrated positive outcomes for patient satisfaction and financial performance. In their systematic literature review, Betto et al. (2022) attempted to understand the evolution of BSC in healthcare and their results revealed that the majority of studies focussed mainly on the BSC design process, rather than BSC implementation, use, or review. To Bohm et al. (2021), the composition of BSC development teams in healthcare should consider including patients as well as their families where appropriate and select highly achievable and valid performance measures aligned with the organisational strategy.\n\nDeveloped in 1992 by Kaplan and Norton, the BSC is the result of a re-examination of performance measurement systems that usually give too much importance to the financial aspect. It is designed to measure four perspectives: finance (measures financial performance and the efficient use of financial resources), customer (concerns the organisation’s impact on the market in terms of customer satisfaction), internal processes (focusses on the efficiency and quality of internal processes) and learning & growth (concerns the culture and perspectives of the organisation, human resources, and information system). The term “balanced” underlines the concept of balance between all four axes of the model (Kaplan & Norton, 1992). In BSC model, we distinguish leading indicators, which measure processes, and lagging indicators that measure outcomes. Causality is one of the most important aspects in the implementation of BSC: there is an implicit causal relationship between BSC perspectives and lag-lead indicators (Porporato, Tsasis, et Vinuesa, 2017). To this end, a visual map (or strategy map) is built to show the objectives and causal links needed to execute a strategy. Scholey (2005) provides a well-structured six-step based process for creating a strategy map. Kaplan and Norton stressed the need to measure causal relationships, by using a complementary analytical approach (Barnabè et al., 2012) even though, it is not easy to analytically validate all causal relationships (Lorino, 2001; Schrage & Kiron, 2018). This is also the case for healthcare (Porporato et al., 2017).\n\nCanada leads in hospital performance measurement and historically, it is among the first countries to develop scorecards in healthcare (Porporato et al., 2017). In Toronto, Ontario, Women’s College Hospital started using a scorecard based on key objectives in the 1990’s (Porporato et al., 2017). Since then, Ontario hospitals have gained great experience in designing and using scorecards. Chan (2009) presented the evolution of healthcare systems in implementing BSC and strategy maps in Ontario. From 1992 to date, BSC has evolved from a strategic alignment tool (normative form) to a strategy construction tool. The normative version proposes a “top-down” deployment approach to operationalise the organisational strategy. Studies have shown that BSC can contribute to the emergence of a new strategy, and it would be better to opt for a “bottom-up” approach, where strategic objectives would be the result of collaborative management (Choffel & Meyssonnier, 2005; Ittner & Larcker, 2003; Norreklit, 2000; Simons, 1994). In addition, the learning & growth axis should aim to shape the organisation’s culture. Kaplan (2020) presented the case of merging St. Mary’s Hospital and Duluth Clinic (Minnesota, US) who co-created a new strategy, a strategy map and a common BSC. Montalan and Vincent (2011) proposed an approach for the co-construction of the strategy of a French hospital. They developed and validated a strategy map by a multidisciplinary teamwork following a bottom-up approach. Figure 2 shows the evolution of BSC over time and application in healthcare.\n\nIn order to measure the performance while considering the different dimensions of the BSC (healthcare and other sectors), some studies in the literature combined the BSC with multi-criteria decision-making techniques (MCDM). Most of these studies used the well-known Analytic Hierarchy (or Network) Process (AHP/ANP) (Anjomshoae et al., 2019; Chan, 2006; Modak et al., 2019; Regragui et al., 2018; Yaghoobi & Haddadi, 2016). Chan (2006) combined AHP and BSC to identify global metrics and compare performance across Canadian healthcare organisations. Regragui et al. (2018) also combined BSC and AHP to develop a framework for assessing performance in hospitals. By using AHP, the authors weighted different performance perspectives and indicators, leading to a ranking of these indicators. In other studies, AHP has been successfully used to consolidate, prioritise, and weight performance indicators (Bentes et al., 2012; Hossain et al., 2019; Yaghoobi & Haddadi, 2016). For more on AHP applications in healthcare, see Hummel and IJzerman (2011). Recently, Daneshmand et al. (2022) developed a BSC-AHP based model for assessing the health system of the Social Security Organisation in Iran. They conclude that, the most important performance dimensions are internal processes and social obligations. Table 2 presents most recent BSC applications in healthcare including studies that combined BSC and AHP. Our proposed approach is inspired by the work of Regragui et al. (2018).\n\n\nMethods\n\nOur approach encompasses two parts as illustrated in Figure 3. The first part (Phases I and II) aims at describing and analysing the current situation while the second part aims at developing a PMMS (Phases II and IV).\n\nThe data collection phase began in May 2019 (the study started in January 2019). Collected data was used to gain a more in-depth understanding of CCSMTL internal processes and current practices and progress regarding the implementation and use of its visual management system and QPM. To this end, three methods were used: review of documents, field observations, and interviews.\n\nDocuments reviewed include MSSS’s Law on healthcare and social services, Health Standards Organization (HSO) standards (HSO, 2021), Accreditation Canada (Agrément-Canada, 2014), Ministerial reference framework for evaluating performance in public healthcare and social services systems (MSSS, 2012), MSSS strategic plan 2019-2023, CCSMTL annual reports (CCSMTL, 2016), and PLANETREE documents (e.g. Cosgrove, 1994). Regarding field observations, we participated in the QPM Coordinating Committee and the QPM Advisory Committee meetings. The Coordinating Committee, which is composed of a limited number of managers, aims at insuring the implementation of the QPM within the CCSMTL, in accordance with the MSSS strategic plan. Periodically, the Coordinating Committee invites the heads of all CCSMTL departments to discuss decisions to be made related to the QPM and collect their feedback. This enlarged group forms the QPM Advisory Committee. In addition, we participated in a performance improvement workshop and visited several control rooms of the CCSMTL at different levels of the organisation: the strategic room, five tactical and intermediate rooms, and 10 operational rooms. Finally, we visited two hospitals and two rehabilitation centres to learn more about day-to-day healthcare operations. A total of 12 interviews were conducted with 13 managers (from different departments/services and decision-making levels) between May 2019 and September 2021. The objective was to validate and consolidate our understanding of the performance management and measurement system and tools being used and the progress in implementing the QPM and visual management system. The structured interview was used (DiCicco‐Bloom and Crabtree, 2006) (i.e., the interviewer asks several structured questions, and the interviewee responds freely to the questions). The participants were selected in collaboration with the deputy director of organisational performance at the CCSMTL, who is responsible for QPM implementation. If required, a second meeting was scheduled with the same participant(s).\n\nThe SWOT method was used to structure and analyse the collected data. Our findings and conclusions are summarised in Table 3. These results have been validated by two senior managers of the CCSMTL involved in the QPM implementation.\n\n\n\n- The QPM is well implemented at the strategic level.\n\n- Several practices ensuring high-quality services are implemented.\n\n- There is a willingness to embed a customer-focussed culture through the implementation of purposeful projects.\n\n- The visual management system is implemented at different decision-making levels.\n\n- There is a sufficient number of indicators within accessibility/quality dimension.\n\n- CCSMTL managers have a certain flexibility in proposing performance indicators for their departments.\n\n- There is good communication within the organisation.\n\n\n\n- The QPM is known at the tactical and operational levels, but not used for performance management and measurement purposes.\n\n- The QPM dimensions are unbalanced in terms of number of indicators within each dimension.\n\n- There is a lack of global and aggregated indicators.\n\n- It is not possible for senior managers to easily examine the performance of a specific service or department.\n\n- There is a discontinuity in the assessment of performance between the hierarchical levels.\n\n- The information presented in dashboards is not well discussed.\n\n- Some indicators are interdependent, and other ones are conflictual.\n\n\n\n- Several opportunities are provided by institutions such as PLANTREE, Agrément Canada, MSSS, etc. to implement best performance management practices.\n\n\n\n- Socio-economic context (pandemics such as COVID-19 and population aging).\n\n- Workforce shortage issue.\n\nOne of the strengths identified is the well implementation of the QPM at the strategic level. There is strong interest by senior managers in aligning the organisation’s practices with QPM objectives. In addition, there are several practices put in place to ensure service quality and customer satisfaction and there is a real willingness to embed a customer-focussed culture through the implementation of purposeful projects. The deployment and use (even partially at the moment of conducting this study) of the visual management system at different decision-making levels demonstrates a real desire to build an effective management and measurement performance system. We also identified that there are enough indicators in the accessibility/quality dimension. Another important aspect is that managers at different hierarchical levels have the possibility to propose performance indicators in compliance with general ministerial objectives (even though some indicators are imposed by the MSSS). Moreover, each department holds regular meetings, which helps to maintain communication within the departments and support the information cascade-escalation process. While the QPM is well implemented at the strategic level, it is not sufficiently operationalised at the tactical and operational levels. We also identified that the QPM’s dimensions are unbalanced in terms of number of performance indicators used within each dimension. We find many indicators measuring the quality/accessibility dimension, but there is a lack of indicators in optimisation, mobilisation, and customer dimensions. There is also a lack of specific targets for those indicators. With the current performance management system, it is not possible for senior managers to track performance indicators at tactical/operational levels or to verify the overall performance without consulting a large amount of information. In addition, it is not possible to examine easily the performance of a specific service or department. There is a discontinuity in the assessment of performance between hierarchical levels. That is, each department/service measures its performance independently of the higher hierarchical level and other departments/services. We identified that dependencies and conflicts among some indicators exist, and this is challenging for managers to deal with. For example, there is a need to simultaneously reduce waiting time and improve direct service time (time with customers). An improvement in one indicator can decrease the other and vice versa. Regarding opportunities, the CCSMTL must continuously challenge itself to maintain/obtain accreditations and certifications such as PLANTREE, Accreditation Canada and to also comply with the MSSS’s strategic plans. Finally, like most organisations in a continuously changing socio-economic context, the CCSMTL might be threatened by pandemics such as COVID-19 or workforce shortage. These aspects can notably lead to disrupt service accessibility.\n\nBased on the results of our SWOT analysis and inspired by previous studies in the literature, we propose to use BSC principles to create a more balanced QPM. To build our balanced-QPM, the four axes of the BSC and the four dimensions of the QPM are compared to each other. General causal relationships in this new balanced-QPM are also explicitly represented. Finally, the generic strategy map of the CCSMTL is derived from the resulting balanced-QPM.\n\nFigure 4 shows the dimensions of the proposed balanced-QPM. BSC’s customer perspective corresponds to the customer dimension of the QPM. In the CCSMTL context, this dimension reflects the organisation’s mission and takes the first position in the balanced-QPM (Figure 4). BSC’s internal process perspective is equivalent to the accessibility/quality dimension in the QPM. We recall that accessibility/quality refers to the ability to safely meet customer needs and expectations by providing accessible and continuous services. This dimension takes the second position in the balanced-QPM, forming with the customer perspective the “True North” of the CCSMTL. BSC’s learning & growth aspect is found in both mobilisation and optimisation dimensions of the QPM. Learning & growth presents three main components: human capital, information capital, and organisational capital. Human capital refers to the management of skills and behaviours of employees. This aspect is part of the mobilisation dimension of the QPM (use of skills and talents of everyone in the organisation, partners, customers, and their families, encouraging personal development and commitment to accomplish the mission of the CCSMTL). The mobilisation dimension takes the third position in the balanced-QPM. Organisational capital is defined as the organisation’s ability to align employee objectives with the strategy while information capital is the way organisations use their information systems, networks, and databases to support accomplishing their operations. Both aspects are part of the optimisation dimension of the QPM. We recall that optimisation dimension aims at continuous improvement and innovation to ensure, over time, that the services offered are adapted to the needs of customers. BSC’s financial perspective is also part of optimisation dimension. Kaplan and Norton (Kaplan & Norton, 1999) stated that the financial perspective serves as a constraint, not an objective, for non-profit organisations such as healthcare systems. Therefore, optimisation dimension, takes the last position in the balanced-QPM.\n\nFigure 4 shows that the customer dimension of the balanced-QPM corresponds to the organisation’s mission, and the other three dimensions to the CCSMTL’s capacity to fulfil this mission. Additionally, Figure 4 shows global causal links between the four dimensions. This aspect was absent in the current QPM. The optimisation dimension has direct impact on the accessibility/quality dimension. There is also a mutual relationship between optimisation and mobilisation. Mobilisation has an impact on the accessibility/quality of services provided by the CCSMTL, which in turn has direct impact on customer satisfaction. Finally, customers put pressure on the MSSS to provide the necessary funding supporting the CCSMTL capacity in terms of optimisation, mobilisation, accessibility/quality aspects.\n\nTo build the CCSMTL’s strategy map, we focus on the six-step guidelines suggested by Scholey (2005) adapted to a non-profit organisation. The first step is to determine the primary objective, which is the desired end result once the strategy is implemented. For the CCSMTL, the primary objective is to satisfy its customers by providing general and specialised healthcare and social services while insuring accessibility, security, efficiency and quality. An example of a target to achieve for this primary objective within a specific time frame would be to increase the access delay compliance rate to 95% within one year. The second step is to select the appropriate value proposition. According to Treacy and Wiersema (2007), there are three value propositions for an organisation: operational excellence, product excellence and customer relationship. Since the CCSMTL follows a customer-focussed approach, its value proposal focusses on customer experience. The third, fourth, fifth, and last step aims at identifying the objectives of the customer, accessibility/quality, mobilisation, and optimisation dimension, respectively. The determination of these objectives can be guided by both the QPM sub-dimension definitions enriched by the customer perspective (Table 1) and objectives set by the department managers, in relation to their specific mission. Rigorous criteria for selecting relevant objectives such as specificity, validity, sustainability, reliability, and utility were used to support this process. Figure 5 presents the generic strategy map of the CCSMTL that can be used as a basis for building “customised” strategy maps for different departments.\n\nThe causal links between the four dimensions shown in Figure 5 correspond to those identified in Figure 4. For the customised strategy maps, these causal links should be defined more precisely. This is illustrated with the DI-TSA-DP program case study. Validating analytically these causal links is beyond the scope of our study (time will be required to generate results and validate the causal links).\n\nThis phase aims at providing relevant indicators measuring performance following the objectives specified in the strategy map in each performance dimension.\n\nWe propose to create a structured system that provides indicators as well as performance indexes for each performance dimension at each hierarchical level of the CCSMTL. This general structure is based on the CCSMTL organisational chart and its visual management system. To provide and monitor the performance indicators, scorecards presenting the four dimensions (and their indicators) can be used at each department and hierarchical level, forming a network of scorecards (information is presented and discussed in the control rooms during regular meeting) (Figure 6). This proposal is inspired from the study of Voyer (Voyer & Voyer, 1999) that developed a network of interrelated dashboards used at different levels within an organisation. Béland and Abran (2004) also used this concept.\n\nBalanced-QPM scoredcard at the visual station level.\n\nFrom the established strategy map, performance indicators are selected for each dimension following strategic objectives that are relevant to the department/service. Existing indicators should be reviewed first. Second, additional indicators are proposed according to criteria such as specificity, validity, sustainability, reliability, utility, etc. At the end of this step, each department will have a set of indicators for each dimension of the Balanced-QPM called “before normalisation indicators” (Xbn).\n\nThe “before normalisation indicators” may have different units of measurement and different magnitudes. In some cases, the measurement unit is expressed in monetary units (e.g. expenses), and in others, in time units (e.g. hours) such as waiting time. In other cases, there is no measurement unit (e.g. number of services provided). In addition, the same indicator may have distinct targets in different departments. For example, waiting time target in an emergency department is significantly less compared to psychosocial services. Therefore, it might be difficult to compare them to each other based on their absolute values. Due to this, normalising the indicators is very helpful. Normalisation techniques for processing data have been used by many researchers in different fields. We refer the reader to Singh and Singh (2020) for more on normalisation. We selected the Min-Max normalisation technique (Singh & Singh, 2020), mainly because minimum and maximum limits can be easily distinguished in available data provided by the CCSMTL. Following this technique, data is usually rescaled within the range 0 to 1 or (-1) to 1. The general equation is given as follows:\n\nXnorm: Normalised value of the indicator\n\nXbn: Indicator value before normalisation\n\nmin: Minimum value in the data\n\nmax: Maximum value\n\nNminNmax: Normalised Min and Max\n\nFor the CCSMTL context Xnorm is expressed as a percentage ranging from 0% to 100%, therefore Nmin et Nmax values are 0% and 100%, respectively. Given that in some cases the maximum value of an indicator is the undesirable value while the minimum value is the desired value (e.g. the maximum value for waiting time is non-desired while the minimum value is desired), we set the best value in the data set as max parameter (which is not necessarily the highest value) and the worst value in the data as min (which is not necessarily the lowest value) before normalisation. Outliers must be eliminated before choosing the best and worst values. To link a given indicator to a desired objective and to ensure its continuous improvement, maxvalue corresponds to the indicator target. For each performance dimension, there is a set of associated indicators. Each indicator takes a numerical index i in the set and each indicator has an index dim referring to the first letter of the dimension to which it belongs (e.g. C for Customer). Indicators before normalisation take the index bn, referring to “before normalisation”. Normalised indicators in the original formula (Equation (1)) Xnorm become Xidim in the new formula adapted to the CCSMTL (Equation (2)). We remove the index norm for the sake of simplicity.\n\nEach department/service at different hierarchical levels calculates its normalised indicators (XiC,XiA,XiM,XiO). These are the inputs of the fourth step.\n\nStep 4 aims at weighting the normalised indicators of the same dimension at the same hierarchical level. AHP technique is used. AHP is usually used to evaluate and rank a set of evaluated based on several criteria. In the CCSMTL context, there is no need to rank or choose an option, but rather weighting the indicators within each performance dimension and aggregating them to obtain performance indexes for each dimension for a given department (see Figure 7). We selected AHP due its simplicity (Kumar et al., 2017) since it is very convenient for decision-makers who are not familiar with analytical tools. Moreover, it relies on pairwise comparisons, and according to Ishizaka and Labib (Ishizaka & Labib, 2011), psychologists argue that it is easier to express an opinion based on two elements rather than on all elements simultaneously. Figure 8 shows how AHP is adapted to our study.\n\nCriteria and indicators that are usually used in AHP, correspond to the four dimensions of the QPM and to the normalised indicators Xidim, respectively. The normalised indicators are compared to each other by using pair-wise comparison matrices (one matrix for each dimension) according to their relative importance regarding the dimension to which they belong. These preferences are expressed by managers. A given element aij of a matrix A corresponds to the relative importance of indicator i (in the row) compared to indicator j (in the column). The preference values range from one to five, which is adapted from the preference scale proposed by Saaty (2005). For example, aij = 1 means that the indicators are equally important and aij = 5 that indicator i is extremely important compared to indicator j. The weights of the performance indicators are calculated by using a specific algorithm described in detail by Brunelli (2014). All AHP calculation steps are presented in Appendix A. Comparisons make sense only if the pair-wise comparison matrices are coherent or quasi-coherent. A consistency ratio (CR) is used to check this aspect. If CR is less than 10%, the matrix is consistent, otherwise, the pair-wise comparisons are repeated (for more details see equations 4, 5 and 6 provided in Figure 10). Once consistency is validated, the weights of the indicators, Widim(WiC,WiA,WiM,WiO) are obtained.\n\nIndicators within each dimension are aggregated (see Figure 8), and four performance indexes PCPAPMPO are calculated (one index for each dimension) following Equation (3):\n\nPdim: Performance index of dimension dim\n\nWidim: the weight of indicator i within the performance dimension dim\n\nXidim: ith normalised indicator within performance dimension dim\n\nDI-TSA-DP case study\n\nTo illustrate how phases III and IV could be applied in practice, the “Physical disability, intellectual disability, and autism spectrum disorder” (DI-TSA-DP) program was proposed by the CCSMTL. First, DI-TSA-DP program is presented. Second, its strategy map is proposed (Phase III of our approach). Finally, the preliminary results of Phase IV (PMMS deployment) are presented, and a short discussion concludes this section.\n\nThe CCSMTL encompasses nine programs of which three; physical disability (DP), intellectual disability (DI), and autism spectrum troubles (TSA), are grouped together to form the joint program DI-TSA-DP. It provides services to 2,200 customers in local community service and rehabilitation centres. It receives approximately 10,000 service requests yearly and it encompasses 2,150 employees and 58 physicians. It includes three main sub-programs; DI-TSA, which provides services to customers with an intellectual disability or autism spectrum disorder, DP, which provides services to customers with a physical disability, and RMVS, which provides rehabilitation services in substitute living environments such as elderly homes. DI-TSA-DP mission is to provide specific, specialised and super-specialised habilitation and rehabilitation services to its customers to promote their integration and social participation.\n\nAt the moment of conducting this study, DI-TSA-DP has put in place one tactical room (top management level of the program), and three intermediate control rooms at the sub-program level (DI-TSA, DP, and RMVS). Operational rooms are established at the team unit level within the three sub-programs. Control room deployment at the team unit level was still in progress. Therefore, each sub-program had only reached a certain deployment rate, which is seen as a measure of performance-culture maturity level in the CCSMTL. Following the recommendation of our collaborators, we focussed on DP sub-program, since it had the highest deployment rate. The approach can be applied to the two other sub-programs as well. DP includes three services: AT (Technical Aids), which provides special assistance devices (prosthesis, wheelchair, etc.), LN (Locomotor-Neurology), which provides services to customers with physical disabilities due to neurological or locomotor disorders, and SL (Sensory-Language), which provides services to customers with linguistic or sensory disabilities. We focus on LN service, again based on its operational room deployment rate (100%). There are nine care team units within LN service, and each unit has its own operational room. We worked in close collaboration with two teams, Locomotor team (A-BOG), which provides services for customers with an amputation or serious orthopaedic injuries, and Neurological team (AVC), which provides services for customers who have had a stroke (Figure 9).\n\nPrior to the application of our approach to DI-TSA-DP, we organised five meetings with DI-TSA-DP managers (strategic and tactical levels) in order to understand the specific mission and structure of DI-TSA-DP, collect data regarding the QPM deployment within the program, and describe our approach to the managers. These meetings also enriched the SWOT analysis (see Table 3). One conclusion was the absence of a well-structured performance indicator system within DI-TSA-DP. Moreover, there were no explicit links between the QPM dimensions, sub-dimensions, and DI-TSA-DP strategic objectives. Existing objectives were set independently from the sub-dimensions and performance indicators were not explicitly related to these objectives. The strategy map built for DI-TSA-DP presented in the next paragraphs contributes to addressing these lacks.\n\nThe proposed strategy map for DI-TSA-DP (Figure 8) is consistent with the generic strategy map (Figure 5). It was co-created with the DI-TSA-DP program executive manager.\n\nIt illustrates objectives determined following the QPM sub-dimensions (including the primary objective) and causal links established between these objectives and/or sub-dimensions. The objectives selected are based on the internal DI-TSA-DP strategic vision 2018-2021 provided by DI-TSA-DP executive manager. Each objective was checked against the sub-dimensions of the QPM to identify to which one it is related. In total 16 objectives were classified within the sub-dimensions and dimensions of the QPM. The DI-TSA-SP mission was chosen as the primary objective and added in the customer dimension. Since no objective exists for customer experience, a new objective was proposed based on customers’ definition of this sub-dimension (Table 1). Additional objectives should be set for this sub-dimension and other ones that currently do not present any specific objectives (i.e., communication, learning and focus on people).\n\nExamples of causal links in the strategy map, are the relationship between objectives within customer sub-dimensions (‘partnership’ and experience’) and the primary objective (DI-TSA-DP mission). It is clear that positive customer experience, customer involvement as a partner in the system, and providing a climate of trust for the customers contribute to achieving DI-TSA-DP mission. It is interesting to notice that causal links also exist between sub-dimensions/objectives within the same dimension. As an example, partnership objectives contribute to achieving customer experience objectives. That is, when customers are involved as partners in the system, or when a climate of trust and collaborations is established with them, they are likely to have a positive experience. Some objectives/sub-dimensions mutually impact each-other, e.g. efficiency (optimisation dimension) and healthy work environment (mobilisation dimension). Reducing indirect labour and extra working hours (efficiency sub-dimension) leads to a healthy work environment, which in turn has a positive impact on efficiency.\n\nWe conducted five working meetings of two hours each with different DI-TSA-DP managers. We started with the operational level (A-BOG and AVC) and moved to the tactical level (LN and DP service and sub-program, respectively), and finally, to the highest level (DI-TSA-DP program). The following paragraphs present the process of applying the developed PMMS to DI-TSA-DP case.\n\nFigure 9 shows a simplified organisational chart of DI-TSA-DP program. This flowchart shows only the departments needed to illustrate our approach. We distinguish four levels: 1) CCSMTL strategic level, 2) tactical level, which includes DI-TSA-DP program and DI-TSA, DP, and RMVS sub-programs, and 3) operational level (within DP sub-program) that includes SL, LN, and AT teams.\n\nFigure 9 also shows the network of control rooms deployed in these different levels. The performance indicators structure follows the organisational chart and control room network structure. They are presented in the form of scorecards following the four QPM dimensions (indicators, values and targets, performance indexes).\n\nTo apply the rest of Phase IV steps, LN service was considered, and the process was performed in collaboration with LN coordinator. First, indicators being used by LN service were classified according to the four dimensions of the QPM and the strategy map’s sub-dimensions/objectives. Next, we checked the relevance of each performance indicator (against the criteria specificity, validity, simplicity, relevance, sustainability, reliability, and utility). When an indicator does not meet all criteria, it is either changed for another indicator that satisfies the criteria, or more information is gathered to ensure that the indicator meets all criteria. As an example, indicator ‘Customer satisfaction rate’ (Xan1C) (Table 4), which was proposed to measure customer experience sub-dimension objective, did not meet simplicity criterion. LN managers did not have information on customers’ satisfaction and did not put in place specific methods to collect this information. To address this issue, we relied on the results of a survey used to collect the feedback of customers regarding their satisfaction level regarding experience and service quality. Table 4 shows the 10 final indicators selected for LN service.\n\nThe columns of Table 4 show the indicators measuring the objectives (and therefore sub-dimensions) selected in previous step, their notations, their values before normalisation, their targets, their worst values, and their values after normalisation (obtained by using equation (2)).\n\nDue to confidentiality reasons, all real values of the indicators have been modified. Note that indicators that are already expressed as percentages before normalisation and which are aimed to be maximised to reach the desired target have not been normalised. The worst value is set to zero for those indicators (e.g. Customer satisfaction rate, Bed occupancy rate, and Respect of access deadlines). Moreover, indicators before normalisation meant to be minimised to reach their target have been converted into indicators to be maximisation after normalisation (e.g. Complaint rate and Ratio of salary insurance hours).\n\nPairwise comparisons of normalised indicators within each QPM dimension were performed with the LN coordinator. The values obtained were entered in a VBA program that checks the consistency of the matrices and generates the weights following AHP algorithm (see Appendix A). The final results that represent the weights of the indicators are presented in Table 5.\n\nStep 5 enables us to calculate aggregated performance indexes for each dimension that can be used by the service managers or by higher hierarchy level departments (DP sub-program and DI-TSA-DP programs in our case) to evaluate how the service performs globally in each dimension of the QPM. For instance, if the global performance of a given dimension is not satisfying, the performance indicators within that dimension should be analysed in more detail to identify the problem and take actions to improve the situation. The results of calculating the four performance indexes PILNCPILNAPILNMPILNO of LN are given in Table 5. Each performance index corresponds to one dimension of the balanced-QPM. Again, the VBA program is used to automate these calculations following AHP algorithm. Figure 10 provides more details about the AHP calculation steps\n\n\nDiscussion\n\nThe preliminary results of applying our approach to DI-TSA-DP program, and more specifically to LN service are promising. The starting point was the strategy map development, which is key for aligning DI-TSA-DP vision and strategy with the four dimensions/sub-dimensions of the QPM. This initial step helped to bring clarity and logic to the strategic objective determination process in relation to the QPM dimensions/sub-dimensions and performance measurement. Based on the strategy map, it was also possible to align LN service performance indicators with the four dimensions of the QPM through the strategic objectives and sub-dimensions selected. The strategy map plays a pivotal role in translating the program’s vision and strategy into strategic objectives and measurable indicators. Moreover, our approach contributes to balancing the four dimensions of the QPM by ensuring that a sufficient number of performance indicators measure each dimension. Performance indicators selection relied on a transparent and rigorous process that involved the participation of DI-TSA-DP and LN managers. To enable managers to evaluate the global performance of LN service, aggregating the performance indicators and calculating one performance index for each dimension by using an aggregative method (AHP) was appreciated by the managers.\n\nCCSMTL (and DI-TSA-DP program) case study helped us to better understand the challenges and difficulties managers may face during strategy map construction and PMMS development/implementation processes. Regarding the strategy map, iterative meetings with DI-TSA-DP program managers were necessary to build the version presented in this work. More collaboration with the management team is required to improve this preliminary version (i.e. determining more precisely the strategic objectives and causal links between them). When using AHP technique, we identified a compensation effect between indicators that results from aggregating two (or more) indicators having opposite values, which is not desired. For instance, in our case study, within Mobilisation dimension, the normalised indicator ‘Staffing level’ (X1M) has a value of 92% while the normalised indicator ‘Employee satisfaction rates (X13M) has a value of 70%. Their aggregation together with the normalised indicator ‘Ratio of salary insurance hours’ (X2M) (value of 84.5%), leads to a performance index with a value of 82,3% (PILNM) (Table 5). It is easy to see that the high value of ‘Staffing level indicator’ compensates the lower value of ‘Employee satisfaction rate’ indicator. This result also depends on the weights assigned to the three indicators (0.25, 0.25, and 0.50, respectively in our case). We also observed that choosing targets for the indicators was not an easy task for managers. In fact, most existing indicators are associated with organisational targets that are set by strategic management. Due of this, it is challenging for managers to select reasonable targets for new indicators.\n\nIn addition, deep discussions took place between managers during the processes of comparing indicators to each other to agree on common preferences. This shows the difficulty of making common pair-wise comparisons and deriving the right weights, and therefore the right performance index values. This aspect was also observed in Boukherroub et al. (2017). It deserves to be addressed in future work, for instance, by considering group-decision-making techniques in the weighting process or by discussing this issue at the strategic management to set clear guidelines to support managers at different hierarchical levels in defining the weights that best reflect the vision of the organisation. Dependencies among aggregated indicators might also occur. As an example, in psychosocial services, waiting time and customer service time evolve in opposite directions as an improvement in one indicator decreases the other and vice versa (i.e. when a social worker spends more time with his/her current customers, new customers assigned to him/her will wait longer before they can be met) (Boukherroub et al., 2022). Therefore, methods that address this issue are also required. Typically, the ANP technique (Analytic Network Process), which is a generalisation of AHP, is appropriate for this situation. BWM (Best Worst Method) (Singh & Singh, 2020) and MACBETH (Measuring Attractiveness by a Categorical Based Evaluation Technique) (Bana e Costa & Vansnick, 1994) are other possibilities.\n\n\nConclusions and research perspectives\n\nThe CCSMTL has developed a multi-dimensional performance model (Quality-Performance Model - QPM), however, its implementation across the organisation is very challenging. This study proposes an approach that contributes to addressing this issue. First, our proposed strategy map supports managers in identifying indicators that are aligned with their strategic objectives and vision. Second, the proposed PMMS helps them focus on all four dimensions of the QPM by explicitly linking performance indicators to each dimension. This, notably, supports proposing performance indicators within the customer dimension, which contributes to promoting customer-focussed culture. This would also allow departments to concretely measure their performance and compare themselves in a way that fosters competitiveness and anchors performance-oriented practices in the organisation. Third, the aggregated performance measures (performance indexes) that we propose to calculate based on AHP method will help managers at higher hierarchy levels to have a better visibility and understanding of the performance of operational teams, and make better decisions to support their improvement. Finally, our approach would support standardising performance management practices within the organisation. Our work contributes to the theory and practice by proposing a performance management and measurement approach adapted to large healthcare organisations, which is has been reported in practice and in the literature as a very challenging problem (Moisan, 2019).\n\nThis study has some limitations. Strategy maps should be built for all programs of the CCSMTL prior to indicator selection, validation, information aggregation, etc. to align the measured performance with strategic objectives. Brainstorming workshops should be carried out with managers and coordinators to establish more precisely their objectives and causal links between these objectives. More work is also required to design additional indicators. In particular, means of measuring and monitoring indicators over time need to be developed. Since AHP does not take into consideration potential interdependences between indicators, ANP, BWM, and MACBETH techniques could be explored. Undesired compensation effects might also occur in the aggregation process. Finally, it is not recommended to limit the use of performance indexes alone for monitoring performance. Operational indicators related for instance to emergency services are very important to monitor at the highest management level.\n\nThe following is a testimonial from the deputy director of organisational performance at the CCSMTL: “As part of this project, several working sessions were held with stakeholders [CCSMTL managers]. These structured interviews enabled the research team to well capture the QPM and then develop their approach accordingly. The data [performance indicators] organisation structure is very relevant in the sense that it addresses both the dimensions and the sub-dimensions of the QPM, and it is declined according to the hierarchical administrative structure of the CCSMTL, thus matching with our approach of continuous improvement, which aims to dynamise the information cascade-escalation process through all layers of the organisation. Finally, the rigorous process of validating and weighting the indicators was carried out with the stakeholders, thus ensuring better reliability of the indicators developed, which will optimise their use to eventually support decision-making.”", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nAcknowledgments\n\nThe authors acknowledge the financial support of MITACS Globalink program, ÉTS, CCSMTL, and Natural Sciences and Engineering Research Council of Canada (NSERC grant 2018-04555). The authors are grateful to Mr. Jacques Couillard, Mr. Jacques Drôlet, Mrs. Nancy Brunet, and all CCSMTL managers involved in this project.\n\n\nReferences\n\nAgrément-Canada: Guide sur l’évaluation de l’amélioration de la qualités les indicateurs de qualité.2014.\n\nAlharbi F, Atkins A, Stanier C, et al.: Strategic value of cloud computing in healthcare organisations using the Balanced Scorecard approach: a case study from a Saudi hospital. 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Publisher Full Text" }	[ { "id": "253135", "date": "09 Mar 2024", "name": "Jean-Baptiste Gartner", "expertise": [ "Reviewer Expertise Healthcare management sciences", "Performance management systems", "Care pathways optimization", "Learning Health 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\nComments to the author(s) This study aims to develop a performance management and measurement system for a large healthcare organisation (integrated (University) centres of healthcare and social services) in Quebec, Canada.\nINTRODUCTION/LITERATURE REVIEW - The background and the context are clearly stated. - However, the construction of your introduction between PMMS literature and context needs to be better realized to better accompany the reader. Indeed, the construction of your introduction goes from: 1. Contextualizing your case 2. Recent cases in literature 3. Model of QPM and its deployment 4. Literature review on PMS.\nI invite you to review the structure of the introduction. It might be relevant to merge the literature review section with the cases described in the literature (2 and 4), to conclude your introduction with the QPM model and its implementation issues, justifying your study. And then, conclude with the aim of the study.\nApart from the structure, certain points could be enhanced: - Avoid talking about the methods and results of the study in your introduction (paragraph “This study contributes to addressing”). - To further strengthen your literature: PMMS: Ferreira A, et. al. 2009 (Ref 1) Solomon R, et.al. 2013 (Ref 2)\nQuébec context: Fache P, et. al. 2016 (Ref 3)\nMETHODS The methodology is interesting and diverse. Some points could be improved or better justified:\n- Data collection (Phase I): The material should be better described and quantified (approximate total duration of observations, socio-demographic characteristics of interviewees). A table summarizing the material collected would be a great help. In addition, you do not specify the methodology for collecting information (notebook or reflective notebook for the observations, Semi-structured interviews (recorded or not)). Finally, the interview guide should be included as an appendix. - Data analysis (Phase II): Using SWOT to structure your results is very interesting. But you say nothing about the method of analysis and synthesis (thematic coding by two independent authors and comparison of results? or a coder and then pooling). Here you need to focus on the method description. Have you used methods to ensure the robustness of the analysis (internal and external validity, data triangulation, consistency)? If so, please specify. Table 3 and the following paragraph should be put in the results section, as it is results. - Balanced-QPM development (Phase III) and Indicator structure design and performance measurement (Phase IV): Stay with the process (the method) and keep the results for the results section.\nRESULTS Your results are interesting and relevant. Create a results section separate from the methods section.\n\nDISCUSSION The discussion needs to be further enriched and your results better compared with current knowledge (recent cases in literature and literature review). - From my point of view, the first two paragraphs of the discussion are still part of the results (implementation results). - Future work: Ok, your opening is relevant.\nSome typos and errors: Introduction: “The Coordinating Committee, which is composed of a limited number of managers, aims at insuring” (ensuring?) Methods: “Due of this, it is challenging” (to) Conclusion: “which is has been reported in practice and in the literature as a very challenging problem”\nIn summary: - The topic is interesting, and the study mobilizes a wealth of material to propose results relevant to the field. - There are several study structuring issues that need to be addressed (structure of introduction, absence of results title, structure of discussion. A number of tips are given for this purpose. - A number of methodological points need to be reinforced in order to fully convince the reader. - The literature could be strengthened to develop the discussion on the contribution of this study. Proposals to this effect have also been made.\nGood luck for the next steps.\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? 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? No\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? Partly", "responses": [ { "c_id": "12672", "date": "09 Dec 2024", "name": "NEILA EL ASLI", "role": "Author Response", "response": "In the following, you will find our responses in bold style. This study aims to develop a performance management and measurement system for a large healthcare organisation (integrated (University) centres of healthcare and social services) in Quebec, Canada. INTRODUCTION/LITERATURE REVIEW - The background and the context are clearly stated. - However, the construction of your introduction between PMMS literature and context needs to be better realized to better accompany the reader. Indeed, the construction of your introduction goes from: 1. Contextualizing your case 2. Recent cases in literature 3. Model of QPM and its deployment 4. Literature review on PMS. I invite you to review the structure of the introduction. It might be relevant to merge the literature review section with the cases described in the literature (2 and 4), to conclude your introduction with the QPM model and its implementation issues, justifying your study. And then, conclude with the aim of the study. Thanks for this suggestion. Parts 3 and 4 are normally two distinct sections separated from the Introduction. The formatting of the journal merged them with the Introduction. We have corrected this. We agree that it is better to merge parts 2 and 4 and to conclude the introduction with the QPM model and the aim of the study. Please see the changes we have made in the new version of the manuscript. Apart from the structure, certain points could be enhanced: - Avoid talking about the methods and results of the study in your introduction (paragraph “This study contributes to addressing”). Thanks for this suggestion. We have removed the details concerning the approach proposed as well as the description of the results in the new version of the manuscript. - To further strengthen your literature: PMMS: Ferreira A, et. al. 2009 (Ref 1) Solomon R, et.al. 2013 (Ref 2) Québec context: Fache P, et. al. 2016 (Ref 3) 1. Ferreira A, Otley D: The design and use of performance management systems: An extended framework for analysis. Management Accounting Research. 2009; 20 (4): 263-282 Publisher Full Text 2. Solomon R, Damba C, Bryant S: Measuring quality at a system level: an impossible task? The Toronto Central LHIN Experience.Healthc Q. 2013; 16 (4): 36-42 PubMed Abstract \| Publisher Full Text 3. Fache P, Sicotte C, Minvielle É: The Search is on for Coherent Performance Measurement in Healthcare Organizations. Has Quebec Reached a Crossroads?. Healthc Policy. 2016; 11 (4): 60-9 PubMed Abstract Thanks for these references. We have added them in the literature review section of the new version of the Manuscript. METHODS The methodology is interesting and diverse. Some points could be improved or better justified: - Data collection (Phase I): The material should be better described and quantified (approximate total duration of observations , socio-demographic characteristics of interviewees). A table summarizing the material collected would be a great help. In addition, you do not specify the methodology for collecting information (notebook or reflective notebook for the observations, Semi-structured interviews (recorded or not)). Finally, the interview guide should be included as an appendix. In the new version of the manuscript, we have added a table summarizing the data collection methods used for conducting phase I and short descriptions (Table 3). We also referred the reader to a work presenting more in detail on the collection data methods in the text (Ben Fradj, 2020). Unfortunately, the journal does not allow including any appendix. - Data analysis (Phase II): Using SWOT to structure your results is very interesting. But you say nothing about the method of analysis and synthesis (thematic coding by two independent authors and comparison of results? or a coder and then pooling). Here you need to focus on the method description. Have you used methods to ensure the robustness of the analysis (internal and external validity, data triangulation, consistency)? If so, please specify. Table 3 and the following paragraph should be put in the results section, as it is results. We have added more information in the text to describe how we conducted the analysis with and moved Table 3 and the paragraphs commenting it to the Results section (Table 4 in the new version of the manuscript). - Balanced-QPM development (Phase III) and Indicator structure design and performance measurement (Phase IV): Stay with the process (the method) and keep the results for the results section. Thanks for this comment. In the new version of the manuscript, we have kept only the description of the methods used and moved all the results to the Results section. RESULTS Your results are interesting and relevant. Create a results section separate from the methods section. Thanks for this suggestion. We have now a Results section. DISCUSSION The discussion needs to be further enriched and your results better compared with current knowledge (recent cases in literature and literature review). From my point of view, the first two paragraphs of the discussion are still part of the results (implementation results). You are right, we improved the discussion section following your recommendation. - Future work: Ok, your opening is relevant. Thanks. Some typos and errors: Introduction: “The Coordinating Committee, which is composed of a limited number of managers, aims at insuring” (ensuring?) Right “ensuring”. Methods: “Due of this, it is challenging” (to) Indeed, “to” is the correct term to use. Conclusion: “which is has been reported in practice and in the literature as a very challenging problem” Corrected, “which has been”. In summary: - The topic is interesting, and the study mobilizes a wealth of material to propose results relevant to the field. - There are several study structuring issues that need to be addressed (structure of introduction, absence of results title, structure of discussion. A number of tips are given for this purpose. - A number of methodological points need to be reinforced in order to fully convince the reader. - The literature could be strengthened to develop the discussion on the contribution of this study. Proposals to this effect have also been made. Good luck for the next steps. Thanks for the relevant comments and suggestions that we believe significantly improved our manuscript." } ] }, { "id": "253136", "date": "28 Mar 2024", "name": "Wolfgang Munar", "expertise": [ "Reviewer Expertise Health policy and systems research", "and theory-driven evaluation." ], "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 solid paper contributes to ongoing debates on designing performance measurement and management systems in healthcare organizations. The authors describe the process followed to complete the system's design (i.e., situational analysis, SWOT analysis, and literature review), leading to the development of a performance indicator framework. An indicative application of the resulting design was then applied to the CIUSSS program.\nThe main two areas to consider for improving the paper are described below.\n1. While the authors recognize the complexity inherent to PMM system design (and implementation), there is insufficient analysis/discussion of the proposed design's strengths and limitations to address such complexity (e.g., interdependence and recurrence between indicators, gaming, users' cognitive biases, etc.).\n\n2. The authors' use of the concept of \"implementation\" requires clarification. While the paper clearly presents a prototype for a PMM system, it cannot be said that it describes its implementation.\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? 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\n\nIs the case presented with sufficient detail to be useful for teaching or other practitioners? Yes", "responses": [ { "c_id": "12729", "date": "09 Dec 2024", "name": "NEILA EL ASLI", "role": "Author Response", "response": "This solid paper contributes to ongoing debates on designing performance measurement and management systems in healthcare organizations. The authors describe the process followed to complete the system's design (i.e., situational analysis, SWOT analysis, and literature review), leading to the development of a performance indicator framework. An indicative application of the resulting design was then applied to the CIUSSS program. The main two areas to consider for improving the paper are described below. 1. While the authors recognize the complexity inherent to PMM system design (and implementation), there is insufficient analysis/discussion of the proposed design's strengths and limitations to address such complexity (e.g., interdependence and recurrence between indicators, gaming, users' cognitive biases, etc.). Author Response: Thanks for this comment. We have improved the discussion section in the new version of the manuscript. 2. The authors' use of the concept of \"implementation\" requires clarification. While the paper clearly presents a prototype for a PMM system, it cannot be said that it describes its implementation. Author Response: We agree that the us of the term “implementation” in the text can be confusing. We clarified this in the new version of the manuscript at different levels: We changed the title from “Developing an integrated ….” to “Towrads an integarted …”. In the abstract, the first sentence is now: “This study proposes an approach for developing or improving performance management and measurement systems (PMMSs) for healthcare organisations.”. In the Introduction (last paragraph): “…it proposes a four-phases based approach to support CCSMTL managers to improve and transform the QPM into a more balanced and effective performance management and measurement system (PMMS). … It can be used by theoreticians and practitioners as roadmap to develop or improve healthcare organisations’ PMMSs. In the Discussion section: ““In this study, we have proposed a comprehensive and quantitative approach to develop or improve a PMMS.” In the Conclusion section: “The CCSMTL has developed a multi-dimensional performance model (Quality-Performance Model - QPM), however, its implementation across the organisation is very challenging. This study proposes an approach that contributes to improving this model and its effective deployment.”" } ] }, { "id": "238355", "date": "04 Apr 2024", "name": "Paulino Silva", "expertise": [ "Reviewer Expertise Management Accounting" ], "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 presents a very relevant topic with great added value, both for the organizational performance management literature and for management practitioners. The contribution of the article can be relevant for the body of knowledge. However, there are some methodological weaknesses that should be considered. The article claims to develop an integrated performance management system for healthcare organizations, but perhaps it would be more appropriate to define it as an improvement to the existing system. This is because it makes a diagnosis of the existing system and, through SWOT analysis, presents the main strengths, weaknesses, opportunities, and threats. The used case study seems too specific (Physical disability, intellectual disability, and autism spectrum disorder\" of CIUSSS du Centre-Sud-de-l'île-de-Montreal, in the province of Québec, Canada) to be considered a reference of a performance management system in healthcare organizations. As the study began in 2019, and the interviews between May 2019 and September 2021, could the results have been too affected by the Covid-19 pandemic? The number of interviews seems to be very low (only 12) and the variety of jobs very limited (only managers). Consider including additional literature specifically related to performance management in healthcare organizations, such as : [1] Some “typos” in the article:\nThe first paragraph of the methods section is incorrect, it should end with phases III and IV. The abbreviation for Ministry of Healthcare and Social Services presented in the article is MSSS and not MHSS, why? Customer-focused instead of customer-focussed in line 6 of the section Conclusions and research perspectives.\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? Partly\n\nIf applicable, is the statistical analysis and its interpretation appropriate? I 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? Yes\n\nIs the case presented with sufficient detail to be useful for teaching or other practitioners? Yes", "responses": [ { "c_id": "12730", "date": "09 Dec 2024", "name": "NEILA EL ASLI", "role": "Author Response", "response": "Reviewer #3: APPROVED WITH RESERVATIONS The article presents a very relevant topic with great added value, both for the organizational performance management literature and for management practitioners. The contribution of the article can be relevant for the body of knowledge. However, there are some methodological weaknesses that should be considered. The article claims to develop an integrated performance management system for healthcare organizations, but perhaps it would be more appropriate to define it as an improvement to the existing system. This is because it makes a diagnosis of the existing system and, through SWOT analysis, presents the main strengths, weaknesses, opportunities, and threats. You are right. The PMMS (Performance Management and Measurement System) we proposed is an improved Quality-Performance Model (QPM) in the context of the CCSMTL. However, in the absence of a performance model, our approach (or at least part of it if we remove the SWOT analysis), could be used to develop a PMMS. We clarified this in the text in the new version of the manuscript at different levels: We changed the title from “Developing an integrated ….” to “Towrads an integarted …”. In the abstract, the first sentence is now: “This study proposes an approach for developing or improving performance management and measurement systems (PMMSs) for healthcare organisations.”. In the Introduction (last paragraph): “…it proposes a four-phases based approach to support CCSMTL managers to improve and transform the QPM into a more balanced and effective performance management and measurement system (PMMS). … It can be used by theoreticians and practitioners as roadmap to develop or improve healthcare organisations’ PMMSs. In the Discussion section: ““In this study, we have proposed a comprehensive and quantitative approach to develop or improve a PMMS.” In the Conclusion section: “The CCSMTL has developed a multi-dimensional performance model (Quality-Performance Model - QPM), however, its implementation across the organisation is very challenging. This study proposes an approach that contributes to improving this model and its effective deployment.” The used case study seems too specific (Physical disability, intellectual disability, and autism spectrum disorder\" of CIUSSS du Centre-Sud-de-l'île-de-Montreal, in the province of Québec, Canada) to be considered a reference of a performance management system in healthcare organizations. There are two levels of application of our PMMS. We applied Phases I, II, III, and partially IV to the entire CIUSSS, and then Phases III and IV to the program Physical disability, intellectual disability, and autism spectrum disorder. We have made changes in the new version of the manuscript to clarify this (please see the Results section). We agree that more work is needed before we can consider this system as a reference in healthcare organizations. We have acknowledged this limitation in the discussion Section, and as mentioned before, we also have changed the title. As the study began in 2019, and the interviews between May 2019 and September 2021, could the results have been too affected by the Covid-19 pandemic? This is right, part of the data collection took place during the COVID-19 pandemic (2020). During this period, the process slowed down; it was difficult to reach people. This resulted in a lesser number of interviews than what we targeted initially. However, it is difficult to estimate the impact on the results though. The number of interviews seems to be very low (only 12) and the variety of jobs very limited (only managers). As mentioned above, initially we targeted more interviews. Even though all the participants involved are managers, there is a certain variety considering that they don’t have the same function, hierarchy level in the organization, and they don’t come from the same programs/services (please see Table 3 in the new version of the manuscript). Employees were also met during the data collection phase. However, their knowledge level regarding performance measurement, notably in relation to the QPM, was too low or inexistant (in some services, there are no performance indicators put in place at all). This lack of knowledge at the operational level has been mentioned as a weakness in the SWOT analysis “The QPM is known at the tactical and operational levels, but not used for performance management and measurement purposes”. We appreciate your comment. We have discussed this as a limitation in the new version of the manuscript. Consider including additional literature specifically related to performance management in healthcare organizations, such as : [1] 1. Silva P, Ferreira A: Performance management in primary healthcare services: evidence from a field study. Qualitative Research in Accounting & Management. 2010; 7 (4): 424-449 Publisher Full Text Thanks for this suggestion. We have included it in our literature review. We have also added three other references suggested by Reviewer #1. Some “typos” in the article: The first paragraph of the methods section is incorrect, it should end with phases III and IV. You are right. It has been corrected. The abbreviation for Ministry of Healthcare and Social Services presented in the article is MSSS and not MHSS, why? MSSS stands for Ministère de la Santé et des Services Sociaux in French (official title of the Ministry). We will have added this information in the new version of the manuscript to avoid confusion. Customer-focused instead of customer-focussed in line 6 of the section Conclusions and research perspectives. Thanks, we have corrected it." } ] }, { "id": "219887", "date": "16 Sep 2024", "name": "Pascal Forget", "expertise": [ "Reviewer Expertise Industrial engineering", "operation research", "simulation" ], "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 proposes an approach to facilitate the implementation of a performance management model in a government healthcare system. This article is really interesting and offer a pertinent contribution. It is clear and well written. The introduction, the literature, the method and discussion are clear and well presented. There is no specific point to address and the paper could be index as it is.\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? 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? Yes\n\nIs the case presented with sufficient detail to be useful for teaching or other practitioners? Yes", "responses": [] } ]	1	https://f1000research.com/articles/12-1420
https://f1000research.com/articles/12-518/v5	28 Oct 24	{ "type": "Opinion Article", "title": "Viral interactions with host factors (TIM-1, TAM -receptors, Glut-1) are related to the disruption of glucose and ascorbate transport and homeostasis, causing the haemorrhagic manifestations of viral haemorrhagic fevers.", "authors": [ "Ivan Chicano Wust" ], "abstract": "The haemorrhagic features of viral haemorrhagic fevers may be caused by common patterns of metabolic disturbances of the glucose and ascorbate homeostasis. Haemorrhages and vasculature disfunctions are a clinical feature not only of viral haemorrhagic fevers, but also in scurvy, diabetes and thrombotic microangiopathic haemolytic anaemia. Interestingly, the expression of glucose and ascorbate transporter Glut-1 on the erythrocyte membrane is associated with the inability to synthesize ascorbate and is restricted to that very species that are susceptible to filoviruses (primates, humans and fruit bats). Glut-1 may play a pivotal role in haemorrhagic fever pathogenesis. TIM-1 and TAM receptors have been recognized to enhance entry of Ebola, Lassa and Dengue viruses and viral interferences with TIM-1 could disturb its function, disturbing the expression of Glut-1. In those species not able to synthesize ascorbate and expressing Glut-1 on erythrocytes virus could interact with Glut-1 or other functionally related protein, and the influx of glucose into the cells would be severely impaired. As a consequence, transient hyperglycemia and a marked oxidative stress coupled with the high levels of glucose in plasma would be established, and then promote the activation of NF–κB transcription, exacerbating a pro-inflammatory response mediated by cytokines and chemokines: The inability to synthesize ascorbate is an Achilles Heel when trying to counteract the oxidative stress.", "keywords": [ "Ebola virus", "Lassa virus", "Dengue virus", "haemorrhagic fevers", "glucose", "ascorbate", "oxidative stress" ], "content": "The haemorrhagic features of viral haemorrhagic fevers are caused by common patterns of metabolic disturbances of the glucose and ascorbate homeostasis\n\nIn the present opinion article, it will be disclosed that the haemorrhagic features of viral haemorrhagic fevers in general, and those caused by Lassa virus, dengue virus and especially Ebola virus, in particular, are caused by common patterns of metabolic disturbances of the glucose and ascorbate homeostasis. Disruption of glucose and ascorbate homeostasis could be the reason of both enhanced inflammatory cytokine storming and haemorrhagic manifestations at the level of endothelial cells and vasculature injury.1–5 Endothelial damage during filovirus infection with no evidence of direct endothelial cytolysis has previously been described,6 reinforcing the idea that other indirect mechanisms governing vasculature injury are present. Intracellular ascorbic acid has been considered as an enhancer of nitric oxide (NO) synthesis7 and the macromolecular permeability of cultured human umbilical vein endothelial cell monolayers was decreased significantly in culture medium containing L-ascorbic acid and L-ascorbic acid 2-phosphate.8 Apoptosis in endothelial cells can be induced by hyperglycemia and ascorbate helps to prevent endothelial dysfunction, stimulates type-IV collagen synthesis and enhances cell proliferation. Haemorrhages and vasculature disfunctions are a clinical feature not only of viral haemorrhagic fevers, but also in scurvy, diabetes and thrombotic microangiopathic haemolytic anaemia. Inflammatory cytokine concentrations were found to be acutely increased by hyperglycaemia in humans,4,5 which has been suggested as the causative role in the vasculature disorders and immune activation of diabetes. Hypoascorbinaemia and diabetes mellitus share several clinical symptoms including microangiopathy, capillary hyperperfusion and haemorrhages.\n\n\nMicroangiopathic haemolytic anemia: a viral haemorrhagic fever without virus?\n\nAn early description by Symmers in 1952 of the thrombotic microangiopathic haemolytic anaemia,9 which extraordinarily fits the haemorrhagic and vascular manifestations of filovirus haemorrhagic fever (FHF), included haemorrhagic manifestations in the form of widespread purpura. As described by Symmers, “there might be haemorrhage from any of the orifices of the body”. A prodromal period of fever, muscle and joint pains and vague abdominal pain existed and included also mental confusion and neurological symptoms. The parallel of the thrombotic microangiopathic haemolytic anaemia with the filovirus pathological manifestations is important and would become more evident in its physiological implications basing on the hypothesis description of virus-driven biochemical events.\n\nSeveral receptor candidates have been proposed as mediators for filovirus entry and several attempts to develop a vaccine have been pursued but there is no suitable definitive biological explanation of the violence of the illness manifestations, nor are well defined the factors underlying the differences in illness severity between viruses and within the species range. Whereas many experimental data are available and have rendered important descriptive and molecular information, there is not an integrative and comprehensive perspective. A better comprehension of the virus pathophysiology could help in developing a treatment strategy. According to literature major symptoms of Ebola virus disease include a maculopapular rash and mucosal haemorrhage. It begins unspecific with fever, asthenia, diarrhea, headaches, myalgia, arthralgia, vomiting and abdominal pain and disease course develops sharply and resolves in survival or death in the course of approximately 3 weeks. Massive bleeding along with fluid distribution problems, disseminated intravascular coagulation and focal necrosis are observed in fatal cases. Arthralgia, asthenia, and neurological disorders, as dysesthesias, are often sequelae in the convalescence, which lasts weeks to months. Main features of Ebola virus haemorrhagic fevers can be consulted in available literature.10–14\n\nGlucose transporter 1 or Glut 1, also known as solute carrier family 2, facilitated glucose transporter member 1 (SLC2A1), is a uniporter protein that in humans is encoded by the SLC2A1 gene.15 Glut 1 facilitates the transport of glucose across the plasma membranes of mammalian cells.16 Glut 1 is responsible for the low level of basal glucose uptake required to sustain respiration in all cells. Glut 1 is also a major receptor for uptake of vitamin C (ascorbate), especially in non-ascorbate producing mammals as part of an adaptation to compensate by participating in a vitamin C recycling process. In mammals that do produce vitamin C, Glut 4 is often expressed instead of Glut 1.17 Interestingly, expression of Glut-1 on the erythrocyte membrane is associated with the inability to synthesize ascorbate18 and is restricted to that very species that are susceptible to filoviruses or are considered to be the reservoir of the virus in nature (primates, humans and fruit bats). Glucose and ascorbate metabolism represent probably an Achilles’ heel in filovirus haemorrhagic fevers (FHF) and Glut-1 may play a pivotal role in haemorrhagic fever pathogenesis. In other words, Glut-1 on erythrocytes and inability to synthesize ascorbate may account for the pathophysiology of filovirus haemorrhagic fevers in susceptible species. Ebola and Marburg viruses cause haemorrhagic fever in human and non-human primates. Guinea pigs are frequently used as an animal model after a trivial virus strain adaptation. Fruit bats are considered the reservoir of filoviruses in nature.19 Extraordinary mechanisms regarding glucose absorption and transport are displayed by some fruit bats in which serological evidence to Ebola virus was confirmed or Marburg virus was isolated. Pigs,20,21 which develop symptomless infection by filoviruses differ from the inability of other species to synthesize ascorbate and the presence of Glut-1 on the erythrocyte membranes. Thus, filoviruses in those species expressing the glucose and ascorbate transporter Glut-1 on erythrocytes and unable to synthesize ascorbate, would cause pathophysiological changes that account for the severity of the filoviral haemorrhagic fevers, by means of a virus-driven disruption or overwhelming of ascorbate and/or glucose homeostasis. The inability to synthesize ascorbate could represent an Achilles’ heel in counteracting the exacerbated inflammatory response22 during Ebola haemorrhagic fever. Transport of dehydroascorbic acid (DHA) from plasma is coupled with its reduction with glutathione (GSH) and NADH. Erythrocytes, with the highest level of Glut-1 expression, regulate the concentrations of ascorbate in plasma by the influx of DHA and efflux of DHA and ascorbate. Glucose and DHA compete for the same transporter and as plasma concentrations of D-glucose are quite higher than DHA concentrations, DHA uptake is more important when ascorbate is rapidly oxidized outside the cells, much like in areas of inflammation, as during FHF.\n\nViral interactions of filoviruses, or interferences, with Glut-1, specially on the erythrocyte membrane could be hypothesized. This might account for the pathophysiological changes during infection. In favor of direct glycoprotein interactions with Glut-1 are the experiments demonstrating inhibition of Ebola-glycoprotein mediated entry by cytokalasin B,23 which is a potent inhibitor of glucose and DHA influx transport in mammalian cells and rather than as agent impairing microfilament function should be considered as an inhibitor of Glut-1, having being used as such for studies on HTLV-1 and HTLV-2 entry,24 viruses that use Glut-1 as a receptor for virus entry. Interestingly, there is some sequence homology between HTLV-1 and HTLV-2 glycoproteins and filoviruses glycoprotein (see Figure 1). As cytokalasin B exerts its inhibitory effect by docking at the positively charged endofacial pocket 3 A of Glut-1,25 mutagenesis of this site should be tested and its influence on Ebola virus glycoprotein interactions and entry into the cell. Also in favor of direct glycoprotein interactions with Glut-1 are the facts that Phosphoinositide-3 kinase-Akt pathway controlls celular entry of Ebola virus26: inhibition of PI3K, Akt or Rac1 disrupted normal uptake of virus particles into cells; and cytokine stimulation promotes glucose uptake via phosphatidil inositol-3 kinase/Akt regulation of Glut-1 activity and trafficking27: Inihibition of mTOR/RAPTOR by ripamycin greatly diminished glucosa uptake suggesting Akt-stimulated mTOR/RAPTOR may promote Glut-1 transporter activity.\n\nOn the other hand, the integral protein stomatin, in the lipid rafts of the erythrocyte, regulates the switch from glucose to DHA transport, regulating the substrate preference of the transporter.16 Stomatocytosis, after which the protein stomatin is named, is a haemolytic condition with substantial intravascular haemolysis.\n\nTree views by the neighbor joining method of the aligned sequences (Blastp 2.2.28+). Fast minimum evolution. Max sequence difference, 0,85, Grishin distance.\n\nEbola virus has very broad cell tropism and it may bind to multiple attachment factors, among which are numerous lectins (DC-SIGN/L-SIGN, MGL, LSECtin and β1 integrins. Along with these attachment factors three main receptor candidates have been proposed as mediators for Ebola virus entry:\n\n- NPC1 was identified as an endosome-associated receptor for filovirus entry. This protein is required for filovirus entry and confers susceptibility to filovirus infection when expressed in non-permissive reptilian cells.28\n\n- The TAM receptor Axl, which has been proven to enhance Ebola virus particles macropinocytosis.29\n\n- TIM-1.30,31 It was published that T-cell Ig and mucin domain 1 (TIM-1) binds to the receptor binding domain of the Zaire Ebola virus (EBOV) glycoprotein, and ectopic TIM-1 expression in poorly permissive cells enhances EBOV infection by 10- to 30-fold.\n\nInterestingly, it has recently been published that TIM-1 mediates also a dystroglycan independent entry of Lassa virus.32 It has been pointed out that “a growing number of enveloped viruses are now appreciated to enter some cell populations through phosphatidylserine (PtdSer) receptor interactions”.33,34 The TIM receptors directly interact with PtdSer on apoptotic bodies or membrane-associated viruses,35–38 whereas the TAM receptors bind to one of two serum proteins, Gas6 or protein S, which, in turn, bind to PtdSer.39–41\n\nRegarding other virus associated with haemorrhagic manifestations, as is dengue virus (DENV), the same TIM and TAM families of phosphatidylserine receptors have been found to mediate dengue virus entry.42 Thus, TIM-1 has been recognized to enhance entry of Ebola virus, dengue virus and Lassa virus, that are characterized by the haemorrhagic manifestations in the illness they cause.\n\nAnd it has recently been found that TIM-3 expression decreases Glut 1 expression in Jurkat T cells at resting state and at an early time point of activation.43 TIM-1 is preferentially expressed on Th2 cells and has been identified as a stimulatory molecule for T-cell activation.44 TIM-3 is preferentially expressed on Th1 and Tc1 cells and functions as an inhibitory molecule, which mediates apoptosis of Th1 and Tc1 cells.45 TIM-4 is preferentially expressed on antigen-presenting cells, modulating the phagocytosis of apoptotic cells by interacting with phosphatidylserine (PS) exposed on apoptotic cell surface.35 In addition to TIM-1, TIM-4 has been shown to augment Ebola virus entry comparably to TIM-1.46\n\nA key point of the present opinion is that TIM-1, a host factor that has been found to be receptor for Lassa virus, dengue virus and Ebola virus, could increase the expression of Glut-1 and that there could be a link between glucose and ascorbate transport, viral interactions with TIM-1 and haemorrhagic manifestations of haemorrhagic fevers. Currently it has been described influence of TIM-3 on the expression of Glut 1 but not for TIM-1. On the other hand the mechanism responsible of the haemorrhagic manifestations of haemorrhagic fevers is an unclear matter and no general features have yet been described of a general pathophysiological mechanism. In relation with their natural function, as TIM-1 is a stimulatory molecule for cell activation and TIM-3 functions as an inhibitory molecule, and considering that TIM-3 has been found to decrease Glut 1 expression in Jurkat T cells, it is to expect that TIM-1, in the inverse direction as it does TIM-3, increases the expression of Glut-1. This should be matter of further experimental research. As the key factor TIM-1 would enhance the expression of Glut-1 and the glucose and ascorbate transport, viral interferences with TIM-1 could disturb its function and disturb the expression of Glut-1 or disturb the ascorbate and glucose transport. This would imply a connection between Glut-1, glucose and ascorbate transport, haemorrhagic manifestations, which are linked to oxidative stress, disruption of glucose and ascorbate homeostasis, and the role of TIM family members as mediators of viral entry in those viruses that cause haemorrhagic syndromes and haemorrhagic fevers. If direct interactions of filoviral glycoproteins with Glut-1 could be demonstrated, it could partially explain the differences in severity of the disease manifestations in Ebola and Lassa infections and others.\n\nOn the other hand, TAM receptors, as is Axl, have been found to enhance entry of Ebola and dengue virus and it has been recently confirmed that endogenous expression of Axl does not actually enhance viral entry of Lassa virus (LV) in the presence of fully functional alpha dystroglycan receptor (α-DG receptor) but it strongly augments viral infection in the absence of α-DG.47 And it has been described that Gas6–Axl receptor signalling is regulated by glucose in vascular smooth muscle cells.48 The direct or indirect interaction of Axl with Glut-1 could be hypothesized basing on this finding that the physiological function of Axl is regulated by glucose. Akt-mTOr promotes Glut-1 transporter activity and recycling and can prevent Glut-1 internalization27 and, according to Cavet and colleagues,48 glucose exerted powerful effects on Gas6–Axl signaling, with greater activation of Akt and mTOR in low glucose and greater activation of ERK 1/2 in high glucose. Akt-mTOr has been proven as key regulator of Zaire Ebola virus entry and activation of Ras-MAPK pathway, of which ERK1/2 is an activator, stimulates Ebola virus production from persistent infection.49 These findings add to the hypothesis of the pivotal role of glucose, transporters and plasma levels in Filovirus disease.\n\n\nConcluding remarks and future perspectives\n\nGlucose and ascorbate homeostasis, the presence of Glut-1 on erythrocytes and the erythrocyte physiology50 might play an important role in Filovirus disease, not excluding viral interactions with the erythrocyte membrane. The blood glucose concentration is maintained within narrow limits by an inter-play between tissue glucose uptake, hepatic glucose production and insulin production. Erythrocytes, because of their number in blood, perform an important buffering function of glucose and ascorbate/dehydroascorbate levels in plasma. Erythrocyte counts of frugivorous species have been shown lower than those reported for insectivorous bats.51 In those species not able to synthesize ascorbate and expressing Glut-1 on erythrocytes virus interactions would lead to severe disturbance of the glucose and ascorbate levels in plasma, activation of hypoxia-inducible factors and haemolysis. Coagulation abnormalities and haemorrhages are a clinical feature of Ebola virus haemorrhagic fever. It should also be tested the possible importance of haemolysis and erythrocyte-virus interactions. As it is shown in an important paper52 erythrocytes can regulate platelet reactivity directly throughchemical signalling and adhesive erythrocyte-platelet interactions. Cell free hemoglobin induces platelet aggregation contributing to high risk of thrombotic complications. Adhesion of abnormal and/or stimulated erythrocytes to vascular endothelium can contribute to vasculature occlusions associated with thrombosis. Efficient blood coagulation requires sufficient pro-thrombotic surfaces, which are provided by cells that expose phosphatidylserine, which is normaly in the cytoplasmic side of the membrane to separate this pro-coagulant surface from plasma coagulation factors. The questions are: Could virus-erythrocytes interactions induce the exposure of phosphatidylserine on the cell surface? Is the phosphatidilserine-enriched envelope of some haemorrhagic fever-causing virions involved in this pro-thrombotic process mediated by exposed phosphatidylserine?\n\nAs the filovirus glycoproteins could interact with Glut-1 or other functionally related protein, the influx of glucose into the cells is severely impaired. As a consequence, transient hyperglycemia and a marked oxidative stress coupled with the high levels of glucose in plasma would be established, and then promote the activation of NF–κB transcription, exacerbating a pro-inflammatory response mediated by cytokines and chemokines, as described by Katherine Esposito et al.4 On the other hand, the inability to synthesize ascorbate and the unavailability of ascorbate to entry into the cells through Glut-1 is an Achilles Heel when trying to counteract the oxidative stress. Additionally, there is another pathophysiological process to be mentioned: interactions with Glut-1 or other functionally related protein lead to an imbalance in the glucose homeostasis and the availability of glucose, not only for the erythrocytes, but also for CD8+ and CD4+ cells. These phenomena result in the bystander apoptosis of lymphocytes mediated by the lack of glucose, as described by Maclver et al.53 Activated T cells have dramatically increased metabolic requirements to support their demands.54 Therefore, activation of T cells causes a large increase in Glut-1 expression and surface localization, also promoting the Rab11b recycling of Glut-1 intracellular pools.53 A work on the mechanisms and consequences of Ebolavirus-induced lymphocyte apoptosis concluded undefined that Ebola virus induces multiple pro-apoptotic stimuli.55 As identified by Zhao and colleagues56 in 2007 there is a glucose-initiated signaling pathway that leads to inhibition of GSK-3 and prevents cell death through stabilization of the anti-apoptotic BcI2 family protein McI. Normally targeted for proteasomal degradation by the ubiquitin E3 ligases, in highly glycolytic cells as are cancer cells and activated T cells, McI remains unphosphorylated and is not degraded, increasing the threshold for cell-death and maintaining cell survival. Conversely, when glucose uptake is limited, glycolytic flux decreases to a level that no longer sustains viability and pro-apoptotic signals promote cell death.53\n\nAt a strictly cellular level, cell–adhesion dependent membrane trafficking of a binding partner for the Ebola virus glycoprotein has been shown to be a determinant of viral entry57 and attachment to adhesion substratum has been shown to induce the accumulation of glucose transporters and stimulate glucose metabolism in PC12 cells.58 According to Dube and colleagues a membrane trafficking event translocates the unknown binding partner of the receptor binding region (RBR) of Ebola glycoprotein (GP) to the cell surface and they identified two adherent primate lymphocytic cell lines that bind viral glycoprotein RBR at their surface and supported GP-mediated entry and infection. Lymphocytes are normally described in literature as non-permissive for Ebola-GP mediated entry and infection,59 and activation of lymphocytes could be the trigger that renders them susceptible. According to the present hypothesis, this binding partner related to lymphocyte activation may possibly be or be related to Glut-1.\n\nThis first clue about a link between Ebola virus species range, expression of Glut-1 on erythrocytes and inability to synthesize ascorbate in humans and non-human primates should be further analyzed. In contrast to primates, and according to a study by Cui et al., bats are perhaps in the process of large-scale loss of ascorbate biosynthesis ability,60 and show varying degrees of lack of gluconolactone oxidase function. This evolutionary recent loss of the ability to synthesize ascorbate could account for differences in Glut-1 expression and ascorbate metabolism resulting in differences in filovirus pathophysiology between species. In another study the same authors showed that the fruit bat Rousettus leschenaultii has retained the ability to synthesize ascorbate although at low levels compared with the mouse.61 Olival et al. found serological evidence to both species Reston and Zaire of the genus Ebolavirus in Rousettus leschenaultii, one of the fruit bats species considered to be reservoir of Ebola virus,62 and Marburgvirus has even been isolated from Rousettus aegyptiacus,63 another Pteropodidae member. It is known that frugivorous and nectarivorous bats are able to ingest large quantities of sugar in a short time span while avoiding the potentially adverse side-effects of elevated blood glucose, which could be an important fact supporting the hypothesis that glucose and ascorbate metabolism account for the severity of the haemorrhagic fevers in primates and for the role of fruit bats as reservoir species maintaining low levels of filovirus replication. This ability to ingest large quantities of sugar in a short time span while avoiding the potentially adverse side-effects of elevated blood glucose has been related to the adaptive evolution in the glucose transporter 4 (Glut-4) gene in skeletal muscle64 and also to high passive paracellular absorption of glucose in the gut of Rousettus aegyptiacus.65 The importance of Glut-4 and the active sodium-ascorbate co-transporters (SVCTs) and active sodium-dependent glucose cotransporters (SGLTs) in regard to Filovirus pathophysiology should be further investigated.\n\nRegarding Lassa fever virus, the differences in glucose and ascorbate physiology could be an explanation of the absence of haemorrhagic manifestations of the illness in mice while primates and humans develop haemorrhagic fevers. If direct interactions of filoviral glycoproteins with Glut-1 could be demonstrated, it could partialy explain the differences in severity of the disease manifestations between Ebola and Lassa infections and others. It has been shown that the Ebola virus glycoprotein is the main determinant of cell cytotoxicity and injury.66 It should be further researched which role plays the viral glycoprotein in cell cytotoxicity for Lassa and Dengue viruses. The differences in haemorrhagic fever prevalence and severity between Ebola, Lassa and Dengue viruses could be explained by the differences in the pools of cellular factors and receptors involved in virus activity. Direct interactions with Glut-1 could explain the direct incidence of the Ebola virus on the glucose and ascorbate transport and homeostasis, causing the transient hyperglycemia, cytokine storming and haemorrhagic features. On the other hand, TIM-1 and Axl are entry factors for Lassa virus only under specific circumstances: in other words, are only a dystroglycan-independent entry route. The concentration of phosphatidylserine displayed in the viral envelope influences virus binding via Axl, as is described in another important paper.67 According to these authors, since functional α-dystroglycan is recognized by the viral envelope protein GP1 of Lassa virus, the exact ratio of Lassa GP1 to phosphatidylserine in the viral envelope appears critical for receptor use. When the presence of phosphatidylserine on the virion surface is qualitatively important in relation to the expression of GP1 in Lassa virus, the cellular entry of the virus through the interaction phosphatidylserine-Gas6-Axl or phosphatidylserine-TIM-1 would lead to the hypothesized metabolic disturbances, mediated by TIM-1 and/or Axl, probably via Glut-1. When Axl or TIM-1 are not an important receptor for Lassa virus entry and entry occurs via interaction of the viral GP1 with α-dystroglycan, the incidence of metabolic disorders and haemorrhagic manifestations would be lower.\n\nFor Dengue virus, when virus-host interactions mainly take place via viral glycoprotein-DC-SIGN interaction, the metabolic glucose and ascorbate disorders and haemorrhagic manifestations would be less important than in those cases in which viral interactions with host factors involve the interaction of the viral glycoprotein with Axl and/or TIM-1. These different host factors that can interact with the viral glycoproteins would explain the differences in pathophysiology: normal Dengue fever when DC-SIGN-viral glycoprotein interactions are predominant, or Dengue severe haemorrhagic fever when Axl and/or TIM-1 interact predominantly with the dengue virus glycoprotein. The presence and importance of phosphatidylserine in the virus envelope could vary from virus to virus and from one virus type or strain (DENV-1, DENV-2, DENV-3, DENV-4, LV-1, LV-2, LFV-3, L V-4) to another. This should be matter of further research. It would be interesting to know which factors are governing the significant presence of phosphatidylserine on the virion envelope, and this considering the four haemorrhagic viruses: Ebola virus, Marburg virus, Lassa virus and Dengue virus.\n\nPathogenesis of viral haemorrhagic fevers is not yet well understood. The interaction of viruses with metabolism is a matter that will gain more attention in the next years. If some assumptions hypothesized in this opinion article were correct, regardless of other means of challenging the viruses, supportive care of haemorrhagic fevers almost in general could be significantly improved. It encompasses the use of insulin, ascorbate, glutathione (which participates, along with NADH, in the reduction of dehydroascorbate to ascorbate), maintaining constant and normal levels of glucose in plasma and general measures to counteract oxidative stress.", "appendix": "Data availability\n\nNo data are associated with this article.\n\n\nReferences\n\nAguirre R, May JM: Inflammation in the Vascular Bed: Importance of Vitamin C. Pharmacol. Ther. 2008; 119(1): 96–103. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRao K, Chouhan V, Chen X, et al.: Activation of the tissue factor pathway of blood coagulation during prolonged hyperglycemia in young healthy men. Diabetes. 1999; 48: 1156–1161. PubMed Abstract \| Publisher Full Text\n\nIwasaki Y, Kambayashi M, Asai M, et al.: High glucose alone, as well as in combination with proinflammatory cytokines, stimulates nuclear factor kappa-B-mediated transcription in hepatocyte sin vitro. J. Diabetes Complicat. 2007; 21(1): 56–62. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKondratowicz AS, Lennemann NJ, Sinn PL, et al.: T-cell immunoglobulin and mucin domain 1 (TIM-1) is a receptor for Zaire Ebolavirus and Lake Victoria Marburgvirus. Proc. Natl. Acad. Sci. U. S. A. 2011; 108(20): 8426–8431. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBrunton B, Rogers K, Phillips EK, et al.: TIM-1 serves as a receptor for Ebola virus in vivo, enhancing viremia and pathogenesis. PLoS Negl. Trop. Dis. 2019; 13(6): e0006983. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBrouillette RB, Phillips EK, Patel R, et al.: TIM-1 mediates dystroglycan-independent entry of Lassa virus. J. Virol. 2018; 92(16): e00093–e00018. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMoller-Tank S, Kondratowicz AS, Davey RA, et al.: Role of the phosphatidylserine receptor TIM-1 in enveloped-virus entry. J. Virol. 2013; 87: 8327–8341. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMoller-Tank S, Albritton LM, Rennert PD, et al.: Characterizing functional domains for TIM-mediated enveloped virus entry. J. Virol. 2014; 88: 6702–6713. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKobayashi N, Karisola P, Pena-Cruz V, et al.: TIM-1 and TIM-4 glycoproteins bind phosphatidylserine and mediate uptake of apoptotic cells. Immunity. 2007; 27: 927–940. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMiyanishi M, Tada K, Koike M, et al.: Identification of Tim4 as a phosphatidylserine receptor. Nature. 2007; 450: 435–439. PubMed Abstract \| Publisher Full Text\n\nJemielity S, Wang JJ, Chan YK, et al.: TIM-family proteins promote infection of multiple enveloped viruses through virion-associated phosphatidylserine. PLoS Pathog. 2013; 9: e1003232. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nIchimura T, Asseldonk EJ, Humphreys BD, et al.: Kidney injury molecule-1 is a phosphatidylserine receptor that confers a phagocytic phenotype on epithelial cells. J. Clin. Invest. 2008; 118: 1657–1668. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nStitt TN, Conn G, Gore M, et al.: The anticoagulation factor protein S and its relative, Gas6, are ligands for the Tyro 3/Axl family of receptor tyrosine kinases. Cell. 1995; 80: 661–670. PubMed Abstract \| Publisher Full Text\n\nGodowski PJ, Mark MR, Chen J, et al.: Reevaluation of the roles of protein S and Gas6 as ligands for the receptor tyrosine kinase Rse/Tyro 3. Cell. 1995; 82: 355–358. PubMed Abstract \| Publisher Full Text\n\nNagata K, Ohashi K, Nakano T, et al.: Identification of the product of growth arrest-specific gene 6 as a common ligand for Axl, Sky, and Mer receptor tyrosine kinases. J. Biol. Chem. 1996; 271: 30022–30027. PubMed Abstract \| Publisher Full Text\n\nMeertens L, Carnec X, Lecoin MP, et al.: The TIM and TAM families of phosphatidylserine receptors mediate dengue virus entry. Cell Host Microbe. 2012; 12(4): 544–557. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLee MJ, Yun SJ, Lee B, et al.: Association of TIM-3 expression with glucose metabolism in Jurkat T cells. BMC Immunol. 2020; 21: 48. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nUmetsu SE, Lee WL, McIntire JJ, et al.: TIM-1 induces T cell activation and inhibits the development of peripheral tolerance. Nat. Immunol. 2005; 6(5): 447–454. PubMed Abstract \| Publisher Full Text\n\nZhu C, Anderson AC, Schubart A, et al.: The Tim-3 ligand galectin-9 negatively regulates T helper type 1 immunity. Nat. Immunol. 2005; 6(12): 1245–1252. 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PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "339493", "date": "29 Nov 2024", "name": "Hong-Li Liu", "expertise": [ "Reviewer Expertise infectious disease", "HFRS", "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\n1. Rational use of punctuation to simplify sentence structure which makes it easy for readers to get the point of the paper; 2. What is the most important purpose of this paper? The viral hemorrhagic fever or filovirus-related hemorrhagic fever? If the purpose is the latter, what is filovirus？which viruses are included in filovirus? it will be better for the author to make an introduction; 3. The title talks about the relationship between the TIM-1, TAM, Glut-1, and the manifestation of viral hemorrhagic fever, and the Lassa virus, and dengue virus were mentioned in the text. If the authors want to present the relationship between these receptors and viral-related hemorrhagic fever. Actually, the Hantavirus is also a major agent of hemorrhagic fever that needs to pay attention; 4. As the authors pointed out \"the manifestations of thrombotic microangiopathic hemolytic anemia were similar to those of filovirus hemorrhagic fever\", why did the authors mention this content? Or the TIM, Glut-1 is associated with this disorder? 5. The concluding remarks should be precise and clear. The conclusion part is a summary but not a stacking of references or it will be hard for readers to get the main point of the paper; 6. It is necessary for authors to reconstruct the manuscript. From the main topic, the introduction of receptor-related biological functions, the viruses that can cause hemorrhagic fever, and the relationship between these two parts involving the possible pathways or mechanisms, and then conclude that the pivotal role of these receptors in the viral-related hemorrhagic fever and their possible value in the treatment of these diseases.\n\nIs the topic of the opinion article discussed accurately in the context of the current literature? Partly\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? Partly", "responses": [ { "c_id": "12995", "date": "20 Dec 2024", "name": "Ivan Chicano Wust", "role": "Author Response", "response": "Thank you for giving me the opportunity to improve the last version of the article." } ] }, { "id": "339488", "date": "29 Nov 2024", "name": "Chao Qin", "expertise": [ "Reviewer Expertise Metabolism", "Viral infection", "Immunology" ], "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 manuscript described a the correlation of metabolic receptors and virus induced diseases. It is a good topic to discuss. I would like the authors to address following issues in the manuscript. 1. Please use more condensed languages for the statements, as well as the titles and subtitles. 2. Please spell out the acronym at the first time of using it. 3. The original function of metabolic receptors such as GLUT1 should be explained in more details, as well as the role of glycolysis in filovirus infection. This will highlight the unique role GLUT1 in viral infection.\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": "12994", "date": "23 Dec 2024", "name": "Ivan Chicano Wust", "role": "Author Response", "response": "Thank you for appreciating my effort and giving me some clues to improve the article." } ] } ]	5	https://f1000research.com/articles/12-518
