M. bovis) is the zoonotic bacterium responsible for bovine tuberculosis. An attenuated form of M. bovis, Bacillus Calmette-Guerin (BCG), is a modified live vaccine known to provide variable protection in cattle and other species. Protection for this vaccine is defined as a reduction in disease severity rather than prevention of infection and is determined by evaluation of the characteristic lesion of tuberculosis: the granuloma. Despite its recognized ability to decrease disease severity, the mechanism by which BCG imparts protection remains poorly understood. Understanding the histopathologic differences between granulomas which form in BCG vaccinates compared to non-vaccinates may help identify how BCG imparts protection and lead to an improved vaccine. Utilizing special stains and image analysis software, we examined 88 lymph nodes obtained from BGC-vaccinated and non-vaccinated animals experimentally infected with M. bovis. We evaluated the number of granulomas, their size, severity (grade), density of multinucleated giant cells (MNGC), and the amounts of necrosis, mineralization, and fibrosis. BCG vaccinates had fewer granulomas overall and smaller high-grade granulomas with less necrosis than non-vaccinates. The relative numbers of high- and low- grade lesions were similar as were the amounts of mineralization and the density of MNGC. The amount of fibrosis was higher in low-grade granulomas from vaccinates compared to non-vaccinates. Collectively, these findings suggest that BCG vaccination reduces bacterial establishment, resulting in the formation of fewer granulomas. In granulomas that form, BCG has a protective effect by containing their size, reducing the relative amount of necrosis, and increasing fibrosis in low-grade lesions. Vaccination did not affect the amount of mineralization or density of MNGC.Mycobacterium bovis ( Mycobacterium bovis (M. bovis) is the most host-promiscuous member of the Mycobacterium tuberculosis complex (MTBC), capable of causing tuberculous disease in over 85 species of animals, including some endangered wildlife species ( species . It is tMycobacterium bovis which has been used as a vaccine for protection against Mycobacterium tuberculosis (Mtb) in humans for over 100 years. It has long been accepted as providing variable protection against virulent M. bovis in livestock and wildlife , lower numbers of cattle with gross lesions, and less severe disease than non-vaccinates  is an attenuated strain of wildlife . Importaironment . In natuccinates . Becausesmission . Thus, tsmission .The characteristic lesion of tuberculosis is the granuloma. Granulomas are collections of macrophages with multiple modifiers such as the presence or absence of varying amounts of neutrophils, lymphocytes, multinucleated giant cells (MNGC), fibrosis, necrosis, and mineralization. These are dynamic structures, with cell trafficking resulting from host- and bacterially-originating signals. Until recently, it was believed that the tuberculous granuloma was a structure which benefited both host and pathogen at different points in infection, early on by promoting dissemination of the bacteria as a result of migrating macrophages, and later by “walling off” the offending intruder through fibrosis. Although this paradigm remains popular, new advances in research create challenges for its defense . Understet. al have demonstrated reduced granuloma numbers, size, necrosis, peripheral fibrosis, and MNGC numbers in lymph nodes of BCG vaccinated animals compared to non-vaccinated controls when experimentally infected with virulent M. bovis  and BCG vaccinates (n = 12). Animals were kept on pasture prior to the start of the study. As environmental bacteria are present in our area, and in order to prevent possible confounding responses to BCG vaccination, animals were confirmed non-reactive to Mycobacterium avium (M. avium) via interferon gamma release assay (IGRA) immediately before the start of the study. Animals in the vaccinated group received a subcutaneous injection 1 ml of 5 × 105 CFU of BCG Danish. Three months following vaccination, both vaccinates and non-vaccinates received 5.5 × 102 CFU of M. bovis strain 1,315 via aerosolization as described elsewhere (n = 11) and BCG vaccinates (n = 10). Animals were bottle-raised on a pasteurized milk product and kept in clean pens to reduce exposure to environmental mycobacteria. Due to the cleanliness of the housing situation the risk of exposure to environmental mycobacteria was considered low and IGRA testing for M. avium was not performed. Animals in the vaccinated group received a subcutaneous injection 1 ml of 1 × 106 CFU of BCG Danish at 2 weeks of age. Three months following vaccination, both vaccinates and non-vaccinates received 1 × 103 CFU of M. bovis strain 1,315 via aerosolization. Approximately 18 weeks post-challenge animals were euthanized. In both studies animals were humanely euthanized by intravenous administration of sodium pentobarbital. All animal procedures were approved prior to the experiments by the Institutional Animal Care and Use Committee (IACUC) at the National Animal Disease Center. At necropsy, both experimental groups demonstrated a significant reduction in gross lesions in the lungs of BCG-vaccinates compared to non-vaccinates and a significant decrease in CFU/gram of lymph node tissue between vaccinates and non-vaccinates . Von Kossa Calcium and Masson’s Trichrome staining were performed following manufacturer’s instructions (Newcomer Supply) for calcium and collagen fiber analysis, respectively. Following staining, slides were digitally scanned using the Aperio AT2 whole slide imager, to generate bright-field whole- slide images at 20X (0.5 μm/pixel) and 40X (0.25 μm/pixel) magnification in a 24-bit color pyramid TIFF (.SVS file). The. SVS files were transferred into HALO software  for annotation and analysis.The mediastinal and tracheobronchial lymph node values for each metric were initially evaluated separately to ensure differences were not a result of anatomic location. No significant differences were noted, therefore values from the tracheobronchial and mediastinal lymph nodes were combined and averaged to yield a single value per animal.Digitally scanned images were evaluated at 10X magnification. Using HALO annotation tools, granulomas were outlined in different colors based on grade, and the total granuloma area calculated using HALO’s annotation software. A granuloma was defined as a focus consisting of increased accumulations of macrophages admixed with varying numbers of lymphocytes, neutrophils and MNGCs. If a tissue section lacked granulomas, it was excluded from the remaining histopathologic examinations. Granulomas were separated into two grades: low and high.Low-grade granulomas were defined as singular, with approximately ≤10% necrosis. High-grade granulomas were defined as having greater than 10% of its total area as necrosis, being multicentric, or coalescent with other granulomas. Necrosis was defined as an area of tissue destruction characterized by degenerating, fragmented cells, and structural debris with or without mineralization. When present, fibrous encapsulation was included in the total granuloma area calculated. Care was made to closely follow the directional flow of the collagen fibers when numerous, closely associated but independent granulomas were present. When fibrous bands incompletely separated granulomas or had no identifiable pattern of separation, these were counted as a single, coalescing lesion and considered high grade.2 of non-necrotic area.High grade granulomas were outlined in green while low grade granulomas were outlined in yellow . The absNumerous previous studies utilized a four-stage scoring system developed by Wangoo et al to describe granuloma histopathology . GranuloUtilizing Von Kossa-stained slides, high- and low-grade granulomas were identified in two independent layers, at 10X magnification. Samples without identifiable lesions were excluded. High- and low-grade designations used for the H&E stains were followed to classify the lesions into their respective groups. These outlines served as the boundaries for tissue to be analyzed. A digital filter was formulated to identify calcium-containing areas representative of mineralization. Starting with algorithm Indica labs-Area quantification v2.1.7, a stain color was selected and optimized to identify mineralization. The percent area of mineralization for both high- and low-grade granulomas was calculated using the area of staining identified by the filter divided by the total area analyzed for each high- and low-grade granuloma.Utilizing Masson’s trichrome-stained slides, high- and low-grade granulomas were outlined in two independent layers, at 10X magnification to ensure all granulomas were identified. Samples without identifiable lesions were excluded. High- and low-grade designations used for the H&E stains were followed to classify the lesions into their respective groups. A digital filter was formulated to identify collagenous areas highlighted by the trichrome stain and applied to the entire granuloma. Starting with algorithm Indica labs-Area quantification v2.1.7, a stain color was selected and optimized to identify fibrosis based on the color of collagen found in dense collagenous bundles. The filter was applied to each annotated layer. The percent fibrosis for high- and low-grade granulomas was calculated using the area of staining identified by the filter divided by the total area analyzed for each high- and low-grade granuloma.p-value ≤0.05. Error bars represent standard errors.For all scored granuloma metrics  data were analyzed with a simple linear regression model (lm) in R (version 4.2.1). Each scored metric was evaluated independently, fitting grade, vaccination status, and their interaction as fixed effects. A fixed effect of ‘group’, denoting the two separate experimental cohorts, was evaluated for model fit for each metric and only deemed appropriate to include for analysis of percent fibrosis. A pairwise comparison of Least Squares means (lsmeans package) was utilized to determine significant differences between specific grade and vaccination group contrasts. For all, significance was determined when Graph construction was performed using GraphPad Prism 9 . Values for the mediastinal and tracheobronchial lymph nodes for each animal were combined to yield a single data point per animal. For all analyses performed except in the evaluation of the total number of granulomas, animals without granulomas in either lymph node were excluded. In cases where an animal lacked either high- or low-grade granulomas , these data points were excluded with exception of granuloma count.BCG vaccinates had