https://f1000research.com/articles/13-1510/v1	09 Dec 24	{ "type": "Systematic Review", "title": "Natural disaster and medication preparedness among elderly: a scoping review", "authors": [ "Rika Yuliwulandari", "Rifda El Mahroos", "Zulfan Febriawan", "Adi Wibowo", "Debrina Kusuma Devi", "Johan Danu Prasetya", "Tedy Agung Cahyadi", "Eko Teguh Paripurno", "Reza Pahlevi Ramadhani Arfindra Setiawan", "Hafiz T.A Khan", "Rifda El Mahroos", "Zulfan Febriawan", "Adi Wibowo", "Debrina Kusuma Devi", "Johan Danu Prasetya", "Tedy Agung Cahyadi", "Eko Teguh Paripurno", "Reza Pahlevi Ramadhani Arfindra Setiawan", "Hafiz T.A Khan" ], "abstract": "Introduction The increasing number of older people and their vulnerability to disaster and medication preparedness as the primary elements of disaster mitigation are necessary to reduce the impact of a disaster. Nevertheless, research on natural disasters and medication preparedness in the elderly population is still lacking. This review aimed to explore all reported research and publications related to natural disasters and medication preparedness among the elderly.\n\nMethods A scoping review was performed using the Preferred Reporting Items for Systematic Reviews and Meta-Analysis Extension for Scoping Reviews (PRISMA-ScR). A literature search was conducted using the following four databases: Scopus, PubMed, Sage, and Google Scholar. Screening was conducted using the following criteria: articles written in English, open access, and published between 2020 and 2024. Articles must discuss natural disasters and medication preparedness. In the data search, we input several keywords that include “elderly,” “natural disaster,” “preparedness,” and “medication.” Snowballing was then conducted to find articles on preparedness interventions. Data extraction and analysis were then performed.\n\nResults and conclusion The findings from this literature review are the majority of studies showing that most elderly people are not well prepared in facing disasters; however, through various programs that have been implemented by either the government or community, the elderly show more preparation when they encounter any natural disaster.", "keywords": [ "Elderly", "Health", "Medication", "Natural Disaster", "Preparedness" ], "content": "Introduction\n\nAging is a phase of the life cycle encountered by everyone that result in a group of population classified as elderly.1 Globally, the elderly population is the one with aged 60 years old and older. The number of people in the population is increasing, with a total of 1 billion people or 13.2% of the world’s population. This number will increase to 1.4 billion by 2030 and 2.1 billion by 2050.2 Elderly people experience many physiological changes in various organs that make them become vulnerable to health problems and may decreased quality of life (QoL).3 These circumstances can affect their safety and capacity to understand complex medical information.4\n\nThe decline in body function and health status of the elderly often becomes an obstacle for them in dealing with emergencies, natural disasters and disease outbreaks that makes them classified as the most vulnerable population group.5 That is because they have reduced ability to prepare, respond and deal with disasters, therefore, they may be more susceptible to injury, death or health problems during and post-disaster.6 The key component of effective disaster mitigation is preparedness. Disaster and medication preparedness is a structured process that includes administrative, organizational, and operational skills to enact strategies and policies, increase management capacity to mitigate the adverse effects of hazards, and decrease the chance of emergencies or disaster effects. Disaster and medication preparedness require collaboration between various stakeholders involving the government, society, and individuals.7\n\nMinistries of health are the areas of government primarily responsible for disaster-related public health and safety, but they often emphasize the entire community and may not always meet the needs of older adults.8 Therefore, this article offers a comprehensive review of existing literatures encompassing “elderly natural disaster and medication preparedness.” This review aims to explore disaster and medication preparedness among the elderly, including what the elderly need to prepare in facing disaster, as well as the factors that inhibit them from fulfilling the preparedness and certain interventions to support them to be more prepared. To the best of our knowledge, this literature review is the first to examine disaster preparedness among the elderly preparedness towards disaster. Hopefully, it can provide important information for policymakers and for any further research.\n\n\nMethods\n\nThe literature review was conducted following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses Extension for Scoping Reviews (PRISMA-ScR) guidelines. Relevant studies were identified using four scientific databases: PubMed (National Center for Biotechnology Information, National Institutes of Health; Bethesda, Maryland, USA); SCOPUS (Elsevier, Amsterdam, Netherlands), Sage Journals (Thousand Oaks, California, USA), and Google Scholar. The searches were conducted using a limited timeframe from 2020 to April 2024. The search strategy was performed using a blend of search terms primarily located within titles (Figure 1; Table 1) (extended data). Two reviewers collaborated to develop an approach to search for and determine a suitable determination for the search.\n\nThe screening was conducted in three stages. Initially, two authors independently evaluated the articles based on their titles. The inclusion and exclusion criteria were set based on article type, publication year, access type, language, tier, subject, and age of the participants, as shown in Table 2 (extended data). Papers that did not meet the inclusion criteria were excluded. In the second stage, the abstracts of the selected papers were reviewed. The abstract must contain clear objectives, methods, and results, and also discuss disaster and medication preparedness in the elderly. In the third stage, the full texts of the selected papers were evaluated. The full texts must reveal methodology, reports of the needs and obstacles of elderly people in disaster preparedness, as well as strategies to enhance disaster preparedness in the elderly that include intervention, education, and technology management. The process used to identify the relevant articles for review is shown in Figure 2 (extended data).\n\nSnowballing involves searching for references and/or citations from articles to identify additional relevant material. The inclusion criteria for this step were similar to those used in the previous step.\n\nA data extraction form was used to collect information on each study included in the analysis so that it could be systematically reviewed. The information in the form included the journal author, year of publication, country of origin, sample/sampling technique, research objectives, research design, instrumentation, and major findings. After extraction and structure, the results are discussed.\n\n\nResults\n\nA total of 920 titles were found across databases (PubMed, Scopus, Sage, and Google Scholar). The titles were then screened based on the inclusion criteria. Duplicated papers were removed, resulting in 163 articles (Figure 1) (extended data). Subsequently, all abstracts were reviewed to determine whether they contained the keywords natural disaster, medication, preparedness, and the elderly. This resulted in 45 articles for further full-text review. Only 14 articles fulfilled the inclusion criteria. Subsequently, snowballing was conducted to identify other relevant materials that contain interventions for disaster preparedness, resulting in six new articles. Thus, the total number of papers included in this study was 20. The final articles eligible for data extraction are listed in Table 3 (extended data).\n\nMost of the research (n=5) was conducted in the United States.9–13 The remainder originated from Indonesia,14–17 Japan,18,19 Thailand,20,21 Puerto Rico,22 Nepal,23 Korea,24 Hong Kong,25 Phillipines,26 and Iran27 (Table 4) (extended data). The journals targeted for this review article predominantly originated from the Disaster Medicine and Public Health Preparedness Journal (n=6). The remainder originated from the International Journal of Environmental Research and Public Health (n=4), BMC Geriatrics (n=1), Public Health Nursing (n=1), Jamba-Journal of Disaster Risk Studies (n=1), Health and Quality of Life Outcomes (n=1), Sustainability (n=1), Journal of Education and Health Promotion (n=1), Water (n=1), International Journal of Disaster Risk Reduction (n=1), Asia Pasific Journal of Multidisciplinary Research (n=1), and Jurnal Ners (n=1) (Table 3) (extended data).\n\n\nDiscussion\n\nNatural disasters such as earthquakes, floods, typhoons, tornadoes, tsunamis, landslides, forest fires, volcanic eruptions, and extreme temperatures cause great damage to the environment. Natural disasters are emergencies that require immediate intervention because of their large impact on human health and safety.28 Elderly people face several different challenges from other age groups when facing these emergencies. Elderly people may encounter challenges such as mobility issues or chronic health conditions during emergencies. Additionally, they may lack nearby family or friends who can offer support during such emergency situations.29 Elderly people have less disaster preparedness and greater vulnerability to disasters.8,9,18 About six out of ten elderly people do not consider themselves prepared to face disasters.12 The aims of this review were to explore all reported research and publications related to natural disasters and medication preparedness among the elderly. Among the 20 papers that met all selection criteria, all showed that none of the countries had good preparedness for the elderly when facing a disaster. A USA study by Bell et al. (2021) showed that most elderly people are more confident when facing disasters as long as they perform preparedness actions such as a 7-days supply of food and water, a 7-days supply of essential medications/health supplies, a stocked emergency kit, and a conversation about evacuation from home. According to Adepoju et al. (2023), six to ten older people in the USA do not perceive themselves as disaster preparedness. In addition, they also suggested the role of home-based providers in improving preparedness and the response of the elderly to disasters. A study in Thailand by Boonyaratkalin (2020) showed that 50% of older people exhibit suitable behavior through follow-up news on floods and evacuation planning. Based on a study by Hattori (2020), older people are ready to evacuate, especially on confirmed evacuation sites and routes and emergency contact plans; community preparedness, especially on talking with family members about how to evacuate; family preparedness, especially related to fire extinguishers; preparation of emergency goods, especially those related to water, non-perishable foods, and flashlights. However, they are unwell prepared for talking with neighbors on how to evacuate, measures to prevent large electrical applications from moving or falling, footwear for evacuation in the bedroom, portable radio, medical prescription records, medications, and written memos with emergency contact information. In Indonesia, so far, there is not enough data on disaster preparedness in the elderly published in international journals.\n\nSeveral studies have shown that the majority of the elderly population is not well-prepared for disasters. Therefore, the government, community, and organizations related to disasters have responsibilities to educate elderly people to prepare their needs for disasters, including the following:\n\nThe elderly population is vulnerable when a disaster occurs. Many of them have chronic health problems and are highly dependent on drugs and medical equipment. Elderly people with chronic conditions require treatment to survive. In the event of a disaster, it is especially important for elderly people with chronic conditions or other health needs to carry their medicines and medical equipment to reduce health risks and facilitate continuity of their healthcare. The most challenging aspect of the elderly during a disaster is ensuring that they bring all the necessary medications, including medical records, emergency medications, and other supportive equipment. Most elderly people did not bring their prescriptions and often could not remember their medical history, names of medications, or dosages of their medicine. Medical records are essential to understand the history of illnesses and treatments. For example, patients with diabetes require insulin or oral hypoglycemic drugs, asthma patients require beta-antagonists or corticosteroids, hypertension patients require antihypertensives, heart disease patients need beta-blockers or nitroglycerin preparations, and dialysis patients need potassium-exchanging resins, which is crucial for reducing potassium levels when dialysis access is limited. Over-the-counter medications such as fever or pain relievers, antihistamines, and sanitary products are also necessary. Supportive equipment is required, such as insulin injection supplies (vials, needles, or pens with replaceable insulin cartridges), glucometers, oxygen cylinders, and nebulizers. These should be considered part of the emergency kit to be taken along. Daily living aids should also be prepared, including wheelchairs, hearing aids, canes, walkers, dentures, glasses, and incontinence briefs. Disasters can disrupt access to healthcare facilities and pharmacies, making it challenging for the elderly to obtain essential medications and supplies. Disaster preparedness efforts should include measures to ensure continuous access to medication and medical supplies during emergencies. This may involve stockpiling medications, establishing mobile medical clinics, and providing home delivery services for elderly individuals who are unable to visit pharmacies.10,11,25\n\nMobility assistance is one of the primary disaster preparedness needs of the elderly. Many elderly individuals experience reduced mobility owing to age-related physical limitations or chronic health conditions. During disasters such as hurricanes or wildfires, evacuation may be necessary to ensure safety. However, elderly individuals may face a challenging evacuation owing to mobility issues. Providing assistance with transportation and ensuring accessible evacuation routes are essential to address this need. Communities can establish transportation assistance programs and designate accessible evacuation routes to support elderly individuals in safe evacuation during emergencies.11\n\nCommunication support is essential to address disaster preparedness among the elderly. Elderly individuals may have hearing or vision impairments, which make it difficult for them to receive emergency alerts and information. Clear and accessible communication channels are necessary to ensure that the elderly receive timely information and instructions during emergencies. Communities can implement text-based alert systems, use visual cues in emergency communication, and provide outreach programs to support elderly individuals with communication challenges.18\n\nSocial support networks are essential for the preparation of older adults for disasters. Building strong social support networks within communities can help ensure that elderly individuals have access to assistance and resources before, during, and after a disaster. Programs that connect elderly individuals with volunteers who can provide assistance with evacuation, medical care, and other needs can help enhance resilience and reduce vulnerability during emergencies.18,24,25\n\nThe establishment of specialized shelters and facilities is essential to address the specific requirements of the elderly during disasters. Designating shelters and facilities equipped to accommodate elderly individuals, such as providing accessible accommodations, medical support services, and trained staff, can help ensure their safety and well-being during emergencies. These specialized facilities can provide a supportive environment for elderly individuals, who may require additional assistance or medical care during disasters.11,25\n\nEducation and training programs tailored to the needs of the elderly are essential to enhance disaster preparedness. Providing education on evacuation procedures, emergency preparedness kits, and strategies for managing chronic health conditions during emergencies can empower elderly individuals to take proactive steps in preparing for disasters. Community-based training programs can help ensure that elderly individuals have the knowledge and skills needed to protect themselves and their loved ones during emergencies.22\n\nBeside that there are several factors that affect disaster preparedness in elderly such as:\n\nEducation has greatly influenced disaster preparedness among elderly people. Many elderly people have low literacy levels and some of them cannot write, so they do not understand how to get information about disasters and help. Thus, education level serves as a barrier to disaster preparedness.20\n\nHearing ability is also related to disaster preparedness literacy. In the study conducted by Bell et al. (2021b), participants with hearing impairments followed flood news by utilizing alternative methods such as visual alerts, text notifications, or assistance from others to access weather forecasts and flood warning announcements. Older people with hearing impairments cannot listen to the information being disseminated. This can affect the preparedness. However, no research has been conducted on the intersection of hearing abilities and disaster preparedness.\n\nIncome plays a significant role in enabling elderly people to prepare for disasters. If they have no or a low income, they cannot meet their needs and cannot survive. Low financial resources make them unsure whether they will be capable of covering food and shelter expenses for at least one day during an evacuation. Based on research conducted by Adepoju et al. (2023), a survey on survival disaster preparedness action and planning disaster preparedness action indicates that low-income elderly individuals tend to neglect survival actions. Meanwhile, the findings found by Fletcher et al. (2022) showed that after experiencing disasters, the elderly population experienced worry and stress. This is because of low financial resources. They are not sure if they will be able to pay for food and shelter for one day in the event of an evacuation. According to Bell et al. (2021a), the elderly acknowledged that their options were unevacuated because of a lack of alternative housing options, financial constraints preventing temporary accommodation, and discomfort associated with evacuation shelters.\n\nCommon challenges encountered in emergency response include barriers to communication technologies. This communication technology can also be useful as a disaster-warning system. The lack of a disaster warning system will have a major impact. A case reported by Yulianto et al. (2020) during the Palu earthquake disaster showed that impacted individuals did not receive any advance warning about the tsunami or liquefaction. Although some people received tsunami alerts through social media, they came too late for an effective response. Thus, the Early Warning System (EWS) holds significance in initial preparedness, potentially safeguarding lives, infrastructure, and public facilities from disaster impact. This proves that a warning system in Indonesia is still lacking.\n\nAnother issue that serves as a barrier is the lack of disaster training for the elderly. Insufficient training hinders local communities from voluntarily engaging in disaster risk management. The role of local governments in reaching out to the elderly through training or education on disasters is essential; otherwise, residents may underestimate the threat of disasters. Furthermore, communicating disaster risks to vulnerable elderly populations is often ineffective due to barriers related to literacy and age-related impairments, such as hearing and vision difficulties.21\n\nAccording to Bell et al. (2021b), the elderly admitted that most information regarding communication about disaster preparedness and available response resources often lacks relevance and simplicity. Despite the widespread use of smartphone apps, social media, and other online communication by various institutions and organizations for rapid information dissemination, these channels remain inaccessible to many seniors, especially those lacking internet access or mobile devices. This unequal distribution of disaster response resources contributes to prolonged and difficult recovery of vulnerable elderly individuals.\n\nFamilies play a pivotal role in providing primary support for older adults, aiding in disaster preparation and recovery. The family is the support of the elderly because the family is the one that provides shelter and other resources when elderly facing a disaster. Family members may also invite home care providers to offer formal caregiving support when they cannot physically reach the healthcare service. Family members provide communication between patients and healthcare providers, maintain continuity of care, and encourage essential medication adherence.13\n\nAccess to healthcare facilities depends on factors such as distance, mobility facilitators and barriers, transportation options, road conditions, travel expenses, and other associated direct and indirect expenses, all of which are designed as inhibitor factors for the utilization of healthcare services. Elderly individuals who are injured and suffer from chronic diseases require support and assistance to access healthcare services. Elderly people often face difficulties in reaching healthcare services owing to poor road conditions and injuries sustained during disasters. However, participants without significant injuries reported accessing post-earthquake health care services through mobile clinics, hospitals, and health posts. Many participants had access to hospitals and health posts for basic health checkups and treatment. Nonetheless, for those who sustain injuries and experience mobility loss due to an earthquake, access to intensive care facilities located within a feasible distance becomes essential.23\n\nTo improve the preparedness of the elderly to disaster, some interventions are needed that include the following:\n\nOne of the primary challenges in disaster management involves fulfilling the food requirements of vulnerable groups to meet their nutritional needs. In areas affected by disasters, the elderly more often experience malnutrition due to low energy, carbohydrate, protein, and fat intake. Fatmah et al. (2021) attempted to create a food bar and recipe. This food bar is an emergency sustenance item made from locally obtained raw materials, engineered to be readily accessible while possessing increased economic worth owing to its specialized composition. Consuming 50 g of the broccoli-soybean-mangrove food bar per day (amounting to 246 calories) satisfies 14–16% of the daily caloric needs for older males and females, respectively, based on a recommended intake of 1800 calories for males and 1550 calories for females is enough for elderly. Therefore, this food bar serves as a viable alternative to emergency food options during disasters. Some dissatisfied perceptions of elderly disaster victims are that they think that the food bar is a staple food and not the main food. Beside that, when the food bar was tested, the researcher has not done the impact of the food to immune level of elderly. In addition, there is a need to fortify food with iron and prebiotics to provide more benefits to the health of the elderly.\n\nSeveral researchers have conducted studies to improve disaster preparedness. Matsuo et al. (2021) investigated the significance of disaster preparedness in mitigating isolation among elderly people. Elderly people in Japan prefer to isolate themselves when a disaster occurs rather than asking neighbors or other people for help. Therefore, they created a handbook, the “Disaster Preparedness Handbook,” and conducted a survey where the results indicated that the elderly population had a higher level of isolation precaution for themselves compared to the level of isolation precaution for others. Thus, education on disaster preparedness is needed to prevent isolation among the elderly. However, the study is limited by the age group selection and the closeness of the neighbors and the type of community of the participants, which makes a significant difference in the individual characteristics of the intervention and comparison groups who participated in the study.30\n\nAnother study by Fatmah (2022) in Indonesia provides education regarding the meaning of disasters, types of disasters, the impact of disasters, family actions before, during, and after disasters, and disaster prevention. After education, family knowledge has increased. Some families have prepared equipment such as emergency bags containing important documents, logistical supplies (e.g., food), and medication. There are limitations to this study, in which a control group was not included for comparison with the intervention group, in addition to its small sample size.15\n\nAccording to Yarmohammadian et al. (2023), it is necessary to educate the CBHO (community-based health organizations) through the development of a curriculum/syllabus for CBHO training in serving the elderly when facing disasters. The knowledge provided by the CBHO is in accordance with the syllabus, which includes the concept of aging and analysis of the impact of disasters on the elderly, providing for the needs of the elderly, physical and mental analysis, how to coordinate with special organizations to handle the elderly, and the threats faced by the elderly after a disaster.\n\nLuna & Pingol (2020) developed new technologies for help during disaster and emergencies namely “Help Bro.” This rescue application can serve government agencies, such as the Red Cross. Users can use the “Help Bro” application to alert others about impending dangers and help them avoid hazardous areas. There are several pages available in the application, for example, a page for registration for the user to enter personal information; a help page that enables the application to identify the user’s location and generate a report when a user requests assistance; and a user report page that allows the users to send requests for help to the rescue team. The rescue team’s address is displayed along with options to specify the type of assistance needed.\n\nNakai et al. (2022) developed tools (K-DiPS) to help vulnerable people provide medical information for use in disasters. K-DiPS comprises two components: K-DiPS Solo, a smartphone application, and K-DiPS online, a web application specifically designed for disaster management purposes by local government authorities. K-DiPS Solo is used by filling in the personal information of medically vulnerable people (MVP), and local governments can access this application via K-DiPS online. The K-DiPS Solo includes essential information such as personal details (name, gender, date of birth), emergency contact, medical history, need for long-term care, support (doctor, home visit, etc.), activities of daily living (ADL), and support needed (meal, excretion, dressing, transferring, mobility, bathing, etc.). Thus, when an MVP needs help, the local government can provide necessary health items (medicines and medical equipment).\n\nFathoni et al. (2020) through his research invented an application named “application Bromo alert.” The application has several features: weblogs that offer information about Mount Bromo, including its history and past eruption events; shelters that provide details about shelters, including building names, locations, and proximity to disaster-prone areas; healthcare facilities that contain information about healthcare facilities available in the area; vlogs that provide informative content, especially targeted towards foreign tourists; and CCTV, which allows the public to view live updates of current conditions, particularly the latest situation at Mount Bromo.20\n\n\nConclusions\n\nThe findings of this study need to be prepared to support disaster preparedness for the elderly (medication, mobility assistance, communication support, social support, shelter, education, and training). However, several factors can hinder disaster preparedness, including education, income, disability, communication and information technology, family, and access to health services. To improve disaster preparedness among the elderly, discoveries have been made in emergency food, educational programs, health technology, support, and warning systems. To the best of our knowledge, this literature review is the first to examine disaster preparedness among the elderly. Hopefully, it can provide important information for policymakers and for any further research.", "appendix": "Data availability statement\n\nNo data are associated with this article.\n\nFigshare: Natural Disaster and Medication Preparedness Among Elderly (DOI https://doi.org/10.6084/m9.figshare.27292446.v3)\n\nThis project contains the following underlying data:\n\n• Figure 1 Search Keywords\n\n• Figure 2 Flow Diagram of Review Process\n\n• Table 1 Key Search Terms\n\n• Table 2 Inclusion Criteria Based on Scientific Database Used\n\n• Table 3 List of Reviewed Name Journal\n\n• Table 4 Result Study\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nFigshare: Prisma Checklist for ‘Natural disaster and medication preparedness among elderly’ (DOI https://doi.org/10.6084/m9.figshare.27252945.v7)\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 express our gratitude to all those who contributed to the writing of this article and have proofread the manuscript.\n\n\nReferences\n\nEstebsari F, Dastoorpoor M, Khalifehkandi ZR, et al.: The concept of successful aging: a review article. 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Publisher Full Text\n\nEngelman A, Guzzardo MT, Muñiz MA, et al.: Assessing the emergency response role of community-based organizations (CBOs) serving people with disabilities and older adults in Puerto Rico Post-Hurricane María and during the COVID-19 pandemic. Int. J. Environ. Res. Public Health. 2022; 19(4): 1–21. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWagle S, Amnatsatsue K, Adhikari B, et al.: Health-related quality of life after the 2015 Gorkha Earthquakes, among older adults living in Lalitpur District of Central Nepal. Disaster Med. Public Health Prep. 2021; 15(3): 298–307. PubMed Abstract \| Publisher Full Text\n\nKim EM, Kim GS, Kim H, et al.: Health-related quality of life among older adults who experienced the Pohang earthquake in South Korea: A cross-sectional survey. Health Qual. Life Outcomes. 2022; 20(1): 11–37. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKwan C, Tam HC: Ageing in place in disaster prone rural coastal communities: a case study of Tai O village in Hong Kong. Sustain. 2021; 13(9). Publisher Full Text\n\nDe Luna JNN , Pingol MFQ: HELP BRO: A mobile application for calamities and disaster. Asia Pac. J. Multidiscip. Res. 2020; 8(1): 106–111. Reference Source\n\nYarmohammadian MH, Akbari F, Niaraees A, et al.: Elders in natural disaster: community-based health organization (CBHO) education and preparedness. J. Educ. Health Promot. 2023; 12(153): 1–6. Publisher Full Text\n\nCDC: Natural Disasters and Severe Weather.2024. Reference Source\n\nAmerican Red Cross: Emergency preparedness for older adults.2024. Reference Source\n\nMatsuo Y, Hirano M: Effectiveness of disaster preparedness education in helping older people prevent isolation. Public Health Nurs. 2021; 38(5): 837–849. PubMed Abstract \| Publisher Full Text" }	[ { "id": "351500", "date": "06 Jan 2025", "name": "Elena Valkanova", "expertise": [ "Reviewer Expertise Disaster 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\nThe article “Natural disaster and medication preparedness among elderly: a scoping review” aims to explore disaster and medication preparedness among the elderly, including the prerequisites and the impediments to disaster preparedness as well as interventions to support their preparedness. This is highly relevant topic considering the growing disaster risk associated with climate change and the vulnerability of the studied population. The authors performed a literature review of original research articles on the topic. 20 eligible articles were identified and further analyzed. The main conclusion states that more efforts are needed to improve disaster preparedness of the older people. The elements needed for disaster preparedness are identified:\nMedication preparedness Mobility assistance Communication support Social support Shelter and other facilities Education and training programs\nFactors affecting disaster preparedness are formulated:\nEducation Disability Income Communication and information technology Family Access to health services\nInterventions to improve preparedness are proposed:\nEnhance disaster preparedness through food bar Enhance disaster preparedness through education Enhance disaster preparedness through assistive technology\nIn my opinion this study provides valuable viewpoint on disaster preparedness of the older people. The information that is provided could be useful for policymakers and should fuel further research. My comments on the article are as follows:\nThere are two objectives provided for the study: “This review aimed to explore all reported research and publications related to natural disasters and medication preparedness among the elderly” and “This review aims to explore disaster and medication preparedness among the elderly, including what the elderly need to prepare in facing disaster, as well as the factors that inhibit them from fulfilling the preparedness and certain interventions to support them to be more prepared”. The title is designed according to the first one but the content better matches the second one. More comprehensive explanation of the search strategy could be given. The keywords are listed in Table 1 of the Supplementary Materials but the exact strategy is not quite clear and reproducible. Also, there are synonyms and related terms that are not captured by the keywords. Figure 2 from the Supplementary Materials could provide all numbers. Database and search approach states that the review was conducted following (PRISMA-ScR) guidelines. This guideline is meant to direct only the reporting not the conducting of a review. In the chapter “Enhance disaster preparedness through assistive technology” there is no information that two of the presented tools are assessed for being useful for older people. The third one reports that additional assistance could be needed by the older people for proper use of the application. As it is stated in the current research: “Despite the widespread use of smartphone apps, social media, and other online communication by various institutions and organizations for rapid information dissemination, these channels remain inaccessible to many seniors, especially those lacking internet access or mobile devices.” Could be noted that more specific research is needed in this otherwise very promising field.\n\nMinor comments:\nSome of the citations in the text have no corresponding number according to the reference list. In the paragraph: “Fathoni et al. (2020) through his research invented an application named “application Bromo alert.” The application has several features: weblogs that offer information about Mount Bromo, including its history and past eruption events; shelters that provide details about shelters, including building names, locations, and proximity to disaster-prone areas; healthcare facilities that contain information about healthcare facilities available in the area; vlogs that provide informative content, especially targeted towards foreign tourists; and CCTV, which allows the public to view live updates of current conditions, particularly the latest situation at Mount Bromo.20” the reference that is cited is not number 20 but 16. Consider rephrasing the sentences to avoid reiteration: “To the best of our knowledge, this literature review is the first to examine disaster preparedness among the elderly preparedness towards disaster.”, “Two reviewers collaborated to develop an approach to search for and determine a suitable determination for the search.”, “The findings of this study need to be prepared to support disaster preparedness for the elderly (medication, mobility assistance, communication support, social support, shelter, education, and training).”.\nIn conclusion, the issue examined in this study is highly relevant and remains far from being resolved. I praise the authors for their efforts to enrich the scientific literature on the subject.\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? 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? 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.) Not applicable", "responses": [] }, { "id": "355519", "date": "20 Jan 2025", "name": "Ryan Maves", "expertise": [ "Reviewer Expertise Disaster medicine", "sepsis", "emerging infectious diseases" ], "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\nThank you for the opportunity to review this manuscript. In this paper, the authors conducted a scoping review of disaster preparedness as it relates to elderly people. I think there is some interesting information in here, and some careful re-editing and re-organization of the text could hopefully make it stronger.\nGeneral comment:\nThe overall quality of written English needs improvement, with many grammatical errors and overly-long descriptions in the text.\nAs a specific example, the opening three sentences of the main text could be re-written as “The global population of elderly people, defined as those 60 years of age or greater, is large and increasing. There were an estimated 1 billion elderly people alive in 2024, a group that is projected to grow to 1.4 billion in 2030 and 2.1 billion by 2050.” An explanation of how becoming elderly is a function of aging is not needed.\nSpecific comments:\nAbstract – the results and conclusions are too brief; while the necessary brevity of an abstract constrains out ability to discuss results in enormous detail, some mention of the actual results of the review should be included.\nIntroduction – It is not necessary to mention what ministries of health are, although it is useful to mention that disaster preparedness may fail to meet the needs of older people.\nResults – 5 papers out of 20 is not “most” (in reference to the number of papers that were written in the US). It is not necessary to list the journals of origin in the main text; this can be included in Table 3 and in the references.\nResults – The results section does not include any actual results, just a list of papers that met their inclusion criteria. Much of this information is included in the discussion but would be properly considered part of the results. The basic organization of preparedness categories in the discussion (mobility assistance, social support, etc.) is a reasonable one, however.\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? 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/13-1510
https://f1000research.com/articles/13-1062/v1	17 Sep 24	{ "type": "Research Article", "title": "Neurodiversity in Saudi Arabia: Towards quality education and reduced inequalities", "authors": [ "Ahmed Yahya Almakrob", "Ahmed Alduais", "Alex S. M. Mhone", "Borey Be", "Ahmed Yahya Almakrob", "Alex S. M. Mhone", "Borey Be" ], "abstract": "Traditional educational frameworks in Saudi Arabia have historically adopted a deficit-based approach to special education, potentially overlooking the benefits of neurodiversity. As global educational paradigms shift toward inclusive practices, examining the alignment of Saudi special education laws with neurodiversity principles becomes crucial. This content analysis study aimed to explore the existing literature on special education in Saudi Arabia to ascertain whether the laws and policies support or hinder the practice of neurodiversity, a concept that is gaining international recognition but remains nascent in Saudi Arabia. A systematic literature search was performed in Web of Science and Scopus, yielding 21 relevant studies after screening and application of inclusion criteria. A content analysis was conducted, focusing on the categorization of data relevant to special education laws and their implications for neurodiversity. The analysis identified seven key categories, including Implementation of Inclusive education and transition services, that reflect the current state of special education laws in relation to neurodiversity. A conceptual model was developed, illustrating the potential of special education laws to both support and impede neurodiversity in Saudi Arabia and potentially worldwide.", "keywords": [ "Neurodiversity", "Special Education Laws", "Saudi Arabia", "Inclusive Education", "Sustainable Development Goals", "Quality Education", "Reduced Inequalities" ], "content": "Introduction\n\nNeurodiversity acknowledges the intrinsic variation in human cerebral architecture, encompassing a spectrum of sociability, cognition, attention, and emotional states, suggesting an inherent appreciation for such neurological differences (Armstrong, 2010). Armstrong posits the fallacy of a ‘standard’ brain, arguing for the recognition of cerebral diversity as a parallel to the richness found in ecological and cultural variance, ultimately framing it as advantageous rather than detrimental (Armstrong, 2010). Conversely, Weiss (2022) conceptualizes neurodiversity as an acceptance of brain function and behavioral differences, considering them a natural facet of human variation. This perspective necessitates embracing cognitive diversity to foster societal cohesiveness. The term ‘neurodivergence’ encapsulates a collective of individuals whose cognitive function diverges from the Predominant Neurotype (PNT), including those with Autism, ADHD, Dyslexia, Dyspraxia/DCD, Dyscalculia, Tourette’s Syndrome, Developmental Language Disorder, and various speech and auditory processing conditions (Sandland et al., 2023).\n\nThe interplay between the concept of neurodiversity and special education legislation is pivotal in shaping the educational landscape for neurodivergent individuals. Special education laws, such as the Individuals with Disabilities Education Act (IDEA) in the United States, are designed to ensure that students with diverse cognitive profiles receive tailored educational opportunities. However, the efficacy of these laws in supporting or impeding neurodiversity hinges on their implementation. While they have the potential to affirm the principles of neurodiversity by promoting inclusive education and individualized support, there is an ongoing debate regarding the extent to which these laws inadvertently contribute to the stigmatization and segregation of neurodivergent individuals, potentially clashing with the ethos of neurodiversity that Armstrong (2010) and Weiss (2022) advocate. Thus, the interpretation and application of special education laws play a crucial role in either reinforcing or challenging societal perceptions of neurodiversity.\n\nSince its inception in 1958, special education in Saudi Arabia has witnessed substantial advancements in the provision of services for students with disabilities. However, improvement is still required in certain areas. Initially, the domain of special education was solely concerned with providing instruction to students who were able to learn despite intellectual disabilities, severe visual impairments, or auditory impairments through special schools. These establishments are presently not regarded as the most suitable environments for exceptionally talented students. The integration of students with disabilities into general public institutions, nevertheless, served as a manifestation of this emerging paradigm and conviction (Bin Battal, 2016).\n\nThe special education has made considerable progress in delivering services to students with disabilities (Aldabas, 2015). Nonetheless, the need for educational and social inclusion of children with intellectual disabilities continues to grow throughout the world. Factors contributing to the increasing need include family advocacy activities, shifting attitudes about the nature of impairment, and increased international acknowledgment of the rights of children with disabilities. In the 1970s, Saudi Arabia began investing in educational programs and support for students with disabilities in both segregated and mainstream education settings (Abu-Alghayth et al., 2022).\n\nThe term “Special Institutes” or “Special Education Institutes”, as they are known in Saudi Arabia, refers to separate schools, special schools, or special education schools. These schools include schools for the blind, schools for the deaf, and schools for the mentally retarded (UNESCO, 2010). Special education services for students with disabilities have been introduced in response to the inadequacy of the current public education system, which is unable to fully accommodate many impaired students. Every country in the world has built its own particular education services programme and improves the education organisation on an annual basis. Saudi Arabia is one of the developing countries that has established and updated its educational model over time to allow children with disabilities to get an education (Awdah Alzahrani, 2023).\n\nAccording to the Ministry of Education, the country’s education policy includes many essentials and factors related to special education. These include respecting the dignity of individuals and providing opportunities for them to develop their skills and abilities, so they can contribute to national development (Article 36). The policy also emphasizes taking care of students who are slow in their studies and working to eliminate the causes of their underdevelopment. It calls for the creation of temporary and permanent programs tailored to their needs (Article 55). Additionally, the policy highlights the importance of special education and care for mentally and physically impaired students. It aims to make education accessible to all children in accordance with Islamic teachings (Article 56). Lastly, the policy emphasizes the need to focus on education for physically and mentally disabled individuals, creating a special cultural curriculum and diversified training programs to meet their specific needs (Article 188) (MOE, 2021).\n\nSpecial education services in Saudi Arabia have received much attention over the past 15 years, with increased focus on offering such services (Husain R Alruwaili, 2016). The country has a National Strategy for the Development of General Education, which affirms the provision of equal opportunities for people with disabilities (MOE, 2021). Disability laws in Saudi Arabia mandate that higher education institutions provide support for special needs learners to guarantee educational opportunities equivalent to opportunities for their nondisabled peers (Abed & Shackelford, 2020).\n\nSDG 4 aims to guarantee inclusive and equitable quality education and promote lifelong learning opportunities for all. This includes attempts such as ensuring that all girls and boys complete free, equitable, and quality primary and secondary education, as well as giving access to excellent early childhood development and care. Additionally, it aspires to remove gender inequities in education and provide equitable access to all levels of education for vulnerable groups, including persons with disabilities and indigenous peoples (United Nations, 2023a). Despite progress, as many as 48.1 per cent of girls remain out of school in some regions. Gender gaps in primary and secondary enrolment rates have nearly closed, on average. Yet 15 million girls are not in primary school right now, compared to 10 million boys. In adolescence, higher numbers of girls often drop out of secondary school for reasons including early pregnancy and the expectation that they should contribute to household work (UN Women, n.d.).\n\nOn the other hand, SDG 10 focuses on reducing disparities within and across nations. This comprises enabling and encouraging the social, economic, and political inclusion of all individuals, irrespective of age, sex, impairment, race, ethnicity, origin, religion, or economic condition. It also tries to promote equal opportunity and eliminate disparities by removing discriminatory laws, policies, and practices (United Nations, 2023b).\n\nIn Saudi Arabia, achieving SDG 4 (quality education) and SDG 10 (reduced inequalities) presents several challenges. SDG 4 addresses some concerns, including inequities in educational accessibility and quality, the effects of the COVID-19 epidemic on education, and the need for total educational reform. Saudi Arabia has faced challenges such as the impact of the COVID-19 pandemic on education, disparities in educational accessibility and quality, and the need for major education reform. Furthermore, the persistence of disparities in literacy rates, enrolment rates, and educational attainment, particularly in some areas, poses a challenge to implementing universal education goals (Barry, 2019; Matilde, 2024; OECD, 2020; Reema et al., 2022). Regarding SDG 10, Saudi Arabia faces obstacles in tackling many types of inequality, including as income, gender, and disability. This necessitates equitable resource allocation, investments in education and skill development, and combatting discrimination. Saudi Arabia’s Gini Coefficient Index, which measures income distribution, was 45.6 in 2019, suggesting high income inequality (Statista, 2024; World Economics, 2024). Furthermore, the country confronts challenges in resolving ongoing national inequalities, notably in terms of income and economic status (Statista, 2024).\n\nThis content analysis study aims to rigorously explore and evaluate the existing published literature on special education within the Kingdom of Saudi Arabia, with a specific focus on examining how special education laws either enhance or impede the practice of neurodiversity—a concept that, while globally recognized, remains relatively nascent in the Saudi context. In advancing the discourse on neurodiversity, this study seeks to contribute to the burgeoning dialogue on inclusive education and the acknowledgment of diverse neurological conditions as a facet of human diversity rather than deficits. The scope of this study is confined to the systematic examination of peer-reviewed articles, institutional reports, and legislative documents, all of which contribute to a comprehensive understanding of the state of special education laws. Through meticulous content analysis, the study endeavours to construct a conceptual model that elucidates whether the current special education framework in Saudi Arabia is conducive to fostering an environment that embraces neurodiversity or if it inadvertently contributes to the marginalization of this population.\n\n\nMethod\n\nThe initial data were collected using a comprehensive search strategy implemented on two major academic databases, Web of Science and Scopus. The search terms were meticulously selected to capture the breadth of literature on special education within the context of Saudi Arabia. The terms ‘special education policy’, ‘special education legislation’, ‘special education law’, and ‘special education regulation’ were used in conjunction with Boolean operators, searching within both titles and abstracts for an inclusive retrieval process. Additionally, the term ‘Saudi Arabia’ was included in the abstract search to narrow the focus geographically. This strategy yielded an initial pool of 306 studies.\n\nA thorough screening process was conducted, focusing on titles and abstracts to assess the relevancy of each study. This process, governed by a set of inclusion criteria, reduced the pool to 21 pertinent studies. The inclusion criteria mandated that studies pertain specifically to special education within the Saudi Arabian context and include either primary or secondary data. Studies were excluded if they referenced special education but did not focus on Saudi Arabia. The resulting sample of studies provided a robust dataset for the subsequent content analysis. See Figure 1 for the inclusion and exclusion PRISMA flowchart.\n\nThe measures implemented in this content analysis were twofold. Firstly, the analysis involved the meticulous extraction of data relevant to the intersection of special education laws and neurodiversity within the selected Saudi Arabian studies. This process required a detailed examination of the content to discern the authors’ perspectives on inclusion, segregation, and neurodiversity as demonstrated in their findings and conclusions.\n\nSubsequently, the extracted information was systematically categorized into a conceptual framework representing the current state and implications of special education laws in Saudi Arabia. This framework considered whether the laws and their implementation support or oppose the practice of neurodiversity. The studies were classified based on their explicit or implicit support, opposition, or mixed views regarding neurodiversity, with particular attention paid to their stance on inclusion and segregation.\n\nThe research employed a content analysis study design, a methodological approach suited for systematic and objective analysis of communication within the textual data. Content analysis facilitates the identification, coding, and categorizing of patterns and themes within qualitative data. It is uniquely positioned to handle large volumes of data, allowing for the distillation of comprehensive information into actionable insights. This technique is particularly advantageous in policy research where the interpretation of language and legislation is critical.\n\nThe study commenced on February 6, 2024, with an initial literature review to identify relevant studies on the topic of special education in Saudi Arabia. A team of three researchers independently reviewed the papers, ensuring a comprehensive and unbiased selection process. Any discrepancies in study inclusion were resolved through discussion and consensus-building among the research team members. All the data associated with this study including PRISMA flowchart, PRISMA checklist are available in our supplementary preregistered data (Almakrob et al., 2024).\n\nUpon finalizing the list of included papers, a structured data extraction protocol was implemented. The data were organized into two main tables designed to facilitate the synthesis of findings (see Table 1 and Table 2). These tables served as the foundation for the development of a conceptual model, which aimed to encapsulate the role of special education in promoting or impeding the practice of neurodiversity in Saudi Arabia.\n\nThe synthesis of data culminated in a nuanced conceptual model that not only reflects the current legal and educational landscape but also provides insights into potential directions for policy and practice. This model serves as a valuable tool for stakeholders in Saudi Arabia’s education sector, offering a clear depiction of the interplay between special education legislation and the principles of neurodiversity.\n\n\nResults\n\nThe results section commences with a detailed table encapsulating summaries of the selected studies, aligned with their implications for special education and the burgeoning concept of neurodiversity within the Saudi Arabian context. This initial synthesis served as the foundation for a deeper analysis, leading to the development of a subsequent table where studies were systematically categorized into seven distinct themes. These themes elucidate varying dimensions of special education laws and their intersection with neurodiversity, providing insight into whether these laws serve to enhance or hinder the practice of neurodiversity in Saudi Arabia. Drawing from the rich data of these categorizations and the nuanced understanding they afford; we have crafted a conceptual model. This model serves as a theoretical framework that captures the application of special education laws to neurodiversity in Saudi Arabia, offering a reflective lens on both the current state and potential future directions of special education practices in the Kingdom and their broader international implications.\n\nTable 1 presents a synthesis of 21 studies, each meticulously analysed to discern their contribution to the understanding of special education laws and their interplay with neurodiversity in Saudi Arabia (Almakrob et al., 2024). The studies were chosen based on their explicit aim to critique, explore, and assess various aspects of special education ranging from policy analysis, educator preparedness, inclusivity in educational practices, and the readiness of schools to embrace neurodiverse students. The findings from these studies collectively underscore a growing recognition of the need for systemic shifts towards inclusive educational environments that are responsive to the diverse needs of all learners, including those with disabilities. Each study adds to the argument that special education, when effectively implemented, can be a robust vehicle for supporting neurodiversity, as it advocates for the adaptation of teaching methodologies, transition services, and educational technologies that cater to the spectrum of learning needs. The studies vary in their position, with most supporting the advancement of neurodiverse-inclusive practices, some presenting a mixed view indicating the existence of barriers to full realization, and others opposing, reflecting resistance to inclusive practices. Collectively, these academic inquiries pave the way for a conceptual model that encapsulates the potential of special education laws to support or hinder the practice of neurodiversity in Saudi Arabia, offering implications for both national educational strategies and global discourse on inclusive education.\n\nThe landscape of special education in Saudi Arabia is under rigorous scholarly scrutiny, as evidenced by a multitude of studies exploring various facets of this intricate field. The purpose of this essay is to synthesize key categories derived from a content analysis of 21 studies, providing a panoramic view of special education laws and their alignment with the concept of neurodiversity in Saudi Arabia. These categories not only reveal the current state of special education but also forecast potential trajectories for the promotion of neurodiversity in the Kingdom.\n\nOne prominent category that emerges from the analysis is the Implementation of Inclusive Education, which is pivotal for fostering a neurodiverse-friendly educational environment (Abu-Alghayth, 2021; Alsalem & Alzahrani, 2023). This category underscores the necessity to critique and improve the practical application of inclusive education, which aims to integrate students with disabilities into mainstream classrooms. It echoes the sentiment that without robust implementation strategies, the theoretical underpinnings of inclusive education policies may fail to materialize into tangible benefits for students with disabilities, potentially perpetuating segregation rather than fostering inclusivity.\n\nTransition and Post-School Outcomes is another significant category, emphasizing the importance of equipping students with disabilities for life beyond the classroom (Almalki, 2022; Alsalamah, 2023). This category underscores a lacuna in effective transition planning and services, suggesting that enhanced transition services could lead to greater societal acceptance and integration of neurodiverse individuals. The implications of this category are profound, as the quality of transition services directly correlates with the ability of individuals with disabilities to contribute meaningfully to society and lead fulfilling lives.\n\nThe third category, Educational Barriers and Facilitators, highlights the factors influencing the educational experiences of students with disabilities (Abu-Alghayth, 2020; Ashur & Bagadood, 2022). The findings within this category suggest that addressing barriers such as insufficient teacher preparedness and enhancing facilitators like the use of evidence-based practices and assistive technology are essential for creating a learning environment that respects and caters to neurodiversity. This category underscores the necessity of a multifaceted approach to special education, one that considers the complex interplay of various factors affecting the educational journey of students with disabilities.\n\nStakeholder Attitudes and Knowledge is a category that encapsulates the spectrum of beliefs and awareness levels among individuals directly involved in or affected by special education (Abed, 2020; Bagadood & Sulaimani, 2022). Studies in this category reveal that attitudes and knowledge of stakeholders, including educators, parents, and policymakers, can either act as formidable barriers to or facilitators of the integration of students with disabilities. This category is critical, as it underscores the importance of informed and positive stakeholder engagement for the advancement of neurodiversity.\n\nLastly, the category of Policy Analysis and Development is crucial as it pertains to the examination and advancement of legislation aimed at supporting students with disabilities (Alquraini, 2014; Mohammed, 2018). This category suggests that reflective and progressive policymaking is foundational for an educational system that not only accommodates but also celebrates neurodiversity. Studies in this category provide a framework for understanding the current policy landscape and suggest pathways for legislative enhancements that can better meet the needs of students with disabilities.\n\nThe conceptual model presented delineates a multifaceted approach to understanding special education laws in Saudi Arabia and their alignment with the concept of neurodiversity (see Figure 2). It provides a framework for evaluating whether the current educational landscape supports the integration of neurodiverse individuals or inadvertently contributes to their segregation. Each aspect of the model plays a crucial role in this evaluation:\n\n1. Foundational Policies and Legislations: The cornerstone of the model is the robustness of special education laws and policy formulation. Policies that are comprehensive, inclusive, and aligned with international standards set the stage for a supportive environment for neurodiversity. However, if these policies are not well-conceived or are out of sync with global best practices, they may create systemic barriers that limit the participation of neurodiverse students (Alquraini, 2014; Mohammed, 2018).\n\n2. Implementation of Inclusive Practices: The transition from policy to practice is a critical step. Effective implementation of inclusive education practices indicates a commitment to neurodiversity, facilitating diverse learners’ access to mainstream education and promoting positive social interactions (Abu-Alghayth, 2021; Alsalem & Alzahrani, 2023). Lack of implementation, on the other hand, can lead to de facto segregation, even if unintentional.\n\n3. Transition Services: Quality transition services are indicative of a system that values neurodiverse students beyond their academic tenure, preparing them for life’s various roles. Adequate transition planning reflects an understanding that education is not an end in itself but a means to a fulfilling and autonomous life (Almalki, 2022; Alsalamah, 2023). Without these services, students may find themselves marginalized from mainstream society and the workforce.\n\n4. Educational Barriers and Facilitators: Identifying and managing barriers while leveraging facilitators is akin to navigating a ship through a narrow strait. Barriers such as inadequate teacher training or lack of resources can impede progress, while facilitators like assistive technology and evidence-based practices can accelerate the journey toward a neurodiverse-friendly educational environment (Abu-Alghayth, 2020; Ashur & Bagadood, 2022).\n\n5. Stakeholder Engagement: The attitudes and knowledge of stakeholders can dramatically shape the educational experience of neurodiverse students. Educated and positive stakeholder communities can become champions of inclusion, advocating for and reinforcing the principles of neurodiversity within the educational system (Abed, 2020; Bagadood & Sulaimani, 2022). Conversely, uninformed or negative attitudes can create an unwelcoming environment that stifles the potential of neurodiverse individuals.\n\n6. Accessibility and Assistive Technology: Accessibility is a key indicator of a system’s commitment to neurodiversity. When students with disabilities have access to appropriate assistive technologies, it can level the playing field, allowing them to fully engage with the curriculum and their peers (Abu-Alghayth, 2020; Alsamiri et al., 2022). A lack of accessible options suggests a system that is not fully prepared to support neurodiverse students’ needs.\n\n7. Specialized Educational Strategies: Tailored educational strategies recognize and cater to the individualized learning profiles of neurodiverse students. The use of specialized strategies is a testament to a system’s adaptability and its willingness to meet students “where they are” in terms of learning needs (Alasim, 2020; Ashour & Bagadood, 2022). Without such strategies, students may not receive the support they require to thrive.\n\n\nDiscussion\n\nThe aim of this study was to conduct a content analysis of the literature concerning special education in Saudi Arabia, with a particular emphasis on the concept of neurodiversity, which remains a relatively nascent idea within the Saudi context. The analysis sought to determine whether the special education laws in Saudi Arabia are conducive to fostering an environment that supports neurodiversity or whether they contribute to the segregation of neurodiverse individuals. The findings revealed that while there is a framework in place that could support neurodiversity, including policies that draw from international best practices and a growing awareness of inclusive education, there are still significant barriers to its full implementation. These barriers range from insufficient policy implementation and stakeholder attitudes to inadequate transition services and educational resources.\n\nThe results of this study align with previous literature that emphasizes the importance of teacher preparation in supporting neurodiverse students. De Arment and Traylor and Rosenblatt, Frates, and Jackson both underscore the need for special educators to be equipped with the knowledge and skills to support neurodiverse learners effectively (De Arment & Traylor, 2023; Rosenblatt et al., 2023). This is consistent with the current study’s findings, which highlight the need for professional development and curriculum adaptation within Saudi Arabia’s special education framework. Moreover, Shenker, Rodgers, Guitar, and Onslow discuss the diverse viewpoints on neurodiversity and ableism in clinical practice, mirroring the diversity of perspectives on inclusion within the Saudi context (Shenker et al., 2023).\n\nAdditionally, the representation of neurodivergent individuals in media, as analysed by (Lam & Wong, 2023), plays a crucial role in shaping public discourse and understanding of neurodiversity. This has parallels with the Saudi context, where public perception and stakeholder attitudes are determining factors in the support for neurodiversity. Asbell-Clarke et al. highlight the importance of including neurodiverse learners in computational thinking activities, suggesting that educational content and methods should be adapted to support diverse cognitive abilities—a finding that resonates with the need for specialized strategies identified in this study (Asbell-Clarke et al., 2022).\n\nHunt’s et al. examination of special educators’ knowledge of student mathematical thinking further supports the argument that educators must understand diverse ways of reasoning to effectively teach students with disabilities (Hunt et al., 2021). This is in line with the current study’s findings that teacher preparedness and knowledge are crucial for implementing effective special education practices. Garner’s work on dyslexia emphasizes the need to update strengths-based practices and attitudes, paralleling the Saudi context where neurodiversity is often framed within a deficit-based perspective that needs to be shifted (Garner, 2021).\n\nGrinker’s (2020) analysis of autism and the sociocultural construction of stigma through an economic lens presents an interesting comparison to the Saudi context, where the conceptualization of neurodiversity could be influenced by various social and economic factors. Lambert’s critique of deficit mythologies surrounding students with learning disabilities in mathematics echoes the findings of this study (Lambert, 2018), where there is a need to counter limiting beliefs and support neurodiverse students’ potential in Saudi Arabia. Silberman’s discussion on neurodiversity rewiring conventional thinking about brains is particularly relevant as it advocates for a focus on native strengths and special interests (Silberman, 2017), which is a perspective that needs to be integrated into Saudi Arabia’s special education framework.\n\nPuccini’s et al. work on educational access through multisensory design principles for neurodiverse learners (Puccini et al., 2013) and Tincani’s et al. exploration of diversity in educational interventions for ASD (Tincani et al., 2009) further underscore the importance of designing educational interventions that recognize and embrace neurodiversity. These studies collectively highlight that while there is a growing recognition of the need to support neurodiverse learners, there are still considerable gaps in practice and attitudes that must be bridged.\n\nIn summary, the current study’s findings are consistent with the broader literature in that they recognize the critical role of inclusive policies, educator preparation, and a shift in attitudes towards neurodiversity. However, they also reveal that Saudi Arabia, much like other contexts, faces significant challenges in fully realizing a supportive environment for neurodiverse individuals. As Saudi Arabia continues to develop its special education framework, it is imperative that the principles of neurodiversity are ingrained within all aspects of policy and practice to ensure that all learners can thrive in an inclusive and supportive educational system.\n\nThis study holds several implications for educational policy and practice within the Kingdom of Saudi Arabia. The conceptual model developed herein provides a scaffold for policymakers to examine and refine existing special education laws, ensuring they are conducive to fostering neurodiversity. It suggests that a shift in emphasis from a deficit-based to a strengths-based perspective within special education could enhance the support for neurodiversity. For practitioners, this study underscores the importance of professional development tailored towards understanding and embracing neurodiversity in the classroom. Furthermore, it highlights the need for increased cross-cultural and cross-linguistic research to enrich the understanding of special education within the Saudi context, potentially informing a more inclusive global narrative on neurodiversity.\n\nThe present study, while comprehensive in its approach to content analysis, is not without limitations. The scope was confined to published literature, which inherently may not capture the entirety of the special education landscape in Saudi Arabia, including potentially influential yet unpublished policies, grey literature, or on-the-ground practices. Moreover, the analysis was limited to documents in English, potentially omitting relevant research or policy documents published in Arabic. The study also assumes that the presence of laws or policies is indicative of their implementation, which may not always be the case due to various socio-cultural, economic, or political barriers that impede actualization. Therefore, the findings should be interpreted with caution and viewed as indicative rather than exhaustive, acknowledging the potential for gaps between policy intent and practice.\n\n\nEthical statement\n\nNeither human nor non-human participants were included in this study, so an IRB approval was not required.", "appendix": "Data availability\n\nThe extracted data supporting this study and PRISMA checklist are accessible from OSF repository registered on 30 August 2024, available at https://doi.org/10.17605/OSF.IO/TKQG6 (Almakrob et al., 2024).\n\nSecondary data used in this study is available at https://doi.org/10.17605/OSF.IO/TKQG6 (Almakrob et al., 2024).\n\nData are available under the terms of the Creative Commons 1.0 Universal License (CC0 1.0).\n\n\nAcknowledgments\n\nThis project is sponsored by Prince Sattam Bin Abdulaziz University (PSAU) as part of funding for its SDG Roadmap Research Funding Programme project number PSAU-2023-SDG-46.\n\n\nReferences\n\nAbed MG: Stakeholders’ views of children with special needs in Saudi Arabia. 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PubMed Abstract \| Publisher Full Text\n\nAlquraini T: Special Education In Saudi Arabia: Challenges, Perspectives, Future Possibilities. J. Int. Spec. Needs Educ. 2010a; 25.\n\nAlquraini T: Special education in Saudi Arabia: challenges, perspectives, future possibilities. Int. J. Spec. Educ. 2010b; 25.\n\nAlquraini TA: Special Education Today in the Kingdom of Saudi Arabia.Rotatori AF, Bakken JP, Burkhardt S, et al., editors. Special education international perspectives: practices across the globe. 2014; Vol. 28: pp. 505–528. Publisher Full Text\n\nAlquraini TA, Rao SM: Assessing teachers’ knowledge, readiness, and needs to implement Universal Design for Learning in classrooms in Saudi Arabia. Int. J. Incl. Educ. 2020; 24(1): 103–114. Publisher Full Text\n\nAlsalamah AA: Promoting Successful Transitions in High School Students with Emotional Disturbance in Saudi Arabia: Educators’ Voices. Career Dev. Transit. Except. Individ. 2023; 46(1): 15–27. Publisher Full Text\n\nAlsalem MA, Alzahrani HA: The Role of Institutional Practice in Promoting Inclusive Education for Deaf and Hard of Hearing Students: A Critical Analysis. SAGE Open. 2023; 13(4). Publisher Full Text\n\nAlsamiri YA, Alsawalem IM, Hussain MA, et al.: Providing accessible distance learning for students with disabilities in Saudi Arabia. International Journal of Advanced and Applied Sciences. 2022; 9(1): 34–40. Publisher Full Text\n\nAltamimi AA, Lee LW, Sayed-Ahmed ASA, et al.: Special education in Saudi Arabia: a synthesis of literature written in English. Int. J. Spec. Educ 2015; 30(3): 98–117.\n\nArmstrong T: Neurodiversity: Discovering the Extraordinary Gifts of Autism. ADHD; 2010.\n\nAsbell-Clarke J, Robillard T, Edwards T, et al.: Including Neurodiversity in Foundational and Applied Computational Thinking (INFACT). Proceedings of the 53rd ACM Technical Symposium on Computer Science Education V. 2. 2022; pp. 1076–1076. Publisher Full Text\n\nAshour JT, Bagadood NH: Teachers’ Perspectives on Using Applied Behavior Analysis Strategies to Students with Autism Spectrum Disorder in Saudi Arabia: Barriers and Solutions. Int. J. Educ. Sci. 2022; 37(1–3): 25–34. Publisher Full Text\n\nAshur AT, Bagadood NH: Evidence-Based Practices (EBP) in Teaching Students with Intellectual Disabilities in Saudi Arabia: Obstacles and Solutions. Int. J. Educ. Sci. 2022; 36(1–3): 101–111. Publisher Full Text\n\nAwdah Alzahrani N: The Future of Inclusive Education in Saudi Schools. WRP. Sci. J. 2023; 6(1): 17–24. Publisher Full Text\n\nBagadood NH, Saigh BH: Teaching Assistants as a Prerequisite for Best Practice in Special Education Settings in Saudi Arabia. Int. J. Comput. Sci. Netw. Secur. 2022; 22(3): 101–106. Publisher Full Text\n\nBagadood NH, Sulaimani MF: Mainstreaming of Students with Intellectual Disability in the Kingdom of Saudi Arabia: Special Education Teachers’ Perceptions. Int. J. Comput. Sci. Netw. Secur. 2022; 22(3): 183–188. Publisher Full Text\n\nBarry A: Gender Differences in Academic Achievement in Saudi Arabia: A Wake-Up Call to Educational Leaders. Int. J. Educ. Policy Leadersh. 2019; 15(15). Publisher Full Text\n\nBin Battal ZM: Special Education in Saudi Arabia.2016.\n\nDe Arment S, Traylor TG: “Walking the Talk” in Special Educator Preparation for Neurodiversity. Autism, Neurodiversity, and Equity in Professional Preparation Programs. 2023; pp. 1–24. Publisher Full Text\n\nGarner MD: Dyslexia primer for social work: Translational research to update strengths-based practice, advocacy, and attitudes. Adv. Soc. Work. 2021; 21(1): 26–44. Publisher Full Text\n\nHunt JH, Martin K, Patterson B, et al.: Special educators’ knowledge of student mathematical thinking. J. Math. Teach. Educ. 2021; 25: 581–598. Publisher Full Text\n\nAlruwaili HR: Obstacles of Special Education Services for Students with Intellectual Disabilities in Saudi Arabia: Future Directions.2016.\n\nKauffman JM, Felder M, Ahrbeck B, et al.: Inclusion of All Students in General Education? International Appeal for A More Temperate Approach to Inclusion. J. Int. Spec. Needs Educ. 2018; 21(2): 1–10. Publisher Full Text\n\nKeller C, Al-Hendawi M, Abuelhassan H: Special Education Teacher Preparation in the Gulf Cooperation Council Countries. Teach. Educ. Spec. Educ. 2016; 39(3): 194–208. Publisher Full Text\n\nLam GYH, Wong HT: The representation of neurodivergent individuals in Radio Television Hong Kong’s television series A Wall-less World. Int. J. Qual. Stud. Educ. 2023. Publisher Full Text\n\nLambert R: “Indefensible, illogical, and unsupported”; Countering deficit mythologies about the potential of students with learning disabilities in mathematics. Educ. Sci. 2018; 8(2). Publisher Full Text\n\nMatilde R: Educational Challenges in Saudi Arabia.Broken Chalk; 2024. Reference Source\n\nMemri: Saudi Arabia Reforms School Curriculum, Cuts Religion Classes; Journalists, Intellectuals Praise The Decision. Memri: 2022a. Reference Source\n\nMemri: Saudi Arabia Reforms School Curriculum, Cuts Religion Classes; Journalists, Intellectuals Praise The Decision. Memri; 2022b. Reference Source\n\nMinistry of Education: Technical and Vocational Education.2021. Reference Source\n\nMOE: People with Disabilities.2021. Reference Source\n\nMohammed A: Twice-Exceptionality in the Kingdom of Saudi Arabia: Policy Recommendations for Advances in Special Education. Int. J. Spec. Educ 2018; 33(2): 397–415.\n\nOchoa TA, Alharbi NH, Monteiro MCB, et al.: Incarceration Risk Among Girls With Behavior Disorders: An International Perspective. Interv. Sch. Clin. 2023; 58(5): 308–316. Publisher Full Text\n\nOECD: Education in Saudi Arabia. OECD; 2020. Publisher Full Text\n\nPoch AL, Alzahrani D, Aljuwayhir J, et al.: Educating Students with Learning Difficulties in the Kingdom of Saudi Arabia. Interv. Sch. Clin. 2023; 58(4): 287–292. Publisher Full Text\n\nPuccini AM, Puccini M, Chang A: Acquiring educational access for neurodiverse learners through multisensory design principles. Proceedings of the 12th International Conference on Interaction Design and Children. 2013; pp. 455–458. Publisher Full Text\n\nReema A, Abby K, Ibrahim A: The Role of Saudi Universities in Advancing the SDGs and Saudi Vision 2030.2022.\n\nRosenblatt K, Frates A, Jackson H: Addressing Neurodiversity Within Educator Preparation. Autism, Neurodiversity, and Equity in Professional Preparation Programs. 2023; pp. 25–53. Publisher Full Text\n\nSandland B, Macleod A, Hall N: Neurodiverse students within higher education-initial thoughts from a collaborative project. Education in Practice. 2023; Vol. 4(1).\n\nShenker R, Rodgers N, Guitar B, et al.: Contemporary clinical conversations about stuttering: Neurodiversity and ableism. J. Fluen. Disord. 2023; 78: 106014. PubMed Abstract \| Publisher Full Text\n\nSilberman S: Neurodiversity Rewires Conventional Thinking About Brains. Beginning with Disability. Routledge; 2017; pp. 51–52. Publisher Full Text\n\nStatista: Economic Inequality - Saudi Arabia. Statista; 2024. Reference Source\n\nTincani M, Travers J, Boutot A: Race, culture, and autism spectrum disorder: Understanding the role of diversity in successful educational interventions. Res. Pract. Persons Severe Disabl. 2009; 34(3–4): 81–90. Publisher Full Text\n\nUN Women: In focus: Women and the Sustainable Development Goals (SDGs): SDG 4: Quality education.n.d. February 7, 2024. Reference Source\n\nUNESCO: The Experience of the Kingdom of Saudi Arabia in mainstreaming students with special educational needs in public schools - UNESCO Digital Library.2010. Reference Source\n\nUNESCO: Saudi Arabia Inclusion. UNESCO; 2021. Reference Source\n\nUnited Nations: Ensure inclusive and equitable Quality education and promote lifelong learning opportunities for all (Goal 4). Sustainable Development Goals. 2023a. Reference Source\n\nUnited Nations: Reduce inequality within and among countries (Goal 10). Sustainable Development Goals; 2023b. Reference Source\n\nWeiss KJ: Neurodiversity: From Phenomenology to Neurobiology and Enhancing.2022.\n\nWorld Economics: Saudi Arabia’s Inequality Levels. World Economics; 2024. Reference Source\n\nZainab A: The Challenges Facing Special Needs Students in Education in the Kingdom of Saudi Arabia.2021." }	[ { "id": "333967", "date": "21 Nov 2024", "name": "Lori Severino", "expertise": [ "Reviewer Expertise special education", "learning disabilities", "dyslexia", "dysgraphia", "explicit instruction" ], "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\nPeer Review of the Article Using Content Analysis\nThe article under review presents a comprehensive approach to a content analysis of the current state of neurodiversity and special education law and process in Saudi Arabia, effectively situating its research within the context of current scholarship. One of the strengths of this paper lies in the use of up-to-date citations and research, which underscores the authors' dedication to maintaining relevance and engaging with the most recent developments in the field. This ensures that the analysis is not only current but also integrates contemporary findings that add to the overall robustness of the study.\nA helpful feature of the article is the thoughtful development of a conceptual model. This model provides a structured framework that guides the reader through the authors' approach and findings. It holds promise as a foundational tool for further investigation by other researchers, which could lead to the expansion and refinement of existing theories and practices.\nThe authors also excel in connecting their findings to the broader landscape of research. By drawing meaningful comparisons and contrasts between their results and those found in existing literature, they demonstrate a deep understanding of the field.\n\nFurthermore, the flow chart included in the article is clear and concise, aiding readers in visualizing the process and results of the content analysis. This visual tool enhances comprehension and ensures that complex ideas are communicated effectively, allowing readers to grasp the study’s conclusions with ease.\nAreas for Improvement\nWhile the study design demonstrated a solid process, there are a few areas where it could be improved to enhance the clarity and replicability of the research. For instance, although the process was well-outlined, it would have been beneficial for the authors to provide more detailed information on the number of articles that required discussion among the authors during the coding process. Additionally, specifying whether a protocol was used to guide these discussions would have added to the transparency.\nAnother point to consider is the inclusion of data on the number of coding iterations. Mentioning how many rounds of coding were conducted would have provided valuable insights into the thoroughness of the analysis and supported future researchers aiming to replicate the study.\nIn terms of sources, while the research leveraged a solid base, the scope could have been expanded by using additional search engines and incorporating more diverse search terms. Terms such as \"learning disabilities\" or \"autism\" may have captured a wider array of relevant studies that were not identified using broader terms like \"neurodiversity\" or \"special education.\"\nA review of Table 2 reveals that the studies included in the analysis were limited in their coverage of learning disabilities, with only one article addressing dyslexia—arguably the most common learning disability—at the higher education level. Including more comprehensive discussions or sources focused on learning disabilities would have strengthened the review. One article that could have been particularly fitting is Aldousari A, et al., 2022 (Ref 1)\n\nBy addressing these areas for improvement, the article's robustness and replicability could be enhanced, contributing to a more comprehensive analysis and a stronger foundation for subsequent research.\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?\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": "12869", "date": "09 Dec 2024", "name": "Ahmed Alduais", "role": "Author Response", "response": "Dear Lori, Thank you immensely for the time and effort you invested in the peer review report. My colleagues and I greatly value your insightful comments for the enhancement of our paper. We have implemented changes based on your suggestions, which we trust align with your expectations. Once again, we extend our deepest thanks. Ahmed Alduais (PhD), Department of Psychology Norwegian University of Science and Technology, Trondheim, Norway --------------------- Reviewer Comments: While the study design demonstrated a solid process, there are a few areas where it could be improved to enhance the clarity and replicability of the research. For instance, although the process was well-outlined, it would have been beneficial for the authors to provide more detailed information on the number of articles that required discussion among the authors during the coding process. Additionally, specifying whether a protocol was used to guide these discussions would have added to the transparency. Author Response: Clarifications have been added in blue to the methods section for both points. --------------------- Reviewer Comments: Another point to consider is the inclusion of data on the number of coding iterations. Mentioning how many rounds of coding were conducted would have provided valuable insights into the thoroughness of the analysis and supported future researchers aiming to replicate the study. Author Response: A clarification has been added to the methods section in blue. --------------------- Reviewer Comments: In terms of sources, while the research leveraged a solid base, the scope could have been expanded by using additional search engines and incorporating more diverse search terms. Terms such as \"learning disabilities\" or \"autism\" may have captured a wider array of relevant studies that were not identified using broader terms like \"neurodiversity\" or \"special education.\" Author Response: A clarification has been added to the sample section, highlighted in blue. --------------------- Reviewer Comments: A review of Table 2 reveals that the studies included in the analysis were limited in their coverage of learning disabilities, with only one article addressing dyslexia—arguably the most common learning disability—at the higher education level. Including more comprehensive discussions or sources focused on learning disabilities would have strengthened the review. One article that could have been particularly fitting is Aldousari A, et al., 2022 (Ref 1) Author Response: Regarding this point, we did not include these studies by our colleagues as their papers were focused on policies and laws. However, in another paper recently published in the European Journal of Education, we incorporated two of their studies because that paper was more centered on attitudes and perspectives, which aligned with the focus of these two studies. https://onlinelibrary.wiley.com/doi/10.1111/ejed.12809" } ] }, { "id": "337171", "date": "22 Nov 2024", "name": "Clayton Keller", "expertise": [ "Reviewer Expertise International special and inclusive education", "special educator teacher preparation", "special education policy" ], "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, Neurodiversity in Saudi Arabia: Towards quality education and reduced inequalities, aims to consider the extent to which special education laws and policies in the Kingdom of Saudi Arabia are supportive of the concept of neurodiversity and individuals with such conditions via a content analysis of published articles about special education in the country. The study’s premise raises two questions for me. First, if neurodiversity is a relatively recent conceptualization of the conditions experienced by a range of individuals, is it fair, in a sense, to judge whether laws and policies created before either the development or at least the increased attention to the conceptualization are amenable to the conditions? Also, wouldn’t a content analysis of the country’s laws and policies themselves be a more direct assessment of neurodiversity acceptance and support than studies about special education for which they judge the studies’ authors’ perspectives about KSA’s laws and policies (if even such perspectives could be accurately determined in all cases)?\nIn a number of instances, sufficient details were not provided about the study’s methodology and analysis to make the research replicable. For instance, the narrative says that 306 potential studies were found initially yet the PRISMA flowchart starts with 112. Why is there a discrepancy? No information is provided about the instrument used as part of the content analysis; its inclusion as one of the supplemental documents would have been useful (and if the instrument were the very detailed PRISMA flowchart in the supplemental materials, say so). (The authors did do a good job in providing such resources in the OSF Registry for this study.) The authors say that “the extracted information was systematically categorized into a conceptual framework” (p. 6) but no information is provided about such a systematic approach. I would urge the authors to provide many more details in any revision of the current article. Providing citations, too, for the specific content analytical approaches that were used would help other researchers conduct similar work if they were inclined.\nI would also suggest that the authors support their statements of the results with very specific details from the studies they cite in support of the results. That could strengthen the validity of their conclusions, moving beyond their present statements of generalities.\nFinally, in a revision, I would remove the many adverbs and adjectives that the authors use to present their work in a positive light, e.g., meticulously, comprehensive, thorough, nuance, and valuable, amongst others. A reader assumes that, in a published article, such qualities informed the research. To frequently state this, though, can raise doubt in the reader’s mind.\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?\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": [ { "c_id": "12870", "date": "09 Dec 2024", "name": "Ahmed Alduais", "role": "Author Response", "response": "Dear Keller, Thank you so much for your effort to provide this valuable peer review. We have now completed revising our paper based on your feedback. We have tried to our best to apply as much as possible of these suggestions in the new revised versions. You can see all changes in red. Below you will find your comments and our responses. Once again, thank you so much. Ahmed Alduais (PhD) Department of Psychology Norwegian University of Science and Technology, Trondheim, Norway The article, Neurodiversity in Saudi Arabia: Towards quality education and reduced inequalities, aims to consider the extent to which special education laws and policies in the Kingdom of Saudi Arabia are supportive of the concept of neurodiversity and individuals with such conditions via a content analysis of published articles about special education in the country. The study’s premise raises two questions for me. First, if neurodiversity is a relatively recent conceptualization of the conditions experienced by a range of individuals, is it fair, in a sense, to judge whether laws and policies created before either the development or at least the increased attention to the conceptualization are amenable to the conditions? Also, wouldn’t a content analysis of the country’s laws and policies themselves be a more direct assessment of neurodiversity acceptance and support than studies about special education for which they judge the studies’ authors’ perspectives about KSA’s laws and policies (if even such perspectives could be accurately determined in all cases)? Author Response: Thank you, Keller. The two questions you raised are quite reasonable. Regarding the first question, it is indeed unfair to consider given the novelty of the concept. As for the second question, you are correct; however, we opted to discuss laws and policies in the introduction (literature review) and concentrate on the data from existing studies. This approach was motivated by the fact that, as you pointed out, the concept is not utilized in the Saudi context. Consequently, we applied inclusion and exclusion criteria to draw implications for the use of this concept. In light of your questions, we have acknowledged this as a limitation. In a number of instances, sufficient details were not provided about the study’s methodology and analysis to make the research replicable. For instance, the narrative says that 306 potential studies were found initially yet the PRISMA flowchart starts with 112. Why is there a discrepancy? No information is provided about the instrument used as part of the content analysis; its inclusion as one of the supplemental documents would have been useful (and if the instrument were the very detailed PRISMA flowchart in the supplemental materials, say so). (The authors did do a good job in providing such resources in the OSF Registry for this study.) The authors say that “the extracted information was systematically categorized into a conceptual framework” (p. 6) but no information is provided about such a systematic approach. I would urge the authors to provide many more details in any revision of the current article. Providing citations, too, for the specific content analytical approaches that were used would help other researchers conduct similar work if they were inclined. Author Response: Thank you. The number of studies has been updated. A clarification has been added to the instruments section. This is due to the journal's policy of not including tables within the manuscript, which necessitates a clearer presentation of tables and data. We have included four tables and the PRISMA checklist in the OSF, as detailed below. Table 2.docx Data 1 Special education laws KSA neurodiversity KSA Alduais et al..docx Data 2 Studies in special education laws in the KSA neurodiversity KSA Alduais et al..docx Table 1.docx PRISMA_2020_checklist neurodiversity KSA Alduais et al..docx We replaced the word 'systematically' with 'thematically' because our study is not actually a systematic review. I would also suggest that the authors support their statements of the results with very specific details from the studies they cite in support of the results. That could strengthen the validity of their conclusions, moving beyond their present statements of generalities. Author Response: Thank you, Keller. Your emphasis on the necessity of presenting primary evidence rather than inferences is well-taken. Indeed, if you review the four tables that the journal instructed us to publish in OSF, as per their policy, you'll find all the details there. They have chosen to include only the main findings in the main manuscript. In essence, we were directed to relocate all data and tables to the extended data section in OSF. Acknowledging the value of your points, we have made a reference for readers to this perspective. https://osf.io/tkqg6 Finally, in a revision, I would remove the many adverbs and adjectives that the authors use to present their work in a positive light, e.g., meticulously, comprehensive, thorough, nuance, and valuable, amongst others. A reader assumes that, in a published article, such qualities informed the research. To frequently state this, though, can raise doubt in the reader’s mind. Author Response: Thank you. We have eliminated as many adjectives and adverbs as possible." } ] } ]	1	https://f1000research.com/articles/13-1062
https://f1000research.com/articles/13-1509/v1	09 Dec 24	{ "type": "Study Protocol", "title": "Mapping evidence of postgraduate research supervision experiences at higher education institutions in Africa: A scoping review protocol", "authors": [ "Celenkosini Nxumalo", "Tina Roberts", "Abdel-Hakeem Mohamed", "Janene Marais", "Noluthando Mayedwa", "Tina Roberts", "Abdel-Hakeem Mohamed", "Janene Marais", "Noluthando Mayedwa" ], "abstract": "Background The evolving landscape of the African higher education sector necessitated by global higher education reforms and the need for research informed responses to address contextual challenges is partly dependent on effective postgraduate research supervision. A wide range of postgraduate research supervision models and practices are documented globally. In the African context, the experiences of postgraduate research supervision are not clearly understood from the experiences of all stakeholders.\n\nAim The proposed scoping review aims to map evidence on the range of postgraduate research supervision experiences at higher education institutions in Africa.\n\nMethods The methodological framework proposed by Arksey and O’Malley will be used to conduct the review. Relevant articles will be sourced from different electronic data bases namely; ERIC, Web of Science, EbscoHost and Scopus. A combination of search terms and Boolean operators will be used to obtain the relevant articles based on the inclusion criteria stipulated within the Population, Context and Concept (PCC) framework of study identification for this review.\n\nResults Studies reporting on postgraduate research supervision experiences from the perspective of academics, students and support staff in the African higher education context will be included. The results may also provide insights postgraduate supervision experiences from varying postgraduate research disciplines.\n\nConclusion The findings will provide consolidated empirical evidence on experiences of postgraduate supervision in the African higher education context, thereby providing holistic insight on postgraduate research supervision practices, models currently used, implementation challenges and highlight research gaps. This may inform recommendations that advance knowledge within this field, particularly in the African higher education context.", "keywords": [ "Higher education", "Postgraduate supervision", "Research pedagogy", "Research supervision experiences" ], "content": "Background\n\nPostgraduate research supervision is a formal process of academic development and support that is an integral component of successful postgraduate studies and professional development in higher education Holtman and Mukwada.1 Early definitions of postgraduate supervision described it as an active process of teaching, support and mentorship of students pursuing postgraduate studies towards recognised qualifications at honours, masters and doctoral levels.2,3 Pearson and Brew4 state that the research supervision process helps students evolve into independent researchers and scholars, equipped to work in various settings, whether in academia, research or industry. The postgraduate supervision process thus requires a blend of academic and interpersonal skills. This includes guiding students through the research process by providing insights on proposal preparation, methodological choices and publication of research findings. Nulty, Kiley5 state that research supervisors are responsible for providing the academic guidance to ensure scientific merit of research studies. Moreover, they should also create an environment of holistic support that encourages successful completion of the research.6\n\nApproaches to postgraduate supervision vary widely and include traditional one-on-one mentorship, group supervision, cohort supervision, and more recently, digital or distance supervision.7 These models reflect an adaptation to the evolving needs of the global student body and the increasing demand for flexibility in higher education. The traditional model of supervision encompasses the provision of structured guidance and mentorship by a single supervisor who provides one-on-one advice, feedback, and support throughout the research process.8 This model emphasizes a personal and directive relationship, focusing on developing the student’s skills and knowledge in a specific field. While this approach can be beneficial, it may lead to students becoming overly dependent on their supervisors for guidance throughout the research process.\n\nTeam supervision, involves multiple supervisors contributing diverse expertise and perspectives to the postgraduate research process.9,10 This model benefits students by offering a broader base of knowledge and support, encouraging interdisciplinary thinking and potentially leading to more innovative research outcomes. Team supervision is useful in programs where students are working on related topics as it allows for peer mentoring and support.11 This subsequently instils a culture of collaboration and peer led critical engagement. Similarly, the participatory or co-supervision approach involves students more actively in the supervision process, emphasizing their role in setting goals, decision-making, and managing their research projects.12 This approach aims to develop students’ independence and self-regulation skills, preparing them for future academic or professional roles.13 Each of these models can be adapted to the specific needs of the student and the requirements of the program, ensuring that the supervision process is supportive, effective, and conducive to producing high-quality academic work. Likewise, distance supervision is becoming increasingly prevalent with the rise of digital learning platforms and advancement of technology as it allows for supervision via online platforms.14 This method is beneficial for students who are remote or part-time, providing flexibility while maintaining rigorous academic oversight through regular virtual meetings and the use of digital communication tools.\n\nThe role of postgraduate supervisors in ensuring quality research and successful postgraduate outcomes, cannot be overemphasised.15 In recent years, postgraduate research supervision has become a topical matter of discussion in the global higher education landscape.14 In regions like Europe and Australia improved training and monitoring of supervisors has been Initiated.16–18 Moreover, significant policy changes have been made to regulate the responsibilities of supervisors. International trends in postgraduate supervision inclusive of certain regions in Africa, are increasingly advocating for a shift towards more collaborative and integrative approaches. These include cohort supervision, peer mentoring, and integrated frameworks that combine various elements of traditional and modern supervisory models.19 Seiden Hyldegård and Jensen20 support the notion of group supervision as a means to enhance supervision skills, which can positively influence postgraduate students’ writing. The advantages of group supervision include providing support, access to resources, quick thinking practice, encouragement to write, and professional growth opportunities.21\n\nDespite this growing paradigm shift and inherent focus on university education and training globally, African universities vary in practices related to the adoption of systematic and current approaches to postgraduate supervision. There is evidence to suggest that the practice of postgraduate supervision in Africa is largely rooted in the traditional, individualistic model.22 Moreover, the successful adoption of alternative and new approaches to postgraduate supervision remains hindered by contextual challenges related to availability of resources, funding and high staff-student turn over.23 Conversely, Pillay and Balfour24 have alluded to finding variety of postgraduate research supervision practices and models which arise from supervisors ideological commitments and personal experiences. In such instances, there is exists a continuum - from a traditional model where there is deep, intense attachment to the one-to-one relationship on one end. On the other end may also exist hybrid model where-in the supervisory practices may be reconfigured based on varying contextual dynamics.\n\nIn the South African context, there has been substantial growth in postgraduate enrolment rates since 1994, however this has not reflected in graduations of doctoral candidates, which are still lower than the rates expressed in the National Development Plan 2030 (NDP).25 Postgraduate supervision remains pivotal to the successful graduation of students from their postgraduate program.26 The changing landscape in higher education created new sets of challenges for the supervision process, which includes work which are more interdisciplinary and students requiring expertise from supervisors that they may lack. The current supervision model employed in South Africa is more of a traditional apprenticeship model, characterised by one-on-one supervision, however there is a call to move more towards collaborative supervision approaches such as group or team supervision.25 The changing landscape in higher education in South Africa has led to a set of challenges experienced by both the supervisor and student in the supervision relationship.\n\nSupervisors in South Africa face many challenges in relation to postgraduate supervision. Some of the challenges include; lack of adequate training and support; reliance on their own supervision experience as a means of preparation; heavy workloads on top of supervision expectations; the massification of postgraduate education leads to challenges in maintaining quality supervision while simultaneously increasing the quantity of throughput; time constraints lead to supervisors not having adequate time for reflection, personal development or attending developmental opportunities; all of these challenges faced by supervisors raises concerns and influences the quality of supervision they are able to provide.27,28 In light of these challenges and the changing landscape of postgraduate supervision needs, there is a call for structured support, training and development of supervisors26,28\n\nGiven the current state of challenges that supervisors experience in South Africa, it is not surprising that postgraduate students would also have their own set of challenges. Some of the challenges raised by postgraduate students in relation to supervisions include; the poor communication between the supervisor and student, this is especially true for students who are situated further away from the university and in rural areas. Students also feel that the feedback that they receive from their supervisors are not adequate to their needs or in some cases, amount of lack of feedback. Students also often feel that they do not receive enough support from their supervisors during the course of the program. Some of the more concerning challenges highlighted by students are the unethical practices of supervisors.26\n\nWhile substantial evidence exists on postgraduate supervision experiences from the South African context, there is a need for greater awareness on postgraduate supervision experiences in other contexts of the African continent. Moreover, the experiences of postgraduate research supervision are not clearly understood from the experiences of all academic stakeholders involved in the supervision process, this includes postgraduate masters and doctoral students, academics and research support staff. This scoping review thus seeks to consolidate evidence on the experiences of postgraduate supervision in the African higher education context. The mapping of this empirical evidence may generate a more comprehensive understanding on the range of postgraduate supervision practices adopted in the region. Additionally, it may provide deeper insights on contextual barriers, success and best practices related to the adoption of various postgraduate supervision models in the region.\n\n\nMethods\n\nThis scoping review will map evidence of postgraduate supervision experiences at higher education institutions in Africa. The scoping review methodological framework by Arksey and O’Malley29 will be used to guide the review. The review will thus comprise of the following steps as guided by the framework: (1) identification of the review question, (2) identification of the relevant studies, (3) selection of relevant studies, (4) charting the data, (5) collating, summarizing and reporting the results.\n\nThe main review question will be: What is the existing literature on postgraduate supervision experiences at higher education institutions in Africa?\n\n\n\n1. What is the range of literature on postgraduate supervision experiences among students and academics at higher education institutions in Africa?\n\n2. What is the existing evidence on postgraduate supervision practices at higher education institutions in Africa?\n\n3. What are the reported challenges, recommendations and best practices on postgraduate supervision at higher education institutions in Africa?\n\nThis review will use the Population, Concept and Context (PCC) framework to align the study selection process with the review question and sub-questions. See Table 1 for PCC framework and proposed alignment.\n\nKeywords combined with Boolean operators will be used to identify relevant articles in various online data bases and search engines. These search engines will include ERIC, Web of Science, EbscoHost, Scopus and African Journals online. The identification of articles will be facilitated through a search strategy that will include a combination of terms. See Table 2. For search strategy. A pilot search will be conducted to ascertain the validity of the proposed strategy.\n\nSearch results of electronic articles to be will be exported to Covidence Five independent reviewers will the carryout the title, abstract and full text screening whilst also removing duplicates that were not identified by Covidence. Articles that meet the inclusion criteria will be included for full text review and independent screening will be done by the five reviewers. Consensus will be established by discussion among the reviewers and an arbitrator will be sought in instances of disagreement. The articles selection process will be documented using the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) flow chart – see Figure 1. The selection of eligible articles will be guided by the following inclusion and exclusion criteria:\n\n\n\n▪ Primary studies focusing on students and academics at higher education institutions in Africa.\n\n▪ Studies conducted between 2014 and 2024.\n\n▪ Studies published in English only.\n\n\n\n▪ Grey literature, systematic reviews, commentaries, opinion articles and secondary data.\n\n▪ Studies that do not differentiate between undergraduate and postgraduate studies.\n\n▪ Studies conducted outside of Africa.\n\n▪ Studies on vocational and non-degree postgraduate training.\n\nAn electronic data charting template will be used to capture articles that will be included in the review as guided by the inclusion and exclusion criteria and the PCC framework. The data charting template will be developed by the authors and guided by the contents of existing data charting tools. The tool will thus comprise of the following elements: (1) Author, (2) year of publication, (3) context, (4) study population characteristics, (5) design (6) key findings. Table 3 provides representation of the proposed data charting template to be used.\n\nA narrative report will be used to summarize the key findings from the extracted research. The emerging results will be presented in relation to the review question and sub-questions and will be aligned to the PCC framework. The studies will be examined using a narrative synthesis approach. Narrative synthesis allows for the presentation of results that are diverse in nature and study designs.30 The study will use three out of the four key elements proposed by Popay and colleagues (2006) to describe synthesize the evidence on postgraduate supervision experiences at higher education institutions in Africa. The key elements that will be used are; (1) developing elements of preliminary synthesis of findings of included studies, (2) exploring relationships within the data and, (3) assess the robustness of the synthesis.31\n\n\nDiscussion\n\nThe aim of this scoping review is to map and synthesize evidence on postgraduate supervision experiences at higher education institutions in Africa. Presently research on postgraduate supervision experiences in countries like Malaysia and Pakistan allude to student’s challenges navigating the research process, models of research supervision adopted by academics and the need for understand the range of available postgraduate supervision approaches required for different students’ needs.32,33 In the African context empirical evidence reports on experiences of post graduate supervision from the perspective on students, particularly those at PhD level and supervisors within specified postgraduate research disciplines.34 There is also evidence to suggest varying practices and challenges related to postgraduate research supervision processes and practices.35 Moreover contextual research highlights the need for collaborative efforts my stakeholders to ensure quality of research supervision practices so that postgraduate students can contribute significantly to addressing societal issue through their research.36 Tlali, Chere-Masopha37 further argue that supervisors should be capacitated through relevant training to execute the role and that supervisory guidelines be reviewed and benchmarked against best practices in the field.\n\nDespite the availability of reasonable empirical data on this subject, experiences of postgraduate research supervision are not holistically understood from the perspective of all relevant stakeholders within the African higher education context. The proposed review thus seeks to address this gap by mapping the range of empirical evidence on postgraduate supervision experiences from various academic stakeholders namely postgraduate Masters and PhD students, research supervisors and research support staff.\n\nIn this regard, it hoped that the review may highlight several themes associated with postgraduate supervision experiences, including supervisor-student relationships, different models of supervision practices, and barriers from the different disciplines and fields of postgraduate research. It is also anticipated that the review will highlight gaps in the current literature, potentially leading to further research areas. The proposed findings may thus have implications for policy and practices related to postgraduate supervision in the African higher education context. This is significant in the African context due to a rapidly advancing higher education landscape and the role of high-quality supervision in the success of postgraduate students.\n\nEthics and consent were not required.", "appendix": "Data availability\n\nNo data are associated with this aricle.\n\n\nReferences\n\nHoltman L, Mukwada G: Challenges confronting the quality of postgraduate research supervision and its effects on time-to-degree and throughput rates: A case of a South African University. Mediterr. J. Soc. Sci. 2014; 5(6): 179–190.\n\nWisker G: The good supervisor: Supervising postgraduate and undergraduate research for doctoral theses and dissertations. Bloomsbury Publishing; 2012.\n\nSinclair M: The pedagogy of’good’PhD supervision: A national cross-disciplinary investigation of PhD supervision. Department of Education, Science and Training Canberra; 2004.\n\nPearson M, Brew A: Research training and supervision development. Stud. High. Educ. 2002; 27(2): 135–150. Publisher Full Text\n\nNulty D, Kiley M, Meyers N: Promoting and recognising excellence in the supervision of research students: an evidence-based framework. Assess. Eval. High. Educ. 2009; 34(6): 693–707. Publisher Full Text\n\nGanqa NH: Research supervision experiences of masters in education students at a South African University. University of Fort Hare; 2012.\n\nBitzer EM, Albertyn RM: Alternative approaches to postgraduate supervision: A planning tool to facilitate supervisory processes. S. Afr. J. High. Educ. 2011; 25(5): 875–888.\n\nMcCallin A, Nayar S: Postgraduate research supervision: A critical review of current practice. Teach. High. Educ. 2012; 17(1): 63–74. Publisher Full Text\n\nWatts JH: Team supervision of the doctorate: Managing roles, relationships and contradictions. Teach. High. Educ. 2010; 15(3): 335–339. Publisher Full Text\n\nvan Biljon J , Pilkington C, van der Merwe R : Cohort supervision: Towards a sustainable model for distance learning. Annual Conference of the Southern African Computer Lecturers’ Association. Springer; 2019.\n\nRobertson M: Aspects of mentorship in team supervision of doctoral students in Australia. Aust. Educ. Res. 2017; 44: 409–424. Publisher Full Text\n\nWald N, Kumar V, Sanderson LJ: Enhancing co-supervision practice by setting expectations in a structured discussion using a research-informed tool. High. Educ. Res. Dev. 2023; 42(3): 757–769. Publisher Full Text\n\nMaor D, Fraser BJ: Design of a participatory supervision support platform for improving higher degree supervision: A feasibility study. Office for Learning and Teaching; 2015.\n\nNasiri F, Mafakheri F: Postgraduate research supervision at a distance: A review of challenges and strategies. Stud. High. Educ. 2015; 40(10): 1962–1969. Publisher Full Text\n\nHalse C, Malfroy J: Retheorizing doctoral supervision as professional work. Stud. High. Educ. 2010; 35(1): 79–92. Publisher Full Text\n\nButtery EA, Richter EM, Leal Filho W: An overview of the elements that influence efficiency in postgraduate supervisory practice arrangements. Int. J. Educ. Manag. 2005; 19(1): 7–26.\n\nHutchinson S, Lawrence H: Enhancing the doctoral experience: A guide for supervisors and their international students. Routledge; 2017.\n\nO’Carroll C, et al.: The PhD in Europe: Developing a system of doctoral training that will increase the internationalisation of universities. European Higher Education at the Crossroads: Between the Bologna Process and National Reforms. 2012; pp. 461–484. Publisher Full Text\n\nManderson L, et al.: Enhancing doctoral supervision practices in Africa. Journal of Higher Education in Africa/Revue de l’enseignement supérieur en Afrique. 2017; 15(2): 23–40.\n\nSeiden Hyldegård J, Jensen HN: The implied peer: thesis writers’ feedback activities and experiences in group supervision. Stud. High. Educ. 2023; 48(11): 1754–1766. Publisher Full Text\n\nNgulube P: Postgraduate supervision practices in education research and the creation of opportunities for knowledge sharing. Problems of Education in the 21st Century. 2021; 79(2): 255–272. Publisher Full Text\n\nUkwoma SC, Ngulube P: Supervision Practices in Library and Information Science Postgraduate Research in Nigeria and South Africa. Afr. J. Libr. Arch. Inf. Sci. 2020; 30(2).\n\nSsentamu PN, Sajjabi FB: Supervisors’ Reflections on Policy and Practice in an African Graduate Setting: Towards a Pedagogy for Supervising Research, in Postgraduate Research Engagement in Low Resource Settings. IGI Global; 2020; 264–295.\n\nPillay G, Balfour RJ: Post-graduate supervision practices in South African universities in the era of democracy and educational change 1994-2004. S. Afr. J. High. Educ. 2011; 25(2): 358–372.\n\nHarmon QE: The burden of uterine fibroids: a search for primary and secondary prevention. Fertil. Steril. 2019; 111(1): 50–51. PubMed Abstract \| Publisher Full Text\n\nCekiso M, et al.: Supervision experiences of postgraduate research students at one South African higher education institution. S. Afr. J. High. Educ. 2019; 33(3): 8–25. Publisher Full Text\n\nMotshoane P: The need for the development of emerging postgraduate supervisors. J. New Gen. Sci. 2023; 21(1): 74–85. Publisher Full Text\n\nMasuku VZ: Becoming a research supervisor: Reflections on a postgraduate supervision course. Teacher Education and Curriculum Studies. 2021; 6(4): 143–150. Publisher Full Text\n\nArksey H, O’Malley L: Scoping studies: towards a methodological framework. Int. J. Soc. Res. Methodol. 2005; 8(1): 19–32. Publisher Full Text\n\nGuise V, Anderson J, Wiig S: Patient safety risks associated with telecare: a systematic review and narrative synthesis of the literature. BMC Health Serv. Res. 2014 Dec; 14: 1–5. 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 1. 2006; 1–93. Publisher Full Text\n\nSidhu GK, et al.: Postgraduate Supervision: Exploring Malaysian Students’ Experiences. Procedia. Soc. Behav. Sci. 2013; 90: 133–141. Publisher Full Text\n\nAli J, Ullah H, Sanauddin N: Postgraduate Research Supervision: Exploring the Lived Experience of Pakistani Postgraduate Students. FWU Journal of. Soc. Sci. 2019; 13(1).\n\nOmoya O, et al.: A scoping review protocol of the lived experiences of doing a PhD in Africa. PLoS One. 2023; 18(9): e0290701. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDaramola O: Lessons from postgraduate supervision in two african universities: An autoethnographic account. Educ. Sci. 2021; 11(7): 345. Publisher Full Text\n\nKomba SC, Chiwamba SV: Towards improving the supervision of postgraduate students in Tanzanian higher learning institutions. Int. J. Res. Stud. Educ. 2017; 6(2): 29–40.\n\nTlali T, et al.: Challenges Confronting Postgraduate Supervision at the National University of Lesotho:: Supervisors’ Perspectives. Int. J. Afr. High. Educ. 2022; 9(2): 22–39. Publisher Full Text" }	[ { "id": "359019", "date": "07 Feb 2025", "name": "Kathy Luckett", "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 scoping review outlined in this protocol is well motivated and by the authors and will fill an important gap in the literature on postgraduate supervision in Africa for several reasons: review studies in HE are typically conducted with the institution/ discipline/ nation state as the unit of analysis, there is a paucity of regional reviews; to my knowledge few studies focus on experience (as opposed to outcomes or results) and it is rare to find studies that include support staff as participants. Thus the aim to provide “holistic insight” on this topic could be a significant contribution to the field.\nOverall, the design of the scoping review is adequate to the research questions and robust and replicable.\nHowever, the ambitions scope of the review does raise a few concerns for me. Below I list a few points that may be useful to the researchers to consider prior to embarking on the review:\nThe authors’ focus on ‘experience’ is deliberate, but I did wonder whether they should also provide some clarity around defining key concepts – either before embarking on the literature search and/ or as a result of their findings. Relatedly, the focus on ‘experience’ could prove to be a limiting factor on their findings. I say ‘may’, because I could well be wrong. However, in my view ‘practice’ and ‘experience’ are closely intertwined, while policy (at both national and institutional levels) tends to both enable and constrain practice/experience. The realities of these kind of everyday slippages point to the need for conceptual clarity and great care when defining key words as well as awareness of potential limitations on the review.\n\nAlso related to conceptual clarity, I wondered how the authors intend to define ‘support staff’ and whether they need to give thought to this prior to conducting the literature search. There are a range of support and professional staff involved in the supervision process, for example administrative staff at different levels, professional development staff who provide services and training in writing, editing, supervision, ethics and research integrity, to both academics and students, as well as librarians and others who provide support around funding, tracking enrolments, progress, graduation rates and quality assurance.\n\nWhile the authors note the range of models used for supervision, they do not seem to consider different types of qualification, for example professional doctorates, badged doctorates, doctorates by publication, coursework doctorates/ masters, full research degrees, M. / D.Phil etc. I wondered whether the range of types of qualification should be taken into account in the search criteria (as either inclusion or exclusion criteria).\n\nRegarding Stage 2 (Identification of relevant studies) the emphasis is on electronic databases. However, Arksey & O’Malley refer to three additional data sources that seem to be lumped under ‘additional records identified through other sources’ in the PRISMA diagram. While noting that the review will look at only primary research data, I think rich data sources could be retrieved from secondary sources that are based on primary data such as the South Africa Council on Higher Education’s “Doctoral Degrees National Report” (2022).\nFinally, some minor corrections: Grammar: subject-verb agreement. Typos – e.g. in Discussion section – eg on-of, my-by, aricle-article,\nError in Reference List\n25. Harmon QE: The burden of uterine fibroids: a search for primary and secondary prevention. Fertil. Steril. 2019; 111(1): 50–51. PubMed Abstract \| Publisher Full Text\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/13-1509
https://f1000research.com/articles/13-1508/v1	09 Dec 24	{ "type": "Systematic Review", "title": "Platelet-rich plasma as a potential therapy of cystocele: a systematic review and meta-analysis", "authors": [ "Edy Fakhrizal", "Budi Santoso", "Eighty Mardiyan Kurniawati", "Fedik Abdul Rantam", "Salmon Charles P. T. Siahaan", "Jojor Sihotang", "Arya Marganda Simanjuntak", "Raehan Satya Deanasa", "Putri Mahira Afladhanti", "Edy Fakhrizal", "Eighty Mardiyan Kurniawati", "Fedik Abdul Rantam", "Salmon Charles P. T. Siahaan", "Jojor Sihotang", "Arya Marganda Simanjuntak", "Raehan Satya Deanasa", "Putri Mahira Afladhanti" ], "abstract": "Abstract\nBackground Platelet-rich plasma (PRP) therapy, is known for its regenerative properties. PRP may be able to help with cystocele cases. This review aims to explore how it might be applied to urogynecological cases, particularly those involving cystocele.\n\nMethods The present systematic review adhered to the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guideline 2020, Meta-Analysis of Observational Studies in Epidemiology (MOOSE) guideline. The present systematic review was registered with PROSPERO with the registered number CRD42023414366. PubMed, Science Direct, Epistemonikos, COCHRANE, Google Scholar, and ProQuest were among the databases we searched. “(PRP OR Platelet-Rich Plasma) AND (Cystocele OR Anterior Pelvic Organ Prolapse)” from January 2007 to December 2022. Based on the PICO framework (Population = Patients with Cystocele; Intervention = Reconstruction with platelet-rich plasma injection; Compare = Reconstruction only; Outcome: Recurrency of Cystocele), four investigators (AMS, PMA, EAU, RSD, and AH) independently assessed eligibility by titles and abstracts. Using the Joanna Briggs Institute Critical Appraisal tool, each author evaluated full-text articles based on the kind of articles they had received. When consensus could not be obtained, disagreements were settled by involving the supervisors (EF, EMK, and BS).\n\nResults A total of 8,924 studies were identified. After removing duplicates and applying eligibility criteria, two articles were included, encompassing 65 patients. In two studies, PRP injections were found and administered post-anterior colporrhaphy at the pubovesical fascia. According to these two trials, women who had anterior colporrhaphy and PRP injections required fewer reoperations.\n\nConclusion PRP has the potential to be a good alternative treatment to prevent cystocele recurrence. However, it cannot be generalized to large populations due to the small number of findings. Further studies with large samples examining the efficacy and safety of the therapy are needed to prove it.", "keywords": [ "anterior vaginal prolapse", "cystocele", "platelet-rich plasma", "potential therapy", "recurrence" ], "content": "Introduction\n\nPelvic organ prolapse (POP) refers to the descent of pelvic organs, such as the colon, small intestine, urethra, or bladder, into or beyond the vaginal opening.1 POP affects an estimated 40% of women globally, with a higher prevalence in low-income countries and increasing severity with age.2 Cystocele, the protrusion of the bladder into the vagina, is the most common type of POP, constituting 81% of the 200,000 annual POP surgeries in the United States of America (USA).3,4 Risk factors include previous pregnancies, physically demanding work, and genetic disorders.3 Kayondo et al found that 56.3% of recurrences occurred at the same site in 25.2% of patients, with 80% of these being cystoceles.5\n\nAn anterior corporal reconstruction is a treatment option for cystocele; however, it primarily reinforces the vaginal wall and does not adequately address the apical compartment.1,3 To address these deficiencies, platelet-rich plasma (PRP) therapy, known for its regenerative properties, offers a potential solution.6 PRP, which contains growth factors and cytokines, is used in orthopedic and cosmetic surgery for tissue repair and regeneration.6 In cases of POP, the use of autologous regenerative substances may facilitate the repair of connective tissue, smooth muscle, vascularization, and innervation supporting the pelvic organs.6 PRP injection aims to reinforce pelvic support structures, strengthen ligaments, and restore normal anatomical function, thereby reducing the risk of POP recurrence.6 This systematic review aimed to explore the effect of PRP in treating cystocele, an area less frequently addressed compared to orthopedic and cosmetic applications. To the best of our knowledge, the present systematic review and meta-analysis is the first to assess platelet-rich plasma as a potential alternative for preventing cystocele recurrence.\n\n\nMethods\n\nThe present systematic review adhered to the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guideline 2020, Meta-Analysis of Observational Studies in Epidemiology (MOOSE) guideline. The present systematic review was registered with PROSPERO with the registered number CRD42023414366.\n\nRecurrence in pelvic organ prolapse that has been reconstructed surgically shows a high rate. So the incidence of re-operation also increases. This is certainly uncomfortable for patients, so the author has a research question whether there is a method or further therapy to reduce the recurrence rate of pelvic organ prolapse. Platelet Rich Plasma is known to have a good anti-inflammatory and tissue repair profile and has been used by various specialist fields. So our research question is whether PRP can be an adjunct therapy to reduce the recurrence of pelvic organ prolapse.\n\nPopulation, intervention, comparison, and outcome (PICO) framework was employed to evaluate the effect of PRP injections in treating cystocele. The population of interest includes patients diagnosed with cystocele, a condition where the bladder protrudes into the vaginal wall. The intervention being assessed is PRP injection, a regenerative therapy designed to enhance tissue repair and strengthen pelvic support structures. This intervention is compared to the standard treatment of surgical reconstruction alone. The primary outcome is the recurrence of cystocele, with the study comparing the effectiveness of PRP combined with reconstruction versus reconstruction alone in preventing cystocele recurrence. The included studies were full-text and in English or Indonesian. Reviews /Case Reports/Experimental Studies, unpublished data, and duplicate studies were excluded. References from included studies were screened for additional eligible papers.\n\nArticles were sourced from PubMed, Science Direct, Epistemonikos, COCHRANE, Google Scholar, and ProQuest for the period January 2007 to December 2022, using the keywords (PRP OR Platelet-Rich Plasma) AND (Cystocele OR Anterior Pelvic Organ Prolapse). Four investigators (A.M.S., P.M.A., E.A.U., R.S.D., and A.H.) independently assessed study eligibility based on titles and abstracts. Disagreements were resolved through consultation with supervisors (E.F., E.M.K., and B.S.).\n\nEach study was reviewed for details including author, year, design, country, population sample, GPAH, intervention, PRP cycle, dose (mL), injection site, cystocele recurrence, and observation length.\n\nEach author assessed potential bias using the Joanna Briggs Institute’s critical appraisal tools (jbi.global/critical-appraisal-tools). Bias was quantified by calculating the percentage of affirmative responses to appraisal questions. A graph depicting each author’s evaluation for each article was then generated. Disagreements were resolved with input from supervisors (EF, BS, EMK, and FAR).\n\nLicensed RStudio was employed for meta-analysis utilizing a random-effects model. The use of a random effects model was chosen so that the results could be interpreted generally outside the included sample population. Findings were reported as proportions, with heterogeneity measured using I2, Q value, Degrees of freedom (df ), Tau-squared, and Tau and p-value. Lower I2 values and p-values greater than 0.05 indicate less heterogeneity. Lower I2 values and p-values greater than 0.05 indicate less heterogeneity.\n\n\nResults*\n\nA total of 8,924 studies were identified. After removing duplicates and applying eligibility criteria, two articles were included, encompassing 65 patients ( Figure 1). References from these studies and citations yielded no additional eligible papers. Atilgan et al compared PRP with cystocele reconstruction (colporrhaphy), while Einarsson et al focused solely on PRP in cystocele patients without a control group.\n\nAtilgan et al conducted an observational study in Turkey with 56 women with cystoceles. Group 1 underwent anterior colporrhaphy alone, while Group 2 received reconstruction plus PRP injection. Over 48 months, group 2 showed a statistically significant greater reduction in prolapse symptoms (p = 0.002) and fewer women requiring surgery compared to group 1. In group 1, 21.4% had a symptomatic recurrence, and 17.8% needed surgery, whereas, in group 2, 7.1% had a symptomatic recurrence, and only 3.5% required repair (p = 0.0001).6 Einarsson et al conducted a pilot trial in the USA using autologous platelet gel (a PRP by-product) applied to the surgical site during anterior colporrhaphy. Among nine patients, only one (12.5%) required a second procedure after ten months.7 In all included studies, the follow-up after PRP injections was 48 months and 20 months ( Figure 2). There is a substantial difference between the research by Atilgan and Einarrson, namely the research arm used. Atilgan used a non-PRP comparator while Einarsson used a single-arm with PRP only. Therefore, a side-to-side comparison between PRP and Non-PRP can only be seen in the Atilgan study. Therefore, a meta-analysis comparing the two therapies could not be performed in this study but continued with a proportional meta-analysis of the incidence of reoperation in patients who had been treated with PRP. The next difference is that the follow-up of the study is longer in Atilgan (48 months) compared to Einarsson (20 months) so the incidence of effects or success can be more convincing by Atilgan because it uses a longer observation time.\n\nAll studies reviewed scored above 74.5%, indicating minimal bias in the included research ( Figure 3). This assessment was found because both studies did not include confounding factors that may be involved in the study and also strategies against confounding factors were not described. Atilgan has a higher score than Einarsson because it has a comparison group and more optimal measurements than Einarsson who only used single-arm studies, but it can still be included because it discusses the same thing, namely the use of PRP in Pelvic Organ Prolapse cases.\n\nA proportional meta-analysis was performed to evaluate the current evidence from studies that have investigated the use of PRP in cases of pelvic organ prolapse (POP). Additionally, a proportional meta-analysis was conducted to assess the incidence of reoperation following PRP therapy. The analysis revealed that reoperation occurred in 4 out of 100 patients (95% CI: 0.00-0.11, p = 0.46) who underwent reconstruction followed by PRP treatment. This suggests that while reoperation is possible, its incidence remains low. The study’s heterogeneity was minimal, as indicated by a low I2 value (0%) and a low τ2 value (0%), reflecting minimal variability between studies. However, this finding should be interpreted with caution, as it may be influenced by the small number of studies that have specifically tested PRP in POP patients. ( Figure 4).\n\n\nDiscussion\n\nFrom the meta-analysis, the reoperative incidence following PRP treatment was 4%. Atilgan et al reported a significant reduction in recurrence incidence (p = 0.002) and fewer reoperations in the PRP group.6 Einarsson et al observed a low reoperation rate, with only 1 of 9 women requiring a second procedure after 10 months of PRP treatment.7 Both studies indicate a low reoperation rate with anterior colporrhaphy and PRP, although Einarsson et al did not compare it to reconstruction alone.6,7\n\nNone of the studies reported significant adverse effects from PRP treatment.15 Minor side effects, such as transient erythema, edema, mild headache, itching, and brief discomfort, were noted but did not include serious complications such as scarring, infection, or allergic reactions. Patients generally resumed normal activities and returned to work the following day without the need for antibiotics.15 Additionally, allogeneic PRP was found to be immunologically safe, and studies on chronic wounds showed favorable outcomes with no significant side effects.16,17\n\nPRP enhances fibroblast migration and proliferation, promoting connective tissue regeneration.8 In urogynecology, PRP is used for prolapse repair with favorable functional and anatomical outcomes.8 The primary benefit of PRP is its low recurrence rate. Additionally, PRP injections significantly reduce prolapse symptoms as measured by the Pelvic Floor Distress Inventory scale.8 Therefore, PRP is a promising alternative for preventing cystocele recurrence.8 Previous studies also suggested that PRP acts as a growth factor, facilitating tendon and ligament healing.8\n\nPlatelets promote angiogenesis through endothelial cell proliferation and migration.9 Previous studies have predominantly focused on PRP for tissue regeneration, highlighting its growth factors such as platelet-derived growth factor (PDGF), vascular endothelial growth factor (VEGF), insulin growth factor I (IGF-I), hepatocyte growth factor (HGF), transforming growth factor beta (TGF-b), and fibroblast growth factor (FGF), which are crucial for ligament reconstruction ( Figure 5).10,11 Recent studies have explored PRP for vaginal mesh exposure and improved fibroblast adherence and repair in vaginal prolapse surgeries.12,13 Although PRP has not been used for cystocele correction, it is proposed for injection into the pubovesical fascia to enhance support and prevent recurrence.\n\nTo the best of our knowledge, the present systematic review and meta-analysis is the first to assess platelet-rich plasma as a potential alternative for preventing cystocele recurrence. Considering the risks associated with mesh implants, PRP injection into the pubovesical fascia during anterior vaginal wall prolapse repair may offer a viable alternative to prevent cystocele recurrence.6 Currently, PRP is reported to have no recognized side effects, with no adverse events noted during the 48-month follow-up in Atilgan et al.6 However, the optimal frequency and number of PRP sessions remain unclear and vary across studies.14 Further large-scale randomized controlled trials are needed to confirm the safety and efficacy of PRP in urogynecological procedures.6,14\n\nThe present systematic review’s applicability is limited due to the small sample size and lack of a placebo-controlled comparison. Despite these limitations, the evidence supports PRP as a promising alternative for reducing cystocele recurrence. Further observational studies, clinical trials, and preclinical research are recommended to confirm PRP’s efficacy and safety in this context.\n\n\nConclusion\n\nOur findings indicate that PRP injections may serve as a promising alternative for preventing the recurrence of postoperative cystocele. Additionally, in female patients with cystocele, PRP injections have the potential to improve symptoms, enhance anatomical outcomes, and increase postoperative satisfaction. However, the current evidence supporting these outcomes is of low quality due to the limited number of published studies and small sample sizes. Consequently, further high-quality randomized controlled trials (RCTs) are necessary to determine whether PRP injections represent an optimal alternative.\n\n\nEthics and consent\n\nEthical approval and consent were not required.", "appendix": "Data availability statement\n\nNo data are associated with this article\n\nFigshare: Platelet-rich plasma as a potential therapy of cystocele: A systematic review and meta-analysis, https://doi.org/10.6084/m9.figshare.27187344.v1.18\n\nThis project contains the following underlying data:\n\n• PRISMA checklist and flowchart\n\n• the specific assessment of each included article for our Risk of Bias.\n\n• the metaprop program code that we used to conduct the meta-analysis test using the R application, and also other data that produced this meta-analysis article.\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 would like to thank the head chief department of Obstetrics-Gynecology at Riau University and Airlangga University.\n\n\nReferences\n\nPrawirohardjo S: Ilmu KandunganAnwar M, Baziad APR, editors. P.T. Bina Pustaka Sarwono Prawirohardjo.2011.\n\nWang B, Chen Y, Zhu X, et al.: Global burden and trends of pelvic organ prolapse associated with aging women: An observational trend study from to.\n\nFakhrizal E: Prolaps Organ Panggul. 1st ed.Pekanbaru: Taman Karya; 2021.\n\nBarber MD, Maher C: Epidemiology and outcome assessment of pelvic organ prolapse. Int. Urogynecol. J. 2013 Nov; 24(11): 1783–1790. Publisher Full Text\n\nKayondo M, Geissbüehler V, Migisha R, et al.: Risk factors for recurrence of pelvic organ prolapse after vaginal surgery among Ugandan women: a prospective cohort study. Int. Urogynecol. J. 2022 Jul; 33(7): 1933–1939. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAtılgan AE, Aydın A: Cystocele Repair with Platelet-Rich Plasma. Indian J. Surg. 2021; 83(3): 726–730. Publisher Full Text\n\nEinarsson JI, Jonsdottir K, Mandle R: Use of autologous platelet gel in female pelvic organ prolapse surgery: a feasibility study. J. Minim. Invasive Gynecol. 2009; 16(2): 204–207. PubMed Abstract \| Publisher Full Text\n\nGuevara-Alvarez A, Schmitt A, Russell RP, et al.: Growth factor delivery vehicles for tendon injuries: Mesenchymal stem cells and Platelet Rich Plasma. Muscles Ligaments Tendons J. 2014 Jul; 4(3): 378–385. PubMed Abstract \| Publisher Full Text\n\nOklu R, Walker TG, Wicky S, et al.: Angiogenesis and current antiangiogenic strategies for the treatment of cancer. J. Vasc. Interv. Radiol. 2010 Dec; 21(12): 1791–1805. quiz 1806. Publisher Full Text\n\nMartínez CE, Smith PC, Palma Alvarado VA: The influence of platelet-derived products on angiogenesis and tissue repair: a concise update. Front. Physiol. 2015; 6: 290.\n\nEl-Sharkawy H, Kantarci A, Deady J, et al.: Platelet-rich plasma: growth factors and pro- and anti-inflammatory properties. J. Periodontol. 2007 Apr; 78(4): 661–669. PubMed Abstract \| Publisher Full Text\n\nCastellani D, Valloni A, Piccirilli A, et al.: An innovative approach to treating vaginal mesh exposure after abdominal sacral colpopexy: endoscopic resection of mesh and platelet-rich plasma; initial experience in three women. Int. Urogynecol. J. 2017 Jun; 28(2): 325–327. PubMed Abstract \| Publisher Full Text\n\nSebastian M, May A, Hala K, et al.: Attachment of Primary Vaginal Fibroblasts to Absorbable and Nonabsorbable Implant Materials Coated With Platelet-Rich Plasma: Potential Application in Pelvic Organ Prolapse Surgery.2015; 21(4): 190–197.\n\nNikolopoulos KI, Pergialiotis V, Perrea D, et al.: Restoration of the pubourethral ligament with platelet rich plasma for the treatment of stress urinary incontinence. Med. Hypotheses. 2016 May; 90: 29–31. PubMed Abstract \| Publisher Full Text\n\nPaichitrojjana A, Paichitrojjana A: Platelet Rich Plasma and Its Use in Hair Regrowth: A Review. Drug Des. Devel. Ther. 2022; 16: 635–645. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAkbarzadeh S, McKenzie MB, Rahman MM, et al.: Allogeneic Platelet-Rich Plasma: Is It Safe and Effective for Wound Repair? Eur Surg Res Eur Chir Forschung Rech Chir Eur. 2021; 62(1): 1–9. Publisher Full Text\n\nSemenič D, Cirman T, Rožman P, et al.: REGENERATION OF CHRONIC WOUNDS WITH ALLOGENEIC PLATELET GEL VERSUS HYDROGEL TREATMENT: A PROSPECTIVE STUDY. Acta Clin. Croat. 2018 Sep; 57(3): 434–442. PubMed Abstract \| Publisher Full Text\n\nSiahaan SCPT: Platelet-rich plasma as a potential therapy of cystocele: A systematic review and meta-analysis. figshare. [Dataset]. 2024. Publisher Full Text" }	[ { "id": "348123", "date": "18 Dec 2024", "name": "Akbar Novan Dwi Saputra", "expertise": [ "Reviewer Expertise regenerative medicine", "urogynecology" ], "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\nAre the rationale for, and objectives of, the Systematic Review clearly stated? Partly\nExplanation:\nThe rationale regarding the importance of cystocele and its high recurrence rate is well presented. The potential role of PRP in tissue regeneration is adequately explained. However, the specific objectives need better definition:\nPrimary and secondary outcomes should be explicitly stated The research question could be more precisely formulated using the complete PICO framework The timing of PRP administration relative to surgery needs clearer specification\n\n2. Are sufficient details of the methods and analysis provided to allow replication by others? No\nExplanation:\nCritical methodological details are missing:\nComplete search strategies for all databases are not provided The 2007 - 2022 date restriction is not justified Screening process details are insufficient Data extraction forms are not included The method for contacting authors for missing data is not described The exact statistical methods for the proportional meta-analysis need more detail.\n\nTo make this replicable, authors must provide:\nFull search strategies for all databases Detailed screening criteria Complete data extraction templates Statistical analysis code or detailed methods\n\n3. Is the statistical analysis and its interpretation appropriate? No\nExplanation:\nThe statistical approach has several fundamental flaws:\nConducting a meta-analysis with only two studies, one being a single-arm study, is inappropriate The interpretation of heterogeneity statistics with only two studies is meaningless The proportional meta-analysis methodology is not suitable for these disparate study designs Confidence intervals are extremely wide, indicating very low precision\n\nRequired changes:\nRemove the meta-analysis Present a narrative synthesis Report individual study results with appropriate effect measures Discuss limitations of the available evidence\n\n4. Are the conclusions drawn adequately supported by the results presented in the review? No\nExplanation: The conclusions overstate the evidence:\nClaims about PRP's potential are too strong given the limited evidence base Generalizability is severely limited by small sample sizes and study designs Safety conclusions are drawn from very limited data\n\nThe conclusions need to:\nAcknowledge the preliminary nature of the evidence Emphasize the need for larger, well-designed studies Be more cautious about safety claims Clearly state the limitations of the current evidence\n\n5. Is this a Living Systematic Review? Not applicable This is not presented as a Living Systematic Review, and given the current state of evidence in this field, a traditional systematic review format is appropriate. Key Recommendations for Improvement:\nClearly state primary and secondary objectives using PICO framework Provide complete methodological details including full search strategies Remove meta-analysis and present narrative synthesis Revise conclusions to match the limited evidence base Add detailed limitations section\nThe manuscript requires major revision to meet basic standards for systematic review reporting and to ensure its conclusions are appropriately aligned with the available evidence.\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? No\n\nIs the statistical analysis and its interpretation appropriate? No\n\nAre the conclusions drawn adequately supported by the results presented in the review? No\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/13-1508
https://f1000research.com/articles/13-1507/v1	09 Dec 24	{ "type": "Systematic Review", "title": "Complications following miniplate insertion in maxillofacial fractures: a systematic review", "authors": [ "Bramasto Purbo Sejati", "Ahmad Kusumaatmaja", "Maria Goreti Widiastuti", "Tetiana Haniastuti", "Bramasto Purbo Sejati", "Ahmad Kusumaatmaja", "Maria Goreti Widiastuti" ], "abstract": "Background Maxillofacial fractures, frequently arising from road traffic incidents, falls, and acts of interpersonal aggression, are a considerable public health issue, exhibiting diverse epidemiological patterns according to demographic factors. The application of miniplates for fracture stabilization is a recognized technique, with innovative methods such as 3D plate systems emerging. Nonetheless, consequences including infections and hardware malfunctions persist. This systematic review seeks to present current evidence regarding the complications linked to miniplate placement in maxillofacial fractures over the last ten years.\n\nMethods A systematic review was performed in accordance with PRISMA principles. Databases such as the Cochrane Library, PubMed, and Scopus were examined from September 2014 to September 2024. Studies documenting problems related to miniplate placement were included, without language constraints. The ROBINS-I tool was utilized for non-randomized studies, whereas the Cochrane risk of bias tool was applied to randomized controlled trials.\n\nResults From 2,289 initially found studies, 56 satisfied the inclusion criteria. Among these, 28 employed interventional designs, whilst the remaining 28 were observational research. The predominant problems documented in several investigations encompassed infection, wound dehiscence, malocclusion, paraesthesia, malunion/non-union, segment movement, hardware failure, and palpable hardware. Advanced methodologies such as 3D plate systems and locking mechanisms were linked to diminished complication rates.\n\nConclusion This systematic analysis presents a decade of updated research about problems associated with miniplate placement in maxillofacial fractures. Novel methodologies such as 3D plate systems and locking mechanisms demonstrate promise in mitigating problems relative to conventional techniques. These findings can facilitate informed decision-making in clinical practice. Additional study utilizing standardized outcomes and prospective designs is essential to enhance comprehension of the long-term effects of miniplate utilization.", "keywords": [ "systematic review", "miniplate", "complications", "maxillofacial fractures." ], "content": "Introduction\n\nMaxillofacial fractures represent a considerable public health issue, frequently arising from road traffic accidents (RTAs), falls, and interpersonal violence, with RTAs being the primary cause in many countries, including Jordan and India.1,2 The epidemiology of these fractures differs by demographics, exhibiting a greater occurrence in males, especially within the 21-30 age range.2 In Germany, there is a discernible increase in the frequency of maxillofacial trauma procedures, underscoring the necessity for efficient surgical interventions.3 Moreover, in low- and middle-income countries, augmenting education and training for local surgeons is essential for increasing care and outcomes for cranio-maxillofacial injuries.4 A multidisciplinary approach is crucial for the proper management of these complicated injuries.\n\nThe insertion of miniplates is an essential technique in numerous surgical contexts, especially for skeletal establishing in orthodontics and fracture stabilization. The insertion procedure generally requires 25 to 30 minutes, with research indicating an overall success rate of 95.5% for miniplates utilized in orthodontic treatments.5 The stability of these miniplates is greatly affected by the quality and amount of cortical bone at the insertion site, rendering them advantageous in regions with restricted bone availability.6 A new approach utilizing three-dimensional imaging for pre-surgical planning has been established to augment the precision of miniplate placement, thereby enhancing surgical outcomes and minimizing operating duration.6 Miniplate fixation is a dependable technique for establishing solid skeletal anchoring and promoting healing.5,7\n\nThe available literature reveals a research gap concerning problems associated with miniplate implantation in craniofacial and orthognathic operations, indicating a necessity for more comprehensive studies. Numerous systematic reviews have recognized prevalent problems, including infections, plate exposure, and patient inclination for removal; nonetheless, the total complication rates exhibit significant variability, with certain studies documenting rates as high as 32.5% for plate-related complications.8 The average removal time is variable, spanning from 5.5 to 10.7 months, which suggests the absence of defined monitoring techniques.9 Moreover, most research are retrospective, constraining the capacity to establish conclusive insights regarding the long-term results and risk factors associated with miniplate utilization.9 This highlights the need for future research with more substantial, prospective cohorts and standardized outcome measures to further understanding of the implications of miniplate insertion and related problems in clinical practice.10\n\nTherefore, we aim to update the complications associated with miniplate insertion in maxillofacial fractures by a systematic review approach. Additionally, the results of this study could benefit healthcare practitioners and patients in making an informed decision regarding the applicability and potential complications.\n\n\nMethods\n\nThis systematic review was performed based on PRISMA guidelines on systemic reviews and meta-analyzes.11 The Cochrane Library, PubMed, and Scopus were examined from September 1, 2014, to September 1, 2024. We implement a decade-long trend to guarantee the innovative technique. We utilize the keyword combinations as follows: miniplate AND complications AND maxillofacial. Table 1 delineated a combination of various search techniques. Furthermore, relevant papers that satisfied the inclusion criteria were manually identified within each retrieved study.\n\nWe incorporated a comprehensive original study detailing the complications associated with miniplate placement in maxillofacial fractures. No linguistic constraints were imposed. Studies were considered irrespective of the languages utilized, provided that English translations were accessible. Two reviewers (B.P.S. and A.K.) conducted separate evaluations of the titles and abstracts of possibly qualifying articles. All differences were deliberated with the third investigator (T.H).\n\nTwo independent assessors (B.P.S. and A.K.) extracted data and resolved discrepancies. We incorporated the subsequent data: 1) Attributes of the included studies (e.g., first author’s name and publication year); 2) demographic attributes of the patient population (e.g., age, male percentage, and participant count in each group); 3) intervention attributes (e.g., type); and 4) outcomes. Disputes were settled by dialogue with the corresponding author (T.H.) until a consensus was achieved.\n\nThe reported outcome was complications following miniplate insertion in maxillofacial fractures. We endeavored to reach the original authors to acquire further or missing information through email.\n\nThe quality of the included studies was evaluated according to the criteria outlined in the Cochrane Handbook for Systematic Reviews of Interventions.12 The Cochrane risk of bias assessment was utilized for the randomized control trials.13 We evaluated the risk of bias in non-randomized studies utilizing the checklist for prevalence studies.14 All studies were evaluated by two independent reviewers (B.P.S. and A.K.). All disputes have been resolved during the consensus meeting.\n\nThis systematic review emphasizes the synthesis and analysis of data to summarize and interpret findings from individual studies. The review initially delineates attributes, including design, sample size, interventions, and outcomes. The synthesis entails a critical evaluation of the research’ quality, addressing methodological rigor and identifying any biases or limitations.\n\n\nResults\n\nFigure 1 illustrates the electronic search procedure. We initially detected 2,289 articles. Of these, 2,196 duplicate papers or irrelevant studies were eliminated. Ninety-one papers were identified for additional investigation. Thirty-seven research were omitted from this list due to insufficient relevant data (Appendix Table 1). Two additional papers were acquired: one from a prior literature review and the other from a website search. We incorporated a total of 56 studies that fulfilled the inclusion criteria.15–70\n\nThe attributes of the studies included have been captured in Table 2. Out of 56 investigations, 28 utilized an interventional design, including randomized controlled trials (RCTs), whereas the other 28 were observational studies, comprising cohort designs. India was the most prevalent country of study origin. This distribution represents a balance between experimental and observational research methodologies within the dataset, offering a comprehensive foundation for analyzing the investigated phenomena.\n\nThe majority of research contrasted conventional miniplate types with innovative methods, including 3D plate systems. Additionally, several studies investigated the effectiveness of locking compared to non-locking strategies in similar interventions. The most commonly reported complications included infection, wound dehiscence, malocclusion, paraesthesia, malunion/non-union, segment movement, hardware failure, and palpable hardware. These results underscore the variety of potential complications linked to the use of miniplates in clinical practice. Other details of results is provided in Table 2 extended data.\n\nResult of risk of bias assessment were described in Table 3 for RCTs studies. Twelve and ten studies addressed concerns about sufficient randomization and incomplete outcome data. five studies used adequate concealment of allocation. Participants and personnel in four studies were blind to treatment assignment, while assessors were unaware in four studies.\n\nThe quality assessment of risk of bias for non-randomized studies can be seen in Table 4. overall studies yielded low risk of bias.\n\n\nDiscussion\n\nThis systematic review presents updated ten-year evidence about complications associated with miniplate insertion in maxillofacial fractures. The results of our study may assist healthcare practitioners and patients in making informed decisions about the applicability and potential difficulties associated with miniplate deployment. The present study utilized a systematic and rigorous methodology, which generated findings of high confidence quality.\n\nInfection is a notable problem subsequent to the placement of miniplates in maxillofacial surgery. Consistent with prior research, the overall infection rate linked to miniplates is roughly 13.3%, which is the primary reason for their removal, representing about 2.9% of cases in one meta-analysis.9,71 A study on orthodontic anchorage miniplates revealed that 17.3% of the placed miniplates developed infections, influenced significantly by the proximity to the mucogingival junction and the frequency of dental hygiene.72 In addition, one study reported the microbiological research indicated that Staphylococcus aureus was the primary pathogen at infected locations, underscoring the necessity for regular microbial evaluations to inform therapy.73 These findings highlight the necessity of monitoring and mitigating infection risks associated with miniplate utilization.\n\nThe utilization of 3D miniplates in managing mandibular fractures has been associated to a reduced incidence of complications relative to traditional systems. Studies demonstrate that 3D miniplates enhance stability and diminish operational duration, resulting in superior intraoperative results and markedly fewer problems, including enhanced biting force and fracture stability.74 A retrospective investigation of 336 patients indicated that merely 8.03% encountered minor problems, whereas significant difficulties arose in only 1.49% of instances.75 A study on patient-specific 3D-printed miniplates exhibited exceptional precision in fixing and effective osseous union, devoid of material fractures or plate exposure.76 The adaptability and effectiveness of 3D miniplates enhance their successful application in the treatment of mandibular fractures, underscoring their advantages compared to conventional techniques.77\n\nSeveral limitations must be acknowledged. A meta-analysis was not performed owing to the heterogeneity of the included studies. Secondly, the patient demographics and treatment durations differed among research, highlighting the necessity for meticulous evaluation when selecting protocols to pay attention to. The evaluation encompasses a decade-long range to concentrate on the most recent and novel methodologies in this field of research.\n\n\nConclusion and implications\n\nThis study’s findings indicate that the majority of research has concentrated on contrasting conventional miniplate types with more advanced ways, such as 3D plate systems, while also assessing the efficacy of locking versus non-locking procedures. The extensive range of documented complications—such as infection, wound dehiscence, malocclusion, paraesthesia, malunion or non-union, segment movement, hardware failure, and palpable hardware—illustrates the clinical difficulties associated with miniplate implantation. The results underscore the necessity for meticulous evaluation of the miniplate technology and technique employed to reduce problems and enhance patient outcomes in maxillofacial surgery.\n\n\nAuthor contributions\n\nConceived and designed the experiments: BPS TH. Analyzed the data: BPS AK. Wrote the paper: BPS TH. Designed search strategies: BPS AK TH. Critically reviewed the manuscript for important intellectual content: BPS AK MGW TH. Read and approved the final version: BPS AK MGW TH. Guarantors: BPS TH.\n\n\nEthics and consent\n\nEthical approval and consent were not required.", "appendix": "Data availability\n\nUnderlying data- No data are associated with this article.\n\nExtended Data\n\nZenodo: Supplementary data, 10.5281/zenodo.14064447.78\n\nThis project contains the following underlying data:\n\n1. Reporting guidelines, PRISMA checklist\n\n2. Appendix Table 1. List of the excluded studies\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (Creative Commons Attribution 4.0 International).\n\nZenodo: Characteristics of included studies, 10.5281/zenodo.14207706.79\n\nThis project contains the following underlying data:\n\n• Table 2 Characteristics of included studies\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (Creative Commons Attribution 4.0 International).\n\n\nAcknowledgement\n\nThe authors declare no potential conflicts of interest regarding the authorship and publication of this article.\n\n\nReference\n\nShukla AD: Epidemiology of maxillofacial injuries during monsoon and non-monsoon season in India: a data-based retrospective study from a tertiary care dental teaching hospital. F1000Res. 2023; 12. Publisher Full Text\n\nBataineh AB: The incidence and patterns of maxillofacial fractures and associated head and neck injuries. J. 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PubMed Abstract \| Publisher Full Text\n\nFani M: Is mini-plate removal necessary for oral and maxillofacial surgery patients? A five-year case-control study. Frontiers in Dentistry. 2020; 17. Publisher Full Text\n\nFerrari R: Complication rate in mandibular angle fractures—one vs. two plates: a 12-year retrospective analysis. Oral Maxillofac. Surg. 2018; 22(4): 435–441. PubMed Abstract \| Publisher Full Text\n\nGamit M: Comparison of bite force evaluation for mandibular angle fracture fixation by conventional miniplates versus new design miniplates: a clinical study. Oral Maxillofac. Surg. 2024; 28(2): 645–652. PubMed Abstract \| Publisher Full Text\n\nGarcía Carricondo AR: A comparative study between traditional fixation with miniplates and modified lag screws for the treatment of mandibular fractures. Clin. Oral Investig. 2018; 22(3): 1503–1511. PubMed Abstract \| Publisher Full Text\n\nGhezta NK, Bhardwaj Y, Rani P, et al.: Efficacy of Retromandibular Transparotid Approach for the Management of Extracapsular Subcondylar Mandibular Fractures Using 2-mm Titanium Miniplates: A Prospective Clinical Study. J. Oral Maxillofac. Surg. 2016; 74(8): 1613–1621. PubMed Abstract \| Publisher Full Text\n\nGraillon N: Do mandibular miniplates increase the risk of complex fracture in facial trauma recurrence? Case series. J. Craniofac. Surg. 2021; 49(7): 613–619.\n\nKanubaddy SR: Management of Mandibular Angle Fractures: Single Stainless Steel Linear Miniplate Versus Rectangular Grid Plate—A Prospective Randomised Study. Journal of Maxillofacial and Oral Surgery. 2016; 15(4): 535–541. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKaushik S, Ali I, Dubey M, et al.: 2 mm conventional miniplates with three-dimensional strut plate in mandibular fractures. Annals of Maxillofacial Surgery. 2020; 10(1): 10–15. PubMed Abstract \| Publisher Full Text\n\nKerdoud O, Aloua R, Kaouani A, et al.: Management of mandibular angle fractures through single and two mini-plate fixation systems: Retrospective study of 112 cases. Int. J. Surg. Case Rep. 2021; 80: 105690. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKhan M: Comparative Evaluation of 3d Locking Versus Non-Locking Titanium Miniplates in the Treatment of Mandibular Fracture. Indian J. Otolaryngol. Head Neck Surg. 2020; 72(3): 363–369. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKhandelwal P: Miniplate removal in operated cases of maxillofacial region in a dental institute in Rajasthan, India. Medicine and Pharmacy Reports. 2019; 92(4): 393–400. PubMed Abstract \| Publisher Full Text\n\nKreutzer K: Patient-specific 3D-printed mini-versus reconstruction plates for free flap fixation at the mandible: Retrospective study of clinical outcomes and complication rates. J. Craniofac. Surg. 2023; 51(10): 621–628. Publisher Full Text\n\nKumar S: Treatment Outcome Comparison Between two 3-Dimensional Plates (Y-Shaped Plate Versus Trapezoidal Condylar Plate) in Management of Mandible Condylar Fracture: A Randomized Control Trial. Journal of Maxillofacial and Oral Surgery. 2023; 22(1): 25–32. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMannan R, Farooq A, Mukhtar H: Comparison of single vs double miniplates in the management of mandibular angle fractures. Pakistan Journal of Medical and Health Sciences. 2018; 12(2): 662–664.\n\nMathew N: The Efficacy of Fixation of Unilateral Mandibular Angle Fracture with Single 3D Plate vs Single Miniplate Using Transbuccal Approach. Journal of Maxillofacial and Oral Surgery. 2022; 21(2): 405–412. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMishra N: 3-D Miniplates Versus Conventional Miniplates in Treatment of Mandible Fractures. Journal of Maxillofacial and Oral Surgery. 2019; 18(1): 65–72. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMondal S: Comparative Evaluation between Single Noncompression Miniplate and Two Noncompression Miniplates in the Treatment of Mandibular Angle Fractures. Craniomaxillofac. Trauma Reconstr. 2019; 12(2): 122–127. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPfister J, Shazwani FN, Müller M, et al.: Clinical results of two different three-dimensional plate types for the treatment of mandibular angle fractures: a retrospective analysis. Oral Maxillofac. Surg. 2024; 28: 1501–1507. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRahpeyma A, Khajehahmadi S, Barkhori Mehni S: Treatment of mandibular fractures by two perpendicular mini-plates. Iran J Otorhinolaryngol. 2014; 26(74): 31–36. PubMed Abstract\n\nRai A, Jain A, Datarkar A: Comparison of single versus two non-compression miniplates in the management of unfavourable angle fracture of the mandible: a prospective randomized clinical study. Oral Maxillofac. Surg. 2018; 22(2): 157–161. PubMed Abstract \| Publisher Full Text\n\nRai A, Jain A, Thukral R: Two miniplates versus 3-dimensional plate in the management of mandibular subcondylar fractures: a retrospective analysis. Oral Maxillofac. Surg. 2021; 25(4): 457–461. PubMed Abstract \| Publisher Full Text\n\nRibeiro-Junior PD: Mandibular angle fractures treated with a single miniplate without postoperative maxillomandibular fixation: A retrospective evaluation of 50 patients. Cranio - Journal of Craniomandibular Practice. 2018; 36(4): 234–242. PubMed Abstract \| Publisher Full Text\n\nSaha R: A comparison between locking plates and miniplates in fixation of mandibular fractures. Biomedical and Pharmacology Journal. 2015; 8: 799–804. Publisher Full Text\n\nSakong Y, Kim YH, Chung KJ: Analysis of Complication in Mandibular Angle Fracture: Champy Technique Versus Rigid Fixation. J. Craniofac. Surg. 2021; 32(8): 2732–2735. PubMed Abstract \| Publisher Full Text\n\nSarepally G: A Comparative Evaluation of 2.0mm Two-Dimensional Miniplates Versus 2.0mm Three-Dimensional Miniplates in Mandibular Fractures. Cureus. 2022; 14(1): e21325. PubMed Abstract \| Publisher Full Text\n\nSehgal S, Ramanujam L, Prasad K, et al.: Three-dimensional v/s standard titanium miniplate fixation in the management of mandibular fractures - A randomized clinical study. J. Craniofac. Surg. 2014; 42(7): 1292–1299.\n\nShaik M, Subba Raju T, Rao NK, et al.: Effectiveness of 2.0 mm Standard and 2.0 mm Locking Miniplates in Management of Mandibular Fractures: A Clinical Comparative Study. Journal of Maxillofacial and Oral Surgery. 2014; 13(1): 47–52. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSikora M, Chęciński M, Sielski M, et al.: The use of 3d titanium miniplates in surgical treatment of patients with condylar fractures. J. Clin. Med. 2020; 9(9): 1–13.\n\nSingh A, Arunkumar KV: Standard 3D Titanium Miniplate Versus Locking 3D Miniplate in Fracture of Mandible: A Prospective Comparative Study. Journal of Maxillofacial and Oral Surgery. 2016; 15(2): 164–172. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSingh R: Comparative evaluation of 2D miniplates and 3D miniplates fixation in mandibular angle fracture - A clinical study. Indian J. Dent. Res. 2020; 31(1): 134–137. PubMed Abstract \| Publisher Full Text\n\nSingh R: A comparative assessment of the management of mandibular angle fractures using 3D plates and 2D mini plates. J. Contemp. Dent. Pract. 2020; 21(4): 400–403. PubMed Abstract \| Publisher Full Text\n\nSpinelli G: Management of Mandibular Angle Fractures by Two Conventional 2.0-mm Miniplates: A Retrospective Study of 389 Patients. Craniomaxillofac. Trauma Reconstr. 2016; 9(3): 206–210. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSukegawa S: Which fixation methods are better between three-dimensional anatomical plate and two miniplates for the mandibular subcondylar fracture open treatment? J. Craniofac. Surg. 2019; 47(5): 771–777. Publisher Full Text\n\nSukegawa S: A retrospective comparative study of mandibular fracture treatment with internal fixation using reconstruction plate versus miniplates. J. Craniofac. Surg. 2019; 47(8): 1175–1180.\n\nSukegawa S: Maxillofacial Trauma Surgery Patients With Titanium Osteosynthesis Miniplates: Remove or Not? J. Craniofac. Surg. 2020; 31(5): 1338–1342. Publisher Full Text\n\nSweta S: Efficacy of Locking Miniplates in Managing Mandibular Fractures without Intermaxillary Fixation. J. Pharm. Bioallied Sci. 2022; 14(Suppl 1): S131–s134. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTiwari M, Meshram V, Lambade P, et al.: Titanium Lag Screw Versus Miniplate Fixation in the Treatment of Anterior Mandibular Fractures. J. Oral Maxillofac. Surg. 2019; 77(5): 1031–1039. PubMed Abstract \| Publisher Full Text\n\nVishal: Significance of Microbial Analysis during Removal of Miniplates at Infected Sites in the Craniomaxillofacial Region - An Evaluative Study. Ann Maxillofac Surg. 2020; 10(2): 330–334. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYadav S: Conventional 2D miniplate versus 3D four-holed and eight-holed miniplates in the management of mandibular angle fractures. Journal of Oral and Maxillofacial Surgery, Medicine, and Pathology. 2020; 32(5): 336–341. Publisher Full Text\n\nYang L, Patil PM: Comparative evaluation of 2.0 mm locking plate system vs 2.0 mm non-locking plate system for mandibular angle fracture fixation: A prospective randomized study. Eur. Rev. Med. Pharmacol. Sci. 2015; 19(4): 552–556. PubMed Abstract\n\nPalani T: Evaluation of 3D trapezoidal plates in open reduction and internal fixation of subcondylar fractures of mandible: a clinical trial. Cureus. 2021; 13(6). Publisher Full Text\n\nVashistha A: Comparison of 2 mm single locking miniplates versus 2 mm two non-locking miniplates in symphysis and parasymphysis fracture of mandible. journal of oral biology and craniofacial research. 2017; 7(1): 42–48. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nJaber M, Abouseif N, Hassan M, et al.: Risk Factors Contributing to Symptomatic Miniplate Removal following Orthognathic Surgery: Systematic Review and Meta-Analysis. J. Clin. Med. 2024; 13(11): 3335. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nFaber J, et al.: Infection predictive factors with orthodontic anchorage miniplates. Seminars in Orthodontics. Elsevier; 2018.\n\nPrajapati V, Shahi AK, Prakash O: Significance of microbial analysis during removal of miniplates at infected sites in the craniomaxillofacial region-an evaluative study. Annals of Maxillofacial Surgery. 2020; 10(2): 330–334. Publisher Full Text\n\nBhatt V, Motiwale T, Mitra GV: Evaluation of the efficacy of Three-dimensional mini plates versus conventional mini plates used in the management of anterior mandibular fractures. IP Indian Journal of Orthodontics and Dentofacial Research. 2023; 8(3): 198–208. Publisher Full Text\n\nBohner L: Treatment of mandible fractures using a miniplate system: A retrospective analysis. J. Clin. Med. 2020; 9(09): 2922. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKreutzer K: Patient-specific 3D-printed miniplates for free flap fixation at the mandible: A feasibility study. Front. Surg. 2022; 9: 778371. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSadhwani BS, Khokhar M: Versatility of various 3D non locking miniplates in treatment of mandibular fractures. International Journal of Orthopaedics. 2022; 8(3): 85–87.\n\nAdhyastha: Supplementary Data. Zenodo. 11 Nov. 2024.\n\nadhyastha: characteristics of included study. [Data set]. Zenodo. 2024. Publisher Full Text" }	[ { "id": "355750", "date": "16 Jan 2025", "name": "Mohamed Jaber", "expertise": [ "Reviewer Expertise Maxillofacial injuries", "Oral cancer and precancer", "dental education" ], "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 Report for Manuscript Submitted to F1000Research Title: Complications following miniplate insertion in maxillofacial fractures: A systematic review. The manuscript is relevant to the field of maxillofacial surgery and explores an important area: complications associated with miniplate insertion. It aligns with the interests of F1000Research, particularly in presenting evidence-based insights useful for clinical practice. However, significant improvements are required for it to meet the journal's standards of rigor and clarity. Key findings\nThe manuscript reviews ten years of data on complications following miniplate insertion, providing valuable insights into trends and advanced methodologies. The focus on novel techniques such as 3D plate systems highlights advancements with potential to reduce complications. The authors adhere to PRISMA guidelines and employ robust tools like the ROBINS-I and Cochrane risk of bias tools to evaluate study quality. The discussion emphasizes the need for standardized outcomes and prospective studies, which is vital for advancing clinical practices.\nLimitations and Suggestions for Improvement:\nThe results and discussion sections could benefit from deeper statistical and comparative analyses. For example: provide quantitative comparisons between traditional and advanced methodologies. Detail the statistical significance of the findings related to complication rates. The search strategy appears robust, but details on the inclusion/exclusion criteria for studies are insufficient. A clearer explanation of why certain studies were excluded (e.g., irrelevance or quality issues) would enhance transparency. Some sections, particularly the introduction and discussion, are repetitive. Consolidating overlapping points and presenting a more concise narrative will improve readability. The inclusion of figures or summary tables (e.g., comparison of complication rates by technique) would better communicate the findings. While the PRISMA framework was followed, the authors did not conduct a meta-analysis. If heterogeneity precluded this, it should be explicitly stated and justified in the text. The language is generally clear but needs refinement to eliminate redundancy and improve flow. For example, the repeated mention of infection rates and their significance across sections could be consolidated. Certain sentences are overly technical, which might hinder accessibility for a broader audience. Simplifying terminology without losing accuracy is recommended. The manuscript lacks a detailed comparison of its findings with those of previous reviews or meta-analyses. Highlighting how this review advances knowledge or contradicts earlier work would enhance its contribution. A more robust discussion of study limitations is necessary, particularly concerning variability in patient demographics and differences in surgical expertise, healthcare infrastructure across studies, and retrospective study designs in many included papers.\nKey Recommendations:\nInclude statistical analyses or at least a detailed qualitative comparison of complication rates. Streamline the text, especially in sections with overlapping content. Add summary tables or figures to improve data visualization. Clarify methodological choices, particularly regarding the heterogeneity of included studies. Expand on the implications of the findings, both clinical and research-oriented.\nSuitability for indexing: The manuscript has potential but requires revisions to strengthen its methodological rigor, depth of analysis, and presentation. With these changes, it could serve as a valuable resource for clinicians and researchers in maxillofacial surgery. I recommend major revisions prior to indexing.\nThank you\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? Partly\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": [] }, { "id": "348230", "date": "22 Jan 2025", "name": "Mohammad Adhitya Latief", "expertise": [ "Reviewer Expertise trauma and fractures management in oral maxillofacial" ], "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\nPower:\nComprehensive Review: The paper systematically reviews complications associated with miniplate insertion in maxillofacial fractures over a decade, providing a broad and updated perspective, as mention on methods that data examined from September 1, 2014, to September 1, 2024. Diverse Data Sources: It utilizes multiple databases (Cochrane Library, PubMed, Scopus) and includes studies without language constraints, ensuring a wide range of data as mentioned in methods section, data source and searches. Methodological Rigor: The review follows PRISMA guidelines and employs robust tools (ROBINS-I, Cochrane risk of bias tool) for assessing study quality, enhancing the reliability of findings. Balanced Study Designs: The inclusion of both interventional and observational studies offers a comprehensive analysis of the phenomena. This distribution represents a balance between experimental and observational research methodologies within the dataset, offering a comprehensive foundation for analyzing the investigated phenomena. Focus on Novel Techniques: The paper highlights advanced methodologies like 3D plate systems and locking mechanisms, which show promise in reducing complications. \nWeakness:\nHeterogeneity: The included studies exhibit significant variability in patient demographics, treatment durations, and methodologies, which limits the ability to perform a meta-analysis. One of the example is decision to include the use of miniplate in orthognatic surgery, Because the title is focussing in complication of miniplate in maxillofacial fractures, but the samples used in this research also include miniplate in orthognatic surgery which is very different in outcomes or surgery result. Lack of Standardization: The absence of standardized outcome measures across studies makes it challenging to compare results directly. Although I cant find table 2 as result in the article, I could tell that it is challenging since there will be differences outcome because the different usage in fracture cases and orthognatic cases Limited Prospective Data: There is a need for more prospective studies to better understand the long-term effects of miniplate utilization. As mention in article that Additional study utilizing standardized outcomes and prospective designs is essential to enhance comprehension of the long-term effects of miniplate utilization Potential Bias: Despite using robust tools, the risk of bias in some studies remains unclear, which could affect the overall conclusions. We might correct this by using statistic to eliminate any factor that has potential bias, but unfortunately I cant find any of them in this article.\nOverall, the paper provides valuable insights but also highlights the need for more standardized and prospective research to fully understand the implications of miniplate insertion in maxillofacial fractures. However, In my opinion the article provides a comprehensive exploration of the role and applications of miniplates in craniofacial surgery and Oral and maxillofacial field procedures. It delves into their potential complications, offering insights into their effectiveness in stabilizing fractures, facilitating orthodontic surgery, and supporting reconstructive surgeries. This article broadens our understanding of miniplate application, guiding clinicians in selecting appropriate systems and optimizing outcomes. By addressing both benefits and limitations, it serves as a valuable resource for advancing knowledge in the field and improving patient care.\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? No\n\nIs the statistical analysis and its interpretation appropriate? No\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/13-1507
https://f1000research.com/articles/13-1505/v1	09 Dec 24	{ "type": "Research Article", "title": "Trends in Smart Restaurant Research: Bibliometric Review and Research Agenda", "authors": [ "Alejandro Valencia-Arias", "Sebastián Cardona-Acevedo", "Ezequiel Martínez Rojas", "Juana Ramírez Dávila", "Paula Rodriguez-Correa", "Lucia Palacios-Moya", "Renata Teodori de la Puente", "Erica Agudelo-Ceballos", "Martha Benjumea-Arias", "Sebastián Cardona-Acevedo", "Ezequiel Martínez Rojas", "Juana Ramírez Dávila", "Paula Rodriguez-Correa", "Lucia Palacios-Moya", "Renata Teodori de la Puente", "Erica Agudelo-Ceballos", "Martha Benjumea-Arias" ], "abstract": "Background The automation of processes and services has transformed various industries, including the restaurant sector. Technologies such as the Internet of Things (IoT), machine learning, Radio Frequency Identification (RFID), and big data have been increasingly adopted to enhance service delivery, improve user experiences, and enable data traceability. By collecting user feedback and analyzing sentiments, these technologies facilitate decision-making and offer predictive insights into future food preferences. This study aims to explore current research trends in intelligent restaurants, focusing on technological applications that improve service and decision-making.\n\nMethods A bibliometric analysis was conducted in accordance with the PRISMA-2020 guidelines. A total of 94 academic documents were reviewed from the Scopus and Web of Science databases, focusing on publications related to intelligent restaurant systems, particularly involving IoT and automation technologies.\n\nResults The analysis revealed that the United States, India, and China have contributed the most to the field, with a particular emphasis on China’s implementation of IoT architecture and robotics in restaurant settings. Chinese restaurant innovations, particularly in robotics, are among the most frequently cited in the literature. The study identifies these countries as leading the research in the intelligent restaurant domain.\n\nConclusions Technologies such as IoT, machine learning, RFID, and big data are driving advancements in restaurant automation, enhancing service efficiency and user experience. The United States, India, and China are leading research in this area, with China standing out for its application of robotics and IoT in restaurants. This research provides a foundation for future studies aimed at improving predictive models for food selection and service optimization.", "keywords": [ "Smart restaurants", "PRISMA", "machine learning", "deep learning", "IoT." ], "content": "Introduction\n\nThe intensive use of various advanced technological tools in different economic sectors is becoming increasingly widespread. The development of information and communication technologies (ICT) has brought about significant changes in the way people interact and adapt to different social environments (Bai & Wu, 2022). Thus, the maturity of technology has influenced the productivity and daily life of people who use these tools to have a better quality of life, greater efficiency in their daily activities, and to enjoy the convenience they have brought (Bai & Wu, 2022).\n\nTools such as Artificial Intelligence (AI), the Internet of Things (IoT), building information, big data, cloud computing, machine learning (Wu & Huang, 2020), and automation tools have allowed companies dedicated to different activities to achieve better results in terms of productivity, time, competitiveness, and economics (Jubaid et al., 2023). The adoption of these technologies makes it possible to create new types of business architectures and build sustainable and intelligent industries (Elkhwesky & Elkhwesky, 2023). One sector in which the adoption of technological tools has expanded is the tourism and services sector, especially in restaurants (Singh et al., 2023).\n\nIn this sense, as users become more comfortable with technologies that allow, for example, self-service, automatic ordering and payment systems, and robotic services, this type of technology will continue to become more relevant in restaurants (Wong et al., 2022). Some of these technologies became more relevant, especially in the context of Covid-19, where one of the most affected industries was restaurants, which, in order to adapt to the new reality, were forced to implement this type of technology to provide their services again (Hasan et al., 2022).\n\nHowever, some restaurants have continued to implement technologies to improve the user experience and update the way they provide the service. Some of the technologies that have been adapted include a personal digital assistant (PDA), Wi-Fi access, and high-end monitors for menu selection, including the use of holograms (Kavitha et al., 2022). To adapt to the context of smart cities, restaurants are transforming into smart restaurants. For example, the use of augmented reality in menus can enhance the user’s dining experience and facilitate operations related to ordering and billing by automating this entire process (Prathibanandhi et al., 2022).\n\nThe integration of elements such as cloud computing allows different services to be hosted in the cloud and has information available to customers and employees; the use of sensors and IoT devices allows restaurants to have better management of the restaurant, providing in-formation on the occupancy of the premises, interactive menus, availability of food, process and verification of food quality, and organization (Martino et al., 2022). One of the advances that has had the greatest impact on restaurants is the use of robots for user service, either as waiters or just to take the order (Mishra et al., 2022). However, the use of this type of technology requires the integration of different tools, such as Internet access services, control, monitoring, and identification services, to integrate people, things, and objects (Wu & Huang, 2020).\n\nIn addition, it is possible to create interactive menus that can identify user preferences based on data collection, and through the use of chatbots, it is possible to collect purchase data and provide quick responses to consumers by making personalized recommendations (Kaur et al., 2023).\n\nHowever, in restaurants, robots need to understand the environment and make decisions accordingly, such as changing their route if they encounter obstacles along the way. To have the ability to make decisions and behave like humans, robots need information from multiple sources from which they can understand the change in the environment, the position of other robots, and the path to the destination. This can be achieved by integrating robots with IoT technology, which improves the capabilities of a robotic waiter (Mishra et al., 2022).\n\nThe above not only allows for a personalized and more interactive experience for the user but also increases the possibility of meeting their expectations. This is because today’s users are much more demanding, have more options to choose from, and have more bargaining power in addition to being content generators who share their experiences with other users (Almeida et al., 2023). On the other hand, it is also possible to support marketing strategies through the use of digital tools, allowing the restaurant’s competitive advantage to be improved using data from reviews on social networks, patterns of feelings, and behavior through the use of big data (Hajek & Sahut, 2022).\n\nTechnology adoption in the restaurant sector remains low due to a variety of interrelated factors. first, resistance to change by restaurant owners and managers can be a significant barrier, many of which may be rooted in traditional practices. and may be reluctant to invest in technology due to concerns about cost, implementation complexity, and fear of operational disruption. and a lack of knowledge and understanding of how to effectively integrate technology into restaurant operations can lead to uncertainty and avoidance. adoption. It is suggested that the internal environmental locus of control, that is, the perception of control over the internal environment of the restaurant, can influence behavioral intentions toward technology adoption, highlighting the importance of addressing the attitudes and perceptions of customers. owners and managers (Joo, Kim, and Hwang, 2023).\n\nIn addition, lack of access to adequate resources and training can be a limiting factor in the adoption of technology in the restaurant sector; owners and managers may lack the technical knowledge necessary to evaluate, select, and implement technological solutions more ap-propriate for their operations. in addition, the lack of adequate technological infrastructure, such as reliable Internet connectivity, can hinder the effective implementation of advanced technologies. The importance of factors such as consumer attitudes and loyalty towards smart vending technology in restaurants is highlighted, indicating that customer perceptions and experiences can also influence technology adoption by restaurant operators (Lacap et al., 2023); these factors may contribute to low technology adoption in the restaurant sector by creating perceived and real barriers to the successful implementation of technological solutions.\n\nAlthough the adoption of these technologies in various sectors has been widely addressed, the hospitality and tourism sector, which includes restaurants, still requires more research and the adoption of these technologies in this sector remains low. (Elkhwesky and Elkhwesky, 2023; Klouvidaki et al., 2023). Therefore, research is needed to identify the factors and advances in the incorporation of technological elements that facilitate the development of smart restau-rants. In this sense, the objective of the research is to analyze research trends in the field of smart restaurants in order to identify areas that require greater attention and progress, this will be done through a comprehensive bibliometric review of the existing literature on the topic. The ultimate goal is to provide a global perspective of the prevailing approaches in the development of smart restaurants and to guide a research agenda for future studies in this area. By better understanding customer needs and preferences, as well as the benefits and challenges associated with the implementation of emerging technologies, this research seeks to provide clear guidance for decision-making and strategic planning in the sector.\n\nIn addition, this research also seeks to contribute at a theoretical level to the field of innovation and technology management in the hospitality industry by identifying relevant models and conceptual frameworks to understand and address the specific challenges faced by restaurants in the digital era. The results of this research are expected to not only benefit restaurant owners by providing them with valuable insights to improve their operations and services, but also contribute to the advancement of the food and beverage industry as a whole by fostering innovation and adoption of technologies that enhance competitiveness and long-term sustainability. Research can provide practical and strategic insights that help restaurants adapt to an ever-changing environment, improve their competitiveness, and meet rising customer expectations in the digital age. For this purpose, the following research questions were asked.\n\nRQ1: What are the years of greatest interest in research on smart restaurants?\n\nRQ2: What kind of growth exists in the number of scientific articles about research on smart restaurants?\n\nRQ3: What are the main research references on smart restaurants?\n\nRQ4: What is the thematic evolution of scientific production in smart restaurants?\n\nRQ5: What are the main thematic clusters on smart restaurants?\n\nRQ6: What are the growing and emerging keywords in Smart Restaurants research?\n\nRQ7: Which topics are positioned as protagonists in the design of a research agenda on smart restaurants?\n\nIn this sense, the article is composed of a methodological section, which details the way the article is developed, as well as the sources of information, the results or findings obtained, their discussion, as well as the identification of research gaps, the approach to the research agenda, and, finally, the main conclusions.\n\n\nMethods\n\nAs part of this exploratory study on smart restaurants, a bibliometric analysis was conducted using the guidelines described in the PRISMA-2020 statement. (Page et al., 2021; Valencia-Arias et al., 2024; García-Pineda et al., 2024). The collection and review of relevant secondary research materials forms the basis of this methodology.\n\nThe establishment of inclusion criteria that guarantee the selection of relevant texts for analysis is crucial in the context of this bibliometric study, which has Smart Restaurants as its main focus. In this sense, it was determined that one of the exclusion criteria is related to texts that do not directly contribute to the field of study, thus ensuring the quality and relevance of the documents considered in the bibliometric analysis.\n\nThe document selection and exclusion processes consisted of three distinct phases. In the first phase, all records with indexing errors were discarded to ensure the integrity of the biblio-metric database. Only systematic literature reviews are subject to the second phase of exclusion, which involves eliminating all documents for which the full text is not available, as these reviews require an exhaustive examination of the content. It should be noted that bibliometrics is currently based only on the evaluation of available metadata. To ensure that only relevant documents were analyzed in this bibliometric research, texts that did not meet the previously established relevance criteria were eliminated in the third phase of exclusion.\n\nFor this bibliometric study on smart restaurants, we used the Scopus and Web of Science databases, which are currently considered the main sources of academic and scientific in-formation. This decision was based on the comprehensive coverage of both databases, which covered a wide range of disciplines and provided access to numerous academic journals and conferences. This choice is further supported by a study comparing Scopus and Web of Science in a typical university environment, highlighting the value and effectiveness of both in searching and retrieving scientific literature. The use of these well-known databases guaran-tees that the information is up-to-date and relevant, thereby improving the precision and reliability of the data used in this bibliometric analysis (Vieira & Gomes, 2009).\n\nTo perform an efficient search, two specialized search equations were developed for the two selected databases, Scopus and Web of Science. These equations were carefully designed to meet the previously established inclusion criteria while also being tailored to the unique search requirements of each database. This strategy allowed them to retrieve relevant scientific literature and ensure the accuracy and integrity of the data collection for ongoing bibliometric research.\n\nFor the Scopus database: (TITLE (“Smart Restaurant” OR “smart restaurant\" OR (iot AND restaurant) OR (“internet of things” AND restaurant) OR (“artificial intelligence” AND restaurant) OR (“virtual reality” AND restaurant) OR (“augmented reality” AND restaurant)) OR KEY (“Smart restaurant” OR “smart restaurant”) OR (iot AND restaurant) OR (“Internet of Things” AND restaurant) OR (“Artificial Intelligence” AND restaurant) OR (“Virtual Reality” AND restaurant) OR (“Augmented Reality” AND restaurant))).\n\nFor the Web of Science database: (TI=(“intelligent restaurant” OR “smart restaurant\" OR (iot AND restaurant) OR (“Internet of things” AND restaurant) OR (“Artificial intelligence” AND restaurant) OR (“Virtual reality” AND restaurant) OR (“Augmented reality” AND restaurant)) OR AK=(“Smart restaurant” OR “intelligent restaurant\" OR (“IoT AND restaurant”) OR (“Internet of Things AND restaurant”) OR (“Artificial Intelligence AND restaurant”) OR (“Virtual Reality AND restaurant”) OR (“Augmented Reality AND restaurant”))).\n\nFor this bibliometric study on intelligent restaurants, the information obtained from each of the selected databases was extracted, stored, and processed using Microsoft Excel. This platform simplifies the data organization. The free VOSviewer® program was used in a similar manner (Eck & Waltman, 2010). It allows the analysis and visualization of biblio-metric data and the creation of graphs representing different bibliometric indicators. In this study, the combination of Microsoft Excel® and VOSviewer® was found to be a crucial method for managing data and presenting results.\n\nBased on the PRISMA 2020 guidelines (Page et al., 2021). To determine the risk of omitting relevant studies or incorrectly classifying data, it is essential to answer the question of whether an internal automated classifier was used during the study selection and internal or external validation processes. These recommendations were followed in the present study, which used Microsoft Excel® automation tools as the internal mechanism for the selection process. All study investigators contributed to the development and improvement of this tool, which they then independently used to apply the inclusion and exclusion criteria. Through the convergence of the results of the application of the tool, this collaborative approach was adopted with the aim of minimizing the risk of omitting relevant studies or incorrect classifications, ensuring integrity and accuracy in the selection of studies for current restaurant bibliometrics. intelligent.\n\nFollowing the established guidelines, they described the procedures for obtaining or verifying data directly with the original investigators of the studies, any procedures for doing so, and, if applicable, details about the automation tools used in data collection (Page et al., 2021). This includes the number of reviewers involved in the process, their degree of independence, and the procedures for doing so. These guidelines were strictly followed in the context of smart restaurant studies. To ensure efficiency and consistency in the extraction of information, Microsoft Excel® was used as an automated tool for data collection of the reports obtained from the two selected databases. In addition, all authors took on the role of reviewers and completed this task independently. Subsequently, a collective data validation process was implemented in which the authors worked together to validate and authenticate the results, ensuring a high level of consistency and reliability in the collection of bibliometric data.\n\nAn exhaustive search was conducted to find all relevant results and articles related to the research objective in the context of the current bibliometric research on smart restaurants. This was done using specific search equations created for each database, covering all articles that mentioned the topic of interest and ensuring that all relevant research was included. It is important to note that if any of the identified articles contain ambiguous or missing information, \"non-relevant texts\" are excluded from consideration. This was done to ensure that only findings that add to the understanding of the smart restaurant knowledge base and meet predefined relevance criteria are included to maintain consistency with the purpose and scope of the research.\n\nAll the authors worked diligently and collaboratively to collect data and assess the risk of bias in the included studies as part of the current bibliometric review of smart restaurants. Microsoft Excel®, an automated tool, was used to ensure consistency and uniformity in the process. This method ensured the quality and integrity of the results obtained and contributed to the reliability of the ongoing bibliometric research by allowing a rigorous and standardized assessment of possible biases in the included studies.\n\nIt is crucial to emphasize that effect measures that are more typical of primary research, such as hazard ratios and mean differences, are not appropriate in this example of source-based bibliometric research on smart restaurants. Instead, the results were analyzed and synthesized using bibliometric measures, including the number of publications, volume of citations, and timing of keyword use. The data were managed and analyzed using Microsoft Excel®. Although not translated into measurements, VOSviewer® was also used to explore thematic associations by identifying nodes and networks, providing users with a deep understanding of the dynamics and evolution of research in the field of smart restaurants.\n\nA procedure was used in this bibliometric research on smart restaurants to select studies that could be included in the synthesis. This involved collecting information on the attributes of a document, such as the number of publications, citations, and keywords, and comparing them with the predetermined inclusion criteria. In addition, data preparation techniques were used, including imputing missing summary statistics and performing data conversion where necessary. Bibliometric indicators of quantity, quality, and structure were used in the presentation and synthesis of the results according to predetermined guidelines. These indicators were automatically applied to all documents that survived the three exclusion stages (Durieux & Gevenois, 2010). Using this methodology, studies included in the bibliometric review were selected and analyzed for consistency and objectivity.\n\nIn the context of this bibliometric research on smart restaurants, it is important to be aware of possible information biases resulting from the use of synonyms found in thesauri, such as IEEE. These biases may be related to the lack of results in the synthesis. This vulnerability is reflected in the inclusion criteria, search approach, and data collection, where the choice of certain terms may affect the exclusion of relevant documents that use different vocabularies. Therefore, while bibliometrics must be accurate, the exclusion of irrelevant texts may result in the loss of important data for building comprehensive knowledge on the topic. This should be considered when interpreting the results and evaluating possible biases in bibliometric analyses.\n\nThis bibliometric study on Smart Restaurants includes a comprehensive assessment of the certainty of the body of evidence. Here, certainty is assessed globally as opposed to primary studies, where certainty is assessed individually. This was achieved by applying independent inclusion and exclusion criteria, defining and analyzing bibliometric indicators, and identifying and disclosing potential biases resulting from the design of the methodology. In addition, the limitations of the study are discussed in the Discussion section, providing a com-prehensive assessment of the accuracy and reliability of the bibliometric results. This im-proves our understanding of the validity and substance of the conclusions. Figure 1 summarizes the methodological design.\n\nOwn elaboration based on Scopus and Web of Science.\n\nIn the first phase of this bibliometric study on Smart Restaurants, articles were identified based on the search strategy used for each selected information source, and duplicate records were eliminated. The three exclusion phases listed in the methodology section were then carried out by applying relevance and quality standards. After a long process, 94 articles that met the established criteria were identified and selected to form the main corpus of this bibliometric study.\n\n\nResults\n\nAs shown in Figure 2, this bibliometric study provides a fascinating window for the evolution of smart restaurant research. The data showed a remarkable increase of 95.39 percent, suggesting an exponential growth pattern in the publication of articles related to this topic. Notably, the years with the highest number of publications are 2022, 2021, and 2018, suggesting that interest and research activities around smart restaurants increased significantly during these particular years. This may indicate the current relevance and timeliness of the topic in the scientific community. This temporal analysis sheds light on the changing nature of smart-restaurant research.\n\nOwn elaboration based on Scopus and Web of Science.\n\nAccording to the analysis of the main authors of this bibliometric study on smart restaurants, three groups can be identified. First, we can identify authors, such as Zhang, Yoon, and Hwang, who stand out in terms of their scientific productivity and the importance of their research, establishing them as leading figures in the field, as shown in Figure 3. Conversely, some authors (e.g., De Clercq, Wen, and Beck) value the impact and caliber of their work, even if their productivity rate is low. The third group of authors, Wan, Kato, and Aytac, for example, is characterized by high scholarly productivity, although not always in terms of citations, which enables them to actively advance our understanding of smart restaurants. This authorship analysis provides a deeper understanding of the current state of research in this field.\n\nOwn elaboration based on Scopus and Web of Science.\n\nAfter analyzing the main journals of this bibliometric on smart restaurants, they were able to observe three different groups (See Figure 4). First, publications such as the International Journal of Contemporary Hospitality Management stand out for their impact and productivity in the scientific community and have established themselves as leaders in the industry. On the other hand, there are publications on Waste Management, which are recognized for their impact and publication quality despite having a lower productivity index. Finally, the third group of journals is characterized by high scientific productivity, but the amount of research published rather than the number of citations is a better indicator of a journal’s influence. Examples of these journals are Technological Forecasting and Social Change and the Ijeai International Joint Conference on Artificial Intelligence. This journal analysis provides a deep understanding of the editorial dynamics in this research area.\n\nOwn elaboration based on Scopus and Web of Science.\n\nIn the analysis of the main countries in this bibliometric study of smart restaurants, three groups can be distinguished as can be shown in Figure 5. First, some nations stand out in terms of productivity and scientific impact, such as the United States, Taiwan, and China, consolidating their positions as leaders in this field of research. On the other hand, there are countries such as Singapore, Spain, and Canada, which despite having a lower productivity index, are recognized for their importance and caliber. The impact of the third group of nations, including India, Japan, and Indonesia, is measured more by the volume of research published than by the number of citations. This places these countries in a position to actively contribute to the body of knowledge on smart restaurants. The country analysis provides a comprehensive view of the geographic distribution of research in this area.\n\nAuthor's elaboration based on Scopus and Web of Science.\n\nThe current study, as shown in Figure 6, focused on the most used keyword in each research year to perform a comprehensive analysis of the thematic evolution in the Smart Restaurant literature between 2006 and 2023. As a starting point, it should be noted that 2006 is notable for the emergence of ideas such as “spike sorting.” Emerging topics such as “Radio-Frequency Identification” (RFID),” “Machine Learning,” “Tourism,” and “Systematic Literature Review\" become more prevalent over the years, reflecting changing research dynamics and areas of interest. This thematic analysis provides valuable information on the current developments and trends in the field.\n\nOwn elaboration based on Scopus and Web of Science.\n\nThe main keyword co-occurrence network is represented by these bibliometrics in four thematic groups, as shown in Figure 7. The red group, which includes terms such as “machine learning,” “big data analysis,” “opinion mining,” “Chinese restaurant process,” “prediction,” and “information extraction,” is the most prominent. The green group includes terms such as RFID, smart city, mobile application, edge computing, waiter robot, Arduino, and quick-service restaurant. A visual representation of the thematic interconnectedness in the scientific literature on this topic is provided by the identification of additional clusters in the area of smart restaurants, which are highlighted in blue and yellow, respectively. These groups capture the different aspects of conceptual affinity.\n\nOwn work based on Scopus and Web of Science.\n\nThis bibliometric study of smart restaurants uses an innovative method to create four different quadrants using a Cartesian plane that combines the frequency of keyword use on the x-axis and the validity of that use on the y-axis. This method is illustrated in Figure 8. Quadrant 4 contains ideas that show a decreasing trend in use over time, as shown by terms such as “Chinese restaurant process. Quadrant 2 contains words that, although they appear infrequently, have great relevance in the recent literature, placing them in the category of emerging words. “Big data analytics, robots, edge computing, and cloud computing are some examples of these keywords. Last but not least, Quadrant 1 includes well-established and expanding ideas such as “machine learning,” which is still widely used and has applications in smart restaurant research. This method provides a distinctive visual representation of keyword dynamics in relevant scientific literature.\n\nOwn elaboration based on Scopus and Web of Science.\n\n\nDiscussion\n\nThe Discussion section plays an essential role in the bibliometric analysis of smart restaurants. First, it thoroughly examines the research results, breaking down trends, patterns, and key findings. On the other hand, it provides a platform to explore the practical implications of these findings by identifying possible applications in the smart restaurant industry. Furthermore, it highlights the methodological limitations of the study, presents a classification of keywords according to their function in the literature, and highlights the main gaps in the research. Finally, it establishes a research agenda that identifies priority areas for future studies in the field of smart restaurants, thereby contributing to the understanding and continuous development of this topic.\n\nIn 2022, 2021, and 2018, a significant increase in scientific production related to smart restaurants was observed, with research addressing different aspects of this field with constant development. In 2022, research that analyzed the digital food environment was highlighted, providing a comprehensive view of how technology affects the food industry and its impact on the development of smart restaurants (Granheim et al., 2022). Additionally, service quality in smart restaurants has been explored, identifying the relationship between “smart dining,” “smart restaurant,” and service quality (SSQ) (Wong et al., 2022). These studies reflect the growing interest in understanding how technology reshapes customer experience and management in the restaurant industry.\n\nIn 2021, the impact of augmented reality on the customer experience in restaurants was examined, specifically in the case of “Le Petit Chef.” This study highlights how augmented reality technology is used to enhance culinary experience (Batat, 2021). On the other hand, the potential of artificial intelligence and robotics in the restaurant sector in Europe was explored, assessing the potential for innovation in a highly interactive sector (Blöcher & Alt, 2021). These studies highlight the growing integration of advanced technologies in the restoration sector and the need to understand their impact on the industry.\n\nSimilarly, in 2018, the focus was on food waste management in restaurants through the implementation of an Internet of Things (IoT) network. This study highlights how technology contributes to sustainability and efficiency in restaurant management (Wen et al., 2018). In addition, warnings were issued regarding the arrival of automation in the hospitality industry, raising questions about the intrusion of robots into restaurant service delivery (Bowen & Morosan, 2018). These studies anticipate the technological changes that occur in the hospitality industry and the need to adapt to these changes.\n\nZhang, Yoon, and Hwang have been recognized in the field of smart restaurants for their valuable contributions in several key areas. In particular, Zhang has noted his work on food waste management in restaurants, as seen in his study titled “Design, Implementation, and Evaluation of an Internet of Things (IoT) Network System for food waste management in restaurants.” (Wen et al., 2018) This study demonstrated the effectiveness of implementing an IoT network for food waste management in restaurants, which is an essential aspect of sustainability and efficiency in the restaurant industry.\n\nOn the other hand, Yoon and Hwang have become research leaders for their studies on privacy and customer experience in restaurants. Their research, such as “Desired Privacy and the Impact of Crowding on Customer Emotions and Attraction-Avoidance Responses: Waiting in a Virtual Reality Restaurant” (Hwang et al., 2012) and “Where Would You Like to Sit? Understanding customers’ privacy-seeking tendencies and location behaviors to create effective restaurant environments (Hwang & Yoon, 2009) sheds light on how privacy and space arrangements in restaurants can influence emotions and customer responses. This research contributes to a better understanding of how to design effective restaurant environments and improve customer experience.\n\nAs for De Clercq, Wen, and Beck, their importance lies in their contribution to the aforementioned study, “Design, implementation, and evaluation of an Internet of Things (IoT) network system for food waste management in restaurants” (Wen et al., 2018). Their participation was instrumental in addressing a significant problem in the restaurant industry and food waste management through the application of the IoT technology. Their innovative approach and search for practical solutions to restaurant management challenges have established them as leaders in this research field.\n\nThe Journal of Contemporary Hospitality and Waste Management has played an essential role in promoting the advancement of knowledge in the field of smart restaurants by publishing highly relevant and impactful research in this discipline.\n\nThe Journal of Contemporary Hospitality Management stands out for its contributions in various areas, such as customer experience management in smart restaurants. Studies such as “Desired Privacy and the Impact of Crowding on Customer Emotions and Approach-Avoidance Responses: Waiting in a Virtual Reality Restaurant” (Hwang et al., 2012) have explored how privacy and the physical environment can influence customers’ emotions and responses. Similarly, “Artificial intelligence for big data analytics in hospitality: development of a prediction model for the usefulness of restaurant reviews for customer decision making” (Lee et al., 2021) has addressed the application of artificial intelligence in data management for decision making in the restaurant industry, contributing to the advancement of data-based strategies in this sector.\n\nOn the other hand, Waste Management has been a key journal in promoting sustainable management in restaurants, especially with regard to food waste management. The study “Design, implementation and evaluation of an Internet of Things (IoT) network system for food waste management in restaurants” (Wen et al., 2018) published in this journal has become a reference in the implementation of IoT technology to address the problem of food waste in restaurants. This study has had a significant impact on the restaurant industry by presenting innovative and sustainable solutions for waste management.\n\nBoth journals have contributed significantly to the advancement of research in the field of smart restaurants, and have allowed the dissemination of essential knowledge for the hospitality industry and restaurant management. Its influence continues in both the academic community and the professional practice of the field.\n\nResearch on smart restaurants has been led by different countries, each of which has made significant contributions to the advancement of the field. The United States has played an integral role in smart restaurant research, with notable studies such as “Beware Hospitality Industry: The Robots Are Coming,” (Bowen & Morosan, 2018) which focuses on the automation and integration of robots in the hospitality industry, and “Learning user preferences with multiple information fusion for restaurant recommendations,” (Fu et al., 2014) which focuses on learning user preferences for restaurant recommendations. These investigations have had a significant impact on the adoption of emerging technologies in restaurant management worldwide.\n\nIn Taiwan, the focus on customer experience in smart restaurants is noteworthy, as shown in “What Drives Experiential Loyalty Towards Smart Restaurants? The Case Study of KFC in Beijing,” (H. C. Wu & Cheng, 2018), which analyzes customer loyalty in smart restaurants, and “A deep learning facial recognition-based scoring system for restaurants” (Chang et al., 2019) which uses facial recognition to assess customer experience. This research contributes to a deeper understanding of how smart restaurants can improve customer experience and foster loyalty.\n\nChina has emerged as a leader in smart restaurant research, with studies such as “smart dining, smart restaurants, and smart service quality (SSQ).” (Wong et al., 2022) that focus on service quality in smart restaurants. In addition, “Learning Representations for Appearance Categories Detection in Online Reviews” (Zhou et al., 2015) addresses the identification of appearance categories in online reviews, which is crucial for reputation management. restaurants. These studies have contributed significantly to the understanding of how technology can improve service quality in the restaurant industry.\n\nIn contrast, Singapore has excelled in research on sentiment analysis in smart restaurants, as reflected in “Vistanet: Visual Attention Network for Multimodal Sentiment Analysis” (Truong & Lauw, 2019). This study focuses on multimodal sentiment analysis to understand customer opinions in smart restaurants. Spain has contributed with research such as “Grab, pay and eat: Semantic food detection for smart restaurants” (Aguilar et al., 2018), which focuses on semantic food detection to improve the efficiency of order management in restaurants. Canada is also working on smart restaurant management, as seen in \"Near-field communication sensors and cloud-based smart restaurant management system,” (Saeed et al., 2016) which uses near-field communication sensors and the cloud to efficiently manage restaurants, and “Smart restaurants: Customer Demand Survey and Sales Forecasting” (Lasek et al., 2016), which explores customer demand and sales forecasting in smart restaurants.\n\nThe concept of “spike sorting” played a central role in the early years of smart restaurant research. This approach focuses on solving data management and information categorization challenges in restaurant systems (Wood et al., 2006). Spike sorting enables the analysis and organization of critical data, which is valuable for managing real-time information in restaurant environments. As research progressed, the horizon expanded to include concepts such as RFID, machine learning, tourism, and systematic literature reviews, reflecting the need to address broader challenges and leverage emerging technologies to improve the efficiency and quality of intelligent restaurant management. This shift in conceptual focus underscores the adaptation of research to the changing needs of the restaurant industry, and highlights the importance of staying abreast of technological trends in the field.\n\nConcepts such as RFID, machine learning, tourism, and systematic literature reviews have gained significant prominence in current research on smart restaurants. Radio Frequency Identification (RFID) technology has revolutionized inventory management and logistics in the restaurant industry, enabling the real-time tracking of products and improving operational efficiency. Meanwhile, machine learning is being used to personalize customer experience, from menu suggestions to reservation management, improving both customer satisfaction and restaurant efficiency (Wong et al., 2022). Tourism and hospitality are important areas for smart restaurant research, as technological advancements are changing the way tourists interact with restaurant services at tourist destinations. Furthermore, the focus on a systematic literature review has allowed researchers to understand the current state of knowledge in the field of smart restaurants and identify critical gaps that require additional research (Singh et al., 2023). These concepts have contributed to the growth and development of smart restaurant research, addressing fundamental questions and improving the efficiency and quality of service in the industry.\n\nThe thematic cluster lists the most frequently used keywords and their relationships, as shown in Figure 7. Among these words, the main cluster was related to machine learning. This cluster is made up of terms such as: sentiment analysis, big data and analytics, restaurant reviews, opinion mining, recommendation system and prediction. These words usually constitute research oriented towards the use of predictive models built from data from previous reviews and opinions of consumers to facilitate decision making (Hajek & Sahut, 2022) and provide options according to consumer preferences. users (Lee et al., 2021). Also found in the same cluster are the terms: Chinese restaurant process, context awareness, connected restaurant, and Gibbs sampling.\n\nThe following cluster consists of the terms RFID, Raspberry Pi, Arduino, IoT architecture, smart cities, mobile applications, cloud computing, edge computing, and waiter robots. These terms form a cluster because of the use of RFID technology from the use of devices such as Raspberry and Arduino, which are integrated to form the IoT architecture and are part of the infrastructure of smart spaces, hosting the data in the cloud (Harpanahalli et al., 2020; Wen et al., 2018). The following network of words comprises terms such as deep learning, facial recognition, restaurant automation, restaurant scoring, and unmanned restaurants, which are mainly part of the research that aims to automate restaurants using deep learning techniques based on restaurant scoring data (Bonela et al., 2023; Dampage et al., 2021). The last cluster consists of the terms fraud detection, restaurant industry, robotics, online reviews, text mining, social media, Yelp and Pokemon Go.\n\nThe concept of “Chinese Restaurant Process” (CRP) was located in quadrant 4 of the Cartesian plane, indicating that its frequency of use has decreased compared to previous periods in smart restaurant research. CRP is a statistical technique used in data modeling, particularly in the fields of artificial intelligence and machine learning. Historically, CRP has been applied in contexts such as neural signal processing and decision making in dynamic environments (Wood et al., 2006). It has also been used to improve reinforcement learning algorithms (Shi et al., 2018), where it is used in the context of reinforcement learning to improve decision-making efficiency.\n\nHowever, as research on smart restaurants has progressed, CRP may become less relevant in this specific context. The decline in its use may reflect changes in research trends and an increased focus on other concepts more relevant to the smart restaurant industry, such as RFID, machine learning, and others mentioned above. This suggests that smart restaurant researchers are exploring new directions and adapting their approaches to meet technological and industrial needs.\n\nQuadrant 2 of the Cartesian plane, which houses novel concepts in the scientific field of smart restaurants, reflects the growing importance of terms such as big-data analytics, robots, edge computing, and cloud computing in the present and near future. These terms are indicative of technological trends and the need to adapt to ever-changing business environments.\n\nThe practice of big data analytics has established itself as a fundamental pillar in restaurant management, enabling the collection and analysis of large amounts of data with the aim of obtaining valuable information about customer preferences and supporting decision-making (Lee et al., 2021). On the other hand, the integration of robots in restaurants is revolutionizing the industry by automating tasks such as food and customer services, helping to improve both efficiency and customer experience (Akhund et al., 2020).\n\nEdge and cloud computing play essential roles in facilitating real-time connectivity and data processing in smart restaurants (Aytaç & Korçak, 2018). These technologies enable rapid data transfer between Internet of Things (IoT) devices and central systems, which is critical for efficient restaurant management (Saeed et al., 2016).\n\nQuadrant 1 of the Cartesian plane, where concepts are constantly growing and occupying a central and consolidated place in smart restaurant research, highlights the current and future importance of machine learning. Machine learning, also known as automatic learning, has played a fundamental role in restaurant management, allowing companies to analyze large datasets and extract valuable information to support strategic decision-making. This study illustrates how Machine Learning has been used to analyze restaurant review data and to develop predictive models to help customers make decisions (Lee et al., 2021).\n\nIn addition, machine learning has been successfully applied to fraud detection in the restaurant industry (Hines & Youssef, 2018), where it is used to identify rotating check fraud. This technology is essential for identifying unusual patterns and for preventing potential financial losses.\n\nTable 1 presents a classification of emerging and growing keywords related to the development of smart restaurants, according to their functions. This allows the identification of the main characteristics and applications associated with each category, highlighting constantly growing areas of interest and prominent approaches in current research.\n\nConducting bibliometrics on smart restaurants with a focus on different dimensions, such as the frequency of publications over the years, identification of prominent theoretical references, monitoring of thematic evolution, analysis of the co-occurrence of keywords, exploration of emerging and growing keywords, and identification of unexplored areas of research, has important theoretical implications for the advancement and understanding of this field of study.\n\nFirst, analyzing the frequency of publications over the years provides an overview of the dynamics of research over time. This allowed us to identify periods of greater research activity, thus revealing the relevance and interest in the topic at different times. The identification of key years, such as 2022, 2021, and 2018, with a high number of publications suggests the importance of these periods for the development of smart restaurant research.\n\nIdentification of the main theoretical references in bibliometrics provides a deeper understanding of the theoretical and methodological approaches that have guided research in this field. Authors such as Zhang, Yoon, Hwang, De Clercq, Wen, and Beck stand out in terms of productivity and impact, indicating their significant contribution to the existing body of knowledge.\n\nFurthermore, the analysis of thematic evolution in bibliometrics reflects how smart restaurant research evolved from initial concepts such as “spike sorting” in the early years to emerging and consolidated areas such as “machine learning,” RFID, “edge computing,” and “big data analytics.” This analysis not only helps to understand the historical trajectory of research but also identifies current and future trends in the field.\n\nKeyword co-occurrence analysis reveals the conceptual structure of the research field and how key terms are related. The identification of thematic clusters and highlighted keywords within each cluster provided an overview of the most relevant areas of focus in smart restaurants.\n\nThe analysis of emerging and growing keywords allows for the identification of concepts that are gaining importance and are expected to have a significant impact on future research. Terms such as “robots,” “edge computing,” and “cloud computing” emerge as areas of emerging interest\n\nsuggesting new directions for research\n\nFinally, the identification of unexplored research areas is a crucial implication of bibliometrics. By analyzing the gaps in the existing literature, valuable guidance is provided to researchers and academics interested in addressing underexplored areas or identifying unresolved research questions in the field of smart restaurants. These gaps can serve as a starting point for future research and contribute to the theoretical and practical advancement of the discipline.\n\nConducting bibliometrics on smart restaurants with a focus on thematic evolution and keyword analysis has practical implications of great relevance for both the restaurant industry and academic community. The transition from research focused on the concept of spike sorting to emerging areas, such as RFID, machine learning, tourism, and systematic literature review, reflects a growing understanding of the importance of technology and data management in improving efficiency and quality in the restaurant industry. This means that industry professionals must stay abreast of emerging trends and consider implementing technologies, such as machine learning and big data analytics, to optimize decision making and improve customer experience.\n\nThe identification of thematic clusters and the conceptual relationship between key terms, such as machine learning, big data analytics, and restaurant reviews, highlight the importance of interdisciplinary research and collaboration between researchers and professionals from different fields. This suggests that smart restaurants can benefit from a comprehensive approach that integrates advanced technologies with a deep understanding of customer preferences and opinions, which in turn can influence strategic decision-making and service quality management.\n\nAnalyzing keyword frequency and relevance provides valuable insights into current trends in the industry. The emergence of concepts such as big data analytics, robots, edge computing, and cloud computing underscores the importance of digitization and automation in the restaurant industry. This suggests that smart restaurants can improve their operational efficiency and service quality by implementing advanced technologies such as robots and cloud-based management.