fewer granulomas compared to non-vaccinates. The ratios of high- and low-grade granulomas were similar between vaccinates and non-vaccinates.p < 0.0001; Non-vaccinated animals had higher numbers of granulomas compared to BCG-vaccinates .While the number of granulomas varied markedly between BCG-vaccinates and non-vaccinates, the relative percentages of each grade of granuloma were similar . There wBCG vaccinates had smaller high-grade granulomas and lesions with less necrosis.p = 0.0675) in the average size of granulomas between vaccinates and non-vaccinates, however lesions from non-vaccinated animals tended to be smaller  while non-vaccinates had larger high-grade granulomas compared to low-grade granulomas (p = 0.0064).The average size (area) of granulomas was compared between vaccinates and non-vaccinates. There was no significant difference  less tissue destruction as evidenced by a lower percentage of necrosis than non-vaccinates (p < 0.001 for both). There was no significant difference in the percentage of necrosis within low-grade granulomas of vaccinates compared to non-vaccinates with values being nearly identical  higher percentage of necrosis than high-grade granulomas from BCG-vaccinates.In this study, to be considered low-grade, the amount of necrosis had to be ≤10% the total area. However, high grade granulomas may have ≤10% of necrosis if they were multicentric or coalescing. BCG vaccinates had lesions containing significantly (ccinates . As a reMNGC numbers were not dependent on vaccination status or lesion severity.2 of granuloma area. No significant differences were found in the density of MNGC between vaccination groups (p = 0.0632) for low grade granulomas of non-vaccinates to contain a higher number of MNGC than high grade granulomas of the same group  higher percentage of fibrosis than low-grade granulomas of non-vaccinates  had higher average fibrosis than the younger animals . That being said, there were no significant differences in the percentage of fibrosis per granuloma between vaccinates and non-vaccinates for either experimental group . Collectccinates . When miccinates . Additioccinates . SignifiThis study utilized 88 pulmonary lymph nodes from 44 animals , resulting in the evaluation of a total of 1,770 granulomas. Of these, 417 granulomas were in BCG vaccinates while 1,353 were in non-vaccinates.Previous studies which have focused on granuloma histopathology have utilized a 4-stage scoring system created by Wangoo et al . While oAs expected, and in accordance with previous studies, BCG decreased the lesion burden . VaccinaM. bovis . This was unexpected and warrants further investigation, as it has not been reported previously. A possible explanation for the difference in fibrosis found between the two age groups could be levels of tumor growth factor beta (TGF-β), the cytokine most responsible for stimulating fibrosis. However, according to a study performed in sheep, TGF-β is lower in older animals vs. neonates . This woM. bovis . RegardlM. bovis . A possiM. bovis . When brM. bovis . In humaM. bovis . These sDystrophic mineralization is a common sequela to necrosis and therefore it is not surprising to see levels of mineralization increased in high grade granulomas. Nevertheless, it is interesting to note that the percentage of mineralization between high-grade granulomas were similar between vaccination groups and that small amounts of mineralization were present in the low-grade granulomas. While the Wangoo categorization system attributes mineralization with increased lesion score, and therefore, increased disease severity, studies in humans and non-human primates associate mineralization with healing, similar to fibrosis . Indeed Multinucleated giant cells are associated with persistent antigen in multiple diseases . We hypoOne limitation of this study was in interpreting coalescing lesions which had effaced the majority of the lymph node parenchyma. These cases arose occasionally in both vaccinates and non-vaccinates. This phenomenon is well document in lymph nodes of chronically infected cattle, where it may completely erase any semblance of a normal lymph node . BCG proMycobacterium bovis in liquid culture is difficult due to the propensity of the bacilli to clump  staining for mycobacteria has historically been performed in histopathologic examinations of bTB it was not performed in this study due to limitations of the software and the relative insensitivity of ZN staining in formalin-fixed, paraffin-embedded tissues . As quanM. bovis. Once established, BCG does not prevent the development of severe lesions, and the relative breakdown of high- and low- grade granulomas is similar to that of non-vaccinates. Despite this fact, BCG vaccinates have smaller granulomas with less tissue destruction than non-vaccinates. Fibrosis was higher in low-grade granulomas of vaccinates compared to non-vaccinates but this significance was lost when grade was excluded. Mineralization and MNGC density were similar between vaccinates and non-vaccinates, suggesting these may be vaccine-independent events.Our study provides further support to show that BCG vaccination reduced granuloma formation within the pulmonary lymph nodes of cattle following aerosol challenge with virulent The original contributions presented in the study are included in the article, further inquiries can be directed to the corresponding author.The animal study was reviewed and approved by Institutional Animal Care and Use Committee at National Animal Disease Center.CK, PB, and MP: experiment design. CK and MP: sample collection and experiments. CK and EP: data analysis. CK: manuscript preparation. CK, PB, EP, and MP: manuscript editing. All authors contributed to the article and approved the submitted version.Financial support for these studies were provided by the United States Department of Agriculture, Agricultural Research Service Project (CRIS #5030–32000-222). This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.USDA is an equal opportunity provider and employer. Mention of trade names or commercial products in this publication is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Antimicrobial stewardship interventions are targeted efforts by healthcare organizations to optimize antimicrobial use in clinical practice. The study aimed to explore effective interventions in improving antimicrobial use in hospitals. Literature was systemically searched for interventional studies through PubMed, CINAHL, and Scopus databases that were published in the period between January 2010 to April 2022. A random-effects model was used to pool and evaluate data from eligible studies that reported antimicrobial stewardship (AMS) interventions in outpatient and inpatient settings. Pooled estimates presented as proportions and standardized mean differences. Forty-eight articles were included in this review: 32 in inpatient and 16 in outpatient settings. Seventeen interventions have been identified, and eight outcomes have been targeted. AMS interventions improved clinical, microbiological, and cost outcomes in most studies. When comparing non-intervention with intervention groups using meta-analysis, there was an insignificant reduction in length of stay  and a significant reduction in antibiotics’ days of therapy . There were noticeable reductions in readmissions, mortality rates, and antibiotic prescriptions post antimicrobial stewardship multi-disciplinary team (AMS-MDT) interventions. Studies that involved a pharmacist as part of the AMS-MDT showed more significant improvement in measured outcomes than the studies that did not involve a pharmacist. Clostridioides difficile infection (CDI), adverse effects, and increasing antimicrobial and non-antimicrobial healthcare expenses are all major problems  resistance rates and CDI rates) and other outcomes including antibiotic prescribing rates, antibiotic consumption, and cost. Days of therapy are the number of days in which a patient has received antibiotic therapy, while length hospital stay is the difference in days between patient hospital admission and discharge. The primary investigators established a standard data extraction form using Microsoft Excel®. This data extraction sheet was divided into four tables: Inpatient settings (IP) with a pharmacist as part of the AMS-MDT, IP settings without a pharmacist, outpatient settings (OP) without a pharmacist, and OP settings with a pharmacist. The following data were gathered from the identified studies: author name, year, country, sample size, study design, infection site, intervention type, outcomes, and findings. Data extraction was undertaken by two investigators (A. A. Sadeq and N. AbouKhater).The risk of bias was assessed using Version 2 of Cochrane risk-of-bias tool (RoB2) for randomized control trials . Based oFor non-randomized trials, Risk of Bias in Non-Randomized Studies of Interventions (ROBINS-1) was used for bias risk assessment . The oveThe process of risk of bias assessment was performed independently by two investigators (A. Sadeq and N. AbouKhater) and any disagreements were resolved by discussion and consensus.This review has been recorded in PROSPERO  under the code CRD42022302431.2 statistic was used to look at heterogeneity as it calculates the percentage of overall variation that can be attributed to between-study heterogeneity. I2 values of 25%, >50%, and >75% refer, respectively, to low, substantial, and considerable degrees of heterogeneity. Funnel plots were generated using inverse variance methods to examine the publication bias.RevMan® software version 5.4.1 and MetaXL software version 5.2 were used to conduct the analyses with random-effects model to pool and evaluate data from eligible studies that reported the same outcomes. Pooled estimates were represented as a forest plot with a 95 percent confidence interval (CI) range for risk differences and mean differences. The IThe present review has identified influential antimicrobial stewardship multidisciplinary team interventions in both inpatient and outpatient settings. Twenty-one interventions have been recognized with the most common interventions being prospective audit and feedback with direct intervention, antibiotic restriction and pre-authorization, regular dedicated ID rounds, HCP education, use of clinical decision support system, antimicrobial guidelines implementation, and retrospective audit and feedback. The inclusion of a pharmacist as a part of the multidisciplinary team increased the chances of achieving statistically significant changes in the outcomes. Those interventions were able to improve clinical , microbiological, and cost outcomes when AMS-MDT interventions were applied and compared to non-intervention groups.