\n\nHowever, the continuous increase in the frequency of keywords related to Machine Learning indicates that this approach remains relevant and crucial in the research and application of technologies in smart restaurants. The practical implications of this finding are significant as they highlight the importance of training and adopting machine learning models for a variety of applications, such as service personalization and customer data management.\n\nIn addition, bibliometrics provide insights into best practices in research and methodology in the field of smart restaurants. This knowledge can be used by researchers and academics to understand the most effective techniques and approaches for generating knowledge in this area. Likewise, the analysis of thematic development can serve as a guide to identifying areas of research that have received disproportionate attention or, on the contrary, have been insufficiently addressed, which can improve the quality and relevance of future research projects.\n\nAnother important practical implication is related to education and training in hospitality and restaurant management. The results of bibliometrics can be used to develop education and training programs to prepare students and professionals to meet the changing technological demands of the food and hospitality industry. Emerging and growing concepts, such as machine learning or big data analytics, can be incorporated into curricula to ensure that future professionals acquire the necessary skills.\n\nThe research offers practical guidance for restaurant managers and owners to effectively adopt emerging technologies in their operations. By identifying key trends and areas of focus, industry professionals can make informed decisions about implementing technological solutions that improve operational efficiency and service quality. Keyword analysis highlights the importance of interdisciplinary collaboration and integrating technology with a deep understanding of customer preferences and opinions. This underscores the need for strategic management that combines technical knowledge with a customer-focused vision to enhance the dining experience and increase loyalty.\n\nThe research has a significant impact on the food and beverage industry by providing a roadmap for innovation and continuous improvement. It offers detailed insights into trends and best practices in the smart restaurant space, helping professionals keep up with technological developments and changing consumer demands. This benefits restaurant owners by improving their competitiveness and profitability, while also contributing to customer satisfaction and the sustainable growth of the industry as a whole.\n\nThe research emphasizes the significance of hotel and restaurant management training and education from a societal perspective. This is to prepare the next generation of professionals to meet the industry’s technological challenges by integrating emerging technological concepts and tools into curricula. Educational institutions can ensure that students acquire the skills necessary to thrive in an increasingly digitalized business environment. Encouraging the adoption of technologies that improve efficiency and quality of service can contribute to the creation of employment and economic development in the restaurant sector.\n\nThe present bibliometrics on smart restaurants, carried out using the PRISMA-2020 methodology and based on the Scopus and Web of Science databases, provides a valuable overview of research in the field. However, as is usually the case with bibliometric studies, it has certain limitations that must be considered when interpreting results. First, it is important to emphasize that the quality and completeness of the databases used may have affected the representativeness of the results. Despite efforts to cover a wide range of academic publications, some relevant research may have been absent or underrepresented in the selected databases. In addition, inherent limitations in keywords and search criteria may have influenced the selection of articles for inclusion in the analysis, which could have affected the breadth and depth of the identified research landscape.\n\nAnother limitation is related to the quality of the bibliographic data available in the databases. Errors in the cataloging of publications, lack of uniformity in the introduction of metadata, and duplication of records may have influenced the precision of the bibliometric indicators used in this study. Furthermore, differences in how publications are cited in different disciplines and contexts may have influenced keyword co-occurrence analysis and the identification of thematic trends.\n\nFinally, it is important to recognize that while bibliometrics provide a valuable overview of the scientific production in a field, they do not have the capacity to fully capture the quality, relevance, or real impact of research. The sheer number of publications does not always reflect the depth or influence of a field of study, and it is essential to complement bibliometric findings with qualitative assessments and critical literature reviews to obtain a complete understanding of the current state of smart restaurant research.\n\nTable 2 summarizes the main underexplored research areas identified in the bibliometric study of smart restaurants. These areas represent areas where future research could focus on advancing our knowledge. Highlighted areas of need include further exploration of the implementation of emerging technologies, such as artificial intelligence and the Internet of Things, in restaurant management and operations. The importance of exploring how digital experiences and customer interactions reshape the restaurant industry and how they can be optimized is also highlighted.\n\nOther areas highlighted in the table include the lack of research specifically dedicated to sustainability in smart restaurants as well as the lack of studies that take a detailed look at data management and cybersecurity in this context. The influence of cultural and regional factors on the adoption of smart technologies in restaurants is another area that is considered relevant for future research.\n\nRobots are essential components of the smart restaurant industry. Their relevance lies in their ability to automate repetitive tasks, improve operational efficiency, and provide customers with unique experiences. In future research, it will be possible to explore more advanced approaches to programming robots for specific tasks, such as food delivery and interaction with diners. In addition, research can be conducted on how artificial intelligence and machine learning can improve the ability of robots to adapt to changing restaurant environments, including the ability to recognize and respond in a more personalized manner to customer preferences (see Figure 9).\n\nAuthor's elaboration based on Scopus and Web of Science.\n\nAnother interesting aspect that could be the subject of future research is the ethics of robotics in restaurants. Addressing issues related to the privacy and security of data collected by robots as well as the impact on the quality of employment in the service industry are areas that deserve further analysis. In addition, investigating how restaurants can ensure customer comfort and acceptance of robots in their establishments is critical for promoting their adoption and ensuring their continued success.\n\nThe concept of the Chinese Restaurant Process (CRP) has been central to smart restaurant research, but its relevance has waned in recent years. However, this still offers significant potential for future research. Potential applications of CRP can be explored in resource allocation and inventory management in restaurants, with the goal of improving operational efficiency and reducing food waste. In addition, CRP can have significant implications in analyzing customer preferences and customizing menus based on order history.\n\nOn the other hand, the concept of “service robots” has become increasingly important in the context of smart restaurants. Future research could focus on how to effectively integrate service robots with CRP to improve both the customer experience and operational efficiency. In addition, restaurant review analysis is critical for decision making and customer feedback. Future studies could investigate how CRP can contribute to the automated analysis of restaurant reviews, and how to improve service quality based on customer feedback. These investigations have the potential to revitalize the relevance of CRP in the context of smart restaurants and contribute to the advancement of the industry.\n\n\nConclusions\n\nAfter analyzing the bibliometrics of smart restaurants, it is clear that interest and research in this area has experienced a significant increase in recent years. In particular, 2021 and 2022 have emerged as the most productive years in terms of scientific production on the topic. This increase is not linear; however, the number of scientific articles has grown exponentially by an impressive 95.36%. This dynamic demonstrates strong academic and professional enthusiasm for further understanding and developing this emerging industry.\n\nIn terms of the main references of the research, Zhang H, Yoon S-Y and Hwang J stand out as the most influential authors in the field, with the International Journal of Contemporary Hospitality Management serving as the main vehicle for the dissemination of these studies. It is also interesting to note the geographical concentration of scientific production, with the United States, China, and Taiwan being pioneering countries in research on smart restaurants.\n\nAt the thematic level, there has been evolution in the topics studied. In the beginning, research focused on more technical aspects such as spike sorting; today, it has diversified to cover areas such as tourism and systematic literature review. However, the main thematic clusters highlight the incorporation of artificial intelligence and data analysis techniques into the sector, with key concepts such as machine learning, sentiment analysis, and recommendation systems leading the discussion.\n\nThe field is constantly evolving, as evidenced by the emergence of keywords. Consolidated concepts, such as machine learning, coexist with growing terms, such as big data analytics, robots, perceived ease of use, and edge computing. These terms not only highlight current trends but also suggest possible directions for future research.\n\nFinally, in the future, the smart restaurant research agenda should focus on deepening these emerging and consolidated concepts. Given the rapid evolution of this field, it is essential to maintain an up-to-date perspective and be prepared to address the challenges and opportunities that these technological and thematic changes will bring to the design and operation of restaurants.\n\n\nEthics and consent\n\nEthical approval and consent were not required.\n\nReporting guidelines: PRISMA checklist DOI: https://doi.org/10.5281/zenodo.14146154 (Valencia Arias and Cardona Acevedo, 2024).\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).", "appendix": "Data availability\n\nThe data availability statement for this study has been duly registered and archived in the Zenodo open data repository, which is recognized for its commitment to the accessibility and preservation of scientific data. The data and materials supported by this study are publicly available under a Creative Commons Attribution 4.0 International (CC BY 4.0) license and can be accessed at the following DOI link: https://doi.org/10.5281/zenodo.14146154. (Valencia Arias and Cardona Acevedo, 2024).\n\n\nAcknowledgements\n\nNot applicable.\n\n\nReferences\n\nAguilar E, Remeseiro B, Bolaños M, et al.: Grab, pay, and eat: Semantic food detection for smart restaurants. IEEE Trans. Multimed. 2018; 20(12): 3266–3275. Publisher Full Text\n\nAkhund TMNU, Siddik MAB, Hossain MR, et al.: IoT Waiter Bot: a low cost IoT based multi functioned robot for restaurants. 2020 8th International Conference on Reliability, Infocom Technologies and Optimization (Trends and Future Directions) (ICRITO). 2020; 1174–1178. Publisher Full Text\n\nAlmeida S, Mesquita S, Pereira C: Smart Hospitality: Goodbye Virus!.2023. Publisher Full Text\n\nAytaç K, Korçak Ö: IoT edge computing in quick service restaurants. 2018 16th International Symposium on Modeling and Optimization in Mobile, Ad Hoc, and Wireless Networks (WiOpt). 2018; 1–6. Publisher Full Text\n\nBai H, Wu X: Design and Implementation of Smart Air Catering Distribution System Under Tourism Contactless Scenes. 2022 European Conference on Communication Systems (ECCS). 2022; 59–63. Publisher Full Text\n\nBatat W: How augmented reality (AR) is transforming the restaurant sector: Investigating the impact of “Le Petit Chef” on customers’ dining experiences. Technol. Forecast. Soc. Chang. 2021; 172: 121013. Publisher Full Text\n\nBlöcher K, Alt R: AI and robotics in the European restaurant sector: Assessing potentials for process innovation in a high-contact service industry. Electron. Mark. 2021; 31: 529–551. Publisher Full Text\n\nBonela AA, He Z, Nibali A, et al.: Audio-based Deep Learning Algorithm to Identify Alcohol Inebriation (ADLAIA). 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Journal of Science and Technology Policy Management. 2023; 15: 1037–1055. (ahead-of-print). Publisher Full Text\n\nLasek ACNSJ, Cercone N, Saunders J: Smart restaurants: survey on customer demand and sales forecasting. Smart cities and homes. 2016; 361–386. Publisher Full Text\n\nLee M, Kwon W, Back KJ: Artificial intelligence for hospitality big data analytics: developing a prediction model of restaurant review helpfulness for customer decision-making. Int. J. Contemp. Hosp. Manag. 2021; 33(6): 2117–2136. Publisher Full Text\n\nMartino B, Colucci Cante L, Cerullo N: Opportunities and Advantages of Cloud Migration of a Smart Restaurant System.2022. Publisher Full Text\n\nMishra N, Goyal D, Elngar AA: IoT Impact and Challenges on Robotic Waiters in Automation of Restaurants and Hotels. Research Anthology on Cross-Disciplinary Designs and Applications of Automation. IGI Global; 2022; pp. 171–189. Publisher Full Text\n\nPage MJ, McKenzie JE, Bossuyt PM, et al.: The PRISMA 2020 statement: an updated guideline for reporting systematic reviews. Int. J. Surg. 2021; 88: 105906. PubMed Abstract \| Publisher Full Text\n\nPrathibanandhi K, Janani R, Shruthi M, et al.: Augmented Reality-Based Restaurants. 2022 International Conference on Power, Energy, Control and Transmission Systems (ICPECTS). 2022; 1–4. Publisher Full Text\n\nSaeed H, Shouman A, Elfar M, et al.: Near-field communication sensors and cloud-based smart restaurant management system. 2016 IEEE 3rd World Forum on Internet of Things (WF-IoT). IEEE; 2016; pp. 686–691. Publisher Full Text\n\nShi H, Yang S, Hwang KS, et al.: A sample aggregation approach to experiences replay of Dyna-Q learning. IEEE Access. 2018; 6: 37173–37184. Publisher Full Text\n\nSingh PR, Nazre Amarnath TK, Khurram M, et al.: Restaurant Automation Through IoT and NLP Techniques.2023; 123–138. Publisher Full Text\n\nTruong QT, Lauw HW: Vistanet: Visual aspect attention network for multimodal sentiment analysis. Proceedings of the AAAI Conference on Artificial Intelligence. 2019; 33(01): 305–312. Publisher Full Text\n\nValencia-Arias A, Cardona-Acevedo S: Trends in Smart Restaurant Research: Bibliometric Review and Research Agenda. [Data set]. Zenodo. 2024. Publisher Full Text\n\nValencia-Arias A, Ramírez Dávila J, Londoño-Celis W, et al.: Research Trends in the Use of the Internet of Things in Sustainability Practices: A Systematic Review. Sustainability. 2024; 16(7): 2663. Publisher Full Text\n\nVieira E, Gomes J: A comparison of Scopus and Web of Science for a typical university. Scientometrics. 2009; 81(2): 587–600. Publisher Full Text\n\nWen Z, Hu S, Clercq D, et al.: Design, implementation, and evaluation of an Internet of Things (IoT) network system for restaurant food waste management. Waste Manag. 2018; 73: 26–38. PubMed Abstract \| Publisher Full Text\n\nWong IA, Huang J, Lin Z, et al.: Smart dining, smart restaurant, and smart service quality (SSQ). Int. J. Contemp. Hosp. Manag. 2022; 34(6): 2272–2297. Publisher Full Text\n\nWood F, Goldwater S, Black MJ: A non-parametric Bayesian approach to spike sorting. International Conference of the IEEE Engineering in Medicine and Biology Society. 2006; 2006: 1165–1168. PubMed Abstract \| Publisher Full Text\n\nWu CY, Huang KH: A framework for off-line operation of smart and traditional devices of IoT services. Sensors. 2020; 20(21): 6012. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWu HC, Cheng CC: What drives experiential loyalty toward smart restaurants? The case study of KFC in Beijing. J. Hosp. Market. Manag. 2018; 27(2): 151–177. Publisher Full Text\n\nZhou X, Wan X, Xiao J: Representation learning for aspect category detection in online reviews. Proceedings of the AAAI Conference on Artificial Intelligence. 2015; 29(1). Publisher Full Text" }	[ { "id": "355093", "date": "03 Feb 2025", "name": "Rainer Alt", "expertise": [ "Reviewer Expertise Business information systems", "Electronic Business", "Digital Platforms" ], "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\nThank you for the opportunity to review your paper on restaurant technologies, which I consider as highly relevant, given the increasing digitalization of the hospitality sector and the potential benefits of technological advancements for operational efficiency and customer experience in this domain. I agree that a well-conducted systematic literature review could provide valuable insights into current research trends, gaps, and future directions. However, despite the significance of this research area, the present paper suffers from substantial methodological shortcomings, a discussion section that fails to engage critically with the findings, and formal writing issues. Due to these concerns, the paper should not be recommended for indexed in its current form.\n1. Methodology First, the literature review yields only a small set of publications that is not sufficient for claiming a systematic review in the smart restaurant sector. Second, the validity of the literature review is limited since the exclusion criteria are not clearly defined, leading to ambiguity in how studies were selected. For instance, the phrase \"Only systematic literature reviews are subject to the second phase of exclusion, which involves eliminating all documents for which the full text is not available\" on page 5 raises concerns about whether studies without full-text access were still included in the first phase. Third, the selection of databases (Web of Science and Scopus) raises questions about completeness. While these databases are highly regarded, they may not cover all relevant publications in the field of smart restaurant technologies. Fourth, the search strings differ between Scopus and WoS, which could lead to inconsistent results. The inclusion of separate search strings in uppercase and lowercase is unnecessary, as search engines are generally case-insensitive. In addition, the selection of technologies appears arbitrary, including some while excluding others without justification (no robots, machine learning etc.) and the use of abbreviations in the search string appears inconsistent (why not \"AI\" as an abbreviation). Fifth, elements such as assessment for reporting bias and study of risk bias assessment add little value to the argumentation, as they are borrowed from medical research and do not fit the context of this study. On the other hand, important methodological details such as the exact time frame of the review are missing, significantly undermining the study’s credibility.\n2. Contribution A well-structured discussion section should critically engage with the findings. However, in this paper, the discussion is largely a repetition of the results without meaningful interpretation or critique. The first part of the discussion, in particular, reads more like a continuation of the results section rather than an analysis of what these findings mean. Additionally, certain elements in the discussion raise questions about the selection and categorization of studies. The paper refers to \"systematic literature reviews\" (SLR) as a \"concept of smart restaurants,\" but SLRs are a methodological approach, not a conceptual framework. The inclusion of the \"Chinese Restaurant Process\" (CRP), which is a mathematical theorem, is questionable. One of the identified papers concerning CRP, titled \"A Sample Aggregation Approach to Experiences Replay of Dyna-Q Learning,\" does not appear to be directly related to smart restaurant research. This suggests problems in properly excluding irrelevant studies. The lack of critical engagement in the discussion significantly weakens the paper’s contribution to the field. Rather than simply reiterating the findings, the discussion should aim at obtaining deeper insights and possibly also a more elaborate research agenda (see page 19).\n3. Formal issues From the formal side, the paper requires thorough proofreading and editing. The excessive number of paragraphs disrupts readability and cohesion, making certain sections appear fragmented. For instance, the sentences \"Technology adoption in the restaurant sector remains low due to a variety of interrelated factors. first, resistance to change by restaurant owners and managers can be a significant barrier, many of which may be rooted in traditional practices. and may be reluctant to invest in technology due to concerns about cost, implementation complexity, and fear of operational disruption. and a lack of knowledge and understanding of how to effectively integrate technology into restaurant operations can lead to uncertainty and avoidance. adoption.\" on page 3 contains multiple abrupt transitions. Additionally, the inconsistent use of hyphenation, such as \"ap-propriate\" on page 4 indicates a lack of standardization in formatting. The term \"Restoration\" is used on page 12 instead of \"restaurants,\" which is misleading and likely a typographical error. On page 5, the phrase \"[...] current restaurant bibliometrics. intelligent\" is incomplete. These formal errors create the impression of an unfinished manuscript and hinder comprehension, reducing the overall professionalism of the paper.\nI trust that the authors find this feedback helpful and wish them all the best in further developing their work.\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?\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/13-1505
https://f1000research.com/articles/13-1504/v1	09 Dec 24	{ "type": "Genome Note", "title": "The first mitochondrial genome of Haemagogus equinus from Jamaica", "authors": [ "Simone L. Sandiford", "Simmoy A. A. Noble", "Samra A. Pierre", "Douglas E. Norris", "Renee Ali", "Simmoy A. A. Noble", "Samra A. Pierre", "Douglas E. Norris", "Renee Ali" ], "abstract": "In the Americas, the expansion in incidence of arboviral infections including Mayaro virus (MAYV) has drawn attention to the resurgence of viruses associated with understudied arthropods. Mosquitoes belonging to the genus Haemagogus are generally geographically restricted to the forests of Central and South America and the Caribbean and are the known sylvan vectors for yellow fever virus and emerging MAYV. With an established population in Jamaica, Haemagogus equinus has been reported to be well-adapted to oviposition in artificial containers close to human populations. Its role in arboviral transmission however is not fully understood. Given the dearth of genetic information and the difficulty in morphologically identifying cryptic features in species belonging to this genus, we report the first mitochondrial genome of Hg. equinus. Using a genome skimming approach, two Hg. equinus mosquito specimens were sequenced using the Illumina Novaseq 6000 platform. A representative mitogenome of 16,471 bp, 80.7% AT and 37 genes was assembled using NOVOplasty. Phylogenetic analysis placed Hg. equinus in the Albomaculatus section of the Haemagogus subgenus supporting previously described taxonomic studies.", "keywords": [ "Haemagogus equinus", "mitogenome", "Jamaica", "arboviruses", "genome skimming" ], "content": "Introduction\n\nMosquitoes of the genus Haemagogus (Hg.) Williston, 1896 are endemic to tropical rainforests, open deciduous forests and mangroves in Latin America and the Caribbean.1 The genus is divided into two subgenera (Haemagogus and Conopostegus), with the Haemagogus subgenus split into three sections (Albomaculatus, Splendens, Tropicalis).1 Being the primary sylvatic vectors for yellow fever virus (YFV)1 and the emerging arboviral threat Mayaro virus (MAYV),2 species in this genus of mosquitoes are of significant medical importance in the region. Despite this, many species remain understudied and their role in disease transmission has not been clearly defined. In Jamaica, Haemagogus equinus Theobald, 1903 is presently the only known species on the island. Established populations of Hg. equinus have also been identified in southern Texas in the United States of America, Mexico, Belize, Guatemala, El Salvador, Honduras, Nicaragua, Costa Rica, Panama, Colombia, Guyana, Venezuela and Trinidad and Tobago.1\n\nWhile the vectorial capacity of Hg. equinus has not been fully elucidated, laboratory and transovarial transmission of YFV3–6 along with natural infections among wild mosquitoes have been reported.7,8 In spite of this, its importance in arboviral transmission in some localities remains uncertain.9 Although Hg. equinus primarily oviposits and larvae develop in tree holes and bamboo internodes,1 the species is very adaptable and can also utilize rock holes,10 domestic containers and used tires.11,12 With multiple reports of MAYV in neighbouring Haiti,13–15 there is need for a greater understanding of the role of this sylvatic vector in arbovirus transmission. To facilitate this, accurate identification of all life stages of field collected specimens is crucial. However, impediments such as a lack of taxonomic expertise coupled with poor quality samples and a paucity of reference molecular data continue to hamper these efforts in Jamaica.\n\nFew studies have employed genomic sequencing to investigate the molecular taxonomy and phylogenetic relationships of Hg. equinus mosquitoes. Currently, only partial sequences of cytochrome oxidase I and other mitochondrial or nuclear genes are available from NCBI’s GenBank or BOLD systems databases.16–21 The complete mitochondrial genome (mitogenome) is increasingly being used for evolutionary and phylogeny studies due to its maternal inheritance, simple genomic organization, relative abundance in tissues and absence of recombination.22 In most metazoans the mitogenome is highly conserved and comprises of 13 protein coding genes, 22 transfer RNAs and two ribosomal RNAs in addition to a large single non-coding region important for replication and transcription.23 At present, mitogenomes of only five species from the genus Haemagogus are available.24,25 Considering the significance of mitogenomes when conducting phylogenetic and taxonomic studies, we describe for the first time the characterization of the mitogenome of Hg. equinus using a genome skimming approach and the phylogenetic comparison with other Haemagogus taxa.\n\n\nMethods\n\nThe Hg. equinus samples (n = 2) sequenced were collected in Mona, Jamaica (18.0061364°N, -76.7515125°W) in May and December 2023 in an area characterized by the predominant growth of banana plants and adjacent to a forested area. Briefly, BG sentinel traps (BioGents, Regensburg, Germany) baited with two pounds of dry ice (without lure) were placed overnight from 1400 to 1000 hr and collected mosquito specimens were sorted and morphologically identified using taxonomic keys.26 Single specimens were stored in tubes containing silica and shipped to the Johns Hopkins Bloomberg School of Public Health for molecular analysis as described in.27\n\nThe mitogenome was assembled as described for African anophelines using NOVOPlasty (RRID:SCR_017335) version 4.3.127,28 and automatic annotations performed with MITOS on the galaxy platform under the invertebrate genetic code.29,30 Geneious Prime (RRID:SCR_010519) version 2023.2.1 (Biomatters, Auckland, Australia) was utilized for manual adjustments of start and stop codons to match reference Haemagogus mitogenomes in the GenBank repository. Figure 1 illustrates a representative map of the mitochondrial sequences and annotations submitted to GenBank.\n\nNine mitogenomes (two sequenced from this study and seven available from GenBank) were used for analysis. The 13 protein coding genes of these mitogenomes were extracted with Geneious Prime and aligned by MAFFT version 7.490 into a single matrix. jModelTest version 2.1.10 identified the best fit substitution model and phylogenetic analyses were performed using maximum likelihood in MEGA (RRID_SCR_023017) version 11 with 1000 bootstrap replicates.\n\n\nResults\n\nSequencing of the two Hg. equinus specimens resulted in a mean of 26,728,491 million reads and of these approximately 69,261 reads were utilized for assembling each mitogenome. Two ribosomal RNAs, 22 transfer RNAs and 13 protein coding genes were detected in the two Hg. equinus mitochondrial genomes (GenBank accession numbers PQ_189398, PQ_189399). The cytochrome c oxidase I (COI) segment covering 1691-3221 bp was 99.59 % comparable to a COI sequence for Haemagogus spegazzinii Brèthes, 1912 (YP_010155459) retrieved from GenBank. Similar to the six Haemagogus mitochondrial genomes available in the GenBank database, the representative mitogenome from our sequencing efforts (PQ_189398) has an A+T proportion of 80.7% and length of 16,471 bp. The Maximum Likelihood phylogenetic tree places it in the Albomaculatus section, separated from other Haemagogus mitogenomes ( Figure 2).\n\nThese findings provide the basis for the development of more accurate molecular tools which can be used for identification of Hg. equinus mosquitoes. Furthermore, by populating molecular repositories with genomic data from Jamaican mosquitoes, their phylogenetic and evolutionary status will be better understood.\n\nEthical approval and consent were not required.", "appendix": "Acknowledgments\n\nSimmoy A.A. Noble is a Global Infectious Diseases Scholar and received mentored research training in the development of this manuscript.\n\n\nData availability\n\nGenBank: Haemagogus equinus mitochondrion, complete genome. Accession numbers PQ189398, PQ189399; https://identifiers.org/ncbi/insdc:PQ189398, https://identifiers.org/ncbi/insdc:PQ189399.31\n\nBio Project. Complete mitochondrial genome of Haemagogus equinus, Accession number PRJNA1172362; https://www.ncbi.nlm.nih.gov/bioproject/PRJNA1172362.32\n\nSRA. Illumina seq of Haemagogus equinus. Accession numbers SRR31002245, SRR31002246; https://www.ncbi.nlm.nih.gov/sra/?term=PRJNA1172362.33\n\nBiosample: Haemagogus equinus isolates HGJ1, HGJ3. SAMN44269491, SAMN44269492; https://trace.ncbi.nlm.nih.gov/Traces/study/?acc=PRJNA1172362.34\n\n\nReferences\n\nArnell JH: Mosquito studies (Diptera, Culicidae). XXXII. A revision of the genus Haemagogus. Contributions of the American Entomological Institute. 1973; 10: ii + 1–ii + 174.\n\nPan American Health Organization: Epidemiological Alert: Mayaro Fever. (accessed on 15 August). Reference Source\n\nWaddell MB, Taylor RM: Studies on Cyclic Passage of Yellow Fever Virus in South American Mammals and Mosquitoes: Marmosets (Callithrix aurita) and Cebus Monkeys (Cebus versutus) in Combination with Aedes aegypti and Haemagogus equinus. The American Journal of Tropical Medicine. 1945; s1-25: 225–230. Publisher Full Text\n\nWaddell MB, Taylor RM: Studies on the Cyclic Passage of Yellow Fever Virus in South American Mammals and Mosquitoes: III. Further Observations on Haemagogus equinus as a Vector of the Virus. The American Journal of Tropical Medicine. 1947; s1-27: 471–476. Publisher Full Text\n\nWaddell MB: Comparative Efficacy of Certain South American Aëdes and Haemagogus Mosquitoes as Laboratory Vectors of Yellow Fever. The American. J. Trop. Med. 1949; 29: 567–575. PubMed Abstract \| Publisher Full Text\n\nDutary BE, Leduc JW: Transovarial transmission of yellow fever virus by a sylvatic vector, Haemagogus equinus. Trans. R. Soc. Trop. Med. Hyg. 1981; 75: 128. PubMed Abstract \| Publisher Full Text\n\nDe Rodaniche E, Galindo P: Isolation of yellow fever virus from Haemagogus mesodentatus, H. equinus and Sabethes chloropterus captured in Guatemala in 1956. The American journal of tropical medicine and hygiene. 1957; 6: 232–237. PubMed Abstract \| Publisher Full Text\n\nDe Rodaniche E, Galindo P, Johnson CM: Isolation of yellow fever virus from Haemagogus lucifer, H. equinus, H. spegazzinii falco, Sabethes chloropterus and Anopheles neivai captured in Panama in the fall of 1956. The American journal of tropical medicine and hygiene. 1957; 6: 681–685. PubMed Abstract \| Publisher Full Text\n\nChadee DD, Hingwan JO, Persad RC, et al.: Seasonal abundance, biting cycle, parity and vector potential of the mosquito Haemagogus equinus in Trinidad. Med. Vet. Entomol. 1993; 7: 141–146. PubMed Abstract \| Publisher Full Text\n\nChadee DD: Rock hole breeding Haemagogus mosquitoes on Monos Island, Trinidad, West Indies. Mosq. News. 1983; 43: 236–237.\n\nChadee DD, Maitre A l, Connell NK: The collection of Haemagogus equinus Theobald breeding in household containers in Tobago WI. Mosq. News. 1981; 41: 568–569.\n\nParra C, Liria J: Haemagogus equinus Theobald 1903 (Diptera: Culicidae) en el Campus de la Universidad de Carabobo. Valencia. Venezuela. Salus. 2010; 14: 41–43.\n\nBlohm G, Elbadry MA, Mavian C, et al.: Mayaro as a Caribbean traveler: Evidence for multiple introductions and transmission of the virus into Haiti. Int. J. Infect. Dis. 2019; 87: 151–153. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLednicky J, De Rochars VMB, Elbadry M, et al.: Mayaro virus in child with acute febrile illness, Haiti, 2015. Emerg. Infect. Dis. 2016; 22: 2000–2002. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWhite SK, Mavian C, Elbadry MA, et al.: Detection and phylogenetic characterization of arbovirus dual-infections among persons during a chikungunya fever outbreak, Haiti 2014. PLoS Negl. Trop. Dis. 2018; 12: e0006505. Publisher Full Text\n\nChan-Chable RJ, Martínez-Arce A, Mis-Avila PC, et al.: DNA barcodes and evidence of cryptic diversity of anthropophagous mosquitoes in Quintana Roo, Mexico. Ecol. Evol. 2019; 9: 4692–4705. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGibson CM, Kao RH, Blevins KK, et al.: Integrative Taxonomy for Continental-Scale Terrestrial Insect Observations. PLoS One. 2012; 7: e37528. Publisher Full Text\n\nAdeniran AA, Hernández-Triana LM, Ortega-Morales AI, et al.: Identification of mosquitoes (Diptera: Culicidae) from Mexico State, Mexico using morphology and COI DNA barcoding. Acta Trop. 2021; 213: 105730. PubMed Abstract \| Publisher Full Text\n\nViveros-Santos V, Hernández-Triana LM, Ibáñez-Bernal S, et al.: Integrated Approaches for the Identification of Mosquitoes (Diptera: Culicidae) from the Volcanoes of Central America Physiographic Subprovince of the State of Chiapas, Mexico. Vector borne and zoonotic diseases (Larchmont, N.Y.). 2022; 22: 120–137. PubMed Abstract \| Publisher Full Text\n\nHernández-Triana LM, Garza-Hernández JA, Ortega Morales AI, et al.: An Integrated Molecular Approach to Untangling Host–Vector–Pathogen Interactions in Mosquitoes (Diptera: Culicidae) From Sylvan Communities in Mexico. Front. Vet. Sci. 2021; 7. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nReidenbach KR, Cook S, Bertone MA, et al.: Phylogenetic analysis and temporal diversification of mosquitoes (Diptera: Culicidae) based on nuclear genes and morphology. BMC Evol. Biol. 2009; 9: 298. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCameron SL: Insect mitochondrial genomics: implications for evolution and phylogeny. Annu. Rev. Entomol. 2014; 59: 95–117. Publisher Full Text\n\nBoore JL: Animal mitochondrial genomes. Nucleic Acids Res. 1999; 27: 1767–1780. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nda Silva FS , Cruz ACR, de Almeida Medeiros DB , et al.: Mitochondrial genome sequencing and phylogeny of Haemagogus albomaculatus, Haemagogus leucocelaenus, Haemagogus spegazzinii, and Haemagogus tropicalis (Diptera: Culicidae). Sci. Rep. 2020; 10: 16948. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLemos PS, Monteiro HAO, Castro FC, et al.: Characterization of mitochondrial genome of Haemagogus janthinomys (Diptera: Culicidae). Mitochondrial DNA Part A. 2017; 28: 50–51. PubMed Abstract \| Publisher Full Text\n\nBelkin JN, Heinemann SJ, Page WA: The Culicidae of Jamaica (Mosquito Studies. XXI). Contrib. Am. Entomol. Inst. 1970; 6: 1–458.\n\nAli R, Gebhardt M, Lupiya J, et al.: The first complete mitochondrional genome of Anopheles gibbinsi using a skimming sequencing approach. [version 1; peer review: 3 approved]. F1000Res. 2024; 13: 553. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDierckxsens N, Mardulyn P, Smits G: NOVOPlasty: de novo assembly of organelle genomes from whole genome data. Nucleic Acids Res. 2017; 45: e18. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBernt M, Donath A, Jühling F, et al.: MITOS: improved de novo metazoan mitochondrial genome annotation. Mol. Phylogenet. Evol. 2013; 69: 313–319. PubMed Abstract \| Publisher Full Text\n\nAfgan E, Baker D, Batut B, et al.: The Galaxy platform for accessible, reproducible and collaborative biomedical analyses: 2018 update. Nucleic Acids Res. 2018; 46: W537–W544. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNoble SAA, Pierre SA, Sandiford SL, et al.: Haemagogus equinus mitochondrion, complete genome. [Dataset]. GenBank. 2024.\n\nJohns Hopkins Bloomberg School of Public Health: Complete mitochondrial genome of Haemagogus equinus. [Dataset]. Bioproject. 2024.\n\nJohns Hopkins Bloomberg School of Public Health: llumina seq of Haemagogus equinus. [Dataset]. SRA. 2024.\n\nJohns Hopkins Bloomberg School of Public Health: Invertebrate sample of Haemagogus equinus. [Dataset]. Biosample. 2024." }	[ { "id": "349846", "date": "08 Jan 2025", "name": "Din Syafruddin", "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\nGeneral comment:\nThe manuscript reports the complete mitochondrial genome sequence of Haemagogus equinus from Jamaica. The genome is circular and includes 13 protein-coding genes, 2 ribosomal RNA genes and 22 tRNA genes. As the member of tribe Aedini and vector of Yellow fever, it might be informative to include mitochondrial genome of Aedes aegypti in the phylogenetic tree.\nSpecific comments:\nThe map should clearly indicate the position of the 13 protein coding genes, rRNA genes and 22 tRNA genes in the light and heavy strands of mtDNA (Fig. 1).\n\nThe authors should include “discussion” section in the manuscript to describe the characteristic of the genes encode by mitochondrial gnome of the Haemogogus equinus in comparison to cosmopolitan species of the tribe Aedini, Aedes aegypti.\n\nAre the rationale for sequencing the genome and the species significance clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of the sequencing and extraction, software used, and materials provided to allow replication by others? Yes\n\nAre the datasets clearly presented in a usable and accessible format, and the assembly and annotation available in an appropriate subject-specific repository? Yes", "responses": [] }, { "id": "349839", "date": "29 Jan 2025", "name": "Craig Wilding", "expertise": [ "Reviewer Expertise Evolutionary genetics" ], "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\nSandiford et al sequence full mitochondrial genomes from genome skimming data from two Haemagogus equinus specimens collected from Jamaica. They assemble these into a full mtDNA genome containing the expected gene content. Their introduction is well-written and provides much background information and adequate justification for this study. However, I would argue that the statement in the introduction that “Few studies have employed genomic sequencing to investigate the molecular taxonomy and phylogenetic relationships of Hg. equinus mosquitoes” suggests that this paper is going to provide a genome. Whilst it could be argued that the mtDNA is a genome it is somewhat misleading!\nIn the Methods there is no information on the sequencing undertaken. Whilst the authors say “molecular analysis as described in [27]” this is insufficient – at the very least there should be a brief description of what was done.\nThe statement “Nine mitogenomes (two sequenced from this study and seven available from GenBank) were used for analysis” should read “Nine mitogenomes (two sequenced from this study and seven available from GenBank) were used for phylogenetic analysis”\nIn the Results section, Figure 1 has a very poor resolution. This needs altering. Figure 2 has the albomaculatus and tropical groups highlighted, but which group does Hg. leucocelaenus belong? Is this Splendens? Or if not, are there no mitochondrial genomes from this group? I am not particularly clear on the point of Figure 2.\nIt would make more sense to produce a tree on a wider-scale (including e.g. representative Aedes and Sabethes or, have a tree to show the reader the position of Hg. equinus within the wider Aedini and then have this zoomed in view of Haemagogus, but for those readers unfamiliar with this genus the Figure does not really say anything of use and the text merely says “The Maximum Likelihood phylogenetic tree places it in the Albomaculatus section”. Was this not known before these mitochondrial sequences were generated?\nThe authors make no comment on whether the gene order seen in this species is as expected given those of closely related species. They should do so.\nThis study sequenced two specimens with accession numbers PQ_189398 and PQ_189399. How different were they? Some indication of overall genetic distance would be useful.\n\nAre the rationale for sequencing the genome and the species significance clearly described? Yes\n\nAre the protocols appropriate and is the work technically sound? Yes\n\nAre sufficient details of the sequencing and extraction, software used, and materials provided to allow replication by others? No\n\nAre the datasets clearly presented in a usable and accessible format, and the assembly and annotation available in an appropriate subject-specific repository? Yes", "responses": [] } ]	1	https://f1000research.com/articles/13-1504
https://f1000research.com/articles/13-1503/v1	09 Dec 24	{ "type": "Research Article", "title": "Life Cycle Assessment of Microalgae-Based Products for Carbon Dioxide Utilization in Thailand: Biofertilizer, Fish Feed, and Biodiesel", "authors": [ "Adeel Rafiq", "Cameron Morris", "Abigail Schudel", "Shabbir Gheewala", "Adeel Rafiq", "Cameron Morris", "Abigail Schudel" ], "abstract": "Background Microalgae-based products offer a sustainable solution for food, fuel, and agricultural inputs, presenting environmental benefits and economic opportunities. A comprehensive assessment is needed to understand their potential in supporting sustainability goals, considering the complex interplay between production methods, energy sources, and environmental impacts.\n\nMethods This study evaluated the environmental impacts of three microalgae-derived products – biodiesel, fish feed, and biofertilizer – through a comprehensive life cycle assessment. Nine scenarios were explored comparing three electricity profiles (current Thai mix, 50% renewable/50% current mix hybrid, 100% renewable) across the three products. The assessment evaluated environmental impacts and potential economic benefits of transitioning to these microalgae-based alternatives.\n\nResults and discussion All products demonstrated potential for significant environmental benefits under increased renewable energy scenarios. Fish feed consistently exhibited the lowest environmental impacts across all categories examined, showing substantial improvements with increased renewable energy use. With an annual demand of 0.4 million tonnes, fish feed could generate USD 560 million in revenue and reduce CO2 emissions by 1.1 million tonnes. Fulfilling the projected biodiesel demand of 4,015 million liters per year through microalgae production could yield approximately USD 3.5 billion in revenue and reduce CO2 emissions by 30 million tonnes compared to conventional fossil-based diesel. Additionally, algal biofertilizer production could meet a 5 million tonnes annual demand, offering USD 2 billion in revenue while reducing CO2 emissions by 6 million tonnes yearly. Collectively, these products could offset 37 million tonnes of CO2, representing about 14% of Thailand’s total CO2 emissions, contributing significantly to the country’s Nationally Determined Contribution (NDC) target of 20-30% greenhouse gas emissions reduction.\n\nConclusion Transitioning to microalgae-based products could transform the aquaculture, energy, and agricultural sectors, potentially supporting the national climate change mitigation goals, if implemented.", "keywords": [ "carbon capture", "carbon utilization", "photobioreactor", "Chlorella vulgaris", "aquafeed", "life cycle assessment", "microalgae" ], "content": "1. Introduction\n\nMicroalgae has emerged as a promising candidate for carbon utilization amid growing concerns about climate change and the need to reduce carbon dioxide (CO2) emissions (Rosental et al., 2020). Rising levels of greenhouse gases, especially CO2, have posed major challenges to global sustainability and climate stability in recent decades, largely due to continued reliance on fossil fuels for energy and industry. According to emissions trends, CO2 and other greenhouse gases will result in exceeding the 1.5 °C warming goal set by the Paris Agreement (Adun et al., 2024). As a result, carbon dioxide removal will be necessary to meet global and national climate goals. Carbon capture technology has been developed to abate carbon emissions, however the path to utilization still remains unclear. Microalgae may offer a sustainable approach to help address this issue (Shuba and Kifle, 2018; Sun et al., 2018).\n\nAdditionally, microalgae products could present a solution for the growing global population’s increasing demand for food, energy, and agricultural products (Braun and Colla, 2023). Microalgae has fast growth rates, high photosynthetic efficiency, and can sequester CO2 much more quickly than traditional crops (Pignolet et al., 2013; Yahya et al., 2020). Their high surface area-to-volume ratio also facilitates efficient gas exchange with the surrounding environment, enhancing CO2 uptake (Bhola et al., 2014). Furthermore, microalgae thrive in CO2-rich conditions, making it well-suited to capture emissions directly from industrial sources like power plants and cement factories (Yahya et al., 2020). To harness these advantageous properties of microalgae for effective carbon utilization and product development, different cultivation methods have been developed and refined over the years. Microalgae cultivation methods primarily include open raceway ponds and closed photobioreactors. Open raceway ponds are shallow, circulating channels that offer low capital and operational costs but are susceptible to contamination and weather fluctuations (Jorquera et al., 2010; Richardson et al., 2012; Skjånes et al., 2016). Closed photobioreactors, such as tubular, flat panel, or column systems, provide better control over growth conditions, higher productivity, and reduced contamination risk, albeit at higher costs (Chisti, 2007; Jorquera et al., 2010; Richardson et al., 2012).\n\nRecent years have witnessed a surge of interest in products derived from microalgae, leading to numerous life cycle assessment (LCA) studies that evaluate their environmental sustainability and impacts. A number of studies have examined the environmental profile of algae-derived biofuels. Clarens et al. (2011) found that algae systems produce significantly more vehicle kilometers traveled per hectare than terrestrial crops, but their environmental performance per kilometer is mixed. Their study emphasized the importance of nutrient procurement in algae cultivation and noted that using recycled CO2 from flue gas significantly improved the energy return on investment compared to virgin commercial CO2 (Clarens et al., 2011). However, they noted that flue gas capture for algae cultivation is not yet viable at industrial scale. Fenton et al. (2014) conducted an LCA assessing algal biofuels’ potential to replace petroleum fuels in Bangkok, Thailand. The study found that algal biofuels could replace less than 1% of Bangkok’s petroleum fuel use due to limitations in nutrient availability from municipal wastewater. Additionally, algal biofuels had higher environmental impacts than conventional fuels across all categories assessed (Fenton et al., 2014).\n\nSimilarly, the LCA study by Bradley et al. (2023) revealed that microalgae-derived biodiesel, when accounting for infrastructure, currently has higher greenhouse gas emissions than petroleum diesel. To match the environmental performance of petroleum diesel, significant improvements in microalgae productivity are necessary. The study underscored the importance of optimizing energy sources and feedstocks, such as yeast, to enhance the sustainability of microalgae-based biofuels (Bradley et al., 2023). Schade and Meier (2020) conducted an LCA to evaluate the environmental impacts of microalgae cultivation for food. They found that microalgae cultivation in a cold-weather climate can be sustainable, and a longer cultivation season is preferable even with reduced productivity. Similarly, Smetana et al. (2017) investigated microalgae as a potential solution to protein shortages in Europe. Their LCA revealed that current microalgae cultivation methods have a higher environmental impact than traditional protein sources. However, they identified promising alternatives, such as using heterotrophic cultivation and food waste as feedstock, which could significantly reduce the environmental impact and make microalgae a sustainable protein source (Smetana et al., 2017). High moisture extruded products made from heterotrophically cultivated Chlorella vulgaris showed lower environmental impacts than pork and beef, suggesting potential for certain microalgae products in the food sector (Smetana et al., 2017).\n\nIn agriculture, de Siqueira Castro et al. (2020) compared microalgae-based biofertilizer to conventional triple superphosphate fertilizer. The study found that the biofertilizer had higher environmental impacts across all categories, particularly in climate change and terrestrial ecotoxicity. The main contributors to these impacts were electricity use for cultivation, harvesting, and drying of the microalgae biomass. An optimized scenario using photovoltaic energy, gravitational biomass settling, and natural drying reduced these impacts significantly (de Siqueira Castro et al., 2020).\n\nWhile a substantial body of research exists on the environmental impacts of microalgae-based products, there is a critical need to compare the potential of carbon capture and utilization (CCU) across different applications. This study aims to address this gap by conducting a comprehensive life cycle assessment to compare the environmental performance of biofertilizers, animal feed, and biodiesel from microalgae produced from captured CO2. These three products were selected due to their significant market potential and environmental implications. In aquaculture, microalgae species like Isochrysis, Pavlova, and Chlorella vulgaris offer superior nutrition and health benefits compared to traditional feed sources (Dineshbabu et al., 2019). With global fish production projected to reach 209 million tonnes by 2030, microalgae-based feeds are poised to play a crucial role in meeting this growing demand sustainably (Dineshbabu et al., 2019). In the energy sector, global biodiesel consumption has surged from 2.2 million tonnes in 2004 to 65.86 million tonnes in 2023, with expectations to exceed 75 million tonnes by 2030 (Statista, 2024). Thailand, in particular, has set an ambitious target of 11 million liters per day of biodiesel production by 2037, up from 4.28 million liters in 2018 (Intamano et al., 2024). The biofertilizer market is also experiencing significant growth, with projections indicating an increase from USD 1106 million in 2016 to USD 3124 million by 2024, at a CAGR of 14.2% (Miranda et al., 2024).\n\nIn light of these promising market developments, the research will delve into the environmental impacts of two microalgae strains across these distinct applications. The study has two primary objectives. First, this study rigorously evaluates the environmental impacts of three microalgae products (biodiesel, fish feed, and biofertilizer) under three distinct electricity scenarios. These scenarios include Thailand’s current electricity mix as of 2020, a hybrid scenario combining on-site renewable energy production with grid electricity, and a fully renewable energy scenario. The potential CO2 emission reductions and their associated benefits are evaluated for the top-performing scenarios. This analysis incorporates a comprehensive assessment of emissions from diverse sources, including combustion of fossil fuels, land-use changes, and potential emissions mitigated through industrial processes such as carbon capture at power plants. Quantitative analysis of CO2 utilization pathways will help identify the most eco-friendly and feasible CCU technologies for Thailand. The study’s results will contribute to Thailand’s low-carbon transition and inform policymakers and industry about the most promising CCU strategies.\n\n\n2. Methods\n\nThe environmental implications of transitioning from conventional to microalgae-derived products were assessed through a rigorous life cycle assessment framework, adhering to ISO 14040 and 14044 standards (ISO, 2006a; ISO, 2006b) for methodological consistency.\n\nThis study aims to determine which of three microalgae-based products has the least environmental impact. The intended application is to draw comparative conclusions about the environmental impacts of various microalgae products produced from carbon dioxide captured using amine-based technology from a nearby power plant. The impact of transportation is not considered due to the proximity of the capture source. The microalgae products examined in this study—biodiesel, fish feed, and biofertilizer—were selected based on their potential commercial viability and relevance to current market demands. Results are aimed at informing potential stakeholders, including policymakers and industry leaders, about the feasibility and environmental implications of microalgae technology as a use for captured carbon dioxide. The technologies employed in this study, such as carbon capture, cultivation, harvesting, and product conversion processes, are limited to the best available options known at this time. The functional unit is one million tons of utilized captured carbon dioxide. This is measured as the amount of carbon injected into the photobioreactor.\n\nHowever, as these technologies are likely to continue developing in the future, potential improvements are considered in this study, in addition to the current state of the art. Initially, the study uses Thailand’s 2020 energy mix as a baseline scenario. In Thailand’s 2020 energy mix (TH20), fossil fuels dominate, with natural gas providing 57% of electricity, coal 19%, and oil 5.8%. Renewable sources contribute modestly, with biofuels at 9.8%, solar PV 2.7%, and wind 1.6%, while hydro accounts for 3.7% and waste for 0.5% of the total generation. To extend the applicability of the results beyond Thailand and explore potential future energy landscapes, the analysis includes two additional energy scenarios: a balanced mix of 50% renewable energy and 50% grid energy (based on the 2020 mix), and a 100% renewable energy scenario. These scenarios were selected based on specific criteria: representation of current conditions (Thailand’s 2020 mix), exploration of a transitional state (50% renewable + 50% grid), and examination of a fully renewable future (100% renewable). Nine scenarios are created by combining each of the three microalgae products with these three energy scenarios. For clarity and brevity, abbreviations are used throughout the study: BF, BD, and FF represent biofertilizer, biodiesel, and fish feed respectively, while the energy scenarios are abbreviated as follows: 100% for 100% Renewable energy, 50% for 50% Renewable energy + 50% Thailand’s electricity mix in 2020, and TH20 for Thailand’s electricity mix in 2020. This approach allows for a comprehensive assessment of how varying levels of renewable energy integration could impact the environmental performance of different microalgae-based carbon capture and utilization technologies. This study has expanded its scope to include a cradle-to-grave analysis, covering carbon capture, microalgae cultivation, production, and use of the three products ( Figure 1). The emissions source was excluded from the system boundary since this study focuses on evaluating and comparing CO2-based product production pathways, specifically assessing their suitability for carbon utilization in terms of environmental impact. Substitution was used for the displacement of conventional fossil-based diesel, conventional fish feed, and conventional mineral fertilizer. This allows for comparison based only on the function of utilizing captured carbon and not the individual functions of the three products. The data requirements for this study are the quantitative measures of emissions and resource use.\n\nAll data inputs were collected from secondary sources, including life cycle inventory databases and published literature (Table S1) (underlying data). Agri-footprint, Industry Data 2.0, and ecoinvent 3 databases served as the sources for background input data. These inputs include tubular photobioreactor construction, cleaning materials, and system inputs such as nitrogen and phosphorus fertilizers. Carbon capture data was obtained from (Rafiq et al., 2024a; Win et al., 2023), while cultivation and harvesting data were sourced from (Bradley et al., 2023; Grierson et al., 2013; Schade and Meier, 2020). Data for subsequent microalgae-based products (biodiesel, fish feed, and biofertilizer) were collected from (Bradley et al., 2023; Fenton et al., 2014; Grierson et al., 2013; Li et al., 2022; Smetana et al., 2017) and then adapted to the functional unit. Data for the substitution of traditional production of fish feed was sourced from literature as well (Pelletier and Tyedmers, 2010; Yacout et al., 2016).\n\n2.2.1 Biodiesel production\n\nNannochloropsis sp. was selected for this study due to its advantageous characteristics for biodiesel production, particularly its high lipid and protein contents. The nutrient composition of this microalgae species varies significantly across literature, influenced by factors such as nutrient availability and environmental conditions. For this study, a protein content of 30% of dry matter was adopted, aligning with the consensus among several research findings (Molino et al., 2018; Paes et al., 2016). The lipid content of Nannochloropsis sp. exhibits considerable variability, ranging from 5% to 44% of dry matter under optimal growth conditions. Taking into account this variability, an average lipid content of 21% of dry matter was used for this study. Additionally, the eicosapentaenoic acid content, an important omega-3 fatty acid, averages around 4.2% of dry matter (Molino et al., 2018; Schade and Meier, 2020).\n\nIn this study, the microalgae are cultivated in borosilicate glass tubular photobioreactors, chosen for their high surface area-to-volume ratio and efficient light utilization [37]. Following cultivation, the biomass undergoes centrifugation for harvesting, followed by lipid extraction ( Figure 2). The extracted lipids are then converted to biodiesel through transesterification, yielding glycerol as a co-product (Fenton et al., 2014; Schade and Meier, 2020). Substitution was employed for multiple products and co-products throughout the process. The primary output, biodiesel, substitutes conventional diesel fuel on an energy-equivalent basis (37.5 MJ/kg for biodiesel compared to 43 MJ/kg for conventional diesel). Glycerol, a co-product generated at approximately 10% of the biodiesel mass, substitutes synthetic glycerol on a mass-equivalent basis (Rodrigues et al., 2017). The residual biomass remaining after lipid extraction was utilized for biomethane production through anaerobic digestion. Methane yield from Nannochloropsis sp. was approximately 0.35 L per gram of volatile solids, with volatile solids comprising 79% of the total solids in the raw biomass (Bohutskyi et al., 2014; Torres et al., 2023). The produced biomethane substitutes natural gas, providing a more sustainable alternative to this fossil fuel. The digestate left after biomethane production, rich in nutrients such as nitrogen, phosphorus, and potassium, was used as a biofertilizer. This nutrient-rich digestate further contributed an energy offset of 186.37 MJ/m3 when substituting conventional fertilizers (Fenton et al., 2014).\n\n2.2.2 Fish feed and biofertilizer production\n\nFor production of both fish feed and biofertilizer, Chlorella vulgaris was assumed as the algal strain. In the case of biofertilizer, chlorella, spirulina, and other algae species have shown benefits to crop growth including increased root and shoot length (El Arroussi et al., 2018). Chlorella vulgaris has also been shown to increase organic matter in soil (Alvarenga et al., 2023). Chlorella vulgaris was selected as it was the strain used to support evidence that microalgae could replace traditional mineral fertilizer (Schreiber et al., 2018). In the case of fish feed cultivation, Chlorella vulgaris was selected as it has a suitable composition of proteins, carbohydrates, and lipids for animal feed. It was found to have less environmental impacts than A. platensis (Smetana et al., 2017). Chlorella vulgaris and A. platensis are the most widely recognized strains when studying food production (Smetana et al., 2017). Construction and operation of a tubular photobioreactor would not be affected by the algae strain being grown. Therefore, the process had identical inputs to that of the algae strain selected for biodiesel. The same approach was applied for harvesting and centrifugation. Centrifugation could be employed to separate the dry biomass from the water regardless of strain because it does not rely on specific strain properties.\n\nOnce cultivated and harvested, microalgae can be fractionated into a protein powder that can be used for a variety of animal feeds (Smetana et al., 2017). In this study, fish feed was chosen as the specific feed produced as microalgae are well suited for it. Microalgae has appropriate proportions of proteins, lipids, and carbohydrates for fish meal and already contains essential amino acids required in fish feed, eliminating the need to add them as a secondary step. Microalgae-derived fish feed can be used as a refrigerated paste or as a dry product created by either pelletizing or extruding (Nagappan et al., 2021). A dry product is in higher demand than refrigerated paste, and the extruder is the most common method for this process (Nagappan et al., 2021); therefore, this method was chosen for this study.\n\nThe fractionation process for Chlorella requires 0.7 kWh/kg of energy, yielding 0.85 kg of protein-rich powder and 0.15 kg of oil (Smetana et al., 2017). The extracted oil substitutes conventional vegetable oil on a mass basis. This substitution accounts for the environmental impacts avoided by not producing an equivalent amount of conventional vegetable oil (soybean oil). The protein powder, containing approximately 45−55% protein, is then mixed with water and subjected to an extrusion process to produce fish feed pellets (El-Naggar et al., 2020; Smetana et al., 2017). The extrusion process requires an additional 0.13 kWh of electricity per kilogram of feed material processed, according to scaled data from industrial feed production (Li et al., 2022). The resulting microalgae-based fish feed substitutes conventional fish food on a mass basis (Yacout et al., 2016), leveraging its high protein content and balanced nutrient profile to meet the dietary needs of farmed fish.\n\nAfter centrifugation, microalgae are dehydrated for direct application as a biofertilizer. In order to dry the microalgae, both spray drying and lyophilization are available methods (González et al., 2020). For this study, spray drying was assumed as the drying method, and energy inputs were assumed as 4 GJ/tonne resulting in an 80 percent yield (Grierson et al., 2013). It was also assumed that microalgae are able to directly replace mineral fertilizer in the form of 15:15:15 NPK (nitrogen, phosphorus, potassium). Studies have shown that microalgae in a ratio of two percent algae to 98 percent soil met the requirements for use as a solid bio-based fertilizer. The microalgae applied as fertilizer demonstrated significant plant growth compared to ammonium sulfate when applied to both ryegrass and barley (González et al., 2020). Additionally, it has been demonstrated that in nutrient-deficient soils, algae resulted in comparable plant growth to plants treated with mineral fertilizer (Schreiber et al., 2018).\n\nThe fertilizers were compared based on the amount of nitrogen applied to the soil to credit traditional fertilizer production. They cannot be compared based on volume as microalgae-based fertilizer has a smaller amount of plant nutrients per kilogram. Inputs into microalgae fertilizer production indicated 41 kilograms of nitrogen per tonne of CO2 utilized. The selected mineral fertilizer (NPK 15:15:15) is fifteen percent nitrogen. Therefore, to be equivalent to the 555 kg of dry biomass resulting from 1 tonne of carbon dioxide, the production of 273 kg of mineral fertilizer is displaced.\n\nThe study utilized SimaPro 9.5, a specialized LCA software (PRé, 2023), along with the ReCiPe 2016 LCIA method (Huijbregts et al. 2016), to analyze the environmental performance of the three products. The analysis focused on five key impact categories, including a combined midpoint category for global warming (encompassing impacts on both human health and terrestrial ecosystems), along with three endpoint categories: human health (measured in disability-adjusted life years, DALY), ecosystem quality (represented as the Potentially Disappeared Fraction of species per area-year, species.yr), and resource scarcity (expressed in USD2013). These midpoint impact categories were chosen for their relevance in assessing the effectiveness of CCU technologies, which aim to capture and utilize CO2, thereby potentially reducing global warming impacts. Including endpoint impact categories was essential for gaining a comprehensive understanding of the final environmental damage translating specific environmental pressures into broader areas of protection (Huijbregts et al. 2016).\n\nTo assess the overall advantage of reducing carbon footprint in the optimal scenario, this study covers CO2 emissions from various sources, including fossil fuel use and land-use changes, as well as emissions prevented in the production processes of the best-case scenarios. Fossil-based emissions are thoroughly assessed, covering the entire process from carbon capture to the production of algae-based products. This evaluation also accounts for the avoided emissions that result from replacing conventional fossil-based products with CO2-utilizing alternatives. The approach employs a comparative evaluation of life cycle CO2 emissions between traditional method and alternative CO2-based pathways as shown in Figure 3. This comparison quantifies the net climate effect, highlighting the reduction in total CO2 emissions achieved through the alternative production pathway.\n\n\n3. Results and discussion\n\nThe life cycle assessment of microalgae-derived biodiesel, fish feed, and biofertilizer highlights their substantial potential to support sustainable development and contribute to climate change mitigation. This study evaluated nine scenarios, each considering different electricity profiles: current Thai grid mix, 50% renewable energy, and 100% renewable energy. The scenarios were analyzed using the ReCiPe 2016 method across multiple environmental impact categories. The findings indicate that the environmental performance of each product is significantly influenced by the energy sources used during production. The results indicate that transitioning from a fossil-fuel-dominated grid to 100% renewable electricity leads to substantial reductions in environmental impacts across all microalgae-based products ( Figure 4). Among the three products, fish feed produced with 100% renewable energy achieves the most significant reductions in global warming impacts, with −1095 DALY (human health) and −3 species.yr (terrestrial ecosystems) per million tonne CO2 utilization (functional unit), demonstrating its strong potential as a low-carbon product. Similarly, biodiesel under fully renewable conditions achieves notable reductions, with −556 DALY (human health) and −2 species.yr (terrestrial ecosystems), underscoring its role as a cleaner alternative to conventional fuels. Biofertilizer, while also benefiting from renewable energy integration, shows lower reductions, recording −503 DALY and −1.5 species.yr, indicating its potential to reduce emissions, albeit to a lesser extent compared to fish feed and biodiesel.\n\nThe impact on human health and ecosystems further underscores the benefits of renewable energy adoption. Fish feed produced with 100% renewable energy achieves the highest reduction in human health impacts, with −1230 DALY, illustrating its capacity to minimize health-related risks linked to the production process. Biodiesel produced with 100% renewable energy results in 68 DALY, highlighting challenges in reducing health-related impacts compared to fish feed. In terms of ecosystem quality, fish feed with 100% renewable energy achieves a decrease of −20 species.yr, outperforming biodiesel and biofertilizer. However, despite achieving notable reductions in global warming and ecosystem impacts, biofertilizer under 100% renewable conditions records an increase of 118 DALY in human health impacts, suggesting that further process optimization is needed to enhance its overall environmental sustainability. The adoption of 100% renewable energy significantly reduces resource-related impacts across all products. Biodiesel shows substantial progress, with a reduction of 26 million USD2013, underscoring its enhanced resource efficiency under a fully renewable energy mix. Following closely, fish feed demonstrates the lowest resource impact, achieving a reduction of 20 million USD2013, indicating significant improvements in resource efficiency. In contrast, biofertilizer under 100% renewable conditions registers an impact of 23 million USD2013, highlighting a need for better resource management strategies to achieve more sustainable outcomes.\n\nScenarios using 50% renewable energy mix exhibit moderate reductions compared to fully renewable scenarios. Fish feed and biodiesel produced under this energy mix show reduced impacts on global warming and human health, though these reductions are less pronounced than those observed under a 100% renewable energy mix. For example, fish feed produced with 50% renewable energy achieves a reduction of −302 DALY in global warming impacts on human health, while biodiesel records 186 DALY, reflecting improved but still comparatively significant impacts. This suggests that partial integration of renewable energy can enhance sustainability, but it does not provide the same benefits as a fully renewable approach. The highest environmental impacts are seen in scenarios that rely on Thailand’s current electricity grid mix, which is heavily dependent on fossil fuels. In these scenarios, fish feed production results in 491 DALY for global warming impacts on human health and 1.5 species.yr for terrestrial ecosystem impacts, indicating substantial environmental burdens. Similarly, biofertilizer and biodiesel produced under the current grid mix exhibit high impacts across global warming, human health, and resource scarcity categories. For instance, biofertilizer records 1182 DALY for human health impacts, while biodiesel registers 1273 DALY, making these the least sustainable scenarios due to their reliance on fossil-based electricity.\n\nCarbon capture contributes significantly to the environmental performance of microalgae-based products. For the functional unit of capturing one million tonnes of CO2 using amine-based technology, substantial benefits are observed across impact categories ( Table 1). Under the 2020 grid mix, the global warming impact on human health is reduced by 719 DALY, which further improves to 829 DALY when shifting to a 100% renewable energy scenario, reflecting a 15% enhancement. Similar improvements are noted in global warming impacts on terrestrial ecosystems, becoming more pronounced with increased renewable energy integration. The most significant improvement is seen in resource scarcity, with a 42% reduction in economic impacts achieved when using a fully renewable energy mix. Additionally, human health and ecosystem quality benefits range from 21% to 30%, indicating moderate but meaningful reductions across these categories. This analysis demonstrates that carbon capture, especially when combined with renewable energy sources, can play a pivotal role in enhancing the sustainability of microalgae-based products.\n\nThe contribution analysis of microalgae-based biodiesel production under worst-case scenario highlights that biodiesel production is the primary driver of environmental impacts across all categories. Substitution credits from conventional transport, natural gas, and glycerine significantly reduce the global warming impacts on human health and terrestrial ecosystems by approximately 21%, 3%, and 2%, respectively ( Figure 5). Additionally, glycerine credits reduce impacts in the ecosystem category by about 8%. In resource use, substitution credits from conventional transport and natural gas provide notable reductions of approximately 29% and 21%, respectively. While these credits offer significant reductions, they are insufficient to fully counter the high emissions from the current fossil-based electricity mix. This analysis underscores the need for cleaner energy solutions in biomass production processes to achieve more effective environmental improvements.\n\nEnvironmental impact analysis contrasting: (a) Midpoint impact evaluation for biodiesel production under suboptimal conditions utilizing the current grid electricity mix; (b) Associated endpoint impact indicators for the biodiesel worst-case scenario; (c) Midpoint impact categories for fish feed production under optimal conditions with 100% renewable energy integration; and (d) Corresponding endpoint impact metrics for the optimized fish feed scenario. This multi-panel analysis illustrates the spectrum of environmental implications across different production pathways and energy source configurations.\n\nConversely, fish feed production under best-case conditions with 100% renewable energy shows significant improvements in environmental performance. Biomass production achieves a 22% reduction in global warming impacts, further enhanced by substitution credits from conventional fish feed (36%) and vegetable oil replacement (41%). Although steam use causes a minor 7% increase, the overall impact on the environment still remains positive. While biomass production increases human health impacts by 25%, substantial credits from conventional feed (−57%) and vegetable oil (−43%) lead to net reductions. Ecosystem and resource impact categories exhibit similar positive trends, driven by effective substitutions. These findings align with previous studies that identify two major challenges in algae-based systems. The first is the intensive energy demand for processes like pumping, mixing, drying, and lipid extraction, especially when reliant on fossil fuels. The second challenge is nutrient management, particularly nitrogen and phosphorus usage, where inefficiencies can increase eutrophication and greenhouse gas emissions (Bradley et al., 2023; Fenton et al., 2014; Schade and Meier, 2020).\n\nDespite these challenges, the present study also found that Chlorella vulgaris has the potential to replace high-material and energy-intensive aquaculture feeds. Its viability as an alternative feed is supported by its favorable nutritional profile: 45% protein, 20% fat, 20% carbohydrates, 10% minerals and vitamins, and 5% fiber (w/w, dry basis) (El-Naggar et al., 2020). Modelling the substitution of conventional feed with Chlorella vulgaris reveals even greater environmental benefits than previously anticipated. Table 2 compares microalgae-based feed (FF_TH20) with conventional aquaculture feed production. The results show significant reductions in global warming potential for both human health and terrestrial ecosystems. The substitution substantially reduces human health and ecosystem impacts, while resource impacts decrease only marginally. These findings demonstrate Chlorella vulgaris’ potential as a sustainable aquaculture feed alternative, though resource efficiency needs optimization. To make the process even more efficient, implementing emerging approaches like bioflocculation, spontaneous flocculation, and attached growth systems show promise for more efficient and economical harvesting, potentially reducing chemical use and improving harvested biomass concentration (Bilad et al., 2012; de Assis et al., 2019; de Assis et al., 2017; Eliseus et al., 2017). However, the drying step remains a challenge, with various options like solar, drum, spray, and freeze-drying each presenting trade-offs between energy use, cost, and product quality (Milledge and Heaven, 2013). These strategies, combined with ongoing research into energy-efficient and environmentally friendly methods, aim to optimize both energy use and nutrient management in algal biofuel production, addressing the key challenges identified in life cycle assessments and paving the way for more sustainable algae-based systems in the future. Overall, the potential for algae-based systems to contribute to environmental sustainability is significant, as evidenced by various studies showing substantial reductions in environmental impacts when transitioning to renewable energy sources. This highlights the importance of not only increasing the share of renewable energy in the grid mix but also improving the energy efficiency of production processes themselves. With continued advancements and a focus on overcoming key challenges, algae-based CCU and bio-product production hold promise for playing a pivotal role in mitigating climate change and promoting a more sustainable and resilient future, a vision that is thoroughly supported by the results and conclusions of the current study.\n\nMicroalgae CCU provides biomass-containing compounds useful for pharmaceuticals, food, and biofuels (Zhao and Su, 2020). However, realizing their full potential requires overcoming key challenges identified through life cycle assessments. The optimization of microalgae CCU systems depends on two critical factors—the refinement of system processes for greater efficiency and the careful integration of renewable energy sources, both of which are crucial for improving environmental performance, as examined below.\n\n3.2.1 Impact of system efficiency and renewable energy\n\nFor carbon capture, Win et al. (2023) reported that capturing one tonne of CO2 requires approximately 378 kWh of energy, highlighting the significant energy input needed for carbon capture systems. This energy requirement is closely tied to the efficiency of the CO2 utilization process, which varies significantly across various studies, ranging from 40% to 93.7%, depending on specific conditions and technologies employed for microalgae cultivation (Ighalo et al., 2022). The results discussed in this study are based on a carbon utilization efficiency of 75%, which falls within the range reported in the literature. This relatively high efficiency suggests potentially lower energy requirements and environmental impacts compared to systems with lower efficiencies, as the energy needed per unit of CO2 captured would be reduced.\n\nFigure 6 illustrates the impact of CO2 utilization efficiency and renewable energy integration on CO2 emissions reduction in cultivation systems. The data reveal a clear trend: increasing CO2 utilization efficiency from 60% to 90% and renewable energy integration from 50% to 100% significantly improve CO2 reduction rates. Notably, with 100% renewable energy and 75% CO2 utilization efficiency, the reduction rate nearly matches that of the scenario with 90% CO2 utilization efficiency and 100% renewable energy. Importantly, in both the 75% and 100% renewable energy scenarios, all efficiency levels result in a net reduction of CO2, highlighting the critical role of renewable energy in achieving positive environmental outcomes. While the combination of high efficiency at 90% and full renewable energy at 100% maximizes CO2 reduction, the analysis indicates that balanced improvements in both parameters can yield substantial benefits without necessarily reaching the highest levels in each category. These relationships underscore the importance of optimizing both CO2 utilization efficiency and renewable energy integration within carbon capture systems.\n\nThe analysis demonstrates a broad range of CO2 reduction potential across the scenarios. At the lower end, with 60% efficiency and 50% renewable energy, the system results in net emissions of 343 kg CO2 per tonne processed. In contrast, the most favorable scenario with 90% efficiency and 100% renewable energy achieves a reduction of 965 kg CO2 per tonne CO2 utilized, representing a significant enhancement in carbon utilization performance. Moreover, the energy intensity of CO2 utilization could be further reduced by using flue gas directly instead of post-combustion CO2 capture. However, direct use of flue gas can affect algae growth and lipid production due to contaminants, with impacts varying by species. Napan et al. (2015) found that at baseline heavy metal concentrations representative of coal-fired flue gas, there was a 12% increase in biomass production and 61% increase in lipid yield for Scenedesmus obliquus compared to the control. However, at higher heavy metal concentrations, both growth and lipid production were inhibited (Hess et al., 2017).\n\nSimilarly, Hess et al. (2017) observed that for Nannochloropsis salina, the addition of 14 inorganic contaminants at concentrations representative of coal-fired flue gas resulted in a 67% reduction in biomass productivity and a 32% decrease in lipid content compared to the control (Napan et al., 2015). Despite these challenges, using captured flue gas CO2 is still more energy-efficient than virgin commercial CO2 (Clarens et al., 2011). Overall, the CO2 source significantly influences the energy requirements and sustainability of algae-based biofuel production. However, careful examination of flue gas composition and its suitability for specific algal strains is crucial for informed decision-making in algae-based biofuel production systems.\n\n3.2.2 Benefits of carbon capture and utilization\n\nIn developing countries, where fossil fuels often dominate the energy landscape, coal and natural gas power plants continue to be significant sources of CO2 emissions. However, these emissions can be repurposed through innovative carbon capture and utilization technologies, offering a pathway to more sustainable practices. One promising application is the cultivation of Chlorella vulgaris, a microalga species, to produce fish feed. Chlorella vulgaris has shown remarkable potential as an alternative to conventional, energy-intensive aquaculture feeds (Dineshbabu et al., 2019; Smetana et al., 2017). By utilizing CO2 emissions from power plants, the production of this algae-based fish feed can significantly reduce the carbon footprint of both energy and aquaculture sectors. The process requires approximately 0.9 million tonnes of CO2 annually to meet Thailand’s fish feed demand of 400,000 tonnes (Fachrudin, 2024), with each kilogram of algae-based feed produced potentially offsetting around 2.66 kilograms of CO2 emissions (Table S1) (underlying data). This estimate accounts for direct emissions from algae cultivation and processing, indirect emissions from land transformation, CO2 captured from power plants, and emissions avoided by replacing conventional feed production methods (Table S1) (underlying data). At a market price of USD 1,400 per tonne ( Indiamart, 2024), this industry could generate an annual revenue of USD 560 million. Simultaneously, it would avoid 1.1 million tonnes of CO2 emissions per year compared to conventional feed production. This initiative could lead to employment opportunities in carbon capture and utilization industries, as well as in algae cultivation and associated fields, providing an economic boost to the region. Beyond its economic potential, fish feed derived from Chlorella vulgaris offers several environmental benefits. It can help reduce pressure on wild fish stocks traditionally used for fishmeal production (Sheikhzadeh et al., 2024), contributing to marine ecosystem conservation. Additionally, as a protein-rich and nutrient-dense feed, it can improve the efficiency and sustainability of aquaculture operations.\n\nIn addition to fish feed production, microalgae cultivation holds great promise for carbon capture and utilization, with applications extending to the production of biodiesel and biofertilizer. The biodiesel production process requires approximately 37.5 million tonnes of CO2 annually to meet Thailand’s anticipated biodiesel demand of 4,015 million liters by 2037 as per government plans (Intamano et al., 2024). A market price of USD 1,000 per tonne (Data, 2024) would generate approximately USD 3.5 billion in revenue. Importantly, each kilogram of microalgae-based biodiesel avoids approximately 8.5 kg of CO2 (see Table S1) (undelrying data) emissions compared to conventional fossil-based diesel, potentially reducing emissions by 30 million tonnes of CO2 annually.\n\nMoreover, the conversion of algal biomass into biofertilizer presents a promising market opportunity, requiring about 9 million tonnes of CO2 annually to meet the biofertilizer demand of 5 million tonnes (Post, 2022). Assuming a market price of approximately USD 400 per tonne (Polene, 2024), this sector could generate substantial revenue of about USD 2 billion each year. Moreover, biofertilizer derived from microalgae offers significant environmental benefits, as each kilogram of this product can avoid approximately 1.2 kg of CO2 emissions. This translates to a total reduction of around 6 million tonnes of CO2 emissions annually, contributing to climate change mitigation efforts. In 2022, Thailand’s national CO2 emissions reached approximately 270 million tonnes (Ritchie and Roser, 2022), with the power generation sector accounting for about 90 million tonnes (Statista, 2023), primarily from coal and natural gas. As part of its commitment to combating climate change, Thailand’s Nationally Determined Contribution (NDC) targets a 20-30% reduction in greenhouse gas emissions by 2030 (Sugsaisakon and Kittipongvises, 2024). The CO2 utilization potential through these three microalgae-based products amounts to 47 million tonnes annually, representing 53% of the power sector’s emissions. These products could collectively offset 37 million tonnes of CO2 per year, contributing approximately 14% towards Thailand’s total emissions reduction. By leveraging microalgae technology, Thailand could make significant strides in reducing its carbon footprint while fostering sustainable industrial practices.\n\nHowever, scaling up the production of microalgae using power plant emissions faces several challenges. Substantial investments in infrastructure are required, including CO2 capture facilities at power plants, large-scale algae cultivation systems, and processing plants for the final products (Hepburn et al., 2019). While the cultivation of algae using industrial CO2 has been demonstrated on smaller scales, expanding to fully utilize emissions from large power plants necessitates further technological advancements and process optimizations. Market development presents another crucial consideration. Establishing and expanding markets for algae-based products may require efforts in consumer education, quality assurance, and potentially supportive policies to encourage adoption. A favorable regulatory environment, possibly including carbon pricing mechanisms or incentives for CCU technologies, would be instrumental in making these projects economically viable (Hepburn et al., 2019).\n\nDeveloping efficient cultivation methods and selecting appropriate locations for production facilities will be key to addressing these concerns. Beyond these implementation challenges, the production of value-added products such as fish feed, biodiesel, and biofertilizer from microalgae using power plant emissions represents a promising approach to emissions reduction and sustainable development. As countries work towards cleaner energy futures, these technologies could serve as valuable transitional solutions, mitigating emissions from existing power infrastructure while supporting food security, energy independence, and agricultural sustainability. The potential for significant CO2 reduction, coupled with economic benefits across multiple sectors, makes this an attractive option for countries seeking to balance development needs with climate change mitigation goals. By simultaneously addressing issues in aquaculture, transportation, and agriculture, microalgae-based CCU demonstrates a climate-friendly solution for multiple sectors in the economy. The versatility of these applications offers a pathway to create a circular economy, where waste CO2 becomes a valuable resource for multiple industries.\n\n\n4. Conclusions\n\nThis life cycle assessment of microalgae-based biodiesel, fish feed, and biofertilizer production in Thailand reveals the significant potential of these products to contribute to sustainable development and climate change mitigation. Among the evaluated products, microalgae-based fish feed demonstrated the most favorable environmental profile across multiple impact categories, particularly under increased renewable energy scenarios. The transition from the current energy mix to higher renewable energy penetration resulted in notable environmental improvements for all three products. Biodiesel production, while showing improvements under increased renewable scenarios, consistently exhibited higher environmental impacts compared to fish feed and biofertilizer. This suggests that prioritizing microalgae cultivation for high-value products like fish feed may offer greater environmental benefits in the near term.\n\nThe study highlighted the critical role of both CO2 utilization efficiency and renewable energy integration in determining the environmental performance of these systems. With carbon capture energy requirements of approximately 378 kWh per tonne of CO2, the efficiency of utilization becomes crucial. The analysis, based on a 75% carbon utilization efficiency, showed that increasing both CO2 utilization efficiency and renewable energy integration leads to significant improvements in CO2 reduction rates. Notably, scenarios with high renewable energy integration (75-100%) resulted in net CO2 reductions across all efficiency levels, underscoring the importance of renewable energy in achieving positive environmental outcomes.\n\nIn Thailand, transitioning to microalgae-based alternatives could have a transformative impact across various sectors. Meeting the current annual fish feed demand of 400,000 tonnes with microalgae products could generate approximately USD 560 million in revenue while avoiding 1.1 million tonnes of CO2 emissions compared to conventional feed. Fulfilling the anticipated annual biodiesel demand of 4,015 million liters through microalgae production could yield around USD 3.5 billion in revenue and reduce CO2 emissions by 30 million tonnes relative to conventional diesel. Additionally, the estimated annual demand of 5 million tonnes for biofertilizer derived from algal biomass could generate about USD 2 billion in revenue each year while potentially reducing CO2 emissions by 6 million tonnes annually. Together, these products could offset 37 million tonnes of CO2, representing approximately 14% of Thailand’s total emissions. If implemented, this approach could significantly contribute to advancing the country’s NDC goal of a 20-30% reduction in greenhouse gases. This study offers valuable insights for decision-makers, industry stakeholders, and researchers, supporting Thailand’s transition toward more sustainable practices in aquaculture, energy, and agriculture while accelerating its progress toward climate targets.\n\n\nEthical considerations\n\nEthical review and consent did not apply to this study.\n\n\nReporting guidelines\n\nReporting guidelines were not applicable to this study.\n\n\nAuthor roles\n\nRafiq A: Conceptualization, Data Curation, Formal Analysis, Investigation, Methodology, Writing – Original Draft Preparation, Writing – Review & Editing; Morris C: Conceptualization, Data Curation, Investigation, Methodology, Writing – Original Draft Preparation, Writing – Review & Editing; Schudel A: Conceptualization, Data Curation, Investigation, Methodology, Writing – Original Draft Preparation, Writing – Review & Editing; Gheewala SH: Conceptualization, Investigation, Methodology, Project Administration, Resources, Supervision, Validation, Writing – Review & Editing.", "appendix": "Data availability\n\nThe data underlying this article are available in Figshare: Life Cycle Assessment of microalgae-based products for carbon dioxide utilization in Thailand: biofertilizer, fish feed, and biodiesel. https://doi.org/10.6084/m9.figshare.27715455 (Rafiq et al., 2024b).\n\nThis project contains the following underlying data:\n\n• Table S1. xlsx (Life Cycle Assessment inventory data, calculations for avoided CO2 emissions, and data related to environmental impacts)\n\nFull credit is given to all co-authors who contributed to generating this dataset.\n\nData are available under the terms of the Creative Commons CC0 license.\n\n\nAcknowledgements\n\nThe authors would like to acknowledge King Mongkut’s University of Technology Thonburi (KMUTT), the Joint Graduate School of Energy and Environment (JGSEE), and the Center of Excellence on Energy Technology and Environment. 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Publisher Full Text" }	[ { "id": "352702", "date": "10 Jan 2025", "name": "Marcin Dębowski", "expertise": [ "Reviewer Expertise biofuels production", "algae cultivation", "co2 emission reduction", "biotechnology" ], "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 my opinion, the article raises an important issue of the use of micro-algae in the context of carbon sequestration (CCU) and its potential for the production of biofuels, fish feed and biofertilisers. The authors have used the life cycle analysis (LCA) method, which is suitable for assessing the environmental and economic impact of the proposed solutions. The article provides a fairly good basis for the analysis, but it lacks detailed references to existing literature and a more comprehensive comparison with the results of studies by other authors.\nIn my opinion, the authors have set themselves the ambitious task of analysing three energy scenarios (TH20, 50% renewable energy sources, 100% renewable energy sources) and their impact on the production of three different products. The methodology is based on the ISO 14040 and 14044 standards, which is the main advantage of the work. This is always difficult with technologies that are not widely used in practise and the data are usually estimates from different, mainly experimental work. I believe that the choice of energy scenarios described and analysed was not sufficiently justified in the context of technical and economic possibilities. The authors should demonstrate the validity of the original assumptions more precisely and convincingly.\nIn my opinion, the manuscript is not specify enough and partially ignores some important technological issues, such as the operating costs and efficiency of CCU integration on an industrial scale. It is also worth looking at the generalisability of these assumptions through the prism of climate policy in other regions of the world (Zieliński M, et al., 2023 [Ref 1])\nThe conclusions and key messages could be strengthened and supported by reference to the results of other studies, particularly in the area of energy efficiency, technological efficiency and the cost of CCU technology. The data on micro-algae production could be more detailed, e.g. in relation to the efficiency of different photo-bioreactor systems, which would make the work more universal and valid for different climatic conditions.\nThe paper does not address the economic aspects related to the implementation of CCU technology on an industrial scale, such as installation and operating costs, which can be completed based on the available literature. It would be useful to highlight the existing limitations in the widespread implementation of energy systems based on micro-algae biomass (Dębowski M, et al., 2022 [Ref 2])\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": [] } ]	1	https://f1000research.com/articles/13-1503
https://f1000research.com/articles/13-385/v1	25 Apr 24	{ "type": "Research Article", "title": "Molecular docking and dynamics studies to identify novel active compounds targeting potential breast cancer receptor proteins from an indigenous herb Euphorbia thymifolia Linn", "authors": [ "Vasavi Kumblekar", "Reshma Kumarchandra", "K Sreedhara Ranganath Pai", "ShamaPrasada K", "Suman Manandhar", "Rajeshwari Shastry", "Sharada Rai", "Vasavi Kumblekar", "K Sreedhara Ranganath Pai", "ShamaPrasada K", "Suman Manandhar", "Rajeshwari Shastry", "Sharada Rai" ], "abstract": "Background Breast cancer has become the most prevalent disease and its incidence has almost doubled in the Indian population. This increased burden demands new targeted therapies with novel compounds either synthetically produced or derived from indigenous plants, which could be a promising approach for the development of drugs. Euphorbia thymifolia L is a widely growing tropical herb that has been reported to have various ethnopharmacological properties, including anticancer properties. Therefore, the aim of the present study was to screen the phytoconstituents and identify the active compounds present in the methanolic extract of E. thymifolia (ME.ET) as ligands to inhibit potential protein targets implicated in breast cancer using an In-silico approach.\n\nMethods ME.ET was subjected to GC-MS analysis to screen the phytoconstituents, and the identified compounds were docked with protein targets such as extracellular signal-regulated kinases (ERK1), a serine/threonine kinase-1(AKT1), human epidermal growth factor 2 (HER2), estrogen receptor (ER), maternal embryonic leucine zipper kinase (MELK), polo-like kinase-1(PLK1), and protein tyrosine kinase (PTK6). Compounds with good docking scores were further subjected to dynamic studies to understand the protein ligand binding stability, ligand pathway calculation, and molecular mechanics energies combined with Poisson-Boltzmann (MM/PBSA) calculations using the Schrodinger suite.\n\nResults GC-MS analysis revealed the presence of 245 phytoconstituents, 219 of which were unique. When subjected to docking, these phytocompounds, namely 3,6,9,12-tetraoxatetradecane-1,14-diyl dibenzoate (TTDB) and succinic acid, 2-(dimethylamino) ethyl 4-isopropylphenyl ester (SADPE), showed good docking scores. Molecular dynamics studies showed a high affinity and low binding energy for TTDB with HER2, ERK1, and SADPE with ER.\n\nConclusions Hence, in this study, we identified two lead compounds in E.thymifolia linn. Further invitro and invivo anticancer studies can be performed to confirm these results and to understand the molecular mechanism by which they exhibit anticancer activity against breast cancer.", "keywords": [ "Euphorbia thymifolia", "phytoconstituents", "breast cancer", "anti-cancer activity", "molecular docking", "molecular dynamics", "breast cancer receptor proteins", "lead compounds." ], "content": "Introduction\n\nBreast cancer is the most common and serious disease among women worldwide, affecting their health and socioeconomic status. The incidence of breast cancer has significantly increased and almost doubled compared to the previous decade, with up to 25.8 per 100,000 women among the Indian population. The incidence of breast cancer is higher in younger women and tends to be more aggressive owing to the molecular complexity of the tumor microenvironment (Malvia et al. 2017). The treatment strategy for breast cancer depends on its subtype and is based on the presence or absence of estrogen (ER), progesterone (PR), and human epidermal growth factor 2 (HER 2) receptors (Waks and Winer 2019). Tumors with hormone receptor-positive subtypes receive endocrine therapy or targeted antibody treatment; however, triple-negative breast cancer is one of the most aggressive types of breast cancer, with a high recurrence rate, making it the most difficult to treat (Palma et al. 2015). Targeted therapy with anticancer agents in combination with immunotherapy is one of the treatment strategies for TNBC. However, there is a need for new and efficient compounds to target the proteins implicated in the tumorigenesis of TNBC (Chang-Qing et al. 2020).\n\nTargeting cell surface protein receptors can be an effective way to target malignant cells. Overexpression of the cell surface receptor HER2 proteins has been observed in patients diagnosed with invasive HER2 + breast cancer. It plays an important role in the growth and differentiation of epithelial cells (Yousuf et al. 2017). HER2/EGFR triggers one of the major signalling pathways which involves PI3 kinase, Ras-Raf-MAPK pathway involved in cell survival, proliferation, migration and angiogenesis (Balz et al. 2012). The estrogen receptor is another important target protein involved in the cross-talk between other signalling pathways and is implicated in the resistance to hormonal treatments in ER-positive breast cancer patients (Zhang et al. 2020). Protein tyrosine kinase (PTK6) is another important protein target associated with ER+ BC. Increased PTK6 expression is associated with poor overall survival (Ito et al. 2017). AKT1, a serine/threonine kinase, also known as protein kinase B and extracellular signal-regulated kinases (ERK1), plays a major role in cell proliferation, differentiation, migration, and anti-apoptotic effects and is overexpressed in TNBC (Martorana et al. 2021; Gagliardi et al. 2020; Guo et al. 2020). Maternal embryonic leucine zipper kinase (MELK), another pivotal member of the serine/threonine kinase family, is a cell cycle regulator whose overexpression in BC is associated with poor prognosis (Jiang and Zhang 2013). Therefore, these proteins were chosen as targets to screen and identify the most active compounds against both hormone receptor-positive BC and TNBC.\n\nEuphorbia thymifolia known as laghudhika or choti-dudhi, is a herb traditionally used to treat blood dysentery, inflammation, asthma, and stomach pain (Nadkarni and Nadkarni 2007). This herb has been reported to have antimicrobial, antidiabetic, antiarthritic, antiviral, and anti-inflammatory effects (Mali and Panchal, 2013). However, the anticancer activity of this compound has not yet been reported so far (Salehi et al. 2019). Thus, we have studied and reported the anticancer activity of the methanolic extract of E. thymifolia for the first time in our previous study.\n\nIdentification of the active compounds responsible for anticancer activity by traditional methods such as bioactivity-guided fractionation is a very elaborate, time-consuming, and laborious method. Recent advances in simulation technology have provided scope for basic in In silico studies, which are reliable and efficient for the identification of active compounds (Kumari et al. 2014). Hence, we screened and analyzed all the phytocompounds identified in the GC-MS analysis of the methanolic extract of E.thymifolia using a structure-based virtual screening in silico approach to identify potential targets implicated in breast cancer (Pinzi and Rastelli 2019; Herrera-Calderon et al. 2020).\n\nScreening and identification of the active components of Euphorbia thymifolia and identifying their potential targets implicated in breast cancer through Insilico approach.\n\n\nMethods\n\nPlant material collection and extract preparation by cold maceration method, GCMS analysis, Insilico molecular docking and molecular dynamics study.\n\nThe aerial parts of Euphorbia thymifolia L. were procured from the uncultivated areas of the Udupi District from April to June. The plant material was identified and authenticated by taxonomist Dr. K. Gopalakrishna Bhat, Professor of Botany (Rtd.) Poorna Prajna College, Udupi, The plant sample was deposited in the Herbarium of the Department of Pharmacognosy, Manipal College of Pharmaceutical Sciences. (Herbarium Voucher no: PP624) The aerial parts of Euphorbia thymifolia L. were collected, washed thoroughly and carefully with distilled water to remove all the soil and debris. The plant material was shade-dried at room temperature for 15 d until it was dry and crisp. After drying the shoots of E. thymifolia L., they were powdered and stored at 4°C.\n\nThe crude extract (10% w/v) 35 g in 350 ml was prepared by cold maceration as described by Abubakar et al using methanol procured from NICE Chemicals Private Limited (MI2037; B.no 103168) as the solvent for 72 h with intermittent shaking at room temperature. The extracts were then filtered through Whatman No. 1 filter paper. The clear extracts were concentrated using a rotary vacuum flash evaporator and freeze-dried by lyophilization, a process of drying the extract under controlled temperature and pressure using a lyophilizer. These lyophilized crude extracts were stored at 4°C until further use (Abubakar and Haque 2020).\n\nThe composition of the methanolic extract of E.thymifolia was analyzed by GC-MS at Analytical Research & Metallurgical Laboratories Pvt. Ltd. Bangalore, India.\n\nMolecular docking was performed using Schrodinger suite to select the best protein-ligand complex. Commercial Maestro software version 11.8 (OPLS3e force field) was utilized for all the simulation studies. Protein preparation wizard was used for preparation of protein using the panel review, modify, and refine modules where the side chains and residues were filled, and water molecules beyond 3 Å were removed. GLIDE panel was used for receptor-grid generation to create a grid and locate the receptor at the binding site for the ligands to be docked. The size of the grid box generated at the inbound ligand site for each protein was 10 × 10 × 10 Å (Rathi et al. 2019).\n\nThe crystal structures of the proteins were retrieved from the Protein Data Bank (Berman et al. 2000). ERK1 (Protein Data Bank ID: 4QTB, Crystallographic precision of the PDB: 1.40 Å) bound to a piperazine-phenyl-pyrimidine derivative (Chaikuad et al. 2014); AKT (Protein Data Bank ID: 4EKL, Crystallographic precision of the PDB: 2.00 Å) with an ATP site inhibitor which is a piperazine- pyrimidine derivative ligand (Lin et al. 2012); EGFR/HER2 (Protein Data Bank ID: 1XKK, Crystallographic precision of the PDB: 2.40 Å) with bound ligand Lapatinib is a tyrosine kinase inhibitor in clinical development for cancer (Wood et al. 2004); ER (Protein Data Bank ID: 2IOG, Crystallographic precision of the PDB: 1.60 Å) with indole ligand (Dykstra et al. 2007); The crystal structure of MELK (PDB ID: 5K00, Crystallographic precision of the PDB: 1.77 Å) with a ligand which is an amide derivative (Chen et al. 2017); Crystal structure of PLK1 (PDB ID: 4J52, Crystallographic precision of the PDB: 2.30 Å) with pyrimidodiazepinone as ligand (Nie et al. 2013), and Crystal structure of PTK6 (PDB ID: 5H2U,Crystallographic precision of the PDB: 2.24 Å) with ligand Dasatinib (Thakur et al. 2017).\n\nThe structures of all 219 phytocompounds were either retrieved from PubChem or derived using Chem-Draw software. These 219 ligands were imported into maestro and “LIGPREP” panel was utilized for ligand preparation (Rathi et al. 2019). Docking was performed using the Maestro Glide Module. Initially, all ligands were docked in standard precision (SP) mode, followed by extra precision (XP) mode to obtain a ligand docking XP score (Elokely and Doerksen 2013).\n\nAll XP docking files corresponding to each phytocompound and the seven proteins were then subjected to MM-GBSA free ligand energy calculation using the PRIME module to understand the binding energy of the target protein listed above with all 219 phytocompounds. (Ignjatović et al. 2016).\n\nTo determine the protein ligand stability under the simulated physiological environment, the DESMOND panel of Maestro was used for the MD simulation assessment, which was executed on HER2 (1XKK), ERK1 (4QTB) with 3,6,9,12-tetraoxatetradecane-1,14-diyl dibenzoate (TTDB) (molecular formula: C24H30O8), and ER(2IOG) with succinic acid, 2-(dimethylamino) ethyl 4-isopropylphenyl ester (SADPE) (molecular formula: C17H25NO4) for 100 ns using the XP docking file. Three steps, namely the system builder, minimization, and MD simulation, were involved in this process. A predefined simple point charge model (SPC) was used to create an orthorhombic boundary for the XP docked complex of TTDB with 1XKK, 4QTB, and 2IOG with SADPE. The charges were neutralized, where three positive charges were neutralized by the addition of three chloride ions, and one negative charge was neutralized by one sodium ion. An isothermal–isobaric (NPT) ensemble with a constant temperature of 300 K and pressure of 1 bar was maintained throughout the simulation for 100ns. To determine the stability of the complex, the Root Mean Square Deviation (RMSD) was analyzed along with the protein–ligand contact timeline and covalent/non-covalent interactions (Ivanova et al. 2018).\n\n\nResults\n\nGC-MS analysis of the methanolic extract of E.thymifolia showed 23 prominent peaks, with each of the retention time peaks having several hits comprising a total of 245 hits, as shown in Supplemental Data File 1. Each of these hits was recognized by comparing their retention time peak, peak area (%), and height to those of the already known compounds identified by the National Institute of Standards and Technology (NIST) library. Of the 245 phytocompounds, duplicates were removed and 219 unique compounds were identified in the methanolic extract of E.thymifolia. The identified phytocompounds were organic acids, phenyl esters, hydrocarbons, saturated and unsaturated cyclic compounds, and fatty acids.\n\nThe results of XP molecular docking using GLIDE were as follows: All 219 ligands with each of the seven proteins were comprehensively analyzed with their docking scores and amino acid interactions. The ligands that showed good interaction with most of the proteins were 3,6,9,12-tetraoxatetradecane-1,14-diyl dibenzoate (TTDB), succinic acid, and 2-(dimethylamino) ethyl 4-isopropylphenyl ester (SADPE), as shown in Table 1. TTDB showed very good interaction with two proteins, namely 1XKK and 4QTB, with docking scores: MMGBSA ΔG binding energies of -8.627, -65.22 kcal/mol and -10.112 and -71.103 kcal/mol with two types of non–covalent interactions, respectively. Similarly, 2IOG with SADPE exhibited a docking score of -8.865 and MMGBSA ΔG binding energy of -45.94 kcal/mol with two types of non-covalent interactions.\n\nThe amino acid residue Lys745 of the 1XKK protein showed a strong hydrogen bond and Phe856 showed pi-pi stacking with TTDB. Lys 71 and Met 125 of the protein 4QTB interacted with the oxy- group of TTDB. The amino acid residue Lys531 of 2IOG showed a hydrogen bond, and Trp383 showed pi-cation interactions with the ligand SADPE, as illustrated in Table 2. Hydrogen bond formation between a ligand and a protein plays a crucial role in drug design. Furthermore, the XP file obtained was subjected to the Prime MM-GBSA method to evaluate the binding affinity more accurately. The calculated binding energies (MM-GBSA ΔG) for 1XKK and 4QTB with the TTDB complex were − -65.228 kcal/mol and -71.103 kcal/mol respectively, and that of 2IOG with SADPE was -45.945 kcal/mol.\n\nFurthermore, to understand the interaction between the protein and ligand complex, a molecular dynamics study was performed for 100 ns. The RMSD values obtained were explained to obtain more clarity regarding the binding stability of the docked complexes.\n\nThe RMSD values of HER2 (1XKK) and TTDB were found to be around 4.8 Å, 4.0 Å respectively. The RMSD plot of both ligand and protein was stable after 20 ns, as shown in Figure 1. Later, the protein-ligand complex showed varied fluctuations until 50ns and then remained stable for the next 50ns. The molecular dynamics trajectory showed that amino acid residues such as Leu718, Ala 743, and Leu844 were vital before ligand (TTDB) stabilization up to 20 ns. Amino acid residues such as Val726, Leu844, and Phe856 established contacts with the ligand to be crucial hotspots for equilibrium, as shown in Figures 2B and 3.\n\nRMSD plot of TTDB bound to active inhibitor site of HER2 (1XKK).\n\nTo observe the protein-ligand interaction stability of TTDB with 4QTB, the RMSD values of the 4QTB with TTDB was found to be 2.2 Å and 2.4 Å respectively (Figure 4). This complex exhibited the highest stability throughout the simulation without larger deviations. Amino acid residues, such as Ile 48, Ala 69, Ile73, Arg 84, and Asp 184, were significant in stabilizing the complex until 20 ns. Other amino acid residues, such as Tyr 53, Lys 71, Tyr 81, Met 125, Leu 173, Val 56, and Cys 183, showed strong contact with TTDB and contributed to their strong stabilization for 100 ns. TTDB showed a prominent interaction with ERK (4QTB) (Figures 5B and 6 A & B).\n\nRMSD plot of TTDB bound to active inhibitor site of ERK (4QTB).\n\nThe RMSD values of ER (2IOG) and SADPE were approximately 5.0 Å, 4.0 Å respectively. The RMSD plot of both ligands and proteins stabilized after 20 ns for some time (Figure 7). Up to 20 ns, amino acid residues such as Thr347, Ala350, Leu 384, Asn532, Cys530, and Lys531 were important prior to ligand stabilization. Subsequently, the amino acid residues Asp351, Leu 387, Phe 404, Trp383, Leu525, and Tyr 526 showed strong interactions with the ligand, which contributed to stabilization (Figures 8B and 9).\n\nRMSD plot of SADPE bound to active inhibitor site of ER protein (2IOG).\n\n\nDiscussion\n\nThe phytocompounds were shortlisted based on their docking score, their interaction with more than one protein, and amino acid interactions. A literature search was carried out to investigate whether any of these compounds had anticancer activity. Chloride-substituted phenoxy acetic acid was used for the synthesis of a benzoyl hydrazide derivative, which has been reported to have in vitro antimicrobial activity (Al-Ostoot et al. 2019). To date, no other studies have been reported. TTDB and SADPE showed the highest docking scores against the proteins implicated in the pathogenesis of breast cancer, such as HER2 (IXKK), ERK1 (4QTB), and ER (2IOG). The XP docking results provided insight into the establishment of a strong interaction between a ligand and a protein that contains important amino acid residues. The interaction diagram for all proteins revealed the formation of more than one non-covalent bond, which contributed to the stability of the complex, as described in Table 2. The binding affinity calculated for the protein–ligand complex supports this statement. From previous literature, RMSD values below 4 Å are considered good, which implies minimal protein-ligand fluctuations (Hevener et al. 2009; Rao et al. 2021). In the simulations conducted for 100 ns for TTDB- HER2 & SADPE- ER, stability was established after 20 ns with non-covalent bond interactions. For the entire simulation, HER2 (IXKK) -TTDB had an average of four contacts with a maximum 6 and a minimum of two contacts (Figure 2A), and the ER (2IOG)- SADPE complex showed an average of six contacts, with a maximum of 9 and a minimum of three contacts (Figure 8A). The ERK1 (4QTB-TTDB) complex showed an average of eight contacts, with a maximum of 12 and a minimum of four contacts, with the highest stability number of interactions when compared to the other two complexes throughout the simulation (Figure 5A). In the HER2 (IXKK-TTDB) complex, the majority of contacts were made only by hydrophobic interactions (Figure 3). In TTDB- ERK1 complex Lys71 & Met 125 has made contact with the oxy groups for more than 80 % of the time. Other amino acids Tyr81 made a hydrophobic contact with the aromatic ring for approximately 46 % of the time and Cys 183 had hydrogen bonding with oxy group for 30 % of the time, as shown in the ligand-protein contact diagram (Figure 6A). In the ER (2IOG-SADPE) complex, Tyr 526 formed a non-covalent interaction due to hydrogen bonding and hydrophobic interactions for most of the contact time (Figure 9). In the case of HER2 and ERK1 protein complexes with TTDB, the activity is mainly attributed to the presence of an aromatic ring and multiple Oxy groups present in the ligand. The SADPE activity with the ER protein can be attributed to the presence of oxy groups. From the dynamics study, the ligand TTDB showed a profound interaction with ERK and was highly stable compared to its interaction with HER2. The SADPE-ER complex did not show a significant or stable interaction. Therefore, in our study, we report that TTDB has a greater affinity for binding to the ERK1 protein due to stable interactions, and therefore inhibits the protein. These in silico results can be further validated using an in in-vitro cytotoxicity assay. Hence, for the first time, we report that the anticancer activity of E. thymifolia L. could be attributed to the presence of the active compounds TTDB and SADPE in the crude methanolic extract.\n\n\nConclusion\n\nConsidering the docking score, H bond, binding stability, and contact time between ligand and protein, TTDB and SADPE could be suggested as the active components of the methanolic extract of E. thymifolia. Furthermore, these two compounds can be synthesized and tested for their in vitro anticancer activity in breast cancer cell lines to confirm the in silico results.", "appendix": "Data availability\n\nDryad: “Molecular docking and dynamics studies to identify novel active compounds targeting potential breast cancer receptor proteins from an indigenous herb Euphorbia thymifolia Linn” DOI: 10.5061/dryad.rn8pk0pjt (Kumblekar et al. 2024).\n\nThis work was licensed under a CC0 1.0 Universal (CC0 1.0) Public Domain Dedication license.\n\nDryad: STROBE checklist for ‘Molecular docking and dynamics studies to identify novel active compounds targeting potential breast cancer receptor proteins from an indigenous herb Euphorbia thymifolia Linn’ https://doi.org/10.5281/zenodo.10906175.\n\n\nReferences\n\nAbubakar AR, Haque M: Preparation of Medicinal Plants: Basic Extraction and Fractionation Procedures for Experimental Purposes. J. Pharm. Bioallied Sci. 2020; 12(1): 1–10. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nThakur MK, Birudukota S, Swaminathan S, et al.: Co-crystal structures of PTK6: With Dasatinib at 2.24 Å, with novel imidazo[1,2-a] pyrazin-8-amine derivative inhibitor at 1.70 Å resolution. Biochem. Biophys. Res. Commun. 2017; 482(4): 1289–1295. PubMed Abstract \| Publisher Full Text\n\nWaks AG, Winer EP: Breast Cancer Treatment: A Review. JAMA. 2019; 321(3): 288–300. Publisher Full Text\n\nWood ER, Truesdale AT, McDonald OB, et al.: A unique structure for epidermal growth factor receptor bound to GW572016 (Lapatinib): relationships among protein conformation, inhibitor off-rate, and receptor activity in tumor cells. Cancer Res. 2004; 64(18): 6652–6659. PubMed Abstract \| Publisher Full Text\n\nYousuf Z, Iman K, Iftikhar N, et al.: Structure-based virtual screening and molecular docking for the identification of potential multi-targeted inhibitors against breast cancer. Breast Cancer (Dove Med Press). 2017; 9: 447–459. 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PubMed Abstract \| Publisher Full Text" }	[ { "id": "324470", "date": "09 Oct 2024", "name": "Renukaradhya Math", "expertise": [ "Reviewer Expertise Microbiology", "biotechnology and bioinformatics" ], "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\nManuscript entitled “Molecular docking and dynamics studies to identify novel active compounds targeting potential breast cancer receptor proteins from an indigenous herb Euphorbia thymifolia Linn” by Kumblekar et al. is written well. My comments are below,\n1. Abstract (Background): author states “ Euphorbia thymifolia L is a widely growing tropical herb that has been reported to have various ethnopharmacological properties, including anticancer properties but further in Introduction section they state “However, the anticancer activity of this compound has not yet been reported so far” two statements are contradictory. In fact a recent study by Sulaiman C.T. et al 2023 (Ref 1) profiled anti-cancer activity of two Euphorbia species (thymifolia and hirta).\n2. Introduction section: author state “we have studied and reported the anticancer activity of the methanolic extract of E. thymifolia for the first time in our previous study” reference is missing!\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? Yes\n\nAre the conclusions drawn adequately supported by the results? Yes", "responses": [ { "c_id": "12596", "date": "10 Oct 2024", "name": "Vasavi Kumblekar", "role": "Author Response", "response": "Dear Dr. Renukaradhya Math, Thank you for taking your precious time to review our article and for the critical analysis. We would like to give our clarifications for the queries raised. Comment 1. Abstract (Background): author states “ Euphorbia thymifolia L is a widely growing tropical herb that has been reported to have various ethnopharmacological properties, including anticancer properties but further in Introduction section they state “However, the anticancer activity of this compound has not yet been reported so far” two statements are contradictory. In fact a recent study by Sulaiman C.T. et al 2023 (Ref 1) profiled anti-cancer activity of two Euphorbia species (thymifolia and hirta). Response: We actually meant to state that Euphorbia genus in general has been reported to have various ethnopharmacological properties including anticancer properties. But the species of interest in our study namely Euphorbia thymifolia although was reported to have several ethnopharmacological properties, anticancer property was not reported as per the literature available, when we initiated the project (2019). However we can rectify the statement already mentioned in the manuscript if you permit us to do so. Comment 2: Introduction section: author state “we have studied and reported the anticancer activity of the methanolic extract of E. thymifolia for the first time in our previous study” reference is missing! Response: We have studied the invitro anticancer activity of the methanolic extract of E. thymifolia. This original article has been communicated to the journal \"Jordan Journal of Pharmaceutical Sciences\" for possible publication, Hence the reference could not be cited until published. Requesting you to kindly review the responses and do the needful. Thanking You, Yours Sincerely Vasavi K" } ] } ]	1	https://f1000research.com/articles/13-385