Participants completed self-report questionnaires in English or Sesotho. Results: Participants  had high HIV self-management scores  that corresponded with treatment outcomes . This might be attributed to condition-specific factors, including once-daily doses (100%) and longer duration of treatment (81.4% on ART for more than 10 years). Participants were older (median age 22), and the majority (61.7%) had stable living conditions. Individual strengths were associated with higher self-management scores (p < 0.01) and mental health problems with lower self-management scores (p < 0.05). Most (97.9%) were satisfied with their health care services. Conclusions: Uncomplicated treatment regimens, longer duration of treatment, stable living conditions, individual strengths, good mental health, and satisfaction with healthcare services have a positive influence on self-management.Background: HIV treatment outcomes of adolescents and youth living with HIV (AYLWH) are lagging. One way to improve outcomes is through supporting AYLWH to acquire self-management skills. Although self-management is associated with improved health outcomes, condition-specific, individual/family, and social/environmental contextual factors influence self-management. We aimed to describe factors influencing the self-management of AYLWH in Lesotho. Methods: A cross-sectional survey design was used. AYLWH ( The proportion of adolescents and youth living with human immunodeficiency virus (HIV) globally is growing. In 2020, 410,000 young people between the ages of 10 and 24 were newly infected with HIV. Among these, 150,000 were adolescents between the ages of 10 and 19  nWe had to adapt the original self-management scale to improve the reliability and validity of this sample. Therefore, the required self-management skills of AYLWH and abilities may evolve from adolescence to early adulthood  and shouSelf-management scores in this study were high, with the medical management scale, which included items such as knowledge of viral load, having the highest mean scores. The self-management scale with the lowest mean items scores was the resiliency scale related to emotional management. Participants had a lower mean score for having regular contact with friends. Positive relationships and resources such as family, peers, and friends facilitate AYWH engagement in care .The high self-management scores in this study corresponded with the high adherence and viral suppression rates found in this study. Crowley et al. (2020)  also fouHigh self-management scores may be explained by individual factors such as age, although age was not associated with self-management in our study. Our sample included predominantly older adolescents with apparently stable family conditions. More than half completed grades 10–12. Older adolescents might be more prepared and ready for the transition into adult care, as they become increasingly independent and more capable of self-management . Other iSatisfaction with services may improve self-management. Adolescents reported high satisfaction and liked attending the clinic, although some reported transport problems. Cluver et al. 2018) found that health system factors improving retention in the care of AYLWH included staff who are kind and have time for adolescents . The sat18 found To our knowledge, this is the first study to explore self-management amongst AYLWH in Lesotho. Due to the use of convenience sampling, AYLWH not attending services were excluded, and thus, the results may not accurately represent all AYLWH. Our sample included predominantly females and older youth. There was limited variability in self-management scores and some of the independent variables. Only two facilities were included in the study, and these were facilities that provided adolescent-friendly services. These findings may, therefore, not apply to other settings where such services are not rendered. The findings might be generalizable to settings where adolescent-friendly services are provided to older, predominantly female AYLWH. Future studies should consider using stratified random sampling to ensure representation of gender, age, and level of adherence/clinic attendance. A comparison of settings that provide adolescent-friendly services and those that do not might assist in teasing out the effect of these services on self-management.The results of this study provide further support that uncomplicated treatment regimens, longer duration of treatment, stable living conditions, individual strengths, good mental health, and satisfaction with health care services have a positive influence on self-management. The required chronic disease self-management skills and abilities of AYLWH may be context-dependent and evolve and should be explored further.


Therefore, close monitoring of AI is mandatory, as the prognostic nature of its longitudinal progression is still unclear. Clearly, significant AI during VAD support warrants surgical intervention at the appropriate timing, especially in patients of destination therapy. Nonetheless, such an uncertainty in the progression of AI translates to a lack of consensus regarding the management of this untoward complication. In practice, proposed surgical options are aortic valve replacement, repair, closure, and more recently transcatheter aortic valve implantation or closure. Transcatheter approach is of course less invasive, however, its efficacy in terms of long-term outcome is limited. In this review, we summarize the recent evidence related to the pathophysiology and surgical treatment of AI associated with CF-LVAD implantation.Moderate to severe aortic insufficiency (AI) in patients who underwent continuous-flow left ventricular assist device (CF-LVAD) implantation is a significant complication. According to the INTERMACS registry analysis, at least mild AI occurs in 55% of patients at 6 months after CF-LVAD implantation and moderate to severe AI is significantly associated with higher rates of re-hospitalization and mortality. The clinical implications of these data may underscore consideration of prophylactic aortic valve replacement, or repair, at the time of CF-LVAD implantation, particularly with expected longer duration of support and in patients with preexisting AI that is more than mild. More crucially, even if a native aortic valve is seemingly competent at the time of VAD implantation, we frequently find  Accorc valves –13. Therc valves –18, espeIn practice, proposed surgical options are AVR , 20, AV de novo AI under CF-LVAD support remain controversial, following three factors are likely to be associated with de novo AI: (1) continuous or intermittent AV closure due to the constant increase of aortic diastolic pressure with the decrease in LV end-diastolic pressure, (2) increased transvalvular gradient due to decompression of the LV, leading to stretching or partial prolapse of AV leaflets, and (3) pathologic changes or dilatations in the aortic sinus due to turbulent backflow with high blood velocity from a CF-LVAD outflow . Kainuma et al. suggested such an altered mechanical stress on the AV could be associated with the structural, functional, and histological changes of the aorta and AV  and lower rates of survival  (p = 0.024] (p = 0.038) (p = 0.006)  . Auvil ede novo AI occurred in 15.2% of patients after CF-LVAD implantation and such a de novo AI was not significantly associated with mortality , during median follow-up of 527 days , at 1 year after VAD implantation   in their randomized study  , that thed study , which cThus, there are several conflicting studies to each other, regarding the risk factors of significant AI and its effect on mortality. However, given the results shown in the registry analysis , it woulp = 0.026), but not thromboembolic events  should be addressed with either valve closure, repair, or replacement  Class o. It shouc events . These fc events .p < 0.01). Notably, more CHF-related readmissions were observed in the mild AI group, as compared with no or trace AI   . Their rp = 0.031), leading to no survival difference  (p < 0.0001)  . As for  0.0001) . Althoug 0.0001) . From thWhile AVR with bioprosthetic valve contains a few issues, such as longer ischemic time, valve thrombosis, or commissural fusion , AV repaA largest single-center experience of AV repair by central AV closure at VAD implantation is reported by Fukuhara and colleagues . They con = 40) and AVRs (n = 6)  . The CAVde novo AI after CF-LVAD implantation between those who were on the Impella support and those not, concluding that mild or moderate de novo AI was observed in 82% of patients in the Impella group, as compared 43% in the non-Impella group (p = 0.038)  donor heart allocation system commenced giving a priority to patients supported with non-dischargeable mechanical circulatory support (MCS) devices while awaiting heart transplantation, prompting temporary MCS devices in heart transplant centers being more frequently used in the United States (US) . The Imp= 0.038) . The patantation . They spantation . Thus, ede novo AI after VAD implants   we currently tend to intervene on the AV more aggressively at the time of CF-LVAD implantation and the need of late AV intervention is decreasing, (2) for late de novo AI, less invasive procedures, such as TAVI or percutaneous closure, are more likely to be conducted instead of surgical interventions, and (3) In some countries, urgent heart transplantation is now becoming a feasible option to deal with de novo significant AI. For these reasons, there is no definite agreement on an ideal secondary AV intervention, and the decision should depend on each clinical scenario.To the best of our knowledge, prospective studies on the efficacy of bioprosthetic AVR or AV repair for late de novo AI in VAD patients, especially when significant morphological change in the AV is observed. However, this procedure necessitates longer ischemic time and decent exposure of the aortic root, as compared to AV repair, which may raise a concern for postoperative right ventricular dysfunction in these high-risk candidates , warranting a prudent judgement on the indication of surgical AVR for late de novo AI. Additionally, Doi et al. reported a case of commissural fusion after bioprosthetic AVR after CF-LVAD implantation, casting doubt on bioprosthetic AVR as a desirable option, as compared to AV repair (Apart from the clinical case series above, there is a registry database analysis on the risk of AVR in CF-LVAD patients. Zaidi et al. reported the survival outcomes of AVR and TAVI late after CF-LVAD implantations, using the Nationwide Readmission Database in the US . AlthougV repair . Right vV repair . Based oin-situ CF-LVAD may disturb prosthesis deployment and increase the risk of prosthesis migration and still there is no consensus on how to optimize pump speed to prevent valve migration while deploying the device (Another option for treating AI could be TAVI. Although TAVI is not used routinely as a treatment option for severe AI, as of the year 2022, an international multicenter registry data already demonstrated its feasibility and efficacy in non-VAD patients, especially with new-generation devices . Howevere device . Hopefule device , 73, suce device and J-valThere are three other case series regarding TAVI for AI in CF-LVAD patients. First one is by Belkin et al., reporting 7 patients underwent 9 attempted TAVI procedures . UnfortuAs mentioned previously, concomitant AV closure at CF-LVAD implantation was associated with increased risk of mortality . Moreovede novo AI, still there is no clear consensus on the timing of intervention or the choice of treatment modality. Clearly, symptomatic severe AI in a CF-LVAD patient needs to be addressed, either surgically or percutaneously. Hopefully in the future, TAVI would become a first-line treatment of late de novo AI in CF-LVAD patients, after technical refinements and device improvements. Despite the scarcity of established evidence so far, our continuing efforts are imperative to develop new insights in the future, overcoming this scabrous clinical entity during CF-LVAD support.The present review summarized current updates on CF-LVAD associated AI, in terms of its pathophysiology, incidence, clinical impacts on outcomes, prevention, and surgical or trans-catheter interventions. As a concomitant procedure with CF-LVAD implantation, current guidelines are recommending AV repair or AVR for greater than mild AI, which is well-supported . For milBoth authors listed have made a substantial, direct, and intellectual contribution to the work and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous non-Hodgkin's lymphoma with great clinical challenge. Machine learning (ML) has attracted substantial attention in diagnosis, prognosis, and treatment of diseases. This study is aimed at exploring the prognostic factors of DLBCL by ML.  