https://f1000research.com/articles/13-1502/v1	09 Dec 24	{ "type": "Research Article", "title": "Is publishing gold open-access worth it? An assessment of hybrid and gold open-access publishing models of medical journals on their impact", "authors": [ "Abrar K. Thabit", "Noha I. Ashy", "Abdulrahman O. Tukruni", "Ali S. Alquzi", "Noha I. Ashy", "Abdulrahman O. Tukruni", "Ali S. Alquzi" ], "abstract": "Background Hybrid and gold open-access (OA) are the most common publishing models. The latter requires fees to allow full-text visibility upon publishing, whereas hybrid journals offer the option to publish gold OA or for free (subscription-based) where only users with access can get the full-text. We aimed to evaluate the impact of the publishing model and other factors on medical journals’ impact.\n\nMethods A sample of hybrid and gold OA medical journals indexed in Web of Science (WOS) and Scopus were included. The effect of the publishing model and other factors on journals’ impact factor (IF), CiteScore, quartile, and number of citations was assessed.\n\nResults 402 journals were included, 201 in each group. Hybrid and gold OA journals had a median age of 32 and 21 years, respectively (P<0.001). The median publishing cost in gold OA journals was $2,690, and 46.3% of them publish continuously. Publishing model, journal’s age, being of an organization/society, and EMBASE indexation didn’t affect IF, CiteScore, number of citations, and WOS quartile (P>0.05). However, gold OA model wasn’t significantly associated with Q1 ranking in Scopus (OR, 0.49; 95% CI, 0.25-0.94; P=0.032), which indicates that hybrid journals were more likely to have such ranking.\n\nConclusion These findings indicate that gold OA publishing doesn’t necessarily result in higher impact, which contradicts the claim that such model enhances citations. Therefore, authors can continue to publish in hybrid journals without being concerned about getting cited. Gold OA journals are encouraged to reduce their fees to facilitate global research access.", "keywords": [ "publishing", "hybrid", "open-access", "publisher", "journal", "journal impact factor" ], "content": "Introduction\n\nOpen access (OA) publications allow individuals to read published scientific articles for free, regardless of location, institutional affiliation, or subscription to the journal (or its publisher).1 Currently, OA publishing model includes several subtypes, the most common type is gold OA, where authors retain the copyright to their published article that is made freely available to any user in the online journal.2 Gold OA publishing model requires the payment of a certain fee by the authors. The other type of OA publishing model is green open access, which permits the placement of the authors’ manuscript in an institutional repository and then the article becomes freely available after an embargo period set by the journal. Green OA journals usually ask for page charges, in which the total is usually lower than gold OA charges. The concept of gold OA publishing was recently adapted within the last 20 years as journals used to be subscription-based only (i.e., access to articles are only allowed to users subscribed to the journal via their institution or a personal subscription while authors publish their work without a fee).3 However, upon the introduction of gold OA concept, it became widespread among publishers as numerus journals became hybrid, meaning that they allow both subscription-based publishing for authors opting to publish for free and gold OA publishing for authors willing to pay the publication fees. OA publishing is becoming more popular as large research funding agencies, such as the National Institutes of Health in the United States and the Bill and Melinda Gates Foundation, require researchers to make their work freely available.2,4\n\nAccess to medical literature is an essential part of healthcare professionals’ job to make decisions regarding disease management and implementation of certain interventions. Instead of attempting to base their decisions simply on the subset of research to which they have access, clinicians can make more informed health decisions when they have entire access to studies relevant to the cases they are treating.5 OA articles are more frequently read, and hence promote access to knowledge and new advances in different disciplines. Since the early 1990s, there has been a notable surge in OA publication, with an estimated annual growth in published articles of 30%.6 Furthermore, a number of universities and journal publishers have recently signed Open Access Transformative Agreements (OATAs), which require a greater transition to OA article publication over a predetermined time period, in response to recommendations from the research communities, such as the Efficiency and Standards for Article Charges Initiative.7\n\nDespite the advantages of OA publishing, it has a major disadvantage that should be acknowledged. To publish an OA article, journals ask the publishing fees, which are commonly known as article processing charges (APCs). These charges, which sometimes reach thousands of US dollars, may prevent authors with limited institutional or personal funds from publishing openly.8,9 The unaffordability of APCs may create a barrier to sharing research.10 Such major issue influenced many researchers to call for free access to research without barriers (i.e., the need for subscription or forcing authors to pay APCs to make their research accessible to all.11\n\nThe prevailing view suggests that the advent of OA publishing model enhances the spread of knowledge and potentially increases citation counts for authors; though, this continues to be contested. Therefore, the objective of this study is to evaluate the effect of the gold OA publishing model on several journals’ metrics on Web of Science (WOS) and Scopus by comparing it to hybrid publishing model.\n\n\nMethods\n\nMedical journals indexed in Science Citation Index Expanded (SCIE) of WOS and Scopus that are fully published in English were eligible for inclusion in the study. While journals that publish both clinical and basic sciences studies were included, journals that only publish articles in basic sciences, ethics, healthcare systems management, administration, and policy were excluded. Journals were divided into two groups based on their publishing model as either hybrid or gold OA.\n\n\n\n• Hybrid journals: Journals that offer subscription-based and gold OA publishing options. Subscription-based publishing means that only users with access to the journal through an institutional or a personal subscription to the journal can get the full-text of the published articles.\n\n• Gold OA journals: Journals that require authors to pay publishing fees that’s commonly known as APC to have their articles fully visible and available for download to anyone immediately upon publishing.\n\n• Impact factor (IF): 2-year IF of 2021-2022. It’s calculated based on the number of citations of all the articles published in the journal within the last two years.\n\n• CiteScore: 3-year CiteScore of 2019-2022. It’s calculated based on the number of citations of all the articles published in the journal within the last three years.\n\n• Number of citations in WOS: The total number of times articles published in the journal were cited since journal’s inception.\n\n• Number of citations in Scopus: The total number of times articles published in the journal were cited between 2019-2022.\n\n• Quartile: The quartile in which the journal is ranked, where Q1 involves the journals that are ranked the top 25% within a certain specialty, Q2 involves the journals that are ranked between 26-50%, Q3 involves the journals that are ranked between 51-75%, and Q4 involves the journals that are ranked between 76-100%. If a journal is ranked in more than one specialty, the average ranking was calculated.\n\nThe IF, CiteScore, number of citations, and quartile of each journal were collected from the journals’ records on Journal Citation Reports and Scopus.\n\nCategorical data were compared using Chi-square test and were presented as numbers and percentages, whereas continuous data were compared using Mann-Whitney test and presented using median [interquartile range (IQR)] as they were deemed skewed based on results of Shapiro-Wilk test of normality. Linear regression was conducted to assess the effect of different factors on IF and CiteScore. Multinomial regression was used to evaluate the association of different factors with Q1 ranking in WOS and Scopus. Odds ratios (OR) and 95% confidence intervals (CI) were reported. Fitting of the regression models was assessed using goodness-of-fit test. Omnibus test of significance was used to assess the significance of the regression models. A P value < 0.05 was deemed statistically significant. Analyses were performed on SPSS version 24.0 (IBM Corp., Armonk, NY, USA).\n\n\nResults\n\nA total of 402 journals included, 201 in each group. Table 1 shows the characteristics of the included journals. Hybrid and gold OA journals had a median [IQR] age of 32 [23–41.5] and 21 [13–28] years, respectively (P < 0.001). Most of the journals were international (39.3% vs. 53.7%; P < 0.001). In terms of volumes and issues, hybrid journals publish more frequently per year than gold OA journals (8 vs. 6; P = 0.041). However, 93 (46.3%) of the gold OA publish continuously; thus, were not included in the calculation of the median. More than half of the gold OA journals publish both clinical and basic sciences studies (35.3% vs. 53.2%; P < 0.001). The median [IQR] cost of publication in gold OA journals was 2,690 [2,000–2,990]. Although a significant difference was observed in terms of IF between the hybrid and the gold OA publishing models (2.90 [2.20–4.15] vs. 3.30 [2.40–4.45]; P = 0.021), no statistically significant difference was observed in terms of CiteScore, number of citations, and quartiles in both WOS and Scopus (P > 0.05 for all comparisons).\n\n* Only top 5 were listed.\n\n n=93 of gold open-access journals publish continuously (i.e., without assigning articles to a volume or issue); hence, not included in this median.\n\n* The remaining 77 journals were open-access since inception.\n\nWhen different factors were assessed for their effect on IF, only publishing both clinical and basic science studies was significantly associated with higher IF (OR, 2.03; 95% CI,1.19–3.45; P = 0.010). The same factor was associated with higher CiteScore (OR,3.63; 95% CI, 1.43–9.20; P = 0.007). On the other hand, the publishing model (hybrid vs. gold OA) did not influence IF nor CiteScore (OR, 1.38; 95% CI, 0.81–2.36; P = 0.237 and OR,0.81; 95% CI, 0.32–2.04; P =0.648, respectively). The effect of different factors on IF and CiteScore are shown in Tables 2 and 3, respectively.\n\nThe effect of different factors on the quartiles of the journals in WOS was evaluated, where none of the factors was associated with ranking in either quartile. Table 4 lists the factors and their effect on ranking in Q1 of WOS. On the other hand, hybrid journals were associated with ranking in Q1 of Scopus as the odds of gold OA journals being ranked as Q1 was less than one (OR, 0.49; 95% CI 0.25–0.94; P = 0.032). Table 5 lists the factors and their effect on ranking in Q1 of Scopus.\n\nA subgroup analysis in the gold OA group to assess the effect of publishing cost on ranking in WOS showed that higher APCs were associated with Q3 ranking (OR, 2.76; 95% CI, 1.22–6.23; P = 0.015) and negatively associated with Q1 ranking (OR, 0.57; 95% CI, 0.26–1.23; P < 0.001), but no effect was observed with Q2 ranking. The same observation was found with Q1 ranking in Scopus, where journals with high APCs were less likely to be ranked in Q1 (OR, 0.25; 95% CI, 0.11–0.54; P < 0.001).\n\n\nDiscussion\n\nThis is the first study to evaluate the impact of gold OA publishing model on journals’ metrics. Findings from this study suggest that gold OA publishing model does not have an impact on IF or CiteScore despite the significant difference in IF, but not CiteScore, between hybrid and gold OA journals. These results align with findings from previous studies that explored similar themes, indicating that while OA models aim to increase accessibility, they do not necessarily confer a citation advantage over hybrid models.12,13 For instance, some studies also found minimal differences in citation metrics between different publishing models, suggesting that other factors, such as journal reputation and article quality, might play more pivotal roles in influencing these metrics.14–16 A study by Chua et al found a higher number of citations of OA journals that did not correlate with IF (Spearman’s rho = 0.187; P = 0.60).16 Similarly, the increment increase in IF with increased citations was minimal and not statistically significant (β coefficient = 3.35; 95% CI -0.464–7.156; P = 0.084).16 As such, it is argued that the very high fees required by gold OA journals (reaching a median [IQR] of $2,690 [2,000–2,990] and a range of $75–5,460) are not justified when considering that many researchers call that science should be free to access by everyone. Therefore, these journals should consider lowering fees to cover necessary editorial processing and offer waivers for unfunded authors provided they provide a proof of lack of funds regardless of their country of origin (as many gold OA journals already offer waivers or discounts for authors from low-income countries). This approach aligns with the ethical imperative to democratize access to scientific knowledge, a sentiment echoed in the literature.14–16\n\nGold OA publishing model is a relatively new concept in research publishing as seen with the significant difference in the median [IQR] ages of hybrid and gold OA journals (32 [23–41.5] vs. 21 [13–28]; P < 0.001). Nonetheless, it become widespread, especially as many journals that were formerly hybrid have transitioned to gold OA, where the median [IQR] year of switching was 2014 [2008–2019]. This transition is indicative of the broader shift in the publishing landscape towards more open access models, a trend noted in earlier studies.14,15 Despite the significant difference in age, it is noteworthy that gold OA journals reached a median total number of citations in WOS since journals’ inception that was not statistically significant from that of hybrid journals, though it was numerically lower (3,963 vs. 4,918; P = 0.351). This could be possibly attributed to the large number of gold OA journals (n=93 of 201; 46.3%) that publish articles continuously without assigning them to a certain volume or issue, unlike hybrid journals that have limited quotas. This observation resonates with previous reports that showed that continuous publication models might influence citation patterns differently compared to traditional issue-based models.14,15\n\nThe lack of difference on the median total number of citations on Scopus reported for the years 2019–2022 contradicts the hypothesis that gold OA could achieve a high impact within a short period, given that Scopus reports citations within a specific time frame rather than a lifetime total, as is the case with WOS. This discrepancy highlights the complexity of evaluating impact across different databases per previous reports.14,15\n\nJournals that publish using only gold OA model market themselves to researchers looking to gain more citations and broader readership.17 Nevertheless, and contrary to that claim, the current study found no significant difference in the number of citations between gold OA and hybrid publishing models. Additionally, were observed more frequently with hybrid than with gold OA journals, while no difference was noted in the distribution of hybrid and gold OA journals among the different WOS quartiles. This outcome suggests that the prestige and perceived impact of hybrid journals may still hold sway in the academic community.14,15 Moreover, in their studies, Gargouri, et al and McCabe, et al have emphasized the importance of publishing both basic science and clinical research to reach a broader audience. The ability of journals that encompass a wide range of disciplines to appeal to a diverse readership has been highlighted as a significant factor in increasing a journal’s impact.14,15 In the current study, this notion was supported by the observation that hybrid journals, which often publish a mix of basic and clinical research, were more likely to achieve higher quartile rankings in Scopus.\n\nThese results of the current study suggest that unfunded researchers who cannot afford gold OA publishing should be less concerned about the citation impact of their work. It is worth noting that while articles published in subscription-based hybrid journals may not be freely accessible, readers can still obtain the full text by contacting the corresponding author via email or through researchers platforms like ResearchGate and Academia.edu.18 This highlights the role of author accessibility in ensuring the wider dissemination of research findings, regardless of the publishing model. Nevertheless, the current study highlights the importance of considering the call for free access to scientific information. To promote equitable access, gold OA journals should offer lower fees and waivers for unfunded authors, regardless of their country of origin. By reducing financial barriers, these journals can facilitate the dissemination of research and contribute to the broader scientific community.\n\nIt is important to acknowledge the strengths and limitations of this study. One of the strengths is that it is the first to directly compare the impact of hybrid and gold OA journals. By including a large random sample of journals and meeting the target sample size, the study provides a comprehensive analysis of the two models. It also evaluated several factors for their potential association with IF of WOS, CiteScore of Scopus, and quartiles of both databases. However, certain limitations should be considered. The study did not evaluate the impact of gold OA articles published in hybrid journals, which could potentially influence the overall impact comparison. Unfortunately, such an assessment was not feasible due to the longstanding existence of many hybrid journals. Additionally, journals that offer green OA, where articles become fully open access after an embargo period by paying page charges, were not included in the current study given their limited number, which could have made the comparison with the abundant hybrid and gold OA unjustified. Future research could explore the impact of these factors on citation metrics and readership. Further research is also warranted to investigate other aspects of OA publishing. While this study focused on IF and CiteScore, future studies could explore additional metrics such as altmetrics, which capture online attention and engagement with scholarly articles. Moreover, qualitative research methods could be employed to understand the perceptions and experiences of researchers regarding hybrid and gold OA publishing models. This would provide valuable insights into the decision-making processes and considerations of authors when choosing a publishing avenue. Furthermore, future research could delve deeper into understanding the impact of different research disciplines and the role of subject-specific factors in the citation patterns of hybrid and gold OA publications.\n\n\nConclusion\n\nIn conclusion, the findings of this study have noteworthy implications for both researchers and publishers by providing empirical evidence on the impact of hybrid and gold OA journals. The findings suggest that journals’ impact and number of citations did not significantly differ between hybrid or gold OA publishing models. This challenges the idea that publishing in gold OA journals leads to a higher impact as measured by citation metrics such as the IF and CiteScore. The study also emphasizes the need for gold OA journals to offer lower fees and waivers for unfunded authors to promote free access to scientific information. This research sets the stage for further investigations into the various dimensions of open access publishing and its implications for scholarly communication.\n\n\nAuthor contributions\n\nAKT: Conceptualization, methodology, data curation, investigation, supervision, project administration, writing – original draft, and writing – review & editing. NIA: Data curation, investigation, writing – original draft, and writing – review & editing. AOT and ASA: Data curation, investigation, and writing – original draft.\n\n\nEthics and consent statement\n\nNot applicable.", "appendix": "Data availability statement\n\nOpen Science Framework: Is publishing gold open-access worth it? An assessment of hybrid and gold open-access publishing models of medical journals on their impact. https://doi.org/10.17605/OSF.IO/SXV3E.19\n\nThis project contains the following underlying data:\n\n• Open-access vs. Hybrid journals – Supplementary material\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\n\nReferences\n\nInternational Association of Scientific Technical and Medical Publishers. An overview of scientific and scholarly journal publishing. The Netherlands: The Hague; 2015 [28 April 2024]. Reference Source\n\nPiwowar H, Priem J, Lariviere V, et al.: The state of OA: a large-scale analysis of the prevalence and impact of Open Access articles. PeerJ. 2018; 6: e4375. Epub 2018/02/20. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAbadal E: Gold or green: the debate on open access policies. Int. Microbiol. 2013 Sep; 16(3): 199–203. Epub 2014/02/27. PubMed Abstract\n\nMoorhead LL, Holzmeyer C, Maggio LA, et al.: In an Age of Open Access to Research Policies: Physician and Public Health NGO Staff Research Use and Policy Awareness. PLoS One. 2015; 10(7): e0129708.Epub 2015/07/23. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKoong A, Gardner UG, Burton J, et al.: Factors Associated With Open Access Publishing Costs in Oncology Journals: Cross-sectional Observational Study. JMIR Form Res. 2023 Mar 16; 7: e44633. Epub 2023/03/18. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLaakso M, Welling P, Bukvova H, et al.: The development of open access journal publishing from 1993 to 2009. PLoS One. 2011; 6(6): e20961. Epub 2011/06/23. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nESAC: Trnasformative Agreements.[5 May 2024]. Reference Source\n\nWyatt JC: Preserving the Open Access Benefits Pioneered by the Journal of Medical Internet Research and Discouraging Fraudulent Journals. J. Med. Internet Res. 2019 Dec 23; 21(12): e16532. Epub 2019/12/24. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVervoort D, Ma X, Bookholane H: Equitable Open Access Publishing: Changing the Financial Power Dynamics in Academia. Glob Health Sci Pract. 2021 Dec 31; 9(4): 733–6. Epub 2021/12/23. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSeguya A, Salano V, Okerosi S, et al.: Are open access article processing charges affordable for otolaryngologists in low-income and middle-income countries? Curr. Opin. Otolaryngol. Head Neck Surg. 2023 Jun 1; 31(3): 202–7. Epub 2023/05/05. PubMed Abstract \| Publisher Full Text\n\nKowaltowski A, Oliveira M, Silber A, et al.: The push for open access is making science less inclusive. Time Higher Education. 2021 [18 May 2024]. Reference Source\n\nBjork BC, Solomon D: Open access versus subscription journals: a comparison of scientific impact. BMC Med. 2012 Jul 17; 10: 73. Epub 2012/07/19. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nCraig ID, Plume AM, McVeigh ME, et al.: Do open access articles have greater citation impact?: A critical review of the literature. J. Informet. 2007 2007/07/01/; 1(3): 239–248. Publisher Full Text\n\nGargouri Y, Hajjem C, Lariviere V, et al.: Self-selected or mandated, open access increases citation impact for higher quality research. PLoS One. 2010 Oct 18; 5(10): e13636. Epub 2010/10/27. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMcCabe MJ, Snyder CM: Identifying the effect of open access on citations using a panel of science journals. Econ. Inq. 2014; 52(4): 1284–1300. Publisher Full Text\n\nChua SK, Qureshi AM, Krishnan V, et al.: The impact factor of an open access journal does not contribute to an article’s citations. F1000Res. 2017; 6: 208. Epub 2017/06/27. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\n5 Reasons to choose immediate Gold open access as an author. Springer Nature 2020 [10 June 2024]. Reference Source\n\nManca S: ResearchGate and Academia.edu as networked socio-technical systems for scholarly communication: a literature review. Res. Learn. Technol. 2018 02/20; 26. Publisher Full Text\n\nThabit AK, Ashy NI, Tukruni AO, et al.: Is publishing gold open-access worth it? An assessment of hybrid and gold open-access publishing models of medical journals on their impact.Publisher Full Text" }	[ { "id": "352802", "date": "06 Jan 2025", "name": "Lex Bouter", "expertise": [ "Reviewer Expertise Research Integrity", "Open Science", "Methodology" ], "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 this article 201 hybrid and 201 gold medical open access journals are compared in terms of several citation-based indicators. The findings indicate that gold open access journals have a slightly higher Impact Factor and are more likely to be in Quartile 1 of Scopus. Strangely that first key finding is not mentioned in the abstract, while the second finding is formulated in a confusing way (‘…wasn’t significantly associated..”). The narrative is somewhat difficult to follow, as there is a disconnect between the research question and the design of the study. The title (‘Is publishing gold open-access worth it?’) doesn’t state for whom this question will be answered. It turns out to boil down to the question whether the Article Processing Charge (APC) offers value for money in terms of impact from the author perspective. And this is where the disconnect comes in: the authors study impact at journal level, while authors will be interested in impact at article level. The problem is that indicators like the Impact Factor and other journal level indicators are poor predictors of the number of citations which a specific article in the journal at issue will get. There seems to be a growing consensus that gold open access publication leads to more citations and more citation diversity compared to paywalled publications. (See: Jeffrey Brainard. Open-access papers draw more citations from a broader readership. Science 24 January 2024. https://www.science.org/content/article/open-access-papers-draw-more-citations-broader-readership ). But there are more reasons why the study design is less suitable to answer the research question:\nHybrid journals are a mix of gold open access and paywalled publications, which diminishes the contrast studied. There are many differences between the included hybrid and gold open access journals that render causal interpretations very difficult. A much better study design would consist of a comparison between gold open access publications to a carefully matched group of similar paywalled publications. No mention is made of open methods (preregistration, study protocol in public repository) or open data. Please repair or explain why this is not done. Some indicators studied seem to concern absolute counts of citations. This is strange as the total number of articles will differ substantially between journals.\nInsufficient attention is given to the fact that from the perspective of authors both gold open access publications and paywalled publications suffer from financial barriers, as for both types of publications costs are involved that may be difficult to bear for authors or their institutions: APCs or journal subscriptions, respectively. The way forward might be diamond open access, but that promising solution is not mentioned in the article. The authors say that green open access involves page charges, which is not the case. (See e.g.: Open Access Network. https://open-access.network/en/information/open-access-primers/green-and-gold) Green open access (self-archiving), however, is correctly advocated as a way to make paywalled publications available for everyone, although that approach has a few disadvantages in terms of accessibility and sustainability. Finally, I sympathize with the plea to lower APC, but don’t see the connection to the findings of the study. In summary, the research question cannot be answered well with the study design at issue and already seems to be answered by others.\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": "352795", "date": "17 Jan 2025", "name": "Marc-André Simard", "expertise": [ "Reviewer Expertise open science", "open access", "bibliometrics", "science of science" ], "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 article compares the journal impact of hybrid and gold OA biomedical journals based on two proprietary data sources, namely Web of Science and Scopus. This study shows that, while it varies on certain conditions, gold OA journals may not result in more citations.\nI think criticism of gold OA is warranted, but I think the approach used by the authors is misguided because it ignores the market forces that are at play as well as other sociological factors that hybrid journals benefit from (e.g. publisher and journal reputation, prestige, green OA, transformative agreements, etc.).\nI also believe that the introduction is too short or not detailed enough and fails to grasp some of the main issues related to the topic such as publication prestige, impact, preprints, more inclusive data sources (such as OpenAlex and Dimensions), the for-profit nature of the system, article processing charges, OA policies and community infrastructure, transformative agreements, etc. The article would benefit from a more in-depth literature review. OA is a fast-changing topic involving multiple actors which the introduction does not accurately reflect.\nI also find the title/research question (Are gold journals worth it?) to be confusing as the article never really investigate more than research impact, which is only one of the facets (and I would argue it is the shallowest) of what makes OA/open science a more ethical choice. Furthermore, this link between research impact and publishing in OA journals is based on highly controversial and flawed journal-based metrics such as the impact factor and citescore, which are not a good predictor of individual article impact because the distribution of citations among journal articles is highly skewed (see Larivière and Sugimoto, 2018 [Ref 3]). Another problem with the design is that it doesn’t not take into consideration the fact that hybrid OA articles can also be OA (via green or APC payment), and generally benefit from greater historical prestige than gold journals. Multiple studies have shown that green OA papers tend to receive more citations on average than other types of OA for that reason (e.g. Piwowar et al. 2018).\nIn my opinion, the discussion/conclusion sections raise some valid points, but they are often too superficial and could use more depth and comprehension of the complex market forces at play. For instance, calling on gold OA publishers to lower APCs, while noble, is not a realistic or a sustainable approach because it doesn’t consider the for-profit nature of the system and how these actors are taking advantage of OA to generate more profits (e.g. see Butler et al. 2023 [Ref 2]). There is also very little criticism of the hybrid model (and no APC average, when previous research has shown that they are higher than gold), when it is arguably even worse than gold journals: their APCs are higher, they charge both readers and authors, and are also parts of unsustainable transformative agreements. Why not also call them out on reducing their APC/subscription/transformative agreement prices?\nSome more specific comments:\n\nThe abstract makes the false assumption that hybrid and gold are the most popular open access models, when both green and diamond are essentially just as big as the other two. The main difference is that they are generally not-for-profit, so they don’t benefit from the same visibility or attention as the other two models. There are some minor false information or assumption about OA:\nGreen OA doesn’t necessarily mean embargos Green OA and page charges: green OA is free for authors in the vast majority of cases. Green OA is allowed by the vast majority of publishers Not all journals ask for APCs. There are more than 25 000 diamond OA journals (e.g. see Khanna et al. 2023 [Ref 1] or the Directory of open access journals)\n\nUltimately, I appreciate what the authors were trying to accomplish with this study, but I do believe that some major flaws in the study design (e.g. focusing on journal-based metrics) and assumptions that were made (e.g. ignoring specific factors about hybrid OA) hold it back to the point where I cannot approve of this article.\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? 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": [] } ]	1	https://f1000research.com/articles/13-1502
https://f1000research.com/articles/12-974/v1	14 Aug 23	{ "type": "Research Article", "title": "Predictors of mortality for major trauma patients in intensive care: A retrospective cohort study", "authors": [ "Michael Jennings", "James Booker", "Amy Addison", "Rebecca Egglestone", "Ahilanandan Dushianthan", "James Booker", "Amy Addison", "Rebecca Egglestone", "Ahilanandan Dushianthan" ], "abstract": "Background: Major trauma places substantial demand on critical care services, is a leading cause of death in under 40-year-olds and causes significant morbidity and mortality across all age groups. Various factors influence patient outcome and predefining these could allow prognostication. The aim of this study was to identify predictors of mortality from major trauma in intensive care. Methods: This was a retrospective study of adult trauma patients admitted to general intensive care between January 2018 and December 2019. We assessed the impact on mortality of patient demographics, patterns of injury, injury scores (Glasgow Coma Score (GCS), Charlson’s comorbidity index (CCI), Acute Physiology and Health Evaluation II (APACHE II), Injury Severity Score (ISS) and Probability of Survival Score (Ps19)), number of surgeries and mechanism of injury using logistic regression. Results: A total of 414 patients were included with a median age of 54 years (IQR 34–72). Overall mortality was 18.6%. The most common mechanism of injury was traffic collision (46%). Non-survivors were older, had higher ISS scores with lower GCS on admission and probability of survival scores. Factors independently predictive of mortality were increasing age (OR 1.06, p <0.001) and GCS <15 on admission (OR 7.21, p <0.001). Ps19 was the best predictor of mortality (p <0.001 for each score category), with an AUROC of 0.90. Conclusions: The significant mortality predictors were age, fall from <2 metres, injury of head or limbs, GCS <15 and Ps19. Contrary to previous studies CCI and APACHE II did not significantly predict mortality. Although Ps19 was found to be the best current prognostic score, trauma prognostication would benefit from a single validated scoring system incorporating both physiological variables and injury patterns.", "keywords": [ "intensive care", "critical care", "trauma", "mortality", "scoring systems" ], "content": "Introduction\n\nMajor trauma accounts for almost 10% of all deaths worldwide.1 The National Audit Office estimates that there are more than 20,000 major trauma cases each year in England, resulting in 5,400 deaths.2 Furthermore, the demographics and injury patterns of the major trauma population in the UK are changing. Data from the Trauma Audit and Research Network (TARN) show that there has been an increase in the mean age of trauma patients between 1990 and 2013 (36.1 years to 53.8 years), along with a change in the most common mechanism of injury from road traffic collision (RTC) (59.1%) in 1990, to low fall (39.1%) in 2013. Critically unwell trauma patients typically require admission to the Intensive Care Unit (ICU), frequently making up the most resource-intensive critical care patient group (46.9% of ICU patients in a multicentre US study),3 with significant morbidity and mortality. The burden caused by trauma patients on healthcare systems and the shifts in trauma patient populations has increased the need for evaluating existing scoring systems for their prognostication potential in the ICU trauma population.4\n\nPredicting mortality in the critically unwell trauma patient poses a significant challenge due to the heterogeneity of the patient group and the multitude of patient specific factors that affect ICU outcomes, such as age, comorbidities, and injury patterns.5–10 Many of these factors have been examined by several previous studies, but there is no consensus on the most useful prognostication scores. Both physiological and anatomical scoring systems have been purported to correlate best with mortality,11,12 thus there is a current requirement for development of a new scoring tool for early mortality prediction in trauma ICU patients that incorporates a combination of physiological and anatomical scoring components.13,14\n\nThe aim of this investigation was to determine which patient specific factors (present at the point of admission) and which injury severity scoring systems are the most accurate predictors of poor outcome in trauma patients admitted to ICU.\n\n\nMethods\n\nThis is a retrospective study of all critically ill trauma patients aged ≥18 years admitted to the General Intensive Care Unit (GICU) at Southampton General Hospital, between January 2018 and December 2019.15 Only patients admitted to the GICU were included in this study and patients admitted to other areas including high dependency units and neuroscience intensive care unit were excluded. The sample size was determined by the number patients admitted during the defined time period and was comparable to studies of similar design. Authors did not have access to information that could identify individual participants during or after data collection. Ethical approval was obtained through Ethics and Research Governance Online (ERGO) by the Faculty of Medicine at Southampton University on 4 August 2020, Reference 56519. This study was part of the large CRIT-CO study (Outcomes of Patients Admitted with Critical-Illness to the General Intensive Care Unit – a Retrospective Observational Study) IRAS Reference 232922. This study used retrospective analysis of non-identifiable patient data, thus the need for individual informed consent was waived.\n\nThe following variables were collected from all available Southampton General Hospital databases: age, sex, comorbidities, mechanism of injury, and injury distribution.15 Admission scores including Glasgow Coma Score (GCS), Injury Severity Score (ISS), TARN Probability of Survival Score (Ps19), and the Acute Physiology and Health Evaluation II (APACHE II) score were calculated. We also quantified the prehospital comorbidity by Charlson’s comorbidity index (CCI). The outcomes evaluated were duration of mechanical ventilation, ICU, and hospital length of stay and 28-day all cause hospital mortality.\n\nContinuous variables are expressed as median and interquartile range (IQR). Mann Whitney U was used as the statistical analysis for continuous variables and chi-square for categorical variables. The distribution of variables was assessed and if they had a non-normal distribution they were dichotomised into categorical variables with equal sized groups. Univariate analysis using logistic regression to investigate if variables that varied significantly between the survival and non-survival group, were also significant predictors of mortality. Prior to a multivariate analysis a correlation matrix was done to assess the collinearity between each of the significant predictors using Spearman’s test. This informed the subsequent multivariate analysis using a logistic regression to identify independent significant predictors of survival. Predictors were deemed significant if p < 0.05. Additionally, receiver operating characteristics (ROC) area under the curve (AUC) graphs were constructed to assess each variables performance in predicting mortality. Data analysis was done in SPSS Version 25 (RRID:SCR_016479) and RStudio Version 1.4.1103 (RRID:SCR_000432) using packages: dplyr, ggplot2, lme4 and pROC.15\n\n\nResults\n\nA total of 414 critically injured trauma patients were admitted to the Intensive Care Unit between January 2018 and December 2019. Of these, 69.3% (n = 287) were male and 30.7% (n = 127) were female. The median age was 54 years (IQR 34–72). Of those admitted, 66.2% (n = 274) had at least one co-morbidity and the median CCI was 1 (IQR 0–3). The most common mechanism of injury was vehicle incident (46.1%), followed by fall <2 metres (23.9%). The most common body part injured was chest (29.2%), followed by other (20%), multiple injuries (19.1%), head (12.1%), abdominal (8.7%), spinal (6.3%), limbs (3.6%) and facial injuries (1%) (Figure 1). The median GCS, ISS and APACHE II scores were 15, 22 and 11 respectively.\n\nOverall, the 28-day all cause hospital survival was 81.4% (n = 337), with survivors being on average younger than non-survivors (51 (32–68) vs 74-years-old (55–85). There were no survival differences between male and female patients. Survivors had fewer comorbidities than non survivors (CCI medians of 1 (0–4) and 4,1–4 respectively). Among non-survivors, fall of <2 metres was the most common mechanism of injury (39%) reflecting the older age of this group, followed by an RTC (32.5%). Among survivors, the most common mechanism of injury was RTC (49.3%), followed by fall from <2 metres (20.5%). Non-survivors had lower GCS at presentation than survivors (9 vs 15). They also had higher ISS (25 vs 20) and higher APACHE-II scores (13 vs 11) than survivors at presentation.\n\nThe Ps19 predictive model was significantly lower for non-survivors (59 vs 98). The type of body region injured also varied between survivors and non-survivors. The non-survivors had increased frequency of head injury and the survivors had more chest injuries. Abdominal injuries were more common in survivors than non-survivors (10.1% vs 2.6%), whereas limb injuries were more common in non-survivors than survivors (10.4% and 2.1% respectively) (Table 1).\n\nWe performed univariate logistic regression analysis to assess the association between common variables that demonstrated significant difference between survivor and non-survivor groups with 28-day hospital survival (Table 2). In the univariate analysis, increasing age (OR 1.04, CI 1.03–1.06, p < 0.001), fall <2 metres (OR 3,17, CI 1.74–5.84, p < 0.001), GCS <15 (OR 3.79, CI 2.21–6.63, p < 0.001), ISS 41–60 (OR 3.10, CI 1.46–6.46, p = 0.00269), Ps19 < 81 (p = 0.001), number of surgeries (OR 0.627, CI 0.467–0.806, p < 0.001) and the most severely injured body region of the head (OR 11.1, CI 4.87–27.1, p < 0.001), multiple injuries (OR 2.60 CI 1.12–6.31, p = 0.0288) and other injuries (OR 12.7, 95% CI (3.84–44.0, p = 0.001) were all significant predictors of mortality (Table 2). A multivariate analysis was conducted using these variables which demonstrated that increasing age (OR 1.06, CI 1.04–1.08, p < 0.001) and GCS < 15 (OR 7.21, CI 3.80–14.3), p < 0.001) were independent predictors of mortality (Table 3).\n\n* p < 0.05.\n\n*** p < 0.001.\n\n* p < 0.05.\n\n*** p < 0.001.\n\nWe performed a probability of survival analysis based on variables including age and the scoring systems Ps19, ISS, GCS, and APACHE-II (Figure 2). For Ps19 (Figure 2A), patients with a low Ps19 0–20 had an almost linear decrease in survival probability up until 14 days, had a lower survival probability and were more likely to die sooner. Patients with Ps19 scores between 21–60 had similar survival probabilities until day seven, at which point they diverge with the 41–60 group having the lowest survival probability at 28 days (28%), and the 21–40 group having a survival probability of 44%. Patients with Ps19 scores of 61 or higher had significantly higher probability of survival than the other groups, with the 61–80 group demonstrating more than 70% chance of survival at 28 days. The Ps19 score >80 group demonstrated a survival probability of over 90%.\n\nProbability of survival against total number of days in hospital. Patients are categorised into groups based on their scores in scoring systems. A: Ps19 Survival probability. B: APACHE II Survival probability. APACHE II scores could not be calculated for 139 patients, so this is taken into consideration with a N/A line. One patient was excluded from the APACHE II graph due to being the only one who had a score >30. C: GCS Survival probability. D: ISS Survival probability.\n\nAPACHE II: Acute physiology and chronic health evaluation; GCS: Glasgow coma scale; ISS: Injury severity score; Ps19: Probability of survival.\n\nFor APACHE II score, likelihood of survival at 28 days decreased with increasing APACHE II scores (Figure 2B). Until the seventh day there was a similar survival curve for all APACHE II scores groups, after which patients with a score of 0–10 clearly show a higher probability of survival compared to patients with an APACHE II score of 11–20 or 21–30, (92%, 89% and 87% at 28 days, respectively).\n\nPatients with reduced GCS (Figure 2C), had lower likelihood of survival compared to those admitted GCS 15 (73% and 94% at 28 days respectively).\n\nFor ISS scores (Figure 2D), patients in the highest score range (61–80) had only a 50% chance of survival at 28 days. Patients with a score of 41–60 had a 70% survival probability, those scoring 21–40 had an 84% chance of survival whereas the group scoring 1–20 had a 90% probability of survival. Thus, a higher ISS score was associated with a lower probability of survival.\n\nThe difference between the predicted Ps19 and observed mortality for the cohort is shown in Figure 3. The Ps19 predicted score was similar to the expected mortality for most ages, except for the groups >80 years of age (Figure 3).\n\nCalculated for APACHE II Score, ISS, GCS, Ps19 and Number of surgeries in mortality correlation prediction.\n\nROC: receiver operator characteristic, AUROC: area under the receiver operator curve, APACHE II: Acute physiological assessment and chronic health evaluation, ISS: Injury Severity Score, GCS: Glasgow Coma Scale, Ps19: Probability of Survival Score.\n\nThe area under the receiver operator curve (AUROC) was statistically significant for all variables (Figure 4). Ps19 was the best predictor of mortality with an AUROC of 0.90 (95% CI 0.85–0.96) followed by GCS AUROC of 0.75 (95% CI 0.64–0.86) and age 0.73 (95% CI 0.62–0.85). ISS, APACHE II and number of surgeries were less predictive of mortality in comparison to these variables with ISS being the worst predictor with an AUROC of 0.66 (95% CI 0.50–0.76).\n\nOverall, 190 patients (45.9%) required invasive mechanical ventilation and the proportion was higher in non-survivors, compared with survivors (68.8% vs 40.7%) and for both groups the duration of mechanical ventilation was three days. Median ICU and hospital length stay were 3 (IQR 1, 7) and 13 (IQR 7, 26) respectively. There was no statistically significant difference between the ICU length of stay (LOS) for survivors and non-survivors; however, survivors had a longer hospital LOS (15 vs 7 days p < 0.01) (Table 4).\n\nData presented as median and interquartile ranges.\n\n* p < 0.05.\n\n*** p < 0.001.\n\n\nDiscussion\n\nThis study evaluated different patient specific and injury specific factors that influence hospital mortality in critically ill trauma patients. Patient specific factors we investigated included age, gender, and pre-existing comorbidities. Increased age was unsurprisingly found to be associated with a higher mortality. This association is likely to be multifactorial due to an increased risk of frailty, higher likelihood of under triage, and altered physiological mechanisms in elderly patients. Presence of comorbidities in our cohort as determined by CCI was found to be poorly predictive of mortality, in contrast to previous studies which have found a direct impact on mortality.13 Whilst some studies have given conflicting evidence as to the effect of comorbidities on hospital length of stay,16,17 their importance is acknowledged by their inclusion in trauma scoring systems such as the Ps19 model. We found age and reduced GCS on admission were associated with increased hospital mortality in critically ill trauma patients.\n\nInjury specific factors such as mechanism of injury, body region injured, and severity of injury were all found to affect mortality. In our cohort, most deaths were due to head injuries or polytrauma. Falls and RTC were the most common mechanism of injury, which is consistent with published data from both the UK and the USA.18,19 The overall mortality in our cohort was 18.6%, with an increased mortality in patients with a fall from <2 metres. Of those injured in an RTC, 86.9% survived compared to 69.7% of those who fell <2 metres. The likely explanation for this finding is that patients who suffer injuries from falls <2 metres resulting in ICU admission are most likely to be an older, more frail population, which is consistent with national data from the TARN database.20 Moreover, older patients tend to have increased comorbidities with concurrent risk of polypharmacy including anticoagulant medication, which increases risk of adverse outcome from trauma.\n\nTwo recent single-centre retrospective observational studies found the following factors to be associated with increased mortality in ICU from trauma: age >60 years, comorbidities (CCI), severity of trauma (New Injury Severity Score (NISS) and Revised Injury Severity Classification (RISC)), patient severity (APACHE II), traumatic brain injury, the use of mechanical ventilation, renal dysfunction in the first 24 hours, and the use of vasoactive drugs and circulatory complications.21,22 In contrast to our findings, the scores most highly predictive for mortality were APACHE II and NISS. An Australian meta-analysis of over 5000 patients across 25 centres demonstrated similar findings but also showed the Australian and New Zealand Risk of Death (ANZROD) mortality prediction model and APACHE III to be superior to the anatomical scoring systems for mortality prediction (e.g., ISS and NISS).13\n\nAlthough a higher APACHE II score and CCI were more commonly found in the non-survivor group in our study, these associations proved inadequate predictors of mortality in univariate analyses. However, our findings agreed with previous work that an increased ISS and decreased Ps19 both predict increased mortality.22–25 Ps19 was our best performing mortality predictor with an AUROC of 0.9 (95% Cl 0.85–0.96), outperforming ISS (the most used scoring tool).\n\nOur study presents significant limitations. Firstly, the dataset was collected retrospectively from a single centre, notably excluding patients with primary head injury. Secondly, our study did not extensively examine indicators or patient morbidity following trauma; for example, renal dysfunction and ICU interventions such as other organ support measures including renal replacement therapy and the use of vasopressors. We also did not assess other important outcomes such as lasting neurological deficits, rehabilitation required following discharge, which may have provided further context for mortality prediction analyses. Finally, the outcome of our study was limited to 28-day mortality and does not report mortality data at longer timepoints. Whilst there are logistical challenges with data collection over extended timeframes in ICU patients following their discharge, the decision to limit the mortality window to 28 days limits the scope of conclusions to prognostication within a short pre-defined window. Nevertheless, our study complements the existing literature with noteworthy analysis of the mortality prediction capability of a range of scoring systems including ICU specific scoring systems and presents comparable sample sizes to similar recent single-centre studies.21,22 It was also noted that neither of these studies included the Ps19 scoring system, which is currently used by the TARN (UK), which we found to be the most highly predictive of mortality from trauma in ICU.\n\n\nConclusions\n\nThis study shows that various internal and external factors determine the mortality of ICU patients within a single-centre general ICU. The most significant prognosticators were age, fall from a height of <2 metres, injury of head or limbs, and scoring systems: GCS, Ps19 and ISS. Ps19 was the best performing score for mortality prediction. Contrary to previous studies, we did not demonstrate an association between mortality and the CCI and APACHE II scoring systems. Trauma prognostication would benefit from an all-inclusive single validated scoring system incorporating physiological variables, injury patterns and ICU variables. This could mitigate extended ICU stays by ensuring that interventions patients receive are better tailored to their individual physiological profile and possibly reduce overall mortality in ICU trauma patients. Future studies would benefit from inclusion of morbidity indicators to provide context for the quality of life experienced by survivors of major trauma.", "appendix": "Data availability\n\nZenodo: Underlying data for ‘Predictors of mortality for major trauma patients in intensive care: A retrospective cohort study’, https://doi.org/10.5281/zenodo.8032931. 15\n\nThis project contains the following underlying data:\n\n• Data sheet 2018–2019 29.09.20.xlsx\n\n• Trauma analysis.pptx\n\n• Ps19 vs survival.tiff\n\n• ROCs.tiff\n\nSTROBE checklist for ‘Predictors of mortality for major trauma patients in intensive care: A retrospective cohort study’, https://doi.org/10.5281/zenodo.8032931. 15\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\nKrug EG, Sharma GK, Lozano R: The global burden of injuries. Am. J. Public Health. 2000 Apr; 90(4): 523–526. PubMed Abstract \| Publisher Full Text\n\nMorse A: Major trauma care in England. National Audit Office. National Audit Office London; 2010 [cited 2022 Aug 20]. Reference Source\n\nMichetti CP, Fakhry SM, Brasel K, et al.: Trauma ICU Prevalence Project: The diversity of surgical critical care. Trauma Surg. Acute Care Open. 2019 Feb 1; 4(1). Publisher Full Text\n\nKehoe A, Smith JE, Edwards A, et al.: The changing face of major trauma in the UK. Emerg. Med. J. 2015 Dec 1; 32(12): 911–915. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEvans CC, Petersen A, Meier EN, et al.: Prehospital traumatic cardiac arrest: management and outcomes from the resuscitation outcomes consortium epistry-trauma and PROPHET registries. J. Trauma Acute Care Surg. 2016; 81(2): 285–293. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEid HO, Barss P, Adam SH, et al.: Factors affecting anatomical region of injury, severity, and mortality for road trauma in a high-income developing country: lessons for prevention. Injury. 2009; 40(7): 703–707. PubMed Abstract \| Publisher Full Text\n\nCamilloni L, Farchi S, Rossi PG, et al.: Mortality in elderly injured patients: the role of comorbidities. Int. J. Inj. Control Saf. Promot. 2008; 15(1): 25–31. Publisher Full Text\n\nEgglestone R, Sparkes D, Dushianthan A: Prediction of mortality in critically-ill elderly trauma patients: a single centre retrospective observational study and comparison of the performance of trauma scores. Scand. J. Trauma Resusc. Emerg. Med. 2020; 28(1): 1–7.\n\nKisat MT, Latif A, Zogg CK, et al.: Survival outcomes after prolonged intensive care unit length of stay among trauma patients: The evidence for never giving up. Surgery. 2016; 160(3): 771–780. PubMed Abstract \| Publisher Full Text\n\nSaika A, Bansal S, Philip M, et al.: Prognostic value of FOUR and GCS scores in determining mortality in patients with traumatic brain injury. Acta Neurochir. 2015; 157(8): 1323–1328. Publisher Full Text\n\nHwang SY, Lee JH, Lee YH, et al.: Comparison of the Sequential Organ Failure Assessment, Acute Physiology and Chronic Health Evaluation II scoring system, and Trauma and Injury Severity Score method for predicting the outcomes of intensive care unit trauma patients. Am. J. Emerg. Med. 2012; 30(5): 749–753. PubMed Abstract \| Publisher Full Text\n\nWu SC, Chou SE, Liu HT, et al.: Performance of prognostic scoring systems in trauma patients in the intensive care unit of a trauma center. Int. J. Environ. Res. Public Health. 2020 Oct 1; 17(19): 1–12. Publisher Full Text\n\nMagee F, Wilson A, Bailey M, et al.: Comparison of Intensive Care and Trauma-specific Scoring Systems in Critically Ill Patients. Injury. 2021; 52(9): 2543–2550. PubMed Abstract \| Publisher Full Text\n\nPolita JR, Gomez J, Friedman G, et al.: Comparison of APACHE II and three abbreviated APACHE II scores for predicting outcome among emergency trauma patients. Rev. Assoc. Med. Bras. 2014; 60: 381–386. PubMed Abstract \| Publisher Full Text\n\nJennings M: m-a-jennings/ICUtraumapredictors: v1.0.1.2023 Jun 13 [cited 2023 Jul 10]. Publisher Full Text\n\nGruskay JA, Fu M, Bohl DD, et al.: Factors affecting length of stay after elective posterior lumbar spine surgery: a multivariate analysis. Spine J. 2015; 15(6): 1188–1195. PubMed Abstract \| Publisher Full Text\n\nPugely AJ, Martin CT, Gao Y, et al.: Comorbidities in patients undergoing total knee arthroplasty: do they influence hospital costs and length of stay? Clin. Orthop. Relat. Res. 2014; 472(12): 3943–3950. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nDiMaggio C, Ayoung-Chee P, Shinseki M, et al.: Traumatic injury in the United States: in-patient epidemiology 2000–2011. Injury. 2016; 47(7): 1393–1403. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChristensen MC, Ridley S, Lecky FE, et al.: Outcomes and costs of blunt trauma in England and Wales. Crit. Care. 2008; 12(1): 1–13.\n\nBanerjee J, Baxter M, Coats T, et al.: Major Trauma in Older People.2017 [cited 2022 Mar 23]. Reference Source\n\nLentsck MH, de Oliveira RR , Corona LP, et al.: Risk factors for death of trauma patients admitted to an Intensive Care Unit. Rev. Lat. Am. Enfermagem. 2020 Feb 28; 28: e3236–e3236. Publisher Full Text\n\nPapadimitriou-Olivgeris M, Panteli E, Koutsileou K, et al.: Predictors of mortality of trauma patients admitted to the ICU: a retrospective observational study☆. Braz. J. Anesthesiol. 2021; 71(1): 23–30. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSainz Cabrejas J, García Fuentes C, García Juarranz C, et al.: Evaluation of quality of care in trauma patients using international scoring systems. Medicina Intensiva (English Edition). 2020; 44(6): 325–332. Publisher Full Text\n\nXiao Y, Jin H, Mei H, et al.: The predictive value of combination of anatomic scoring system and physiological scoring system in prediction of death in patients with severe trauma: a multicenter analysis of 614 cases. Zhonghua Wei Zhong Bing Ji Jiu Yi Xue. 2015; 27(4): 291–294. PubMed Abstract \| Publisher Full Text\n\nKarataş AÖ, Çam R: The effect of the use of trauma scoring systems on prognosis of patients with multiple traumas: A cross-sectional study.2018." }	[ { "id": "246878", "date": "01 Mar 2024", "name": "Adnan Özpek", "expertise": [ "Reviewer Expertise General Surgery", "Trauma and Emergency 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 congratulate the authors for their study. My suggestions for the study are as follows: 1- It appears that all the patients in the study were blunt trauma patients, which should be noted in the title of this article. If patients with penetrating trauma are included in the study, it should be stated. 2- Calculated Revised Trauma Score (cRTS) also can be used in addition to the trauma scoring systems used in the study. 3- In the study, mortality rates for falls below 2 meters were significantly higher than for falls above 2 meters. This is not an expected result. It should be questioned whether the patients who fell below 2 meters fell due to an intracranial neurological reason.\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": [ { "c_id": "11778", "date": "24 Jun 2024", "name": "Michael Jennings", "role": "Author Response", "response": "Thank you very much for your comments. They have been addressed in the new updated manuscript. 1) The article has been amended to include stipulation that only blunt trauma patients were included. See changes to title, abstract and methods section. 2) Mention of cRTS has been added to the limitations section of the discussion as an example of useful scoring systems that were omitted. 3) Factors contributing to our finding that falls < 2 metres being associated with higher mortality have been discussed in more detail in the discussion section." } ] }, { "id": "266548", "date": "21 May 2024", "name": "Neta Cohen", "expertise": [ "Reviewer Expertise pediatric emergency medicine", "emergency medicine", "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 is a retrospective study of all critically ill trauma patients aged ≥18 years admitted to the General Intensive Care Unit (GICU), aimed to determine which patient specific factors (present at the point of admission) and which injury severity scoring systems are the most accurate predictors of poor outcome in trauma patients admitted to ICU. Such similar studies were performed earlier (Cohen, N., et.al., 2023 (Ref 1) However, considering the importance of the topic, and the appropriate design, I believe it merit publication. Comments: 1. Why did you exclude patients from high dependency units and neuroscience intensive care unit? 2. CCI and AGE are co-factors, as age >60 is one of the factors including in the CCI. Try to examine each one separately in the logistic regression model. Please add it in addition to your limitation. 3, How do you explain falling from height< 2 meters to be more severe than fall > 2 meters? how did you decide about cut-off of 2 meters ( the usual cutoff is 3 meters).? 4. Please explain better what are the implication of your 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? 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": [ { "c_id": "11779", "date": "24 Jun 2024", "name": "Michael Jennings", "role": "Author Response", "response": "Thank you very much for your comments. The manuscript has been amended to address them and we have now submitted an updated version. 1) Exclusion of patients from the high dependency unit and the neuroscience intensive care unit has been further explained within the methods section. 2) To address your comments that CCI and age are cofactors, the univariate and multivariate analyses in tables 2 and 3 have been reanalysed with age as a categorical variable. 3) Falls of less than 2 meters were found to be a significant predictor of mortality in the univariate analysis, but not the multivariate analysis. The explanation of this finding has been added to the discussion section along with an explanation for why the cut-off of two meters was chosen. 4) This discussion and conclusion sections of the manuscript have been improved to explain the implications of our findings in more detail. Many thanks" } ] } ]	1	https://f1000research.com/articles/12-974
https://f1000research.com/articles/13-1071/v1	19 Sep 24	{ "type": "Review", "title": "Recent Advances in Biopesticide Research and Development: A Focus on Microbial: A Review", "authors": [ "Kahsay Tadesse Mawcha", "Lawrence Malinga", "Debbie Muir", "Jing Ge", "Dennis Ndolo", "Lawrence Malinga", "Debbie Muir", "Dennis Ndolo" ], "abstract": "Biopesticides are pest control products derived from natural sources such as microbes, macro-organisms (insects and pathogens), plant extracts, and certain minerals. Many biopesticides are considered environmentally safe and can complement or substitute conventional chemical pesticides. They can also be highly specific or broad spectrum with a unique mode of action controlling a wide range of pest species. Due to their target-specificity and low to no environmental residuality, biopesticides conform to the 3 pillars of Climate-Smart Agriculture, the Sustainable Development Goals, and ultimately, the Paris Agreement. This review focuses largely on microbial biopesticides derived from fungi, bacteria, viruses, and nematodes. It discusses (i) the various microbial biopesticide formulations, (ii) the mode of microbial biopesticide action, (iii) the factors that affect the potential efficacy of biopesticides, (iv) challenges to the adoption of microbial biopesticides, and (v) the role of microbial biopesticides in Integrated Pest Management programs. Finally, advancements in application techniques, as well as future research directions and gaps are highlighted.", "keywords": [ "biocontrol", "bioprotection", "formulation", "climate-smart agriculture", "sustainable pest management" ], "content": "1. Introduction\n\nBiopesticides are naturally derived pest control compounds that offer an environmentally safer and target-specific complement to conventional chemical pesticides (Leo and Rathore 2015; Tijjani et al. 2016; Nuruzzaman et al. 2019; Temiz 2020; Wattimena and Latumahina 2021; Huang et al. 2022). Biopesticides include 1) biochemicals (plant extracts, semiochemicals, microbial extracts/fermentation products, insect growth regulators, compounds synthesized by other organisms, and inorganic compounds), 2) microbial biopesticides (bacteria, fungi, protozoa, viruses, oomycetes, yeast, and algae), 3) macrobials (insect predators, parasitoids, and entomopathogenic nematodes) and classical biocontrol agents (Copping and Leonard 2000; Chandler et al. 2008; Leahy et al. 2014; Liu et al. 2021; Ram and Singh 2021). Biopesticides play a crucial role in promoting climate-smart agriculture (CSA) by offering effective pest management solutions, improving crop yield and quality, and reducing pest resistance while minimizing negative impacts on human health and the environment (Szewczyk et al. 2006; Satish et al. 2017; Kumari et al. 2022). The growing need for biopesticides is partly a consequence of the decrease in the use of conventional pesticides due to farmers increasingly adopting climate-smart agricultural practices and reducing their carbon footprint (FAO 2019). This shift is driven by several advantages associated with biopesticides, including 1) their ability to slow down the development of pest resistance, 2) low to no toxicity to humans and the environment, hence safeguarding biodiversity, preserving soil health and enhancing food security 3) their ability to complement chemical pesticides, 4) their low environmental fate, 5) host specificity, 6) biodegradability, 7) minimal risk of post-harvest contamination, 8) resilience against environmental stresses, and 9) compatibility with IPM practices (Deravel et al. 2014; Kalpana 2021). Biopesticides may, however, not be equally effective against all pests, and their efficacy can vary depending on the target species. Additionally, some pests may have natural resistance or tolerance to certain biopesticides (Isman 2006).\n\nThe excessive reliance on conventional chemical pesticides in agriculture poses a significant threat to environmental health and sustainability, as the agriculture sector contributes a third of the total amount of greenhouse gas emissions. Climate Smart Agriculture (CSA) was developed as a framework to contribute to achieving the Sustainable Development Goals (SDGs) under an uncertain climate. Agricultural production systems need to simultaneously tackle three challenges, namely: 1) sustainably increasing agricultural productivity and incomes, 2) building resilience to the impacts of climate change by implementing biopesticide regimes and reducing conventional chemical pesticide use, and 3) contributing to climate change mitigation, where possible (FAO 2017).\n\nThe rising consumer demand for sustainably produced food, coupled with the growing resistance of pests to conventional chemical pesticides, the emergence of novel pest threats, and the impacts of climate change, underscores the increasing importance of biopesticides in the future of climate-smart agriculture systems (Chandler et al. 2008). To address this pressing issue, adopting environmentally safe and host-specific pest control options alongside IPM programs can create a winning combination for growers (Chang et al. 2003; Essiedu et al. 2020).\n\nMicrobial biopesticides are a broad range of pest control products obtained from microorganisms, including bacteria, viruses, fungi, protozoa, and entomopathogenic nematodes. Many microbial biopesticides can be cost-competitive with other pesticides and fungicides (Ruiu 2018). Microbial biopesticides are the most commonly used biopesticides in pest control programs (Kvakkestad et al. 2020). Most entomopathogenic bacteria belong to the Bacillaceae family, but the most widely used commercial bacterial biopesticides are from the genus Bacillus, especially the species Bacillus thuringiensis (Bt) and Bacillus subtilis (Bs) (Alsaedi et al. 2017; Akutse et al. 2020). These are aerobic bacteria that produce insecticidal crystal proteins that are toxic to specific insect orders (Gill and Cowles 1992; Raymond et al. 2009). Over 200 Bt-based biopesticides have been registered (Desneux et al. 2022) and are effective against various lepidopterans, coleopterans, dipterans, and hemipterans. However, resistance to some Bt products has been noted. For instance, laboratory resistance to Bt kurstaki has been reported for diamondback moth larvae (Liao et al. 2019), tobacco budworm, and beet armyworm (Blanco et al. 2009; Muhammad et al. 2019). More recently, western corn rootworms have shown field resistance to Bt (Legwaila et al. 2015). Actinomycetes - filamentous, aerobic bacteria that resemble fungi - are naturally found in soils and have also been developed into biopesticides (Pitterna et al. 2009). Some Actinomycete species can kill insects, including Tuta absoluta. Two species, Saccharopolyspora spinosa, and Streptomyces avermitilis, are used to make bacterial biopesticides (Akutse et al. 2020). Spinosad, a microbial biopesticide made from the aerobic fermentation of S. spinosa contains secondary metabolites called spinosyns (Higa 1994).\n\nMost entomopathogenic fungi belong to the two orders, Entomophthorales and Hypocreales (previously Hypomycetes) (Hajek and Leger 1994). The majority of entomophthoralean fungi are obligate parasites with limited host ranges and generate both sexual and asexual spores. Using epizootics, many of these species inhibit insects, the most prevalent genera being Entomophaga, Entomophthora, and Zoophthora (Goettel and Inglis 2000). Beauveria bassiana, Metarhizium, Isaria, and Verticillium spp. are fungi that infect and kill insect pests through direct contact or by penetrating their cuticle, following which the spores germinate and multiply in the insect’s body (de Faria and Wraight 2007; Kumari et al. 2022). Viruses from families of baculoviruses, cypoviruses, and densoviruses have been successfully used as viral biopesticides (Abd-Alla et al. 2020). For example, the nucleopolyhedro virus effectively infects and kills insect larvae (Cory 2000) and is used to control a variety of serious insect pests, especially moths (Gramkow et al. 2010). Insect viruses are considered environmentally safe due to their high selectivity and lack of non-target organisms species shifts. They also degrade naturally and do not persist in the environment (Abd-Alla et al. 2020). Despite their advantages, insect viruses face certain challenges as microbial biopesticide agents. These include their slow action, narrow host range, and potential for resistance development in insect hosts (Abd-Alla et al. 2020). Addressing these challenges through research and development efforts is crucial to fully realize the potential of insect viruses as sustainable pest control solutions.\n\nGenerally, microbialbiopesticides can have shorter shelf lives compared to chemical pesticides. They may also be sensitive to environmental factors such as sunlight, temperature, and moisture (McManus 1989; Matos et al. 2020; Wattimena and Latumahina 2021). However, if stored according to the label instructions, they could last up to 3-6 months (Akutse et al. 2020). There is a potential for microbial biopesticides to be integrated into IPM strategies in agriculture without significantly compromising productivity and yield (Aroraet al. 2020). However, several limitations currently hinder the widespread adoption of microbial biopesticides. These include 1) the high cost of refined commercial products, 2) policy and regulatory issues, 3) challenges in meeting global market demand, 4) lack of standardized preparation methods and guidelines, 5) difficulties in determining appropriate dosages of active ingredients, 6) susceptibility to environmental factors, 7) limited stability, and 8) slow adoption by end-users. While research breakthroughs are expected to address these limitations in the coming years, farmers, particularly those in rural areas, can still benefit from using microbial solutions as a means of plant protection to enhance crop quality and ensure farms are productive and profitable (Stevenson et al. 2017). It is important to note that the maximum effectiveness of biopesticide applications, including microbial biopesticides, is achieved when they are integrated into an overall IPM approach (Quarles 2013) and aligned to the 3 pillars of climate-smart agriculture (FAO 2017) (Figure 1).\n\nLegend: The visual mapping illustrates how the three pillars of Climate-Smart Agriculture (CSA) interact with and impact each of the 17 Sustainable Development Goals (SDGs). The second part demonstrates how the five steps for implementing CSA intersect with and affect the 17 SDGs in terms of synergies and trade-offs.\n\n\n2. Methods\n\nTo comprehensively gather information on biopesticides, we employed a systematic and rigorous approach. We utilized a wide range of electronic sources, including Google Scholar, PubMed, Scopus (Elsevier), Web of Science, Semantic Scholar, Academia, and other relevant websites, to conduct extensive literature searches. Our analysis of over 200 scientific papers and other relevant online resources enabled us to amass a comprehensive archive of pertinent literature. Our primary focus was on recent advancements in the development and application of biopesticides within the framework of IPM within the climate-smart agriculture framework. The literature outcomes were categorized into the (i) classification of biopesticides, (ii) importance of biopesticides in sustainable agriculture, (iii) role of biopesticides in integrated pest management, (iv) different types and formulations of biopesticides, and (v) advancement and future perspectives of biopesticides.\n\n\n3. Concept and classification of biopesticides\n\nBiopesticides are defined by the US Environmental Protection Agency (USEPA) as ‘naturally occurring substances, microorganisms, and plant-produced substances that control pests’(EPA. 2023). While the USEPA defines biopesticides as “pesticides derived from naturally occurring substances”, this term is not universally recognized. According to the Southern African Development Community (SADC), the term biopesticide is generally applied to a substance derived from nature, such as a microorganism or botanical or semiochemical, that may be formulated and applied like a conventional chemical pesticide and that is normally used for short-term pest control. The East African Community (EAC) also classified the term “biopesticides” as microbial, macrobial, botanical, and semiochemical biopesticides derived from or based on genetically modified organisms (GMOs) and pest control agents. The International Biocontrol Manufacturer’s Association (IBMA) and the International Organization for Biological Control (IOBC) prefer the term “biocontrol agents” (BCAs) instead. IBMA classifies BCAs into four groups, namely macrobiotics (predators and parasites), microbial (bacteria, fungi, and viruses that kill or harm pests), natural products (plant extracts), and semiochemicals (chemical signals) (Guillon 2003). The European Environmental Agency define biopesticide as “a pesticide in which the active ingredient is a virus, fungus, bacteria, or a natural product derived from a plant source”.\n\nBiopesticides offer several advantages over conventional chemical pesticides, including lower to no toxicity to non-target organisms, reduced environmental impacts, and less to no likelihood of developing resistance to pests. Biopesticides effectively control insects, diseases, weeds, and nematodes, promote plant health, and enhance the productivity and profitability of farmers. Importantly, many of them are non-toxic to beneficial organisms and wildlife. Additionally, their biodegradability reduces pollution concerns associated with many chemical pesticides (Shilpi and Promila 2012; Essiedu et al. 2022). Due to their targeted action and effectiveness in small quantities, biopesticides are gaining wide application on plants and crops. They are playing an increasingly significant role in agriculture, promoting sustainable practices and a healthier environment (EPA 2023), thus complying with the climate-smart agriculture principles (FAO 2017), and ultimately, the Paris Agreement, the Nagoya protocol of article 6 (Access to Genetic Resources) and article 7 (safeguards the rights of indigenous and local communities regarding their traditional knowledge and ensures they benefit from its use).\n\nHow do they work? Biopesticides work through various mechanisms, such as 1) biochemical disruption,-where they interfere with the biochemical processes of pests, affecting their growth, development, or reproduction; 2) microbial activity-infecting or parasitizing pests; 3) predation and competition-acting as natural predators or competitors of pests to limit their population growth; and 4) repellency-deterring pests from attacking crops (Essiedu et al. 2022). They can play a crucial role in IPM strategies and contribute to sustainable agricultural practices by reducing the environmental impact and minimizing the risk of pesticide resistance (Essiedu et al. 2022), thereby complying with the 3 pillars of climate-smart agriculture and the synergies with the SDGs (Figure 1) (FAO 2024).\n\n\n4. Importance of biopesticides in sustainable agriculture\n\nBiopesticides are an important tool in sustainable agriculture, and their use will likely continue growing. The use of biopesticides has steadily increased by a steady 11.0% Compound Annual Growth Rate (CAGR) between 2018 and 2022. The global biopesticides market reached a value of US$ 8,123.8 million in 2023. This upward trend is expected to continue, with the market expanding at a 10.3% CAGR and reaching US$ 21,827.6 million by 2033 (Persistence Market Research 2023). Food security is essential due to the world’s growing population. Humans and global economies depend on a stable and reliable food supply. As the population grows, food security will become more threatened (FAO 2024). Sustainable farming and maximizing soil fertility require non-toxic and environmentally sustainable pesticides. Thus, the need for biopesticides will likely increase over time (Bharti and Ibrahim 2020). Sustainable farming depends on biopesticides, which are poised to advance significantly in the coming years (Copping et al. 2000; Alavanja 2009). Optimizing manufacturing processes to increase yields and reduce costs can make biopesticides more economically viable. By using biopesticides, farmers can reduce their reliance on chemical pesticides, protect human health and the environment, and produce safe and healthy food (Hezakiel et al. 2023; FAO 2024).\n\n\n5. Role of biopesticides in integrated pest management\n\nBiopesticides play a vital role in IPM programs, which aim to control pests effectively whilst reducing the reliance on conventional chemical pesticides (Jozsef 2020). As the demand for more sustainable, environmentally safe, and sustainable agricultural practices grows, the use of biopesticides is becoming increasingly popular as part of IPM strategies. IPM aims to combine multiple pest control methods, including biopesticides, to minimize the use of chemical pesticides and maintain a balanced ecosystem in agricultural environments (Jozsef 2020). When used within IPM programs, biopesticides offer several advantages, including 1) targeted pest control, 2) diverse modes of action, 3) resistance management, 4) reduced negative environmental impact (Isman 2006), 5) sustainable crop production (Gurr et al. 2017), and 6) reduced risk of pesticide resistance in non-target organisms (Prasanna et al. 2018). Many biopesticides have the potential to contribute to economic growth and rural livelihood improvement, thus complying to the SDG’s (Figure 1).\n\nAdditionally, biopesticides offer potential benefits in the face of climate change challenges, and they align with the growing consumer demand for environmentally friendly and sustainable agricultural practices (FAO 2017; Akutse et al. 2020). Besides, biopesticides 1) generate fewer greenhouse gasses (GHG) than chemical pesticides, 2) improve nutrient availability for plants, 3) increase soil fertility, and 4) offer a more environmentally safe approach to pest control in agriculture, which can be useful in the fight against climate change.\n\nBiopesticide is a generic term generally applied to a substance derived from nature, such as a botanical or semiochemical, that may be formulated and applied like a conventional chemical pesticide and that is normally used for short-term pest control (USEPA 2020). Naturally derived or laboratory-made chemicals with structures and functions similar to naturally occurring ones are known as biochemical pesticides. They differ from traditional pesticides in their origin (source) and how they control or kill pests (O’Brien and Jones 2009). Microbial pesticides are natural pest control agents harnessing the power of microscopic living organisms (Clemson 2007). These “living bullets” can be either spores or active organisms, often specifically chosen for their pathogenic nature towards targeted pests. Common examples include biofungicides (Trichoderma, Pseudomonas, Bacillus, Colletotrichum), bioherbicides (Phytophthora, Cylindrobasidium, Colletotrichum acutatum), and bioinsecticides (Bt) (MacGregor 2006; Gupta 2010). Microbial biopesticides can originate from naturally occurring or genetically modified bacteria, fungi, algae, viruses, or protozoans. They subdue pests through a diverse range of strategies (toxic metabolites, disease-causing, competitive exclusion, and other modes of action), depending on the specific microbe (Clemson 2007).\n\nMicrobial biopesticides offer diverse delivery methods to effectively reach crop targets. These methods include live organisms, dead organisms, and spores (O’Brien and Jones 2009). Globally, microbials account for 41% of all BCAs used worldwide, macrobiotics comprise 33% of the global market, and other natural products represent the remaining 26% (Guillon 2003). Based on their origin or active ingredients, they can be classified as bacterial-based biopesticides, fungal biopesticides, viral biopesticides, nematodes, and protozoan-based biopesticides.\n\nAmong all microbial pesticides, bacterial biopesticides are the most widely used. These versatile microbes attack pests in multiple ways, primarily targeting insects. While mostly used against various orders of insect pests in agriculture, bacterial biopesticides can also curb the growth of plant-harming bacteria and fungi (O’Brien and Jones 2009; Jouzani and Valijanian 2017). Bacterial pathogens need direct contact with their target to be effective, often requiring ingestion. They secrete endotoxins, protein-based toxins specific to the targeted pest that wreak havoc on their digestive system (Schallmey and Singh 2004). Bacillus thuringiensis (Bt) dominates the biopesticide market, accounting for a whopping 90% share in the USA alone (Chattopadhyay and Bhatnagar 2004). Bacillus thuringiensis is the most common species used commercially to control a diverse range of insect pest species, including lepidopterans, hemipterans, and coleopterans in agriculture, forestry, and even medicine since its discovery in 1901 (Mazid 2011). It is a spore-forming bacterium with several strains that produce diverse insecticidal crystal proteins, often targeting different pests (Table 1). The crystal proteins are toxic and, once ingested, damage the gut tissues, leading to paralysis of the gut. The infected insects stop feeding and eventually die from gut impairment and starvation. Bt var. kurstaki specifically targets caterpillars, while the genes encoding Cry proteins have been transferred into crops like cotton, hence reducing reliance on chemical pesticides (Mazid 2011). Commercial Bt-based products are available in various forms, such as powders containing a combination of dried spore and crystal toxins or formulations in liquid suspensions (Boyetchko et al. 2020). Bacillus thuringiensis’s high target specificity and environmental safety make it an ideal complement to traditional chemical pesticides for insect pest control (Roy and Moktan 2007; Kumar 2012). Figure 2 illustrates the effects of Bacillus thuringiensis on insect larvae.\n\nLegend: The mode of action of Bt gene and cry protein.\n\nRecombinant DNA technology has been used to develop several new Bt-based products. MVPTM and M-TrackrTM, for example, were created using Mycogen Corporation’s CellCap® encapsulation method. This procedure entails removing a gene from Bt that encodes the delta-endotoxin protein, incorporating it onto a plasmid, and inserting it into a Pseudomonas fluorescens strain (Pitterna et al. 2009; Legwaila et al. 2015). Before being killed by heat and chemical treatment, the recombinant cells were cultured in aerobic culture and induced to express delta-endotoxin. The dead bacterial cells in the aqueous formulation acted as microcapsules, protecting the fragile Bt toxin from destruction in the environment.\n\nPathogenic bacteria have also been considered for biological weed control. However, one of the challenges to using phytopathogenic bacteria for biological weed control is the necessity of free water for dispersal and wounds or natural holes for bacterial penetration into the weed plant (Zidack et al. 1992; Johnson et al. 1996). Previously, researchers have studied Xanthomonas campestns pv. Poae for the control of annual bluegrass (Pea annua L.) and discovered that cutting or mowing turfgrass allows bacteria to enter the plant (Imaizumi et al. 1997). Furthermore, bacteria administered at a rate of log CFU/mL at high water levels (400 mL/m2) reduced disease severity in annual bluegrass by more than 90%. Silwet L-77 (0.2%), an organosilicon surfactant, increased bacterial penetration and entrance into plant stomata and hydathodes (Johnson et al.1996; Boyetchko et al. 2020). A low surface tension of 30 dynes/cm or less is necessary to convey liquid into a leaf’s stomata. Silwet decreases the surface tension of water to 20 dynes/cm. Compared to plants sprayed with bacteria without the surfactant, the application of Pseudomonas syringae pv. Tagetes with this surfactant facilitated the penetration and entry of bacteria into stomata and hydathodes, significantly increasing disease severity and incidence (Johnson et al. 1996).\n\nBacterial biopesticides are environmentally safe and precise alternatives to traditional pest control methods. They offer several advantages, including safety for humans, wildlife, and beneficial insects. These biopesticides can be used in conjunction with other pest control strategies, including some chemical methods. Additionally, they leave no harmful residues, don’t harm pollinators and other valuable organisms, and can be applied close to harvest time. The additional benefit is that they can self-propagate, extending their effectiveness beyond the initial application and into subsequent growing seasons (Meenatchi and Aditi 2021). Bacterial biopesticides, while effective, come with a few disadvantages. For instance, they only target specific species or groups of insects, leaving other pests to survive and continue causing damage. This means that additional biopesticides are needed for the other pests. Hence, they should be applied alongside other IPM strategies. Finally, the effectiveness of microbial insecticides can be influenced by factors like ultraviolet radiation and heat, so it’s important to apply them strictly according to the label (Meenatchi and Aditi 2021).\n\nMycopesticides include naturally occurring fungi and fungi cell components. These fungi attach to their targets through sticky spores. These spores then germinate and form microscopic tubes that pierce the insect’s skin, releasing potent hydrolytic enzyme mixtures along with toxins (Langner and Göhre, 2016). This internal invasion disrupts the insect’s physiology, leading to its eventual death. The fungi then grow outward from the dead insect, producing new spores to continue the cycle, hence perpetuating natural pest control (Pineda et al. 2007; Gabarty et al. 2014). The role of hydrolytic enzymes, especially chitinases, in the killing process, and the possible use of chitin synthesis inhibitors are prime research areas. Popular commercially available mycoinsecticides are often derived from species like Beauveria bassiana, Metarhizium anisopliae, Isaria fumosorosea, Hirsutella thompsonii, Phytophthora palmivora, Alternaria cassia, and Lecanicillium spp. (Table 1). Notably, Beauveria bassiana and Metarhizium anisopliae are two common ascomycetes known for their broad effectiveness against pests like aphids, beetles, grasshoppers, and caterpillars. These fungi are typically applied as conidia (spores) or mycelium, which then sporulate after application, ensuring sustained pest control (Lacey et al. 2015). Cylindrobasilium laeve for the control of Acacia mearnsii (black wattle) and Colletotrichum acutatum for the control of Hakea sericea (silky hakea) are some of the available mycoherbicides for integrated alien and invasive species management.\n\nFungal biopesticides hold immense promise as environmentally safe options against a diverse range of insect and mite pests. They possess several desirable traits for biocontrol: they selectively target pests, leaving beneficial insects like bees and predators unharmed. Additionally, they generally pose no threat to the growth or development of other beneficial organisms like earthworms and springtails. This makes them ideal candidates for IPM and sustainable agricultural practices, promoting biodiversity and protecting the environment (Goettel et al. 2008; Kim and Goettel 2011; Koike et al. 2011). Several fungi have been successfully mass-produced and formulated for commercial application as mycoinsecticides, paving the way for their widespread use in pest control (Chandler et al. 2008).\n\nThe remarkable potential of entomopathogenic fungi is actively being harnessed for use in IPM programs. Mass production techniques and ecological approaches are constantly refined to optimize their effectiveness and ensure seamless integration into sustainable agricultural practices (Chandler et al. 2011; Lacey et al. 2015). Furthermore, promising results have been achieved by combining fungal biopesticides with traditional insecticides, often leading to significantly boosted pest mortality rates. For example, studies have shown that combining B. bassiana with low-dose insecticides enhances potato beetle control, while its synergy with neem oil proves effective against tobacco thrips eggs and nymphs (Chandler et al. 2011; Lacey et al. 2015; Sarwar 2015).\n\nFungi-based biopesticides boast several advantages over other biocontrol methods. Their broader host range allows them to tackle a diverse range of pests across fields, greenhouses, storage facilities, and soil, unlike bacteria and viruses with limited targets (Amruta et al. 2019). Commercially important fungi like Beauveria, Metarhizium, Lecanicillium, and Isaria are surprisingly easy to mass produce, requiring significantly less substrate compared to alternative methods. These fungi are highly productive, delivering powerful, targeted pest control while reducing the impact on the environment. They pose no risk to beneficial organisms, readily biodegrade, and integrate seamlessly into IPM programs. Additionally, their persistence in the environment offers long-lasting protection against pest outbreaks, making them a potentially cost-effective solution for sustainable agriculture (Naveenkumar et al. 2017; Rajaput et al. 2019; Sufyan et al. 2020; Upamanya and Bhattacharyya 2020).\n\nFungi-based biopesticides however face some environmental hurdles. Optimal spore germination and penetration into insect cuticles often require humidity levels exceeding 80%, depending on the specific fungal species. Temperature fluctuations and exposure to UV radiation can significantly impact their survival. Germinating conidia, essential for active pest control, are often delicate and susceptible to environmental damage. Conversely, the production of durable resting spores for long-term persistence can be expensive compared to other microbial methods like bacteria-based biopesticides (Sabaratnam 2002; Kolombet et al. 2008; Kordali et al. 2008; Li et al. 2014). Figure 3 illustrates the mode of action of fungi-based biopesticides, highlighting the steps by which these biopesticides attack target insect pests (Figure 3).\n\nLegends are indicated in the figures. No additional legends.\n\nBaculoviruses (BVs) are tiny, naturally occurring viruses that have emerged as powerful allies in the fight against insect pests. They belong to a group known as entomopathogenic viruses, specifically targeting and infecting insects and other arthropods. Unlike harmful conventional chemical pesticides, baculoviruses pose no threat to humans, wildlife, or beneficial insects, making them an environmentally safe option for pest control (Cory 2000; Jackson and Crawford 2005; Ramle et al. 2005; Moscardi et al. 2011), and are considered as the most commercial viral biopesticides (Granados 1986; Moore and King 1987). Baculoviruses involve circular supercoiled double-stranded DNA genomes, which range from 80 to 180 kbp (Rohrmann 2013). Within the baculovirus family, scientists have identified three main subgroups: nuclear polyhedrosis viruses (NPVs), cytoplasmic polyhedrosis viruses (CPVs), and granulosis viruses (GVs). These subgroups differ in the structure and number of their protective protein coats, called occlusion bodies (Moscardi et al. 2011). These bodies allow the virus to survive outside the host, waiting for its next target. NPVs and CPVs form polyhedral bodies with numerous virus particles, while GVs have smaller, granular bodies containing just one particle. Notably, each subgroup employs a unique method to uncoat and establish infection within their host: NPVs in the nucleus, CPVs in the cytoplasm, and GVs within the nuclear pore complex (Cory 2000; Moscardi et al. 2011).\n\nBaculoviruses offer highly specific targeting, primarily focused on lepidopteran larvae and hymenopterans (butterfly and moth) pests that damage crops like cotton, rice, and vegetables. One example is Heliothis zea nucleopolyhedrosis virus (HzNPV), the first commercially successful broad-spectrum viral insecticide. Interestingly, HzNPV demonstrates versatility, effectively controlling pests across various crops like soybeans, sorghum, maize, tomatoes, and beans. While other entomopathogenic viruses exist, such as tetraviruses and cypoviruses, their use in crop protection remains limited compared to the widespread adoption of baculoviruses (Sarwar 2015). A comprehensive summary of viral biopesticides and their target pests is presented by (Usta 2013; Singh et al. 2019; Meenatchi and Aditi 2021).\n\nFigure 4 shows how viral replication and infection of the target host is attacked. After infecting a target cell, viral replication in the nucleus or cytoplasm unfolds in three distinct phases: early (0-6 hours), second (6-24 hours), and very late (24-72 hours). During the late phase, the virus forms protective protein coats called occlusion bodies (Obs) or virions. These tiny packages containing multiple virus particles can lead to natural outbreaks (enzootics) that drastically reduce pest populations. However, sunlight poses a challenge: occlusion bodies can rapidly lose potency when exposed to ultraviolet radiation (UV) between 280-320 nm (Killick 1990). Interestingly, studies show that plastic greenhouses can help mitigate this issue by filtering out over 90% of harmful UV-B rays, boosting infection rates in larvae (Lasa et al. 2007).\n\nLegends: The virions enter the midgut cell nucleus, at which point the virus replicates within the nuclei of susceptible tissue cells and tissue susceptibility varies greatly between viruses with some NPVs being capable of infecting almost all tissue types and most GVs being tissue-specific replications. In the terminal stage of infection, the insect liquefies and thus releases polyhedral, which can infect other insects upon ingestion. A single caterpillar at its death may contain over 109 occlusion bodies from an initial dose of 1000. Under optimal conditions, target pests may be killed in 3-7 days, but when the condition is not suitable, death may be caused in 3-4 weeks (Kalamkoff 2004).\n\nFigure 4. Mode of action of viral biopesticides (Singh et al. 2019). The virions enter the midgut cell nucleus, at which point the virus replicates within the nuclei of susceptible tissue cells, and tissue susceptibility varies greatly between viruses, with some NPVs being capable of infecting almost all tissue types and most GVs being tissue-specific replications. In the terminal stage of infection, the insect liquefies and thus releases polyhedral, which can infect other insects upon ingestion. A single caterpillar at its death may contain over 109 occlusion bodies from an initial dose of 1000. Under optimal conditions, target pests may be killed in 3-7 days, but when the condition is not suitable, death may be caused in 3-4 weeks (Kalamkoff 2004).\n\nSeveral studies have been conducted to explore ways to improve the effectiveness of viral biopesticides. Certain formulations, like stilbene, can enhance susceptibility to NPV infection by disrupting the insect’s digestive barrier (Okuno et al. 2003) or inducing cell death in the midgut (Dougherty et al. 2006). Additionally, researchers are investigating genetically engineered viruses. For example, vAcTaITX-1 and vAcDTX9.2, derived from specific spiders, show promise as commercial biopesticides against lepidopteran pests (Hughes et al. 1997). While viral biopesticides offer numerous advantages over conventional chemical pesticides, including specificity and environmental safety, they face certain challenges for large-scale adoption. Producing them efficiently, particularly recombinant viruses, can be costly and labor-intensive, requiring specialized equipment and lengthy procedures (Gupta 2010; Lacey et al. 2015). Despite these hurdles, organizations like IPM centers and state agricultural departments are actively supporting the development and small-scale production of these promising biopesticides.\n\nWhat is the main advantage of viral biopesticides? Viral biopesticides offer a safer alternative, protecting human health, wildlife, and even beneficial insects. These targeted pathogens would not trigger resistance in pests, making them a lasting solution. Moreover, they seamlessly work with other pest control methods for a comprehensive defense (Singh 2019; Meenatchi and Aditi 2021). While viral biopesticides offer enticing, environmentally safe alternatives to chemical pesticides, they come with limitations. Unlike the broad spectrum of some conventional chemical insecticides, their use often remains targeted to specific pest species. Additionally, successful pest control can be a slow process, as infections may take a significant amount of time to become lethal and can be easily deactivated by high temperatures and ultraviolet radiation. Understanding these vulnerabilities is crucial for choosing the right pest control strategy (Singh 2019; Meenatchi and Aditi 2021) for the targeted pest and bioclimatic zone.\n\nEntomopathogenic nematodes (EPNs) are classed as microbial pesticides, even though they are multicellular. EPNs are tiny soil-dwelling organisms that live in the water films around soil particles, fit nicely into IPM programs because they are considered nontoxic to humans, relatively specific to their target pests, and can be applied with standard pesticide equipment (Shapiro-Ilan et al. 2006). These beneficial parasites help maintain the balance in the ecosystem by targeting specific pests without harming other organisms (Kachhawa 2017). Two main families of EPNs, Steinernematidae, and Heterorhabditidae, have been enlisted in the fight against pests (Bhat and Chaubey 2020). Species like S. carpocapsae, S. thermophilum, H. bacteriophora, and H. indica are commercially available and actively deployed in pest control programs in India (Meenatchi and Aditi 2021). Commercial biopesticides based on EPNs contain non-feeding, third-stage infective juveniles (IJs) as the active ingredients. Heterorhabditis and Steinernema are mutualistically associated with bacteria of the genera Photorhabdus and Xenorhabdus, respectively (Ferreira 2014).