In total, 1211 DLBCL patients were retrieved from Huaihai Lymphoma Working Group (HHLWG). The least absolute shrinkage and selection operator (LASSO) and random forest algorithm were used to identify prognostic factors for the overall survival (OS) rate of DLBCL among twenty-five variables. Receiver operating characteristic (ROC) curve and decision curve analysis (DCA) were utilized to compare the predictive performance and clinical effectiveness of the two models, respectively.  The median follow-up time was 43.4 months, and the 5-year OS was 58.5%. The LASSO model achieved an Area under the curve (AUC) of 75.8% for the prognosis of DLBCL, which was higher than that of the random forest model (AUC: 71.6%). DCA analysis also revealed that the LASSO model could augment net benefits and exhibited a wider range of threshold probabilities by risk stratification than the random forest model. In addition, multivariable analysis demonstrated that age, white blood cell count, hemoglobin, central nervous system involvement, gender, and Ann Arbor stage were independent prognostic factors for DLBCL. The LASSO model showed better discrimination of outcomes compared with the IPI and NCCN-IPI models and identified three groups of patients: low risk, high-intermediate risk, and high risk.  The prognostic model of DLBCL based on the LASSO regression was more accurate than the random forest, IPI, and NCCN-IPI models. Diffuse large B-cell lymphoma (DLBCL) is the most common histological subtype of non-Hodgkin's lymphoma, manifesting highly heterogeneity in genetic and phenotypic characteristics. The International Prognostic Index (IPI) and enhanced International Prognostic Index (NCCN-IPI) are widely used prognostic models mainly based on clinical variables such as age, stage of disease, and performance status, which are challenged due to improved treatment options, pathobiology, and life expectancy of patients with DLBCL –3. AnothMachine learning (ML) is widely defined as a computational strategy and a branch of artificial intelligence (AI). It automatically determines methods and parameters to obtain an optimal solution to the problem. The learning process assumes that it simulates an aspect of human intelligence and can be used for superficial intelligent purposes . ML clasLASSO is a regression-based methodology permitting for a large number of covariates in the model, which introduces regularization function to punish excessive fitting on the basis of logistic regression, making it compress some regression coefficients and make the coefficients with smaller absolute values to 0, so as to automatically remove unnecessary/uninfluential covariables, and can simultaneously select variables and estimate parameters , 14, 15.Few studies have explored the prognostic factors of DLBCL using ML based on clinical variables , 21. Then = 848) and a validation cohort . All pathological biopsies were double blinded and reviewed by at least two pathologists. Patients included in this study were treated with rituximab-based immunochemotherapy. Exclusion criteria is as follows: (1) patients with other tumors and (2) special types of lymphoma . Ethics approval was obtained from independent Ethics Committees of each participating center in HHLWG. This study was conducted in accordance with the declaration of Helsinki.We retrospectively collected 1211 newly diagnosed DLBCL patients from August 2008 to January 2021 from 7 medical centers of the Huaihai Lymphoma Working Group (HHLWG). Patients were randomized into a training cohort , presence of bulky disease (≥7.5 cm), B symptoms, albumin (ALB), white blood cell count (WBC), hemoglobin (HB), platelets (PLT), total cholesterol (TC), lymphocyte count (LYC), red blood cell count (RBC), neutrophil count (NE), height, weight, Ann Arbor stage, cell of origin, and immunological markers . GCB or non-GCB phenotypes were determined by the Hans algorithm.Follow-up was conducted by reviewing inpatient medical records and making phone calls. All patients were followed up until July 28, 2021, or until death, whichever came first. Overall survival (OS) was calculated as the interval between the time of diagnosis and death from any cause or the last follow-up. The survival status of all patients was confirmed with death records or a telephone call to the patients themselves or to the next of kin of the patient (if patient died during the follow-up).U-test. Continuous variables were transformed into categorical variables by MaxStat analysis .Data were presented as numbers (percentages) for categorical variables and median  for all continuous variables. Clinical factors between the training and validation cohorts were compared using the Chi-squared test and the Mann–Whitney λ. Random forest regression model for random forest regression analysis was constructed based on Breiman's random forest algorithm, and the Cox proportional hazards model was used to analyze the multivariable association between prognostic factors, identified in random forest regression analysis, and the OS of DLBCL. The discrimination ability of the LASSO-cox and Random forest regression models were evaluated by the receiver operator characteristic (ROC) curve analysis and Harrell's concordance index. Area under the curves (AUCs) of different models were compared using DeLong's test. For clinical usefulness, net benefit was examined against the training and validation cohorts using the decision curve analysis (DCA). Kaplan–Meier analysis was used to estimate the survival rate of DLBCL, and the log-rank test was performed for the difference between groups. All statistical analyses were performed by R software .We utilized the “glmnet” package to fit the LASSO-cox regression and used tenfold cross-validation to select the penalty term, U test and Chi-squared test showed that there was no significant difference in age, gender, WBC, Ki-67, ECOG PS score, and IPI between the training cohort and the validation cohort  with complete data were included in the final analysis. The training cohort consisted of 848 patients, and the validation cohort consisted of 363 patients. The median follow-up time was 43.4 months and the 5-year OS was 58.5%. Mann–Whitney In the random forest model, the error rate was relatively low and stable when the number of survival trees was 490 . The impP < 0.05). Multivariable Cox model based on random forest showed that age, WBC, HB, CNS involvement, ALB, and ECOG were indicators for the survival of DLBCL patients . This result was not changed in the validation cohort .The LASSO model achieved an AUC of 75.8% (95% CI: 71.4%-80.3%) for predicting the prognosis of DLBCL in the training cohort, which was higher than the random forest model . The C-index of the LASSO model was higher than that of IPI  and NCCN-IPI .All patients have complete data for the variables required to calculate the IPI and NCCN-IPI scores, and the survival curves are shown in Supplementary Figure P < 0.05). Based on the LASSO model, we used a maximum of 6 scoring points for categorized age (≥70), WBC (≥8.02), HB (<104), male, the presence of CNS involvement, and Ann Arbor stage III-IV, each having a score of 1. Four stratification risk groups were formed based on KM analysis: low risk , low-intermediate risk , high-intermediate risk , and high-risk . The LASSO model showed better discrimination of outcomes compared with the IPI and NCCN-IPI model and identified an LR group, HIR group, and HR group (According to the maximal Chi-squared method, 70, 104, and 8.02 were the optimal cut-off points for age, HB, and WBC, which distinguished two prognostic groups most effectively (HR group .In this retrospective multicenter study, we proved that the LASSO model is superior to the random forest model in predicting the prognosis of DLBCL. In addition, the model based on LASSO regression showed better discrimination of outcomes compared with the IPI and NCCN-IPI and identified a low-risk group, high-intermediate risk group, and high-risk group more precisely.Predictive analysis is an important application of ML. For example, ML has been used to predict the prognosis of many diseases, including COVID-19, lung cancer, and stroke –24. HoweMultivariable Cox proportional regression analyses using prognostic factors identified from LASSO models showed that older age, male sex, higher white blood cell level, lower hemoglobin level, and CNS involvement were risk factors of DLBCL. This is consistent with previous studies –30. FemaThe current prognostic model was developed using LASSO regression based on clinicopathological variables and increased the accuracy to stratify the low-risk, high-intermediate risk, and high-risk groups in newly diagnosed DLBCL, compared to the IPI and NCCN-IPI models. Compared to the IPI model, the NCCN-IPI scoring model applied a refined classification of age and normalized LDH to better predict the risk of death . In thisAccording to the variables screened by LASSO regression, we established a prognostic model with the highest integral at six points, and divided the patients into four risk groups. The most widely used prognostic models, IPI and NCCN-IPI, both included five clinical predictors and identified four risk groups for DLBCL by traditional regression analysis. The 5-year OS of high-risk group identified by IPI and NCCN-IPI were 39.8% and 35.3%, respectively. By contrast, the high-risk group defined by the LASSO model was 22.1%, suggesting that the LASSO model was more accurate in identifying DLBCL patients at high risks than the IPI and NCCN-IPI models. Therefore, clinical applications of the LASSO model may improve the prognosis of DLBCL patients.In summary, in this retrospective study of real-world data, we found that LASSO model was superior to random forest in predicting the prognosis of newly diagnosed DLBCL, although both were superior to the IPI and NCCN-IPI models. More importantly, the prognosis model based on LASSO was more accurate in identifying low-risk, low-intermediate risk, and high-risk patients than the IPI and NCCN-IPI models.


Coronary artery disease (CAD) is the most common cardiovascular disease (CVD), and previous studies have shown a significant association between N-glycosylation, a highly regulated posttranslational modification, and the development of atherosclerotic plaques. Our aim was to determine whether the N-glycome of immunoglobulin G (IgG) is associated with CAD, as N-glycans are known to alter the effector functions of IgG, which may enhance the inflammatory response in CAD. Therefore, in this study, we isolated IgG from subjects with coronary atherosclerosis (CAD+) and from subjects with clean coronaries (CAD−). The purified IgGs were denatured and enzymatically deglycosylated, and the released and fluorescently labelled N-glycans were analysed by ultra-high performance liquid chromatography based on hydrophilic interactions with fluorescence detection (HILIC-UHPLC-FLR). Sex-stratified analysis of 316 CAD− and 156 CAD+ cases revealed differences in IgG N-glycome composition. The most notable differences were observed in women, where the presence of sialylated N-glycan structures was negatively associated with CAD. The obtained chromatograms provide insight into the IgG N-glycome composition in CAD as well as the biomarker potential of IgG N-glycans in CAD. Cardiovascular diseases (CVDs), a group of diseases affecting the heart and blood vessels, are the most common non-communicable diseases globally, responsible for an estimated 18.6 million deaths in 2019 , yet peoGlycosylation is one of the most common and important posttranslational modifications of proteins. It involves a series of enzymatic reactions in the synthesis of carbohydrate structures (glycans) attached to the protein backbone . Unlike A growing body of research shows that alterations in protein glycosylation are involved in the development of cardiovascular disease through various molecular mechanisms and thus have significant biomarker potential for disease development and progression as well as for therapeutic monitoring . For exaImmunoglobulin G (IgG) is the most abundant antibody in human blood and a very important effector molecule in the immune response. IgG Fab fragment (antigen-binding fragment) specifically recognizes and binds various antigens, while the Fc fragment  performs the effector function by binding to various Fc receptors . The IgGConsidering previous research and the importance of glycosylation in CVD and the fact that there is no prospective evidence of altered IgG N-glycosylation in CAD, in this study, we performed an IgG N-glycome analysis of CAPIRE participants with angiographically diagnosed coronary atherosclerosis (CAD+) and participants with clean coronaries (CAD−). Our primary objective was to investigate the cross-sectional association between IgG N-glycome composition and CAD, as well as differences in IgG N-glycosylation occurring during a two-year follow-up period after CAD diagnosis.CAPIRE  is a multicentre, prospective, observational study aimed at identifying new mechanisms promoting or protecting against coronary atherothrombosis; in its longitudinal phase, subjects are being followed for eight years. Details of the study have been reported . BrieflyThe whole study population had a peripheral venous blood sample taken at the inclusion point and after two years of follow-up. The samples were treated to obtain 0.5 mL whole blood, plasma, and serum aliquots and stored in a freezer at −70 °C. Samples of each subject enrolled in the study were collected in a single dedicated biological bank . Personnel unaware of participant characteristics measured all biomarkers in a central laboratory in a single batch.IgG was isolated from the plasma samples by affinity chromatography, as described previously . In brieDeglycosylation of IgG was performed as previously described . IsolateFluorescently labelled N-glycans were analysed by ultra-high performance liquid chromatography based on hydrophilic interactions with fluorescence detection (HILIC-UHPLC-FLR) on a Waters Acquity UPLC H-class instrument consisting of a quaternary solvent manager, sample manager, and an FLR fluorescence detector. The instrument was under the control of Empower 3 software, build 3471 . Samples were kept at 10 °C before injection, and the separation temperature was 60 °C. Chromatographic separation of glycan structures was performed on Waters UPLC Glycan bridged ethylene hybrid (BEH) amide chromatographic columns . Solvent A was 100 mM ammonium formate, pH 4.4, while solvent B was LC-MS grade acetonitrile (ACN). Separation was performed with a linear gradient of 25–38% solvent A at a flow rate of 0.40 mL/min in a 29-min analytical run. Excitation and emission wavelengths were set at 250 and 428 nm, respectively. Data processing was performed using an automatic processing method with a traditional integration algorithm, after which each chromatogram was manually corrected to maintain the same intervals of integration for all the samples. Each chromatogram was separated into 24 N-glycan peaks , and theTo remove experimental variation from the measurements, we performed normalization and batch correction on the UHPLC glycan data. To make measurements across samples comparable, we performed normalization by total area was performed. Prior to the batch correction, normalized glycan measurements were log-transformed due to right-skewness of their distributions and the multiplicative nature of batch effects. Batch correction was performed on log-transformed measurements using the ComBat method (R package sva), where the technical source of variation  was modelled as batch covariate. To correct measurements for experimental noise, we subtracted the estimated batch effects from log-transformed measurements.Association analyses between CAD status and baseline glycomic measurements were performed using a regression model. Analyses included glycan measurement as dependent continuous variable, CAD status was included as independent variable, with age and sex included as additional covariates. Additionally, interaction analyses were performed using a regression model where baseline glycan measurement was included in model as dependent continuous variable, CAD status, and interaction between CAD status and sex (CAD:sex) were included as independent variables, with age and sex included as additional covariates. To gain insight into associations between IgG N-glycome and CAD separately in women and men, we performed sex-stratified analyses. For sex-stratified analyses of association between CAD status and baseline glycomic measurements, two regression models were used. In model 1, glycan measurements were included as dependent continuous variable; CAD status was included as independent variable, with age included as additional covariate. In model 2, glycan measurements were included as dependent continuous variable, and CAD status was included as independent variable, with age, BMI, smoking, and diabetes included as additional covariates. For sex-stratified two-time points analyses of samples through their observation period, two linear mixed-effects models (LMM) were implemented. In first LMM model, glycan measurements were included as dependent continuous variable, time, CAD status, age, and interaction between time and CAD (CAD:time) status were modelled as fixed effects, while individual sample ID was modeled as a random intercept. In second LMM model, glycan measurements were included as dependent continuous variable, time, CAD status, age, BMI, smoking, diabetes, BMI:time, smoking:time, diabetes:time, and CAD:time were modelled as a fixed effects, while individual sample ID was modeled as a random intercept. Prior to analyses, glycan variables were all transformed to standard Normal distribution  by inverse transformation of ranks to Normality . Using rank transformed variables in analyses makes estimated effects of different glycans in different cohorts comparable as transformed glycan variables have the same standardized variance. False discovery rate was controlled using Benjamini–Hochberg procedure (function p.adjust(method = “BH”)).Data were analyzed and visualized using R programming language (version 4.0.2).IgG N-glycome composition was analysed in CAPIRE participants classified by CCTA into CAD−  and CAD+ , whose samples were collected at the inclusion point and after the two-year follow-up period. Descriptive information on the included participants at the inclusion point is presented in To gain insight into the IgG N-glycome composition of participants at the time of inclusion in the study, we first compared the baseline glycomic measurements between the CAD+ and CAD− cases for both women and men. At the inclusion point, statistically significant differences were observed when comparing CAD+ and CAD− women, while the same was not observed in men. Based on model 1, in women, IgG sialylation was significantly negatively associated with CAD+ . In partp-value < 0.05, see Sequentially, we investigated whether the same trend of associations between IgG N-glycosylation and CAD continued during the two-year follow-up period after CAD diagnosis. Differences in IgG N-glycome composition observed during the follow-up period in CAD+ and CAD− cases are shown in This study represents the first comprehensive analysis of IgG N-glycosylation in coronary artery disease (CAD). To investigate the association between coronary atherosclerosis and the composition of IgG N-glycome, we performed the N-glycomic analysis of IgG isolated from plasma of subjects with angiographically diagnosed coronary artery disease (CAD+) and subjects with clean coronary arteries (CAD−) collected at inclusion and after a two-year follow-up.As previous studies demonstrated evidence of sexual dimorphism in CVD  as well In men, however, the differences were minor and revolved mainly around the presence of bisecting GlcNAc. At the inclusion point, GP24 was significantly positively associated with CAD. Furthermore, results after the two-year follow-up period demonstrated only a marginally positive association between core-fucosylated N-glycan structures with bisecting GlcNAc and CAD. However, this association was previously observed in a study that found sex-stratified associations between IgG N-glycans and the incidence of CVD . InteresHowever, even though in this study, we found an association between IgG N-glycome and CAD, we were not able to illuminate the mechanism behind those associations. Thus, it is known that in addition to direct effects on the immunomodulatory function of IgG, alterations in the glycosylation profile have also been associated with cardiovascular risk factors , which aThis study, to our knowledge, is the first study to investigate the association of total IgG N-glycome with coronary artery disease. Nevertheless, there are limitations to our study. First, despite the multicenter design of the CAPIRE study, only European centers with Caucasian patients were involved. Therefore, our conclusions should not be extrapolated to non-European populations because the results may not reflect the general population of other countries. Second, patients might have changed their medication regimen or lifestyle after coronary CTA; we were not able to assess the impact of this possible bias on our results. Finally, our study lacks a validation cohort; however, we are planning to set up another study to validate our results.In conclusion, our study demonstrated several significant cross-sectional associations between IgG N-glycosylation and CAD. Further research on IgG glycosylation as a potential biomarker for CVD prevention and early diagnosis is encouraged.


We present the unified method and use it to integrate the ion sound and Langmuir waves (ISLW) model to retrieve optical soliton solutions. Some new dynamical optical solitons involving the combo of rational, trigonometric, and hyperbolic function solutions are added in this study. The derived optical soliton solutions display various properties such as beat pattern and oscillation with increasing, decreasing, and simultaneously increasing and decreasing amplitudes. Moreover, kink, dark bell, singular kink, single breather, multiple breathers, dark-, bright-, and dark bright periodic waves are founded. Finally, some dynamical characteristics of the acquired solutions are depicted. Unified method; Ion sound and Langmuir waves (ISLW) model; Optical soliton; Beat pattern We cannot imagine the operating of optical fiber, email, internet, mobile phones and many other communications except the idea of a solitary wave 2Q signifies the electrostatic field excitations, R denotes the normalized density perturbation. In engineering and applied sciences, especially in high-frequency cases, the ISLW model is applicable for the impact of ponderomotive force.The ISLW model has the following form 3w are real parameters need to be measured afterward. Combining Eq. ψ, we haveAssume the subsequent relation for solving Eq. 4Suppose the trial solution of Eq. Case-01: Hyperbolic function (when M is negative):Case-02: Trigonometric function (when M is positive):Case-03: Rational function  by x. We see that (a) displays oscillations with rapidly growing amplitudes at (b) illustrates oscillations with rapidly reducing amplitudes, then drops to 0, and then again oscillations with rapidly growing amplitudes at (c) exhibits oscillations with rapidly reducing amplitudes at (a-d) by (a-c). We see that for (a)); for (b)), it remains the same; and for (c)). The solutions (a-d) by (a-f). When (a-c)). When (d-f)). The solutions . . . The solutions  by the solution  by (a-c). This solution represents a dark periodic wave if (a)), a dark-bright periodic wave if (b)), and a bright periodic wave if (c)). We can say that the results acquired for the ISLW model in the unified approach are unprecedented and newest.The solutions 6Fig. 6. Single breather and multiple breather waves are shown in Fig. 7. Moreover, dark, dark-bright, and bright solitons are shown in This paper has presented the unified method to integrate the ISLW model to retrieve optical solitons that can be used in birefringent fibers. Dulal Chandra Nandi: Analyzed and interpreted the data; Contributed reagents, materials, analysis tools or data. Mohammad Safi Ullah: Conceived and designed the experiments; Wrote the paper. Harun-Or-Roshid; M. Zulfikar Ali: Performed the experiments.This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.No data was used for the research described in the article.The authors declare no conflict of interest.No additional information is available for this paper.