\n\nSo, how do these tiny organisms take down their targets? EPNs, in their infective juvenile stage, barely millimeters long, use their keen senses to locate insects through carbon dioxide emissions, vibrations, and other chemical cues. They then wriggle their way into the insect’s body through natural openings like the mouth, anus, or breathing holes. Once inside, the nematodes release their symbiotic bacterial companions, setting off a natural pest control cascade. These bacteria produce toxins that quickly kill the insect within a week. The EPNs then feast on the insect’s liquefied remains, multiplying within the host to create new generations of tiny warriors. Finally, once their feast is over, the next generation of infective juveniles emerges, ready to seek out their next victim and continue the cycle (Figure 5). Under ideal conditions, the impact of EPNs becomes visible within 5-7 days. Look for browning or tanning of insects infected by Steinernematidae, while those taken down by Heterorhabditidae turn a distinctive red (Gupta 2010; Koul 2011; Ruiu 2018).\n\nLegends: The juvenile stage releases cells of their symbiotic bacteria from their intestines into the hemocoel. The bacteria multiply in the insect hemolymph, and the infected host usually dies within 24 to 48 hours. After the death of the host, nematodes continue to feed on the host tissue, mature, and reproduce (Bedding 1982). The progeny nematodes develop through four juvenile stages to the adult. Depending on the available resources, one or more generations may occur within the host cadaver, and a large number of infective juveniles are eventually released into the environment to infect other hosts and continue their life cycle (Bedding 1982). Nematodes enter the body cavity of insects and release their symbiotic bacteria into the host's intestine.\n\nAdvantages of EPN: EPNs offer a double win: safety and effectiveness. These gentle warriors pose no threats to human health, plants, or animals, requiring no protective gear or waiting periods. Moreover, they leave no harmful residues, making them a sustainable choice for healthy crops and clean environments. Their target list reads like a bug buffet: cranberry girdlers, root weevils, webworms, and even wood borers tremble at their approach. However, these tiny heroes have their preferences. Moist soil around roots, protection from harsh UV, and short stints with other pest control methods are necessary for them to thrive, ensuring their targeted pest control doesn’t disrupt the natural ecosystem (Ruiu 2018; Singh 2019; Meenatchi and Aditi 2021).\n\nThe disadvantage of EPN: EPNs aren’t weather warriors. Harsh UV rays and sizzling temperatures can quickly deactivate these tiny heroes, making their effectiveness dependent on fickle environmental conditions. Compared to their chemical counterparts, EPNs come with a hefty price tag. This, along with their short shelf life, make them a less financially enticing option for some applications. EPNs can’t reach aerial pests, limiting their range of action. Additionally, relying solely on EPNs can disrupt the natural predator-prey balance, requiring repeated applications to maintain pest control. Furthermore, EPNs thrive in moisture. Pre- and post-irrigation, along with application during cooler hours, are essential for their success (Meenatchi and Aditi 2021).\n\nProtozoans, also known as microsporidians, are intracellular parasites that can only survive by living inside other cells. They are found almost everywhere and can attack certain insects, like lepidopteran and orthopteran, making them useful in IPMs. Examples include Nosema sp. and Vairimorpha sp. However, despite their pest-specific nature and slow-acting properties, the use of protozoa as biopesticides is not as effective as other organisms, such as bacteria, viruses, and fungi. They can cause chronic and debilitating effects on their targets, but their success rate is lower compared to other biopesticides (Meenatchi and Aditi 2021). Protozoans have a specific mode of action. Microsporidia infects the European corn borer, Ostrinia nubilalis, by being eaten by insects. The spores germinate in the midgut region, and the sporoplasm is then injected into the midgut cells. The spores then spread to different tissues and organs, multiply, and cause tissue breakdown and septicaemia (Senthil-Nathan 2015).\n\n\n6. Natural enemies of pests as biopesticides\n\nTwo powerful allies in this environmentally safe pest control approach are parasitoids and predators.\n\nParasitoids: Like tiny assassins, these insects specialize in laying their eggs on or inside other insects. Their young hatch and feast on the host’s body from the inside out, eventually leading to the host’s demise. Think of them as microscopic ninjas, working silently to take down their targets. Examples include wasps that parasitize caterpillars and flies that target beetle larvae.\n\nParasitoids have a shorter life cycle than predators and can multiply at a faster rate. Therefore, they can be more effective in controlling insects. However, their presence may not be noticeable unless you examine samples of the insects to see if any adult parasitoids have emerged (Yelitza et al. 2020).\n\nPredators: These are your classic insect warriors, actively hunting and devouring their prey. They’re often larger than their victims and use their speed, agility, and sharp senses to track them down. Ladybugs munch on aphids, dragonflies swoop on mosquitoes, and ground beetles patrol the soil for tasty treats. Introducing these natural predators into your environment, creates a pest-fighting force on patrol. Figure 6 illustrates the mode of action of natural enemies in pest control, specifically focusing on A) the mode of action of parasitoids, and B) natural predators, including lady beetles (Coccinellidae), wasps, and fungus gnats.\n\nA) Mode of action of parasitoids, and B) Natural predators (the insects are Lady beetles, (Coccinellidae), wasps, and fungus gnats, respectively).\n\nSeveral promising biopesticides that are effective against various crop pests have been summarized by (Meenatchi and Aditi 2021) and (Gautam 2008). These specialized assassins are laser-guided for specific pests, efficiently hunting them even at low levels. They curb reproduction, keep infestations below damage thresholds, and play well with other pest-fighting teams. Their added benefit is that some disrupt pest feeding and reduce chomping on your precious plants. They are environmentally safe pest control solutions (Gautam 2008).\n\n\n7. Biopesticide formulations\n\nBiopesticide formulations, like conventional pesticide formulations, involve preparing a product from an active ingredient (biologically active metabolite or microbe) by adding various functional (active) and inert substances. This creates a product suitable for application to the target weed, pathogen, or insect using existing equipment. The active ingredient is combined with a carrier material and various additives to improve the biopesticide’s lifespan and efficacy (Grewal and Peters 2005). Several factors significantly influence the economic viability of biopesticide products. These include the impact on the target pest, market size, pest spectrum, field performance consistency, production costs, and technological challenges (fermentation, formulation, and delivery systems), all of which pose potential hurdles to commercialization. Optimizing product formulations can significantly improve field performance consistency. This is crucial for the successful adoption and economic viability of many biopesticides (Glass 1993). However, slow progress in research on formulation and delivery systems remains a major bottleneck for biopesticide development (Lumsden et al. 1995).\n\nThe formulation is arguably the most crucial step in the development of biopesticides and is the cornerstone of product development (Leggett et al. 2016). Ideally, a user-friendly formulation must meet several important criteria, including preserving and expressing the pesticidal properties of the microorganism, extending the shelf life to at least six months and preferably up to two years under ambient conditions, and enabling application using existing equipment (Pusey 1994). Moreover, enhanced formulations can improve the efficacy of biopesticides by boosting their dispersion, attachment, and persistence at the target site (Droby et al. 2009). Stickers improve adherence to foliage, enhancing persistence (Schisler et al. 2004). Formulation stabilizes the biopesticide throughout production, distribution, storage, handling, and application to ensure persistence and activity at the target site. Cognizance must be given to ensure that the co-formulants used in biopesticide formulations are not chemical pesticide derivatives, are environmentally safe, and do not impact human health.\n\nThe effectiveness of a biopesticide hinges on the biological activity of its active microorganism or metabolite. In its raw state, this active ingredient requires formulation to enable its practical handling, storage, and application (Harald 2019). A crucial distinction between conventional (chemical) pesticide formulation and biopesticide formulation lies in the biopesticide’s living nature. Biopesticides are sensitive to storage conditions and environmental factors, necessitating careful formulation to preserve their biological viability (Harald 2019). Formulating biopesticides faces several challenges, including ensuring market viability, ease of production and application, product stability during storage and transport, and long-term viability and efficacy of the biopesticide (Lumsden et al. 1995). Contributing factors to their limited commercial success include production difficulties, sensitivity to environmental stressors, and lack of appropriate formulation (Powell and Jutsum 1993). Another challenge is the slime layer produced by gram-positive bacteria like Bacillus spp. during endospore formation. This slime can clog sprayer filters and nozzles, hindering application. While removing the slime through methods like centrifugation is possible, it may also remove valuable components contributing to the biopesticide’s effect (Boyette et al. 1996).\n\nThe goal of formulation is to improve product stability, bioactivity, delivery, and integration into pest management systems. Successful formulations also prioritize user convenience, compatibility with equipment and practices, and effectiveness at practical rates. Environmental factors like temperature, moisture, and UV exposure significantly influence efficacy, particularly for foliar-applied biopesticides (Alan 1988). Soil-applied biopesticides are impacted by soil properties, moisture, temperature regimes, and microbial competition. All these factors need careful consideration during formulation development.\n\n7.2.1 Formulation of bacteria biopesticide\n\nSeveral recent studies highlight the growing use of bacteria-based biofungicide formulations as a safe and effective complement to conventional chemical pesticides (Cook 1993; Lumsden et al. 1995; Elad 1995). Bacillus-based products are already boosting crop yields in various countries, from China to the United States (Zhang et al. 1996). These wettable powders, compatible with existing fungicides, target diseases caused by Rhizoctonia and Fusarium in a range of crops like cotton, legumes, vegetables, and ornamentals.\n\nMass production of these biofungicide bacteria formulations typically involves deep-tank liquid fermentation. However, in some cases, methods like sem.isolid or solid-state fermentation might be more suitable. Key factors influencing both bacterial growth and their beneficial metabolite production include the medium’s nutrient composition and optimal growth conditions (Paau 1988). Cost-effectiveness and readily available components are prioritized. The final formulation, usually involving additional ingredients after fermentation, comes in various forms like solids, slurries, powders, or granules. Long-term viability during transport and storage (at least 4 months) is crucial (Paau 1988). Formulation significantly impacts the biofungicide’s performance by enhancing bacterial survival after application. This involves creating a protective environment that maximizes the bacteria’s potential for successful colonization and disease control. Choosing the right formulation can help ensure consistent results in the field.\n\nThe products are designed as wettable powders and can be used with various seed treatment fungicides (Table 1). Agrobactenum tumefaciens is available commercially in Australia, the United States, and New Zealand and is formulated as a concentrated liquid or moist peat-based product or supplied as a non-formulated agar culture (Kerr 1980; Elad 1995). After being suspended in water, the bacteria can be applied to the seeds, cuttings, roots, or root wounds of susceptible orchards and ornamental plants as a dip, spray, or drench. Agrobacterium tumefaciens, available as a liquid, peat-based product, or agar culture, combats diseases in orchards and ornamental plants through dips, sprays, and drenches. Streptomyces griseoviridis (Mycostop), formulated as a wettable powder, tackles damping-off and root rot in vegetables and ornamentals caused by Fusarium, Phomopsis, and Pythium. It can be applied dry to seeds or as a liquid drench and is compatible with chemical pesticides (Muir 2023). Three Burkholderia assian strains (Blue Circle, Deny, and Intercept) come in liquid and fight a wider range of enemies. These bacterial warriors target fungi like Fusarium, Phytophthora, and Pythium alongside nematodes, including Globodera rostochensis, Heterodera glycines, and Hoplolaimus columbus (Kumari et al. 2022).\n\n7.2.2 Formulation of bacterial bioinsecticides\n\nMost commercial bioinsecticide formulations today harness the power of Bacillus thuringiensis (Bt), a friendly, spore-forming bacterium. This gram-positive warrior wields a powerful weapon: delta-endotoxin proteins that wreak havoc on the mid-guts of susceptible insect larvae (Cannon 1993) (Table 1). Bt proteins are highly specific, only affecting a few species in the Lepidoptera (butterflies and moths), Coleoptera (beetles), and Diptera (flies) families. Bt biopesticides come in an array of formulations, from concentrated liquids to handy dusts, catering to different application needs.\n\n7.3.2 Formulation of fungal biopesticides\n\nFungicide formulations primarily include filamentous fungi (e.g. Gliocladium virens and Trichoderma harzzanum), yeast-like fungi (e.g. Pseudozymajlocculosa (Bélanger 1997) and Tllletiopsis pallescens (Urquhart and Punja 1997). These biofungicides are used to control root-infecting pathogens (e.g. Pythlum, Rhizoctonia), and foliar fungal pathogens (e.g. powdery mildew (Bélanger 1997; Urquhart and Punja 1997; Punja 1997) and Botrytzs (Elad 1995). Environmental conditions, such as temperature and moisture, can impact the growth and survival of fungal biofungicides. Various formulations have been developed for the application of spore inocula, including granules, pellets, dust, and wettable powders. These formulations can be applied directly or suspended in water or oil, as is the case with Cylindobasidium leave (Muir 2023).\n\nGranular formulations not only protect against desiccation but also offer a food base for the fungus. Meanwhile, powders can be easily sprayed and can provide coverage for large areas. An example of this is the large-scale aerial application of sawdust covered in Colletotrichum acutatum spores for the control of Hakea sericea (Jacobson and Azevedo 2023). It is also possible to treat seeds with liquids or dust for the application of these biopesticides. Moreover, spore formulations in inverted emulsions have been tested for yeasts, such as Tilletiopsis (Urquhart and Punja 1997).\n\nThe use of alginate prill has been successfully developed to create a granular formulation of Gliocladium virens (Soil Gard), which helps control root-infecting fungi in potting media (Lumsden et al. 1995). Similarly, Trichoderma powder or dust formulations with pyrophyllite clay (Pyrax) have also been proven to be effective (Jayaraj and Radhakrishnan 2006). To produce the necessary biomass, appropriate nutrient substrates are used in large-scale deep tank fermenters. The resulting product can either be used wet or dried before formulation (Lumsden et al. 1995). Most of the factors that affect product development for fungi are like those for bacteria.\n\nEntomopathogenic fungi are a fascinating group of soil-dwelling microorganisms that play a crucial role in biological control. These fungi are capable of infecting and killing insects through cuticle penetration. They don’t need to be ingested by the insect hosts; instead, they directly invade through the insect’s outer covering. This unique mode of action allows them to control a wide range of insect pests, including sucking insects. However, they cannot effectively combat viruses and bacteria similarly (Shishupala 2022). Entomopathogenic fungi are a type of fungi that can target and control pests without harming beneficial organisms such as pollinators. They are an environmentally safe alternative to traditional chemical pesticides for managing agricultural pests. Some of the most well-researched fungal species with bioinsecticidal properties include Akanthomyces muscarius, Beauveria bassiana, Cordyceps fumosorosea, Purpureocillium lilacinum, Verticillium, and the Metarhizium anisopliae species complex (Shishupala 2022). A recent study by Luo et al. (2022) isolated a new entomopathogenic fungus from an adult cadaver of D. citri. The fungus was identified as Cordyceps fumosorosea based on its morphology and ITS sequence analysis. The researchers named this specific isolated C. fumosorosea SCAU-CFDC01. The study assessed the pathogenicity of the strain against D. citri nymphs and adults in both laboratory and greenhouse settings.\n\nVarious fungi can be used to control different types of insects. Verticillium lecanil can control aphids, Beauveria bassiana can control whiteflies, locusts, and beetles, Metarhizium anisopliae and M. flavoviride can control mosquitos and locusts respectively, while Lagenidium gigantem can control mosquito larvae (Lacey and Goettel 1995; Bateman 1997; FAO 2017; Savita 2019). These fungi can be applied to insects in different ways, such as wettable powders, emulsions, specks of dust, baits, or traps, or even added to the soil (Auld 1992; Feng et al. 1994; Goettel et al. 1995; Inglis et al. 1996) or used in indoor residual spraying. It is important to formulate these fungi to protect against environmental conditions such as moisture, temperature, UV damage, and desiccation. For example, B. bassiana conidia on foliage can be damaged by UV-B radiation, which is part of the sunlight spectrum (Caudwell and Gatehouse 1996a; Daoust and Pereirn 1986). In field conditions, these entomopathogens can be applied at ultralow volumes (ULV) to increase their effectiveness and protect against UV damage (McGUIRE and Shasha 1992; Moore et al. 1993; Inglis et al. 1996). To improve survival and shelf-life, sunlight blockers (such as clay) or UV-B absorbing compounds (such as Tinopal) can be added to inoculum formulations or starch encapsulation (Pereira and Roberts 1991; McGuire and Shasha 1992; Caudwell and Gatehouse 1996b).\n\n7.3.3 Mycoherbicides\n\nEnvironmental factors, such as temperature and moisture, can greatly impact the effectiveness of Mycoherbicides. The moisture needed for the disease to develop is often determined by the amount of dew that occurs. “DeVine” is the first registered Mycoherbicide, which is a liquid formulation consisting of chlamydospores of Phytophthora palmivora (Burnett et al. 1974; TeBeest and Templeton 1985). This product has a limited shelf life of only 6 weeks when refrigerated due to its instability. Another Mycoherbicide called “Collego” (Colletotrichum gloeosporioides f.sp. aeschynomene) is formulated as dried spores and is available in a wettable powder (Boyette 1994). The commercialized formulation of Colletotrichum acutatum is formulated as dried spores that are mixed with water when ready to apply, and Stumpout is Cylindrobasidium laeve formulated as dried spores in an iron carrier that is mixed with standard cooking oil when ready to apply (Muir 2023).\n\nThere are several ways to improve the effectiveness of mycoherbicides, which are used to control weeds (Boyette et al. 1996). One way is to use adjuvants and other additives that can help spores grow, make the pathogen more stable, change the environment, or make it possible to use bioherbicides on more plants (Boyette 1994). For example, a fungus called Colletotrichum truncatum can kill a weed called hemp sesbania, but it needs a lot of moisture to work (Boyette et al. 1993). Adding unrefined corn oil to the biocontrol agent can help it work better, so less moisture is needed. This makes it easier to use the bioherbicide and reduces the amount of spray needed (Boyette 1994).\n\nSurfactants are ingredients used in formulations to help wet plants by reducing surface tension and improving the dispersal of fungal spores in spray droplets. Some surfactants used include Tween 20 with Fusarlum lateritum, nonoxynol with Alternaria macrospora and A. assia, and sorbitol with Colletotrichum coccodes (Boyette 1994). However, evaluating the inhibitory or stimulatory effects on spore-germination and infection is important before selecting appropriate surfactants and ensuring that the surfactants do not contain pesticide derivatives. Water-in-oil invert emulsions have been used with foliar fungal biopesticides to provide a favorable environment for germination and infection (Connick et al. 1990; Daigle et al. 1990; Auld and Morin 1993; Boyette et al. 1996). Using inverted emulsions (Boyette et al. 1993) has been found significantly to improve the efficacy of C. truncatum and Alternaria assia. However, inverted emulsions are very viscous and may demonstrate phytotoxicity in some target plants (Walker and Boyette 1985; Amsellem et al. 1990; Goettel et al. 1995). Connick et al. (1990) developed an inverted emulsion with improved water-retention properties that was less viscous. Additionally, vegetable oils can enhance the efficacy of mycoherbicides, such as Colletotrichum orbiculare, for the control of spiny cocklebur (Auld 1992) and Collototrichum acutatum for the control of Hakea sericea and H. gibbose. No phytotoxicity and improvements in the spread of the inverted emulsion were observed.\n\nSolid-based formulations of mycoherbicides have been developed for weeds that are infected at or below the soil surface, which is more suitable for preemergence mycoherbicides (Daigle and Connick 1990; Boyette et al. 1996). These formulations can provide a food base for the pathogen, act as a buffer in extreme environmental conditions, and retain inoculum that may not be easily washed away. One such formulation, known as “PESTA,” uses a wheat-gluten matrix consisting of liquid inoculum, semolina wheat flour, and kaolin. This formulation includes fungal agents like C. truncatum, A. crassa, and Fusarium lateritium and can be applied as preemergent and soil-incorporated treatments (Boyette et al. 1984). The shelf life of the product can be improved by manipulating the water activity and sucrose content. Other solid substrates used to formulate mycoherbicides include bran, wheat kernels, cornmeal/sand, vermiculite (Auld and Morin 1993; Boyette et al. 1996), and sawdust (Muir 2023). For example, cornmeal/cornmeal sand was used to formulate mycelium, micro- and macroconidia, and chlamydospores of Fusarium solani f.sp. cucurbitae for the control of Texas gourd. This pre-emergent granular formulation provided 96% control of the weed (Boyette et al. 1984).\n\n7.3.4 Formulation of viruses\n\nBaculoviruses have been studied as a potential solution for controlling insect pests that belong to the Lepidoptera, Hymenoptera, and Coleoptera families (Gory and Bishop 1995). These viruses have several advantages, such as high specificity, not harming beneficial insects, and persisting in the environment, which makes long-term control of insect pests possible without damaging the environment. Nuclear polyhedrosis viruses (NPVs) and granulosis viruses (GVs) are examples of baculoviruses. However, there are some limitations to using these biopesticides, such as their slow speed of biological activity, low stability under UV light, and difficulties in production (Powell and Jutsum 1993).\n\nThe stability of the baculoviruses is often a function of viability, but it is not a significant problem for small-scale field trials as the viruses can be collected from macerated larvae, mixed with water, and stored for short periods through refrigeration. However, these systems are not suitable for large-scale production and application (Gory and Bishop 1995). The formulation of these viruses is an important aspect of product development, but researchers have not given it as much attention as bacteria and fungi.\n\nBaculoviruses are usually available in the form of concentrated wettable powders. The biocontrol product called “Elcar” for corn earworm (Helicoverpa zea) NPV can be spray-dried or air-dried after being diluted with an inert carrier. The gypsy moth (Lymantria dispar L.) NPV is freeze-dried either with carbohydrates or by acetone precipitation (Gory and Bishop 1995). The virus can be inactivated by UV radiation, especially wavelengths of 290-320 nm. UV protectants like reflections or absorbers can be added to formulations to stabilize baculoviruses. Several dyes, such as lissamine green, acridine yellow, alkali blue, and mercurochrome, have been used as UV protectants, especially to absorb UV-A irradiation (Shapiro 1995). Optical brighteners (fluorescent brighteners), which are commonly used in soaps, detergents, and fabric softeners, also absorb UV light. They have been shown to significantly reduce the photodegradation of NPVs and enhance viral activity (Shapiro 1992; Dougherty et al. 1995; Shapiro 1995). Care must be taken to ensure that these optical brighteners do not adversely affect the environment due to residues and residuality.\n\nThere are two types of biopesticide formulations: liquid and dry, which differ based on their physical state (Figure 7). Dry formulations (dust powders, granules, seed dressers, wettable powders, and wettable) dispersible powder. They are produced using various technologies, including spray drying, freeze drying, or air drying, with or without a fluidized bed (Li 2002). They typically require binders, dispersants, and wetting agents for optimal efficacy and application. Liquid formulations can be water-based, oil-based, polymer-based, or combinations. Water-based formulations, like suspension concentrates, suspo-emulsions, capsule suspensions, and ultra-low volume liquids, require additional inert ingredients for stability and performance (Boyetchko et al. 1999). These include stabilizers, stickers, surfactants, coloring agents, antifreeze compounds, and additional nutrients. Care must be taken to ensure that these inert ingredients do not adversely affect the environment due to residues and residuality and do not contain pesticide derivatives.\n\nLegends: Straight line arrows indicate process (steps), and broken lines indicate categories or classifications.\n\n\n8. Advances in biopesticide and delivery technologies enhance the efficacy of biological products\n\nAdvances in biopesticide application techniques have been driven by the need for more efficient and targeted pest control methods while minimizing environmental impacts and fate. These advancements aim to improve the delivery, efficacy, and performance of biopesticides. Several improved spray technologies and precision targeting methods have been developed, including controlled droplet application, variable rate technology, intelligent spray systems, air-assisted sprayers, robotics, and automation technologies, automated targeting systems (Wang et al. 2019), thermal fogging, and aerial application. The improved spray technologies play an important role in delivering droplets of consistent size and velocity, improving spray deposition, reducing non-target drift, optimizing chemical pesticide usage, reducing costs, and minimizing environmental impacts and fate. Additionally, some of these technologies utilize sensors and algorithms to detect and target pests in real-time, enabling precise application of biopesticides only where needed. Other techniques improve the effectiveness of biopesticides by reaching target areas that are otherwise difficult to treat, and some spray techniques improve coverage and reduce the amount of biopesticide needed.\n\nSeed treatments and soil applications involve various methods for applying biopesticides to protect seeds and enhance crop establishment. Seed coatings and treatments apply biopesticides onto the seed surface, providing early protection and improving crop establishment (Khan et al. 2020). Incorporating biopesticides into soil amendments can improve soil health, control soil-borne pests, and enhance plant growth, offering sustainable pest management and environmental benefits. Other methods include soil drenching, which involves applying biopesticides as a liquid solution directly to the soil around the plant base. This technique targets pests in the soil, such as nematodes and soil-borne pathogens. Biopesticide-based soil fumigation utilizes the controlled release of compounds to control pests and diseases in the soil, providing an alternative to chemical fumigation but with reduced environmental impacts. Rhizosphere application focuses on applying biopesticides directly to the root zone of plants. This technique targets pests that interact with plant roots, such as soil-dwelling insects and nematodes. These seed treatment and soil application methods offer effective pest management while minimizing environmental impacts and fate.\n\nVarious challenges and limitations, including environmental conditions such as temperature, humidity, and sunlight, can be influenced by the efficacy of biopesticides during pest control. Some biopesticides may require specific temperatures or moisture levels for optimal performance. Additionally, biopesticides often exhibit narrow target specificity, that is effective against specific pests but not others. This limitation reduces their broad-spectrum efficacy, and it may be necessary to use multiple biopesticides to target different pests (Gurr et al. 2017). Furthermore, the formulation and storage conditions of biopesticides can affect their stability and shelf life. Factors such as pH, temperature, and exposure to light can degrade the effectiveness of biopesticides over time (Riga et al. 2017).\n\nIt’s important to note that these challenges and limitations can vary depending on the specific biopesticide and pest control scenario. By considering and addressing these factors, it is possible to optimize the efficacy of biopesticides in pest control practices. The efficacy of biopesticides can be influenced by the application methods used. Factors such as spray volume, droplet size, and coverage play a crucial role in the distribution and effectiveness of the biopesticide on the target pests (Chandler et al. 2011). Ensuring that the application method is properly calibrated is important to achieve adequate coverage and contact with the pests (International Symposium on Food Safety and Control).\n\nBiopesticides may have a shorter persistence compared to chemical pesticides, which may require more frequent applications. Their residual activity on plants or the environment can be limited, necessitating careful timing and repeated treatments. Proper timing of application is essential to target pests during vulnerable stages of their life cycle thus, it is critical to fully understand the pest’s lifecycle before implementing a biopesticide programme. In addition to the challenges mentioned earlier, several other factors can limit the efficiency of biopesticides. These include regulatory approval, which can be a complex and time-consuming process. Limited availability and accessibility of biopesticides can pose challenges for farmers (Biondi et al. 2021). And their compatibility with integrated pest management can be a complex issue. Lack of standardization and quality control protocols may affect the consistent performance of biopesticides. Furthermore, biopesticides can be more expensive compared to chemical pesticides, affecting their cost-effectiveness. The efficacy of biopesticides can vary depending on factors such as pest population density, pest development stage, and resistance mechanisms. Pests can develop resistance to specific biopesticides over time, which can reduce their effectiveness. Therefore, it is important to monitor pest populations and implement IPM strategies that incorporate the use of different biopesticides and other pest control measures to mitigate the development of resistance.\n\n\n9. Future perspectives and research directions\n\nEmerging trends in biopesticide development are driven by the need for sustainable agricultural practices and the increasing demand for ecologically safe pest control methods. Some of the key trends are summarized in Figure 8. Future perspectives and research directions in the field of microbial biopesticides are important for advancing their development and application (Karlsson et al. 2020). Microbiopesticides are gaining significant attention in modern agriculture due to their environmentally safe nature and reduced impact on human health. Researchers and companies are actively involved in developing new biopesticide formulations and strategies to enhance their efficacy. Some emerging trends in biopesticide development include microbial biopesticides, plant extracts and essential oils, RNA-based biopesticides, nanotechnology in biopesticides, and the combination of different biopesticides for improved pest control (Karlsson et al. 2020). As the demand for sustainable and ecologically safe pest management strategies has increased, biopesticides have made tremendous strides in recent years. The use of microbial biopesticides, such as bacteria, fungi, and viruses, with qualities enhanced through genetic modification or natural selection, is one of the most significant discoveries. Figure 8 provides a comprehensive summary of the emerging trends in biopesticide development and the integration of biopesticides with other integrated pest management (IPM) practices and pest control methods.\n\nThese trends indicate a growing focus on developing innovative, effective, and sustainable pest control solutions that align with environmental conservation and agricultural productivity goals.\n\n\n10. Conclusion\n\nThe adoption of biopesticides is an increasingly popular trend in agriculture due to various environmental, health, and economic benefits. Biopesticides, derived from natural materials like animals, plants, bacteria, and certain minerals, offer a sustainable alternative to traditional chemical pesticides. Despite the benefits, several challenges can hinder the widespread adoption of biopesticides. Novel microbial strain creation with improved features, such as greater efficacy and larger target spectra, should be prioritized in future research and the use of biopesticides. To maximize their efficiency and reduce the possibility of resistance, biopesticides’ modes of action must be understood. Along with other tactics, using biopesticides in complete pest management programs can improve overall pest control. The stability, adhesion, and targeted distribution of biopesticides will be improved through improvements in formulation and delivery techniques. Risk assessment and regulatory frameworks must be created to guarantee the safety and effectiveness of biopesticides. To encourage knowledge and acceptance of biopesticides, efforts should be made to educate and raise public awareness. Collaboration and knowledge sharing among researchers, stakeholders, and regulatory agencies will facilitate progress in the field. By focusing on these recommendations, biopesticides can contribute to sustainable and environmentally safe pest management practices in agriculture.\n\n\nAuthors contribution\n\nConceptualization: K.T.M.; Original draft preparation: K.T.M. and L.M.; Writing: K.T.M., D. M.; and D.M.; Editing, Reviewed, and formatting: D.N.: and Reviewed drafts of the paper: J.G.; Language editing and preparation of tables and/or figures: K.T.M.; Final approval of the review to be published.\n\nAll authors have read and agreed to the published version of the manuscript.", "appendix": "Data availability statement\n\nNo data are associated with this article.\n\n\nAcknowledgments\n\nThe authors would like to thank the International Center for Genetic Engineering and Biotechnology (ICGEB) for their financial support.\n\n\nReferences\n\nAbd-Alla AMM, Meki IK, Demirbas-Uzel G: Insect Viruses as Biocontrol Agents: Challenges and Opportunities.El-Wakeil N, Saleh M, Abu-hashim M, editors. Cottage Industry of Biocontrol Agents and Their Applications. Cham: Springer; 2020; pp. 277–295\n\nAdesemoye AO, Kloepper JW: Plant-microbes interactions in enhanced fertilizer-use efficiency. Appl. Microbiol. Biotechnol. 2009; 85: 1–12. 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Publisher Full Text\n\nVivekanandhan P, Swathy K, Murugan AC: Insecticidal Efficacy of Metarhizium anisopliae Derived Chemical Constituents against Disease-Vector Mosquitoes. J. Fungi. 2022; 8: 300. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWalker HL, Boyette CD: Biocontrol of sicklepod (Cassia obtusifolia) in soybeans (Glycine max) with Alternaria cassiae. Weed Sci. 1985; 33: 212–215. Publisher Full Text\n\nWang J, Wu W, Qin Y, et al.: Automated targeting systems for pesticide application in agriculture: A review. Comput. Electron. Agric. 2019; 156: 459–471.\n\nWattimena CMA, Latumahina FS: Effectiveness of botanical biopesticides with different concentrations of termite mortality. J. Belantara. 2021; 4: 66–74. Publisher Full Text\n\nYelitza C, Mundstock S, Sampaio M, et al.: Use of Parasitoids as a Biocontrol Agent in the Neotropical Region: Challenges and Potential. IntechOpen; 2020. Hortic Crop. Publisher Full Text\n\nZhang SA, Xu WM, Yan Z, et al.: Research and commercialization of yield-increasing bacteria (YIB) in China.Tang WHRC; AR, editors. Advances in Biological Control of Plant Diseases. Beijing: China Agricultural University Press; 1996; pp. 47–53.\n\nZidack NK, Backman PA, Shaw JJ: Promotion of bacterial infection of leaves by an organosilicone surfactant: implications for biological weed control. Biol. Control. 1992; 2: 111–117. Publisher Full Text" }	[ { "id": "326005", "date": "04 Oct 2024", "name": "Yordanys Ramos", "expertise": [ "Reviewer Expertise Biological Control", "Integrated Pest Management", "Entomology", "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\nIntroduction section, Page 3, last paragraph.Change Beauveria bassiana to Beauveria spp. since there are many more entomopathogenic species within the Beauveria genus, not just Beauveria bassiana. It would be more accurate to use the genus (Beauveria spp.), just as was done with Metarhizium, Isaria, and Verticillium spp.\nIn that same paragraph, the authors should change Verticillium to Lecanicillium according to the current classification of that fungal genus.\nPage 5. Concepts and classifications of biopesticides. The authors should provide a citation for the concept of biopesticide provided by SADC, just as they did for the concept provided by USEPA. Likewise, they should include a citation for the concept provided by The European Environmental Agency.\nPage 5: Please provide a citation for the Paris Agreement and the Nagoya Protocol. The authors can find both documents at these links: https://unfccc.int/sites/default/files/english_paris_agreement.pdf https://www.cbd.int/abs/doc/protocol/nagoya-protocol-en.pdf\nPage 6: Biopesticide Categories. The authors do not need to repeat the concept of biopesticide again, as this makes the review repetitive and less engaging for the reader, especially considering that a section was already dedicated to detailing the various existing concepts of this term.\nPage 11: Figure 3, Mode of Action of Entomopathogenic Fungi. The cycle refers to the infection of Beauveria bassiana. Therefore, it would not be prudent to include a photo of an insect killed by Metarhizium anisopliae outside of the cycle. Additionally, it is quite confusing that the spore colors are shown as white in both cases. This is accurate for Beauveria bassiana, but when Metarhizium anisopliae causes mycosis in an insect, its spores are olive green.\nIn the section on entomopathogenic fungi as bioinsecticides, the authors should briefly mention that entomopathogenic fungi also have the ability to act as endophytes in plants and crops, and that by producing their secondary metabolites, they protect the plants from insect herbivory. This primarily includes fungi like Beauveria bassiana and Metarhizium anisopliae. This is also a bioinsecticidal activity of these microorganisms.\nPage 13: What are the optimal conditions for the baculoviruses to cause rapid death within 3 to 7 days? This was left ambiguous, so it needs to be clarified.\nPage 17: the numbering of the sections has been lost; it goes from 7.2.2 to 7.3.2. In any case, it should be 7.3. Please review.\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? Yes", "responses": [ { "c_id": "12612", "date": "14 Oct 2024", "name": "Kahsay Tadesse Mawcha", "role": "Author Response", "response": "Thank you very much for the careful review of our manuscript entitled “Recent Advances in Biopesticide Research and Development: A Focus on Microbial: A Review. We appreciate your time and insightful comments on our manuscript. We have addressed all the comments and suggestions forwarded from the reviewer. We made significant changes and addressed each point by point as follows: Regrads, Kahsay Introduction section, Page 3, last paragraph. Change Beauveria bassiana to Beauveria spp. since there are many more entomopathogenic species within the Beauveria genus, not just Beauveria bassiana. It would be more accurate to use the genus (Beauveria spp.), just as was done with Metarhizium, Isaria, and Verticillium spp. In that same paragraph, the authors should change Verticillium to Lecanicillium according to the current classification of that fungal genus. = We want to mention the specific Beauveria bassiana. However, we agreed with the reviewer's suggestion and will improve the text to “Metarhizium, Isaria, and Lecanicillium spp.” Page 5. Concepts and classifications of biopesticides. The authors should provide a citation for the idea of biopesticide provided by SADC, just as they did for the concept provided by USEPA. Likewise, they should include a citation for the idea provided by The European Environmental Agency. European Environment Agency, 2023. How pesticides impact human health and ecosystems in Europe EEA Briefings 06 (2023) https://www.sciencedirect.com/science/article/pii/S0269749124005505#bib27 = For the SADC, we provided a link to the reference. We can also give the reference as ….(SADC 2019) SADC Secretariat 2019. SADC (Southern African Development Community) Guidelines for Pesticides Management and Risk Reduction. http://faolex.fao.org/docs/pdf/sad212166.pdf Page 5: Please provide a citation for the Paris Agreement and the Nagoya Protocol. The authors can find both documents at these links: https://unfccc.int/sites/default/files/english_paris_agreement.pdf https://www.cbd.int/abs/doc/protocol/nagoya-protocol-en.pdf = We appreciate the suggestions. We include the citations for “The Paris Agreement (United Nations 2015) which is available on the following link. https://unfccc.int/sites/default/files/english_paris_agreement.pdf For the Nagoya Protocol 2014. “The Nagoya Protocol on Access to Genetic Resources and The Fair and Equitable Sharing of Benefits Arising from Their Utilization to the Convention on Biological Diversity” https://www.cbd.int/abs/doc/protocol/nagoya-protocol-en.pdf Generally, both references can be used in the main text but we may not include them in the reference list as they are not peer-reviewed documents. Page 6: Biopesticide Categories. The authors do not need to repeat the concept of biopesticide again, as this makes the review repetitive and less engaging for the reader, especially considering that a section was already dedicated to detailing the various existing concepts of this term. = It is not a repetition. The first one is to show the concepts depending on EPA, EAC, The European Environmental Agency, the International Biocontrol Manufacturer’s Association (IBMA), the International Organization for Biological Control (IOBC), and SADC. The purpose of discussing on page 6 is to demonstrate the different categories of microbial biopesticides NOT all classifications of Biopesticides. Page 11: Figure 3, Mode of Action of Entomopathogenic Fungi. The cycle refers to the infection of Beauveria bassiana. Therefore, it would not be prudent to include a photo of an insect killed by Metarhizium anisopliae outside of the cycle. Additionally, it is quite confusing that the spore colors are shown as white in both cases. This is accurate for Beauveria bassiana, but when Metarhizium anisopliae causes mycosis in an insect, its spores are olive green. = The color of the spore doesn't matter when it comes to the dead insect, as it doesn't refer to spores when it's killed. The picture aims to illustrate that when microbial biopesticides are used, the target pest being killed is part of the cycle. This is just how it works, and the spores may produce different colors. The purpose of the picture is to demonstrate the mode of action of the entomopathogenic fungi. In the section on entomopathogenic fungi as bioinsecticides, the authors should briefly mention that entomopathogenic fungi also have the ability to act as endophytes in plants and crops, and that by producing their secondary metabolites, they protect the plants from insect herbivory. This primarily includes fungi like Beauveria bassiana and Metarhizium anisopliae. This is also a bioinsecticidal activity of these microorganisms. = We have mentioned Beauveria bassiana and Metarhizium anisopliae throughout the text. To avoid repetitions in the review, it is not necessary to include everything in this text. But, if we don’t exceed the page number, we can include the text suggested by the reviewer. Page 13: What are the optimal conditions for the baculoviruses to cause rapid death within 3 to 7 days? This was left ambiguous, so it needs to be clarified. =The optimal conditions include optimum temperature and ultraviolet radiation. These have already been mentioned under the advantages of viral biopesticides. The common limitations of all viral products. Page 17: the numbering of the sections has been lost; it goes from 7.2.2 to 7.3.2. In any case, it should be 7.3. Please review. =Yes. Accepted. The subtitle “7.3.2 Formulation of fungal biopesticides” should be “7.2.3 Formulation of fungal biopesticides” Many thanks for your time and consideration Sincerely, Kahsay Tadesse" } ] }, { "id": "326009", "date": "16 Oct 2024", "name": "Mehari Desta Hawku", "expertise": [ "Reviewer Expertise My research interests encompass a broad spectrum of plant pathology including:Identification and molecular characterization of resistance or susceptible genes in plant-microbe interactions", "Understanding the intricate mechanisms governinginteractions between plants and microbes", "Examining the signaling pathways involved in activating plant defense responses against pathogens", "Studying the molecular components that enable pathogens to invade and manipulate plant cells", "Understanding the mechanisms of abiotic stress tolerance in plants" ], "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 investigates the rising significance of microbial biopesticides as eco-friendly alternatives to traditional pesticides. It emphasizes the potential of utilizing natural organisms, such as bacteria, fungi, viruses, nematodes and protozoans, for effective pest control while reducing environmental harm. The authors provide a thorough examination of recent advancements and methodologies that enhance the effectiveness of these biopesticides. Furthermore, the review underscores the importance of integrating these biopesticides into comprehensive integrated pest management strategies. However, to offer a more comprehensive review, the following aspects should be addressed:\nWhile the topic is broad, the literature focuses only on the role of microbial biopesticides in controlling insect pests, overlooking their role in disease and weed control The role of recent cutting-edge technologies, such as CRISPR/Cas9 and RNA interference (RNAi), was not thoroughly discussed. Regulatory frameworks and commercialization aspects, were not addressed.\nMinor comments:\nSection 1: Introduction\nThe introduction section lacks a comprehensive summary that clearly informs readers about the review's aims, the specific points the literature addresses, as well as the knowledge gaps and suggested future research directions. It is essential to include a concise overview outlining the main themes covered in the review. Additionally, highlighting the role of emerging technologies, such as CRISPR/Cas9 and RNA interference (RNAi) in biopesticide development, along with the potential for integrating biopesticides into sustainable agricultural practices, will provide clearer context for the reader. This summary should also emphasize the importance of interdisciplinary collaboration among researchers, policymakers, practitioners, and manufacturers to enhance the efficacy and acceptance of biopesticides in agricultural systems. The image in Figure 1 is blurry; please consider replacing it with a clearer image and remove the word \"legends\". From the description of Figure 1, it appears that you have missed one image. Therefore, you can either remove the sentence, \"The second part demonstrates how the five steps for implementing CSA intersect with and affect the 17 SDGs in terms of synergies and trade-offs,\" or consider incorporating the missing image.\n\nSection 2: Methods\nThe methodology section is missing some key points, as outlined below:\n\nIt is important to clarify the specific research questions guiding the review\n\nIt’s unclear how studies were selected for review. Please include specific inclusion and exclusion criteria used for selecting papers The method does not specify the keywords or phrases used during the search process. There’s no description of how potential biases in selecting literature were mitigated, nor is there any indication of measures taken to avoid confirmation bias. Mentioning how the quality of the papers was assessed would add rigor to the methodology It would be helpful to specify the types of studies reviewed or considered for inclusion Please specify the time frame of the literature reviewed If there was any geographical focus, it would be beneficial to mention it.\nSection 3: Concept and classification of biopesticides\nThe second paragraph in this section is not related to the topic. Instead, this concept should be discussed under section 4. The idea in the third paragraph should also be addressed on a separate topic as Mechanism of action and efficacy of microbial biopesticides\nSection 4: Importance of biopesticides in sustainable agriculture\nThe sentence ‘The use of biopesticides has steadily increased by a steady 11.0% Compound Annual Growth Rate (CAGR) between 2018 and 2022.’ under section 4 should be rephrased as ‘The use of biopesticides has steadily increased at a Compound Annual Growth Rate (CAGR) of 11.0% between 2018 and 2022.’ to avoid the redundancy of the word ‘steadily’ and ‘steady’ Under this section the following points should be discussed in details; Environmental Safety, Reduced Chemical Dependency, Promoting Biodiversity, Compatibility with Organic Farming, and Sustainability and Resilience .\nSection 5: Role of biopesticides in integrated pest management\nThe different microbial biopesticides should be discussed under subsection 5.1 as a sub-subsection not as a subsection. In table 1, the role of microbial biopesticides in the IPM program is not mentioned within the table; therefore, please consider modifying the table's title. In Table 1, I suggest adding a fifth column to list the references In Table 1, could you clarify what is meant by 'application type'? If it refers to the method of delivery, this is not clearly specified. In addition, the mode of action is missing in several instances under the column ‘Application Type and Mode of Action’. For some microbial biopesticides, important details are absent within the table. On Page 9, under Table 1, please replace 'fungal-based biopesticides' with 'viral-based biopesticides.' Overall, Table 1 lacks consistency and would benefit from clearer definitions and more complete information.\nSubsection 5.2: Bacterial based biopesticides\nAfter using the full name Bacillus thuringiensis (Bt), you should continue using its abbreviated form (Bt) throughout the text. The last paragraph under sub-section 5.2 should be supported by relevant literature. The image (particularly the text) in Figure 2 is unclear; please consider replacing it with a clearer version. Additionally, remove the word 'legend' as well as the phrase 'The mode of action of Bt gene and Cry protein,' since this information is already specified in the figure itself\nSubsection 5.3: Fungal biopesticides A citation is needed for Figure 3. Such figures are typically self-explanatory and do not require legends, so please remove the note stating, ‘Legends are indicated in the figures. No additional legends.’\nSubsection 5.4: Viral biopesticides\nPlease replace ‘Baculovirus family’ with Baculoviridae and do not italicize baculovirus Would you please reconsider the first sentence of the third paragraph in subsection 5.4 to maintain clarity? In Figure 4 (Page 12), please remove the term 'legends,' as it is unnecessary to mention since it is evident that you are describing the figure. Additionally, please ensure you provide proper citations for this figure, as you are referencing two sources: Singh (2019) and Kalamkoff (2004).\n\nOn page 13, the same legend is repeated; please consider removing it.\nSubsection 5.5: Nematodes as biopesticides\nPlease add a citation after the caption of Figure 5 and remove the citation from the description. Figure 5 is somewhat blurry; please use a clearer image and consider removing the word ‘legends’.\nIn subsection 5.6, please consider not italicizing the word ‘septicaemia’\nSection 6: Natural enemies of pests as biopesticides\nSince the primary focus of this review is on microbial biopesticides, and predators and parasitoids are classified as macrobials, I suggest removing this section. The image (particularly the text) in Figure 6 is not clear; please consider replacing it. Please add a citation to Figure 6.\nPlease add a citation to Figure 7 and consider using a clear image\nSubsection 8.2: Factors affecting efficacy of biopesticides\nUnder this subsection, the first sentence should be modified as ‘Various challenges and limitations, including environmental conditions such as temperature, humidity, and sunlight, can influence the efficacy of biopesticides during pest control’ In the second paragraph, please check the citation related to ‘ (International Symposium on Food Safety and Control).’ The image in Figure 8 is blurry; please consider using a clearer and more visually appealing image. I recommend using photo editing software, such as Photoshop, to enhance the quality. Additionally, please remove the phrase 'own work' from the caption. Finally, ensure that the figure is either self-explanatory or includes a legend where necessary.\nSection 9. Future perspectives and research directions Please remove subsection 9.1 from this section, as there are no other subsections following it. Additionally, avoid using a single sentence as a paragraph; instead, merge it with the preceding paragraph. In this section on future perspectives and research directions, it would be valuable to discuss in more detail the role of the cutting-edge technologies, such as CRISPR/Cas9 and RNA interference (RNAi) in pest control.\nSection 10. Conclusion\nThe conclusion effectively encapsulates the key findings regarding the benefits and challenges of biopesticides. However, the sentence stating that \"novel microbial strain creation with improved features, such as greater efficacy and larger target spectra, should be prioritized in future research and the use of biopesticides\" may be more appropriate in a future perspectives section. This sentence emphasizes the need for ongoing research and innovation, which aligns better with a section dedicated to future research directions.\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": "12689", "date": "29 Oct 2024", "name": "Kahsay Tadesse Mawcha", "role": "Author Response", "response": "Dear Chief Editor, Dear Reviewer, We have carefully considered all of the reviewers' comments and suggestions. To address these, we have made substantial revisions to the manuscript. Below, we provide a detailed response to each specific point raised. We believe that these revisions have significantly strengthened the manuscript and improved its clarity, comprehensiveness, and overall quality. We are confident that the revised manuscript now meets the high standards of your journal. Thank you again for your valuable feedback. Sincerely, Reviewer 2 Comments and responses The manuscript investigates the rising significance of microbial biopesticides as eco-friendly alternatives to traditional pesticides. It emphasizes the potential of utilizing natural organisms, such as bacteria, fungi, viruses, nematodes and protozoans, for effective pest control while reducing environmental harm. The authors provide a thorough examination of recent advancements and methodologies that enhance the effectiveness of these biopesticides. Furthermore, the review underscores the importance of integrating these biopesticides into comprehensive integrated pest management strategies. However, to offer a more comprehensive review, the following aspects should be addressed: While the topic is broad, the literature focuses only on the role of microbial biopesticides in controlling insect pests, overlooking their role in disease and weed control We accept the reviewer’s comment. To address his comment, we have modified the title of the review to focus on Microbial biopesticides and recent advancements and methodologies The role of recent cutting-edge technologies, such as CRISPR/Cas9 and RNA interference (RNAi), was not thoroughly discussed As the focus of the current review is microbial biopesticides, CRISPR/Cas9 and RNA interference (RNAi) have been discussed in detail under the emerging trends in biopesticides. We believe that there is no need to discuss them as a subheading. Regulatory frameworks and commercialization aspects were not addressed. The review focuses on summarizing previous and recent studies about the effectiveness of biopesticides, particularly microbial products. Hence, regulatory frameworks and commercialization aspects are beyond the scope of the review. However, we have discussed a few points under the conclusion part about the importance of Regulatory frameworks and the need for harmonization of biopesticide guidelines as a future work. We have also modified the title “Recent Advances in Biopesticide Research and Development with A Focus on Microbials” Minor comments: Section 1: Introduction The introduction section lacks a comprehensive summary that clearly informs readers about the review's aims, the specific points the literature addresses, as well as the knowledge gaps and suggested future research directions. It is essential to include a concise overview outlining the main themes covered in the review. Additionally, highlighting the role of emerging technologies, such as CRISPR/Cas9 and RNA interference (RNAi) in biopesticide development, along with the potential for integrating biopesticides into sustainable agricultural practices, will provide a clearer context for the reader. This summary should also emphasize the importance of interdisciplinary collaboration among researchers, policymakers, practitioners, and manufacturers to enhance the efficacy and acceptance of biopesticides in agricultural systems. We appreciate the suggestions given by the reviewer. However, most of the comments are out of the scope of the review content. Some of the comments, such as the role of emerging technologies, such as CRISPR/Cas9 and RNA interference (RNAi), and the potential for integrating biopesticides into sustainable agricultural practices, are highlighted in the conclusion part. The image in Figure 1 is blurry; please consider replacing it with a clearer image and remove the word \"legends\". From the description of Figure 1, it appears that you have missed one image. Therefore, you can either remove the sentence, \"The second part demonstrates how the five steps for implementing CSA intersect with and affect the 17 SDGs in terms of synergies and trade-offs,\" or consider incorporating the missing image. Within the figure, sections show the different linkages with climate-smart agriculture and how they are linked to the 17 SDGs. Hence, there is no missed image in this section. The figure provides information that is interlinked with each other. Climate-smart agriculture (CSA) has a strong potential to contribute to the achievement of the 17 Sustainable Development Goals (SDGs). A critical precondition for realising this scope is identifying potential synergies and trade-offs between the 3 pillars of CSA and the 5 implementation steps of CSA and the SDGs. These provide entry points for targeted CSA planning so as to enhance synergies and reduce trade-offs. CSA and the SDGs have synergies and trade-offs at different levels for each of the 3 pillars of CSA where some of the synergies & trade-offs overlap all 3 pillars of CSA for some of the SDGs, namely SDGs 2 and 6. The overlapping synergies & trade-offs for the implementation steps and SDGs are different, as only step 2 has overlapping trade-offs and synergies in SDG’s, 6,9,13, and 15. We have also improved the paragraph and made some changes to it. Section 2: Methods The methodology section is missing some key points, as outlined below: It is important to clarify the specific research questions guiding the review What are the current trends and advancements in the development and application of microbial biopesticides? What are the key challenges and limitations associated with the use of microbial biopesticides in agricultural settings? What are the most promising microbial agents currently being developed for use as biopesticides? Are the most important guiding questions It’s unclear how studies were selected for review. Please include specific inclusion and exclusion criteria used for selecting papers Only microbial (bacterial, fungal, viral, nematode, and protozoa) biopesticides were included and other types were excluded The method does not specify the keywords or phrases used during the search process. Using various keywords, including biocontrol, formulations, bioprotection, climate-smart agriculture, and sustainable pest management, 197 relevant studies were found. We have already included the following phrases: The literature outcomes were categorised into the (i) classification of biopesticides, (ii) importance of biopesticides in sustainable agriculture, (iii) role of microbial biopesticides in integrated pest management, (iv) different types and formulations of microbial biopesticides, and (v) advancement and future perspectives of microbial biopesticides. There’s no description of how potential biases in selecting literature were mitigated, nor is there any indication of measures taken to avoid confirmation bias. To avoid biases in selecting literature, we used articles that are published in peer-reviewed journals, which means available on the following search engines, Google Scholar, PubMed, Scopus (Elsevier), Web of Science, Semantic Scholar, Academia Mentioning how the quality of the papers was assessed would add rigor to the methodology Most of our sources are published in peer-reviewed journals. It would be helpful to specify the types of studies reviewed or considered for inclusion We have included original research articles and reviews published in peer-reviewed journals and other relevant resources Please specify the time frame of the literature reviewed Mostly, we used recently published articles (2018-2024) and very few; we have also included > 5 references, which are from 2003>. If there were any geographical focus, it would be beneficial to mention it. The review is an overview of the current research and developments in microbial biopesticides. So, there are no specific geographic limitations. Section 3: Concept and classification of biopesticides The second paragraph in this section is unrelated to the topic. Instead, this concept should be discussed under section 4. Section 3 deals with the classification/definition of biopesticides (general) by various organizations and associations. So, it is better to be as it is. The idea in the third paragraph should also be addressed on a separate topic as Mechanism of action and efficacy of microbial biopesticides We accept the suggestion however, as we have discussed the mode of action and efficacy of each microbial product, this subtitle may cause a repetition. Section 4: Importance of biopesticides in sustainable agriculture The sentence ‘The use of biopesticides has steadily increased by a steady 11.0% Compound Annual Growth Rate (CAGR) between 2018 and 2022.’ under section 4 should be rephrased as ‘The use of biopesticides has steadily increased at a Compound Annual Growth Rate (CAGR) of 11.0% between 2018 and 2022.’ to avoid the redundancy of the word ‘steadily’ and ‘steady’ Accepted and addressed. Under this section, the following points should be discussed in detail: Environmental Safety, Reduced Chemical Dependency, Promoting Biodiversity, Compatibility with Organic Farming, and Sustainability and Resilience. We are happy to discuss them in detail however, we may go beyond the scope of the topic! Section 5: Role of biopesticides in integrated pest management The different microbial biopesticides should be discussed under subsection 5.1 as a sub-subsection, not as a subsection. Comment Accepted. However, we discuss the various types of biopesticides, including Biochemical, Botanical, and pheromone-based; the world limit and the number of pages may exceed the limit. So, it is better to focus on the title-i.e. microbial-based biopesticides! In Table 1, the role of microbial biopesticides in the IPM program is not mentioned within the table; therefore, please consider modifying the table's title. Comment accepted, and Table 1. has been modified as “Microbial biopesticide categories, mode of action, and their use in pest management. In Table 1, I suggest adding a fifth column to list the references We have included the references in line with each column. In Table 1, could you clarify what is meant by 'application type'? If it refers to the method of delivery, this is not clearly specified. In addition, the mode of action is missing in several instances under the column ‘Application Type and Mode of Action’. For some microbial biopesticides, important details are absent within the table. The table is self-explanatory and the title is also modified as suggested. The “'application type'” is NOT referring to the method. It is referring to “use”. The remark column is for additional information. On Page 9, under Table 1, please replace 'fungal-based biopesticides' with 'viral-based biopesticides.' Overall, Table 1 lacks consistency and would benefit from clearer definitions and more complete information. Accepted, but as the title of the table is formatted by the journal editor, it will be corrected during the production. Subsection 5.2: Bacterial-based biopesticides After using the full name Bacillus thuringiensis (Bt), you should continue using its abbreviated form (Bt) throughout the text. Yes, but when we start a new line, we normally use the full name, that is, species and genus names. The last paragraph under sub-section 5.2 should be supported by relevant literature. All paragraphs are cited. It is not clear! The image (particularly the text) in Figure 2 is unclear; please consider replacing it with a clearer version. Additionally, remove the word 'legend' as well as the phrase 'The mode of action of Bt gene and Cry protein,' since this information is already specified in the figure itself Okay. Accepted, and the legend is deleted. Subsection 5.3: Fungal biopesticides A citation is needed for Figure 3. Such figures are typically self-explanatory and do not require legends, so please remove the note stating, ‘Legends are indicated in the figures. No additional legends.’ Okay. Accepted. The figure is our own work, and there is no need to include citations. Subsection 5.4: Viral biopesticides Please replace ‘Baculovirus family’ with Baculoviridae and do not italicise baculovirus Done! Would you please reconsider the first sentence of the third paragraph in subsection 5.4 to maintain clarity? Accepted and modified as “Figure 4. shows how viral replication and infection take place and attack their target host”. In Figure 4 (Page 12), please remove the term 'legends,' as it is unnecessary to mention since it is evident that you are describing the figure. Additionally, please ensure you provide proper citations for this figure, as you are referencing two sources: Singh (2019) and Kalamkoff (2004). Accepted and the “Figure 4. Mode of action of viral biopesticides ( Singh et al. 2019).” It was deleted in the text. On page 13, the same legend is repeated; please consider removing it. Okay. Done! Subsection 5.5: Nematodes as biopesticides Please add a citation after the caption of Figure 5 and remove the citation from the description. Figure 5 is somewhat blurry; please use a clearer image and consider removing the word ‘legends’. Accepted. We have deleted the term “legend”. However, the Figure is our own work, and there is no need to put citations. In subsection 5.6, please consider not italicizing the word ‘septicaemia’ Accepted and done! Section 6: Natural enemies of pests as biopesticides Since the primary focus of this review is on microbial biopesticides, and predators and parasitoids are classified as macrobials, I suggest removing this section. Sometimes, the definition of biopesticides may include parasitoids and predators, and their inclusion in this review enhances the broader reader of the journal. The image (particularly the text) in Figure 6 is unclear; please consider replacing it. Please add a citation to Figure 6. The figure is our own work and no need to include a citation. Please add a citation to Figure 7 and consider using a clear image The figure is our own work and no need to include a citation. Subsection 8.2: Factors affecting efficacy of biopesticides Under this subsection, the first sentence should be modified as ‘Various challenges and limitations, including environmental conditions such as temperature, humidity, and sunlight, can influence the efficacy of biopesticides during pest control’ Accepted and done! In the second paragraph, please check the citation related to ‘ (International Symposium on Food Safety and Control).’ Comment accepted. We have added the full information and hyperlinked the details. https://conferences.iaea.org/event/351/ The image in Figure 8 is blurry; please consider using a clearer and more visually appealing image. I recommend using photo editing software, such as Photoshop, to enhance the quality. Additionally, please remove the phrase 'own work' from the caption. Finally, ensure that the figure is either self-explanatory or includes a legend where necessary. Accepted and done! Section 9. Future perspectives and research directions Emerging trends in biopesticide development Please remove subsection 9.1 from this section, as there are no other subsections following it. Additionally, avoid using a single sentence as a paragraph; instead, merge it with the preceding paragraph. In this section on future perspectives and research directions, it would be valuable to discuss in more detail the role of cutting-edge technologies, such as CRISPR/Cas9 and RNA interference (RNAi), in pest control. Accepted and addressed! By combining CRISPR/Cas9 and RNAi, researchers can develop microbial biopesticides with enhanced specificity, efficacy, and environmental safety. These technologies offer exciting possibilities for the future of pest control and sustainable agriculture. Additionally, we have added “Future perspectives and research directions in the field of microbial biopesticides, including key research directions include Genetic engineering to improve microbial agents, synergistic interactions with other pest control methods, advanced formulation and delivery techniques, resistance management strategies, environmental impact assessments, integration into sustainable agriculture, and public awareness and adoption are essential for advancing their development and application. The future of microbial biopesticides looks promising, with ongoing research focusing on enhancing their efficacy, specificity, and environmental safety”. Section 10. Conclusion The conclusion effectively encapsulates the key findings regarding the benefits and challenges of biopesticides. However, the sentence stating that \"novel microbial strain creation with improved features, such as greater efficacy and larger target spectra, should be prioritized in future research and the use of biopesticides\" may be more appropriate in a future perspectives section. This sentence emphasizes the need for ongoing research and innovation, which aligns better with a section dedicated to future research directions. Yes but the conclusion part also needs to show the future perspectives in the area, so we keep this section as it is. Many thanks for the time you took to improve the paper. Sincerely, Kahsay Tadesse" } ] } ]	1	https://f1000research.com/articles/13-1071
https://f1000research.com/articles/13-270/v1	15 Apr 24	{ "type": "Research Article", "title": "Study anti-viral drugs for their efficiency against multiple SARS CoV-2 drug targets within molecular docking, molecular quantum similarity, and chemical reactivity indices frameworks", "authors": [ "Alejandro Morales-Bayuelo", "Jesús Sánchez-Márquez", "Ricardo Vivas-Reyes", "Savaş Kaya", "Jesús Sánchez-Márquez", "Ricardo Vivas-Reyes", "Savaş Kaya" ], "abstract": "The study focused on drug discovery for COVID-19, emphasizing the challenges posed by the pandemic and the importance of understanding the virus’s biology. The research utilized molecular docking and quantum similarity analyses to explore potential ligands for SARS-CoV-2 RNA-dependent RNA polymerase.\nDocking Results Docking outcomes for various ligands, including Oseltamivir, Prochloraz, Valacyclovir, Baricitinib, Molnupiravir, Penciclovir, Famciclovir, Lamivudine, and Nitazoxanide, were presented. Interactions between ligands and specific residues in the RNA-dependent RNA polymerase were analyzed.\n\nReactivity Descriptors Global parameters, such as electronic chemical potential, chemical hardness, global softness, and global electrophilicity, were computed for the ligands. For the local reactivity descriptors, the Fukui Functions were used. Fukui functions, representing electrophilic and nucleophilic sites, were calculated for selected ligands (Valacyclovir and Penciclovir). Nucleophilic character assignments for specific molecular regions were discussed, providing insights into potential charge-donating interactions.\n\nResults and Discussion Challenges in COVID-19 drug discovery, such as virus mutability, rapid evolution, and resource limitations, were summarized. Progress in vaccine development and the need for ongoing research to address variants and breakthrough cases were emphasized.\n\nOverlap Operator Analysis Higher MQSM between Lamivudine and Molnupiravir (0.5742) indicates structural and electronic similarity. Lowest MQSM between Oseltamivir and Prochloraz (0.2233) implies structural dissimilarity.\n\nCoulomb Operator Analysis Higher MQSM between Lamivudine and Molnupiravir (0.9178) suggests both structural and electronic similarity. Lowest MQSM between Baricitinib and Famciclovir (0.6001) indicates greater structural diversity. Measurements above 0.5 in Table 3 suggest electronic similarity, emphasizing the electronic aspects in molecular analysis. In this sense, it study employed a multi-faceted approach combining molecular docking, quantum similarity analyses, and chemical reactivity assessments to explore potential drug candidates for COVID-19. The findings provide valuable insights into ligand interactions, reactivity patterns, and the challenges associated with drug discovery in the context of the global pandemic.", "keywords": [ "SARS-CoV-2 virus", "COVID-19 treatments", "molecular docking", "molecular quantum similarity", "chemical reactivity indices", "Density Functional Theory." ], "content": "1. Introduction\n\nCOVID-19 has caused a significant global public health crisis, with millions of confirmed cases and deaths worldwide. Healthcare systems in various countries faced unprecedented challenges, including shortages of medical supplies, overwhelmed hospitals, and strain on healthcare professionals. While vaccines offered a pathway out of the pandemic, challenges such as vaccine hesitancy, distribution issues, and access disparities in lower-income countries are significant global concerns. The virus responsible for Coronavirus Disease (COVID-19), known as SARS-CoV-2, exhibits varying degrees of symptoms among patients, mailto:1.https://covid19.who.int/. While most individuals undergo mild to moderate symptoms and recover without specific treatment, some progress to severe cases necessitating medical attention.1 Transmission of the virus occurs through microscopic liquid particles expelled from an infected person’s mouth or nose during activities such as coughing, sneezing, speaking, singing, or breathing. These particles, ranging from small aerosols to larger respiratory droplets, can be transmitted through close contact with an infected person or by touching contaminated surfaces and subsequently touching the face.2\n\nAs an RdRp (RNA-dependent RNA polymerase), the virus heavily relies on this enzyme for the replication and transcription of its genome. This characteristic makes it an appealing target for the study of its biology and the development of antiviral drugs.3\n\nIn the effort to combat COVID-19, multiple drugs are under investigation, and thus far, the FDA has granted approval to only one—remdesivir (Veklury). This antiviral medication is utilized in the treatment of COVID-19 among adults and adolescents aged 12 and above, typically administered intravenously for hospitalized patients.4,5 Recognizing the significance of comprehending how these drugs stabilize the active site of the receptor structure, this study utilized Molecular Docking, Molecular Quantum Similarity (MQS), and global and local reactivity indices to assess remdesivir and other associated compounds. Examples include Oseltamivir, Prochloraz, Valacyclovir, Baricitinib, Molnupiravir, Valacyclovir, Penciclovir, Famciclovir, Lamivudine, and Nitazoxanide.\n\nThe MQS concept, introduced by Carbo-Dorca and colleagues, analyzes molecular similarities among different compounds. In this research, Density Functional Theory (DFT) was employed to bridge the gap between Molecular Quantum and Quantum Chemistry, combining molecular docking with chemical reactivity indices.5 This approach provided valuable insights into potential alternative treatments for COVID-19 and shed light on the interactions between approved drugs like remdesivir and other potential ligands.\n\nMolecular docking plays a crucial role in drug discovery for COVID-19 by facilitating the identification and design of potential therapeutic compounds. Here are some key aspects highlighting the importance of molecular docking in the context of COVID-19 drug discovery: Target Identification and Validation: Molecular docking helps identify potential drug targets within the SARS-CoV-2 virus, such as viral proteins critical for its replication and survival. It aids in validating the chosen targets by predicting the binding affinity and interaction patterns of small molecules with these targets.\n\nDocking simulations provide insights into the binding mechanisms between potential drug candidates and viral proteins. This information is crucial for understanding how a drug may interfere with the virus’s life cycle. Molecular docking predicts the binding affinity of a ligand to a target protein. Compounds with high binding affinity are more likely to have therapeutic effects, making them promising candidates for further experimental validation. Rational Drug Design: Docking studies guide rational drug design by providing a structural basis for modifying existing drugs or designing new compounds that specifically target essential viral proteins. Antiviral Drug Development: The identification of potential drug candidates through molecular docking contributes to the development of antiviral drugs targeting specific proteins crucial for the virus’s replication or entry into host cells. Cost and Time Efficiency: Computational approaches like molecular docking significantly reduce the time and costs associated with drug discovery by prioritizing the most promising compounds for experimental validation.\n\n\n2. Methods\n\nIn the docking experiment, the receptor structure was derived following specific protocols based on the crystal structure of SARS-CoV-2 RNA-dependent RNA polymerase with PDB code 6m71. Adjustments to the structure were made using the protein preparation wizard module from the Schrödinger suite 2017-1. These adjustments included:\n\ni) Optimization of the hydrogen bond (H-bond) network and refinement of the protein structure.\n\nii) Determination of protonation states at physiological pH using the PropKa utility.\n\niii) Execution of restrained molecular minimization through the Impact Refinement (Impref) module, with heavy atoms constrained to a low root-mean-square deviation (RMSD) from the initial coordinates (chrome-extension://efaidnbmnnnibpcajpcglclefindmkaj/https://www.modekeji.cn/wp-content/uploads/2019/08/gli55_user_manual.pdf).6–8\n\nConversely, the molecular structures of the compounds were constructed using the Maestro Editor (Maestro, version 11.1, Schrödinger, LLC). Subsequently, 3D conformations were generated using the LigPrep module, and ionization/tautomeric states were predicted under physiological pH conditions using Epik. Finally, energy minimization was performed with the Macro model using the OPLS2005 force field.\n\nGlide (https://newsite.schrodinger.com/platform/products/glide/)9,10 with default parameters and Standard Precision (SP) model was used for docking investigations. The docking grid was created using default settings, with the co-crystallized ligand in the center. For the van der Waals radii of the nonpolar protein atoms, a scaling factor of 0.8 was applied to facilitate the binding of larger ligands. Extra precision (XP) was also utilized to expand alternate receptor conformations appropriate for binding ligands with unusual orientations via induced fit docking (IFD); this method allows the protein to undergo side-chain, backbone, or both movements upon ligand docking. All results were redocking and RMSD were performed. The binding pocket of the RdRp—GLY616, TRP617, ASP618, TYR619, LEU758, SER759, ASP760, ASP761, ALA762, LYS621, TYS799, TRP800, GLU811, PHE812, CIS813, and/or SER814—was found using Glide.9,10\n\nThe docking process involves four precise steps, relying on Glide’s scoring function and Prime’s advanced conformational refinement to ensure accuracy:\n\n(i) Initial docking using Glide is executed on the rigid receptor to generate a set of poses.\n\n(ii) The side-chain prediction module of the Prime module is employed to sample the protein, followed by structural minimization for each pose of the protein/ligand complex.\n\n(iii) Redocking of the ligand into the low-energy induced-fit structures from the previous step is conducted using Glide’s default parameters (without vdW scaling).\n\n(iv) The binding energy (IFDScore) is estimated, considering the docking energy (GScore), receptor strain, and solvation terms (Prime energy).\n\nTo further assess the interactions of the ligands in the active site, the extent of residue movement induced by the IFD computation is considered. For both the most and least active ligands, all poses are compared within the molecular set. Molecular dynamics calculations over 30ns are employed to predict the best poses and analyze their stabilization in the active site.\n\n\n3. Quantum Similarity Analyses\n\nA Molecular Quantum Similarity Measure (MQSM) amid two A and B systems, known as ZAB, compares two molecules that may be created using their respective Density Functions (DFs).\n\nBoth DFs can be multiplied and integrated in terms of their electronic coordinates, which are then weighted using a predetermined positive operator Ω(r1, r2)11–13:\n\nThe operator used in Equation 1 plays a crucial role in determining the information being compared and serves as the measure of similarity between the two systems. For example, when the operator chosen is the Dirac delta function, it proves to be an efficient approach for functions with high peak values, like the electronic density. Moreover, it provides a similar mathematical abstraction as a charge or point mass, i.e., Ω(r1, r2) = δ(r1 - r2). One of the first similarity metrics employed is the overlapping MQSM; another widely used alternative is the Coulomb operator, i.e., Ω(r1, r2) = \|‌‌r1 - r2\|‌‌−1, resulting in a Coulombic MQSM. Even if the two molecules are equivalent, a similarity measure can be employed for any two molecular systems; this measurement is known as a self-similarity measure (ZAA for the case of molecule A).12\n\nFor a given group of N molecules, we can derive a measure of similarity for each molecule concerning the others in the group, including itself. These similarity measurements can then be used to construct a symmetric matrix. The i-th column of this matrix represents a compilation of all similarity measurements between the i-th molecule and every constituent in the group, including itself. Consequently, each vector (matrix column) serves as a discrete N-dimensional representation of the i-th structure. These vector sets can be demonstrated as a set of chemical descriptors. However, this set of similarity matrix columns goes beyond merely representing another set of molecular descriptors, as commonly done for theoretical molecule description; each descriptor possesses unique properties.12–20\n\ni) It is universal, deriving from any collection of molecules and any individual molecule within that group.\n\nii) It is impartial, as there are no other possibilities available in the construction process than those dictated by the density functions and similarity measurements involved.\n\n\n\nCarbó’s similarity index between two molecules I and J are constructed from Equation 2. Because this index is also known as the cosine similarity index, it corresponds to the cosine of the angle included by the density functions involved when considered as vectors. For any pair of compared molecules, this Carbo QSI has a value between 0 and 1, depending on the similarity between the two molecules (when I = J, the index approaches 1).13–28\n\nTaking into account the similarity of equation 3:\n\nIt is simplified to the so-called Euclidean distance index when k = x = 2. Index 3 of the form can also be defined as follows:\n\nThis Equation 4 forms the distance index of infinite order.18–30\n\nThe distribution of Dirac’s delta, Ω (r1, r2) = δ (r1, r2), is the most typical and intuitive choice for such a positively defined operator. These selections transform the broad definition of MQSM to compute the overlap MQSM that obtains measurements of the volume by both electronic density functions when they are superimposed.17–20\n\nThe Dirac delta function is derived instinctively from the physical definition, and it is computationally compliant. The MQSM calculates the degree of overlap between molecular comparisons using information about the electron concentration in the molecule.16–21\n\nWhen the operator (Ω) is replaced with the Coulomb operator, Ω (r1, r2)= 1│r1−r2│, the coulomb MQS is generated, which defines the electrostatically repellent coulomb energy between two charge densities20,21:\n\nThe coulomb operator affects the overlap density functions. When considering molecular density functions as an electron distribution in space, this equation is simply an extension of the coulomb operator for the distribution of continuous charge, thus can be used as electrostatic potential descriptors in some instances. This operator is correlated to electrostatic interactions and obtains the measurement of electrostatic repulsion between electronic distributions.30–37\n\nAnother major transformation that can be expressed in terms of the classical distance is:\n\nHere Δxj=xaj−xbj is the distance between a and b, and k = 2 is the definition of distance, respectively. Subsequently, the Euclidean distance between A and B as two quantum objects are defined by17–21:\n\nOccasionally it is written as: DAB=ZAA+ZBB+ZZAB, where DAB has values in the range of [0,∞) but for earlier circumstances where the compared items are identical, it converges to zero between them.17–21:\n\nThe norm of the differences in the density functions of the compared objects can be used to interpret this index geometrically. The distance or dissimilarity index can be used to define the Euclidean distance index, which can also be represented as21–25:\n\nIn this investigation, the TGSA (Typical Geometry Superposition Algorithm) approach was utilized for data alignment. Devised by Gironés, TGSA operates under the assumption that the optimal way to align molecules involves superimposing them onto a shared skeleton, considering solely the atomic types and interatomic bonding interactions based on the atomic number coordination.23–28\n\nThe program initiates by scrutinizing pairs of atoms in the molecules, aligning their common substructure for a group of molecules using topological and geometrical considerations. Notably, the superposition achieved is distinctive and unaffected by the choice of similarity measure.28–34\n\nInitially, the program organizes molecular coordination into bases based on the reduction of atomic numbers, defining a path for the number of hydrogens in the molecule (excluding hydrogen atoms for computational efficiency). Subsequently, atomic pairs are delineated, specifying the involved atoms and their respective distances.34–37 Translocations are identified through changes in the conformations’ spine caused by substitutions in the molecules.23 Bones not aligning with the skeletons are eliminated during this process.\n\nThe program then assembles atomic triads by incorporating three atoms from the compared pairs, forming a triangle in the plane representing the chemical box’s efficacy.23,24 The triangles generated for two molecules are compared using their respective interatomic and translational distances. Triads meeting the classification criteria are retained, superimposed, and dictate the molecular alignment result.23\n\nIt’s worth noting that since TGSA characterizes molecules as rigid structures without flexibility (no vibration or rotation in bond distances and angles), it may not yield optimal results for diverse molecular structures due to the restricted alignment with the common recognition skeleton. Nevertheless, this method consistently aligns with chemical intuition and is favored for its accessibility and lower computational requirements.26–37\n\n\n4. Chemical Reactivity Analysis\n\nStudies in the field have demonstrated an established link between quantum similarity and descriptors of chemical reactivity.38,39 Both quantum similarity and DFT employ the density function as a key element in examining similarity indices. Specifically, the Coulomb index can be associated with electronic aspects that influence chemical reactivity. To determine global reactivity indices such as chemical potential (μ),27 hardness (ɳ),38 and electrophilicity (ω),39,40 Frontier Molecular Orbitals (FMO) and the energy gap will be utilized for computation. These indices (Equations 11-13) offer valuable insights into the stability of systems, with chemical potential gauging electron tendency to depart from the equilibrium system,41 while chemical hardness assesses a chemical species’ resistance to altering its electronic configuration.\n\nThe mathematical definition of the electrophilicity index (ω) is related to the stabilization energy of a system when it becomes saturated by electrons from the external environment.39,40:\n\nIn this research, the local reactivity descriptors under consideration were the Fukui functions. The Equations42,43 illustrate the system’s electronic density response to variations in the global charge, representing the derivative of the electronic density concerning the electron count under a consistent external field.\n\nThe terms f+r→ and f−r→ have been employed to denote nucleophilic and electrophilic attacks, respectively.31–33 This approach utilizes both global and local reactivity descriptors to examine quantum similarity within the molecular set. All computations were conducted using the B3LYP method44 with the 6-311XXG(d,p) basis set,45 which involves an improvement to the 6-311G(d) basis set. This enhancement allows for calculations of electronegativity, hardness, reactivity indices, and frontier molecular orbitals at a quality level comparable to much larger basis sets like Aug-cc-pVQZ and Aug-cc-pV5Z. The Gaussian 16 package46 was employed in conjunction with this method/basis set combination.\n\n\n5. Results and Discussion\n\nDrug discovery for COVID-19 presents several challenges, and researchers worldwide have been working diligently to address these issues. Some of the key challenges include: Virus Mutability: SARS-CoV-2, the virus responsible for COVID-19, can mutate, leading to the emergence of new variants. This mutability poses a challenge in developing drugs that can effectively target different strains of the virus. Rapid Evolution of the Pandemic: The rapid spread of the virus and the urgent need for effective treatments make it challenging to follow traditional drug development timelines. Accelerated timelines can compromise thorough testing and validation processes. Lack of Pre-existing Therapies: Unlike some other infectious diseases, there were no pre-existing drugs specifically designed to target SARS-CoV-2. Developing new drugs from scratch is a time-consuming process.\n\nComplexity of the Virus Life Cycle: Understanding the intricate details of the virus’s life cycle and the host-pathogen interactions is essential for developing targeted therapies. This complexity requires a deep understanding of virology and immunology. Drug Safety: Ensuring the safety of potential treatments is crucial. Some drugs may show promise in early stages but could have adverse effects or interactions with other medications, requiring extensive testing for safety. Drug Delivery Challenges: Designing effective drug delivery systems to ensure that the drug reaches the target tissues in sufficient concentrations is a significant challenge. This is especially important for antiviral drugs targeting the respiratory system. Antibody Resistance: The virus may develop resistance to certain antiviral drugs or antibodies over time. This highlights the need for ongoing research to identify multiple targets for drug development and combination therapies. Global Collaboration: International collaboration is crucial for sharing data, resources, and expertise. However, coordinating efforts across borders and overcoming logistical and political challenges can be complex. Vaccine Success and Impact: The success and widespread distribution of COVID-19 vaccines have been crucial in controlling the pandemic. However, ongoing research is needed to address vaccine effectiveness against new variants and to develop treatments for breakthrough cases. Resource Limitations: Drug discovery requires significant financial and human resources. The COVID-19 pandemic has strained healthcare systems globally, and prioritizing and allocating resources for research and development can be challenging. Despite these challenges, the scientific community has made remarkable progress in a short time, developing vaccines and exploring various therapeutic approaches. Continuous research and collaboration will be essential for addressing the evolving nature of the pandemic and improving our ability to manage and treat COVID-19. For these reasons, in this study are obtained new insights for a serie of ligands, based on the crystal structure of SARS-CoV-2 RNA-dependent RNA polymerase with PDB code 6m71. Please, see Figures 1 and 2.\n\nIn Figure 1 are shows the docking outcomes for Oseltamivir. The main interaction is with the residues GLU166 (-H, 1.323 Å), GLN189 (-H, 1.185 Å) and GLY143 (-H, 1.054 Å). However, in Figure 2 for Prochloraz has a stacking (also called π–π stacking, (1.213 Å)) with the residues HIE41.\n\nThe Figure 3 shows the docking results for Valacyclovir, this ligand shows interactions with the residues CYS145 (-H, 1.452 Å), SER144 (-H, 1.114 Å), GLY143 (-H, 1.156 Å) and HIS163 (-H, 1.254 Å).\n\nIn Figure 4 show the interaction for Baricitinib, this Figure shows interactions with the residues GLU166 (-H, 1.568 Å), GLY143 (-H, 1.365 Å). Unlike, in the Figure 5 we can see interactions for Molnupiravir with the residue CYS145 (-H, 1.248 Å and 1.238 Å).\n\nIn Figure 6, shows the interactions for Valacyclovir with the residue HIS163 (-H, 1.156 Å) and CYS145 (-H, 1.256 Å). The Figure 7, shows the interactions for Penciclovir with the residues HIS164 (-H, 1.005 Å) and π-π interactions with the residues HIE41 (-H, 1.269 Å)\n\nIn the Figures 8 and 9, shows the interactions with the residues HIE41 (-H, 1.567 Å) for Penciclovir and with the residues GLY143 (-H, 1.456 Å), CYS145 (-H, 1.436 Å) and HIS163 (-H, 1.485 Å).\n\nFinally, in the Figures 10 and 11 we can see the interactions with the residues GLU166 (-H, 1.054 Å) and GLY143 (-H, 1.158 Å) for Lamivudine and with the residues GLU166 (-H, 1.254 Å), HIE41 (-H, 1.266 Å) and GLY143 (-H, 1.354 Å) for Nitazoxanide.\n\nMQSM helps in the identification of molecules with similar quantum properties, which can be useful for finding potential drug candidates. Similar quantum properties suggest similar chemical behaviours. MQSM involves the calculation of quantum descriptors, which are numerical representations of molecular properties derived from quantum mechanical calculations. These descriptors may include electronic density, electron localization function, and others. Various similarity measures, such as Carbo’s similarity index or Euclidean distance, are employed to quantify the degree of similarity between quantum descriptors of different molecules MQSM can aid in lead optimization by identifying compounds with quantum properties like known drug candidates. This assists in designing molecules with improved pharmacological profiles.\n\nThe MQSM can be used to understand the interactions between ligands and their target receptors at a quantum level. This is crucial for rational drug design. It is often used in conjunction with molecular docking studies. While docking predicts the binding affinity and geometry of ligands with target proteins, MQSM provides insights into the quantum properties that influence these interactions.\n\nMQSM offers a detailed and atomistic understanding of molecular properties, allowing for a more nuanced analysis of chemical similarity. Unlike traditional structural similarity measures, MQSM considers the quantum properties of molecules, providing a more comprehensive comparison.\n\nIn the context of antiviral drug discovery, MQSM can be employed to identify molecules with similar quantum properties to known antiviral drugs, aiding in the search for new therapeutics. Therefore, Molecular Quantum Similarity analysis is a valuable tool in drug discovery that leverages quantum mechanical principles to assess the similarity of molecular properties. This approach contributes to the rational design of novel drugs and the optimization of lead compounds for improved pharmacological profiles.\n\nIn Tables 1 and 2. the higher MQSM is between the compounds Lamivudine and Molnupiravir using the Overlap operator 0,5742 with a euclidean distance of 4,2364, see Table 2. The lowest MQSM is between the compounds Oseltamivir and Prochloraz (0,2233) with a eclidean distance of 5,5841 (see Table 2). These measurements are below 0.5, according to the range of carbon indices (0.1]. So we can say that structurally they are quite different.\n\nIn Tables 3 and 4 the higher MQSM using the Coulomb operator is between the compounds Lamivudine and Molnupiravir (0,9178) with a euclidean distance of 22,5461, see Table 4. However, the lowest MQSM is between the compounds Baricitinib and Farmciclovir (0,6001) with a euclidean distance of 44,3298. The comparison between the compounds Lamivudine and Molnupiravir has the higher values in both case using the Overlap and Coulomb operators. Unlike to the MQSM using the overlap operator, the measurements of the Table 3 are above 0.5, according to the range of carbon indices (0.1]. Therefore, we can say that although structurally they are not so similar, electronically they are. Because the electronic similarity index is higher with respect to the overlap, we have analyzed in depth the electronic properties of the analyzed ligands.\n\nThe investigation explored into global and local chemical reactivity descriptors through DFT computations. This part contrasts the reactivity of the ligands within the study, encompassing both overarching parameters and locally descriptive functions of reactivity. Electrophilicity values hold potential significance in stabilizing the active site of ligands engaged in non-covalent interactions. Illustrated in Figure 12 are the computed global parameters—chemical potential, chemical hardness, global softness, and global electrophilicity—offering a comparative analysis of the ligand sample’s chemical reactivity. As depicted in Figure 12, Nitazoxanide emerges as the most reactive molecule, displaying the lowest values of electronic chemical potential (μ) and chemical hardness (η), alongside the highest global softness and global electrophilicity. Conversely, the remaining compounds exhibit descriptor values that display less pronounced differences. Consideration of solely global descriptors might suggest comparable reactivity. The values obtained in each case are detailed in Table S1 in the accompanying supplementary information.\n\nSince the analysis of the global parameters is limited, we will complete it with the comparison of some local descriptor functions. The electrophile and nucleophile Fukui functions (as a measure of reactivity) were then compared using the Frontier Molecular Orbital (FMO) approach. The electrophilic-nucleophilic character of the following functions also shows those molecular areas that are most likely to form charge-donating interactions (basically by charge delocalisation). These types of interactions are important and difficult to determine using docking analysis. Figure 13 shows Fukui function f−r→ calculated under the FMO approximation (HOMOr→2) for the compounds Valacyclovir and Penciclovir (A and B respectively), it can be noted that f−r→ is similar in both compounds, and in both cases, the function assigns the most nucleophilic character to the condensed rings. In Figures S1-S8, see Underlying Data, the f−r→ functions for all ligands in the study can be seen. Comparing the HOMOr→2 function from Figure 13-B with Figure 8, it can be observed that the interactions of Penciclovir with GLY 143, HIE 41, and LEU 141 obtained through docking are compatible with the f−r→ function.\n\nIsovalue was 0.008 in both cases. The figure was created using GaussView 5.0.\n\nFigure 14 the functions f+r→ calculated under the FMO approximation (LUMOr→2) for compounds A) Prochloraz, B) Molnupiravir and C) Lamivudine. It can be noted that f+r→ is similar in the three compounds; in all three cases, the function assigns the most nucleophilic character to various carbons in the six-carbon ring. The ring may have some ability to rotate and orient itself towards where it can form the strongest interactions. Comparing Figure 14 with Figure 2, it can be seen that f+r→ is compatible with the interaction of Prochloraz with HIE 41. A similar conclusion is drawn when comparing Figure 14 with Figure 5, in this case, the interaction is formed between Molnupiravir and PHE 140. Lastly, comparing Figure 14 with Figure 10 highlights the interaction between Lamivudine and GLU 166.\n\nThe isovalue was 0.008 in all cases. The figure was created using GaussView 5.0.\n\nFigure 15 shows the functions f+r→ calculated under the FMO approximation (LUMOr→2) for compounds A) Valacyclovir and B) Penciclovir. It can be noted that f+r→ is similar in the two compounds, and in both cases, the function assigns the most nucleophilic character to the -NH2 group in the condensed rings. In Figures S1-S8, see Underlying Data, mailto:https://doi.org/10.7910/DVNs/7KFPUT, Harvard Dataverse, V1., the f+r→ functions for all ligands in the study can be seen.\n\nThe isovalue was 0.008 in both cases. The figure was created using GaussView 5.0.\n\n\n6. Conclusions\n\nThe study on ligands based on the crystal structure of SARS-CoV-2 RNA-dependent RNA polymerase with PDB code 6m71 provides valuable insights into potential drugs for COVID-19. The conclusions highlight the challenges in drug discovery for COVID-19, including virus mutability, the rapid evolution of the pandemic, lack of pre-existing therapies, complexity of the virus life cycle, drug safety, drug delivery challenges, antibody resistance, global collaboration, vaccine success, and resource limitations. Despite these challenges, the scientific community has made remarkable progress in developing vaccines and exploring therapeutic approaches.\n\nThe docking results for various ligands, including Oseltamivir, Prochloraz, Valacyclovir, Baricitinib, Molnupiravir, Penciclovir, Famciclovir, Lamivudine, and Nitazoxanide, reveal specific interactions with key residues of the SARS-CoV-2 RNA-dependent RNA polymerase. These interactions provide valuable information about the potential efficacy of these ligands in inhibiting the virus.\n\nThe global reactivity descriptors analysis, including electronic chemical potential, chemical hardness, global softness, and global electrophilicity, offers a comparative analysis of the ligands’ chemical reactivity. Nitazoxanide emerges as the most reactive molecule, displaying the lowest electronic chemical potential and chemical hardness, along with the highest global softness and global electrophilicity. The other compounds show less pronounced differences in their reactivity.\n\nThe Fukui function comparison provides additional insights into the local chemical reactivity of the ligands. For example, the comparison of Fukui functions for Valacyclovir and Penciclovir shows similar nucleophilic character in both compounds, particularly in the condensed rings.\n\nThe study is comprehensive, integrating docking results with global and local reactivity descriptors to assess the potential of various ligands as drugs for COVID-19. The information presented contributes to our understanding of the interactions between ligands and the SARS-CoV-2 RNA-dependent RNA polymerase, aiding in the development of targeted therapies.\n\nThe higher MQSM is observed between Lamivudine and Molnupiravir, indicating electronic similarity. However, the comparison between Oseltamivir and Prochloraz suggests lower electronic similarity, implying structural dissimilarity. The Coulomb operator reveals higher electronic similarity between Lamivudine and Molnupiravir, consistent with the overlap operator. The lower electronic similarity between Baricitinib and Famciclovir suggests greater structural diversity. In both cases, the electronic similarity indices using the Coulomb operator are higher than those using the overlap operator, emphasizing the electronic aspects in the molecular analysis. Further investigation into the electronic properties of the ligands is warranted. These findings enhance our understanding of the molecular properties and guide future drug discovery efforts based on both structural and electronic considerations.", "appendix": "Data availability\n\nHarvard Dataverse: Replication data for Study of a series of ligands used as inhibitors of the SARS-CoV-2 virus.\n\nMorales-Bayuelo, Alejandro, 2023, “Replication data for Study of a series of ligands used as inhibitors of the SARS-CoV-2 virus”, https://doi.org/10.7910/DVN/7KFPUT. 47\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\nAMB thanks to the Universidad del Sinú, Seccional Cartagena.\n\n\nReferences\n\nConsulted 18-12-2023. Reference Source\n\nBanerjee A, Kulcsar K, Misra V, et al.: Bats and Coronaviruses. Viruses. 2019; 11. pii: E41. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYang D, Leibowitz JL: The structure and functions of coronavirus genomic 3′ and 5′ ends. Virus Res. 2015; 206: 120–133. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSong Z, Xu Y, Bao L, et al.: From SARS to MERS, Thrusting Coronaviruses into the Spotlight. Viruses. 2019; 11. pii: E59. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\n[(a)] Graham RL, Donaldson EF, Baric RS: A decade after SARS: strategies for controlling emerging coronaviruses. Nat. Rev. Microbiol. 2013; 11: 836–848. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\n[(b)] Morales-Bayuelo A, Sánchez-Márquez J: New findings on ligand series used as SARS-CoV-2 virus inhibitors within the frameworks of molecular docking, molecular quantum similarity and chemical reactivity indices. F1000Res. 2023; 11: 914. 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Molecular Similarity and Reactivity: From Quantum Chemical to Phenomenological Approaches. Carbó R, editor. Vol. 14. . Amsterdam: Kluwer Academic Publishers; 1995; pp. 3–30. Understanding Chemical Reactivity. Publisher Full Text\n\n[(a)] Morales-Bayuelo A, Matute RA, Caballero J: Understanding the comparative molecular field analysis (CoMFA) in terms of molecular quantum similarity and DFT-based reactivity descriptors. J. Mol. Model. 2015; 21: 156. PubMed Abstract \| Publisher Full Text\n\n[(b)] Morales-Bayuelo A: Ricardo Vivas-Reyes. J. Quant. Chem. 2014; 2014: 1–12. Article ID 850163. Publisher Full Text\n\n[(c)] Morales-Bayuelo A, Vivas-Reyes R: Topological model to quantify the global reactivity indexes as local in Diels–Alder reactions, using density function theory (DFT) and local quantum similarity (LQS). J. Math. Chem. 2013; 51: 125–143. Publisher Full Text\n\n[(d)] Morales-Bayuelo A, Vivas-Reyes R: Theoretical model for the polarization molecular and Hückel treatment of PhosphoCyclopentadiene in an external electric field: Hirschfeld study. J. Math. Chem. 2013; 51: 1835–1852. Publisher Full Text\n\n[(e)] Morales-Bayuelo A, Valdiris V, Vivas-Reyes R: J. Theor. Chem. 2014; 14: 1–13.\n\n[(f)] Morales-Bayuelo A, Torres J, Baldiris R, et al.: Theoretical study of the chemical reactivity and molecular quantum similarity in a series of derivatives of 2‐adamantyl‐thiazolidine‐4‐one using density functional theory and the topo‐geometrical superposition approach. Int. J. Quantum Chem. 2012; 112: 2681–2687. Publisher Full Text\n\n[(g)] Morales-Bayuelo A, Torres J, Vivas-Reyes R: Quantum molecular similarity analysis and quantitative definition of catecholamines with respect to biogenic monoamines associated: Scale alpha and beta of quantitative convergence. Int. J. Quantum Chem. 2012; 112: 2637–2642. 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Publisher Full Text\n\nSchipper PRT, Gritsenko OV, van Gisbergen SJA , et al.: Molecular calculations of excitation energies and (hyper)polarizabilities with a statistical average of orbital model exchange-correlation potentials. J. Chem. Phys. 2000; 112: 1344–1352. Publisher Full Text\n\nCarbó-Dorca R, Leyda L, Arnau M: How similar is a molecule to another? An electron density measure of similarity between two molecular structures. Int. J. Quantum Chem. 1980; 17: 1185–1189. Publisher Full Text\n\nCarbó-Dorca R, Gironés X: Foundation of quantum similarity measures and their relationship to QSPR: Density function structure, approximations, and application examples. Int. J. Quantum Chem. 2005; 101: 8–20. Publisher Full Text\n\nBultinck P, Gironés X, Carbó-Dorca R: Molecular Quantum Similarity: Theory and Applications. Rev. Comput. Chem. 2005; 21: 127. Publisher Full Text\n\nConstans P, Amat L, Carbó-Dorca R: Toward a global maximization of the molecular similarity function: Superposition of two molecules. J. Comput. Chem. 1997; 18: 826–846. Publisher Full Text\n\nCarbó-Dorca R, Mercado LD: Commentaries on quantum similarity (1): Density gradient quantum similarity. J. Com. Chem. 2010; 31: 2195–2212. Publisher Full Text\n\nGironés X, Carbó-Dorca R: Modelling Toxicity using Molecular Quantum Similarity Measures. QSAR Combinator. Sci. 2006; 25: 579–589. Publisher Full Text\n\nCarbó-Dorca R, Besalú E, Mercado LD: Communications on quantum similarity, part 3: A geometric‐quantum similarity molecular superposition algorithm. J. Com. Chem. 2011; 32: 582–599. PubMed Abstract \| Publisher Full Text\n\nCarbó-Dorca R, Gironés X: Foundation of quantum similarity measures and their relationship to QSPR: Density function structure, approximations, and application examples. Int. J. Quantum Chem. 2005; 101: 8–20. Publisher Full Text\n\nCarbó-Dorca R, Besalú E: Communications on quantum similarity (2): A geometric discussion on holographic electron density theorem and confined quantum similarity measures. J. Comput. Chem. 2010; 31: 2452–2462. PubMed Abstract \| Publisher Full Text\n\nMorales-Bayuelo A, Ayazo H, Vivas-Reyes R: Europ. J. Med. Chem. 2010; 45: 4509.\n\nMorales-Bayuelo A, Torres J, Vivas-Reyes R: Quantum molecular similarity analysis and quantitative definition of catecholamines with respect to biogenic monoamines associated: Scale alpha and beta of quantitative convergence. Int. J. Quantum Chem. 2012; 112: 2637–2642. Publisher Full Text\n\nMorales-Bayuelo A, Torres J, Baldiris R, et al.: Theoretical study of the chemical reactivity and molecular quantum similarity in a series of derivatives of 2‐adamantyl‐thiazolidine‐4‐one using density functional theory and the topo‐geometrical superposition approach. Int. J. Quantum Chem. 2012; 112: 2681–2687. Publisher Full Text\n\nMorales-Bayuelo A, Torres J, Vivas-Reyes R: HÜCKEL TREATMENT OF PYRROLE AND PENTALENE AS A FUNCTION OF CYCLOPENTADIENYL USING LOCAL QUANTUM SIMILARITY INDEX (LQSI) AND THE TOPO-GEOMETRICAL SUPERPOSITION APPROACH (TGSA). J. Theo. Comp. Chem. 2012; 11: 223–239. Publisher Full Text\n\nMorales-Bayuelo A, Vivas-Reyes R: Topological model to quantify the global reactivity indexes as local in Diels–Alder reactions, using density function theory (DFT) and local quantum similarity (LQS). J. Math. Chem. 2013; 51: 125–143. Publisher Full Text\n\nMorales-Bayuelo A, Vivas-Reyes R: Theoretical model for the polarization molecular and Hückel treatment of Phospho-Cyclopentadiene in an external electric field: Hirschfeld study. J. Math. Chem. 2013; 51: 1835–1852. Publisher Full Text\n\nMorales-Bayuelo A, Baldiris R, Vivas-Reyes R: Scale Alpha and Beta of Quantitative Convergence and Chemical Reactivity Analysis in Dual Cholinesterase/Monoamine Oxidase Inhibitors for the Alzheimer Disease Treatment using Density Functional Theory (DFT). J. Theor. Chem. 2013; 2013: 1–13. Publisher Full Text\n\nMorales-Bayuelo A, Vivas-Reyes R: Theoretical Calculations and Modeling for the Molecular Polarization of Furan and Thiophene under the Action of an Electric Field Using Quantum Similarity. J. Quant. Chem. 2014; 2014: 1–10. Article ID 585394. Publisher Full Text\n\nMorales-Bayuelo A, Vivas-Reyes R: Topological Model on the Inductive Effect in Alkyl Halides Using Local Quantum Similarity and Reactivity Descriptors in the Density Functional Theory. J. Quant. Chem. 2014; 2014: 1–12. Article ID 850163. Publisher Full Text\n\nMorales-Bayuelo A, Valdiris V, Vivas-Reyes R: Mathematic analysis on a series of 4-Acetylamino-2-(3,5-dimethylpyrazol-1-yl)-6-pyridylpyrimidines: a simple way to relate quantum similarity with local chemical reactivity using the Gaussian orbitals localized theory. J. Theor. Chem. 2014; 14: 1–13.\n\nMorales-Bayuelo A, Vivas-Reyes R: Understanding the Polar Character Trend in a Series of Diels-Alder Reactions Using Molecular Quantum Similarity and Chemical Reactivity Descriptors. J. Quant. Chem. 2014; 2014: 1–19. Article ID 239845. Publisher Full Text\n\nParr RG, Pearson RG: Absolute hardness: companion parameter to absolute electronegativity. J. Am. Chem. Soc. 1983; 105: 7512–7516. Publisher Full Text\n\nGeerlings P, De Proft F, Langenaeker W: Conceptual density functional theory. Chem. Rev. 2003; 103: 1793–1874. Publisher Full Text\n\nChattaraj PK, Sarkar U, Roy DR: Electrophilicity index. Chem. Rev. 2006; 106: 2065–2091. Publisher Full Text\n\nParr RG, Szentpaly L v, Liu S: Electrophilicity Index. J. Am. Chem. Soc. 1999; 121: 1922–1924. Publisher Full Text\n\nContreras RR, Fuentealba P, Galván M, et al.: A direct evaluation of regional Fukui functions in molecules.Chem. Phys. Lett.1999; 304(5–6): 405–413. ISSN 0009-2614. Publisher Full Text\n\nYang W, Mortier WJ: The use of global and local molecular parameters for the analysis of the gas-phase basicity of amines. J. Am. Chem. Soc. 1986 Sep 1; 108(19): 5708–5711. PubMed Abstract \| Publisher Full Text\n\nFuentealba P, Pérez P, Contreras R: On the condensed Fukui function. J. Chem. Phys. 2000; 113: 2544–2551. Publisher Full Text\n\nZhao Y, Truhlar DG: The M06 suite of density functionals for main group thermochemistry, thermochemical kinetics, non-covalent interactions, excited states, and transition elements: two new functionals and systematic testing of four M06-class functionals and 12 other functionals. Theor. Chem. Accounts. 2008; 120: 215–241. Publisher Full Text\n\nFree Software: Gaussian 16, Revision B.01Frisch MJ, Trucks GW, et al.: GaussView 5.0.Wallingford, E.U.A.: Gaussian Inc.; 2016.\n\nMorales-Bayuelo A: Replication data for Study of a series of ligands used as inhibitors of the SARS-CoV-2 virus. [Dataset]. Harvard Dataverse. 2023; V1. Publisher Full Text" }	[ { "id": "292812", "date": "13 Jul 2024", "name": "Sajjan Rajpoot", "expertise": [ "Reviewer Expertise Inflammation and Immunity", "Drug Designing", "Cancer Immunotherapy", "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\nThe manuscript discussed the SARS-CoV-2 RdRp as the drug target for COVID-19 and attempted to study a set of anti-viral drugs including Oseltamivir, Prochloraz, Valacyclovir, Baricitinib, Molnupiravir, Penciclovir, Famciclovir, Lamivudine, and Nitazoxanide to highlight their different binding pattern with RdRp through molecular docking, and performed the drug structures comparison through molecular quantum similarity measure (MQSM) and their chemical reactivity and Fukui function comparison using multiple parameters. The manuscript suggest that ligand parameters are crucial in drug designing and these parameters of anti-viral drug including structural basis and their interaction pattern gives an insight to be considered for future drug designing. The effort of the authors is appreciated. Manuscript has valuable information for the readers. However, the following are the concerns and my opinion which is further suggested for authors to implement.\nMajor\nTitle:\nThe tile of the manuscript needs revision. It has mentioned multiple SARS-CoV-2 drug targets which is confusing for readers. Do authors mean multiple target sites within RdRp or multiple target proteins/enzymes of SARS-CoV-2? The title can also include “SARS-CoV-2 RdRp” to be specific as target protein.\n\nAbstract:\nThe Abstract needs revision. Either write the abstract as a single paragraph running text mentioning all the key sections of the study or structure well in category like background, methods, results and discussion and conclusion. Please include the key results of the manuscript and conclusive statement at the end.\n\nSpell out all the abbreviations used in abstract.\n\nIntroduction: Overall, the introduction needs re-organization.\nCite more recent and relevant references on COVID-19 epidemiology, symptoms in first paragraph. Authors should briefly include structural introduction for SARS-CoV-2 such as genome size, their key structural and non-structural proteins and further emphasize on their target protein RdRp in detail (such as its protein size, domains, active site, etc.) and why it is important to target. SRAS-CoV-2 RdRp has been studied well in recent and many in-silico studies are performed including molecular docking and molecular dynamic simulation. Authors should cite some previous studies and write how their study differ from it. Third paragraph, please cite few references with drugs under investigation. In few lines, summarize the study plan, what you have performed and what question this study is answering at the end of the introduction as closing remark. Overall, include more citations for each paragraph in introduction.\n\nMethods:\nSection 2.1-Please add the hyperlink for PDB ID 6m71 next to it for easy access. Section 2.1- Please mention the total number of compounds used in the study and list out all the compounds either in main manuscript or a table/excel sheet in supplementary data.\n\nResults and Discussion\nOverall, the results in section 5.1 are not well written and discussed. Authors needs to focus more on their outcomes-based text in the result and discussion section. Authors should give the rational for molecular docking study in the beginning and conclude their docking result with their finding before moving to next study in section 3.1. As mentioned above, write how many compounds were total docked with RdRp and what analysis led to identify the best compounds and binding pose of the complex? Are the representative figures (Fig 1-11) being the best binding ligands? Are interacting residues of RdRp with ligands mentioned in manuscript are crucial in limiting the RdRp function? Include these in your results. Section 5.2- please co-relate how the outcome of the molecular docking complement with your subsequent study in the MSQM section. Does the similarity analysis of ligands co-relate with the binding pose and binding strength/affinity of the ligands within the RdRp?\n\nFor Fig 1-11, it has two sub-figures for each, as 3D and 2D model. Authors should label them as A, B.. or i, ii.. as per journal format. For Fig 1-11, besides the main figure title, add well explained figure legend for each sub-section along with color codes, the visualization tool and its version used for representative figure preparation. For Fig 1-11, authors may also add the color label obtained from their docking visualization tool defining the type of interactions between ligand and target protein. Authors have mentioned about the binding energy (IFDScore) and docking energy (GScore) in methods. How does these energies were useful in your study to understand the binding strength of the ligands and comparison between them besides the interaction with RdRp residues. Please write in your results. Also, the energies for the docking complex can be summarized and presented in a table for better understanding. Also, this will help to understand the binding strength in comparison to structure similarity. In continuation to above comment (f), considering Section 2.2 and section 5.1, authors have mentioned about the molecular dynamics calculation over 30s to predict the best poses. Please elaborate on that. Does it mean the molecular dynamics simulation performed on the RdRp and drug complex? Also, the same should be explained in result section. Please write the result of the calculation that how it supported in finding the best pose. Section 5.3- Fig.12, The graph needs improvement. If possible, the authors are suggested to use Prism-GraphPad/Origin or other similar software for representative graph preparation. Overall, please make sure the graphs are uniform in terms of representation-like the x-axis and y-axis font size should be same. Fig.12A, the x-axis label (drugs name) is below the top x-axis line, please correct. Add axis lines for each graph, remove the inside grid lines. Increase the font size, it’s not clearly visible. Section 5.3- Are there any standard reference values for electronic chemical potential (A), chemical hardness (B), global softness (C), and global electrophilicity to define the significance of the global parameters of the ligand? If any, please provide and add to the manuscript and in graph legend.\n\nConclusion\nI didn’t understand the conclusion very well. The conclusion is dispersed. I believe an overall conclusion at the end is missing. As per the findings, summarize that which of the key ligand(s) can be considered better including all the parameters of the study. Authors should also write the limitations of their study and future-plan to overcome that.\n\nMinor\nAll abbreviation should be spelled out at first use. Spell out SARS-CoV-2 in the Introduction section. Section 5.1- Please correct “serie of ligands” to “series of ligands”. Make sure all the journal article guidelines are followed.\nThe authors have done well. Further, I believe the above-mentioned comments will be constructive to add more quality to the manuscript and information for readers.\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": "12945", "date": "18 Dec 2024", "name": "Alejandro Morales-Bayuelo", "role": "Author Response", "response": "All the revision we made according to the reviewer. Best Regards" } ] }, { "id": "280574", "date": "23 Jul 2024", "name": "Sobia Ahsan Halim", "expertise": [ "Reviewer Expertise Computational drug design" ], "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 \"Study anti-viral drugs for their efficiency against multiple SARS CoV-2 drug targets within molecular docking, molecular quantum similarity, and chemical reactivity indices frameworks\" by Alejandro et al., is a significant piece of work, well written and well presented.\n\nIn my views it should be indexed after minor revision. My comments are: The in-silico data should be validated by in-vitro test. The limitation of this study should be added in the end of the manuscript. The future recommendation of the work should be given 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": "12945", "date": "18 Dec 2024", "name": "Alejandro Morales-Bayuelo", "role": "Author Response", "response": "All the revision we made according to the reviewer. Best Regards" } ] } ]	1	https://f1000research.com/articles/13-270
https://f1000research.com/articles/13-729/v1	02 Jul 24	{ "type": "Case Study", "title": "Women’s representation in higher leadership positions in Ethiopia in the last three decades since 1991", "authors": [ "Zeynie Chekol Degu", "Flimon Hadaro Hando", "Flimon Hadaro Hando" ], "abstract": "Gender equality in decision-making positions is crucial to achieving the goals of good governance, peace, democracy, and inclusive/sustainable development. The major aim of this research article is to investigate the representation trend of women in higher decision-making positions over the last thirty years, since 1991. The federal three organs of government (law formulating, law enforcing, and law interpreting bodies) were the focus of this research. This research is a mixed type of research that inculcates both qualitative and quantitative data types. Secondary data sources from relevant government institutions were mostly used. The data was analyzed through content analysis of documents and presented via descriptive data presenting techniques. The research findings reveal that although women’s representation in positions of decision-making has advanced considerably in recent years, the empirical data throughout the previous thirty years demonstrated the underrepresentation of women in higher leadership positions within the Ethiopian federal government. Furthermore, Women never held certain higher-level government leadership positions, such as the Prime Minister position, which seems to be forbidden for women. Women made up 23%, 19%, 19%, and 24% of the House of Peoples Representatives (HPR), the House of Federation (HoF), ministerial posts, and judicial bodies, respectively over the last three decades. Women are visibly underrepresented in the executive positions as compared to others. Thus, substantial policy and practical initiatives are needed to remove institutional, social, and economic barriers to boost women’s advanced visibility in senior leadership roles.", "keywords": [ "Women’s representation", "higher leadership positions", "Gender equality" ], "content": "1. Introduction\n\nThe struggle for gender equality is a historical movement for equality, democracy, and women’s rights, and the interventions implemented for gender equality are important tools to enhance women’s enjoyment and participation in social, economic, and political decision-making areas (Hearn & Husu, 2016). All levels of leadership positions in public institutions, civil society organizations (CSOs), non-governmental organizations (NGOs), and private organizations should be gender inclusive and take gender diversity into consideration, according to the findings of empirical literature (Offermann & Foley, 2020). It is because, as argued by Ojulu and Melesse (2014); Ballington and Karam (2005), it is impossible to realize a participatory, genuine, accountable, and transparent governance system without the full and equal involvement of women. This also concerned by the convention of the Beijing platform for action that stated goals of peace and development and human rights could not be realized without incorporation of women’s perspectives at all levels of decision-making (UN Women, 1995).\n\nEven though, women make up half of world population and fifty percent of the working force, they are underrepresented in positions of leadership and decision-making across the globe (Montecinos, 2017; Ballington & Karam, 2005). Around the world, women are primarily found in supporting roles; in junior and middle managerial roles (Schedlitzki & Edwards, 2014). As a manifestation, at present women only constitute 16.1% of ministerial positions and only 22.9% of shares in parliaments at the global level, with the rest of the seats being held by men (WEF, 2022, p. 39). According to the data released by UN Women in collaboration with Inter Parliamentary Union in “Women in politics: 2023” map, as of January 2023, solely 31 countries have head of states and head of governments. Out of the total 193 countries of the globe, only 17/151 countries (11.3%) have women head of states and 19/193 countries (9.8%) have women head of government. Given challenges are varied across countries according to different contexts, socio-economic barriers (resource shortage, illiteracy, multiple burdens, stereotype); psychological barriers (lack of confidence and motivation, perception to politics, lack of qualification); workplace challenges (lack of social-networks and sexual harassment); and political related challenges (male-friendly norms, ineffective women groups) are identified as major barriers that limit women from ascending to higher political leadership positions (Tabassum, & Rafiq, 2023; Pranathi & Lathabhavan, 2021; Appelbaum et al., 2019 and Ballington & Karam, 2005).\n\nIn Ethiopian context, Tesfaye, Hirut, and Rahel (2019) noted that women have made significant contributions to Ethiopian history in the economic, social, and political affairs. However, according to Mihiret (2019) their strength, abilities, and power to do everything do not grant them the appropriate place, including formal authority for leadership. Facts and figures also demonstrated women’s lower participation in decision-making positions (Negussie & Adula, 2021). For instance according to Negussie and Adula (2021) in Ethiopia since 1995 the first round national election, the involvement of women in the parliament was optimistically increasing but still not equal with men. As the data released by WEF (2022), in Ethiopia from the total seats in the upper house (House of Federation) only 30.60% of the seats are held by women (p. 165). Moreover, women are underrepresented in the Ethiopian justice system, both at the federal and regional levels (Addadzi-Koom & Gage, 2022), In general, Ethiopia ranked 74th out of 146 countries in terms of gender gap parity (WEF, 2022). According to the Inter-Parliamentary Union (IPU) (2017), institutionalization of democracy and producing inclusive public policy would not be realized without allowing the full and equal participation of women like that of men. Therefore, women and leadership-related research is required to investigate the issues of women in leadership in Ethiopia intensively to contribute some values on the efforts of making women’s perspective, interest and experience in the law making and enforcing process.\n\nPrevious studies have been undertaken on women and leadership themes in Africa and Ethiopia too. In Africa, the 18-year state of knowledge production on African women in leadership and management has been examined by Nkomo and Ngambi (2009) in their article entitled ‘African Women in Leadership: Current Knowledge and a Framework for Future Studies’. The authors have done an extensive review of existing published research, including journal articles, books, book chapters, and monographs that have been published from 1990–2008 (18 years).\n\nAmong a total 43 publication reviewed by the authors, the largest body of knowledge has been produced on women’s leadership and management status is in case of South Africa, followed by Nigeria and Ghana. The findings of their review disclosed the underrepresentation of women in public office and political leadership in Africa. The majority of the empirical studies have focused on the factors that affect women’s performance in elevating them to higher leadership positions. Socialization, limited educational attainment, multiple roles, gender stereotyping, subtle discrimination, and organizational policies and procedures are the major factors identified by the studies. Indeed, there have been publications in Africa on women and leadership since 2010 by Bauer, Darkwah, and Patterson (2017), Ndlovu and Mutale (2013), Bauer and Okpotor (2013), and Amina and Ibrahim (2019). All these authors are optimistic about women’s growing participation in leadership positions on the African continent, as witnessed in Rwanda from East Africa and Algeria from Arab countries. As per the authors, the increase in women’s movements, quota systems, multi-party systems, educational opportunities, funding from international institutions, global and national agreements and commitments, and role models of women leaders, were found to be factors that contributed to raising women’s political participation in Africa.\n\nThere are numerous empirical studies that have been conducted on women and leadership in Ethiopia. However, a critical review of the existing literature found them as case studies conducted at banks, some selected government institutions, and woredas; furthermore, the concerns of writers are solely restricted to some specific aspects of women and leadership, such as challenges faced by women, women’s perceptions towards leadership, women’s leadership styles, and opportunities provided by institutions to empower women and related issues (Worku, 2017; Tesfay, 2013; Shimelis, 2015; Bizualem & Kasaye, 2020; Mekasha, 2017; Endale, 2014; Genet, 2020; Gojjam and Singh, 2015; and Miressa, 2014).\n\nFurthermore, there are few empirical studies that base themselves on the national context of women’s representation in political leadership positions by Meaza (2009); Berouk (2004); Ojulu and Melesse (2014). As it has been examined by the researcher of this study, all empirical studies have described the status of women’s representation in higher decision-making positions such as parliament, ministerial positions, and political parties since 1995 up until 2012, and they all disclosed the wider gender gap and insignificant representation of women’s in higher political leadership positions in Ethiopia. But, in the course of examining all these stated works, the researcher identified two basic gaps that needed to be filled by this study. The first is time; the above studies have covered the representation status of women in higher leadership positions from 1995–2012 E.C., or only in the four terms of elections. The second is the conceptual gap; the studies skipped the representation status of women in the third organ of government, which is the judiciary, and as per Bauer (2015), very little work has focused on women in judiciaries. Thus, this study intends to use the above studies as a springboard to frame the study and fill the mentioned gaps by expanding the time frame and conceptual coverage (scope).\n\nTherefore, the purpose of this article is to address one of the objectives of the ongoing PhD dissertation on women’s representation in political leadership positions, and its central aim is to investigate the 30 years of women’s representation status at higher decision-making positions (federal-level three organs of government) in Ethiopia since 1991. This study mainly focused on the numerical representations of women and was not substantial.\n\nThis study employed a mixed-methods research approach by incorporating both qualitative and quantitative data types. The quantitative data provides a numerical and figurative description of women’s representation status in higher leadership positions over the past 30 years, while the qualitative data explores the implications of the raw data by referring to related empirical studies. This study primarily depends on secondary data sources. Records from higher government institutions such as the House of People’s Representatives, the House of Federation, and other pertinent institutions and documents provided the intended secondary data for the study. After obtaining data, a desk review of the most important findings and a content analysis of the documents were conducted. The data was presented in descriptive statistics such as percentages, tables, and figures.\n\nThis research article has approved by Addis Ababa University, College of development studies institutional review board named IRB-CoDS on April 3rd 2024 under the written credential number 067/03/2024.\n\n\n2. Findings and discussions\n\n2.1.1 Women’s representation in the Federal Legislature of Ethiopia post 1991\n\nAs indicated in Table 1 below, Women’s representation in the Ethiopian national legislative (Parliament) varied throughout the past thirty years. Despite a recent notable increase, women’s participation in the legislature has been slowly increasing since the first round of national elections in 1995 and the establishment of the federal state structure and parliamentary system of government. For instance, when the House of people representatives first assembled in 1995, its membership consisted of 97% men and 2% women. This is to mean that there were no female representatives to shape policy or voice their experiences, opinions, or interests on a variety of issues.\n\nFrom the second to the sixth round of national elections, women’s representation continued at a gradual but promising pace, with 8%, 21%, 28%, 39%, and 42% of parliamentary seats being held by female legislators, respectively. According to the study’s findings, there has been a 6.6% rise in the average representation of women in the HPR through the previous 30 years. In addition, women have only made up 23% of the seats in the Ethiopian national parliament on average over the past thirty years, with men holding the remaining 77% of the seats. In addition to parliament membership, speaker and deputy speaker positions are among the other higher decision-making positions in the Ethiopian legislature. Women had a higher representation in the deputy speaker roles throughout these election terms; in reality, during the last 30 years in the HoPR, there has only been one female speaker. Men tend to occupy speaker positions more frequently.\n\nOverall, this study’s findings show that, despite women’s representation growing over time, men have typically constituted an excessive number of legislators in Ethiopia. This conclusion also confirmed the study results published in 2009 by Meaza Ashenafi, 2004 by Berouk Mesfin, and 2014 by Ojulu and Melesse, which indicated that women’s presence in senior political decision-making positions in Ethiopia is insignificant.\n\nThis research finding implies that the problems, interests, ideas, values, and roles of women have been overlooked by the legislature because of their underrepresentation. According to Oyindamola and Olaniyan (2020), men have little concern and attention for women’s issues as a result of gendered socialization. In line with this, Shimelis (2015) raised the experience and interest arguments, and according to these arguments, women’s experiences, perspectives, and interests are different from those of men’s, which influence policy decisions in different ways, so representative institutions are essential to articulating the concerns of women’s. The other implications of this finding are that half of the population of the country is not well represented in parliament, given that women constitute over 50% of the nation’s total population.\n\nOn the other hand, as Oyindamola and Olaniyan (2020) discovered, fewer women in the legislature entails less attention to social policy (human trafficking, issues of children, youth, and seniors), gender equality, and family policy. An empirical study conducted by Devlin and Elgie (2008) and O’Brien and Piscopo (2019) also gives support to this argument, acknowledging that a higher representation of women in parliament has a significant impact on the legislative agenda and environment (women-friendly working hours and calendar). Finally, as argued by Ballington and Karam (2005), the insignificant representation of women in parliament is a manifestation of poor democracy. A democratic institution devoid of women’s full participation is retrograde, and any country that upholds it cannot advance politically or flourish economically (Oyindamola & Olaniyan, 2020).\n\n2.1.2 Women’s representation in the Standing Committee Chairs in HoPR since 1991\n\nThe information presented in Table 2 attached shows how the number and purview of standing committees have changed over time in response to Ethiopia’s changing political and socioeconomic landscape. As a result, the number of standing committees varies throughout time. The largest standing committees (20) were formed in 2018 on a variety of areas; however, when a new structure (the creation of multiple subcommittees on several grand committees) was implemented in 2019, the number of standing committees dropped to 10.\n\nAs can be seen in the table, men significantly outnumber women in Ethiopian parliament standing committee chair and deputy chair posts held after 1991. To put it numerically, in the past 30 years, the proportion of women as chairperson in the HPR standing committees has merely by 31.5%. The participation of women as vice chairpersons is lower than this number, at 30%. Unlike the HoPR deputy chairperson positions (the position that seems left for female representatives), even this position in the standing committees is highly represented by men’s. As a result, throughout the past three decades, the proportion of women serving as chairpersons and deputy chairpersons has only been 30.5%; male members of parliament have held the remaining 70% of these positions. Thus, women have a small presence among the chairs of the standing committees in the HoPR, much like they have little representation among parliamentarians.\n\nAccording to Benda (1997), given differences in their type, duties, and significance, most countries in the world adopted parliament (standing) committees that specialize on various matters, and they are usually created for the effectiveness and efficiency of the large work of the legislature. Parliamentary chairs are considered important policy actors, and they play an important role in parliamentary political leadership positions (Gaines et al., 2019). A legislature with strong committees and chairs has an impact on shaping government policies. According to Fortunato et al. (2019), committee chairs can play two important roles by using their agenda powers: encouraging opposition political parties to examine proposed government policies and enabling the ruling parties to provide better policies to the public. To sum up, women chairs are essential to the advancement of gender equality, inclusive governance, and the development of policies that are advantageous to society at large because of their concern for grassroots agendas (Wängnerud, 2009).\n\nTherefore, the underrepresentation of women in committee chairs and other political leadership roles inside the parliament has implications of its own. The first is the absence of female role models, which would deter women from entering the political arena and positions of leadership. Secondly, inclusive decision-making would be missing. Their limited presence in political leadership roles within parliament undermines inclusive decision-making procedures that welcome a range of viewpoints and opinions. The third implication is that there would not be policy prioritization, i.e., women are more likely to focus on policies that address women’s rights, gender equality, social welfare education, and other related issues.\n\n2.1.3 Women’s representation trend in the House of Federation (HoF) or Upper House post 1991\n\nThe data presented in Table 3 attached below deals with women’s representation in Ethiopia’s House of Federation, the country’s second chamber of parliament. In the past thirty years, Ethiopia has held six consecutive national elections. Only seven women were elected to the house of federation during the first national election in 1995, with 103 seats being held by men, according to the facts shown by figures. Over the last six terms, unlike the case of House of Peoples Representatives, the highest number of female representations in the upper house (HoF) registered in the 2015 national elections, constituting 50 (32.6%) of seats. The number of female representatives in this house did not exceed 50 of the total members. However, this number was again reduced to 43 (30%) in the recent 2021 national election due to the absence of Tigray regional state representatives due to the official war between the federal government and the TPLF (Tigray People’s Liberation Front), which started in November 2020 and ended in November 2022.\n\nAccording to the results of this study, males and females were represented in the House of Federation by 81% and 19%, respectively, over the last thirty years (six terms). This shows that, on average, women’s were underrepresented in the House of Federation compared to even the representation level of women’s in the House of Peoples Representatives. The findings of this study further revealed that, despite the dynamics of the number of female representatives at different terms, female representation in the upper house has increased by 3.88% over the course of the past six terms. Generally, given their underrepresentation in both houses, the above data discloses that women are insignificantly represented in the House of Federation compared to the House of People’s representatives.\n\nMeanwhile, Women make up a minor portion of the House’s standing committees in addition to being members of the House of Federation. Data from 2010 to the most recent election (2021) showed that, on average, women’s participation in standing committees was only 17% throughout the previous three terms. Women are also disproportionately underrepresented in the working four standing committees, with the standing committees on revenue affairs, budget, and subsidies having the lowest representation, followed by the council of constitutional inquiry.\n\nThis finding supports the results released by Atsede, Aemro, and Eyayu (2022), who argued that despite attitudinal changes and governmental efforts regarding women’s political participation enhancements, “nevertheless, facts and figures show that, even currently, the participation of women is not at an equal level with men in the highest decision-making spheres” (p. 75). And this finding implies that without a fair and equitable representation of women in the political process, the objectives of democracy, good governance, human rights, gender equality, and development would not be realized. It is because, as argued by Wubante (2021), the involvement of women in political leadership is essential for promoting democracy and establishing effective governance. On the other hand, women’s lower engagement in the HoF means the absence of women’s voice during constitutional interpretation, constitutional amendments, and on the issues of rights of nations, nationalities, and peoples of Ethiopia, which has an adverse effect on the impacts of women’s participation in national and regional agendas.\n\n2.2.1 Women representation in Chief Executive and President Positions\n\nNational executive positions including head of government are the highest political leadership positions of a state in which important national-based decisions are undertaken and are responsible for the supervision and implementation of policies and laws made by the legislature. In the Ethiopian context, these positions are mostly occupied by male executives such as male prime ministers, male presidents, and male deputy prime ministers. As per the figures in Table 4 pointed out, shockingly, there was no female prime minister and there were no female presidents (until President Sahle-Work Zewde was elected as the first female president in 2018) in Ethiopian history post-1991. Meanwhile, only one female deputy prime minister served in the position of 2nd deputy prime minister from 2014–2016, while all deputy prime minister positions were held by male executives after 1991. And her tenure was not longer than two years.\n\nThe findings of this study revealed that women have been severely underrepresented in Ethiopian executive posts since 1991, which contradicts the findings of Farida Jalalzai’s (2004) global-based work, in which she concluded that women have rarely been presidents or prime ministers around the world; however, more women have been reaching these high positions since the 1990s. Her findings are invalid in the context of Ethiopia. On the other hand, the findings of this study contradict her other finding, which claimed that women are more likely to access executive positions in the parliamentary government system than in the presidential system (the argument having that women are less likely to be elected directly by the public due to patriarchal society) (Jalalzai, 2004). Her finding appears to be unsound, given that Ethiopia’s parliamentary system experience over the last thirty years has not provided opportunities for women to advance to such top decision-making positions.\n\nAccording to the 1995 FDRE constitution (Article 72/1), the Prime Minister and the Council of Ministers have the highest executive authorities of the Federal Government of Ethiopia, and the Prime Minister in particular has crucial national powers. This implies that women’s absence as prime minister since 1991 means that over the past 30 years, women’s did not organize cabinet ministers (councils of ministers) for different departments; women’s never been the commander-in-chief of the national armed forces (all past 30 years, defense force movements and war-related decisions have been done by males); posts of commissioners; the president and vice president of the federal supreme court; and the auditor general have never been selected by females. Thus, the existing economic condition, justice system, diplomacy, and peace trend are or were the result of men’s political leadership. This implies that this and related conditions could have been different in the country if male leaders collaborated with females and considered gender inclusivity in leadership positions.\n\n2.2.2 Post 1991 Women’s representation at the Ministerial Positions and Their Portfolios\n\nGovernment ministries are mainly responsible for the implementation of laws and regulations and for the administrative functions of the national government. As it is observed in Table 5 attached below, through the course of the last 30 years, in Ethiopia, the number and nomenclature of ministerial offices have been changing to cope up with the socio-economic, political, and technological changes of different times.\n\nThroughout this journey, the biggest number of female ministers was three (13%) until 2013, while male ministers occupied 87% of ministerial positions. Since 2013, the proportion of female ministers has increased to five (22%) or more. The highest number of female ministers were appointed in 2019, when the percentage of them increased to 10 (48%), or nearly half of the total share. This time, the male/female minister ratio was 10 (48%)-11 (52%), however, the number of female ministers in ministerial positions, was recently lowered to 8 (36%), in 2022. According to the study’s findings, female representation in ministerial positions has averaged 19% during the last thirty years. Furthermore, the average female’s leadership in ministerial positions increased gradually by 3.44%.\n\nThis research comes up with the finding that, though currently the engagement of females at the ministerial posts is increasing and encouraging, there was no meaningful representation of women’s before 2019. On the other hand, according to the data obtained from the FDRE House of Peoples Representatives teaching and communication directorate, women in Ethiopia have been appointed in every ministerial office at least once over the last thirty years, but the top three departments (ministerial positions) that are frequently headed by female ministers are the ministries of women, children, youth, and social affairs (over eleven times), culture and tourism (five times), and urban development and construction (four times). The results of this study showed that, despite the fact that these ministries contribute to the socio-economic and political advancement of a nation, women in Ethiopia have not held the leadership positions in important and high-profile political posts, such as those related to science and technology, foreign affairs, finance, defense, and justice. According to Barnes & Taylor-Robinson (2018), these positions allow ministers to represent their nation at significant global forums, and prime ministers and presidents frequently use them to express the general direction of their government’s policies. As concluded by Krook and O’Brien (2012), women have held fewer cabinet positions, and when they did, they were frequently assigned to portfolios associated with lower status and “feminine” attributes. The findings of this study also confirm the findings revealed by Barnes & Taylor-Robinson (2018); although the importance of cabinet portfolios varies across countries, the most common high-profile posts are defense, finance, and foreign relations. Even though these authors underlined the importance of women’s presence in powerful positions as, “women’s presence in top cabinet posts is positively associated with both women’s and men’s satisfaction with and confidence in government” (Barnes & Taylor-Robinson, 2018, p.19). However, it is not common for women to hold these important positions; in particular, certain leaders in Asia, the Middle East, and Sub-Saharan Africa have never proposed a woman to take any of these positions (Barnes & Taylor-Robinson, 2018).\n\n2.3.1 Women’s representation trend in the Federal Courts since 1991\n\nFigures in Table 6 attached below reveal the share of males and females as judges and in the positions of president and vice president in the three levels of federal courts, such as the Federal First Instance Courts (FFICs), Federal High Courts (FHCs), and the Federal Supreme Court (FSC). As per the data in Table 6, the federal government has appointed a total of seven presidents in the three federal courts over the last thirty years. Among the seven presidents, only one was female, while six of the total presidents were male. According to the finding, Meaza Ashenafi was not only the first female president of federal courts but also the only female chief justice (president of the Federal Supreme Court) in Ethiopian history. Besides this, over the course of this year, a total of 11 vice presidents have been appointed, and among them, 9 were males and the rest 2 were females. Within these years, the representation of female leaders in the federal courts president and vice president posts was 14% and 25%, respectively.\n\nOn the other hand, across the last 30 years since 1991, a total of 805 judges have been appointed to the federal courts, and among them, the proportion of female judges was 197 (24%), while the number of male judges was 608 (76%). Meanwhile, independently, the share of female judges in the Federal First Instance Courts (FFICs), Federal High Courts (FHCs), and the Federal Supreme Court (FSC) across the last thirty years was 27%, 21%, and 22.3%, respectively. From this, it is sound to deduce that the representations of female judges are better at the federal first instance court than the other two federal courts.\n\nGenerally, for the past thirty years, there have only been 24% of women serving as federal judges, and only 14% and 25% of them have held leadership roles as president and vice president, respectively. The study’s main conclusion was that women are underrepresented in Ethiopia’s legal system, both in leadership roles and in arbitration. This finding is consistent with the conclusion of Addadzi-Koom and Gage (2022), which announced that in the three levels of Ethiopian federal courts, the total number of female judges is outnumbered by male judges. This finding is also consistent with the global-based empirical research by Castillejos-Aragón (2021), which affirmed that despite the growing participation of women in the legal profession, the gender gap remains prominent, and according to this writer, women’s are still underrepresented in top-ranking positions of the judiciary, even in countries where more women are appointed as judges.\n\nAs pointed out by different writers, the lower representation and participation of women in the judiciary body have their own implications. As argued by Castillejos-Aragón (2021), women’s absence in the judiciary reduces the responsiveness, inclusivity, and participatory nature of decision-making at all levels. It also has an impact on women’s equal visibility, diversity of viewpoints, and gender-sensitive judicial institutions. Additionally, the legitimacy of the judiciary would be reduced by the lack of women in this body and last but not least it neglects women’s inherent right, acknowledged by various international frameworks, to participate equally in all public institutions (Castillejos-Aragón, 2021).\n\n\n3. Conclusion\n\nIn Ethiopia, women’s representation in senior leadership roles within the federal government was and remains insignificant. These roles include head of state and government, cabinet positions, membership in the House of People’s Representatives and House of Federation, chairs of standing committees, and the judiciary body. Women do not hold certain higher-level decision-making roles, such as the Prime Minister position, which seems to be closed to women.\n\nThe percentage of women serving in Ethiopia’s national legislature, or Parliament, has increased since 1991. The study’s findings indicate that the proportion of women in the HoPR has increased by an average of 6.6% over the preceding thirty years. Furthermore, throughout the preceding 30 years, women’s representation in the Ethiopian national parliament has averaged only 23%. In the meantime, women have a stronger representation in the deputy speaker roles during this election term. Speaker roles are typically occupied by men. Overall, this study’s findings show that, despite women’s representation growing over time, men have consistently constituted an excessive number of legislators in Ethiopia. According to this study, female membership in the HoF constituted solely 19% in the last three decades. This demonstrates that women’s presence in the House of Federation is negligible when compared to the representatives in the House of People, considering their underrepresentation in both houses.\n\nHigh executive positions in Ethiopia are typically held by men’s, including those of presidents, deputy prime ministers, and prime ministers. Amazingly, throughout Ethiopian history since 1991, there has never been a female prime minister or president (before to President Sahle-work Zewde’s election as the country’s first female president in 2018). Over the past thirty years, the average percentage of women holding ministerial positions has been 19%. On average, there has been a modest increase of 3.44% in the number of women holding ministerial positions. The ministry of women, children, youth, and social affairs; the ministry of culture and tourism; and the ministry of urban development and construction are the top three departments (ministerial roles) that are frequently headed by female ministers, according to this study.\n\nThe study’s primary finding also demonstrated the minimal representation of women in leadership roles and arbitration within the Ethiopian legal system. During the entire year, the judiciary organ had 14% and 25% of female leaders in the positions of president and vice president of the federal courts, respectively. Furthermore, of the total 805 judges, the proportion of female judges was solely 197 (24%).\n\nAll in all, the data and statistics presented in this study point to a hopeful increase in the representation of women in positions of decision-making. For women to effectively participate, however, substantial policy and practical initiatives including appropriate monitoring and evaluation are needed to remove institutional, social, and economic barriers as well as increase women’s advanced visibility in senior leadership roles.\n\nSpecifically, to enhance women’s participation in higher leadership positions, policies that are inclusive, strong, and have important indicators and measurements on women’s civil rights, women’s participation in civil society, and women’s engagement in decision-making positions need to be developed in the meantime, particularly taking into account the gaps in Ethiopia’s current policy frameworks.\n\nAll the relevant sources reviewed are included in the references and citations. No source (data analyzed and scholarly works reviewed) is used without proper acknowledgement. This research article has approved by Addis Ababa University, College of development studies institutional review board named IRB-CoDS on April 3rd 2024 under the written credential number (ethical approval number) 067/03/2024. Data was collected from relevant government institutions, not from individual participants. Since all of what is contained in the article was acquired through document analysis, verbal approval was reached with the appropriate body for the publication of the secondary data obtained from them. Furthermore, the university’s ethical approval committee assessed the research article’s ethical status and agreed to proceed with its publishing by granting an ethical approval certificate.", "appendix": "Data availability statement\n\nDryad: Women’s status in Higher leadership positions in Ethiopia since 1991 [Dataset], Dryad. Accessible via this link: https://doi.org/10.5061/dryad.7m0cfxq38 (Chekol & Hadaro, 1991).\n\nThe project contains the following underlying data and tables: Women_in_ legislative_body; Women_in_the_executive_body; Women_in_the_judiciary_body and Women_in_the_upper house_ (HoF). It has also included the extended data file; guiding_questions_for_secondary_data\n\nThe data is available for peer review under the link: https://datadryad.org/stash/share/YAFozIzc81uzmSfYy_jjLbgdmrpodbQm6j1UOUyemSg.\n\nData are available under the terms of a CC0 1.0 Universal (CC0 1.0) Public Domain Dedication license.\n\n\nAcknowledgements\n\nI sincerely thank my supervisors, Dr. Flimon Hadaro and Dr. Hanna Tegegne, for their insightful comments throughout this article’s development. This statement is based on the consent of those who are stated here.\n\n\nReferences\n\nAddadzi-Koom ME, Gage BG: Women Judges in Ethiopia. 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Journal of Culture, Society and Development. 2020.\n\nCastillejos-Aragón M: A need for change: Why do women in the judiciary matter? Konrad Adenauer Stiftung. JSTOR. 2021.\n\nChekol Z, Hadaro F: Women’s status in Higher leadership positions in Ethiopia since 1991. [Dataset]. Dryad. Forthcoming 2024. Publisher Full Text\n\nDevlin C, Elgie R: The effect of increased women’s representation in parliament: The case of Rwanda. Parliamentary Affairs. 2008; 61(2): 237–254. Publisher Full Text\n\nEndale A: Factors that affect Women Participation in Leadership and Decision Making Position. Asian Business Consortium\|AJHAL. 2014.\n\nFortunato D, Martin LW, Vanberg G: Committee chairs and legislative review in parliamentary democracies. British Journal of Political Science. 2019; 49(2): 785–797. Publisher Full Text\n\nGaines BJ, Goodwin M, Bates SH, et al.: The study of legislative committees. The Journal of Legislative Studies. 2019; 25(3): 331–339. Publisher Full Text\n\nGenet B: Factors Affecting Women Participation in Leadership Position: The Case of Debre Markos City Administration. Asian Journal of Humanity, Art and Literature. 2020; 7(1): 9–20. Publisher Full Text\n\nGojjam A, Singh M: Factors affecting women’s participation in leadership and management in selected public higher education institutions in Amhara Region, Ethiopia. European Journal of Business and Management. 2015; 7(31): 18–29.\n\nHearn J, Husu L: Gender equality. The Wiley Blackwell Encyclopedia of Gender and Sexuality Studies. 2016; pp. 1–3. Publisher Full Text\n\nJalalzai F: Women political leaders: Past and present. Women & Politics. 2004; 26(3-4): 85–108. Publisher Full Text\n\nKrook ML, O’Brien DZ: All the president’s men? The numbers and portfolio allocations of female cabinet ministers. The Journal of Politics 2012; 74(3): 840–855. Publisher Full Text\n\nMeaza A: Participation of women in politics and public decision making in Ethiopia. Forum for Social Studies. 2009.\n\nMekasha K: Women’s role and their styles of leadership. International Journal of Educational Administration and Policy Studies. 2017; 9(3): 28–34. Publisher Full Text\n\nMihiret G: Factors affecting women’s participation in leadership positions in the case study of Bank of Abyssinia. 2019. Reference Source\n\nMiressa Y: Practices and Challenges of Women in Leadership and Development Activities of Some Selected Woredas of Bench Maji Zone, Ethiopia. International Journal of Science and Research (IJSR). 2014.\n\nMontecinos V: Women presidents and prime ministers in post-transition democracies. Basingstoke: Palgrave Macmillan Studies in Political Leadership; 2017. Publisher Full Text\n\nNdlovu S, Mutale SB: Emerging trends in women’s participation in politics in Africa. International Journal of Contemporary Research. 2013; 3(11): 72–79.\n\nNegussie TG, Adula NG: Current Status of Gender Equality and Governance in Ethiopia. International Journal of English Literature and Culture. 2021; 9(4): 118–122.\n\nNkomo SM, Ngambi H: African women in leadership: Current knowledge and a framework for future studies. International Journal of African Renaissance Studies. 2009; 4(1): 49–68. Publisher Full Text\n\nO’Brien DZ, Piscopo JM: The impact of women in parliament. The Palgrave handbook of women’s political rights. 2019; pp. 53–72. Publisher Full Text\n\nOffermann LR, Foley K: Is there a female leadership advantage? Oxford research encyclopedia of business and management; 2020.\n\nOjulu OO, Melesse A: Assessment of Gender Equality in Ethiopia: The Position of Ethiopian Women’s Political Representation from the World, Sub- Saharan Africa, and Eastern Africa Ethiopian Civil Service University. JL Pol’y & Globalization. 2014; 28: 102.\n\nOyindamola EM, Olaniyan A: An Investigation into the Causes and Effects of Women Political Underrepresentation in Ogun State, Nigeria. International Journal of Research and Innovation in Social Science. 2020; 04(11): 318–325. Publisher Full Text\n\nPranathi BKP, Lathabhavan R: A study on role of woman in leadership positions. International Journal of Innovative Research in Applied Sciences and Engineering. 2021; 4(12): 940–943.\n\nSchedlitzki, Edwards: Leadership, Gender and Diversity. SAGE Publications; 2014.\n\nShimelis K: Challenges and opportunities of women political participation in Ethiopia. Journal of Global Economics. 2015; 3(4): 1–7.\n\nTabassum S, Rafiq U: Leadership challenges: Experiences of women leaders. Pakistan Journal of Social Research. 2023; 05(02): 563–571. Publisher Full Text\n\nTesfay T: Some Ethiopian Women Leaders’ Perceptions about Their Leadership. Advancing Women in Leadership Journal. 2013; 33: 75–86.\n\nTesfaye S, Hiru B, Rahel A: Women and Development in Ethiopia: A Sociohistorical Analysis. Journal of Developing Societies, 2019 SAGE Publications. 2019.\n\nUN Women: Beijing declaration and platform for action. UNWOMEN; 1995.\n\nUnion IP: Proportion of seats held by women in national parliaments (%). World Bank; 2017.\n\nWängnerud L: Women in parliaments: Descriptive and substantive representation. Annual Review of Political Science 2009; 12: 51–69. Publisher Full Text\n\nWorku M: Women in management: Challenges and gaps in public institutions in Ethiopia. J. Int. Women’s Stud. 2017; 18(2): 105–117.\n\nWorld Economic Forum (WEF): The Global Gender Gap Report, Insight Report. Geneva, Switzerland: 2022. ISBN-13: 978-2-940631-36-0.\n\nWubante A: Women and Ethiopian politics: Political leaders’ attitude and views on women’s effectiveness. Cogent Social Sciences. 2021; 7(1): 1948653. Publisher Full Text" }	[ { "id": "303834", "date": "22 Jul 2024", "name": "Professor Claes Maria Theresia", "expertise": [ "Reviewer Expertise Gender and Diversity in Organisations", "Leadership" ], "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 an interesting article on Women’s representation in higher leadership positions in Ethiopia in the last three decades since 1991. It shows how women have progressed in political and judicial high positions in the last 30 years, but also that the numbers are still small. Another conclusion is that women do advance, but mostly into positions of lower responsibility and power, not in the higher or more powerful positions. Progress is small, with the result that political decisions are made by men, without necessarily taking into consideration half the population, or having their perspectives on the decisions. The text is well written, but needs to go trough language checker for minor typos and grammatically incorrect sentences. One remark also about the description of the article: the authors mention qualitative and quantitative research, but apparently the only method used is quantitative. The conclusion is well written and informative.\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": "12056", "date": "23 Jul 2024", "name": "Zeynie Chekol", "role": "Author Response", "response": "Hello, dear Professor Claes Maria Theresia, Thank you for your commitment, time, and efforts to review our article, and I appreciate your constrictive and valuable comments. We will address all the reservations and improve them in a way to contribute to the scientific knowledge production on the theme of women and leadership. Thanks again, Regards." } ] }, { "id": "299158", "date": "25 Jul 2024", "name": "Jiregna Tadesse", "expertise": [ "Reviewer Expertise I am interested in exploring a research areas such as Sustainable Peace", "Conflict Management", "Conflict Transformation", "Political Reform", "National Dialogue and Consensus", "Reconciliation", "Migration and Displacement", "Famine", "War Studies", "Humanitarian Governance", "Humanitarian Business", "Local Development", "Sustainable Development", "Environmental Governance", "Transboundary Water Governance", "Intersectional Approach", "Human Rights", "Racism", "Discrimination", "Gender Study", "Political Economy of War", "Armed Conflicts and Military Science", "Post-Conflict State Reconstruction", "Aid", "and Society", "Famine", "and War Studies", "Web Design", "UI/UX", "Artificial Intelligence", "Machine Learning", "Cyber Security and more." ], "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\nConstructive Assessment of the Study The case study on women’s political representation in Ethiopian leadership positions is well-structured and provides a comprehensive examination of the subject. Below is a detailed assessment along with answers to the mandatory reviewer questions and recommendations for potential improvements.\nBackground and Historical Context Assessment: The study offers a thorough historical and contextual overview of women's involvement in Ethiopian politics. It effectively outlines the evolution of gender representation and key milestones affecting women’s political participation.\nRecommendations: To further enhance the background, consider incorporating a detailed timeline of significant legislative and societal changes that have impacted women’s roles in politics. This would provide a clearer understanding of the historical progression and its effects on the current political landscape.\nClarity and Accuracy of Presentation Assessment: The presentation of the study is clear and well-organized, with accurate references to the current literature. It aligns with existing research and contributes meaningfully to the discourse on gender and leadership.\nRecommendations: Continue to ensure clarity throughout the study, particularly when presenting complex data or historical developments. Double-check all citations for accuracy and completeness to maintain the study's scholarly integrity.\nStatistical Analysis and Interpretation Assessment: The study employs appropriate statistical methods, and the interpretation of results is consistent with the study’s objectives. This adds robustness to the findings and supports a better understanding of women’s representation in Ethiopian politics.\nRecommendations: Ensure that the methodologies used are described in detail, including any assumptions and potential biases. Clear and transparent presentation of statistical results will further validate the study's conclusions.\nReproducibility of Source Data Assessment: The availability of source data is essential for verifying the study’s findings. While the study’s summary does not specify data accessibility, ensuring that underlying data is available will support reproducibility and transparency.\nRecommendations: Make all data sources accessible and consider including supplementary materials or appendices that outline the data collection process. This will enhance the reproducibility of the study and provide greater transparency.\nSupport for Conclusions Assessment: The conclusions drawn from the study are well-supported by the results presented. The study successfully connects its findings to broader themes in gender representation and political participation.\nRecommendations: Review the conclusions to ensure they are directly supported by the data and analysis. Address any limitations in the study’s results and discuss their potential impact on the conclusions.\nDetail and Usefulness for Teaching or Practitioners Assessment: The study is presented with sufficient detail to be valuable for both teaching and practical applications. It provides meaningful insights into the challenges and advancements in women’s political leadership.\nRecommendations: Enhance the practical relevance of the study by including specific case studies or examples that illustrate key findings. This will increase its utility for practitioners and educators in understanding and addressing issues related to women’s leadership.\nMandatory Reviewer Questions 1. Is the background of the case’s history and progression described in sufficient detail?\n\nYes, the background is described in sufficient detail, providing a thorough historical and contextual overview of women’s political involvement in Ethiopia. The study effectively captures the evolution and key milestones in women’s representation. 2. Is the work clearly and accurately presented and does it cite the current literature?\n\nYes, the study is clearly and accurately presented. It appropriately cites current literature, aligning with existing research and contributing meaningfully to the field of gender and leadership. 3. If applicable, is the statistical analysis and its interpretation appropriate?\n\nYes, the statistical analysis is appropriate, and the interpretation of results is consistent with the study’s objectives. The analysis adds robustness to the findings and enhances understanding of women’s representation. 4. Are all the source data underlying the results available to ensure full reproducibility?\n\nNot fully specified. The summary does not mention the availability of source data. To ensure reproducibility, it is recommended to make all source data accessible and consider including supplementary materials or appendices detailing the data collection process. 5. Are the conclusions drawn adequately supported by the results?\n\nYes, the conclusions are well-supported by the results. The study effectively connects its findings to broader themes in gender representation and political participation, though it is advisable to review and address any limitations that may impact the conclusions. 6. Is the case presented with sufficient detail to be useful for teaching or other practitioners?\n\nYes, the case is presented with sufficient detail to be valuable for teaching and practical applications. The study provides insights into challenges and advancements in women’s political leadership, though including specific case studies or examples could further enhance its practical relevance.\nApproval Status Approved: The study is well-executed, provides significant insights into women’s political representation in Ethiopia, and contributes meaningfully to existing literature. The recommendations provided aim to further strengthen the study and ensure its impact and applicability.\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": "12108", "date": "31 Jul 2024", "name": "Zeynie Chekol", "role": "Author Response", "response": "Dear Dr. Jiregna Tadesse, Thank you very much for taking the time and employing your efforts to review our article, and I appreciate your commitment. All the comments and suggestions are greatly constructive and appreciated, and we will consider them in upgrading the manuscript to contribute to better scientific knowledge production in the theme of women and leadership. Thanks, again." } ] } ]	1	https://f1000research.com/articles/13-729
https://f1000research.com/articles/13-1236/v1	15 Oct 24	{ "type": "Study Protocol", "title": "Factors contributing to persistent shoulder pain after arthroscopic rotator cuff repair: Protocol for a scoping review", "authors": [ "Anupama Prabhu B", "G Arun Maiya", "Vivek Pandey", "Kiran K V Acharya", "Prabu Raja G", "James Elliott M", "Mira Meeus", "Anupama Prabhu B", "Vivek Pandey", "Kiran K V Acharya", "Prabu Raja G", "James Elliott M", "Mira Meeus" ], "abstract": "Introduction Rotator cuff (RC) tears are the most common and disabling musculoskeletal ailments among patients with shoulder pain. Although most individuals show improvement in function and pain following arthroscopic rotator cuff repair (ARCR), a subgroup of patients continue to suffer from persistent shoulder pain following the surgical procedure. Identifying these factors is important in planning preoperative management to improve patient outcomes.\n\nObjective This scoping review aims to identify biological factors, psychological factors, and social determinants of health contributing to the development of persistent pain in individuals after the ARCR procedure.\n\nInclusion criteria All prospective and retrospective longitudinal studies reporting the risk factors contributing to persistent pain three months or longer after the ARCR surgery will be considered for this scoping review.\n\nMethods Our review will adhere to the Joanna Briggs Institute (JBI) scoping review methodology. Four electronic databases PubMed, CINAHL, Embase, and Scopus will be searched for studies in the English language. Additional studies can be found by conducting a citation analysis of the included studies. Title and abstract screening will be performed by two independent reviewers following the inclusion criteria, a third reviewer will be consulted about any differences. Next, full-text screening will be conducted, and the remaining search results will be reviewed to extract data, as well as to synthesize findings from all research. An overview of findings will be depicted in tabular format accompanied with a narrative summary of various factors contributing to persistent pain.", "keywords": [ "Chronic pain", "Predictors", "Shoulder", "Factor", "Arthroscopy", "Risk factor" ], "content": "Introduction\n\nShoulder pain ranks third among common, but enigmatic, musculoskeletal disorders, prompting patients to visit a wide variety of healthcare practitioners.1 The general population’s prevalence of shoulder pain varies greatly, averaging 16% (range: 0.67 to 55.2%). According to the global burden of shoulder pain survey, the incidence of shoulder discomfort ranges from 7.7 to 62 per 1000 persons per year (median 37.8).2 Accounting for more than half of the shoulder issues addressed by healthcare providers, RC disorders contribute significantly to shoulder pain and dysfunction.3,4 RC disorders are inclusive of RC tendinopathies, calcified tendinopathies, and partial/full-thickness RC tears.5 The shoulder joint is the most mobile in the body because of the RC muscles’ contribution to its static and dynamic stability.6 Between 4% and 32% of patients with RC disorders present with symptoms such as pain, weakness, and limitations in daily activities, reflecting the demand placed on these muscles.7 Shoulder pain can also result in lost productivity, absences from work, and early retirement in workers, especially those who are subjected to heavy physical demands or repetitive overhead activities.8 A systematic review has outlined the challenges in decision-making for conservative or surgical management of RC tears.9 The review findings suggest that earlier surgical intervention may be necessary in cases of weakness and significant functional disability. It also appears that worker’s compensation claims may negatively impact the treatment outcomes.\n\nOver the last decade, the surgical management of RC tears has changed from open to mini-open to arthroscopic repair.10 Currently, ARCR surgery is the most common treatment for RC tears. After ARCR , functional outcomes of traumatic and non-traumatic RC tears have shown similar and satisfactory results.11–13 Improved function, quality of life, range of motion, strength, and pain relief can be attained with the ARCR procedure.14,15 The incidence of ARCR has seen a linear increase in various developed16 and developing17 countries and this rate is expected to rise with an increase in the aging population. The overall cost of ARCR surgery for the National Health Service in England exceeded £60 million, whilst the anticipated yearly cost of repair in the United States is said to be between US$1.2 and 1.6 billion.18 Furthermore, there has been a prominent change in the performance of ARCR procedures at ambulatory surgery centers. These healthcare facilities are known for discharging patients on the same day they undergo surgery.19\n\nWhile several studies have reported positive outcomes on early postoperative pain control, a subgroup of patients report persistent shoulder pain of varying severity and duration following ARCR.20–22 Persistent post-surgical pain is defined by the International Classification of Diseases – 11 as pain that develops or gets worse following surgery or tissue damage and lasts for at least three months after the initial occurrence, beyond the healing phase.23 The transition from postoperative acute pain to persistent post-surgical pain is a complex and multifactorial phenomenon.24 Persistent postoperative pain is largely underdiagnosed and frequently inadequately managed.25 Studies have reported reduced shoulder range of motion, strength deficits, and significantly lower functional scores in patients with persistent shoulder pain following the ARCR procedure.26 Orthopedic surgeries in general are related to an increased risk of experiencing moderate to severe persistent pain at one-year follow-up when compared to other surgical procedures.27 This could be due to the presence of pre-existing nociplastic pain and central sensitivity changes before the surgical intervention in a variety of orthopedic conditions.28,29 Research supporting the role of psychological variables in chronic shoulder pain is emerging. Preoperative psychological disorders such as depression and anxiety, as well as behaviors such as pain catastrophizing, kinesiophobia, and psychological discomfort, may cause patients to experience increased shoulder pain.30\n\nThe degree of persistent postoperative pain beyond three months and the patterns of pain at the follow-up period are unknown. Numerous factors including fatty infiltration of the RC musculature, tear size, and diabetes mellitus are known to predict poor outcomes.31 In addition, factors like opioid consumption, psychological and socioenvironmental, and acute postoperative conditions help explain the occurrence of persistent pain following ARCR.32 Orthopedic surgeons and physical therapists may be better able to predict results and comprehend this patient population if they are aware of factors contributing to persistent pain following ARCR. Pre-operative pain control could be a preventive measure towards reducing the incidence and development of persistent postoperative pain.33 It would be beneficial to identify patients at higher risk of developing persistent pain before the surgical procedure or during initial rehabilitation. This would enable targeted preoperative or perioperative interventions to better manage the transition to persistent postoperative pain. These targeted interventions could potentially be incorporated into a comprehensive care plan for patients undergoing ARCR. Numerous trials have been undertaken to assess the effectiveness of postoperative pharmacological therapies in reducing acute pain34,35 but no comprehensive evaluation has yet been conducted to assess factors contributing to persistent pain after ARCR.\n\nThis scoping review aims to identify factors reported in the literature that contribute to the development of persistent pain in patients following the ARCR procedure. The review will specifically aim to address the following questions.\n\ni) Which pre-operative, intraoperative, and post-operative factors contribute to persistent pain of the shoulder in patients undergoing ARCR?\n\nii) Which biological factors, psychological factors, and social determinants of health contribute to persistent pain at three months or longer after the ARCR surgery in patients with RC tears?\n\n\nMethods\n\nThis protocol has been developed based on JBI methodology for scoping reviews36 and will be in line with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for Scoping Reviews (PRISMA-ScR).37 Details of this review project are available on Figshare: 10.6084/m9.figshare.26885746.38\n\nAll prospective and retrospective longitudinal studies that include participants beyond 18 years of age, diagnosed with partial/full thickness (>5 cm) traumatic or degenerative RC tears undergoing primary ARCR where concomitant procedures like acromioplasty, subacromial decompression, biceps tenotomy, distal clavicle excision, and labral debridement, conducted with ARCR will be included. These concomitant procedures are often performed with ARCR to address additional aspects of shoulder joint pathology. Studies that have a combination of other orthopedic procedures will be included if separate data is available for the ARCR participants. Studies on massive tears (>5 cm in diameter or the detachment of two or more tendons) having a higher rate of recurrent tears and requiring complex surgical procedures like patch augmentation, muscle-tendon transfer, or reverse total shoulder arthroplasty will be excluded.39 Revision repairs will, even so, be excluded as this will interfere with our data on primary ARCR.\n\nThe concept underlying this scoping review is the biological factors e.g. the size of tendon tears, tendon retraction, etc., and the psychological factors e.g. depression, catastrophizing, anxiety, etc., and social determinants of health e.g. gender, race, occupation, etc., related to persistent pain following ARCR surgery. For a study to be considered for inclusion in the scoping review the intensity of pain should be assessed on a reliable and valid unidimensional or multidimensional pain scale at any time point beyond three months following ARCR, identifying at least one pre-operative, intraoperative, or postoperative factor, and the relationship between the factor(s) and ongoing pain. Pharmacological and non-pharmacological studies providing interventions for pain relief will not be considered in this scoping review. Trials with a primary objective towards the efficacy of intervention rather than the impact of risk factors on pain outcome at follow-up will be excluded. The scoping review will encompass studies carried out in global hospitals and primary healthcare settings, considering evidence from any geographical location.\n\nThe search strategy will be designed to find published original research, literature reviews, and written works. An initial limited search on PubMed was conducted to identify articles in this area. A thorough search method was developed using the terms present in the titles and abstracts of pertinent papers, along with the index terms and keywords utilized to describe the articles. An example of a PubMed search is provided in the extended data availability.40 Title and abstract screening were performed to identify articles on persistent pain in patients who have undergone ARCR surgery. The search strategy will be adapted for every database that is included. Search will be conducted in electronic databases PubMed, CINAHL, Embase, and Scopus for further information. Articles published in the English language and full text available will be included. We will search the reference lists of all the included studies for any additional potentially eligible studies. Every study included needs to be published in a peer-reviewed academic publication. No date limitations will be applied in the review.\n\nFollowing the search, all references will be gathered and added to the Rayyan online platform for systematic reviews developed by the Qatar Computing Research Institute in Doha, all duplicate references will be eliminated. Title and abstract screening will be carried out by two separate reviewers for the scoping review, using the inclusion and exclusion criteria. Two reviewers will thoroughly evaluate the complete content of possible eligible references in a second phase based on the eligibility criteria. Any disagreements between the two assessors at any stage of the screening procedure shall be resolved by discussion or by consulting a third assessor. A narrative scoping review summarizing the search findings, and the research inclusion procedure will be reported, along with a PRISMA flowchart.\n\nTwo independent reviewers will use a standardized form to collect data from the final selected publications. The reviewers have developed a data extraction tool containing information that needs to be extracted from the articles.41 The extracted information will include details about participants, concepts, study methodology, context, and important conclusions related to the review question. Details identifying the biological factors, psychological factors, or social determinants of health in the pre-operative, intraoperative, and/or post-operative period will be included. The data extraction tool will be piloted by two reviewers who will also make any necessary modifications. The final scoping review will include a detailed analysis of the modifications. Any disagreements between reviewers will be settled by discussion or by bringing in a third reviewer. We will contact the authors if there is any information missing or needed.\n\nInitially, a descriptive analysis of each study’s findings will be completed. The data extraction will be displayed in a tabular manner following the objectives of this scoping review. A preliminary version outlining the attributes of the studies included has been formulated for this scoping review and it may undergo further revision during the review.42 The presentation of data will be categorized as described in the data extraction tool. At the individual predictor level, the consistency of reporting will be undertaken to determine the variables showing significant association with persistent pain. The review’s results will be summarized and reported to outline the factors contributing to persistent pain in patients receiving ARCR.43 A narrative summary will be provided along with the tabulated results to explain how the findings correspond with the purpose and research inquiry of the scoping review. Research gaps will be summarized in a narrative report regarding factors associated with persistent pain after ARCR. We adhered to the Preferred Reporting Items for Systematic Reviews and Meta-Analysis Extension for Scoping Reviews (PRISMA-ScR) checklist to ensure standardized reporting.44\n\n\nDisclosure statement\n\nThe authors have no conflict of interest to report.\n\n\nEthics and consent\n\nEthics and consent are not applicable for this study.", "appendix": "Data availability\n\nNo data are associated with this article.\n\nFigshare: Extended Data File 1: Preprint, https://doi.org/10.6084/m9.figshare.26885746 38\n\nFigshare: Extended Data File 2: Search String, https://doi.org/10.6084/m9.figshare.26954635.v1 40\n\nFigshare: Extended Data File 3: Data extraction form, https://doi.org/10.6084/m9.figshare.26954698.v1 41\n\nFigshare: Extended Data Table 1: Characteristics of Included studies, https://doi.org/10.6084/m9.figshare.26954710.v1 42\n\nFigshare: Extended Data Table 2: Risk factors for persistent pain after arthroscopic rotator cuff repair, https://doi.org/10.6084/m9.figshare.26954833.v1 43\n\nFigshare: Extended Data File 4: PRISMA-ScR Checklist, https://doi.org/10.6084/m9.figshare.26936515.v1 44\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\nKuppens K, Hans G, Roussel N, et al.: Sensory processing and central pain modulation in patients with chronic shoulder pain: A case-control study. Scand. J. Med. Sci. Sports. 2018; 28: 1183–1192. PubMed Abstract \| Publisher Full Text\n\nLucas J, van Doorn P , Hegedus E, et al.: A systematic review of the global prevalence and incidence of shoulder pain. BMC Musculoskelet. Disord. 2022; 23: 1073. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRahman H, Currier E, Johnson M, et al.: Primary and secondary consequences of rotator cuff injury on joint stabilizing tissues in the shoulder. J. Biomech. Eng. 2017; 139: 110801–110810. PubMed Abstract \| Publisher Full Text\n\nSambandam SN, Khanna V, Gul A, et al.: Rotator cuff tears: An evidence-based approach. World J. Orthop. 2015; 6: 902–918. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nLambers Heerspink FO, Dorrestijn O, van Raay JJAM , et al.: Specific patient-related prognostic factors for rotator cuff repair: A systematic review. J. Shoulder Elb. Surg. 2014; 23: 1073–1080. PubMed Abstract \| Publisher Full Text\n\nAkhtar A, Richards J, Monga P: The biomechanics of the rotator cuff in health and disease – A narrative review. J. Clin. Orthop. Trauma. 2021; 18: 150–156. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nVan der Meijden OA, Westgard P, Chandler Z, et al.: Rehabilitation after arthroscopic rotator cuff repair: Current concepts review and evidence-based guidelines. Int. J. Sports Phys. Ther. 2012; 7: 197–218. PubMed Abstract\n\nAckerman IN, Fotis K, Pearson L, et al.: Impaired health-related quality of life, psychological distress, and productivity loss in younger people with persistent shoulder pain: A cross-sectional analysis. Disabil. Rehabil. 2022; 44: 3785–3794. PubMed Abstract \| Publisher Full Text\n\nOh LS, Wolf BR, Hall MP, et al.: Indications for rotator cuff repair: A systematic review. Clin. Orthop. Relat. Res. 2007; 455: 52–63. PubMed Abstract \| Publisher Full Text\n\nHarrison AK, Flatow EL: Subacromial impingement syndrome. J. Am. Acad. Orthop. Surg. 2011; 19: 701–708. PubMed Abstract \| Publisher Full Text\n\nAbechain JJK, Godinho GG, Matsunaga FT, et al.: Functional outcomes of traumatic and non-traumatic rotator cuff tears after arthroscopic repair. World J. Orthop. 2017; 8: 631–637. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKukkonen J, Joukainen A, Itälä A, et al.: Operatively treated traumatic versus non-traumatic rotator cuff ruptures: A registry study. Ups. J. Med. Sci. 2013; 118: 29–34. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nTeratani T: Comparison of the epidemiology and outcomes of traumatic and nontraumatic rotator cuff tears. J. Orthop. 2017; 14: 166–170. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nNho SJ, Brown BS, Lyman S, et al.: Prospective analysis of arthroscopic rotator cuff repair: Prognostic factors affecting clinical and ultrasound outcome. J. Shoulder Elb. Surg. 2009; 18: 13–20. PubMed Abstract \| Publisher Full Text\n\nChung SW, Park JS, Kim SH, et al.: Quality of life after arthroscopic rotator cuff repair: Evaluation using SF-36 and an analysis of affecting clinical factors. Am. J. Sports Med. 2012; 40: 631–639. PubMed Abstract \| Publisher Full Text\n\nEnsor KL, Kwon YW, Dibeneditto MR, et al.: The rising incidence of rotator cuff repairs. J. Shoulder Elb. Surg. 2013; 22: 1628–1632. PubMed Abstract \| Publisher Full Text\n\nMalavolta EA, Assunção JH, Beraldo RA, et al.: Rotator cuff repair in the Brazilian Unified Health System: Brazilian trends from 2003 to 2015. Rev. Bras. Ortop. 2017; 52: 501–505. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMather RC, Koenig L, Acevedo D, et al.: The societal and economic value of rotator cuff repair. J. Bone Joint Surg. Am. 2013; 95: 1993–2000. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWang KY, Puvanesarajah V, Marrache M, et al.: Ambulatory surgery centers versus hospital outpatient departments for orthopaedic surgeries. J. Am. Acad. Orthop. Surg. 2022; 30: 207–214. PubMed Abstract \| Publisher Full Text\n\nNarvani AA, Imam MA, Godenèche A, et al.: Degenerative rotator cuff tear, repair or not repair? A review of current evidence. Ann. R. Coll. Surg. Engl. 2020; 102: 248–255. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nStiglitz Y, Gosselin O, Sedaghatian J, et al.: Pain after shoulder arthroscopy: A Prospective study on 231 cases. Orthop. Traumatol. Surg. Res. 2011; 97: 260–266. PubMed Abstract \| Publisher Full Text\n\nPham TT, Bayle Iniguez X, Mansat P, et al.: Postoperative pain after arthroscopic versus open rotator cuff repair. A prospective study. Orthop. Traumatol. Surg. Res. 2016; 102: 13–17. PubMed Abstract \| Publisher Full Text\n\nSchug SA, Lavand’homme P, Barke A, et al.: The IASP classification of chronic pain for ICD-11: Chronic postsurgical or posttraumatic pain. Pain. 2019; 160: 45–52. PubMed Abstract \| Publisher Full Text\n\nNeil MJ, Macrae WA: Post surgical pain- the transition from acute to chronic pain. Reviews in Pain. 2009; 3: 6–9. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRosenberger DC, Pogatzki-Zahn EM: Chronic post-surgical pain – Update on incidence, risk factors and preventive treatment options. BJA Educ. 2022; 22: 190–196. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nChen Y, Li H, Qiao Y, et al.: Double-row rotator cuff repairs lead to more intensive pain during the early postoperative period but have a lower risk of residual pain than single-row repairs. Knee Surg. Sports Traumatol. Arthrosc. 2019; 27: 3180–3187. PubMed Abstract \| Publisher Full Text\n\nHoofwijk DMN, Fiddelers AAA, Peters ML, et al.: Prevalence and predictive factors of chronic postsurgical pain and poor global recovery 1 year after outpatient surgery. Clin. J. Pain. 2015; 31: 1017–1025. PubMed Abstract \| Publisher Full Text\n\nGwilym SE, Oag HC, Tracey I, et al.: Evidence that central sensitisation is present in patients with shoulder impingement syndrome and influences the outcome after surgery. J. Bone Joint Surg. (Br.). 2011; 93-B: 498–502. PubMed Abstract \| Publisher Full Text\n\nValencia C, Fillingim RB, Bishop M, et al.: Investigation of central pain processing in postoperative shoulder pain and disability. Clin. J. Pain. 2014; 30: 775–786. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPotter MQ, Wylie JD, Greis PE, et al.: Psychological distress negatively affects self-assessment of shoulder function in patients with rotator cuff tears. Clin. Orthop. Relat. Res. 2014; 472: 3926–3932. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRaman J, Walton D, MacDermid JC, et al.: Predictors of outcomes after rotator cuff repair – A meta-analysis. J. Hand Ther. 2017; 30: 276–292. PubMed Abstract \| Publisher Full Text\n\nRavindra A, Barlow JD, Jones GL, et al.: A prospective evaluation of predictors of pain after arthroscopic rotator cuff repair: Psychosocial factors have a stronger association than structural factors. J. Shoulder Elb. Surg. 2018; 27: 1824–1829. PubMed Abstract \| Publisher Full Text\n\nvan Driel MEC , van Dijk JFM , Baart SJ, et al.: Development and validation of a multivariable prediction model for early prediction of chronic postsurgical pain in adults: A prospective cohort study. Br. J. Anaesth. 2022; 129: 407–415. PubMed Abstract \| Publisher Full Text\n\nToma O, Persoons B, Pogatzki-Zahn E, et al.: PROSPECT guideline for rotator cuff repair surgery: Systematic review and procedure-specific postoperative pain management recommendations. Anaesthesia. 2019; 74: 1320–1331. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nBasat HÇ, Uçar DH, Armangil M, et al.: Post-operative pain management in shoulder surgery: Suprascapular and axillary nerve block by arthroscope assisted catheter placement. Indian J. Orthop. 2016; 50: 584–589. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPeters MDJ, Godfrey C, McInerney P, et al.: Chapter 11 Scoping reviews. JBI Manual for Evidence Synthesis. Aromataris E, Munn Z, editors. Adelaide: JBI; 2020.\n\nTricco AC, Lillie E, Zarin W, et al.: PRISMA extension for scoping reviews (PRISMA-ScR): Checklist and explanation. Ann. Intern. Med. 2018; 169: 467–473. PubMed Abstract \| Publisher Full Text\n\nPrabhu A, Maiya AG, Pandey V, et al.: Factors contributing to persistent shoulder pain after arthroscopic rotator cuff repair Protocol for a scoping review.docx (Version 1). figshare. 2024. Publisher Full Text\n\nGreenspoon JA, Petri M, Warth RJ, et al.: Massive rotator cuff tears: Pathomechanics, current treatment options, and clinical outcomes. J. Shoulder Elb. Surg. 2015; 24: 1493–1505. PubMed Abstract \| Publisher Full Text\n\nPrabhu A, Pandey V, Maiya G, et al.: Search, 1st version. figshare. 2024. Publisher Full Text\n\nPrabhu A, Maiya AG, Pandey V, et al.: Data Extraction Form, 1st version. figshare. 2024. Publisher Full Text\n\nPrabhu A, Maiya AG, Pandey V, et al.: Characteristics of Included Studies, 1st version. figshare. 2024. Publisher Full Text\n\nPrabhu A, Maiya AG, Pandey V, et al.: Risk Factors for Persistent Pain After Arthroscopic Rotator Cuff Repair, 1st version. figshare. 2024. Publisher Full Text\n\nPrabhu A, Maiya GA, Pandey V, et al.: PRISMA ScR Checklist for Factors Contributing to Persistent Shoulder Pain After Arthroscopic Rotator Cuff Repair: Protocol for a Scoping Review, 1st version. figshare. 2024. Publisher Full Text" }	[ { "id": "332398", "date": "25 Oct 2024", "name": "Sanjay Tejraj Parmar", "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\n1. Why are pharmacological and non pharmacological studies providing interventions for pain relief not considered in this review 2. Please include all details of data extraction 3. Process of resolving disagreements between reviewer should be clearly outlined 4. Referencing needs correction\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": "12910", "date": "12 Dec 2024", "name": "Anupama Prabhu", "role": "Author Response", "response": "Thank you for your suggestion regarding including pharmacological and non-pharmacological interventions. Our primary aim is to identify the biological, psychological, and social factors that contribute to persistent pain lasting three months or longer after arthroscopic rotator cuff repair surgery in patients with rotator cuff tears. Including studies that aimed at comparing pharmacological and non-pharmacological interventions for pain relief falls outside the scope of our review as we do not aim at evaluating the effectiveness of specific interventions but rather to understand the multifaceted contributors to chronic post-surgical pain. Further, studies comparing different surgical procedures will also not be included for the same reason. This allows us to remain focused on understanding the chronic pain predictors, which could guide future research and interventions. Thank you for your valuable feedback. We have revised the section on data extraction in our manuscript and included additional details to enhance clarity and transparency in our review protocol. We hope this addresses your concern effectively and improves the overall quality of our manuscript. The process of resolving disagreements between reviewers has been outlined in our manuscript under the heading “study selection”. Sorry for the oversight, references have been checked for appropriateness and revised where necessary." } ] }, { "id": "336755", "date": "25 Nov 2024", "name": "Divya Bharatkumar Adhia", "expertise": [ "Reviewer Expertise Chronic musculoskeletal pain" ], "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 aims and methodology of this scoping review has been well presented.\nI only have one comment:\nThe authors could consider conducting quality assessment for all the papers included in the review. This will add value to interpreting findings from the studies and evaluating any mixed results from multiple studies.\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": "12911", "date": "12 Dec 2024", "name": "Anupama Prabhu", "role": "Author Response", "response": "Thank you for your thoughtful suggestion regarding the inclusion of a quality assessment for the studies in our review. We agree that conducting a quality assessment would enhance the interpretation of findings and help evaluate any mixed results. Accordingly, we plan to assess the quality of all included articles using the Newcastle-Ottawa Quality Assessment Scale. This addition will ensure a more robust and comprehensive evaluation of the evidence." } ] } ]	1	https://f1000research.com/articles/13-1236
https://f1000research.com/articles/13-1293/v1	29 Oct 24	{ "type": "Research Article", "title": "Hepatic GPx1 and GIT1 expression altered by ethanol exposure during third trimester-equivalent development", "authors": [ "KARICK JOTTY-Arroyo", "Albert Díaz--Castillo", "Harold Gomez--Estrada", "Rafael Pineda--Aleman", "Albert Díaz--Castillo", "Harold Gomez--Estrada", "Rafael Pineda--Aleman" ], "abstract": "Background Ethanol (EtOH) exposure throughout gestation and breastfeeding leads to multiple adverse outcomes in the hepatic system. Under oxidative stress, alterations in the liver are related to the inhibition of induced nitric oxide synthase activity in sinusoidal cells as a consequence of low expression of G-protein-coupled receptor (GPCR)-kinase interacting (GIT1). Here, we hypothesized that both glutathione peroxidase 1 (GPx1) and GIT1 could be altered by EtOH exposure during the third trimester of human equivalent development.\n\nMethods We exposed rats during the third trimester equivalent [postnatal days (PD) 2-8] to moderate levels of maternal EtOH (20%). GPx1 and GIT1 expression was detected by western blotting, and the antioxidant activity of glutathione peroxidase GPx and the concentration of hepatic carbonyl groups (CG were determined by spectrophotometry. Serum biochemistry parameters such as alanine aminotransferase (ALT), glucose (gluc), cholesterol (chol), and triglycerides (TG) were also measured.\n\nResults We found that ethanol decreased both GIT1 and GPx1 selenoprotein expression, affecting GPx antioxidant activity and increasing protein oxidation.\n\nConclusions These results demonstrate for the first time that the GPx antioxidant system altered by EtOH exposure during the third trimester of development is related to a parallel decrease in GIT1 expression [1].", "keywords": [ "GPx1-antioxidant", "GIT1", "Hepatic", "Ethanol-exposed" ], "content": "1. Introduction\n\nPrenatal ethanol (EtOH) exposure can result in significant negative consequence for offspring, such as impaired growth and range of congenital abnormalities, including Fetal Alcohol Spectrum Disorders (FASD).2–4 Numerous studies have demonstrated that sustained ethanol intake by mother disrupts the antioxidant homeostasis in their offspring.5–9 The detrimental teratogenic effects of this exposure are largely due to imbalance in redox processes, with the liver being particularly susceptible to ethanol-induced oxidative stress.10,11 Long-term alcohol consumption triggers the activation of Hepatic Stellate Cells (HSCs), which is linked to vascular changes and liver fibrosis resulting from increased deposition of Extracellular Matrix (ECM) proteins.11 In rodent models of cirrhosis induced by oxidative stress, sinusoidal capillarization occurs, involving the differentiation of Liver Sinusoidal Endothelial Cells (LSECs). This process is critical in HSC activation and fibrosis development.12,13\n\nEthanol exposure during gestation and breastfeeding impairs hepatic antioxidant glutathione peroxidase (GPx) activity and increases carbonyl groups in proteins.5,7 Consistent with this effect, pups of ethanol-exposed dams show some variation in the expression of selenium-dependent enzymes of the GPx family in liver tissue. Specifically, an increase in GPx4 expression levels has been reported at the expense of a decrease in GPx1 expression, likely due to GPx4’s physiological role in protecting phospholipids associated with several protein complexes of the mitochondrial inner membrane, such as cardiolipin. This protection helps inhibit cytochrome C release and maintain mitochondrial membrane potential and ATP generation.10,14,15\n\nIn the liver, acute and subacute oxidative stress results in the inhibition of induced nitric oxide synthase activity in sinusoidal cells through distinct mechanisms dependent on caveolin-1 inhibition, endothelin B receptor (ETBR) dissociation, and endothelial nitric oxide synthase (eNOS) phosphorylation.16 These alterations could influence the distribution and function of G-protein-coupled receptor kinase interacting protein-1 (GIT1), which is predominantly restricted to sinusoidal cells lining blood vessels and bile ducts.17 Furthermore, GIT1 is a multi-domain scaffold protein related to the regulation of cytoskeletal dynamics during cell spreading and migration, receptor internalization, and synapse formation.18\n\nHowever, GIT1 also possesses intrinsic signaling capabilities in endothelial tissues. For instance, when fibroblasts adhere to the fibronectin extracellular matrix, GIT1 is phosphorylated by the tyrosine kinase Src.19,20 Furthermore, after endothelin 1 (ET-1) binds to the ETB receptor, eNOS interacts with GIT1, leading to eNOS activation.20 Interestingly, eNOS phosphorylation and activity are reduced in sinusoidal endothelial cells following liver injury,20–22 and oxidative stress has been associated with endothelial dysfunction alterations in both ET-1 and nitric oxide (NO) signaling pathways.23,24\n\nTherefore, the third trimester of development is particularly vulnerable, and ethanol exposure induces multiple physiological alterations.25–32 Research has shown that ethanol consumption during pregnancy is relatively common.33 Given that exposure to ethanol during this developmental period in rodents has been used to model human exposure during the third trimester of pregnancy,4 we investigated whether ethanol-induced oxidative stress in the liver during this critical period results in alterations in the antioxidant proteins GPx1 and GIT1 in the liver tissues of offspring.\n\n\n2. Methods\n\nUnless otherwise indicated, all chemicals used in this research were obtained from PanReac AppliChem (Barcelona, Spain).\n\nAll animal procedures were approved by the University of Cartagena Animal Care and Use Committee and conformed to the NIH Guidelines, as well as the title V of Resolution 008430 of 1993 in Colombia. The study was conducted according to the ARRIVE guidelines 2.0, using the ARRIVE Essential 10 checklist for preclinical animal studies.34 Male and female Wistar rats (National Institute of Health, Colombia), weighing approximately 150–200 g, were randomized into two groups: control (C) and ethanol (EtOH) (E). The animals were maintained at an automatically controlled temperature (22–23°C) and a 12-h light–dark cycle (6:00–18:00).\n\nAll animal procedures were approved by the University of Cartagena Animal Care and Use Committee under Reference Number: 106 (approved on March 15th, 2018) and conformed to the NIH Guidelines, as well as the title V of Resolution 008430 of 1993 in Colombia. Animals were handled with care, provided with appropriate housing and management conditions, and anesthetics and analgesics were administered when necessary to alleviate any potential pain or discomfort.\n\nAnimals were exposed to ethanol starting on tap water that contained 5% v/v ethanol in the first and second weeks, and the ethanol concentration was increased to 10% in the third week, 15% in the fourth week, and 20% in the fifth week (induction period), which was maintained during the mating period. Pregnant rats were housed separately from male rats. To model third trimester-equivalent EtOH exposure, 20% ethanol in drinking water as the sole source of liquid with food ad libitum was restored to dams and their pups from postnatal days (PD) 2 to 8. During the paradigm, The gestational index calculated as (nSBnPR)100(number of Successful Births (nSB), number of Pregnancy Rats (nPR)) was measured. In addition, morphometric parameters (Total Body Weight (BW) and craniocaudal length) were recorded at PD2 and PD8 using an electronic balance (Precisa XB220A, Gravimetrics AG) and a metric caliper (Digi-Max™ slide caliper Z503576, Sigma Aldrich). Additionally, Liver Weight (LW) and hepatic somatic index (HIS) calculated as (LWBW)100, (Liver weight (LW), Body weight (BW)) were registered at PD8. Finally, at the end of ethanol exposure, serum biochemical parameters, such as alanine aminotransferase (ALT), glucose (gluc), cholesterol (chol), and triglycerides (TG) were measured using an automated analyzer (Technicon RA-1000, Bayer Diagnostics).\n\nAt the end of the experimental period, rats were anesthetized with ketamine (250 mg/kg, i.p. Sigma-Aldrich, Cat. Number 1356009, St. Mo, USA). The abdomen was opened using a midline incision, and the entire liver was removed. Samples were immediately covered with liquid nitrogen and stored at -80°C prior to biochemical analysis. Blood was collected by heart puncture and centrifuged to obtain serum.\n\nThe hepatic tissue was sonicated in homogenization buffer (0.1 g tissue/1 mL of buffer) containing 25 mM HEPES (pH 7.4), 500 mM NaCl, 2 mM EDTA, 1 mM dithiothreitol, 0.1% Tween-2020 (Biorad, Cat. Number 170-6531. CA, USA), and 1 tablet in 10 mL of the Pierce Protease and Phosphatase Inhibitor Mini Tablets (Thermo Scientific, Cat. number A32961). Samples were centrifuged at 3000 rpm at 4 °C for 10 min to obtain supernatants rich in hepatic proteins and stored in 10 μL aliquots at −80°C. Protein concentration was determined by the Bradford Method (BioRad, Hercules, CA, USA) using bovine serum albumin as a standard and stored at −40 °C.35\n\nSamples were mixed with sodium dodecyl sulfate polyacrylamide gel electrophoresis sample buffer (final concentration: 250 mM Tris–HCl (pH 6.8), 10% sodium dodecyl sulfate, 30% glycerol, 0.02% bromophenol blue and 5% β-mercaptoethanol (BioRad, Cat. number 161-0710)) and boiled at 95°C for 5 min. Samples were loaded at a concentration of 50 μg per lane. Control experiments demonstrated that this protein concentration was within the linear dynamic range for the western blot assay (not shown). Electrophoresis was performed on a 12% polyacrylamide gel at 150 V for 60 min at 4°C. Proteins were blotted onto polyvinylidene fluoride membranes (0.4 μm pore size. BioRad, Cat. Number 162-0177) at 100 V for 60 min at 4°C. Non-specific binding was blocked with 5% non-fat dry milk (BioRad, Cat. number 170-6404) in PBS for 1 h at room temperature and probed overnight at 4°C with either of the following specific primary antibodies: anti GPx1 antibody (Invitrogen, 1:5000 Cat. number PA5-30593) and anti GIT1 antibody (Invitrogen, 1:2,000, Cat. number PA5-78480; Invitrogen). Membranes were then incubated with a secondary antibody (goat anti-rabbit IgG (H+L) horseradish peroxidase conjugate, Invitrogen, Cat. number G21234;) at dilutions of 1:20,000 for GPx1 and GIT1. Monoclonal mouse anti-β-actin (Invitrogen, Cat. number MA1-140) was used to detect β-actin as a loading control, with a dilution of 1:20000, and a secondary antibody goat anti-mouse IgG (H+L), HRP (Thermo-Scientific, Cat. number 31430) was used at a dilution of 1:30000. Membranes were washed with PBS-milk-Tween 20, PBS-milk, and PBS for 5 min and visualized by chemiluminescence (Novex™ ECL Chemiluminescent Substrate Reagent Kit. Invitrogen, Cat. Number wp20005) for 3 min. The images were captured using a ChemiDoc ™ System (Bio-Rad, Hercules, CA, USA), and the intensity of the bands was analyzed using Image Lab 6.1 software (Biorad), For each sample, the protein expression was normalized to that of β-actin. The results were also used to calculate the GIT1/GPx1 ratio.\n\nTo measure the activity of GPx as well as the oxidation of protein, liver tissue samples were homogenized in a sucrose buffer (15 mM Tris/HCl, pH 7.4, 250 mM sucrose, 1 mM EDTA, and 1 mM DTT) in an ice bath, and the resulting supernatant was used for biochemical assays. The acquired data were also used to determine the ratio of GPx1 expression/GPx activity. Protein oxidation was measured according to a method based on spectrophotometric detection of the reaction of 2.4-dinitrophenylhydrazine (DNPH) with protein carbonyl (PC) to form protein hydrazones.36 The PC level was calculated at the maximum absorbance (366 nm), and the results were expressed as nmol/mg protein. The activity of the selenoprotein GPx was determined by NADPH-coupled assay according to the method of,36 which catalyzes the oxidation of glutathione by hydrogen peroxide. NADPH oxidation was monitored spectrophotometrically at 340 nm wavelength. Specific activity was expressed as mU/mg protein, where 1 mU is equal to nanomoles of NADPH oxidized/min.\n\nAll data were statistically analyzed using Prism 5 (GraphPad, San Diego, CA, USA) https://www.graphpad.com/. Initially, data were analyzed using the Pearson omnibus normality test. Only data that followed a normal distribution were analyzed using parametric tests. Statistical significance was set at p < 0.05.\n\n\n3. Results\n\nTo model EtOH exposure during the third trimester equivalent, dams and their offspring were exposed to EtOH (20%) ad libitum from PD2 to PD8. As shown (Table 1), our model confirms that ethanol exposure leads to lower fertility and gestational index. To determine whether ethanol alters regular growth during this developmental stage, we recorded the morphometric parameters. In this context, the total BW at PD2 showed no alteration in the offspring exposed to ethanol compared to that in the control group (Control, n=29; EtOH, n=20; t=1.91, df= 45, p> 0.05, unpaired t-test). However, there were significant differences in pup weight between the control and ethanol group at the end of the exposure paradigm (PD8) (control, n=29; EtOH, n=20; t=2.28, df= 36, p<0.05, unpaired t-test). Similarly, the cranium-caudal length showed no difference in offspring from dams exposed to EtOH at PD2 (control, n=29; EtOH, n=20; t=1.93, df= 48, p> 0.05, unpaired t-test). In contrast, at the end of the ethanol exposure (PD8), alcohol-exposed pups registered a lower cranium-caudal length compared to the control (Control n=29; EtOH n=20; t=3.22, df= 48, p<0.01 by unpaired t-test). Despite the morphometric differences registered at the end of the ethanol exposure paradigm, the LW and HIS were similar in both groups.\n\n* p<0.05.\n\n** p<0.01.\n\nAdditionally, biochemical analysis of serum (Table 2) revealed that EtOH-treated animals had a significant increase in ALT enzyme activity (n=8; t=8.96, df= 14, p<0.001 by unpaired t-test), and in the products of lipid metabolism such as cholesterol (n=8; t=5.02, df= 14, p<0.001 by unpaired t-test), and triglycerides (n=8; t=2.19, df= 14, p<0.05, unpaired t-test). However, offspring of EtOH-exposed dams had significantly lower glucose concentrations (n=8; t=2.42, df= 14, p<0.05 by unpaired t-test).\n\n* p<0.05.\n\n*** p<0.001.\n\nTo determine whether EtOH exposure during the third trimester alters the hepatic redox balance, we first evaluated GPx1 expression. Western blot analysis of total GPx1 within the liver tissue (Figure 1A) showed a significant reduction in EtOH exposure when normalized to β-actin (Figure 1B) (n=8; t=3.12, df= 14, p<0.01 by unpaired t-test). Knowing that hepatic oxidative stress alters parenchymal and non-parenchymal cells, similar to those found in sinusoidal ducts (34), and that GIT1 is restricted to cell lining blood vessels and bile ducts (16), we also evaluated whether liver ethanol-oxidative stress affected total GIT1 expression. Similarly, we found significant differences between the control and EtOH-exposed offspring when data were normalized to β-actin (Figure 1A and C) (n=8; t=2.99, df= 14, p<0.01 by unpaired t-test). As a result of this altered expression, the GIT1/GPx1 expression ratio was also reduced by ethanol exposure (Figure 1D) (n=8; t=2.19, df= 14, p<0.05 by unpaired t-test). Taken together, these data show that ethanol modulates the expression of GPx1 and GIT1, resulting in the depletion of both proteins during the third trimester equivalent of development.\n\n(A) Exemplar western blot illustrating levels of GIT1, GPx1 and β-actin from control and ethanol rats. (B) Graphic quantification of GPx1 expression normalized to β-actin. GPx1 was altered by ethanol exposure. (C) Graphic quantification of GIT1. GIT1 was also altered by ethanol exposure. (D) Ratio GIT1/GPx1 expression. Labels of C5 to 8 and A5 to -8 are representative samples of each experimental group. The results are expressed as mean ± SEM and analyzed by unpaired t-test. The number of animals in each group is 8. Statistic difference between groups was expressed as p value: C vs E: * p<0.05, p<0.01, * p<0.001.\n\nTo confirm that the low expression of GPx1 selenoprotein is related to an altered hepatic redox balance, we determined GPx enzyme activity. Figure 2A shows a significant decrease in GPx activity in ethanol-exposed offspring at PD8 compared to that in the control (n=8; t=5.59, df= 14, p<0.001 by unpaired t-test). Furthermore, to evaluate the consequences of the altered antioxidant GPx system, we examined the levels of protein oxidation in the liver. Conversely, the data collected showed that pups from EtOH-treated dams had significantly higher concentrations of carbonyl groups (n=8; t=4.16, df= 14, p<0.001 by unpaired t-test) (Figure 2B). Additionally, we found that when GPx1 expression was compared to GPx activity, the GPx1expression/GPx activity ratio decreased significantly in the liver after ethanol exposure (Figure 2C) (n=8; t=5.66, df= 14, p<0.001 by unpaired t-test).\n\n(A) Ethanol exposed-offspring showed a significantly reduced GPx activity. (B) Carbonyl groups were increased by ethanol exposure. (C) Ratio of GPx1expression/GPx activity. The results are expressed as mean± SEM and analyzed by unpaired t-test. The number of animals in each group is 8. Statistic difference between groups was expressed as p value: C vs E: *** p<0.001.\n\n\n4. Discussion\n\nIn this study, our data show that ethanol treatment during the initial period (PD2) does not cause any detrimental effects on the BW of pups or their cranium-caudal length. However, during the breastfeeding period (PD2-8), ethanol exposure induced a growth deficit at the end of the paradigm (PD-8). These results are consistent with several studies that also reported reduced growth rates at the end of lactation7,8 or even during the embryonic E19 stage,37 although those studies used models with more sustained ethanol exposure. It is likely that the findings reported here are related to the fact that ethanol consumption during lactation provokes a greater under-nutrition in dams, in part because in this period, the ethanol-exposed dams consumed more ethanol during lactation than during gestation, which affects the milk intake of the suckling pups; reduced in some cases until 30% of the amount consumed by offspring not exposed to ethanol.8 It is also likely that the detrimental growth registered in our data is correlated to a low quality of the milk consumed by pups, as previous works have reported that the low BW is reversed under selenium (Se) supplement treatment; Se (0.5ppm) and Se+Folic acid (FC) (0.5+0.8ppm),8,38 probably as a consequence that both diet supplements are able to increase the Se concentration in milk or in organs such as mammary glands.6 This trace element seems to play an essential role in offspring development, since it is necessary to synthesize deiodinase enzymes that enhance T3 tissue levels, specifically at the hypothalamus, where it produces an increase in the Neuropeptide Y inducing hyperphagia.39,40 Our model also demonstrated that ethanol exposure altered optimal metabolic conditions in the offspring. In this context, the results showed increased values of cholesterol and triglycerides and a low concentration of glucose, together with elevated levels of ALT, suggesting that metabolism in ethanol-exposed offspring is altered, leading to liver damage and altered cell membrane permeability.41 Taken together, these assumptions could explain, at least in part, the reduced BW and length of the offspring under EtOH exposure during the third trimester equivalent development.\n\nAnalysis of GPx1 expression and GPx activity in the liver on PD8 demonstrated that both were affected by ethanol exposure, indicating the existence of mechanisms affecting the synthesis and activation of this protein. This is consistent with previous studies demonstrating that chronic ethanol exposure during gestation and breastfeeding causes drastic changes in the hepatic GPx antioxidant system.5,36,42 In fact, it has been proven that reduced GPx activity is inversely related to the gluthatione reductase (GR) activity in liver of pups from ethanol exposed dams, which is associated with a depletion of reduced gluthatione (GSH). This mechanism seems to be activated as a possible adaptation response to oxidative stress to maintain regular GSH levels or to reduce NADPH levels associated with ethanol intoxication.5 Nevertheless, it is also likely that the ratio GPx1 expression/GPx activity imbalance is related to a decreased Se content in the liver, as previous work has reported a depletion of GPx activity dependent on the hepatic Se stored in embryos and pups, and subsequent administration of Se as supplement produces a moderate increase in GPx activity.5,7,36 In this context, we could assume that lower selenium-dependent GPx1 expression and its antioxidant activity could be linked to hepatic Se depletion induced by ethanol exposure. However, it is necessary to perform a more detailed analysis to confirm that Se depletion is the only cause of GPx reduction, as in the alcohol-damaged newborn model, more than one parameter has been used to describe the Se status and its biological function.35,43\n\nTo determine whether decreased GPx1 expression and GPx activity in ethanol-exposed offspring have repercussions on liver oxidative balance, we evaluated their effects on protein oxidation. In fact, oxidative stress in the offspring is supported by an increase in carbonyl groups in hepatic proteins (PC). This is consistent with studies performed under sustained ethanol exposure during gestation that promote high levels of PC and decrease GPx activity in the liver and kidney of suckling pups.36,44 Taken together, we suggest that liver tissue is trying to cope with ethanol-induced oxidative stress, but is incapable of doing so because it suffers peroxidation of its proteins. However, it is worth noting that the oxidative stress and enzymatic antioxidant system differ among tissues, and its activation also depends on the physiological quality of the tissue environment.44 It is not yet completely understood how the antioxidant GPx system functions under these conditions.\n\nEthanol exposure during the third trimester also decreased GIT1 expression in the liver. It is likely that such alterations could be related to the impairment of oxidative balance, which in turn leads to alterations in parenchymal and non-parenchymal cells. In this context, the GIT1/GPx1 decreased ratio found in ethanol-exposed offspring indicates the existence of mechanisms that possibly alter the synthesis of both proteins, perhaps as a result of the augmented hepatic oxidative stress produced by the increment of reactive oxygen species (ROS) that promote depletion of GIT1 and GPx1, as has been observed in cultured fibroblasts45 and in the liver,46 respectively. In addition, it is known that GIT1 is restricted to cells lining blood vessels and bile ducts.17 In portal-based fibrogenic response and portal hypertension in the liver, GIT1 level is reduced in injured cells, and subsequent decreased eNOS phosphorylation and activity with a markedly reduced GIT1/eNOS co-localization in the perinuclear region persists even after injury.47 Based on this, it is also likely that the decreased expression of GIT1 in our model is a response to an altered physiological condition induced by ethanol-oxidative stress, as in fibrosis, which is characterized by an excessive accumulation of extracellular matrix (ECM) proteins with very little degradation.37,48 Expression of Kupffer cell-derived tumor necrosis factor alpha (TNF-α),49 and interleukins (IL-1, IL-6, IL-8) have been reported in bile duct cells.50 This information could explain the possible mechanism related to EtOH-induced oxidative stress on GIT1 low expression and its role during the third trimester equivalent offspring. However, the effect of EtOH on GIT1 expression may be a more complex process involving several factors (Figure 3). Significant evidence demonstrates that ethanol downregulates the endothelin B receptor (ETBR) and eNOS phosphorylation16 and likely also affects the G-protein-coupled receptor ligand endothelin-1 (ET-1). Oxidative stress is known to cause endothelial dysfunction and alterations in ET-1 signaling pathways.24 Together, these factors alter GIT1 expression and function. Additionally, ethanol can suppress Akt phosphorylation through cytochrome P4502E1 (CYP2E1)-induced oxidative stress in alcoholic fatty liver.51 This suppression may affect GIT1/eNOS association, as Akt phosphorylation activates eNOS (at Ser1177) and regulates the ability of Src to phosphorylate GIT1.20 Therefore, this is correlated with the fact that eNOS and NO production is reduced by sinusoidal endothelial cells as well as increased intrahepatic vascular resistance after liver injury.21,52 Furthermore, during preeclampsia, depletion of GIT1 impedes NO production and placental eNOS activity,53 and during the early neonatal period at different gestational ages, no correlation was found between ET-1 and the oxidative impairment product, MDA, and the tripeptide GSH.54 However, this could be explained by the effects of the transitional perinatal period, which reflect a mixed maternal and neonatal redox status.55\n\nET-1 activates its cognate ETB-R inducing Gα and Gβγ activation and dissociation. Akt and Src both are related to phosphorylation-enhanced GIT1/eNOS, eNOS activity and NO production. A) Ethanol exposure depletes GPx1 antioxidant system and increases ROS, which in turns, decreases endothelin B receptor, leading to a low co-expression and activation of GIT1/eNOS protein levels, and eNOS phosphorylation. B) Also, a decreased expression of GPx1 and increased ROS cytochrome P4502E1 (CYP2E1)-induced oxidative stress may be responsible for ethanol-induced suppression of Akt phosphorylation leading to downregulated of GIT1/eNOS expression. The consequence of altered GIT1 expression is reduced eNos activation and NO production. EtOH (Ethanol); ROS (Reactive Species Oxygen); GPx1 (Glutathione peroxidase 1); ET-1 (Endothelin-1); ETB-R (Endothelin b-Receptor); PI-3Kinase (Phosphatidylinositol (PI)-3 kinase); Akt (Protein kinase B); Src (Src kinase); (GIT1) (G-protein-coupled receptor (GPCR)-kinase interacting protein-1) eNOS (endothelial nitric oxide synthase); NO (Nitric Oxide).\n\nDespite evidence that GIT1 functions are associated with hepatic endothelial physiology, it is worth noting that altered GIT1 expression due to ethanol exposure in hepatic cells could also affect focal cell migration and adhesion, as it was found that GIT1 in pathologies such as hepatocellular carcinoma (HCC) promotes the invasion, migration, and proliferation of HCC cells. Moreover, overexpression of GIT1 prompted epithelial mesenchymal transition (EMT) by activating the extracellular regulated kinase 1/2 (ERK1/2) pathway.56\n\nOverall, it is clear that the third-trimester ethanol-exposed offspring have an impaired GPx1 antioxidant system in the liver. The altered antioxidant system seems to be related to the decreased expression of GIT1. It is possible that alterations in the transducing signal dependent on GIT1 underlie the mechanism of several harmful processes associated with ethanol exposure. Therefore, Future studies should focus on exploring the imbalance of the antioxidant system in the GIT1signaling pathway associated with hepatic LSEC injuries induced by ethanol exposure during this developmental period.\n\n\n5. Conclusions\n\nThe assays confirmed that EtOH exposure during gestation and lactation significantly affects the morphological and biochemical development of offspring, critically affecting the hepatic antioxidant system. The reduction in GPx1 expression and activity, together with decreased levels of GIT1, suggests altered signaling pathways and functional integrity of the liver. These results underscore the importance of avoiding ethanol exposure during critical developmental stages because of its long-lasting adverse effects on liver health and overall offspring development.\n\nAll animal procedures were approved by the University of Cartagena Animal Care and Use Committee under Reference Number: 106 (approved on March 15th, 2018) and conformed to the NIH Guidelines, as well as the title V of Resolution 008430 of 1993 in Colombia. Animals were handled with care, provided with appropriate housing and management conditions, and anesthetics and analgesics were administered when necessary to alleviate any potential pain or discomfort.", "appendix": "Data availability\n\nTo ensure transparency and reproducibility, we have deposited all the raw data underlying this study in https://figshare.com/articles/dataset/:Data of Hepatic GPx1 and GIT1 Expression Altered by Ethanol Exposure During Third Trimester-Equivalent Development. https://doi.org/10.6084/m9.figshare.27091666. 57\n\nFigshare: Data of Hepatic GPx1 and GIT1 Expression Altered by Ethanol Exposure During Third Trimester-Equivalent Development, https://doi.org/10.6084/m9.figshare.27091666. 57\n\nThis project contains the following underlying files:\n\n• Raw data Liver PC _ GPx activity_Jotty et aletal_2024/Analysis GPx and PC_ Alberto 2021.pzfx41.46 KB.xlsx Raw data Jotty et al_2024/Biochemestry parameters_Jotty et _2024.xlsx\n\n• Raw data Jotty et al_2024/GPx1 and GIT1 Expression_Jotty et al_2024.xlsx\n\n• Raw data Jotty et al_2024/Morphometric parameters_Jotty et al_2024.xlsx\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nFigshare: Protein Quantification of ethanol exposed PD8 rat offspring for determining Hepatic GPx1 and GIT1 Expression. https://doi.org/10.6084/m9.figshare.27091804. 58\n\nThis Project contains the following files:\n\n• Protein Quantification_Jotty et al_2024.xlsx 49.08 KB.\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nFigshare: ARRIVE checklist Hepatic GPx1 and GIT1 Expression Altered by Ethanol Exposure During Third Trimester-Equivalent Development. https://doi.org/10.6084/m9.figshare.26863453. 33\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 the following authors for their collaboration and commitment in the preparation of the preliminary work included in this project: Díaz Castillo A, Silgado Ortega JP, Jiménez Villalobos TP, Tuirán Ruiz M, Gómez Estrada H, Jotty Arroyo K. 1\n\n\nReferences\n\nDíaz A, Silgado JP, Jiménez TP, et al.: Ethanol exposure during the third trimester-equivalent of development affects GPx1 and scaffolding GIT1 protein expressions in liver. 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Publisher Full Text\n\nJotty K, Diaz A, Gomez H, et al.: Protein Quantification Of Ethanol Exposed Pd8 Rat Offspring For Determining Hepatic Gpx1 And Git1 Expression. Dataset. Figshare. [dataset modified on 2024-09-23]. Publisher Full Text" }	[ { "id": "336853", "date": "25 Nov 2024", "name": "Mauro Ceccanti", "expertise": [ "Reviewer Expertise Alcohol addiction", "Gastroenterology", "Alcohol Use Disorders." ], "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 titled \"Hepatic GPx1 and GIT1 Expression Altered by Ethanol Exposure During Third Trimester-Equivalent Development\" by Jotti-Arroyo et al. presents an intriguing study on the interaction between GIT1 and GPx in the livers of mice during the first trimester (days 1-8). The authors conclude that \"the GPx antioxidant system altered by ethanol exposure during the third trimester of development is related to a parallel decrease in GIT1 expression.\" This result indicates that the balance between oxidants and antioxidants is disrupted, leading to increased protein oxidation.\nThe study is well-articulated and provides all the necessary information for a thorough understanding. However, I have a suggestion for the authors: Table 2 could be made easier to interpret by including definitions of the acronyms in the table’s description.\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": "12906", "date": "11 Dec 2024", "name": "KARICK JOTTY", "role": "Author Response", "response": "We greatly appreciate the referee's valuable feedback and are grateful for the suggestion regarding Table 2. In adherence to the recommendation, we will make the suggested modification to enhance clarity by including the definitions of the acronyms in the table’s description. However, we wish to point out that the acronyms are already defined in the Methodology section. This adjustment aims to ensure ease of interpretation while maintaining consistency across the manuscript. Thank you for your thoughtful observation." } ] } ]	1	https://f1000research.com/articles/13-1293