Musca sorbens can act as a vector for ocular Chlamydia trachomatis, causing trachoma, yet there has been very little research on control measures. We investigated whether insect repellent products, specifically insecticide-treated clothing, could provide personal protection to the user from eye-seeking flies.The eye-seeking fly We first conducted a series of phase I laboratory studies to inform our choice of field intervention. We then conducted a phase II randomised controlled trial testing the efficacy of permethrin-treated scarves (PTS) in reducing fly-face contact in Oromia, Ethiopia. Children aged 4-10 years in full health and with no known adverse reactions to permethrin or other insecticides were allocated to either arm using restricted randomisation. Intervention arm children wore Insect Shield® versatile wraps (as PTS) for 28 days. The primary outcomes, fly-eye, -nose and -mouth contact, were assessed on the first day (0/30/60/180 minutes), on day 7 and on day 28. All participants present per timepoint were included in analyses. This trial was registered with ClinicalTrials.gov (NCT03813069).Participants were recruited to the field trial between 29/10/2019 and 01/11/2019, 58 were randomised to test or control arm. More fly  contacts were observed in the PTS arm at baseline. After adjusting for baseline contact rates, across all timepoints there was a 35% decrease in fly-eye contacts in the PTS relative to control arm . Similar cross-timepoint reductions were seen for fly-nose and fly-mouth contacts . All children were included on day 0. Two in the control arm were absent on day 7, one left the study and four were excluded from analysis at day 28. No adverse events occurred in the trial.Musca sorbens flies are sufficiently repelled by PTS to reduce fly-eye contacts for the wearer, thus possibly reducing the risk of trachoma transmission. Permethrin-treated scarves may therefore an alternative to insecticide space spraying for protection from these flies.Wellcome Trust. Musca sorbens in the transmission of the blinding eye disease trachoma, there are scant evidence-based measures for their control. Insect repellents and insecticide-treated clothing can provide personal protection to the user, and many arthropod species are susceptible to their active ingredients. We searched MEDLINE, Global Health, Embase Classic and Embase with no restrictions on date or language, combining all terms and synonyms for M. sorbens with search terms related to insect repellents, insecticides and insecticide treated clothing. We retrieved 38 publications, none of these studies tested the use of insect repellents or insecticide-treated clothing against M. sorbens.Despite the role of eye-seeking Musca sorbens populations, like other species of filth fly, can increase exponentially given favourable breeding conditions. For this reason, fly control by population suppression, including insecticide spraying, may not be an optimal control tool. However, personal protection from fly-eye contacts may be useful and appropriate in the trachoma-endemic context. In our phase II study in Oromia, central Ethiopia, children aged 4-10 years who wore permethrin-treated headscarves experienced fewer fly-eye contacts relative to those in a control group given no intervention. To our knowledge, this is the first study of the use of insecticide-treated clothing against M. sorbens. Our findings support further large-scale studies of the use of permethrin-treated headwear against eye-seeking flies.Permethrin-treated clothing should be further tested in large-scale phase III trials with both epidemiological (trachoma) and entomological (fly-eye contact) outcomes. Such studies will inform whether the distribution of permethrin-treated clothing within communities can reduce fly-eye nuisance at scale and bolster trachoma elimination programmes.Alt-text: Unlabelled boxChlamydia trachomatis (Ct) initially causes inflammation, which over time can lead to scarring, entropion, and blindness.Chlamydia trachomatis is an intracellular bacterium, whose unusual lifecycle involves stages that are adapted for extracellular life, called elementary bodies (EB). These are shed by an infected person in ocular and nasal secretions. From here they are transmitted to another person mainly via close contact, or contamination of surfaces. Routes of transmission are, therefore, multiple, and their relative importance is presumed to vary by setting.Trachoma is a progressive eye disease. It is the most common infectious cause of blindness globally,Musca sorbens is a synanthropic ‘filth’ fly which feeds on ocular discharge and from mucous membranes of humans.M. sorbens to become contaminated with Ct EBs, then transport them from one person to another, acting as a mechanical vector of disease.M. sorbens to Ct transmission is likely to be dependent on the local fly population density, as well as possible local variation in fly behaviour, biology or ecology. Partly because of this variability, the contribution of M. sorbens to trachoma transmission is not fully characterised nor understood. Further, practical control methods for this disease vector are lacking, with space spraying with pyrethroid insecticides being the only evidence-based option.,Schinus mole, are laid down in houses to deter flies. An alternative method of protection could be the use of insecticide-treated clothing (ITC). Insecticide-treated clothing incorporates insecticides that have spatially repellent properties or are contact irritants, and may be a practical and contextually appropriate means to deter eye-seeking flies. While ITC has been shown to provide protection from malaria and leishmaniasis in some contexts,,Personal protection from the aggressive eye-seeking behaviour would both protect against disease transmission and alleviate distress. Insect repellents are used world-wide to prevent nuisance biting by both vector and non-vector arthropod species. Commercially available topical repellents are rarely used by people in low-income countries with endemic arthropod-borne diseases, because of availability, cost and the impracticality of a product that requires repeat application. In some regions, plants with repellent properties are used, either by burning leaves or laying out fresh foliage,M. sorbens. We anticipated that in areas with high fly density, an immediate benefit of reduced fly-face contact may encourage uptake of this intervention. We first screened the use of the topical repellents DEET , IR3535 (ethyl 3-[acetyl(butyl)amino]propanoate), Picaridin (butan-2-yl 2-(2-hydroxyethyl)piperidine-1-carboxylate) and delta-Undecalactone , as well as Craghopper and Insect shield ITC garments (both containing permethrin (3-phenoxyphenyl)methyl 3--2,2-dimethylcyclopropane-1-carboxylate), against M. sorbens fly-skin contact in laboratory bioassays. The screening process led to our testing topical IR3535 (Jungle Formula Kids) and permethrin-treated scarves (PTS) in a small-scale laboratory-based trial, which in turn informed our choice of intervention for the field trial. We then conducted a randomised controlled trial in Oromia, Ethiopia, testing whether PTS could protect against fly-face  contact in children aged 4-10 years.We hypothesised that insect repellents may provide personal protection against the eye-seeking M. sorbens, we screened a range of these for possible efficacy . This work led to a small-scale preliminary laboratory study in which six participants tested the topical repellent Jungle Formula Kids® , and permethrin-treated scarves , for protection from skin contact by M. sorbens .We first conducted a series of phase I studies to select our repellent intervention. After designing arm-in-cage laboratory bioassays for testing repellent products against the eye- and skin-seeking M. sorbens,The results of this preliminary study informed our choice of intervention for the subsequent phase II randomised controlled field trial , in whichttps://doi.org/10.17037/DATA.00002423) was approved by the LSHTM ethics committee (LSHTM Ethics Ref: 15049‑01), the National Research Ethics Regulatory Committee Ethiopia/Ministry of Science and Higher Education and the Food, Medicine and Healthcare Administration and Control Authority of Ethiopia (02/25/33/06).All participants in all studies were free to withdraw at any time without giving a reason; no participant withdrew. The study protocol  or nothing (control arm) and were allocated to the two trial arms by the trial statistician using restricted randomization. The factors on which allocation was restricted were age, where the difference in mean age was restricted to be within 0.5 years in the two groups, and sex, where the difference in numbers of males between the two groups could not be more than one. A total of 100,000 possible permutations of allocations to arms were generated and those permutations that did not meet the restriction criteria were discarded. Of the remaining acceptable permutations, one was chosen using a computer-generated random number. Neither data collectors  nor participants were masked to arm allocation.2, total amount 102 [PTS102] and 204 [PTS204] mg/scarf). These candidate products were tested for their protection against fly-skin contact using modified arm-in-cage bioassays to calculate the protective efficacy . The permethrin-treated scarves used as the field trial intervention were Insect Shield® versatile wraps (referred to as PTS) , and hand-dipped permethrin-treated scarves  (permethrin 0.017 and 0.034 mg/cm as PTS) . These w as PTS)  while ma. The control arm participants were only asked to wear a placebo scarf during the 10-minute fly-face observation periods.The primary outcomes were fly-eye, -nose and -mouth contact, measured over several time points. On day 0 of the study, baseline measurement of fly-eye, -nose and -mouth contact was made. We collected data on several personal and environmental variables including ocular and nasal discharge , participant biometrics  and environmental variables (ambient temperature (°C), light intensity, relative humidity (RH%), presence of rain or wind) as secondary outcomes. For baseline primary outcome measurement, all participants wore a placebo scarf while ten minutes of fly-face contact were recorded: participants sat on a chair, facing entomological field workers who tallied fly-eye, -nose and -mouth contacts from a distance of approximately 1.5 m. We defined (1) fly-eye contact as a fly touching the eye, lid edge or eyelashes; (2) fly-nose contact as a fly touching the nostril area; (3) fly-mouth contact as a fly touching the lips or lip margin. Repeat contacts by the same fly were recorded as new contacts, as differentiating between new and repeat contacts was not always possible. Observation periods were also filmed using a tripod-mounted camera (Nikon D7200 single-lens reflex). After baseline measurements, participants were given scarves , then another ten minutes of fly-face contact was immediately recorded for all participants . Fly-face contact measurements (10-min sessions) were then repeated at 30 minutes (T30), 1 hour (T60) and 3 hours (T180) later, with intervention arm participants continuing to wear their PTS from T0 onwardsAfter the first day, participants in the intervention arm were encouraged to wear their PTS throughout each day for the 28-day duration of the study, which was governed by a compromise between maximising weeks of follow-up to measure continued efficacy and time constraints for study completion. Observations for all participants were repeated on day seven (D7) and day 28 (D28). On those follow-up days, fly-face contact measurements were done as per T0 measurements with both a control measurement  and a test measurement . Secondary outcome measures were repeated on follow-up days. We encouraged adherence to the study (continued wearing of the PTS from day 0 to day 28 in intervention arm) at each visit by emphasizing that PTS could protect from trachoma and diarrhoeal disease transmission by flies. Between these visits, we made phone calls to a nominated local person who had a mobile phone, to encourage the participant's caregivers to support the child in adhering to protocol.PE = 100*(1-(T/C))).Study data were electronically captured in encrypted case report forms using tablets  programmed with the open-source survey tool kit ODK Collect. These were uploaded to a secure server as close to daily as possible. Protective efficacy (PE) against fly-face contact per participant was the planned effect measure, with PE calculated as the proportion of fly-face  contact after application of the PTS (T) in relation to contacts before application of the PTS at baseline (C)  contacts, as we observed a large imbalance in fly-face contacts at baseline. On follow-up days D7 and D28, control observations (10 minutes with placebo scarf) were performed for all participants as well as test observations (10 minutes with scarf according to intervention arm). We compared fly-face contact in these control observations using the same method.To obtain an average estimate of the effect of trial arm on fly-eye, -nose and -mouth contact across all timepoints, a random-effects negative binomial regression model was used with fly  contacts in ten minutes as the outcome, trial arm as the primary exposure and including timepoint as a categorical exposure. The same model was used to test whether the effectiveness of the scarf changed over time, also including an interaction term between day and trial arm. To assess the association of other personal and environmental exposures with the fly-face outcomes at baseline, negative binomial regression was used with exposure variables included one at a time. Associations were checked for potential confounding by adjusting for each of the unused variables one by one and the effect estimate checked for a substantial (∼10%) change in size. If multiple confounders were identified, then all were included in the model, however, because of the relatively small sample size we limited multiple adjustments to no more than four.All statistical analyses were done in STATA version 15.1. The trial is registered with ClinicalTrials.gov (NCT03813069).The funder of the study had no role in study design, data collection, data analysis, data interpretation, or writing of the report. All authors had full access to all the data in the study and had final responsibility for the decision to submit for publication.M. sorbens when applied to cover all exposed skin . This would not be an appropriate intervention for protecting the peri-ocular area. Mixed levels of protection were observed for permethrin-treated clothing . In the preliminary laboratory trial, PTS provided protection from fly contact that increased over time, from no observable protection in the first minute , to good protection in the eighth minute  (Our repellent screening tests indicated that higher concentrations (15-20%) of topical repellents were effective against  - 74·6) A. Surpri - 74·6) B. This w - 74·6) , despiteSixty-seven participants consented and were recruited between Oct 29, 2019 and Nov 01, 2019 in the Kubi Guta kebele of Shashemene woreda, Oromia. Sixty-four were screened for eligibility, three found to be non-eligible, and the oldest three remaining children excluded to achieve the sample size (58). These participants, aged 4-10 years, were randomised to test or control arm , Table 1We observed a greater number of fly-eye, -nose and -mouth contacts in the permethrin arm than in the control arm at baseline prior to any intervention  , Table 2P=0·04). Children in the intervention arm continued to wear their PTS throughout the first day of testing during which the adjusted results were fairly consistent: at follow-up timepoints T0, T30, T60 and T180, there were estimated reductions in fly-eye contacts of between 40 and 50% . Across all timepoints, we found a 35 % reduction in fly-eye contacts in the intervention arm relative to the control arm , and we found no evidence that the effect of the intervention changed over time (P=0·38).After adjustment for the baseline value, evidence was found that children in the intervention arm immediately  experienced almost 50% fewer fly-eye contacts compared to children in the control arm , and thereafter, estimated reductions of 30-50% (P=0.12). Our analysis of protection against fly-eye contact as measured by protective efficacy provides some evidence of protection in the intervention arm at all timepoints  . The unadjusted cross-timepoint analysis indicated a reduction of 24% of fly-nose contacts , but no overall evidence of a reduction in fly-mouth contacts .Fly-nose and fly-mouth contact rates followed similar trends to fly-eye contact rates . After aP=0·05 [adjusted analysis]; RR 0·4, 95% CI 0·16-1.02, P=0·06 [unadjusted analysis])  (On follow-up days (D7 and D28), placebo scarf observations were made for participants in both arms. On D7, we found evidence that children in the intervention arm were afforded ‘residual’ protection from fly-eye contact  . There walysis]) , and no alysis]) . ApproxiOverall, flies were observed to visit eyes more frequently than the nose or mouth, with fly-eye contacts made approximately 1.8 and 2.8 times more frequently than fly-mouth and fly-nose contacts respectively. In all participants at baseline (prior to scarf distribution) we found no evidence that individuals with ocular discharge experienced more fly-eye, -nose or -mouth contact  contacts in our baseline (pre-intervention) measures, with greater number of contacts in the PTS arm. Increased fly contacts are indicative of either higher fly density or increased host-seeking/aggression in the locality of the PTS arm children. Within 60 minutes, adjusted and unadjusted analyses show little difference between arms, indicating that either the source of imbalance was redressed, or the intervention effect was sufficient to overwhelm it. As such, adjusting for baseline values of fly contact has a large impact before 60 minutes, but little impact thereafter. While both analyses are given here, we present adjusted analyses as the primary results on the basis that it is correct to take the baseline imbalance into consideration.M. sorbens in our PTS laboratory bioassays. An insect's response to an insecticide is dependent on intrinsic and extrinsic factors and is dose-dependent, but can be broadly characterised in its action as toxic, contact irritant or spatial repellent.M. sorbens, that is, these flies are stimulated to move away without making physical contact with the fabric. This mode of action allows the PTS to protect the user's skin even when not directly covered with fabric. In our laboratory studies, the action of permethrin developed over 8-minute test bioassays and continued to exert an effect for at least 8 minutes after the impregnated garment had been removed (control ‘after’ bioassays). This time-delay for the effect may explain the variable protection afforded by commercial ITC in screening tests (Appendix 1) in which fly contact was measured at minute 4-5. The ‘immediate’ effect of the PTS in the field study is consistent with this short time delay, as it took five to ten minutes to set up and prepare for the T0 measurement. Another study found that the full effect of permethrin on mosquitoes took several hours to develop.Our field trial tested permethrin-treated scarves, treated with the pyrethroid insecticide permethrin. Permethrin is a neurotoxin, although it is important to note that synthetic pyrethroids are some of the least toxic-to-mammals insecticides in use,,,In the field trial intervention arm, across all timepoints we found a 35% decrease in fly-eye contact relative to the control arm. Insecticide space spraying was associated with a 96% and 88% reduction in fly-eye contact in two studies conducted in The Gambia,M. sorbens. Insect repellents have been used successfully against the Australian bush fly M. vetustissima  (Muscidae),M. sorbens complex.M. domestica (Linneus) (Muscidae) and 20% M. vetustissima) in an area 1 metre from treated surfaces.Stomoxys calcitrans (Linneus) (Muscidae).27To the best of our knowledge, to date this is the first trial of the use of any repellent product against African P values should be taken in the context of multiple testing. However, fly-eye, -nose and -mouth contacts on an individual are highly correlated outcomes  and so cannot be considered independent outcomes. We therefore report the P values as calculated rather than adjusting for multiple testing. Finally, fly contacts were measured by trained entomological fieldworkers, and while this is an accepted method of measuring nuisance from eye-seeking flies,Despite preparatory phase I studies, modifications were made to the study protocol for data analysis after fieldwork had commenced. Protocol modifications were made appropriately, in response to evolving knowledge of how permethrin affected the flies and seasonal changes in the field setting (i.e. increased fly density as the study progressed into the hot season). It is important to note that while the effect measure for the primary outcomes, protective efficacy, was replaced with analysis of the rate ratio, this alteration did not change the underlying parameters and assumptions of the study design. Both effect measures compare fly contact rates between arms. The performance of multiple tests (three primary outcome measures at six timepoints) could be considered a further limitation of the analysis, therefore It will be important to replicate and measure the longevity of PTS repellency, ideally in a large-scale phase III study. Insect shield® items are factory treated with permethrin such that the repellency should last 70 washes. While the assocation between trachoma prevalance and water scarcity may reduce the number of washes garments experience, high levels of UV light in trachoma-endemic environments may contribute to rapid degradation of permethrin. It might be useful to also trial other permethrin-treated items, for example hand-dipped scarves, which could be cheaper (but might have reduced longevity of repellency). We found evidence that on D7 of the trial children in the intervention arm still benefitted from fewer fly-eye contacts in the absence of the PTS. This is supportive of the possibility that residual permethrin on the skin continued to repel flies,The phase II studies described here show that PTS can protect against eye-seeking flies. Larger, phase III trials with epidemiological outcomes are warranted to corroborate and extend these findings. Permethrin-treated clothing could be an effective weapon against the transmission of trachoma.AR, JGL, AL and MJB conceived the study, AR and JGL designed the study, AR, LROG, OSA, WA, GS, EM, MG, DL, EE, KT, KHK, SML, SLW, EH, DA, FS and MAA implemented the study, AR and DM verified the underlying data, carried out the statistical analysis and interpreted the results, AR did the literature search and wrote the first draft of the manuscript, JGL, MJB, AL and AWS supervised the study, VS, JGL and MJB obtained funding for the study. All authors reviewed and approved the final manuscript before submission.The authors have nothing to disclose.https://doi.org/10.17037/DATA.00002423).De-identified individual participant data on which statistical analysis including summary figures and tables are based, an accompanying data dictionary defining each field, and related documents including the study protocol, participant information sheets and consent forms will be made available from the point of, and for 10 years after, the acceptance for publication of the main findings from the final dataset. Datasets will be publicly available at LSHTM Data Compass (