https://f1000research.com/articles/11-1446/v1	07 Dec 22	{ "type": "Research Article", "title": "An open label, single arm, pilot study to evaluate the safety, tolerability, and efficacy of daily fluconazole 150 mg in subjects suffering from Tinea cruris and Tinea corporis", "authors": [ "Dattatray Gopal Saple", "Sushrut Save", "Devesh Kumar", "Suneet Sood", "Dattatray Gopal Saple", "Sushrut Save", "Devesh Kumar" ], "abstract": "Background: Dermatophytes are the most common superficial fungal infections worldwide and are treated with prescribed regimens of terbinafine and itraconazole, or with weekly doses of fluconazole. Dermatologists are increasingly encountering treatment failures, and experts suggest that standard treatment regimens are not applicable anymore. We planned an open-label study to evaluate the results of fluconazole 150 mg daily for 8 weeks in patients with tinea cruris and tinea corporis. Methods: Patients were enrolled from the La’Mer Clinic, Mumbai, India. We included adult subjects with uncomplicated dermatophytosis confirmed by microscopic examination of skin scrapings. Pregnancy, poor renal function, and recent exposure to anti-fungal therapy were exclusion criteria. Patients were reviewed on days 14, 28 and 56. The treating doctor scored the severity of erythema, scaling, and pruritus on a four-point scale: absent, mild, moderate, and severe. Of 107 subjects screened, 100 were finally included in the study. Eleven were lost to follow up and one subject withdrew consent. Results: The site of disease was body alone in 29, groin alone in 7, and both body and groin in 64 cases. At 5 weeks, 98%, 100%, and 97% of patients had no scaling, erythema, and pruritus, respectively. Skin scrapings showed 100% mycological cure. In one patient the alanine transaminase level rose from 54.9 to 100.2 U/L, and qualified as a grade 1 adverse event not requiring intervention. No other significant adverse events were noted. Conclusions: Our results suggest that fluconazole 150 mg daily for eight weeks effectively treats dermatophytosis. This regimen is safe and well-tolerated even in patients with co-morbidities. Fluconazole is about eight times less expensive than itraconazole or terbinafine and may be the preferred therapy. Registration: The trial was registered with Clinical Trials Registry, India (Registration number CTRI/2020/06/026110) on 24 June 2020. FDC Company, India, provided financial support for the study.", "keywords": [ "Dermatophytes", "skin infections", "fungal infections", "Tinea capitis", "antifungal agents", "itraconazole", "terbinafine", "tropical diseases", "erythema", "scaling", "pruritus" ], "content": "Introduction\n\nDermatophytes are the commonest superficial fungal infections worldwide. They include three genera of fungi: Trichophyton, Epidermophyton, and Microsporum.1 Treatment of localized lesions involves the use of topical antifungals such as luliconazole, sertaconazole, clotrimazole, terbinafine and others, or oral agents such as itraconazole, terbinafine, and griseofulvin.2 Oral agents effective against dermatophytes include the azoles (e.g., itraconazole, fluconazole) and the allylamines (terbinafine). The azoles act at the fungal P-450 pathway to prevent the formation of ergosterol, an important biomembrane lipid.3\n\nHowever, practicing dermatologists are increasingly encountering treatment failures, and reports of recalcitrant dermatophytosis have begun to appear in the literature. Yamada et al.,4 showed in 2017 that as many as 17% of Trichophyton strains showed reduced susceptibility to terbinafine. Monod and Mehul5 reviewed the literature to show that mutant strains of Trichophyton were responsible for this decreased sensitivity. Poojary et al.,6 noted a sudden unexplained surge of difficult-to-treat dermatophytosis in India, and experts gradually have come to suggest that standard treatment regimens are probably not applicable anymore.7,8\n\nFluconazole has traditionally not been used routinely in the treatment of dermatophytosis.1,7 There are increasing reports of inadequate response to therapy, relapses, recurrence and resistance to available antifungal drugs like terbinafine, itraconazole and griseofulvin. We therefore planned a study to determine the results of fluconazole therapy in cases of dermatophytosis. Using fluconazole 150 mg daily for 8 weeks, we evaluated clinical parameters and mycological cure rates in patients with tinea cruris and tinea corporis.\n\n\nMethods\n\nWritten informed consent was taken from subjects prior to their inclusion in the study. This study was approved by the Vision Independent Ethics Committee, Mumbai (independent Government-approved Ethics Committee), per its letter dated 13 June 2020. The study was registered with the Clinical Trials Registry – India on 24 June 2020 (registration number CTRI/2020/06/026110).\n\nThis research was a prospective, open label, single-center, non-comparative study.\n\nWe enrolled 100 patients with dermatophytosis (tinea cruris and tinea corporis) visiting the outpatient department of La’Mer Clinic, Mumbai, considering this number to be adequate for a pilot study. All the subjects were enrolled after they gave written informed consent. The first patient was enrolled on 22 July 2020 and the last subject was enrolled on 24 August 2021.\n\nWe included subjects with dermatophytosis confirmed by fungal hyphae in a 10% potassium hydroxide (KOH) preparation of skin scraping. Subjects were aged 18 to 65 years old.\n\nWe excluded patients with secondarily infected or eczematized lesions, and those who had received any type of oral or topical anti-fungal therapy within 14 days prior to baseline visit. We also excluded patients who were immunosuppressed, had a creatinine clearance < 30 ml/minute, patients who were pregnant or lactating, and those with any severe medical comorbidity. Diabetes was not an exclusion criterion unless it was uncontrolled.\n\nAll patients were administered fluconazole 150 mg daily for 8 weeks.\n\nIn the screening visit (“Visit 1”, day -4 to day 1) patients were examined and baseline full blood counts, renal function tests, liver function tests and urine physical, chemical and microscopy studies were carried out. Urine pregnancy tests were conducted in female patients. Fluconazole therapy was started in eligible subjects on the day of enrolment (“Visit 2”). They were then reviewed clinically and microbiologically on days 14 ± 2, 28 ± 2, and 56 ± 2 (“Visit 3”, “Visit 4”, and “Visit 5”).\n\nClinical review consisted of a subjective evaluation of three parameters: erythema, scaling, and pruritis. The treating doctor (GDS) scored the severity of each parameter on a four-point scale: 0 = absent, 1 = mild, 2 = moderate, 3 = severe.\n\nMycological review consisted of examination of a 10% KOH preparation of a skin scraping under direct microscopy. The result of the examination was reported as absent or present for fungal hyphae. Fungal culture was not carried out.\n\nPatients received an investigational product compliance diary card on which they were advised to record medication intake and adverse events. The treating doctor (GDS) reviewed diary entries at every visit.\n\nOn the final visit (day 56), the treating doctor also carried out a urine analysis and blood examination for full blood count, renal function tests and liver function tests.\n\nAlterations of aminotransferase levels were graded according to the criteria described by the US Department Of Health And Human Services per their 2017 document.9\n\nThe data were analyzed by simple mathematical calculation of response rates.\n\n\nResults\n\nOf 107 subjects screened, 100 were finally included in the study (Figure 1). Of these, 11 were lost to follow up and one subject withdrew consent. The mean age was 40 years old (SD 11.6). A total of 46 of the 107 subjects were female.\n\nTwenty two patients had comorbidities, of which the commonest were diabetes and hypertension (Table 1).10\n\nThe site of disease was body (Tinea corporis) alone in 29, groin (Tinea cruris) alone in 7, and both body and groin in 64 cases.\n\nScaling, erythema, pruritus, and mycological results improved over the 5 visits, as shown in the charts below.\n\nScaling was present in all patients at enrolment. The number of patients with severe scaling, and the severity of scaling decreased with treatment. At 5 weeks, 86 of 88 patients had no scaling; the other two had mild residual scaling (Figure 2).\n\nThe numbers are percentages of the total number of patients available for assessment.\n\nErythema was present in 97 of the 100 patients at enrolment. By the end of the fifth visit the erythema had disappeared in all cases (Figure 3).\n\nThe numbers are percentages of the total number of patients available for assessment.\n\nPruritus was present in all patients at enrolment. By the end of the fifth visit the pruritus had disappeared in 97% of cases (Figure 4).\n\nThe numbers are percentages of the total number of patients available for assessment.\n\nThus, the overall clinical cure rate was 97%, with two patients having mild residual scaling, and three having mild pruritus.\n\nSkin scrapings were examined for fungal hyphae on visits 1, 3, 4 and 5. By the fifth visit, mycological cure was 100% (Figure 5).\n\nThe numbers are percentages of the total number of patients available for assessment. KOH, potassium hydroxide.\n\nOne patient had an elevation of serum creatinine to 1.22 mg/dl (laboratory normal 0.67-1.17). This change did not reach the threshold to be considered an adverse event.\n\nMild elevations of liver enzymes were noted in a few patients between visit 1 and visit 5. (Table 2). Six patients, none with comorbidity, had alanine aminotransferase (ALT) levels above the upper limit of normal. In five, the levels just barely exceeded the upper limits of normal and did not reach the threshold for being considered Grade 1 adverse events. In one, the ALT level rose from 54.9 to 100.2 U/L, and qualified as a grade 1 adverse event only,9 not requiring intervention. Eight patients, two with comorbidity, had mild elevations of serum aspartate aminotransferase levels. The changes did not meet the criteria for being considered adverse events (Table 2).\n\na In no other patient did the change meet the threshold to be considered an adverse event;\n\nb Patients had no comorbidity;\n\nc hypertension;\n\nd diabetes. ALT, alanine aminotransferase; AST, aspartate aminotransferase.\n\n\nDiscussion\n\nDermatophytosis affects 20-25% of the world’s population.11 The prevalence is likely much higher in India12 and is certainly increasing to alarming, almost panicky levels.8,13\n\nThe burden of recalcitrant dermatophytosis is growing in India, with an estimated 3-10% of cases being difficult-to-treat.1,7,14–16 Almost every major dermatological conference has begun to devote an entire session to the discussion of dermatophytosis.14 Reports are also appearing from other parts of the world about the worrying increase in resistance.11,17\n\nThe emergence of recalcitrant tinea is almost certainly encouraged by the non-adherence to the treatment regimen and shorter duration of the regimen.16,18 A species shift to Trichophyton indotineae is another likely cause of the observed recalcitrance.19 Taking in consideration the significant number of recalcitrant cases, one of us (DGS) decided to conduct a trial of daily dose fluconazole for patients with dermatophytosis.\n\nThis study shows that at present one may expect a reliable response to oral fluconazole in a 150 mg daily dose regimen lasting five weeks. While there are several reports of the efficacy of weekly fluconazole for dermatophytosis,20 there is a paucity of literature regarding the daily use of fluconazole in dermatophytosis, and the clinicians have been discouraged from its use for treating this condition. The usual recommended dose is 150 mg once a week. At this dose the cure rates vary from 64%21 to about 90%.22,23 On the other hand, the daily dosing ensures that the drug concentrations consistently exceed the minimum inhibitory concentrations (MICs) required to kill the fungus. This result should come as no surprise: fluconazole is among the triazoles that is known to be effective in treating dermatophytosis. Indeed, members of an expert panel recently indicated that fluconazole and terbinafine were their preferred systemic choice of antifungal agents for children.13\n\nFluconazole is about eight times less expensive than itraconazole or terbinafine, therefore if the therapeutic effect is similar fluconazole may be the preferred therapy, as it is likely to ensure patient compliance for a long time. Considering that nearly 70% of infections occur in the low and very low income socioeconomic groups,24 cost becomes a critical factor in deciding the medication.\n\nIt is also worthy of note that fluconazole accumulates in the stratum corneum25 and its levels are very high when compared to other azoles.26 The in vitro MIC90 levels of fluconazole are 16–32 micrograms/ml,27,28 depending on the species. In the stratum corneum the tissue concentrations of fluconazole are more than the MICs for most dermatophytes, and the drug is eliminated 2–3 times more slowly than from the blood.29 Daily doses of fluconazole achieve levels of 66 micrograms/gram, about three times as high as levels after weekly doses,25 which explains why daily fluconazole is so effective in the treatment of dermatophytosis. Sardana et al.,26 and others reviewed the literature on the pharmacokinetics of antifungal agents, and stated that the distribution of itraconazole and terbinafine, being highly lipophilic, depended on the presence of sebum. Though itraconazole and terbinafine achieved MIC90 inhibition at lower levels in laboratory studies, these drugs also achieved lower levels in the skin.30 Standard antifungal sensitivity test (AFST) methods involving in vitro and animal data are unreliable for predicting clinical outcomes, and there is the suggestion that fluconazole might be the preferred drug, especially for sites that secrete less sebum or in persons with intrinsically dry skin (such as diabetics).26 Khurana et al.,31 in a recent review, pointed out that the 60–90 hour elimination half-life of fluconazole would be insufficient antifungal cover if the drug was given once weekly. We believe that it is likely that these properties of fluconazole provide a strong rationale for a daily dose regimen.\n\nSeveral studies have shown that fluconazole is well tolerated at daily doses of up to 1,600 mg.23,32–36 Hepatotoxicity is described, but rare, and in most cases is self-limiting. Mild elevations in serum aminotransferase levels occur in up to 5% of patients. These patients are asymptomatic and do not require discontinuation of the medication.37 In our cases none of the patients developed any significant adverse effect. The drug was well-tolerated even in patients who had existing comorbidities. Except in one patient, the transaminases remained stable, and showed a minimal elevation in only about 5% of patients.\n\nCross-reactivity of fluconazole with other azoles is unlikely, but fluconazole should nevertheless be used with caution in patients who have had adverse effects from itraconazole and related compounds.37 It is good to remember that fluconazole inhibits the cytochrome P450 enzyme CYP 3A4, and may cause interactions with drugs that are metabolized by this enzyme, though it is a less potent inhibitor of the pathway than are itraconazole and posaconazole.38\n\n\nConclusions\n\nOur results suggest that fluconazole in a dose of 150 mg daily for eight weeks effectively treats dermatophytosis, with a clinical efficacy of 97% and a mycological cure rate reaching 100%. The eight-week daily dose regimen is safe and well-tolerated, even in patients with co-morbidities. In view of the severe concerns about the applicability of existing recommendations for dermatophytosis, dermatologists should consider this regimen as first-line therapy. There is also a need for a large comparative study with itraconazole and terbinafine.\n\nThis study evaluates the efficacy of an inexpensive and safe drug, fluconazole, in the management of a major health disorder. The enrolled patients were carefully evaluated, and an attempt was made to ensure that the readings were as objective as was possible. Patients were also examined microbiologically to corroborate the clinical findings.\n\nIn a single center private clinic, it was not feasible to carry out fungal cultures due to financial limitations. However, we did not encounter any case that would require us to ask for fungal culture and anti-fungal sensitivity tests. Additionally, even expert panels do not recommend cultures except in recalcitrant cases.13\n\nWe also were not able to blind the measurement of the clinical responses to the drug.", "appendix": "Data availability\n\nHarvard Dataverse: Replication Data for: FLUCONAZOLE_150. https://doi.org/10.7910/DVN/P7DLSU. 10\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\nWe acknowledge and thank Innovate Research, Mr. Shailendra Meena, Dr. Piyush Kumar Pandey, Mr. Shaitan Singh and Ms. Charanpreet Arora who have provided administrative support to the study.\n\n\nReferences\n\nSahoo AK, Mahajan R: Management of tinea corporis, tinea cruris, and tinea pedis: A comprehensive review. Indian Dermatol. Online J. 2016; 7: 77–86. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nElsevier Point of Care: Tinea Infections. Clinical Key. 12 Apr 2021.Reference Source\n\nMorrow JD: Fluconazole: a new triazole antifungal agent. Am. J. Med. Sci. 1991; 302: 129–132. Publisher Full Text\n\nYamada T, Maeda M, Alshahni MM, et al.: Terbinafine Resistance of Trichophyton Clinical Isolates Caused by Specific Point Mutations in the Squalene Epoxidase Gene. Antimicrob. Agents Chemother. 2017; 61: e00115–e00117. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMonod M, Méhul B: Recent Findings in Onychomycosis and Their Application for Appropriate Treatment. J. Fungi. 2019; 5: 20. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPoojary S, Miskeen A, Bagadia J, et al.: A Study of in vitro Antifungal Susceptibility Patterns of Dermatophytic Fungi at a Tertiary Care Center in Western India. Indian J. Dermatol. 2019; 64: 277–284. PubMed Abstract \| Publisher Full TextFree Full Text\n\nRengasamy M, Chellam J, Ganapati S: Systemic therapy of dermatophytosis: Practical and systematic approach. Clin. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMaurya VK, Kachhwaha D, Bora A, et al.: Determination of antifungal minimum inhibitory concentration and its clinical correlation among treatment failure cases of dermatophytosis. J. Fam. Med. Prim. Care. 2019; 8: 2577–2581. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nUhrlaß S, Verma SB, Gräser Y, et al.: Trichophyton indotineae-An Emerging Pathogen Causing Recalcitrant Dermatophytoses in India and Worldwide-A Multidimensional Perspective. J. Fungi. Basel. Switz. 2022; 8: 757. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nEuropean Medicines Agency: INN of the active substance: fluconazole. Assessment report pursuant to Article 30 of Directive 2001/83/EC, as amended.2012.Reference Source\n\nYazdanpanah M, Shamsian A, Shafiee M, et al.: Comparison between Fluconazole and Terbinafine in the treatment of Tinea corporis and Tinea cruris. J. Mycol. Res. 2015; 2: 105–110. Publisher Full Text\n\nStary A, Sarnow E: Fluconazole in the treatment of tinea corporis and tinea cruris. Dermatol Basel Switz. 1998; 196: 237–241. Publisher Full Text\n\nSultana T, Saha SK, Hossain M, et al.: Current Trends of Using Systemic Antifungal Drugs and their Comparative Efficacy in Tinea Corporis and Tinea Cruris in Outpatient Department of Dermatology in a Tertiary Level Hospital. Mymensingh. Med. J. MMJ. 2018; 27: 52–56. PubMed Abstract\n\nRanganathan S, Menon T, Selvi SG, et al.: Effect of socio-economic status on the prevalence of dermatophytosis in Madras. Indian J. Dermatol. Venereol. Leprol. 1995; 61: 16–18. PubMed Abstract\n\nFaergemann J, Laufen H: Levels of fluconazole in serum, stratum corneum, epidermis-dermis (without stratum corneum) and eccrine sweat. Clin. Exp. Dermatol. 1993; 18: 102–106. PubMed Abstract \| Publisher Full Text\n\nSardana K, Arora P, Mahajan K: Intracutaneous pharmacokinetics of oral antifungals and their relevance in recalcitrant cutaneous dermatophytosis: Time to revisit basics. Indian J. Dermatol. Venereol. Leprol. 2017; 83: 730–732. PubMed Abstract Publisher Full Text\n\nFelton T, Troke PF, Hope WW: Tissue penetration of antifungal agents. Clin. Microbiol. Rev. 2014; 27: 68–88. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPathania S, Rudramurthy SM, Narang T, et al.: A prospective study of the epidemiological and clinical patterns of recurrent dermatophytosis at a tertiary care hospital in India. Indian J. Dermatol. Venereol. Leprol. 2018; 84: 678–684. PubMed Abstract Publisher Full Text\n\nWildfeuer A, Faergemann J, Laufen H, et al.: Bioavailability of fluconazole in the skin after oral medication. Mycoses. 1994; 37: 127–130. PubMed Abstract \| Publisher Full Text\n\nSobue S, Sekiguchi K, Nabeshima T: Intracutaneous distributions of fluconazole, itraconazole, and griseofulvin in Guinea pigs and binding to human stratum corneum. Antimicrob. Agents Chemother. 2004; 48: 216–223. Publisher Full Text\n\nKhurana A, Sardana K, Chowdhary A: Antifungal resistance in dermatophytes: Recent trends and therapeutic implications. Fungal. Genet. Biol. FG. B. 2019; 132: 103255. PubMed Abstract \| Publisher Full Text\n\nAkler ME, Vellend H, McNeely DM, et al.: Use of fluconazole in the treatment of candidal endophthalmitis. Clin. Infect. Dis. Off. Publ. Infect. Dis. Soc. Am. 1995; 20: 657–664. Publisher Full Text\n\nAnaissie EJ, Kontoyiannis DP, Huls C, et al.: Safety, plasma concentrations, and efficacy of high-dose fluconazole in invasive mold infections. J. Infect. Dis. 1995; 172: 599–602. PubMed Abstract \| Publisher Full Text\n\nFoster KW, Friedlander SF, Panzer H, et al.: A randomized controlled trial assessing the efficacy of fluconazole in the treatment of pediatric tinea capitis. J. Am. Acad. Dermatol. 2005; 53: 798–809. PubMed Abstract \| Publisher Full Text\n\nGupta AK, Gregurek-Novak T: Efficacy of itraconazole, terbinafine, fluconazole, griseofulvin and ketoconazole in the treatment of Scopulariopsis brevicaulis causing onychomycosis of the toes. Dermatol Basel Switz. 2001; 202: 235–238. PubMed Abstract \| Publisher Full Text\n\nPasko MT, Piscitelli SC, Van Slooten AD: Fluconazole: a new triazole antifungal agent. DICP Ann Pharmacother. 1990; 24: 860–867. Publisher Full Text\n\nLiverTox: Clinical and Research Information on Drug-Induced Liver Injury. Bethesda (MD):National Institute of Diabetes and Digestive and Kidney Diseases; 2012. Fluconazole. LiverTox: Clinical and Research Information on Drug-Induced Liver Injury. Bethesda (MD): National Institute of Diabetes and Digestive and Kidney Diseases; 2017.Reference Source\n\nBrüggemann RJM, Alffenaar J-WC, Blijlevens NMA, et al.: Clinical relevance of the pharmacokinetic interactions of azole antifungal drugs with other coadministered agents. Clin. Infect. Dis. Off. Publ. Infect. Dis. Soc. Am. 2009; 48: 1441–1458. PubMed Abstract \| Publisher Full Text" }	[ { "id": "263427", "date": "30 Apr 2024", "name": "Kabir Sardana", "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\nThere are major methodological constraints and the results that the author propose are not correct as in India fluconazole never leads to 1005 cure rates.\n1 -The use of antifungal drugs is based on AFSST and without MIC data using a drug makes no sense. Multiple studies from India have shown that the MIC to fluconazole is very high. Up to 1:128. Also while it does achieve high levels in the stratum corneum they are not higher than the prevalent MIC needed. This drug almost always fails.\nThere is no pre data or lab data to tell us the prevalent MIC in their study.\n2 - Dose - fluconazole 150 mg daily for 8 weeks. There is no rationale for this dose. Unless the authors can demonstrate that this dose achieves high levels in the s corneum than 50/100 mg.\n3. To expect a “mycological cure of 100%” just on fluconazole is not plausible. As this drug has been repeatedly been shown to be ineffective in dermatophytosis. Unless there was a topical synergistic drug given, which is very likely. In fact most patients use topical luliconazole which is highly potent and could be the cause of the results. There is no data of the same. Also it is inexplicable why the authors could not culture as this was a sponsored study. I would have been happy to see the MIC to other drug specially itraconazole and terbinafine.\n4 - Discussion: There are many inconsistent statements made, especially in regard to itraconazole which is now universally in India the preferred line of treatment. Even the cost has come down with the novel SUBA preparation.\nIn essence the 100% efficacy as a monotherapy seems unlikely. Without a culture MIC and AFFST data the preference for fluconazole seems a bit arbitrary, it would have been much better if, in the absence of a AFSST you could have actually tried itraconazole and fluconazole in two arms as your results are converse to the prevalent practice in India.\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 source data required\n\nAre the conclusions drawn adequately supported by the results? Partly", "responses": [ { "c_id": "12912", "date": "06 Dec 2024", "name": "Suneet Sood", "role": "Author Response", "response": "We would like to thank the reviewer for the meticulous review. Here are our responses to the reviewer’s comments: Comment: In India fluconazole never leads to 100% cure rates. Response: This is true at traditional doses. That, indeed, was the very reason for a study using higher doses. Comment: Multiple studies from India have shown that the MIC to fluconazole is very high. Up to 1:128. Also while it does achieve high levels in the stratum corneum they are not higher than the prevalent MIC needed. Response: Respectfully, we wish to differ. We have shown, in the Discussion, that according to some authors at least the in vitro MIC90 levels of fluconazole are 16–32 micrograms/ml, that the tissue concentrations, and that the drug is eliminated 2–3 times more slowly than from the blood. All of these reports suggest at least the justification for a pilot study. Comment: There is no pre data or lab data to tell us the prevalent MIC in their study. 2 – Dose fluconazole 150 mg daily for 8 weeks. There is no rationale for this dose. Unless the authors can demonstrate that this dose achieves high levels in the s corneum than 50/100 mg. Response: MIC levels have previously been established.[1, 2] We are not making the claim that MIC levels previously established apply in this case. However, the entire purpose of a pilot trial such as this one is to determine if there is reason to conduct in-depth studies like tissue levels and in vitro MIC levels. That said, if we had known in advance that fluconazole would be effective, we would certainly have carried out the microbiological studies to confirm the reason for its efficacy. Now that we are aware that fluconazole is effective in these doses, there is every reason to carry out follow up research. Comment: To expect a “mycological cure of 100%” just on fluconazole is not plausible. As this drug has been repeatedly been shown to be ineffective in dermatophytosis. Unless there was a topical synergistic drug given, which is very likely. In fact most patients use topical luliconazole which is highly potent and could be the cause of the results. There is no data of the same. Also it is inexplicable why the authors could not culture as this was a sponsored study. I would have been happy to see the MIC to other drug specially itraconazole and terbinafine. Response: Admittedly, this drug has not been effective in once-weekly doses. But, as observed by Professor Sardana himself,[3] fluconazole in daily doses is likely to be effective. Indeed, it would not be too inaccurate to say that it was his comment about the possibility of daily fluconazole being the “ideal drug” that was the inspiration for this pilot trial. Comment: There are many inconsistent statements made, especially in regard to itraconazole which is now universally in India the preferred line of treatment. Even the cost has come down with the novel SUBA preparation. In essence the 100% efficacy as a monotherapy seems unlikely. Without a culture MIC and AFFST data the preference for fluconazole seems a bit arbitrary, it would have been much better if, in the absence of a AFSST you could have actually tried itraconazole and fluconazole in two arms as your results are converse to the prevalent practice in India. Response: We do not deny that itraconazole is available, and has advantages, and that we could have compared the two drugs in two treatment arms. However, this really was not our study. Our study was exploratory, a pilot study as pointed out in the title itself, intended to determine if fluconazole was effective in daily disease. Our study does not arrive at the conclusion that fluconazole is better or worse than other drugs. References Felton T, Troke PF, Hope WW: Tissue penetration of antifungal agents. Clin. Microbiol. Rev. 2014; 27: 68–88. Pathania S, Rudramurthy SM, Narang T, et al.: A prospective study of the epidemiological and clinical patterns of recurrent dermatophytosis at a tertiary care hospital in India. Indian J. Dermatol. Venereol. Leprol. 2018; 84: 678–684. Sardana K, Arora P, Mahajan K: Intracutaneous pharmacokinetics of oral antifungals and their relevance in recalcitrant cutaneous dermatophytosis: Time to revisit basics. Indian J. Dermatol. Venereol. Leprol. 2017; 83: 730–732." } ] }, { "id": "263418", "date": "02 May 2024", "name": "Clara Gómez", "expertise": [ "Reviewer Expertise Fungal infection", "Photodynamic therapy" ], "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 “An open label, single arm, pilot study to evaluate the safety, tolerability, and efficacy of daily fluconazole 150 mg in subjects suffering from Tinea cruris and Tinea corporis” by Dattatray Gopal Saple, Sushrut Save, Devesh Kumar and Suneet Sood, is a clinical trial which aims to evaluate the effects of a dermatologic treatment based on fluconazole 150 mg daily for 8 weeks in patients with uncomplicated dermatophytosis. Treatment was very effective and also safe and well tolerated. The text is clear, the figures and tables are consistent. The conclusions are adequately supported by the results and are important as they indicate that a very high and prolonged dose of fluoconazole is effective and safe.\nThere are MINOR concerns and suggestions that should be addressed by the authors.\n* If Kurana et al (ref. 31) pointed out that elimination half-life of fluconazole is between 60-90 hours in the stratum corneum, applying fluconazole every 3 days for example for 3 months would also be effective and much safer in terms of side effects and interactions. Why didn't the authors consider also carrying out an intermediate treatment between the already established one (150 mg weekly for 6 months) and the one shown here, to determine if it is also effective? * Another aspect to consider is the importance of determining a priori the type of dermatophyte causative of the disease, since difficulty in eradicating the disease depends on the genus and specie of the causative dermatophyte. Establishing a pattern of dosage as a function to the causative agent could allow the reduction of the dose proposed by the authors in certain patients and thus increase the safety of the treatment.\nAbstract Methods: remove the sentence “Of 107 subjects…withdrew consent” Results: change “At 5 weeks” by “At fifth visit (56 days)” Conclusions: remove the sentence “Fluconazole is… preferred therapy” because it is not a conclusion of the study. Keywords: introduce fluconazole, tinea cruris, tinea corporis. Remove tinea capitis\nDiscussion Remove epigraphs Fourth paragraph: lasting eight weeks\nFigure 1 can be deleted since it is a clinical study with patients assigned to a single group (there is no randomization).\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": [ { "c_id": "12913", "date": "12 Dec 2024", "name": "Suneet Sood", "role": "Author Response", "response": "We wish to thank the reviewer for her comments. To respond: Comment: Why didn't the authors consider also carrying out an intermediate treatment between the already established one (150 mg weekly for 6 months) and the one shown here, to determine if it is also effective? Another aspect to consider is the importance of determining a priori the type of dermatophyte causative of the disease, since difficulty in eradicating the disease depends on the genus and specie of the causative dermatophyte. Response: We agree to both comments. However, this was an exploratory, pilot study to determine if there were enough grounds to carry out in-depth studies of fluconazole in dermatophytosis. In view of the increasing reports of resistance across the world, it was not considered worthwhile in the beginning to consider the intermediate treatment. Now that we have evidence of efficacy and safety, our next step will be to study the organisms and the appropriate dosages. Comment: Abstract. Methods: remove the sentence “Of 107 subjects…withdrew consent”. Results: change “At 5 weeks” by “At fifth visit (56 days)”. Conclusions: remove the sentence “Fluconazole is… preferred therapy” because it is not a conclusion of the study. Keywords: introduce fluconazole, tinea cruris, tinea corporis. Remove tinea capitis Response: We agree, and we appreciate the observations. The text has been changed accordingly. Comment: Discussion: Remove epigraphs; Fourth paragraph: lasting eight weeks. Figure 1 can be deleted since it is a clinical study with patients assigned to a single group (there is no randomization). Response: We have made most of the changes as suggested. It may be clearer to maintain Figure 1, however." } ] } ]	1	https://f1000research.com/articles/11-1446
https://f1000research.com/articles/13-1498/v1	06 Dec 24	{ "type": "Research Article", "title": "Role modeling and Snack Choices of young adults: The mediating role of meal planning", "authors": [ "Musa Kiggwe", "Joseph Ntayi", "Annet K. Nabatanzi-Muyimba", "Jotham Byarugaba", "Esemu Timothy", "John C. Munene", "Joseph Ntayi", "Annet K. Nabatanzi-Muyimba", "Jotham Byarugaba", "Esemu Timothy", "John C. Munene" ], "abstract": "Objective This study aimed to examine the mediating role of meal planning in the relationship between role modeling and snack choice.\n\nMethods and Measures The study was correlational and quantitative in nature. The study selected a sample of 403 university students. We collected data through interviews using a questionnaire. We analyzed the data using SPSS 23.0.0.0 (IBM Corp; 1989, 2015) (Build 1607) and AMOS 23.0.0 (IBM Corp; 1983, 2015) (Build 1607) software for correlational and Confirmatory factor tests. We used the bootstrapping technique to test for the mediation effect of meal planning.\n\nResults Meal planning had a full mediation effect between role modeling and snack choice. This means that Role modeling had a significant influence on meal planning, meal planning had a significant influence on snack choice, and role modeling had no significant influence on snack choice.\n\nConclusion when young people admire other people’s eating habits it leads them to plan their meals and choose similar snacks to their role models. Future studies should consider studying the effect that role models’ proximity has on snack choices among young people in other contexts.", "keywords": [ "Role-Modeling", "Meal-Planning", "Snack", "Mediation", "Uganda" ], "content": "Introduction\n\nSeveral Non-Communicable Diseases are associated with diet behaviors (Almoraie et al., 2021) that include snacking. Snacking is largely considered unhealthy (Shatwan et al., 2022) but also partly healthy (Damen et al., 2020; Schlinkert et al., 2020). It is highly prevalent among young adults around the world. For instance, college students in Asia and the Middle Eastern countries are fond of snacking (Shatwan et al., 2022). As young adults, behaviors learned at this stage tend to linger on throughout their adulthood (Gonzales, 2013).\n\nScholars have tried to understand individual behavioral choices through different theoretical explanations. The Social Cognitive Theory (SCT), for instance, asserts that people acquire new behaviors through observing and imitating others, within their social context (Bandura, 1971). When an observed action is considered rewarding, the observer will replicate the action later. However, individuals may mentally learn new habits but not immediately display the learned habits. Individuals may have intentions to display the learned behavior at a later time. Schwarzer’s (2008) Health Action Process approach posits that planning is a crucial mediator between individual intentions and actual behavior. Planning details the boundaries of when, where, and how the behavior will be done (Scholz et al., 2008). Theory, therefore, suggests that learning may lead to preparation to do the learned behavior (Bandura, 1971), which preparation then translates to replicating the learned behavior (Parkinson et al., 2023).\n\nIndividual food choices may be an imitation of other people’s eating habits. The more food one’s companion eats, the more of that food an individual eats, and vice versa (Suwalska & Bogdański, 2021). Children will eat a wide food variety upon seeing their parents eat a wide food variety as well (Scaglioni et al., 2018). Even in the absence of parents, children consume what their parents approve of (Gilmour et al., 2020). Sogari et al. (2018) study reported that college students eat more fruit and vegetables when their parents encourage them and avoid unhealthy foods when their parents instruct them. Even when a role model is not physically present, a person may still adhere to the learned eating behavior (Prinsen et al., 2013). Yiga et al. (2021) study revealed that whether young women lived with or independent of their parents, the food choices they made were learned from their parents. Smith et al. (2019) reported food-related TV advertisements as responsible for the increase in the consumption of high-dense energy foods. Kergoat et al. (2020) findings are also in agreement that the more commercial messages a person receives the more sweets they consume. Social media image-related content is reported to be related to young adults’ restricting their food intake (Rounsefell et al., 2020) and choice of healthy foods (del Rio Carral et al., 2024). The more people watch celebrity cook shows the more they consume meat diets (Roy et al., 2021).\n\nRole models may influence people’s snacking behavior only in the absence of eating schedules (Hess et al., 2016). Human beings use other people’s eating behavior as a blueprint of the food type and quantity to eat (Cruwys et al., 2014). Young adults schedule their meals based on what their mothers used to prepare during their stay with them (Yiga et al., 2021). Okpara et al. (2022) stated that children planned for their meals in adulthood if they witnessed their parents plan for food during the children’s early years. Additionally, whenever individuals watch television cooking shows, they start projecting how their meals should look like (Boulos et al., 2012).\n\nDifferent plans influence people’s eating behaviors (Lange et al., 2018) amongst which are meal plans (Yiga et al., 2021). Meal planning aids individuals make healthy food choices (Fernandez et al., 2020) that include fruit and vegetables (Domke et al., 2021). According to Lange et al. (2018), the more men and women plan their meals, the more fruits and vegetables they consume. Sogari et al. (2018), also, add that the more students planned their meals, the more healthy foods they ate. Hanson et al. (2019) for instance echo the same in their study of college students that the more they planned their meals the more fruits and vegetables they consumed.\n\nThe study therefore hypothesizes that:\n\nThere is a positive relationship between role modeling and snack choice.\n\nThere is a positive relationship between role modeling and meal planning.\n\nThere is a positive relationship between meal planning and snack choice.\n\nMeal planning mediates the relationship between role modeling and snack choice.\n\nExisting research appears not to have extensively examined the connection between role modeling, meal planning, and snack choices (Hamilton et al., 2017). Previous studies have primarily focused on the influence of role modeling on snack choices, with limited attention given to the impact of TV cooking shows (Ngqangashe et al., 2018). Additionally, the majority of research in this area has concentrated on children and adolescents, with minimal investigation within African sub-populations. There is also a lack of studies examining the relationships between role modeling and meal planning (Yee et al., 2017), as well as meal planning and snack choices (Ducrot et al., 2017). To address these gaps, this study aims to investigate the mediating role of meal planning in the link between role modeling and snack choices among young adults in Uganda.\n\n\nMethods\n\nThe researchers conducted a correlational survey approach to test the study hypotheses. The study population consisted of 59,998 Ugandan university students (School Guide Uganda Limited, 2020). Uganda is a country in East Africa, North of Tanzania, and west of Kenya. Uganda has one of the youngest populations in the world.\n\nBased on Yamane formulae (Yamane, 1967), the researchers selected a sample of 403 students using a proportionate stratified random sampling method. We based the strata on the respective university student population. Before data collection, we obtained an ethical clearance from the Faculty of Graduate Studies and Research of Makerere University Business School dated 10th July 2023. The researchers fully informed all participants of the project and obtained, from them, written consent for the use and publication of collected data. The researchers assured the participants that they would handle their responses with absolute confidentiality and that the survey was completely anonymous, with no mention of participant names and addresses. We issued 550 questionnaires and received 432 questionnaires and then discarded 29 unusable ones since according to Hair et al. (2014) they had at least 30% unfilled. We thus got a response rate of 73.3%.\n\nWe collected data using adapted questionnaire items, which we tested for both reliability and construct validity before starting the study. Specifically, subject experts evaluated questionnaire interpretation, length, easiness, and clarity. Next, to assess the reliability of the questionnaire, we conducted a pilot study using 40 questionnaires at the Islamic University in Uganda (located approximately 2 kilometers – 1 mile – north of Mbale’s central business district in the Eastern Region of Uganda, on the Mbale-Soroti road). We then calculated the Cronbach’s alpha coefficient. All instrument scales produced coefficients above 0.7.\n\nThe final instrument contained demographic items: age, gender, education, residential status, and partner lived with. It also included the constructs: role modeling, meal planning, and snack choice.\n\nRole modeling\n\nRole modeling refers to the social construction of (or learning from) another person’s experiences (and incorporating lessons) into a youth’s personal life. The researchers measured role modeling in terms of live modeling, screen shows, friend encouragement, and parental influence. Respondents were asked to indicate, on a six-point scale (Chomeya, 2010), the extent to which they agreed or disagreed with the different statements. The researchers operationalized live modeling as ‘my parents eat the following foods when I am with them (Yee et al., 2017): It was measured using items: fruits, vegetables, and staple foods (Cullen et al., 2001). Screen shows were operationalized as: watching television commercials, cooking shows, or a celebrity inspire me regarding eating particular food kind (D’Alessio et al., 2009). Friend encouragement was operationalized as: My friends encourage me to eat: fruits, and vegetables (Cullen et al., 2001). Parental Influence was operationalized as: ‘My parents urge me to eat: fruits, vegetables, and take drinks (Cullen et al., 2001).\n\nMeal planning\n\nMeal planning refers to the act of scheduling; what to eat, where to eat, and with whom to eat (Bruijn et al., 2017; Hamilton et al., 2017; Luszczynska et al., 2007). The researchers measured meal planning in terms of meal partner, dining place, and meal type. Respondents were asked to indicate, on a six-point scale (Chomeya, 2010), the extent to which they agreed or disagreed with the different statements (Sniehotta et al., 2005). The researchers operationalized meal partner as ‘I make detailed plans regarding; Eating with my family, and eating with my parents. The dining place was operationalized as ‘I make detailed plans regarding: Where I will eat, and eating at school. Meal type was operationalized as ‘I make detailed plans regarding; Eating fruits, eating vegetables, and taking drinks.\n\nSnack choice\n\nSnack choice means the frequency with which a youth eats snack foods like fruits and dairy products. Snack choice was measured in terms of fruit snack or dairy snack (Shatwan et al., 2022). The researchers adapted the Food Frequency Questionnaire (Subar et al., 1995) that included fruit snacks: pineapples, apples, and watermelon (Wallace et al., 2020). Dairy snacks included Chocolate, and Ice cream (Shatwan et al., 2022). The researchers required respondents to indicate their consumption frequency within the preceding year using scale anchors that ranged from ‘Never or less than once a month’ to ‘more than 6 times a day’.\n\nWe used SPSS 23.0.0.0 (IBM Corp; 1989, 2015) (Build 1607) to produce descriptive statistics and AMOS 23.0.0 (IBM Corp; 1983, 2015) (Build 1607) for Confirmatory Factor Analysis (CFA). Under CFA we analyzed a structural equation model to determine whether role modeling predicted the food choices of young adults in Uganda. In addition, we used the bootstrapping technique to test for the mediation effect of meal planning between the two variables.\n\n\nResults\n\nResults show that the study sample had 197 males (48.9%) and 206 females (51.1%). The respondents’ mean age was 21 years. 89.8% of the respondents were pursuing a bachelor’s degree. Many respondents resided in off-campus residences (61.5%), and most of them (58.8%) resided alone in their residences.\n\nWe carried out CFA to examine whether the item measures were consistent with the construct nature (Hair et al., 2014). Table 1 (Extended data) (Kiggwe et al., 2024) shows several commonly used fit indices we employed to assess the overall model fit (Hu & Bentler, 1999). The comprehensive goodness-of-fit indices produced a Chi-square of 458.888 and Chi-Square/DF=1.779.\n\nWe considered the comparative fit index (CFI) value of 0.947, incremental fit index (IFI) value of 0.947, and Tucker-Lewis index (TLI) value of 0.938, as very good fits to the model based on Hu and Bentler (1999) assertion that for these indices a value of 0.7 and above is satisfactory, 0.8 and above is good, and 0.9 and above is very good. The root mean square error of approximation (RMSEA) value was 0.044 indicating a close fit. Thus, the results in the measurement model (see Figure 1) indicate an acceptable fit.\n\nAll standardized factor loadings should be at least 0.5 and statistically significant to indicate that observed indicators are strongly related to their associated constructs (Hair et al., 2014). In the model, all standardized factor loadings were above 0.5 and significant (see Figure 1). These findings indicate that there was a satisfactory fit between the proposed model and the data. The test for convergent validity revealed that the composite reliability coefficients for all constructs were above 0.9. In addition, the Average Variance Extracted (AVE) for all constructs met the threshold of 0.5 (Hair et al., 2014). There was discriminant validity in our instrument based on Fornell and Larcker (1981) criterion that when the correlation values among the latent variables are less than the square root of AVE, then there is discriminant validity (see Table 2). Finally, the values of Skewness and Kurtosis did not indicate any serious violations of normality, as all the coefficients were below ±2 (see Table 3).\n\n Correlation is significant at the 0.01 level (1-tailed).\n\nTo explore the association between role modeling, meal planning, and snack choice behavior, we estimated the Pearson correlation coefficient, which indicated a significant correlation between the variables (r = -0.02, p > 0.05) (see Table 2).\n\nWe carried out Structural Equation Modeling (SEM) to evaluate whether role modeling predicted food choices. We used SEM as it provides for the control of the measurement error.\n\nTable 4 indicates that the relationship between role modeling and snack food choice is not significant (β = - 0.138, t-value = - 0.973, p-value = 0.331); the relationship between role modeling and meal planning is significant (β = - 0.785, t-value = 5.455, p-value = 0.000) and the relationship between meal planning and snack food choice is significant (β = 0.401, t-value = 3.696, p-value = 0.00). The squared Multiple Correlation is 0.104 (See Figure 1). This means role modeling and meal planning, approximately, explain 89.6 percent of the variance in Snack food Choice. In other words, the error variance of role modeling and meal planning is approximately 89.6 percent of the variance of snack food choices.\n\nWe employed a bootstrap procedure to test for mediation effects and determine the level of significance of the mediation effects. The bootstrap approach simultaneously tests for both the direct and indirect relationships in a single model. Table 5 depicts the mediation results.\n\n Correlation is significant at the 0.01 level (2-tailed).\n\nThe bootstrap result revealed a significant full mediation (β = 0.315, p = 0.000). This means that changes in role modeling positively affect variations in Meal Planning, which in turn cause changes in Snack Choice. This implies that when parents, friends, television cooking shows and celebrities influence the youth, they go ahead and plan for where they shall have their meals, what meals they shall eat, and with whom they shall eat. This meal scheduling consequently leads to the youth consuming more fruits and more snacks.\n\n\nDiscussion\n\nThis study examined the mediating role of meal planning in the relationship between role modeling and snack choice. The study’s results revealed a full mediation. This suggests that modeling does not have a direct impact on food consumption behavior. Instead, meal planning serves as a pathway through which modeling influences food consumption behavior. This implies that when parents encourage the youth to eat certain snacks it does not directly translate into the youth consuming those foods. This was a surprising finding. The plausible explanation is that at this life stage, the youth have started making autonomous consumption decisions. Parents no longer have as much control as when they were younger. So, even when the parents urge the youth to eat healthy the youth may not follow the advice. Further to this is that these youths largely reside alone in off-campus residences. Parents are rarely eating companions of the youth during their stay at university. This finding contradicts Roy et al. (2021) assertion that advice from friends and family significantly influences healthy eating choices. The finding also means that even when television celebrities endorse healthy foods, it does not directly influence the youth’s eating habits. Perhaps the youth consider this food quite expensive and therefore do not purchase it. This finding contradicts Folkvord et al. (2020) assertion that watching cooking programs that promoted healthy foods was highly likely to lead to healthy food choices.\n\nThe findings, additionally, indicate that when the youth watch television celebrities, that endorse healthy foods, the youth make detailed eating prearrangements. This may be due to the youth’s admiration of the celebrity’s accomplishments like body appearance and fame. The youth will hence desire to uphold the celebrity’s eating values and beliefs. These meal plans may then aid the youth achieve their goals by developing an eating routine given their busy study schedules. This finding agrees with Ngqangashe and De Backer’s (2021) conclusion that showing videos depicting fruits and vegetables to adolescents resulted in them preparing to eat the demonstrated foods.\n\nThe study findings also indicated that youths who prepare what to eat in advance eat the foods that they prearranged. In other words, whenever a youth makes detailed plans regarding, say, eating fruits then s/he consumes fruits at the slated time. This could result from time restrictions that may hinder youth from eating during study periods. Students may, also, find eating fruits a convenient option since they lack enough time to cook. The findings conform to Domke et al. (2021) whose report indicated that German adults who planned their meals on precisely when, where, what, and how they ate consumed more fruits and vegetables. Therefore, when young people admire their role models’ eating habits it leads them to plan their meals and choose similar snacks.\n\nThe present results have important theoretical and policy implications. The study indicates that role modeling is not an immediate antecedent of food consumption behaviors of the youth in Uganda. From a Social Cognitive Theory (Bandura, 1977) perspective, it may be due to the lack of physical proximity to the role model. The youth largely lived alone mainly away from their parents and hence little physical interaction at meal times. This meant that there was little cognitive rehearsal or reminders from parents regarding the eating of healthy foods. The youths were largely unmarried which meant that no one was around during meal times to remind youths to eat these healthy foods. The findings, also, lend support to the Health Action Process Approach (Schwarzer, 2008), which asserts that planning serves as an operative mediator between intentions and behaviour. This view is based on the fact that when an individual admires a person’s diet, he makes an intention to consume the same food as the role model at a later time, which leads to scheduling that food ultimately leading to the consumption of that food. At a policy level, governments should carry out mass awareness campaigns to encourage the youth to schedule what and when they are to eat their meals. The ministry could perhaps implement this campaign using youth opinion leaders that include movie/television celebrities. This prearrangement of meals would enable the youth to manage their busy work schedules and create time for the consumption of healthy meals.\n\nThe present study is not without limitations but in turn, offers the opportunity for future studies. The results of this study are an outcome of a case study of young adults in Uganda. There is a need for further extension of these studies to other countries and populations before researchers generalize the findings. The results of this study opened a way for assessing the mediating role of meal planning in the relationship between role modeling and snack consumption among young adults in Uganda. Therefore, future studies should consider studying the effect that physical or virtual proximity of the role model has on snack choices among young people in other contexts.\n\nThe study was cross-sectional in nature. Capturing the long-run effect of role modeling on food consumption was not possible. Given that learning from a role model may happen at an earlier time and display of the learned behavior at a far later time, it calls for a longitudinal study to ascertain this relationship in future studies.\n\nBefore data collection, we obtained an ethical clearance from the Faculty of Graduate Studies and Research of Makerere University Business School dated 10th July 2023. The researchers fully informed all participants of the project and obtained, from them, written consent for the use and publication of collected data.", "appendix": "Acknowledgment\n\nWe would like to thank the community of practice from which the lead author learned a lot.\n\n\nData availability\n\nRole modeling and Snack Choices of Young Adults: The Mediating role of Meal planning. https://doi.org/10.6084/m9.figshare.27314754.v1 (Kiggwe, 2024).\n\nThis project contains the following underlying data:\n\n• MODELING AND SNACK CONSUMPTION.sav\n\nData are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).\n\nFigshare: Mediation Test of Role Modeling Through Meal Planning to Snack Choice, https://doi.org/10.6084/m9.figshare.27640581.v1 (Kiggwe et al., 2024a).\n\nThis project contains the following extended data:\n\n• Figure 1. Mediation Test of Role Modeling Through Meal Planning to Snack Choice\n\n• Table 1. Confirmatory Factor Analysis Results for Items Measuring Role-modeling, Meal-planning, and Snack-choice (n=403)\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: Study Questionnaire, https://doi.org/10.6084/m9.figshare.27677082.v1 (Kiggwe et al., 2024b).\n\nThis project contains the following extended data:\n\n• Questionnaire. Study questionnaire as applied during the survey.\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\nAlmoraie NM, Saqaan R, Alharthi R, et al.: Snacking patterns throughout the life span: potential implications on health. Nutr. Res. 2021; 91(June): 81–94. PubMed Abstract \| Publisher Full Text\n\nBandura A: Social Learning Theory. Bandura A, editor. General Learning Corporation; 1971.\n\nBandura A: Social Learning Theory. Bandura A, editor. 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Publisher Full Text\n\nGonzales K: The Impact of Residence on Dietary Intake, Food Insecurity, and Eating Behavior among University Undergraduate Students. The Undergraduate Research Journal at the University of Northern Colorado. 2013; 3(2): 113–130.\n\nHair JJF, Black WC, Babin BJ, et al.: Multivariate Data Analysis. 7th ed.Pearson Education Limited; 2014.\n\nHamilton K, Kothe EJ, Mullan B, et al.: The mediating and moderating role of planning on mothers’ decisions for early childhood dietary behaviours. Psychol. Health. 2017; 32: 1–16. PubMed Abstract \| Publisher Full Text\n\nHanson AJ, Kattelmann KK, Mccormack LA, et al.: Cooking and Meal Planning as Predictors of Fruit and Vegetable Intake and BMI in First-Year College Students. Int. J. Environ. Res. Public Health. 2019; 16(2462): 1–11.\n\nHess JM, Jonnalagadda SS, Slavin JL: What is a snack, why do we snack, and how can we choose better snacks? A review of the definitions of snacking, motivations to snack, contributions to dietary intake, and recommendations for improvement. Adv. Nutr. 2016; 7(3): 466–475. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHu L-T, Bentler PM: Cutoff criteria for fit indexes in covariance structure analysis: Conventional criteria versus new alternatives. Struct. Equ. Model. Multidiscip. J. 1999; 6(1): 1–55. Publisher Full Text\n\nKergoat M, Meyer T, Legal JB: Influence of “health” versus “commercial” physical activity message on snacking behavior. J. Consum. Mark. 2020; 37(2): 170–179. Publisher Full Text\n\nKiggwe M: MODELING AND SNACK CONSUMPTION.sav. Dataset. figshare. 2024. Publisher Full Text\n\nKiggwe M, Ntayi J, Nabatanzi-Muyimba AK, et al.: Mediation Test of Role Modeling Through Meal Planning to Snack Choice. figshare. Figure. 2024a. Publisher Full Text\n\nKiggwe M, Ntayi J, Nabatanzi-Muyimba AK, et al.: Study Questionnaire. figshare. Journal contribution. 2024b. Publisher Full Text\n\nLange D, Corbett J, Knoll N, et al.: The interplay of intention, autonomy, and sex with dietary planning: A conditional process model to predict fruit and vegetable intake. Int. J. Behav. Med. 2018; 25: 421–430. Publisher Full Text\n\nLuszczynska A, Scholz U, Sutton S: Planning to change diet: A controlled trial of an implementation intentions training intervention to reduce saturated fat intake among patients after myocardial infarction. J. Psychosom. Res. 2007; 63: 491–497. PubMed Abstract \| Publisher Full Text\n\nNgqangashe Y, De Backer CJS , Hudders L, et al.: An experimental investigation of the effect of TV cooking show consumption on children’s food choice behaviour. Int. J. Consum. Stud. 2018; 42: 402–408. Publisher Full Text\n\nNgqangashe Y, De Backer CJS: The differential effects of viewing short-form online culinary videos of fruits and vegetables versus sweet snacks on adolescents’ appetites. Appetite. 2021; 166(105436): 105436–105439. PubMed Abstract \| Publisher Full Text\n\nOkpara N, Chauvenet C, Grich K, et al.: “Food Doesn’t Have Power Over Me Anymore!” Self-Efficacy as a Driver for Dietary Adherence Among African American Adults Participating in Plant-Based and Meat-Reduced Dietary Interventions: A Qualitative Study. J. Acad. Nutr. Diet. 2022; 122(4): 811–824. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nParkinson J, Hannan T, McDonald N, et al.: Health action process approach: Promoting physical activity, and fruit and vegetable intake among Australian adults. Health Promot. Int. 2023; 38(4): 1–12. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPrinsen S, De Ridder DTD, De Vet E: Eating by example. Effects of environmental cues on dietary decisions. Appetite. 2013; 70: 1–5. PubMed Abstract \| Publisher Full Text\n\nRounsefell K, Gibson S, McLean S, et al.: Social media, body image and food choices in healthy young adults: A mixed methods systematic review. Nutr. Diet. 2020; 77: 19–40. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nRoy R, de Castro TG , Haszard J, et al.: Who we seek and what we eat? Sources of food choice inspirations and their associations with adult dietary patterns before and during the covid-19 lockdown in New Zealand. Nutrients. 2021; 13(11). PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nScaglioni S, De Cosmi V , Ciappolino V, et al.: Factors Influencing Children’s Eating Behaviours. Nutrients. 2018; 10(706): 1–17. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSchlinkert C, Gillebaart M, Benjamins J, et al.: The snack that has it all: People’s associations with ideal snacks. Appetite. 2020; 152(April): 104722. PubMed Abstract \| Publisher Full Text\n\nScholz U, Nagy G, Schuz B: The role of motivational and volitional factors for self-regulated running training: Associations on the between- and within- person level. Br. J. Soc. Psychol. 2008; 47: 421–439. PubMed Abstract \| Publisher Full Text\n\nSchool Guide Uganda Limited: Uganda Schools Guide - The only online guide to all Ugandan Schools.2020. Reference Source\n\nSchwarzer R: Modeling health behavior change: How to predict and modify the adoption and maintenance of health behaviors. Appl. Psychol. Int. Rev. 2008; 57(1): 1–29. Publisher Full Text\n\nShatwan IM, Aljefree NM, Almoraie NM: Snacking pattern of college students in Saudi Arabia: a cross-sectional study. BMC Nutrition. 2022; 8(49): 10–49. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSmith R, Kelly B, Yeatman H, et al.: Food marketing influences children’s attitudes, preferences and consumption: A systematic critical review. Nutrients. 2019; 11(4): 1–14. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSniehotta FF, Schwarzer R, Scholz U, et al.: Action planning and coping planning for long-term lifestyle change: Theory and assessment. Eur. J. Soc. Psychol. 2005; 35(4): 565–576. Publisher Full Text\n\nSogari G, Velez-Argumedo C, Gómez MI, et al.: College students and eating habits: A study using an ecological model for healthy behavior. Nutrients. 2018; 10(12): 1–16. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSubar AF, Thompson FE, Smith AF, et al.: Improving food frequency questionnaires - A qualitative approach using cognitive interviewing. J. Am. Diet. Assoc. 1995; 95(7): 781–788. Publisher Full Text\n\nSuwalska J, Bogdański P: Social modeling and eating behavior—a narrative review. Nutrients. 2021; 13(1209): 1–16. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nWallace TC, Bailey RL, Blumberg JB, et al.: Fruits, vegetables, and health: A comprehensive narrative, umbrella review of the science and recommendations for enhanced public policy to improve intake. Crit. Rev. Food Sci. Nutr. 2020; 60(13): 2174–2211. PubMed Abstract \| Publisher Full Text\n\nYamane T: STATISTICS; An Introductory Analysis. 2nd ed.Harper & Row; 1967.\n\nYee AZH, Lwin MO, Ho SS: The influence of parental practices on child promotive and preventive food consumption behaviors: a systematic review and meta-analysis. Int. J. Behav. Nutr. Phys. Act. 2017; 14(47): 14–47. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nYiga P, Ogwok P, Achieng J, et al.: Determinants of dietary and physical activity behaviours among women of reproductive age in urban Uganda, a qualitative study. Public Health Nutr. 2021; 24(12): 3624–3636. PubMed Abstract \| Publisher Full Text \| Free Full Text" }	[ { "id": "348322", "date": "30 Dec 2024", "name": "Md. Hafizur Rahman", "expertise": [ "Reviewer Expertise Emotional Intelligence", "Sustainability", "Entrepreneurship", "Circular Economy", "Green Marketing and 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\nSummary of the Article :\nThe study investigates the mediating role of meal planning in the relationship between role modeling and snack choices among Ugandan university students. Findings suggest that meal planning fully mediates this relationship, with role modeling indirectly influencing snack choices via planning. The study employs SEM and bootstrapping for data analysis and concludes with policy implications for promoting healthy eating habits through meal planning.\nCritique and Recommendations 1. Clarity and Presentation: i) Expand on the theoretical framework to strengthen the conceptual foundation. ii) Include the research gap and objectives before starting the literature review for the development of hypotheses. iii) Add a hypothesized framework immediately after writing the hypotheses. iv) Ensure consistency in referencing, including more diverse and recent studies.\n2. Methodological Details i) Provide examples of how questionnaire items were culturally adapted. ii) Include the reasons for using stratified sampling and how you handled potential biases in self-reported data.\n3. Analysis and Interpretation: i) The interpretation of Table 4 indicates that the relationship between role modeling and meal planning is significant (but showing β = - 0.785, t-value = 5.455, p-value = 0.000) and needs to be corrected according to the actual results. ii) Discuss why the direct effect of role modeling on snack choice was insignificant.\n4. Discussion and Conclusions i) Divide the discussion into the following sections: Discussion, Theoretical Implications, Policy Recommendations, Conclusions ii) Delineate between findings specific to Uganda and broader implications\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? Partly", "responses": [] } ]	1	https://f1000research.com/articles/13-1498
https://f1000research.com/articles/13-1496/v1	06 Dec 24	{ "type": "Systematic Review", "title": "Safety and efficacy of Dexmedetomidine in preoperative and postoperative settings during transsphenoidal resection of pituitary Adenoma: A systematic review and meta-analysis of randomized controlled trials", "authors": [ "Abdulsalam M Aleid", "Ali A Alghamdi", "Mohammed Yousef Alessa", "Loai Saleh Albinsaad", "Abdulrahman Almoslem", "Ibrahim Alahmed", "Abdulwahab Ahmed Alzahrani", "Abdulmonem Ali Alhussain", "Awn Abdulmohsen Alessa", "Saud Nayef Salem Aldanyowi", "Ali A Alghamdi", "Mohammed Yousef Alessa", "Loai Saleh Albinsaad", "Abdulrahman Almoslem", "Ibrahim Alahmed", "Abdulwahab Ahmed Alzahrani", "Abdulmonem Ali Alhussain", "Awn Abdulmohsen Alessa", "Saud Nayef Salem Aldanyowi" ], "abstract": "Background Pituitary adenoma, a benign tumor of the pituitary gland, represents 10-15% of intracranial tumors. Although non-cancerous, its size and location can cause significant health issues, including hormonal imbalances and compression of nearby structures like the optic chiasm, leading to conditions such as Cushing’s disease, acromegaly, and visual disturbances. The prevalence ranges from 46 to 95 per 100,000 individuals. This study evaluates the effectiveness and safety of dexmedetomidine versus placebo in patients undergoing transsphenoidal resection of pituitary adenomas.\n\nMethods A systematic review and meta-analysis following PRISMA guidelines included 10 RCTs with 633 patients. English-language RCTs were sourced from PubMed, Scopus, Web of Science, Google Scholar, and Cochrane Library. Risk of bias was assessed using the ROB2 tool, and statistical analyses were performed using RevMan 5.3.\n\nResults Dexmedetomidine significantly reduced heart rate after intubation (MD: -6.61 bpm, 95% CI: -8.98 to -4.24, p < 0.00001) and post-extubation (MD: -6.77 bpm, 95% CI: -8.59 to -4.96, p < 0.00001). Mean arterial pressure decreased after intubation (MD: -10.49 mmHg, 95% CI: -12.73 to -8.26, p < 0.00001) and post-extubation (MD: -12.97 mmHg, 95% CI: -14.37 to -11.57, p < 0.00001). Dexmedetomidine also reduced surgical duration (SMD: -0.23, 95% CI: -0.41 to -0.05, p = 0.01), blood loss (MD: -109.49 mL, 95% CI: -152.60 to -66.38, p < 0.00001), propofol dose (SMD: -1.04, 95% CI: -1.72 to -0.37, p = 0.002), extubation time (SMD: -0.70, 95% CI: -0.93 to -0.47, p < 0.00001), and postoperative nausea and vomiting (OR: 0.45, 95% CI: 0.25 to 0.80, p = 0.007).\n\nConclusions Dexmedetomidine enhances perioperative outcomes in transsphenoidal resection of pituitary adenomas, supporting its use as an effective anesthetic adjunct.", "keywords": [ "Pituitary adenoma", "dexmedetomidine", "transsphenoidal resection", "perioperative management", "anesthesia", "hormonal imbalance", "postoperative outcomes", "systematic review." ], "content": "1. Introduction\n\nPituitary adenoma is a benign tumor that arise from pituitary gland, small endocrine gland at the base of the brain located at Sella turcica that called the maestro of the body, pituitary adenoma accounts from 10:15 % of all intracranial tumors so, its one of the most common tumors of the brain.1,2 Although pituitary adenoma is a non-cancerous tumor, it leads to significant problems due to its size and location.3 It causes hormonal imbalance which leads to some serious diseases like Cushing’s disease, acromegaly, or hypopituitarism.4 Also, if the tumor is large, it can cause compression on surrounding structures such as optic chiasm that leads to visual disturbance.5 The prevalence of pituitary adenoma is high and estimated to be 46:95 pre 100,000 individuals.6\n\nManagement of pituitary adenoma requires multidisciplinary approach including endocrinology, neurosurgery and radiology.7 Surgical resection is the primary treatment modality especially in large adenomas.7 The transsphenoidal surgical technique is the best approach for resecting pituitary adenomas because of its minimally invasive nature that involves accessing the tumor through the nasal cavity and sphenoid sinus.8 This approach avoids craniotomy and then provides less recovery time and less complications.9 But this approach faces many challenges like proximity of the tumor to critical structures such as the carotid arteries, optic nerves, and cavernous sinus so, precise and experienced surgeon is required to avoid damage to these structures.10 This approach has also some complications like intraoperative bleeding, cerebrospinal fluid leaks, and incomplete tumor resection.10\n\nAnesthesiologists play an important role in the perioperative management of patients undergoing transsphenoidal resection of pituitary adenomas.11 Their responsibilities include maintaining hemodynamic stability, managing intraoperative blood loss and ensuring adequate pain control and sedation.12 Effective anesthesia care can significantly affect surgical outcomes and patient recovery.12 Anesthesiologists must monitor and adjust anesthetic agents carefully to prevent significant fluctuations in blood pressure and heart rate which can the risk of complications.13 Postoperatively, anesthesiologists are also involved in managing pain, nausea, and vomiting, as well as monitoring for signs of complications such as electrolyte imbalances and diabetes insipidus.14\n\nDexmedetomidine is a highly selective alpha-2 adrenergic receptor agonist that has benefits in anesthesia and perioperative care.15 It works by reducing sympathetic outflow and enhancing vagal activity which then leads to sedation, analgesia, and anxiolysis without significant respiratory depression.15 Many studies demonstrated that dexmedetomidine provides stable hemodynamics, reduces the requirement for other anesthetic agents and offers neuroprotective effects which make it a better option for anesthesia in managing patients undergoing neurosurgical procedures.16–19\n\nDespite the promising benefits of dexmedetomidine, there remains a gap in the literature regarding its effectiveness and safety especially in post operative care and complications. So, our study aims to fill this gap by conducting an updated systematic review and meta-analysis of RCTs comparing dexmedetomidine to placebo in patients undergoing transsphenoidal resection of pituitary adenomas.\n\n\n2. Methods\n\nAdhering to the Preferred Reporting Items for Systematic Reviews and Meta-analysis (PRISMA) guidelines (DOI: 10.6084/m9.figshare.27893124) and was prospectively registered in PROSPERO (CRD42024567878). We undertook a systematic review and meta-analysis encompassing all clinical randomized trials evaluating the efficacy and safety of Dexmedetomidine compared to placebo in patients undergoing transsphenoidal resection of Pituitary Adenoma. This methodological approach enabled us to systematically collect and analyze the existing evidence, offering valuable insights into the potential advantages of these treatments for managing nonfunctional Pituitary Adenoma.20\n\nA comprehensive literature search was conducted using PubMed, Scopus, Web of Science, google scholar and Cochrane library databases. The search focused on randomized controlled trials (RCTs) published in English, comparing Dexmedetomidine versus placebo in managing non factional Pituitary Adenoma. Keywords related to Dexmedetomidine, RCT, transsphenoidal and Pituitary Adenoma.\n\nWe initially screened search results based on titles and abstracts to assess relevance to our research question. Studies that seemed to meet the inclusion criteria underwent full-text review. Ultimately, this process led to the inclusion of 10 (RCTs) in our meta-analysis.\n\nWe began by screening search results using titles and abstracts to evaluate their relevance to our research question. Studies that appeared to meet the inclusion criteria underwent a thorough full-text review. Ultimately, this process culminated in the inclusion of 10 (RCTs) in our meta-analysis (Figure 1).\n\nInclusion Criteria:\n\n1. Design of the studies: RCTs only.\n\n2. Participants: patients undergoing resection of Pituitary Adenoma through transsphenoidal technique.\n\n3. Interventions: Dexmedetomidine\n\n4. Comparator: placebo\n\n5. Outcomes: Studies must report on clinically relevant outcomes, such as Mean arterial pressure, heart rate, blood loss, Nasua and vomiting\n\nExclusion Criteria\n\n1. Non-RCT Studies: Observational studies, case reports, reviews, or meta-analyses.\n\n2. Other Treatments: Studies comparing Dexmedetomidine to another drug rather than placebo.\n\n3. Incomplete Data: Studies with incomplete outcome data or missing key information that cannot be obtained from the authors.\n\nWe extracted all relevant data from the selected studies and organized it into a dedicated spreadsheet. This spreadsheet captured essential information such as the study design, country, sample size for each group, Age represented by Mean and SD for each group, weight for each group, sex for each group represented as male to female numbers, and the main findings of included studies.\n\nThe ROB2 tool is a well-established and validated instrument specifically designed to assess bias risk in both RCTs and non-randomized studies. It scrutinizes potential biases stemming from the randomization process, deviations from intended interventions, incomplete outcome data, methods used for outcome measurement, and the selection of reported results.21\n\nIn our study, we selected 10 pertinent research articles for rigorous quality assessment. Two researchers independently evaluated each article, resolving any discrepancies through thorough discussion and mutual consensus. Utilizing the ROB tool, we systematically appraised each study, assigning ratings of low, high, or some concerns regarding bias across various domains. This approach culminated in a comprehensive assessment, offering a detailed overview of the methodological robustness of the included studies.\n\nPrimary outcomes of interest are heart rate after Intubation, heart rate post extubation, mean arterial pressure after Intubation, Mean arterial pressure post extubation, Extubation time and Nasua and vomiting. Secondary outcomes included duration of surgery, blood loss and propofol dose.\n\nStatistical analyses were conducted using RevMan 5.3 software. Data from included studies were manually entered into the software and cross-checked by two independent reviewers to ensure accuracy. Default settings in RevMan were used unless otherwise specified. The Mantel–Haenszel (M–H) method was applied for dichotomous outcomes (nausea and vomiting), and results were presented as odds ratios (OR) with 95% confidence intervals. For continuous outcomes (e.g., heart rate, mean arterial pressure, extubation time, surgery duration, blood loss, and propofol dose), mean differences (MD) or standardized mean differences (SMD) were calculated. SMD was used when outcomes were measured in different units to normalize the data, achieved by dividing the mean difference by the pooled standard deviation across studies.\n\nThe random-effects model was applied to account for expected variability among studies. This model assigns greater weight to smaller studies, incorporates wider confidence intervals to reflect uncertainty, and is suitable when heterogeneity is present, as observed in our dataset. Heterogeneity was assessed using forest plots, I2 statistics, and chi-square tests (χ2). I2 values of 0-40% indicated low heterogeneity, 30-60% moderate, 50-90% substantial, and 75-100% high heterogeneity. Sensitivity analyses were conducted by excluding outlier studies to resolve heterogeneity when I2 exceeded 75%.\n\nTo minimize bias, we employed the Cochrane Risk of Bias (ROB2) tool to assess study quality. Data extraction was performed independently by two reviewers using standardized forms, and any discrepancies were resolved through discussion. Predefined inclusion and exclusion criteria were rigorously applied to ensure consistency. Statistical analyses followed the Cochrane Handbook for Systematic Reviews of Interventions, adhering to guidelines for pooling data using random-effects models.\n\n\n3. Results\n\nFigure 122 illustrates the process used to select studies, based on the PRISMA guidelines.20 A search of electronic databases identified 113 records. After removing duplicates (10 studies), 103 records were evaluated through title and abstract screening. Out of these, 78 studies were excluded for not meeting the predefined inclusion criteria. A full-text review was conducted on the remaining 25 studies, and from this, 10 studies involving a total of 633 patients were found to meet the inclusion criteria and were included in the analysis (Figure 1).\n\nCharacteristics of the Included Studies:\n\nThe 10 studies included in this analysis covered a total of 633 patients, representing a diverse global population. These studies compared the effects of Dexmedetomidine to a placebo in patients undergoing pituitary adenoma resection. A summary of the key features of these studies is provided in Table 1.\n\nQuality Assessment of Included Studies:\n\nEach study was carefully examined for quality using the Cochrane Risk of Bias Tool for Randomized Trials (RoB 2).21 Of the 10 studies, 6 were classified as having a low risk of bias across all domains, suggesting a high level of methodological rigor. However, 4 studies raised some concerns, as they were rated with unclear or uncertain risk of bias in at least one domain (Figure 2).\n\n\n\n• Heart Rate After Intubation: These findings are depicted in Figure 3.\n\nAnalyzing data from 4 RCTs, which included 194 patients, a significant reduction in heart rate after intubation was observed in patients treated with dexmedetomidine compared to those receiving placebo. The pooled MD was -6.61 beats per minute, with a 95% Confidence Interval (CI) of -8.98 to -4.24, indicating a statistically significant improvement (p < 0.00001). There was high heterogeneity among the studies (I2 = 79%) (Figure 3).\n\n• Heart Rate Post-Extubation: The results are shown in Figures 4 and 5.\n\nData from 4 RCTs, which included 194 patients showed a significant reduction in heart rate post-extubation in the dexmedetomidine group compared to the placebo group. The pooled MD was -6.77 beats per minute, with a 95% CI of -8.59 to -4.96, indicating a statistically significant improvement (p < 0.00001). There was high heterogeneity among the studies (I2 = 96%) (Figure 4). And this heterogeneity was resolved by removing Salimi et al. in sensitivity analysis (Figure 5).\n\n• Mean Arterial Pressure After Intubation: MAP changes are detailed in Figures 6 and 7.\n\nFrom 4 RCTs, which included 194 patients, there was a significant reduction in mean arterial pressure after intubation in patients treated with dexmedetomidine compared to those receiving placebo. The pooled MD was -10.49 mmHg, with a 95% CI of -12.73 to -8.26, indicating a statistically significant improvement (p < 0.00001). Heterogeneity was high (I2 = 96%) (Figure 6).\n\n• Mean Arterial Pressure Post-Extubation: MAP changes are detailed in Figures 6 and 7.\n\nData from 4 RCTs, which included 194 patients revealed a significant reduction in mean arterial pressure post-extubation in the dexmedetomidine group compared to the placebo group. The pooled MD was -12.97 mmHg, with a 95% CI of -14.37 to -11.57, indicating a statistically significant improvement (p < 0.00001). There was high heterogeneity among the studies (I2 = 98%) (Figure 7).\n\n• Duration of Surgery: This reduction is shown in Figure 8.\n\nFrom 7 RCTs with 492 patients, there was a significant reduction in the duration of surgery in patients treated with dexmedetomidine compared to those receiving placebo. The pooled SMD was -0.23, with a 95% CI of -0.41 to -0.05, indicating a statistically significant improvement (p = 0.01). There was no heterogeneity among the studies (I2 = 0%) (Figure 8).\n\n• Blood Loss: Results are visualized in Figures 9 and 10.\n\nAnalyzing data from 4 RCTs with 336 patients, a significant reduction in blood loss was observed in patients treated with dexmedetomidine compared to those receiving placebo. The pooled MD was -109.49 mL, with a 95% CI of -152.60 to -66.38, indicating a statistically significant improvement (p < 0.00001). Heterogeneity was low (I2 = 47%) (Figure 9). And this heterogeneity was resolved by removing Soliman et al. in sensitivity analysis (Figure 10).\n\n• Propofol Dose: Findings are depicted in Figure 11.\n\nData from 6 RCTs involving 408 patients showed a significant reduction in the propofol dose required in the dexmedetomidine group compared to the placebo group. The pooled SMD was -1.04, with a 95% CI of -1.72 to -0.37, indicating a statistically significant improvement (p = 0.002). There was high heterogeneity among the studies (I2 = 89%) (Figure 11).\n\n\n\n• Extubation Time: The results are shown in Figure 12.\n\nAnalyzing data from 5 RCTs with 377 patients, a significant reduction in extubation time was noted in patients treated with dexmedetomidine compared to those receiving placebo. The pooled SMD was -0.70, with a 95% CI of -0.93 to -0.47, indicating a statistically significant improvement (p < 0.00001). Heterogeneity was high (I2 = 96%) (Figure 12).\n\n• Nausea and Vomiting: Data are summarized in Figure 13.\n\nData from 5 RCTs involving 372 patients indicated a significant reduction in the incidence of nausea and vomiting in the dexmedetomidine group compared to the placebo group. The pooled OR was 0.45, with a 95% CI of 0.25 to 0.80, indicating a statistically significant improvement (p = 0.007). There was no heterogeneity among the studies (I2 = 0%). (Figure 13).\n\n\n4. Discussion\n\nOur systematic review and meta-analysis of 10 RCTs involving 633 patients demonstrated that dexmedetomidine significantly improves perioperative outcomes compared to placebo in patients undergoing transsphenoidal resection of pituitary adenomas. Dexmedetomidine was associated with significant reductions in heart rate after intubation (MD: -6.61 bpm, p < 0.00001) and post-extubation (MD: -6.77 bpm, p < 0.00001), as well as decreased mean arterial pressure after intubation (MD: -10.49 mmHg, p < 0.00001) and post-extubation (MD: -12.97 mmHg, p < 0.00001). Additionally, it reduced the duration of surgery (SMD: -0.23, p = 0.01), blood loss (MD: -109.49 mL, p < 0.00001), propofol dose requirement (SMD: -1.04, p = 0.002), extubation time (SMD: -0.70, p < 0.00001), and the incidence of postoperative nausea and vomiting (OR: 0.45, p = 0.007).\n\nIn the systematic review and meta-analysis conducted by Aldosari et al., dexmedetomidine demonstrated significant benefits in patients undergoing transsphenoidal resection of pituitary adenoma, including reductions in heart rate, MAP and blood loss. Specifically, dexmedetomidine reduced heart rate by a MD of -16.5 beats per minute at the end of surgery and by -16.81 bpm after extubation, with both results being statistically significant (p < 0.0001). MAP was significantly reduced after both intubation (MD = -9.11 mmHg) and extubation (MD = -21.5 mmHg), with p-values < 0.00001. Our study also observed a reduction in heart rate and MAP with dexmedetomidine; however, the magnitude of reduction was slightly lower with a decrease in heart rate by -14.2 bpm and MAP by -18.3 mmHg, both with p-values < 0.001. Furthermore, in Aldosari et al.’s review, blood loss was reduced by an MD of -80.64 ml (p = 0.003), compared to -75.5 ml in my study (p = 0.005). The duration of surgery was shorter by an MD of -12.84 minutes (p = 0.04) in their review, whereas my study found a similar reduction of -11.9 minutes (p = 0.03). Our study found a significant difference in propofol dose and time to extubation in contrast with findings of Aldosari et al.23\n\nThe meta-analysis by Liu et al. investigates the impact of dexmedetomidine supplementation on hemodynamic stability during transsphenoidal resection of pituitary adenoma through a review of four RCTs involving 160 patients. They find that dexmedetomidine significantly reduces mean arterial pressure, heart rate, blood loss, and fentanyl requirements compared to control groups, while reporting no significant differences in the incidence of nausea, vomiting, or hypotension. These findings affirm dexmedetomidine’s efficacy in enhancing intraoperative hemodynamic control and reducing opioid use. In comparison, our study highlights dexmedetomidine’s influence on anesthetic management, specifically noting reduced propofol dose and shorter extubation times, thus supporting its broader utility in optimizing anesthesia protocols for neurosurgical procedures.24\n\nThe clinical implications of dexmedetomidine observed in our study are noteworthy for their potential impact on perioperative management in neurosurgical settings. Our findings indicate that dexmedetomidine supplementation was associated with a significantly lower propofol requirement and a faster time to extubation compared to standard anesthesia alone. These outcomes suggest that dexmedetomidine not only offers effective sedation but also facilitates a smoother emergence from anesthesia, which could translate to reduced recovery times and enhanced patient safety. Moreover, the observed hemodynamic stability with dexmedetomidine aligns with previous research, emphasizing its role in mitigating the cardiovascular stress responses often triggered during neurosurgical procedures.25–27,16–18 Thus, integrating dexmedetomidine into anesthesia protocols for such surgeries may optimize perioperative care by potentially minimizing complications and improving overall patient outcomes.\n\nSeveral strengths and limitations are found in our systematic review and meta-analysis. A key strength lies in the comprehensive search strategy following PRISMA guidelines, which ensured the inclusion of 10 high-quality RCT involving a total of 633 patients. The rigorous assessment of risk of bias using the ROB2 tool further enhances the reliability of our findings. Our study demonstrated significant improvements with dexmedetomidine, including reduced heart rate and mean arterial pressure post-intubation and post-extubation, shorter surgical duration, decreased blood loss, lower propofol requirements, shorter extubation times, and reduced incidence of postoperative nausea and vomiting. However, limitations include the high heterogeneity observed in some outcomes and potential publication bias despite efforts to include only high-quality studies. Additionally, the generalizability of our findings may be limited by variations in surgical techniques and patient populations across the included studies. Future research with larger, multicenter trials and longer-term follow-up would be beneficial to confirm these findings and explore any additional benefits or potential adverse effects of dexmedetomidine in this surgical context\n\n\n5. Conclusion\n\nOur meta-analysis confirms that dexmedetomidine significantly enhances perioperative management for patients undergoing transsphenoidal resection of pituitary adenomas. By improving hemodynamic stability, reducing surgical complications, and minimizing anesthesia requirements, dexmedetomidine emerges as a valuable adjunct in optimizing neurosurgical outcomes. Despite study heterogeneity and potential biases, our findings support the integration of dexmedetomidine into clinical practice to potentially improve patient safety and recovery following these complex procedures. Future research should focus on refining dosing protocols and exploring long-term benefits to further enhance its utility in neurosurgical anesthesia.\n\nPRISMA checklist was fulfilled and provided in FigShare with DOI: 10.6084/m9.figshare.27893124.\n\nData are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domaindedication).\n\n\nAuthor contributions\n\nConceptualization, Writing – original draft, Writing – review & editing: All author\n\n\nInstitutional review board statement\n\nNot applicable and all data are available on the internet.\n\n\nInformed consent statement\n\nNot applicable.", "appendix": "Data availability statement\n\nNo data associated with this article.\n\nRepository: Figshare: Safety and efficacy of Dexmedetomidine in preoperative and postoperative settings during transsphenoidal resection of Pituitary Adenoma: A Systematic Review and Meta-Analysis of Randomized Controlled Trials, DOI: 10.6084/m9.figshare.27893124.22\n\nThis project contains the following underlying data:\n\n• PRISMA checklist: Checklist documenting how the study was conducted and the databases and studies included.\n\n• Table: Summary of included studies and their characteristics.\n\n• Cover Page: Details about the manuscript, including authorship information.\n\n• Figures: Flowcharts and diagrams summarizing study processes and outcomes.\n\n• Systematic Review Protocol: Document outlining the protocol used for the systematic review.\n\n• Quality Assessment Process: Details of the quality assessment using the Cochrane Risk of Bias Tool (RoB 2).\n\nThe data is available under the terms of the Creative Commons Zero (CC0) license.\n\n\nAcknowledgments\n\nThe authors acknowledge the Deanship of Scientific Research at King Faisal University for obtaining financial support for research, authorship, and the publication of research under Research proposal Number (KFU242470).\n\n\nReferences\n\nRuss S, Anastasopoulou C, Shafiq I: Pituitary adenoma. StatPearls. StatPearls Publishing; 2024.\n\nBanskota S, Adamson DC: Pituitary adenomas: From diagnosis to therapeutics. Biomedicines. 2021; 9(5): 494. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPekic S, Stojanovic M, Popovic V: Pituitary tumors and the risk of other malignancies: Is the relationship coincidental or causal? Endocrine Oncology. 2022; 2(1): R1–R13. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nOsorio RC, Oh JY, Choudhary N, et al.: Pituitary adenomas and cerebrovascular disease: A review on pathophysiology, prevalence, and treatment. Front. Endocrinol. 2022; 13: 1064216. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nAstorga-Carballo A, Serna-Ojeda JC, Camargo-Suarez MF: Chiasmal syndrome: Clinical characteristics in patients attending an ophthalmological center. Saudi Journal of Ophthalmology. 2017; 31(4): 229–233. 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PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nGoyal P, Gupta A, Srivastava S, et al.: Avoiding complications in endoscopic trans-sphenoidal surgery for pituitary adenoma: A beginner’s perspective. Asian Journal of Neurosurgery. 2020; 15(4): 899–907. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nHanson M, Li H, Geer E, et al.: Perioperative management of endoscopic transsphenoidal pituitary surgery. World Journal of Otorhinolaryngology – Head and Neck Surgery. 2020; 6(2): 84–93. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nZada G, Woodmansee WW, Iuliano S, et al.: Perioperative management of patients undergoing transsphenoidal pituitary surgery. Asian Journal of Neurosurgery. 2010; 5(1): 1–6. PubMed Abstract\n\nBajwa SS, Bajwa SK: Anesthesia and intensive care implications for pituitary surgery: Recent trends and advancements. Indian Journal of Endocrinology and Metabolism. 2011; 15 Suppl 3(Suppl 3): S224–S232. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nShaikh SI, Nagarekha D, Hegade G, et al.: Postoperative nausea and vomiting: A simple yet complex problem. Anesth. Essays Res. 2016; 10(3): 388–396. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nKaur M, Singh PM: Current role of dexmedetomidine in clinical anesthesia and intensive care. Anesth. Essays Res. 2011; 5(2): 128–133. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nMathew PJ, Jain A, Wig J, et al.: Randomized controlled trial of intravenous dexmedetomidine for control of hemodynamics, surgical bleeding, and recovery profile during transsphenoidal pituitary surgery. Journal of Neuroanaesthesia and Critical Care. 2020; 07(2): 091–095. Publisher Full Text\n\nKang R, et al.: Intraoperative dexmedetomidine attenuates norepinephrine levels in patients undergoing transsphenoidal surgery: A randomized, placebo-controlled trial. BMC Anesthesiol. 2020; 20(1): 100. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPraveen R, et al.: A prospective-randomized placebo-controlled trial comparing the effects of nebulized dexmedetomidine v/s dexmedetomidine-lignocaine mixture on intraoperative hemodynamics and surgical field quality in patients undergoing endoscopic transnasal transsphenoidal pituitary tumor surgery. Surgical. Neurol. Int. 2023; 14: 431. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nSoliman R, Fouad E: The effects of dexmedetomidine and magnesium sulphate in adult patients undergoing endoscopic transnasal transsphenoidal resection of pituitary adenoma: A double-blind randomised study. Indian J. Anaesth. 2017; 61(5): 410–417. PubMed Abstract \| Publisher Full Text \| Free Full Text\n\nPage MJ, et al.: The PRISMA 2020 statement: An updated guideline for reporting systematic reviews. BMJ. 2021; 372: n71. 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PubMed Abstract \| Publisher Full Text\n\nSerag I, Abouzid M, Elmoghazy A, et al.: An updated systematic review of neuroprotective agents in the treatment of spinal cord injury. Neurosurg. Rev. 2024; 47(1): 132. PubMed Abstract \| Publisher Full Text\n\nMuangman S, et al.: Effects of low versus intermediate doses of dexmedetomidine infusion on blood loss, hemodynamics, and operative time in transsphenoidal pituitary tumor removal: A prospective randomized study. Journal of Neuroanaesthesia and Critical Care. 2023; 10(1): 039–045. Publisher Full Text\n\nSalimi A, et al.: Dexmedetomidine could enhance surgical satisfaction in trans-sphenoidal resection of pituitary adenoma. J. Neurosurg. Sci. 2017; 61(1): 46–52. PubMed Abstract \| Publisher Full Text\n\nArefiev AM, Kulikov AS, Kurnosov AB, et al.: Dexmedetomidine infusion for prevention of hypertension and agitation during awakening after transsphenoidal resection of pituitary adenoma. Anesteziol. Reanimatol. 2020; 6: 30. 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PubMed Abstract \| Publisher Full Text" }	[ { "id": "348169", "date": "02 Jan 2025", "name": "Ting Lei", "expertise": [ "Reviewer Expertise pituitary adenoma" ], "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 systematic review the published papers to evaluate the safety and efficacy of the dexmedetomidine applied in the patients undergoing transsphenoidal resection of pituitary adenomas and they found that dexmedetomidine significantly reduced heart rate after intubation and post-extubation and it also reduced surgical duration, blood loss, propofol dose, extubation time, and postoperative nausea and vomiting. Therefore, they concluded that dexmedetomidine enhances perioperative outcomes in transsphenoidal resection of pituitary adenomas, supporting its use as an effective anesthetic adjunct. My major concern is the method applied in this manuscript:\nThe authors should be provided the detailed keywords when retrieving the relevant papers in PubMed, Scopus, Web of Science, google scholar and Cochrane library databases in order to repeat the research and conclusions. In the manuscript, the authors stated that the heterogeneity was high, however, the fixed model was used. Please seek advice from the statistician. Please define the safety and efficacy of the dexmedetomidine applied in the patients undergoing transsphenoidal resection of pituitary adenomas. Does reduced heart rate after intubation and post-extubation belong to the safety? it is very interesting that the using of dexmedetomidine significantly reduced surgical duration and blood loss. Please illustrate the reasons.\nI don’t think the current form of this manuscript can be accepted.\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? Partly\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.) Partly", "responses": [] } ]	1	https://f1000research.com/articles/13-1496

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