Children of lower-educated parents and children in schools with a relatively high percentage of peers with lower-educated parents  are more likely to develop emotional and behavioural problems compared to children in higher-educated households and schools. Universal school-based preventive interventions, such as the Good Behaviour Game (GBG), are generally effective in preventing the development of emotional and behavioural problems, but information about potential moderators is limited. This study examined whether the effectiveness of the GBG in preventing emotional and behavioural problems differs between children in lower- and higher-educated households and schools.Mage=6.02 towards the end of kindergarten) from 31 mainstream schools  were followed annually from kindergarten to second grade (2004–2006). The GBG was implemented in first and second grades.Using a longitudinal multi-level randomized controlled trial design, 731 children . No moderation by household-level parental education was found.Overall, the GBG prevented the development of emotional and behavioural problems. However, for emotional problems, the GBG-effect was slightly more pronounced in higher parental education schools than in lower parental education schools (Studies into universal school-based preventive interventions, and in particular the GBG, should consider and incorporate school-level factors when studying the effectiveness of such interventions. More attention should be directed towards factors that may influence universal prevention effectiveness, particularly in lower parental education schools. Poor mental health among school-aged children, including emotional and behavioural problems, is a global public health concern.The GBG has previously been referred to as a ‘behavioural vaccine’ due to its cost-effectiveness and its ability to prevent mental health problems across diverse cultures and populations.,Across nations, a robust risk factor of poor child mental health at both the household and school levels is low socioeconomic status (SES).,,,,The majority of the school-based intervention studies on children’s emotional and behavioural problems have not included household- or school-level parental education or only included SES as a descriptive or a study variable.Participants were recruited from the first 31 elementary schools in rural and urban areas of the Netherlands that agreed to participate in the research project. Schools could participate if they were willing to implement the GBG (if randomly selected in the intervention arm) or if they were willing to be on a waiting list (if randomly selected in the control arm).Mage=6.02, SD= 0.46) to second grade (in spring). Inclusion criteria were (i) active parental consent, (ii) data on school-level parental education and (iii) at least two out of three completed waves of teacher-reported data on emotional and behavioural problems. In total, out of 825 children who were initially included in the study, 731 (50% girls) fulfilled these criteria  or the GBG intervention arm . See Participating schools were randomly assigned, with an oversampling of intervention schools, to either the control  is extended. More information regarding the intervention strategy, implementation and teacher training is described elsewhere.Household-level parental education was based on the highest education level per household, obtained by the (two) parent/caregiver(s). Parental education levels were ranked according to the Dutch Standard Education Classification,School-level parental education levels were determined by the per-school percentage of children of low-educated parents. In the Netherlands, school-level socioeconomic inequalities are measured by children’s parents’ education levels. The Netherlands Inspectorate of Education calculates the percentage of low parental education levels of each school to identify schools that qualify for additional governmental resources.www.duo.nl).Teacher ratings of individual children’s behavioural and emotional problems were assessed by the Problem Behaviour at School Interview (PBSI).Intervention status was dummy-coded .Covariates included gender  and cluster size. Cluster size (i.e. number of participating children per school) was grand-mean centred and included to account for unequal cluster sizes . Baseline differences in kindergarten were controlled for because—despite randomization—children in the GBG arm had moderately higher levels of emotional  and slightly higher levels of behavioural problems  than children in the control arm.A parallel latent growth curve (LGM) model with two-level time-nested-within-individual data structure , in which the development of emotional and behavioural problems was conceptualized by latent growth parameters (intercept and a linear slope), was used to test the main effects and potential moderation by household- and school-level parental education of the GBG in preventing the development of emotional and behavioural problems. The intercept represented the initial level in kindergarten (baseline) and the slope represented change over time (from kindergarten to second grade).nc−1)ICC].The analyses were conducted in three steps. All models were fitted in Mplus version 8.0.N = 25 imputed datasets) and (ii) by testing the models on a subsample (N=596) with complete household-level parental education data.Model fit indices for multi-level latent growth models were used to determine model fit at both the household and school levels. For specifics, see Supplementary appendix Bwww.trialregister.nl). Signed parental informed consent was obtained from parents. Parents and children could revoke participation at any time.This study was approved by the Medical Ethics Committee of the Vrije Universiteit Amsterdam Medical Center and was registered with the ‘Netherlands Trial Register’ [Trial NL470 (NTR512)] (t(1)=2.75, P=0.006, Cohen’s d=0.24.Descriptive statistics of household- and school-level parental education of the whole sample are presented in M=18.61%, SD=23.97%) and intervention arms , t(29)=0.44, P=0.666, Cohen’s d=0.17. The correlation between household- and school-level parental education in our sample was positive and of moderate magnitude .The per-school percentage of children of low-educated parents was not significantly different between the schools in the control . In the control arm, there was a significant yearly increase of emotional problems  and a borderline significant yearly increase of behavioural problems . This indicates that without the GBG, emotional  problems tended to increase from kindergarten to second grade.Results from the unconditional LGMs table S2 B=−0.208, 95% CI , P = 0.003]. In addition, the GBG was also effective in preventing behavioural problems from kindergarten to second grade .Results of main effects  showed tHousehold level: Results showed no significant cross-level interaction between household-level parental education and the GBG-effect on individual-level emotional problem development, B=0.010, 95% CI , P=0.765 , P = 0.005 , P = 0.016] and at 0.50 SD below the mean of school-level parental education . For behavioural problems, no moderation between school-level parental education and the GBG was found, B=0.002, 95% CI , P = 0.382  percentage of children of lower-educated parents, albeit only for emotional problems.,To our knowledge, this study provides preliminary evidence that school-level parental education may impact the effectiveness of the GBG in reducing emotional problems. Previous studies mainly tested household/individual-level factors, such as gender, initial risk status and behaviour type as moderators of universal school-based programmes like the GBG.,The characteristics of lower and higher parental education schools may explain why the GBG was less effective in lower parental education schools for emotional problems. Lower parental education schools may have fewer resources, less effective school management, less teacher support and teachers who are insufficiently prepared to deal with such schools’ diverse populations.The following limitations should be noted. First, and most importantly, we did not have implementation fidelity data. It is possible that there were no major differences in implementation fidelity based on school-level parental education since the interaction effect between school-level parental education and the GBG on behavioural and emotional problems differed. Our study should be considered as an effectiveness trial and an exploratory study meant to stimulate further investigation. It is important to study, for instance, whether the GBG’s weaker effect on emotional problems in lower parental education schools is due to (i) specific school-level factors, (ii) possible problems with implementation or (iii) to lack of components more directly targeting emotional problems. Second, our sample was not randomly drawn from the Dutch population of elementary schools. Third, we used teacher-reports and teachers were not blinded to condition. Self-reports and observational data, which could have provided additional insights, were not available. Fourth, sample size at the between-level was relatively small with 31 schools. Due to this we did not, for example, have enough power to test a three-way interaction of the GBG, household- and school-level parental education. Finally, we used parental education as an index of broader SES. Future replication studies are encouraged to use broader SES indices.Despite these limitations, our results have implications for research and practice. Testing implementation fidelity and school-level moderators that relate to lower parental education schools would result in determining the specific factors to be addressed, such as teacher support and training or implementation infrastructure in schools. Furthermore, if lower parental education schools need more support in preventing emotional problems, more intensified or selective interventions that target high-risk populations could be implemented in these schools. Nevertheless, it is noteworthy that for general prevention efforts the GBG was equally effective in preventing behavioural and emotional problems irrespective of household-level parental education and in preventing behavioural problems irrespective of school-level parental education. Although results suggested that the GBG was less effective in lower parental education schools, it still was an effective tool for preventing the development of emotional problems in these schools. School-based universal interventions reduce the potential that children who may be at risk of developing mental health problems or who may be otherwise difficult to reach will be overlooked. For instance, despite the need for mental health services, it has been shown that the majority of low SES children do not receive treatment.EURPUB online.This study was supported by a grant from the Netherlands Organization for Health Research and Development (ZonMw) (project No. 531003013) and by ZonMw Grants #26200002 and #120620029.T.A.J.H. was funded through a grant awarded by the Norwegian Research Council (project number 288638) to the Centre for Global Health Inequalities Research (CHAIN) at the Norwegian University for Science and Technology (NTNU).Conflicts of interest: None declared.Key pointsThe effectiveness of the GBG in preventing the development of behavioural and emotional problems did not differ between children of lower- and higher-educated parents from kindergarten to second grade.The GBG is equally effective in preventing the development of behavioural problems in schools with higher and lower percentages of children with lower-educated parents, but less effective in preventing the development of emotional problems in lower parental education schools than in higher parental education schools.When testing intervention effectiveness, school-level variables as moderators should be included in study designs.More attention should be directed towards schools with a higher percentage of children with lower-educated parents.ckac143_Supplementary_DataClick here for additional data file.